INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

1738-P increase in tumor size and pulmonary metastasis is observed, compared Sustained Action of Ceramide on Insulin Signaling in Muscle Cells: to wild type mice. In this study, we aimed to determine the mechanisms Implication of the Double-Stranded RNA Activated Protein Kinase through which hyperinsulinemia and the canonical IR signaling pathway drive RIMA HAGE HASSAN, ISABELLE HAINAULT, AGNIESZKA BLACHNIO-ZABIELSKA, tumor growth and metastasis. 100,000 MVT-1 (c-myc/vegf overexpressing) RANA MAHFOUZ, OLIVIER BOURRON, PASCAL FERRÉ, FABIENNE FOUFELLE, ERIC cells were injected orthotopically into 8-10 week old MKR mice. MKR mice HAJDUCH, Paris, France, Białystok, Poland developed signifi cantly larger MVT-1 (353.29±44mm3) tumor volumes than Intramyocellular accumulation of fatty acid derivatives like ceramide plays control mice (183.21±47mm3), p<0.05 with more numerous pulmonary a crucial role in altering the insulin message. If short-term action of ceramide metastases. Western blot and immunofl uorescent staining of primary tumors inhibits the protein kinase B (PKB/Akt), long-term action of ceramide on insulin showed an increase in vimentin, an intermediate fi lament, typically expressed signaling is less documented. Short-term treatment of either the C2C12 cell in cells of mesenchymal origin, and c-myc, a known transcription factor. Both line or human myotubes with palmitate (ceramide precursor, 16h) or directly vimentin and c-myc are associated with cancer metastasis. To assess if insulin with ceramide (2h) induces a loss of the insulin signal through the inhibition and IR signaling directly affects the expression these markers, in vitro studies of PKB/Akt. Extended periods of treatment with palmitate (48h) or ceramide were performed on MVT-1 and human MCF7 cells. 10nM of insulin signifi cantly (16h), however, shows an inhibition of insulin signaling through increased-IRS1 increased the expression of c-myc at 60 minutes and vimentin at 48 hours serine 307 phosphorylation. The double-stranded RNA-dependent protein in MVT-1 cells. Silencing the IR and inhibiting IR signaling decreased c-myc kinase (PKR) could play a central role to mediate long-term ceramide effects on expression in both cell lines. These results imply that the hyperinsulinemia IRS1, as recent studies showed that PKR acts as a key modulator of metabolic may drive tumor growth and metastasis through the IR by increasing vimentin infl ammation, insulin sensitivity and glucose homeostasis in obesity. Here, we and c-myc expression. These observations allow us to begin to understand the show that both PKR mRNA and PKR phosphorylation are increased in muscle different key players that contribute to tumor progression in the setting of T2D. of high fat diet fed mice compared to control mice as well as in myotubes These downstream elements could open the fi eld to new targets for therapy to from diabetic patients compared to human control myotubes. These results improve survival in women with breast cancer and hyperinsulinemia. are confi rmed in vitro in both human and C2C12 myotubes in response to either Supported By: ADA (1-13-BS-108) palmitate or ceramide. Pre-treatment of C2C12 or human myotubes with PKR inhibitors prevents the inhibitory effect of either palmitate or ceramide on IRS1. Finally, we show that c-Jun kinase (JNK) mediates ceramide-activated INTEGRATED PHYSIOLOGY—INSULIN SECRETION PKR inhibitory action on IRS1. Altogether, our data show that ceramide inhibits IN VIVO effi ciently insulin signalling by targeting in a time dependent manner two important actors, e.g. PKB/Akt and IRS1 in myotubes. Guided Audio Tour: Dynamic Regulation of Insulin Secretion (Posters: Supported By: Société Francophone du Diabète 1741-P to 1748-P), see page 13. 1739-P & 1741-P Defective Autophosphorylation of the Insulin Receptor with a Novel A Novel Long-Acting Analogue of Xenin-25 with Promising Anti- 3-basepair In-frame DeletionWITHDRAWN (ΔArg1027 Glu1028) Found in a Patient diabetic Potential with Severe Insulin Resistance NIGEL IRWIN, CHRISTINE MA MARTIN, PETER R. FLATT, VICTOR A. GAULT, Coler- BOLORMAA ENKHTUVSHIN, HIROYUKI TAMEMOTO, SHUICHI NAGASHIMA, aine, United Kingdom NAOKO SAITO, KENT SAKAI, MANABU TAKAHASHI, DAISUKE YAMAMURO, Xenin-25 is a peptide co-secreted with glucose-dependent insulinotropic JUN-ICHI OSUGA, SHIN-ICHI TOMINAGA, SHUN ISHIBASHI, Shimotsuke, Japan polypeptide (GIP) from intestinal K-cells following a meal. Xenin-25 is Type A insulin resistance (IR) syndrome is caused by genetic mutations believed to play a role in glucose homeostasis and possibly potentiate the in the insulin receptor (INSR) gene. We aimed to identify the molecular biological action of GIP. We have investigated the effects of sub-chronic mechanism of severe IR in a 35-years-old nonobese woman who was administration of a longer-acting xenin-25 analogue, xenin-25[Lys13PAL], in diagnosed as having type 2 diabetes at age 31 during pregnancy. She mice fed a high fat (45%) diet. Initial acute studies confi rmed the superior presented acanthosis nigricans, hyperglycemia (fasting plasma glucose- persistent glucose lowering (p<0.05) and insulin releasing (p<0.05) actions of 173mg/dl, HbA1c-10.3%), severe hyperinsulinemia (fasting insulin-66µU/ xenin-25[Lys13PAL] compared to native xenin-25. Twice daily intraperitoneal ml), high C-peptide (2.1ng/ml), normal lipid profi le and absence of polycystic injection of xenin-25[Lys13PAL] for 14 days had no signifi cant effect on energy ovary syndrome. Tests for insulin and INSR antibodies were both negative. intake or body weight. Circulating plasma glucose and insulin levels were To explore the underlying molecular defect of IR, we sequenced the INSR also unchanged. However, on day 14, overall plasma glucose levels during gene. We found a 3-basepair in-frame deletion in exon 17 of one allele of the a glucose tolerance and oral nutrient challenge were signifi cantly (p<0.05) INSR, which is predicted to substitute Glutamine for Arginine and Glutamic lowered by xenin-25[Lys13PAL] treatment. These changes were accompanied acid (ΔArg1027 Glu1028) in the kinase domain. The same mutation was by signifi cant enhancement of intraperitoneal (p<0.05) and oral (p<0.001) found in father’s INSR, who also showed severe IR with diabetes. To clarify nutrient-stimulated insulin concentrations when compared to controls. how the mutation impairs the receptor function, we generated plasmid Moreover, xenin-25[Lys13PAL] treated high fat mice had markedly improved vectors expressing the wild-type (WT) and ΔArg1027 Glu1028 mutants under insulinotropic (p<0.01) and glucose-lowering (p<0.01) actions in response to CMV promoter and transfected them to CHO-KI cells. Blasticidine was used exogenous GIP administration on day 14 when compared to saline controls. for selecting stable transformants. Clones with comparable (expression However, no appreciable change in insulin sensitivity was observed with levels) insulin binding levels (as demonstrated by Western blot analyses) xenin-25[Lys13PAL] treatment. Finally, ambulatory activity was signifi cantly revealed by Insulin binding assays were chosen for further experiments. (p<0.05 to p<0.001) increased during the dark phase in xenin-25[Lys13PAL] Scatchard analyses of insulin binding assay using 125I-insulin showed that mice compared to controls. These data indicate that sustained administration Bmax and Kd were comparable between the two receptors. Western blot of a stable analogue of xenin-25 exerts a spectrum of benefi cial metabolic analysis revealed that the expression levels and proteolytic processing effects in high fat fed mice. This demonstrates the utility of long-acting of the INSR were comparable. Treatment with insulin robustly increased analogues of xenin-25 as novel treatments for type 2 diabetes. autophosphorylation of WT, but not of the mutant INSR. Supported By: ERDF Integrated

POSTERS In conclusion, we found defective autophosphorylation of the INSR with ΔArg1027 Glu1028 mutation, which might cause severe IR.

Physiology/Obesity & 1742-P 1740-P Insulin-like Growth Factor Binding Protein (IGFBP1)-1: Favorable In Insulin Increases the Expression of Vimentin and C-Myc, and Vivo Actions in Glucose Regulation Enhances Breast Cancer Metastasis NATALIE J. HAYWOOD, NADIRA Y. YULDASHEVA, AMIR AZIZ, JESSICA SMITH, ZARA ZELENKO, MARILYN STASINOPOULOS, ROSALYN FERGUSON, RAN PAUL A. CORDELL, MARK T. KEARNEY, STEPHEN B. WHEATCROFT, Leeds, United ROSTOKER, DEREK LEROITH, EMILY J. GALLAGHER, New York, NY, Haifa, Israel Kingdom Women with Type 2 diabetes (T2D) have a 49% increase in breast cancer In humans, the circulating concentration of IGFBP1 has been proposed as related mortality compared with women without T2D. Epidemiological a marker of insulin sensitivity. In prospective studies, low circulating levels studies report that increased endogenous insulin levels and increased insulin of IGFBP1 predict the development of type 2 diabetes. IGFBP1 can impact on receptor (IR) expression are associated with poor survival in breast cancer cellular functions via an RGD (α5β1 integrin binding) motif independent of IGF patients. We used the non-obese female MKR mouse to study the effects of binding. However, whether IGFBP1 is causally implicated in glucose counter- hyperinsulinemia on breast cancer progression. In the MKR mice a signifi cant regulation and could be exploited therapeutically remains unexplored.

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A450 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

We carried out metabolic profi ling of 8-week-old mice over-expressing vs. 21.5±1.2 mg/kg/min) and liraglutide vs. vehicle (22.5±1.6 vs. 16.7±1.9 mg/ human IGFBP1 to determine whether IGFBP1 plays a direct modulatory kg/min) treated NHPs. Our results show a signifi cant increases in glucose role in glucose regulation. To examine whether the RGD motif of IGFBP1 stimulated insulin secretion AUC with exenatide when compared to vehicle is mechanistically implicated in glucose regulation, we administered the (1740±269 vs. 710±227 ng/ml*min, p<0.05) and liraglutide when compared to synthetic hexapeptide GRGDTP (which binds α5β1-integrin) to C57BL/6 mice vehicle (1260±95 vs. 276±316 ng/ml*min, p<0.05). Our results show that at with diet-induced obesity. therapeutic plasma concentrations of liraglutide and exenatide we are able hIGFBP1-tg mice had increased plasma concentration of hIGFBP1 compared to predict insulin secretagogue effi cacy in the NHP, demonstrating the value to wild type littermates (128 v 7ng/ml P≤0.01). Body weight and fasting blood of this model for the optimization of future GLP-1 receptor agonists, as well glucose levels did not differ signifi cantly but glucose tolerance was improved as the characterization other classes of antidiabetic agents. in hIGFBP1-tg mice (AUC of GTT 137 v 213 mmol/L/120min; P≤0.05). Plasma insulin concentration after a 1mg/g glucose bolus was greater in hIGFBP1-tg & 1745-P mice (2.74 v 1.33ng/ml P≤0.05). Acute administration of GRGDTP (40µg IP) Improvements in AIRglu, SI, and DI Continue 2 Years after Gastric signifi cantly improved glucose tolerance in mice with diet-induced obesity Bypass in Patients With and Without Type 2 Diabetes: Results from (AUC of GTT 173 v 264 mmol/L/120min; P≤0.05). the Longitudinal Assessment of Bariatric Surgery Study IGFBP1 has direct actions in glucose regulation and pancreatic insulin JONATHAN Q. PURNELL, BRET H. GOODPASTER, DAVID E. KELLEY, MYRLENE secretion. For the fi rst time, we have shown that the integrin-binding domain STATEN, KAREN FOSTER-SCHUBERT, RONALD L. PRIGEON, GEOFFREY S. JOHN- of IGFBP1 may play an important role in glucose regulation and represents a SON, ABDUS WAHED, DAVID CUMMINGS, PETER J. HAVEL, ANITA P. COUR- putative therapeutic agent in the fi eld of type 2 diabetes. COULAS, DAVID R. FLUM, BRUCE WOLFE, Portland, OR, Pittsburgh, PA, Rahway, NJ, Bethesda, MD, Seattle, WA, Baltimore, MD, Davis, CA & 1743-P Patients with type 2 diabetes (T2DM) have reduced islet secretor y capacity Gut Microbial Metabolite Trimethylamine Inhibits the Toll-Like (ISC) and disposition index (DI) compared to compensatory hypersecretion in Receptor (TLR) Infl ammation Pathway obese insulin resistant non-DM (Controls). We hypothesized that ISC would JULIEN CHILLOUX, JANE F. FEARNSIDE, ALICE R. ROTHWELL, RICHARD H. improve in T2DM undergoing gastric bypass (GBP) but decrease in Controls BARTON, CLAIRE L. BOULANGÉ, JAMES SCOTT, JEREMY K. NICHOLSON, (as insulin sensitivity improved). DOMINIQUE GAUGUIER, NIGEL GOODERHAM, MARC E. DUMAS, London, United 39 subjects with T2DM and 22 matched Controls and underwent a Kingdom, Oxford, United Kingdom, Paris, France frequently sampled intravenous tolerance test before and 6 and 24 months Obesity is associated with systemic low-grade infl ammation characterised after GBP to measure insulin sensitivity (SI), acute insulin response (AIR), by increased levels of circulating proinfl ammatory cytokines such as IL-6. This glucose effectiveness (SG), and DI. low-grade infl ammation can lead to type 2 diabetes through insulin resistance At 6 and 24 months, weight loss was similar in both groups (24% and 29% and/or β-cell failure. We developed a unique approach investigating the for T2DM; 26% and 31% for Controls). SI improved from baseline to 6 and pathophysiological response of 5 inbred mouse strains to a high-fat diet (HFD) 24 months by 210% (from 0.9 to 2.8 10^-5 pM^-1 min^-1, P<0.05) and 380% to characterise their untargeted urinary metabolic signature by 1H-NMR and (from 0.9 to 4.3, P<0.05) in T2DM; and by 160% (from 1.6 to 4.2, P<0.05) and identify metabolic biomarkers of HFD-induced impaired glucose tolerance and 210% (from 1.6 to 5.0, P<0.05) in Controls. At 6 months, AIR increased from diabetes and their underlying molecular mechanisms through pharmacological baseline 83% in T2DM (P<0.05) and decreased by 41% in Controls (P<0.05). target screening of metabolic biomarkers. For instance, we show that the urinary At 24 months, AIR remained increased in T2DM (87%, P<0.05), yet remained excretion of gut microbial metabolite trimethylamine (TMA) is anticorrelated 36% below baseline in Controls (P<0.05). DI increased in T2DM at both 6 with insulin secretion patterns in HFD-fed mice. We then screened (+350%, P<0.05) and 24 months (+790%, P<0.05), and by 60% (P<0.05) and pharmacological targets for TMA and identifi ed a specifi c inhibition of a key 150% (P<0.05) in Controls. In both groups, SG was not different from baseline kinase integrating signals from various TLRs and regulating the transduction at either follow-up visit. pathway for low-grade infl ammation. To further characterise the role of TMA Remission of T2DM following GBP after 6 months is associated with in lowering infl ammation we confi rmed immunomodulatory properties both robust improvements in AIR and SI, with continued improvement after 2 in vitro and in vivo. Our results demonstrate that a gut microbial metabolite years, indicating restoration of ISC in T2DM. Controls also improved SI and is able to hamper LPS-induced infl ammatory response in monocyte cells and DI, with alleviation of compensatory ISC. Our data indicate islet plasticity in mice. In this study, using an original integrated approach, we demonstrate as ISC is preserved not only in non-diabetic obese but also in T2DM, with that urinary TMA is anticorrelated with insulin secretion in HFD mice and that the latter showing progressive improvement for at least 2 years after GBP. TMA inhibits TLR mediated infl ammation, which altogether could participate Longer term follow-up of the LABS diabetes cohort will determine duration in lowering insulin secretion patterns in HFD-fed mice. This work opens new of these benefi ts and whether deterioration in SI, AIR, or both leads to T2DM avenues for the role of gut microbial metabolism in type 2 diabetes. recurrence after GBP. Supported By: MetaCardis Supported By: NIH (DCC-U01-DK066557, M01RR-00037, UL1RR024153, U01DK 66555, UL1TR00012) & 1744-P Development of a Hyperglycemic Clamp in Non-Human Primates to & 1746-P Assess GLP-1 Mediated Insulin Secretion Mitochondrial Pyruvate Carrier 2 Hypomorphism in Mice Leads to BARBARA L. BERNARDO, DAVID A. GRIFFITH, MARGARET S. LANDIS, DEBBIE Defects in Glucose-stimulated Insulin Secretion WANAPUN, DAVID A. TESS, THOMAS S. MCDONALD, AMIT S. KALGUTKAR, KYLE S. MCCOMMIS, PATRICK A. VIGUEIRA, GEORGE G. SCHWEITZER, MARIA JOSEPH B. KUZMISKI, NICHOLAS J. EDMUNDS, Groton, CT, Cambridge, MA S. REMEDI, XIAORONG FU, WILLIAM G. MCDONALD, SERENA L. COLE, JERRY R. To facilitate the research of biopharmaceutical approaches to therapeutic COLCA, ROLF F. KLETZIEN, SHAWN C. BURGESS, BRIAN N. FINCK, St. Louis, MO, modulation of glucose homeostasis, we developed a hyperglycemic Dallas, TX, Kalamazoo, MI clamp model in the non-human primate (NHP). The hyperglycemic clamp Carrier-facilitated pyruvate transport across the inner mitochondrial is a standard technique utilized both clinically and preclinically to assess membrane plays an essential role in anabolic and catabolic intermediary the impact of therapeutic interventions on glucose-stimulated insulin metabolism. The recently identifi ed mitochondrial pyruvate carrier 2 Integrated release. However, very little data exist in the public domain describing this (MPC2) is believed to form a carrier complex with a related protein (MPC1) POSTERS methodology in the NHPs. To characterize the model, we used the GLP-1 to facilitate mitochondrial pyruvate import. We generated Mpc2 knockout receptor agonists exenatide and liraglutide in healthy, adult cynomolgus mice by using zinc fi nger nuclease technology. Constitutive and complete Physiology/Obesity macaques. IV infusions of the GLP-1 receptor agonists were designed to Mpc2 defi ciency resulted in early embryonic lethality in mice, while achieve clinically relevant therapeutic plasma concentrations; plasma heterozygotes appeared visually and phenotypically normal. However, a concentration levels of exenatide were maintained at 0.5-1.5 ng/ml, second line of Mpc2 targeted mice expressed an N-terminal truncated MPC2 liraglutide levels were maintained at 550-650 ng/ml. 30 minutes following protein missing 16 amino acids (Mpc2Δ16) due to an alternative start codon. initiation of drug infusion, a priming dose of glucose was infused to reach Mpc2Δ16 mice were viable but exhibited reduced MPC2 and MPC1 protein steady state hyperglycemia of 160 mg/dl which was maintained by variable abundance, potentially due to diminished interaction between MPC1 and adjustment to the infusion rate every 5 minutes for 2 hours. Plasma insulin MPC2 Δ16 proteins. The Mpc2Δ16 mutation also resulted in reduced capacity and c-peptide were measured every 15 minutes throughout the experiments. for mitochondrial pyruvate oxidation, suggesting decreased mitochondrial The glucose infusion rate required to maintain the targeted hyperglycemia pyruvate transport. Consistent with this, metabolic studies demonstrated was signifi cantly increased (p<0.05) in both exenatide vs. vehicle (30.2±1.2 exaggerated blood lactate concentrations after pyruvate, glucose, or insulin

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A451 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

challenge in Mpc2Δ16 mice. Compared to WT controls, Mpc2Δ16 mice exhibited (P≥0.11 for all). In conclusion, subgroups of pre-diabetes and diabetes exhibit normal insulin sensitivity but elevated blood glucose after bolus pyruvate reduced GLP-1 release following OGTT, indicating that early GLP-1-based or glucose injection. This was attributable to reduced plasma insulin therapy could be particularly relevant for these individuals. concentration during glucose challenge and decreased glucose-stimulated insulin secretion (GSIS) by isolated islets. Defective GSIS in Mpc2Δ16 mice was corrected by glibenclamide, a sulfonylurea KATP channel inhibitor, in both isolated islets and in vivo during glucose challenge. Collectively, these data are consistent with a role for Mpc2 in mitochondrial pyruvate import and suggest that Mpc2 defi ciency results in defective pancreatic beta cell glucose-sensing.

& 1747-P Effect of Diabetes Duration on Improvement of Glucose Metabolism After Gastric Bypass Surgery EVA SVEHLIKOVA, THOMAS R. PIEBER, VERA HÖLLER, BARBARA OBERMAYER- 1749-P PIETSCH, OANA FREISINGER, FRIEDRICH TADLER, BARBARA ERNST, BRITTA Lack of Synergy for Glucagon and GLP-1 on Acute Beta-Cell Function WILMS, MARTIN THURNHEER, BERND SCHULTES, Graz, Austria, St. Gallen, and Insulin Sensitivity in Mice Switzerland GIOVANNI PACINI, BO AHRÉN, Padova, Italy, Lund, Sweden Gastric bypass surgery improves glycemic control but the underlying Combined activation of glucagon and GLP-1 receptors is a potential mechanisms are incompletely understood. The aim of the study was to weight reducing treatment. A synergistic action on beta cell function would compare the changes in beta cell function and insulin sensitivity after add a value of the combination in subjects with diabetes, but would risk gastric bypass in morbidly obese patients with short- and longstanding type hypoglycemia in non-diabetic subjects. We therefore examined acute 2 diabetes (DM). effects of glucagon+GLP-1 on beta cell function and insulin sensitivity in Before, 8 to 21 days and 1 year after the surgery, 32 DM patients (16 with model experiments in mice. Anesthetized mice underwent intravenous diabetes duration ≥8 years) underwent an oral glucose tolerance test (OGTT) glucose tolerance test with glucose alone or with glucagon, GLP-1 or their and a botnia clamp combining an intravenous glucose tolerance test (IVGTT) combination. Insulin secretion was evaluated as area under the curve with a subsequent hyperinsulinemic-euglycemic clamp. (AUCINS), incremental acute (fi rst-phase) insulin response (ΔAIRg) and beta In the whole group, glucose tolerance improved after the surgery cell sensitivity (BCF=AUCINS/AUCGLUCOSE). Insulin sensitivity (SI) was obtained (p<0.001). In the OGTT total insulin secretion (AUCCP: p<0.001) as well as from modeling analysis. the early insulin response (p<0.001) displayed an early increase followed AUCINS, ΔAIRg and BCF were all augmented by both glucagon and GLP- by a decline or constant values respectively. At 1 year we observed an 1 compared with glucose alone whereas glucagon+GLP-1 did only increase improved C-peptide secretion pattern with a shorter time to peak C-peptide ΔAIRg and not AUCINS or BCF. SI was reduced by glucagon and not altered by in OGTT (p<0.01). Insulin response to the IVGTT did not improve. Insulin GLP-1 or the combination.We conclude that there is no synergistic action on sensitivity (M-value: p<0.001) and the disposition index (p<0.01) increased beta cell function by glucagon and GLP-1 in combination, whereas glucagon- postoperatively. mediated reduction in insulin sensitivity is protected by concomitant GLP-1. Longstanding DM showed a greater glucose AUC during OGTT before and Thus, short-term combinatorial use of glucagon+GLP-1 is not benefi cial for after surgery (p<0.01) but a comparable surgery-induced improvement. At beta cell function, but might have the potential to reduce risk for insulin- baseline, longstanding DM patients had lower C-peptide (peak and AUC) mediated hypoglycemia when used in non-diabetic subjects. and early insulin response (all p<0.05). After the surgery, these indices Effects of Glucose Alone and Together with Glucagon, GLP1 and their Combination. were comparable between DM subgroups. Changes in insulin sensitivity were independent of diabetes duration. The disposition index remained Glucose only Glucose+Glucagon Glucose+GLP1 Glucose+Glucagon+GLP1 lower in longstanding diabetes (p<0.05) until 1 year after the surgery, the 35mg; n=83 10µmol/kg; n=18 3µmol/kg; n=35 n=10 postoperative increase was comparable between DM subgroups. AUCins (nmol/L min) 11.9±0.4 19.7±2.5*** 13.0±0.6* 10.6±0.3 The progressive improvement of glucose metabolism in DM patients ΔAIRg (nmol/L) 1.0±0.5 3.1±0.5*** 1.5±0.1*** 1.3±0.3* after gastric bypass relies on both improved insulin sensitivity and beta- cell function. Despite of worse baseline status patients with longstanding BCF (µmolINS/molGLUC) 21±1 32±4*** 26±4*** 18±3 diabetes appear to dispose of a comparable recovery potential after gastric SI [10-4min-1/(pmol/L)] 1.05±0.07 0.66±0.07* 1.23±0.09 1.11±0.18 bypass as do the patients with short disease duration. * P<0.05; ** P<0.01; *** P<0.001. Supported By: EFSD/MSD 1750-P & 1748-P The Serum Level of Soluble CD26/DPP-4 Increases in Response Reduced Release of GLP-1 in Subgroups of Prediabetes and Type 2 to Acute Hyperglycemia After an Oral Glucose Load in Healthy Diabetes Subjects: Associations with High Molecular Weight Adiponectin KRISTINE FÆRCH, SIGNE S. TOREKOV, NANNA B. JOHANSEN, DANIEL R. WITTE, and Hepatic Enzymes ANNA E. JONSSON, OLUF PEDERSEN, TORBEN HANSEN, TORSTEN LAUR ITZ EN, TAKANORI TOMOTSUNE, TERUO JOJIMA, KUNIHIRO SUZUKI, TOSHIE IIJIMA, ANNELLI SANDBÆK, JENS J. HOLST, DORTE VISTISEN, MARIT E. JØRGENSEN, KIKUO KASAI, TOSHIHIKO INUKAI, YOSHIMASA ASO, Mibu, Japan, Koshigaya, Gentofte, Denmark, Copenhagen, Denmark, Strassen, Luxembourg, Aarhus, Denmark Japan The incretin effect is reduced in type 2 diabetes, but it still remains A soluble form of CD26/dipeptidyl peptidase-4 (sCD26/DPP-4), which unclear whether the defect is caused by a reduction in the release of incretin participates in degradation of incretin hormones, is found in the serum hormones and if so, whether this reduction occurs prior to the development and it has DPP-4 enzymatic activity. Serum levels of sCD26/DPP-4 also of type 2 diabetes. In the large ADDITION-PRO study, the release of GLP- are associated with insulin resistance. We investigated whether the Integrated POSTERS 1 and GIP was measured at 0, 30 and 120 min after an OGTT (Fig. 1A) in serum level of sCD26/DPP-4 was infl uenced by the oral glucose tolerance 1,448 individuals with complete information on fasting and 2-hour glucose test (OGTT) in healthy subjects. The serum sCD26/DPP-4 level increased Physiology/Obesity concentrations. 47% were women and mean (SD) age was 66 (7) years. signifi cantly from 824.5 (699.0, 1050) ng/ml at baseline to a peak of 985.0 Total area under the curve (AUC) and incremental AUC (iAUC) for GLP-1 and (796.5, 1215) ng/ml during the OGTT (P<0.0001). The peak sCD26/DPP-4 GIP was calculated for people with normal glucose tolerance (NGT, n=776), level was positively correlated with the baseline age, body mass index, isolated impaired fasting glucose (i-IFG, n=280), isolated impaired glucose fasting plasma glucose (FPG), homeostasis model assessment of insulin tolerance (i-IGT, n=109), combined IFG and IGT (IFG+IGT, n=122), and screen- resistance (HOMA-IR), triglycerides (TG), alanine aminotransferase (ALT), detected diabetes by fasting glucose (F-DM, n=81), 2-hour glucose (2h-DM, and γ-glutamyl transpeptidase (GGT), while it was negatively correlated n=43) or both criteria (F-2h-DM, n=37). Before and after adjustment for age, with high-density lipoprotein (HDL) cholesterol and the serum levels of sex and BMI, iAUC for GLP-1 was reduced in individuals with i-IFG, IFG+IGT, total and high molecular weight (HMW) adiponectin. Stepwise regression and F-DM (P<0.05), but not in people with i-IGT, 2h-DM or F-2h-DM (P≥0.19 analysis was done with forward selection of variables, including age, FPG, for all; Fig. 1B). Total AUC for GLP-1 was only decreased in the IFG+IGT HOMA-IR, TG, HDL cholesterol, uric acid, GGT, C-reactive protein, and group (P=0.02). Neither AUC nor iAUC for GIP differed between the groups HMW adiponectin. In a model that explained 57.5% of the variation of the

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A452 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO peak sCD26/DPP-4 level, GGT (β=0.382, P=0.007) and HOMA-IR (β=0.307, fi ndings suggested that vildagliptin enhanced insulin synthesis and basal P=0.034) were independent determinants of the peak serum level of sCD26/ secretion regardless of constitutive autophagy function, but autophagy was DPP-4. Serum HMW adiponectin decreased signifi cantly from 4.43 (2.80, necessary for the augmentation of glucose-induced insulin secretion which 6.65) µg/ml at baseline to 4.17 (2.48, 6.96) µg/ml at 120 min after the oral is well-known function of DPP-4 inhibitors. glucose load (P<0.0001). The baseline serum level of sCD26/DPP-4 showed Supported By: Innovative Research Institute for Cell Therapy (A062260) a signifi cant negative correlation with the % change of HMW adiponectin during the OGTT. In conclusion, the serum level of sCD26/DPP-4 increased 1753-P acutely after an oral glucose load in apparently healthy subjects. The abrupt The β-Cell Response to Acute Insulin Resistance in Normal and increase of serum sCD26/DPP-4 after a glucose load may be associated with Impaired Glucose Tolerance insulin resistance. MEERA SHAH, FRANCESCA PICCININI, CHIARA DALLA MAN, PAULA D. GIESLER, ROBERT A. RIZZA, CLAUDIO COBELLI, ADRIAN VELLA, Rochester, MN, Padova, 1751-P Italy Trajectories of Fasting Plasma Glucose among Healthy Black and In euglycemic individuals, decreases in insulin action (Si) increase insulin White Adults with Parental Type 2 Diabetes: The Pathobiology of secretion to maintain glucose tolerance. Whether this extends to prediabetes Prediabetes in a Biracial Cohort (POP-ABC) Study has been controversial given the suggestion that increased fasting glucose SOTONTE EBENIBO, CHIMAROKE EDEOGA, JIM WAN, SAMUEL DAGOGO-JACK, is associated with signifi cant β-cell loss. We studied 49 individuals with Memphis, TN normal glucose tolerance (NGT) and 43 with impaired glucose tolerance (IGT) The Diabetes Prevention Program showed no racial disparities in on 2 occasions in random order using an oral challenge. On one occasion, free progression from prediabetes to type 2 diabetes (T2D). To determine whether fatty acid (FFA) elevation, was achieved by infusion of intralipid + heparin. disparities occur more proximally, we studied plasma glucose trajectories On the other study day the same amount of glycerol present in the intralipid -4 among initially normoglycemic black (n=217) and white (n=159) offspring of was infused. FFA decreased Si in NGT (14 ± 1 vs. 22 ± 3 10 dl/kg/min per µU/ parents with T2D during for 5.5 yr of follow-up. Serial assessments included ml, p<0.01) and IGT (12 ± 2 vs. 18 ± 3 10-4 dl/kg/min per µU/ml, p=0.02). This OGTT, anthropometry, body fat, insulin sensitivity, beta-cell function, and increased peak (10.5 ± 0.3 vs. 9.9 ± 0.2 mmol/L, p<0.01) and integrated (4.67 ± chemistries. The mean age of the cohort was 44.2 years; BMI was 31.4 kg/ 0.02 vs. 3.20 ± 0.02 Mol per 6 hr, p<0.01) glucose concentrations in NGT. m2. The White vs. Black data at baseline were: FPG 93.2 + 6.51 mg/dl vs. In IGT, peak (11.1 ± 0.6 vs. 10.8 ± 0.2 mmol/L, p=0.09) concentrations were 90.9 + 6.67 mg/dl (P=0.003); 2hrPG 125 + 23.4 mg/dl vs. 123 + 27.5 mg/dl unaffected, while integrated concentrations (6.27 ± 0.04 vs. 4.43 ± 0.03 Mol (P=0.72); BMI 28.8 + 6.80 kg/m2 vs. 31.2 + 7.40 kg/m2 (P=0.0008). 101 (52 per 6 hr, p<0.01) increased. These changes were accompanied by increased black, 49 white) out of 343 subjects (192 black, 151 white) with evaluable peak (402 ± 40 vs. 345 ± 30 pmol/L, p=0.02) and integrated (50.0 ± 5.0 vs. data developed prediabetes, and 10 subjects (6 black, 4 white) developed 30.3 ± 3.5 nmol per 6 hr, p<0.01) insulin in NGT and in IGT (476 ± 46 vs. 379 ± T2D during 810 person-years (mean 2.5 yr) of follow-up. Analyzing fasting 35 pmol/L, p=0.03 and 52.3 ± 6.2 vs. 40.9 ± 4.9 nmol per 6 hr, p=0.01). β-cell plasma glucose (FPG) as a continuous variable, the change during follow- responsivity in NGT (47 ± 3 vs. 58 ± 5 10-9 min-1, p=0.06) and in IGT (46 ± 3 -9 -1 up ranged from -24mg/dl to 83 mg/dl. The FPG remained stable ( +5mg/dl vs. 53 ± 3 10 min , p=0.09) was expressed as a function of Si. The resulting from baseline) in 50.7% of Whites and 50.5% of Blacks, decreased by > 5 Disposition Index (DITotal) was decreased by FFA in both NGT and IGT. Hepatic mg/dl in 18% of Whites and 17.7% of Blacks, and increased by > 5 mg/dl in extraction of insulin decreased in NGT (0.37 vs. 0.53) but not in IGT (0.57 vs. 31.3% of Whites and 27.6% of Blacks (P=NS). The 10th and 90th percentiles 0.63). Suppression of glucagon was unaffected by FFA in NGT (52 ± 3 vs. 48 ± of delta FPG from baseline were -7.0 mg/dl vs. -8.0 mg/dl and +11.5 mg/dl 2 ng/l, p=0.06), but was impaired in IGT (57 ± 3 vs. 52 ± 2 ng/l, p<0.01). We vs. +11.0 mg/dl, in Whites vs. Blacks, respectively. In multiple regression, conclude that the ability of β-cells to adapt to decreased Si does not differ the signifi cant predictors of delta FPG were , HbA1c, total body fat, trunk qualitatively between NGT and IGT, suggesting that adaptation of α-cell fat, HOMA-IR, and HOMA-B. Age, resting energy expenditure, lipid profi le, secretion and hepatic insulin extraction, are important in the evolution of hsCRP and adiponectin were not signifi cant predictors of glycemia. Thus, prediabetes. during a mean follow-up of 2.5 yr, ~ 70% of free adults with parental T2D Supported By: NIH (R01DK078646) experienced either a decrease or minimal change in FPG, whereas ~30% experienced increases of >5-80 mg/dl, without evidence of racial disparities 1754-P in glycemic trajectories. A “Healthy” High-Fat Diet Protects from Insulin Resistance Despite Supported By: ADA (7-07-MN-13); NIH/NIDDK (R01DK067269) Reducing Insulin Secretion JOSIANE L. BROUSSARD, ISAAC ASARE BEDIAKO, REBECCA L. PASZKIEWICZ, 1752-P CATHRYN M. KOLKA, EDWARD W. SZCZEPANIAK, LIDIA S. SZCZEPANIAK, Effects of Long-term Vildagliptin on Insulin Synthesis and Seretion RICHARD N. BERGMAN, Los Angeles, CA in β-Cell-specifi c Autophagy Defi cient Mice High fat diets cause insulin resistance and hyperinsulinemia. However, the EUN ROH, MIN JOO KIM, MIN KYEONG KIM, DONG HWA LEE, CHANG HO AHN, effects of alternative dietary fats on insulin resistance, β-cell function and KYONGYEON JUNG, EU JEONG KU, OK KYONG CHOI, KYUNG SIL CHAE, SEON- fat deposition are not well studied. It is therefore important to examine the YOUNG SHIN, SOO-HEON KWAK, JI WON YOON, BO KYUNG KOO, MIN KYONG effects of so-called “healthy” fats on metabolic parameters as compared MOON, YOUNG MIN CHO, SOO LIM, SUNG SOO CHUNG, DONG-SIK HAM, JI- to “unhealthy” fats with comparable caloric content. We hypothesize that WON KIM, KUN-HO YOON, MASAAKI KOMATSU, KEIJI TANAKA, KYONG SOO lard fat will induce more severe insulin resistance than a diet composed of PARK, HYE SEUNG JUNG, Seoul, Republic of Korea, Tokyo, Japan salmon oil. Pancreatic β cell-specifi c Atg7 knockout (Atg7 Δβ cell) mice showed Dogs were fed a control diet (40% CHO, 28% protein, 32% fat), which was suppression of constitutive autophagy in the β cells resulting in insulin then supplemented with 6g/Kg of lard (n=8) or salmon oil (n=4) to achieve insuffi ciency and hyperglycemia. There have been several evidences 52% fat. An IV glucose tolerance test was conducted at baseline (BL), 2 on autophagy defi ciency in the β cells of type 2 diabetes. Therefore, we weeks (W2) and 6 weeks (W6) of fat feeding to determine insulin sensitivity evaluated the role of adequate autophagy on the long-term effects of a (SI) and acute insulin response to glucose (AIRg). At each time point, a full dipeptidyl peptidase-4 (DPP-4) inhibitor, vildagliptin, using the Atg7 Δβ cell body MRI was conducted to determine fat mass. Integrated mice. At 8 weeks of age, Atg7 Δβ cell mice and wild-type (Rip-Cre+) mice were Fat feeding induced comparable weight gain in both groups. Lard feeding POSTERS divided into vildagliptin and control group. Vildagliptin was administrated was associated with a signifi cant reduction in SI (Fig 1). Interestingly, while through drinking water (0.3mg/mL). After 12 weeks of treatment, body dogs fed lard were able to compensate for insulin resistance by increasing Physiology/Obesity weights and random blood glucose levels in vildagliptin group were not AIRg, dogs on salmon oil showed a paradoxical reduction of AIRg. Thus, different from those in the control group. However, vildagliptin treatment lard induced the expected changes in SI and AIRg. In contrast, salmon oil caused delayed glucose improvement after 60min during intraperitoneal signifi cantly reduced the β-cell response to glucose, suggesting different glucose tolerance test (IPGTT) in the Atg7 Δβ cell mice, while fasting and early mechanisms of insulin stimulation depending on dietary fat composition. improvement before 30min in the wild-type mice. Vildagliptin induced insulin synthesis in the islets of Atg7 Δβ cell mice, where it was suppressed compared to the wild-type mice. Therefore, fasting serum insulin levels signifi cantly increased after vildagliptin treatment. However, high glucose-induced insulin secretion was not augmented by vildagliptin in the Atg7 Δβ cell mice, both in vivo and ex vivo. Therefore, acute response during IPGTT was blunted. These

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A453 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

increased signifi cantly (in each group, p < 0.001). The DI of total patients increased from 0.043± 0.034 to 0.077 ± 0.051 (p < 0.0001). The resolution of glucotoxicity through long-term CSII therapy may contribute to restoration of beta-cell function and disposition capability in type 2 diabetic patients.

Supported By: Swiss Federal Institute of Technology (Zurich)

1755-P Hepatic Insulin Extraction as Measured by a Novel Physiologic Model Is Defective in Prediabetes FRANCESCA PICCININI, CHIARA DALLA MAN, ADRIAN VELLA, CLAUDIO COBELLI, Padova, Italy, Rochester, MN 1757-P Estimating hepatic insulin extraction (HE) during a meal is important for Insulin-Resistant Rats Display Enhanced Rewarding Effects of developing an understanding of the pathogenesis of prediabetes. Previously Nicotine available models provide a non-physiologic, qualitative estimate of HE. We JANELL R. RICHARDSON, JOSEPH A. PIPKIN, LAURA E. O’DELL, ARBI NAZARIAN, therefore set out to develop a physiologically-based model to measure HE Pomona, CA, El Paso, TX in prediabetes. Tobacco use among persons with Type II diabetes exponentially increases To this purpose, we studied subjects with normal or impaired fasting negative health consequences and mortality rates. It is especially troubling glucose (NFG / IFG respectively) and normal, impaired or diabetic levels of that diabetic persons who smoke have a greater diffi culty with tobacco glucose tolerance (NGT, IGT, DM respectively). Accordingly 62 subjects (9 cessation as compared to non-diabetic smokers. Diabetes is a metabolic IFG/DM, 16 IFG/IGT, 7 IFG/NGT, 12 NFG/IGT and 18 NFG/NGT) underwent a syndrome that consists of insulin resistance (IR) due to disruptions in insulin mixed meal with frequent sampling of plasma glucose, insulin and C-peptide signaling. We have previously shown that insulin depletion enhances the concentrations. The model assumes that C-peptide kinetics is described motivational effects of nicotine. The present study expands our previous by a 2 compartment model, insulin kinetics by a 3 compartment model, work by examining whether IR, produced by a high-fat diet (HFD) regimen, and insulin secretion is made up of 2 components, i.e. one proportional enhances the rewarding effects of nicotine, as measured by the conditioned to glucose rate of change and one proportional, with a constant delay, to place preference (CPP) paradigm. Rats were placed on either a regular diet glucose concentrations. HE suppression is assumed to be linearly dependent (RD) or a HFD for 5 weeks, after which they were assessed for IR via blood on plasma glucose concentrations. Basal (HE ) and total (HE ) HE indices b tot glucose measurements after an insulin challenge. The results revealed were obtained from the model; in addition, a new index quantifying HE that HFD produced IR and non-IR animals. Interestingly, the magnitude of sensitivity to glucose (HE ) was derived. G nicotine CPP was larger in IR versus RD rats. Nicotine CPP was absent in The model optimally fi tted C-peptide and insulin data in all subjects. non-IR animals. A similar increase in body weight was observed in IR and Comparison among groups highlighted that HE is higher in NFG than IFG b non-IR rats as compared to RD rats. This suggests that neither the increased subjects (64% vs. 55%, p=0.02), while HE shows the opposite trend (0.12 vs. G body weight nor the HFD per se in the IR rats contributed to the enhanced 0.16 l/mmol, p=0.03); moreover HE and HE are signifi cantly higher in NGT b tot nicotine reward. These fi ndings also suggest that IR rats undergo unique than IFG/IGT (HE =63% vs. 53%, p=0.01; HE =61% vs. 49%, p=0.02). b tot neurobiological changes related to a disruption in insulin signaling that In conclusion, we have developed a new model for the measurement of promotes the rewarding effects of nicotine. HE. We subsequently demonstrate that HE is impaired in prediabetes, likely Supported By: ADA (7-12-BS-135); Western University of Health Sciences due to an impaired ability of plasma glucose to suppress HE. Supported By: DK78646, FIRB2008 1758-P Peroxisome Proliferator-Activated Receptor Gamma (PPARG) Modu- 1756-P lates Free Fatty Acid Receptor 1 (FFAR1) Dependent Insulin Secretion The Disposition Index Is Increased by the Long-term Continuous ROBERT WAGNER, NORBERT STEFAN, SUSANNE ULLRICH, FAUSTO MACHICAO, Subcutaneous Insulin Infusion Therapy in Type 2 Diabetics HANS-ULRICH HÄRING, ANDREAS FRITSCHE, Tübingen, Germany SOO BONG CHOI, KYUNG-JIN KIM, HYUN JU AN, EUN KYONG JEON, YUN H. Free fatty acid receptor 1 (FFAR1, also known as G-protein coupled receptor NOH, Seoul, Republic of Korea, Chungju, Republic of Korea 40, GPR40) is an emerging drug target to amplify glucose-induced insulin To see if the disposition capability can be improved in type 2 diabetic secretion. We recently showed that common genetic variation in FFAR1

Integrated patients through long-term continuous insulin infusion (CSII) therapy, we POSTERS modulates insulin secretion in humans dependent on plasma free fatty acid examined changes in C-peptidogenic Index (CI), Matzda Index (MI) and (FFA) concentrations. We previously demonstrated markedly lower insulin Physiology/Obesity disposition Index (DI) for 2 years. Two hundred twenty one type 2 diabetic secretion in minor allele carriers of the peroxisome proliferator-activated patients (age, 59.3 ± 9.1 years; male 130, female 91; duration, 11.9 ± 7.8 2 receptor gamma gene’s (PPARG) Pro12Ala variant (rs1805192) in the context years; body mass index, 24.4 ±3.2 Kg/m ; HbA1c 9.1 ± 1.9 %) were treated by of elevated FFAs, but the mode of action could not been revealed. According CSII. Blood samplings were performed at base, 6 month,1 year,and 2 years at to a recent study, PPARG activation causes FFAR1 gene upregulation. We the time of fasting and 2 hours after ingestion of 500 kcalories mixed meal. now tested the hypothesis that the effect of PPARG on insulin secretion is After the 2 year, HbA1c decreased from 9.1 ± 1.9 % to 7.2 ± 1.2 % (p < 0.001) mediated by FFAR1 in humans. and serum C-peptide level increased from 4.6 ± 1.9 to 5.7 ± 1.7 ng/ml (p < In a population of individuals with increased risk of diabetes who under- 0.001) and CI increased from 0.011 ± 0.008 to 0.017 ± 0.009 (p < 0.0001) at 2 went oral glucose tolerance tests, 1920 participants were genotyped for 7 hours after meal ingestion. MI of total patients did not change. We divided tagging SNPs in FFAR1 and the Pro12Ala variant in PPARG. patients into high MI group (insulin sensitive group) and low MI group (insulin Using the insulinogenic index as outcome variable for insulin secretion, resistant group) according to median value of initial MI. In insulin sensitive the FFAR1 SNPs rs12462800 and rs10422744 demonstrated interactions group the MI decreased signifi cantly and in insulin resistant group the MI

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A454 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO with PPARG (p=0.01 and 0.005, respectively). In both models, PPARG also Chronically increasing AA supply in the PI-IUGR fetus improves GSIS, interacted with fasting FFA (p<0.05). In subgroup analyses, rs12462800 perhaps due to higher glucagon concentrations. Longer therapeutic use of and rs10422744 were associated with reduced insulin secretion only in AAs to correct defi cient secretion of insulin, a fetal anabolic hormone, may participants concomitantly carrying the PPARG minor allele and having high improve fetal growth and prevent the increased risk for developing diabetes fasting FFA (p=0.0006 and 0.001, respectively, n=232). in cases of placental insuffi ciency. These results suggest that the minor allele of the PPARG Pro12Ala variant Supported By: NIH (R01DK088139, K08HD060688) exposes its carriers to modulatory effects of FFAR1 on insulin secretion. This subphenotype may defi ne altered responsiveness to FFAR1-agonists, and 1761-P should be investigated in further studies. No Acute Cephalic Insulin Response in Humans Supported By: German Federal Ministry of Education and Research SIMON VEEDFALD, ASTRID PLAMBOECK, CAROLYN DEACON, BOLETTE HART- MANN, JENS J. HOLST, FILIP K. KNOP, TINA VILSBØLL, Copenhagen, Denmark 1759-P The notion of cephalic insulin secretion has been revisited by investigators Fasting Glucose and Insulin Secretion but Not Insulin Sensitivity for years. However, data are inconsistent and methods diverse. In the Are Impaired in Non-Diabetic Men with Prostate Cancer Compared present study we aimed to elucidate the presence of an acute effect of vagal to Controls activation on insulin secretion elicited by modifi ed sham feeding (MSF) at a STEFAN Z. LUTZ, KONSTANTINOS KANTARTZIS, JÖRG HENNENLOTTER, ROBERT permissive level of plasma glucose. WAGNER, MARTIN HENI, TILMAN TODENHÖFER, HARALD STAIGER, CHRISTIAN Normoglycemic healthy males (n = 8) were studied on separate occasions SCHWENTNER, ARNULF STENZL, ANDREAS FRITSCHE, HANS-ULRICH HAERING, by hyperglycemic clamp (6 mmol/L) held between t = 15 and 45 min with or Tübingen, Germany without a 15-minute MSF (performed between t = 45 to 60 min). The meal Patients with prostate cancer (PCa) have a poorer prognosis with reduced stimulus consisted of an appetizing breakfast serving including pancakes survival in case of coexistent type 2 diabetes (T2DM), however studies with jam, fried bacon, egg omelet, yoghurt with muesli and syrup, fruit salad, on the role of T2DM for development of PCa are inconsistent. Metabolic a bun with cheese, orange juice and coffee. The participants were instructed mechanisms which may account for the poorer prognosis are yet unclear. to sample and chew all elements but to spit out all including saliva. We hypothesized that patients with PCa will exhibit alterations in insulin Plasma glucose was held for the duration of clamp at 6.0±0.2 and 6.0±0.3 sensitivity and glucose tolerance compared to healthy controls. For this mmol/L (mean±SD) (P = NS) on the clamp and clamp+MSF day, respectively. purpose, we phenotyped 41 non-diabetic patients diagnosed with PCa (aged The MSF procedure elicited an immediate and robust pancreatic polypeptide 62±7 y, BMI 26.5±3.2, mean±SD) and 45 non-diabetic males as controls (aged response demonstrating effective vagal activation. Incremental insulin 61±6 y, BMI 26.7±2.2, mean±SD) from the Tuebingen Family Study, matched levels corrected for plasma glucose stimulation between t=45 to 60 min for age and BMI. Glycemic traits (plasma glucose levels, insulin sensitivity were similar (P = NS). Incremental C-peptide levels during the same period index by Matsuda/deFronzo and secretion indices) were determined by a (not corrected for glucose) were also similar (P = NS). 5 point oral glucose tolerance test. Compared to the control group, PCa In conclusion, we were unable to identify an acute cephalic insulin patients had no differences in postload glucose levels (120 min) but higher response to vagal activation in humans. Despite a robust experimental model, fasting blood glucose levels (105±2 vs. 97±2 mg/dl, p=0.0004). In parallel, employing a permissive level of glucose to prime the beta cells, and to mimic they also had lower insulin secretion as determined by AUC C-peptide 0-30 initial meal-related glucose absorption combined with a mouthwatering min/AUC glucose 0-30 min (p=0.0139) and AUC C-peptide 0-120 min/AUC breakfast serving giving rise to a robust pancreatic polypeptide response, glucose 0-120 min (p=0.0045), adjusted for age, BMI and insulin sensitivity. there was no demonstrable cephalic secretory phase for insulin. There was no difference in insulin sensitivity between PCa patients and controls (p=0.5542). Moreover, serum triglyceride and HDL cholesterol levels 1762-P were lower in PCa patients (p=0.0158 and p=0.0379, respectively), while Relationships among 12-HETE Concentrations, Indices of Insulin NEFA and LDL cholesterol were similar. In summary, PCa patients display Action, and Beta-Cell Function in Prediabetic Adults higher fasting blood glucose levels due to impaired insulin secretion, without COREY A. RYNDERS, JUDY WELTMAN, BOYI JIANG, JERRY L. NADLER, MARC D. changes in insulin sensitivity. Further investigations are needed to clarify BRETON, EUGENE J. BARRETT, ARTHUR WELTMAN, Norfolk, VA, Charlottesville, the molecular links between PCa, impaired insulin secretion and T2DM and VA to develop novel therapeutic approaches. Infl ammation is central to the development of type 2 diabetes. The pro- infl ammatory oxidized lipid 12-hydroxyeicosatetraenoic acid (12-HETE) has 1760-P been linked to reduced insulin secretion and greater insulin resistance. Chronically Increased Amino Acids Improve Glucose-stimulated The present study examined the relationships among urinary 12-HETE Insulin Secretion in Growth Restricted Fetal Sheep concentrations, glucose disposal, insulin secretion, insulin sensitivity, PAUL J. ROZANCE, SANDRA WAI, MELISSA DAVIS, STEPHANIE R. THORN, SEAN and beta-cell function in a sample of prediabetic adults. Plasma glucose, W. LIMESAND, WILLIAM W. HAY, LAURA D. BROWN, Aurora, CO, Tucson, AZ insulin, and C-peptide were measured every 5-10 min during a 180-min oral Placental insuffi ciency produces intrauterine fetal growth restriction (PI- glucose tolerance test (OGTT) administered to 16 prediabetic adults with IUGR) due to reduced amino acid (AA), glucose and oxygen supply to the ages BMI and fasting glucose averaging 49yrs; 32.4 kg/m2; and106 mg/dl, fetus. These defi ciencies produce decreased fetal glucose stimulated insulin respectively. Urinary HETE concentrations (normalized to creatinine) were secretion (GSIS). PI-IUGR fetuses also have increased risk for developing obtained immediately following the OGTT and quantifi ed by HPLC and RIA. type 2 diabetes as adults. Glucose area under the curve (AUC) was calculated using the trapezoid Our hypothesis was that chronically increasing fetal AA supply in experi- rule. Insulin secretion rate (ISR) was derived from deconvolution of plasma mental PI-IUGR fetal sheep will improve GSIS. C-peptide. Insulin sensitivity was estimated using the oral minimal model. Singleton, chronically catheterized PI-IUGR sheep fetuses were given an Beta-cell function was calculated as the ratio between the areas under intravenous infusion of a complete AA mixture (IUGR-AA, n=6) for 10.7±1.3 the ISR curve to that of the glucose curve normalized to the level of insulin days; targeted to increase fetal branched chain AA concentrations (BCAA) by resistance (inverse of insulin sensitivity). Relationships among the variables ~50%. Fetal arterial plasma concentrations of BCAA, insulin, glucose, oxygen, were tested using Pearson correlations. The late phase, but not early phase Integrated glucagon, cortisol, norepinephrine, and IGF-1 were measured. At the end of the of insulin secretion was positively related to urinary 12-HETE concentrations POSTERS infusion on 133.7±0.4 days gestational age (term=148 days) GSIS was measured (early phase ISR 0-60min, r=-0.02, p=0.94; late phase ISR 60-180min, r=0.57, with a square-wave fetal hyperglycemic clamp. Results were compared to saline p=0.02). The present data provide evidence that 12-HETE is involved in Physiology/Obesity infused IUGR (IUGR-SAL, n=6) and control (CON, n=6) fetuses. regulating glucose-stimulated insulin secretion in prediabetic adults. Fetal plasma BCAA concentrations increased in the IUGR-AA group only Supported By: Commonwealth Health Research Board (52%, P<0.005). Insulin, glucose, oxygen, and IGF-1 were lower, and cortisol and norepinephrine higher, in both IUGR groups compared to CON (P<0.05), but were not different from each other. Glucagon increased in the IUGR-AA group only (375%, P<0.01). GSIS was greater in IUGR-AA compared to IUGR- SAL (P<0.0001) and was comparable to CON (hyperglycemic clamp insulin concentrations 1.76±0.19, 0.75±0.17, and 2.02±0.16 ng/ml in IUGR-AA, IUGR- SAL, CON, respectively). Fetal weights in the IUGR groups were not different from each other.

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A455 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

1763-P urinary glucose excretion. Serum C-peptide concentrations are often P11187, a Potent and Novel GPR40 Agonist Potentiates Glucose- used as markers of insulin secretion and beta cell function using standard Stimulated Insulin Secretion, and Improves Glucose Tolerance in assumptions that C-peptide clearance is unaltered by treatments. However, Rodent Models of Type 2 Diabetes because C-peptide is primarily cleared by the kidney and canaglifl ozin acts RAGHURAM ANUPINDI, AMOL DIXIT, NITIN DESHMUKH, PARIKSHIT GAIKWAD, directly on the kidney, it is possible that canaglifl ozin treatment alters MOHAN PATIL, VIVEK KHATANGALEKAR, GANGADHAR HARI, JULIE BOSE, C-peptide kinetics and clearance. Thus, we evaluated whether canaglifl ozin CHIRAG MEHTA, SNEHAL RASAL, SANJAY KUMAR, VISHAL MAHAJAN, RAJIV treatment impacts C-peptide clearance in Sprague-Dawley (SD) rats. SHARMA, VEENA AGARWAL, Mumbai, India Overnight fasted male SD rats (BW = 417 ± 8 g) were treated with GPR40, a G-protein coupled receptor highly expressed in the human canaglifl ozin (10 mg/kg) or vehicle via oral gavage three hours prior to a and rodent pancreatic β-cells, mediates the free fatty acid induced insulin 5 nmol/kg intravenous injection of rat C-peptide-2. Blood glucose levels secretion in a glucose-dependent manner. P11187 is an oral, highly selective were measured at multiple time points before and after treatment. Serum and potent partial agonist of human, rat and mouse GPR40 receptors with an C-peptide levels were measured after injection and fi t to a one-compartment EC50 of 14.4, 2.53 and 4.25 nM, respectively (CHOK1 cell-based assays). In an model to determine clearance and other kinetic parameters. The plasma level intra-peritoneal glucose tolerance test (IPGTT) in Sprague-Dawley (SD) rats of canaglifl ozin was measured at 6 hours after dose. Our results showed that: and C57BL/6J mice, single oral administration of P11187 improved glycemic (1) canaglifl ozin signifi cantly lowered blood glucose levels at all timepoints control (reduction in area under the curve of blood glucose levels - AUCg) compared to vehicle (P<0.05) with its plasma exposure level 1361 ± 326 ng/ with an ED50 of 0.28 and 0.62 mg/kg (p<0.001) respectively and potentiated ml at 6 hr post-dose; (2) following injection, the serum C-peptide profi les insulin secretion. In a type 2 diabetic model of neonatal streptozotocin were similar in both the vehicle and canaglifl ozin groups; (3) C-peptide (nSTZ) rats, administration of P11187 in an IPGTT resulted in a 77% reduction clearance was similar in vehicle and canaglifl ozin groups (29.7 ± 3.1 vs. 27.8 ± in the AUCg (p<0.001, ED50:1.20 mg/kg). Chronic administration of P11187 in 4.5 ml/min/kg in vehicle and canaglifl ozin groups, respectively, p=0.33). diet-induced obese mice (1 to 10 mg/kg, p.o., 3 weeks) caused a signifi cant In conclusion, a single oral dose of canaglifl ozin at 10 mg/kg signifi cantly reduction in the blood glucose levels as demonstrated in an IPGTT (44%, lowers blood glucose but does not affect C-peptide clearance in SD rats p<0.001). In a hyperglycemic clamp study in SD rats, P11187 exhibited and therefore, endogenous serum C-peptide concentrations can be used a signifi cant (p<0.001) increase in the AUC of glucose infusion rate (~1.5- as indices of insulin secretion in the presence of SGLT2 inhibition in this fold) and plasma insulin (~4-fold). P11187 was not associated with any model. hypoglycaemia up to a dose of 100 mg/kg p.o. in normal SD and Zucker fa/ fa rats. P11187 is currently in Phase-I clinical trials in the USA (Clintrials.gov 1766-P identifi er - NCT01874366). The preclinical data suggest that P11187, a novel How Well Does OGTT Detect Hyperinsulinemic Compensation and GPR40 agonist, can be a potential therapy for type 2 diabetes. Decreases in Beta-Cell Function? VIORICA IONUT, DARKO STEFANOVSKI, ANA VALERIA B. CASTRO, ORISON O. 1764-P WOOLCOTT, JOSIANE BROUSSARD, MALINI IYER, RICHARD N. BERGMAN, Los Ethnic Variation in the Secretion of Insulin and Incretin Hormones Angeles, CA Following Intraduodenal Glucose Infusion in Healthy Humans Increased B-cell function (BCF) is a normal, dynamic response to insulin CHINMAY MARATHE, MICHELLE BOUND, SCOTT STANDFIELD, JUDITH resistance. Also BCF is a hallmark of prediabetes and diabetes. To assess WISHART, KAREN L. JONES, MICHAEL HOROWITZ, CHRISTOPHER K. RAYNER, BCF, the oral glucose tolerance test (OGTT) is a well-accepted method. Adelaide, Australia, South Australia, Australia However, the ability of the OGTT to detect increases or decreases in BCF East Asians generally secrete less insulin than Caucasians, for reasons remains unknown. Therefore, we assessed the accuracy of OGTT-derived that are unclear. The incretin hormones, GIP & GLP-1, drive postprandial indices to quantify the fat-feeding compensatory increases in BCF and the insulin secretion in health, and their secretion could vary with ethnicity, beta-cell injury induced decrease (STZ-induced diabetes).BCF assessments which might account for differences in insulin secretion. Gastric emptying from the intravenous glucose tolerance test, and OGTT were performed in determines duodenal glucose delivery, varies widely between individuals (1-4 20 adult canines. In each animal, the acute insulin response from IVGTT and kcal/min), and infl uences postprandial glucose, insulin and incretin secretion; several indices from OGTT were performed in: a) lean, with normal BCF (wk0); therefore inter-ethnic comparisons are best made with a standardized rate b) after fat-fed induced insulin resistance with compensatory increase in BCF of duodenal glucose delivery. (wk10); and c) decreased BCF post STZ treatment (week 15). To assess AIRg We studied 10 Caucasian (C) (age 47 ± 3 yrs; BMI 29.3 ± 1 kg/m2) and as a gold standard, the measurement was directly compared with insulin 11 Han Chinese (HC) (age 25 ± 1 yrs; BMI 25.1 ± 2 kg/m2) healthy men immunostaining of the pancreatic islets.There was a very strong correlation during intraduodenal glucose infusion at 4 kcal/min for 120 min, following (r2=0.99) between AIRg and % insulin staining of the islets, confi rming the an overnight fast. Blood was sampled frequently for glucose, insulin, and use of the AIRg as a gold standard for beta-cell response.After fat feeding total GIP and GLP-1 assays. Data are mean ± SEM. Fasting, peak values, animals gained weight and became insulin resistant (26% decrease in and incremental areas under the curves (iAUC) were compared by unpaired IVGTT-SI). IVGTT AIRg increased by 30%. Of the OGTT derived indices, AUC t-tests. Ins0-120/AUC Gluc0-120 best described hyperinsulinemic compensation, Neither fasting (5.7 ± 0.1 vs. 5.4 ± 0.1 mmol/L), peak (11.7 ± 0.6 vs. 10.1 with r2 =0.56, p<0.01, while the classic insulinogenic index (IGI)( ΔI0-30/ ± 0.5, mmol/L), nor iAUC (238 ± 34 vs. 188 ± 31 mmol/L.min) for glucose ΔG0-30) and the IGI at15 min performed less well (r2=0.47, p=0.01 and differed between C and HC. Fasting insulin (33.5. ± 12.8 vs. 5 ± 0.8, mU/L, p r2=0.45, p=0.02). The OGTT performed better in detecting decreases in BCF. After STZ, AIRg decreased by 70%, change that was captured by the IGI30 = 0.03), peak insulin (820 ± 185 vs. 192 ± 26, p = 0.002) and iAUC for insulin (29870 ± 7129 vs. 6992 ± 1116 mU/L.min, p = 0.004) were all substantially min (r2=0.58, p=0.002) and by the AUC Ins0-120/AUC Gluc0-120 (r2=0.52, greater for C than HC. Fasting GIP was comparable (21.2 ± 2.4 vs. 16.2 ± p<0.01).OGTT-derived indices can be useful as a tool in clinical studies to 1.3 pmol/L), but peak GIP (85.6 ± 8.4 vs. 57.9 ± 3.5 pmol/L, p = 0.005) was detect changes in BCF, though early vs. latter indices appear to be more greater in C than HC, while the iAUC tended also to be greater in C (2880 ± accurate for assessment. However, the OGTT does not provide an excellent 356 vs. 2108 ± 186 pmol/L.min, p = 0.06). Fasting GLP-1 (20.3 ± 2.6 vs. 25 ± index of BCF as compared to the more accurate AIRg. 3.3 pmol/L), peak GLP-1 (63.5 ± 9.7 vs. 56.5 ± 5.2 pmol/L) and iAUC for GLP-1 Integrated POSTERS (1447 ± 292 vs. 1139 ± 185 pmol/L.min) did not differ between the groups. 1767-P In conclusion, both fasting and glucose-stimulated insulin were much The Basal Oscillating Secretory Release of Insulin from β-Cells Is Physiology/Obesity greater in healthy C than HC. This could not be accounted for by differences Not Driven by a Feedback Loop with Either Glucose or Glucagon in GLP-1 secretion, although modest differences in GIP secretion might CASSANDRA M. NICOTRA, WILLIAM J. LEEDS, YANHUA PENG, JENNIFER D. contribute in part. NEWCOMB, ELLEN H. LINDEN, BARBARA C. HANSEN, Tampa, FL Supported By: NHMRC The basal oscillatory pattern of insulin secretion from β-cells in vivo was identifi ed in both humans and nonhuman primates (NHPs), however, 1765-P the characteristics and mechanism of synchrony of these oscillations and Canaglifl ozin and C-Peptide Clearance in SD Rats specifi cally their relation to in vivo glucose and glucagon levels have never FUYONG DU, MICHAEL K. HANSEN, JIANYING LIU, DAVID POLIDORI, KEITH been determined. The insulin molecule in humans and NHPs are identical, DEMAREST, YIN LIANG, Spring House, PA, La Jolla, CA but released in much higher amounts in NHPs, thus providing the ideal Canaglifl ozin is a potent and selective sodium-glucose co-transporter-2 model for examination of the regulation of this basal secretory pattern. (SGLT2) inhibitor for the treatment of type 2 diabetes that acts via increasing The present study examined basal (overnight fasted) insulin levels (mean

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A456 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

71.8 ± 4.9 µU/ml) and basal glucose (mean 69.7 ± 0.63 mg/dl) in 25 rhesus were fed a standard (10 kcal % fat) or a HFD (60 kcal % fat) for 90 days. monkeys (N=114 experiments). Nine monkeys were subsequently overfed Indirect calorimetry, Echo-MRI, insulin sensitivity assessment and gene (OF) for 6 to 14 weeks at various percentages of their own baseline food expression analysis (qRT-PCR, immunoblots) were performed. intake (100-175%) (24 OF experiments). Signifi cant sinusoidal patterns of The Keap1-hypo mice were partially protected from HFD-induced obesity, insulin secretion with a frequency of 8-11 minutes were observed in 80% were more glucose tolerant, had lower glucose levels after starvation, of total baseline experiments. The remaining 20% of experiments had a developed less hepatic steatosis and tended to have higher energy power spectral density max < 3.5%, indicating no detection of a signifi cant expenditure than the WT. Gluconeogenic (G6Pase, PEPCK) and lipogenic periodicity. Mean insulin levels were strongly correlated with half amplitudes (FASN, ACC1) gene expression was repressed in the livers of the Keap1-hypo (r=0.476, p<0.001), with constant relative amplitudes. A more irregular mi c e o n H F D. I n v i t r o glu c o s e p r o d u c t io n f r o m K e a p1- h y p o p r im a r y h e p a t o c y t e s oscillatory pattern was observed for glucose. Cross-correlation revealed was also repressed. This repressed gluconeogenesis and lipogenesis can that the lag between insulin and glucose was nearly zero, with neither be attributed to the increased p-AMPKα (Thr172) levels in the Keap1-hypo insulin nor glucose leading the other, and thus, providing no evidence of the mice. AMPK activation led to increased phosphorylation of TORC2 (Ser171) previously hypothesized glucose feedback loop. Glucagon was found to be and SREBP-1c (Ser372) and resulted in the suppressed expression of their asynchronous with both insulin and glucose oscillations and did not have a gluconeogenic and lipogenic target genes, correspondingly. consistent frequency. In the OF monkeys, mean levels of insulin increased The activation of the Nrf2 pathway appears to protect against the signifi cantly at 140% OF and the regular oscillations of insulin were often metabolic effects of obesity. disrupted at the highest percentages of OF (150-175%), while glucose and Supported By: NIH (R01CA94076); IOF (PIOF-GA-2012-329442 to D.V.C.) glucagon were unaffected. We conclude that neither glucose nor glucagon levels drive the basal oscillatory release of insulin from the pancreas. & 1770-P Supported By: NIH (HHSN2632008000022C) Direct Evidence for Transient Adaptation of Hepatic Mitochondria to Obesity and Steatosis in Humans CHRYSI KOLIAKI, JULIA SZENDROEDI, TOMAS JELENIK, KIRTI KAUL, PETER INTEGRATED PHYSIOLOGY—LIVER NOWOTNY, FRANK JANKOWIAK, MAREN CARSTENSEN, CHRISTIAN HERDER, MARKUS KRAUSCH, WOLFRAM TRUDO KNOEFEL, MATTHIAS SCHLENSAK, Guided Audio Tour: Hepatic Insulin Resistance and Steatosis (Posters: MICHAEL RODEN, Düsseldorf, Germany 1768-P to 1775-P), see page 13. Lower mitochondrial capacity in muscle associates with insulin resistance (IR) and non-alcoholic fatty liver (NAFL). The role of hepatic mitochondrial & 1768-P function in human IR and NAFL remains unclear. Obese humans without diabetes undergoing bariatric surgery (age 40±2 yrs, BMI 48±2 kg/m2) A Small-Molecule Inhibitor of Novel Protein Kinase C Protects were classifi ed into NAFL+ (n=5) and NAFL- (n=10) based on histologically- Against Lipid-induced Hepatic Insulin Resistance determined liver fat. Normoglycemic humans undergoing elective abdominal MAX C. PETERSEN, STEPHEN G. DANN, BLAS A. GUIGNI, MICHAEL J. JURCZAK, surgery (CON; n=9, age 44±3 yrs, BMI 26±1 kg/m2) served as controls. All VARMAN T. SAMUEL, GERALD I. SHULMAN, New Haven, CT, La Jolla, CA participants underwent euglycemic-hyperinsulinemic clamps, intra-operative Non-alcoholic fatty liver disease (NAFLD) is strongly associated with liver biopsies and ex-vivo high-resolution respirometry of liver tissue. Oxidative hepatic insulin resistance and type 2 diabetes (T2D). However, the molecular stress was assessed from thiobarbituric acid reactive substances (TBARS) and mechanisms linking hepatic lipid metabolism and hepatic insulin resistance mitochondrial content by citrate synthase activity (CSA) and mitochondrial DNA are poorly understood. To examine this question, we studied the effect of a (mtDNA). Both NAFL+ and NAFL- featured lower whole-body insulin sensitivity small-molecule inhibitor (PF-555) of novel protein kinase C (nPKC) on lipid- than CON (M-value; 3.0±0.3 vs. 3.9±0.5 vs. CON 7.0±0.9 mg.kg-1min-1, p<0.01). induced hepatic insulin resistance in awake mice. Two days of a high fat diet NAFL- presented higher maximal respiration related to glycolysis and Krebs (HFD) in vehicle-treated control (CON) mice was suffi cient to induce hepatic cycle activities than CON (3.6±0.4 vs. 1.9±0.1 pmol/s/mg/CSA, p 0.001), while insulin resistance as assessed by a hyperinsulinemic-euglycemic clamp. This ≤ NAFL+ displayed 50% lower respiration linked to -oxidation (1.9±0.2 vs. was associated with a >70% increase in hepatic triacylglycerol (TAG) (P<0.05) β 4.1±0.6 pmol/s/mg/CSA, p<0.01) despite similar mtDNA. The higher oxidative and diacylglycerol (DAG) (P<0.01) content. PF-555-treated (50 mg/[kg-day]) capacity in NAFL-related to hepatic overexpression of -oxidation gene long- mice were protected from lipid-induced hepatic insulin resistance as refl ected β chain acyl-CoA dehydrogenase, whereas the lower respiration of NAFL+ by greater suppression of endogenous glucose production (EGP) during the related to higher serum interleukin-6 (p=0.009), higher hepatic TBARS (303±87 hyperinsulinemic-euglycemic clamp (PF-555: 71±16% vs. CON: 18±15%, P<0.05) vs. 152±40 µmol/mg protein, p=0.04) and lower peroxisome proliferator- despite similar food intake, body weight, and hepatic TAG/DAG content. activated receptor gamma co-activator 1a mRNA (p=0.03). In conclusion, Consistent with the increased suppression of EGP during the hyperinsulinemic- hepatic mitochondrial capacity is upregulated in obese persons without NAFL. euglycemic clamp, insulin-stimulated Akt Ser473 phosphorylation was also The lower mitochondrial capacity in obese NAFL+ is likely linked to hepatic increased ~2-fold in PF-555-treated HFD-fed mice (P<0.05). oxidative stress and systemic infl ammation, providing the fi rst direct evidence Conclusion: These fi ndings support a key role for DAG-induced nPKC activation for adaptation of liver mitochondria to changing bioenergetic demands in in the pathogenesis of lipid-induced hepatic insulin resistance and suggest that obesity and steatosis. nPKC inhibition may be a potential therapeutic strategy for the treatment of Supported By: EFSD/Lilly European Diabetes Research Fund; German Center for NAFLD-associated hepatic insulin resistance in patients with T2D. Diabetes Research; Helmholtz Alliance ICEMED Supported By: NIH (DK-40936) & & 1769-P 1771-P Targeted Disruption of the ApolipoproteinJ Selectively in Liver Keap1/Nrf2 Pathway Activation Represses Hepatic Gluconeogene- Causes Insulin Resistance and Glucose Intolerance sis and Lipogenesis JI A. SEO, INES S. LIMA, MICHELLE CHUNG, MIN JAE KIM, YOUNG-BUM KIM, DIONYSIOS V. CHARTOUMPEKIS, STEPHEN L. SLOCUM, JOHN J. SKOKO, NO- Boston, MA BUNAO WAKABAYASHI, SUSAN AJA, MASAYUKI YAMAMOTO, THOMAS W. ApolipoproteinJ (ApoJ, also called clusterin) is a disulfi de-linked hetero-

KENSLER, Pittsburgh, PA, Baltimore, MD, Sendai, Japan Integrated dimeric protein that is widely distributed in the various tissues and body POSTERS Nrf2 is a transcription factor that regulates the adaptive response to fl uids. Although ApoJ has not been suspected to be involved in energy electrophilic and oxidative stresses. Keap1 is a cytoplasmic protein that Physiology/Obesity metabolism, recent work demonstrates that hypothalamic ApoJ regulates binds Nrf2 and facilitates its ubiquitination and subsequent proteasomal food intake and body weight homeostasis. degradation. Extrinsic or intrinsic stimuli can modify reactive cysteines in However, the knowledge of hepatic ApoJ action in the context of Keap1 and render it incapable of facilitating Nrf2 degradation. Hence, Nrf2 glucose metabolism remains incomplete.To determine the physiological accumulates, translocates to the nucleus and induces the transcription of role of hepatic ApoJ in the regulation of glucose homeostasis and insulin cytoprotective and antioxidant genes. sensitivity, mice lacking ApoJ in liver were studied. Immunoblotting analysis As oxidative stress increases in livers of obese subjects and Nrf2 regulates indicated that ApoJ protein in serum was detected ~75 KDa in control mice. the expression of antioxidant genes and as well as cross-talks with metabolic Surprisingly, no serum ApoJ was found in liver-specifi c ApoJ-defi cient mice, pathways, we hypothesized that Nrf2 activation can ameliorate the diabetic suggesting that liver is a major sourcing organ of circulating ApoJ. At ~27 and obesity phenotype resulting from a high-fat diet (HFD). weeks of age, blood glucose and serum insulin levels in the fasted state To this end, 8 week old C57BL6J wild-type (WT) male mice or mice with were signifi cantly increased in liver-specifi c ApoJ-defi cient mice compared hypomorphic Keap1 alleles (a model of gain of Nrf2 function-“Keap1-hypo”)

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A457 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

to control mice. However, triglyceride and cholesterol levels were unaltered 1774-P by loss of hepatic ApoJ. Mice lacking hepatic ApoJ displayed insulin resistance, as evidenced by the failure of blood glucose levels to decrease WITHDRAWN after insulin injection. Moreover, glucose tolerance was markedly impaired in the absence of hepatic ApoJ. These effects were independent of changes in adiposity. Defi ciency of hepatic ApoJ had no effect on oxygen consumption and carbon dioxide production in mice. These data suggest that hepatic ApoJ could play an important role in regulating whole glucose metabolism and further implicate that targeting hepatic ApoJ could be a therapeutic option for the treatment of metabolic disorders. Supported By: ADA (7-12-BS-094)

& 1772-P Eosinophils Maintain Hepatic Insulin Sensitivity ELIZABETH A. JACOBSEN, JAMES J. LEE, KATIE R. ZELLNER, KARL KAIYALA, NANCY A. LEE, ELENA A. DE FILIPPIS, LAWRENCE J. MANDARINO, Scottsdale, AZ, Seattle, WA Activities by resident eosinophils in adipose tissue and other metabolically important sites have been hypothesized to contribute to the maintenance of systemic glucose homeostasis and insulin sensitivity. This hypothesis was tested by placing 12 week old wild type (WT), ΔdblGATA eosinophil-defi cient (GATA) and hyper-eosinophilic (NJ1638) mice on standard (STD) and 60% high fat (HFD) diets for 6 weeks. All mice were on a C57BL/6J background. Following these 6 week diets, animals were placed in metabolic cages to measure energy expenditure, and dual-catheterized, unrestrained conscious mice had euglycemic clamps with 3H-glucose infusion to measure glucose turnover. Mice on HFD gained weight similarly across genotypes, although 1638 gained less fat-mass than WT or GATA (1.8±0.2 vs. 3.8±0.6 or 3.7±0.6g, respectively, P<0.05) and 1638 had greater body mass adjusted energy expenditure on HFD (9.3±0.1 vs. 8.6±0.1 or 8.2±0.1cal/min, P< 0.01). On STD GATA mice required lower rates of glucose infusion during hyperinsulinemia to maintain euglycemia in comparison to WT or 1638 mice (39±6 vs. 58±4 or 57±4µg/(g·min), respectively, & 1775-P P<0.05). However, on STD or HFD insulin stimulated glucose disposal did not Early Activation of Hepatic Glucose Output in Hypoglycemic Fetuses differ among genotypes, and the presence of excess eosinophils in 1638 did Is Supported by Changes in Amino Acid Metabolism not protect against HFD-induced insulin resistance for glucose disposal in SATYA HOUIN, PAUL J. ROZANCE, LAURA D. BROWN, WILLIAM W. HAY, peripheral tissues. The decreased glucose infusion after STD in GATA mice RANDALL B. WILKENING, STEPHANIE R. THORN, Aurora, CO was due to lower suppression of endogenous glucose production by insulin Hepatic glucose output (HGO) is normally absent in the fetus due to compared to WT or 1638 mice (53±16 vs. 94±6 or 76±9% suppressed from trans-placental maternal glucose supply. We have found an early activation basal, respectively, P<0.05). We conclude that (1) Hypereosinophilia results of HGO in sheep fetuses with hypoglycemia alone and during placental in lower fat gain during HFD, possibly by increasing energy expenditure, and insuffi ciency. This early activation of HGO may predispose offspring to (2) Eosinophils appear to be necessary for the maintenance of hepatic insulin diabetes. We hypothesized that changes in fetal liver amino acid (AA) sensitivity and glucose homeostasis due to either previously reported indirect metabolism support HGO. To test this, studies were done prior to (basal) and systemic effects maintaining healthy adipose tissue or unknown direct effects after fetal hypoglycemia (d1, d5) produced by maternal hypoglycemia with of eosinophils within the liver. insulin infusion. Fetal sheep (n=6) had catheters placed in the umbilical vein, fetal artery, and left hepatic vein to measure net fetal and liver specifi c blood & 1773-P fl ow and substrate fl uxes. Concentrations and uptakes of individual AA were Defi ciency of PDK2 Ameliorates Hepatic Steatosis and Insulin measured and their sum represents total. Fetal glucose concentration and Resistance in Mice Fed a High-Fat Diet fetal glucose uptake from placenta decreased 50% (d1, d5; P<0.01). The fetal AH REUM KHANG, JUN-HWA HONG, BO-YOON PARK, YOUNGHOON GO, SEUNG liver transitioned from net glucose uptake (basal, -13.1 +/- 3.3 umol/min/kg) HEE CHOI, CHAE-MYEONG HA, KEUN-GYU PARK, JUNG-GUK KIM, IN-KYU LEE, to net output by d5 (13.3 +/- 7.2 umol/min/kg; P<0.05). Under basal conditions, SUNG-RAE CHO, NAM HO JEOUNG, ROBERT A. HARRIS, Daegu, Republic of the fetal liver had net uptake of all AA except Ser and Glu. On d1, hepatic Korea, Changwon, Republic of Korea, Indianapolis, IN total AA uptake increased 3-fold (P<0.05) and was 2-fold higher on d5. This Hepatic steatosis is rapidly evolving as a major medical problem in the was driven by increased Thr, Gly, Ala, and Lys uptake and decreased Glu world. It involves a variety of lipid abnormalities including enhanced fatty output. Fetal total AA uptake from placenta was modestly increased (P=0.1). acid infl ux from the adipose tissue, increased de novo lipogenesis, reduced Interestingly, fetal total AA concentrations and branched chain AA (BCAA: fatty acid oxidation and ketogenesis. The pyruvate dehydrogenase kinases Leu, Ile, Val) increased by >20% on d1 and d5. BCAA were positively related (PDKs) regulate pyruvate oxidation by controlling the activity of the pyruvate with HGO (R2>0.4; P<0.05). Thus the source of AA for hepatic uptake may dehydrogenase complex (PDC). We examined whether PDKs were increased come from the placenta or release by fetal tissues like muscle. In addition, in the liver of HFD-fed mice and how they regulate hepatic steatosis and treating fetal sheep hepatocytes with a mixture of AA increased glucose insulin resistance. The physiological importance of regulation of PDC activity production 3-fold (P=0.09) and PCK1 mRNA 10-fold (P<0.01), supporting AA by PDK isoenzyme 2 was assessed by comparing PDK2 knockout (PDK2 KO) mediated HGO. In summary, changes in hepatic AA metabolism, including Integrated POSTERS mice with wild type mice fed a high fat diet (HFD) and an isocaloric low fat increased AA uptake and a re-direction of carbon from Glu to glucose output, diet (LFD). Body weight gain and hepatic steatosis were attenuated by PDK2 support an early activation of HGO in the hypoglycemic fetus. Physiology/Obesity defi ciency in the HFD fed mice. Fasting blood glucose, serum insulin, and liver Supported By: NIH (K01DK090199) pyruvate, lactate, oxaloacetate, citrate, diacylglycerols, and triacylglycerols were also reduced. Hepatic glucose production was also reduced and insulin 1776-P sensitivity was increased in the HFD fed PDK2 KO mice. The hepatic enzyme Deletion of ChREBP Gene in Mice Did Not Improve Glucose Intoler- capacity for fatty acid oxidation and ketogenesis was increased while the ance in Insulin-Defi cient State capacity for lipogenesis was decreased. In spite of this but consistent with KATSUMI IIZUKA, WUDELEHU WU, HIROYUKI NIWA, HIROMI TSUCHIDA, KOU greater PDC activity, the respiratory exchange ratio was higher in the PDK2 DOKUKO, YUKIO HORIKAWA, JUN TAKEDA, Gifu, Japan knockout mice. Energy expenditure was increased without changes in physical Carbohydrate response element binding protein (ChREBP) plays an activity. Increased hepatic insulin sensitivity and improved glucose tolerance important role in glucose and lipid metabolism by regulating de novo lipo- correlated with reduced PKCε phosphorylation. The fi ndings support the case genesis in the liver. It was previously reported that deletion of ChREBP for PDK2 as a promising target for hepatic steatosis and insulin resistance. in ob/ob mice improves obesity, glucose tolerance, and fatty liver. In this

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A458 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER study, we clarify the role of ChREBP in the glucose and lipid metabolism We found that IHF signifi cantly reduced with three treatments (exenatide of insulin defi cient streptozocin (STZ) injected mice. STZ (200mg/BW × 5 Δ=-68%, insulin Δ=-58%, pioglitazone Δ=-49%). Exenatide also reduced VF times) or vehicle-only control (Vehicle) were administrated intraperotoneally (Δ=-36%) and SF (Δ=-13%), pioglitazone decreased VF (Δ=-30%) with no to 8-week-old wild type (WT) and ChREBP homozygous knockout (KO) mice. impact on SF, whereas insulin had no impact on VF or SF. After interventions, After 4 weeks of STZ injection, all experiments were performed. Both WT- HbA1c declined in all groups. Body weight and serum triglycerides decreased STZ and KO-STZ showed similar glucose intolerance. Fed plasma insulin with exenatide. Levels of TNFα (exenatide/insulin/pioglitazone) decreased, levels of WT-STZ and KO-STZ were much lower than those in WT-Vehicle and adiponectin (exenatide/pioglitazone) and irisin (exenatide/insulin) and KO-Vehicle, and as low as fasted plasma insulin levels. Fasted plasma increased. Analysis showed that ΔIHF correlated with ΔHbA1c and Δweight. free fatty acid and triglyceride levels in WT-STZ and KO-STZ mice were ΔIHF correlated with Δtriglycerides and ΔTNFα, but the correlations fell lower than those in WT-Vehicle and KO-Vehicle. Hepatic triglyceride content short of signifi cance after BMI adjustment, and no correlation was shown in KO-STZ was similar to that of WT-STZ, but much lower than that of WT- between ΔIHF and Δadiponectin or Δirisin. By linear regression analysis, Vehicle and KO-Vehicle. Hepatic glycogen content in KO-STZ was similar to ΔHbA1c alone explained 40.5% of the variance of ΔIHF and ΔHbA1c+Δweight that of WT-STZ and WT-Vehicle, but lower than that of KO-Vehicle. Finally, explained 53.4% of the variance. mRNA levels of liver type pyruvate kinase (pklr) and fatty acid synthase In conclusion, liver fat content can be signifi cantly reduced irrespective (fasn), ChREBP target genes, were decreased in livers from WT-STZ mice as of using exenatide, insulin, and pioglitazone. Metabolic control plays an compared with those in WT-Vehicle mice. In livers of both KO-Vehicle and important role in slowing progression of fatty liver in T2DM. KO-STZ mice, glucose induction of pklr and fasn mRNA was diminished. In conclusion, these fi ndings suggest that insulin is indispensable for ChREBP 1779-P action in vivo. DEPTOR, an Inhibitory Protein of mTOR, Contributes to Both Anti- cancer and Antidiabetic Effects of Metformin in Liver 1777-P AKIO OBARA, YOSHIHITO FUJITA, ABULIZI ABUDUKADIER, TORU FUKUSHIMA, Association between GCKR and Glucose Effectiveness (SG) Is YASUO OGURI, MASAYA HOSOKAWA, NOBUYA INAGAKI, Kyoto, Japan Modifi ed by Dietary Fructose in Mexican Americans (MA) Metformin is one of the most commonly used antidiabetic drugs for type 2 RICHARD M. WATANABE, HOOMAN ALLAYEE, ENRIQUE TRIGO, ANNY H. XIANG, diabetes. Metformin has recently received much attention regarding its anti- THOMAS A. BUCHANAN, Los Angeles, CA, Pasadena, CA cancer effect in various types of cancers. Although the main target tissue GCKR is a diabetes risk locus associated with decreased glucose and of metformin’s anti-diabetic action is liver, the drug also reduces the risk of increased triglycerides (TRIG). In vitro studies suggest GCKR variation liver cancer, but the mechanism of its suppression of cell proliferation in liver alters GCKR regulation of glucokinase activity. There is no in vivo evidence remains unknown. To elucidate the anti-cancer effect of metformin in liver supporting this mechanism. We hypothesized the GCKR effect can be c anc er c ells, we ex amine d t he ef fe c t of mTOR signaling on t he c ell pr olifer ation refl ected in SG, since hepatic glucose uptake (HGU) is a large fraction of rate in HepG2 cells. Metformin activated AMPK, suppressed phospho-p70 SG at fasting. Fructose-1-phosphate (F1P) regulates GCKR activity, thus S6 kinase, a downstream target of mTOR, and suppressed cell proliferation we hypothesized dietary fructose alters hepatic F1P levels that interact in HepG2 cells. To clarify the mechanism of the suppressing effect of mTOR with variants to modify GCKR activity and affect SG. We tested these signaling by metformin, we evaluated the expression levels of DEPTOR, an hypotheses in up to 1785 non-diabetic individuals (61.3% female; age: 37±10 endogenous substrate of mTOR suppression degraded by the proteasome years, BMI: 29.4±5.6 kg/m2; mean±SD) from MA families of probands with system. Metformin attenuated proteasome activity, and the protein levels or without a previous diagnosis of GDM. The association between GCKR of DEPTOR were increased in the presence of metformin. The suppressing rs780094 and diabetes-related traits was tested by variance components effect of metformin on cell proliferation and the attenuating effect on (SOLAR V4.3.1) adjusting for age, sex, percentage body fat, and alcohol proteasome activity disappeared by AMPK silencing. The suppressing effect intake. Interaction between rs780094 and dietary fructose (Willet’s food of metformin on cell proliferation also disappeared by DEPTOR silencing. frequency questionnaire) was tested by a joint 2-df test; SNP main effect These fi ndings strongly suggest that regulation of the protein level of and SNP×dietary fructose interaction. rs780094 A allele was associated DEPTOR by the proteasome degradation system has an important role in -5 with increased SG (p=0.0041), increased TRIG (p=7.2×10 ), increased the tumor-suppressing effect of metformin in human liver cancer cells. In cholesterol (CHOL; p=6.9×10-4), and showed a trend for association with addition, we investigated whether the protein levels of DEPTOR are involved decreased fasting glucose (p=0.088). The GCKR A allele and its interaction in the anti-diabetic effect of metformin in liver. Metformin also attenuated -4 -4 with dietary fructose was associated with SG (p=4.9×10 ), TRIG (p=2.7×10 ), the proteasome activity in hepatocytes isolated from BL/6 mice, and the CHOL (p=0.0023), and marginally associated with aspartate transaminase suppressing effect of metformin on gluconeogenesis, a main target of its (p=0.05). SG increased with each copy of the A allele in the lowest dietary action, disappeared by DEPTOR silencing. We conclude that the DEPTOR fructose tertile (19.0±7.9 g), did not signifi cantly change in the middle dietary level, which is regulated by proteasome system, has a critical role in the fructose tertile (32.7±7.8 g), and decreased with each copy of the A allele anti-cancer and the anti-diabetic effects of metformin in liver. in the highest dietary fructose tertile (52.3±16.5 g). We provide the fi rst in vivo evidence suggesting variation in GCKR alters HGU, in turn altering 1780-P fasting glucose and TRIG and that these effects can be modifi ed by dietary Elevated Endogenous Asymmetric Dimethylarginine Contributed to fructose. Insulin Resistance Through the Induction of Endoplasmic Reticulum Supported By: ADA (1-05-RA-140); NIH (DK-061628) Stress in the Liver of High-Fat-Fed-induced Type 2 Diabetic Rats YAN XIONG, YI-PING LENG, QIN-FENG QIN, NI QIU, WEI-JIN FANG, ZHI-MIN HE, 1778-P Guangzhou, China Effects of Exenatide, Insulin, and Pioglitazone on Liver Fat Content Increasing evidence suggested that endoplasmic reticulum (ER) stress and Body Fat Distributions in Newly Diagnosed Subjects with Type contributes to insulin resistance (IR), a leading factor in the development 2 Diabetes of type 2 diabetes mellitus (T2DM). Accumulation of asymmetric dimethyl- YAN BI, HUIJIE YANG, DALONG ZHU, JIANPING WENG, Nanjing, China, Guang- arginine (ADMA), an endogenous nitric oxide synthase (NOS) inhibitor, is zhou, China closely associated with T2DM and diabetic cardiovascular complications, Integrated Previous studies showed that anti-diabetic agents of exenatide, insulin, although the mechanisms have not been elucidated. This study was to POSTERS or pioglitazone could improve glucose control and concomitantly decline determine whether elevated endogenous ADMA are involved in ER stress, intrahepatic fat (IHF). However, most of those reports were placebo promoting the development of insulin resistance in the liver of high-fat- Physiology/Obesity controlled studies; still unclear is whether these benefi cial effects are due fed rats. Expressions of immunoglobulin binding protein (Bip) and C/EBP to either the agent itself or the effects of simply eliminating glucotoxicity homologues protein (CHOP) genes as well as the splicing of X box-binding by achieving glycaemic control and whether these effects differ between protein-1 (XBP-1) mRNA were measured to refl ect the ER stress. Oral agents. glucose tolerance test, insulin sensitivity index, uptaking of [3H]-2-deoxy- Here, thirty-two newly-diagnosed T2DM patients (age 52.7±1.7 years, glucose, gene expression and protein phosphorylation of insulin signaling HbA1c 8.7±0.2%, BMI 24.5±0.5 kg/m2) were randomized to exenatide, molecules were detected to evaluate insulin resistance. The results showed insulin humalog Mix25, or pioglitazone for 6 months. IHF, visceral fat (VF), that hepatic ER stress was provoked in type 2 diabetic rats and coupled and subcutaneous fat (SF) were measured using proton nuclear magnetic with IR in parallel with the elevation of endogenous ADMA concentration, resonance spectroscopy. Plasma TNFα, adiponectin, and irisin were assayed suppression of nitric oxide (NO) generation and NOS activities. Exposure by ELISA. of rat hepatocytes to exogenous ADMA or high glucose also induced ER

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A459 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

stress and IR, which was associated with the inhibition of NO production 1783-P and increase of oxidative stress. Treatment of hepatocytes with antioxidant Activation of Hepatic ROCK1 Promotes Insulin Resistance and pyrrolidine dithiocarbamate not only reduced oxidative stress but also Hepatosteatosis in Diet-induced Obese Mice decreased ADMA-induced ER stress and IR. These results indicated that INES S. LIMA, SEUNG-HWAN LEE, MICHELLE CHUNG, MIN JAE KIM, MARIA elevated endogenous ADMA contribute to hepatic IR through the induction P. MACEDO, YOUNG-BUM KIM, Boston, MA, Seoul, Republic of Korea, Lisbon, of ER stress, and the mechanism underlying ADMA-induced ER stress may Portugal relate to oxidative stress via NO synthase uncoupling. Obesity is a risk factor for non-alcoholic fatty liver disease and is strongly Supported By: Natural Science Research Foundation of China (81170778) associated with insulin resistance. Our previous data demonstrated that liver-specifi c deletion of Rho-kinase 1 (ROCK1) resulted in hypersensitivity 1781-P to insulin and protected against hepatic steatosis in diet-induced obese Protective Effects of Rotenone on High Fat/High Fructose Diet- mice (Diabetes 2012; 61;A464). To further defi ne the physiological role of induced NAFLD in Obese Mice hepatic ROCK1 in regulating glucose and lipid homeostasis, mice expressing JA YOUNG JEON, SEUNG JIN HAN, JONG-GAB JEONG, SUNG-E CHOI, EUN SUK a constitutively active (CA) mutant of ROCK1 in liver were studied. Under HA, TAE HO KIM, SO-YEON AHN, HAE JIN KIM, YUP KANG, DAE JUNG KIM, a high-fat diet, liver-specifi c CA-ROCK1 mutant mice exhibited insulin KWAN WOO LEE, Suwon, Republic of Korea, Incheon, Republic of Korea resistance, as revealed by the failure of blood glucose levels to decrease after Rotenone and metformin are well known compounds that inhibit in insulin injection. However, glucose tolerance was normal. These effects mitochondrial complex I. However the effects and precise mechanism were accompanied by rises in adiposity, serum insulin and glucose levels. of Rotenone are not fully elucidated in fatty liver disease, obesity, and Histological analysis indicated that hepatic steatosis by high-fat feeding diabetes. In this study, we examined the benefi cial effects of Rotenone on was greatly increased in liver-specifi c CA-ROCK1 mutant mice compared high fat/high fructose (HF/HF) diet-induced liver steatosis and malregulation with control mice. Consistent with this, hepatic triglyceride content in of glucose-insulin homeostasis. In addition, we investigated whether liver-specifi c CA-ROCK1 mutant mice was also increased by ~40%, along Rotenone reduces high fat/high fructose (HF/HF) diet-induced ER-stress with an elevation in serum triglyceride and cholesterol levels. Moreover, and infl ammation in fatty liver disease. Seven week old male mice were activation of ROCK1 in mice in liver caused an increase in gene expressions randomly divided into three groups (n=10 in each qroup): normal chow diet of key lipogenic enzymes, including FAS and ACC. However, overexpression (placebo), high fat/high fructose (i.e., 60% fat, 30% fructose) diet (placebo), of hepatic CA-ROCK1 had no effect on gene expression involved in fatty and high-fat/high fructose diet with Rotenone treatment for 10 weeks. acid oxidation and fatty acid uptake. These data demonstrate that selective Rotenone (1mg/kg, n=10) and placebo (n=20) were treated every other day. activation of hepatic ROCK1 is suffi cient to promote insulin resistance and After treatment, intraperitoneal glucose tolerance tests, insulin tolerance hepatic steatosis in diet-induced obese mice. Thus, our studies identify tests, and immunohistological examinations were performed. The normal hepatic ROCK1 as a key regulator of fuel metabolism and further suggest chow diet group test results were as expected. The HF/HF group developed that targeting hepatic ROCK1 could be a novel therapeutic option for the obesity, hyperglycemia, and insulin resistance, whereas the Rotenone treatment of obesity-linked metabolic disorders. treated group strongly recovered obesity-related hyperglycemia, insulin Supported By: ADA (1-09-RA-87) resistance, and insulin signaling. It also reduced hepatic triglyceride content and ameliorated hepatic steatosis. In addition, Rotenone treated mice had 1784-P decreased HF/HF diet-induced infl ammatory and ER-stress related signaling Hepatic Acetyl-CoA Carboxylase Expression and Triglyceride in the liver. Thus these results demonstrate Rotenone has anti-diabetic and Content Are Increased in Mice after Ovariectomy anti-fatty liver effects in HF/HF diet-fed obese mice. GREGORY C. HENDERSON, SARA C. CAMPBELL, MARC A. TUAZON, New Bruns- wick, NJ 1782-P Accumulation of triglyceride (TG) in the liver is associated with insulin CREB3L3 Governs Hepatic Metabolic Genes and Energy Responses resistance, and both liver TG accumulation and insulin resistance increase in in Starvation prevalence after menopause. Therefore, we sought to determine molecular YOSHIMI NAKAGAWA, AOI SATOH, TAKASHI MATSUZAKA, NAOYA YAHAGI, mechanisms of a link between ovarian hormone defi ciency and dysregulated NOBUHIRO YAMADA, HITOSHI SHIMANO, Tsukuba, Japan metabolism. We performed ovariectomy (OVX) or Sham surgery on female Transcriptional regulation of metabolic genes in the liver is the key to C57BL/6J mice (6 per group) at 14-16 weeks of age. Body composition maintaining systemic energy homeostasis. In an energy-depleted state, assessments by magnetic resonance were performed 6 weeks after surgery hepatic transcription factors and co-activators, such as peroxisome followed by blood and liver tissue collection. Blood glucose was measured by proliferator-activated receptor α (PPARα), hepatic nuclear factor 4α a glucometer, hepatic TG was assessed biochemically, and western blotting (HNF4α), forkhead box O1 (Foxo1), peroxisome proliferative activated was performed on liver tissue. Blood glucose concentration was signifi cantly receptor, gamma, coactivator 1α (PGC-1α) and CREB regulated transcription elevated in OVX mice vs. Sham mice (P<0.05). Body weight, gonadal fat mass, coactivator 2 (CRTC2) regulate lipid and glucose metabolism correspondingly. and percent body fat were each signifi cantly elevated in OVX vs. Sham mice Here we demonstrate that a membrane-bound transcription factor, cAMP (P<0.05). Additionally, hepatic TG content was signifi cantly higher in OVX responsive element binding protein 3-like 3 (CREB3L3), is activated during than Sham mice (P<0.05). Next, in order to identify a molecular mechanism fasting, which mediates a wide spectrum of metabolic responses to for accelerated hepatic TG accumulation, we tested hepatic acetyl-CoA starvation. Creb3l3 promoter had binding sites for and activated by PPARα, carboxylase (ACC) protein expression and phosphorylation. Expression of CREB3L3 itself, and HNF4α. total ACC1 and ACC2 were signifi cantly higher in OVX than Sham mice (P < Inversely, Ppara promoter is activated by CREB3L3. CREB3L3 mRNA is 0.05). The ratio between phosphorylated to total ACC were unchanged for downregulated in Ppara KO mice, whereas PPARα mRNA is downregulated both isoforms. Thus, the content of active (unphosphorylated) ACC would in CREB3L3 KO mice. Nuclear CREB3L3 mutually amplifi es PPARα activity in have increased in proportion to the total ACC content. We conclude that the a positive feedback fashion and induces other established fasting upstream increases in hepatic TG and the related metabolic dysregulation may be a factors, leading to the induction of systemic lipolysis, energy expenditure, result of altered ACC expression following withdrawal of ovarian hormones. hepatic ketogenesis and insulin sensitivity. The results indicate that ovarian hormone defi ciency in females may lead to Integrated POSTERS CREB3L3 Consequently, adenoviral and transgenic overexpression of altered regulation of hepatic lipid metabolism, and thus a propensity toward CREB3L3 reduces plasma lipid and glucose levels and body weight in hepatic TG accumulation and ensuing insulin resistance. The results may have Physiology/Obesity both normal and diabetic obese mice. CREB3L3 activates insulin-induced implications for the increased risk for nonalcoholic fatty liver disease after phosphorylation of IRS-2 and Akt in liver. Hepatic CREB3L3 induction menopause in women and the associated increase in risk for type 2 diabetes. activates the thermogenesis related genes in brown adipose tissue and O2 consumption in whole body. CREB3L3 directly transactivates fi broblast 1785-P growth factor 21 (FGF21), which improves metabolic syndrome, and its plasma Chronic Sucralose Treatment Alters Hepatic Glucose Metabolism level, which is at least partially attributed to these effects of CREB3L3 on in C57BL/6J Mice the systemic energy homeostasis. Therefore, CREB3L3 is a crucial regulator WEINA CONG, HUAN CAI, RUI WANG, RUIN MOADDEL, WILLIAM H. WOOD III, of starvation responses. KEVIN G. BECKER, STUART MAUDSLEY, BRONWEN MARTIN, JOSEPHINE EGAN, Baltimore, MD To ameliorate the current obesity epidemic, artifi cial sweeteners are widely used to provide sweet taste without adding extra caloric load. Similar

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A460 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER to nutritive sweeteners, sweet sensation induced by artifi cial sweeteners 1787-P is also mediated by lingual T1R2-T1R3 G protein-coupled receptor. As the Chronic HMG-CoA Reductase Inhibitor Treatment Contributes to most popular artifi cial sweeteners, sucralose(SUA) is 600 times as sweet Dysglycemia by Up-Regulating Hepatic Gluconeogenesis through as sucrose, and recognized as safe according to the American Dietetic Autophagy Induction Association, although its chronic actions on energy metabolism remain HYE JIN WANG, JAE YEO PARK, EUN YEONG CHOE, OBIN KWON, YONG-HO unclear. We therefore investigated the peripheral metabolic effects of LEE, GYURI KIM, JAEHYUN BAE, SE HEE PARK, YU JUNG YUN, MIJIN YUN, protracted exposure (40 weeks) to SUA in normal C57BL/6J mice. We found CHUL HOON KIM, HYANGKYU LEE, YOUNG BUM KIM, EUN SEOK KANG, Seoul, the chronic treatment of SUA (5mM) led to a profound and stable SUA Republic of Korea, Boston, MA exposure (~191uM) in circulation. In contrary to the popular belief, SUA- We investigated the molecular mechanism of the diabetogenic effect of treated mice developed signifi cantly higher body mass (p<0.05) than water- 3-hydroxy-3-methyl-glutaryl-CoA (HMG-CoA) reductase inhibitors, or statins, treated Controls. However, the SUA-treated mice displayed a starvation-like in the liver by treating human hepatoma (HepG2) cells with statins for 24 hr metabolic status including signifi cantly lower fasting glucose, increase and by feeding fi ve-week old mice a high fat diet and treating with statins for of plasma glucagon and enhancement of ketone bodies levels. Most 16 weeks. HepG2 cells treated with statins exhibited increased expression importantly, the hepatic glycogen content was also dramatically reduced of key gluconeogenetic enzymes, including glucose-6-phosphatase (G6Pase) by the chronic SUA treatment (p<0.001). Further mechanistic studies in and phosphoenol pyruvate carboxykinase (PEPCK), as did the livers of mice both liver and hepatocytes demonstrated the depletion of hepatic glycogen treated with statins. Statins induced autophagy by inhibiting mammalian is related to the continuous activation of PKA-mediated glycogenolysis target of rapamycin (mTOR) and p70 ribosomal S6 kinase (p70S6K). Statin and profound inhibition of Glucose-6-pase protein expression. Our study treatment increased light chain 3 beta (LC3b) conversion and decreased demonstrates that chronic SUA treatment in normal C57BL/6J mice not nucleoporin p62 (p62) substrate levels. Chloroquine (CQ) attenuated only failed to maintain/lose bodyweights, but also predispose body into a autophagy by reversing the increase in G6Pase and PEPCK expression caused starvation-like metabolic condition with hepatic glycogen depeletion. Our by statin treatment. Beclin shRNA or LC3b shRNA treatment reducedG6Pase work represents a novel and important contribution to the fi eld of biological and PEPCK expression levels as well as glucose production. Mice treated science as well as a direct translational implication for human health. with statins gained more weight than control mice and exhibited elevated Supported By: NIH fasting blood glucose levels starting 12 weeks after treatment began. However, there was no signifi cant difference in glucose tolerance. Pyruvate 1786-P tolerance and insulin tolerance test results suggested that statin treatment A New Methodology for the Reproducible Measurement of Hepatic increased hepatic gluconeogenesis. Liver tissue from statin-treated mice De Novo Lipogenesis in Humans exhibited more prominent autophagosomes than that of control mice. These CARINE BEYSEN, SCOTT TURNER, SANTOS CARVAJAL-GONZALEZ, CLARE data suggest that chronic statin treatment contributes to the development of BUCKERIDGE, MARC HELLERSTEIN, WILLIAM P. ESLER, GABRIELE E. SONNEN- diabetes by upregulating hepatic gluconeogenesis via autophagy induction. BERG, Emeryville, CA, Cambridge, MA Supported By: Yonsei University Inhibitors of hepatic de novo lipogenesis (DNL) are potential therapies for T2DM and hepatic steatosis but variability in DNL rates make treatment 1788-P effects diffi cult to quantify. We evaluated a reproducible method for the Intermittent High Glucose Increased Hepatocytes Apoptosis in assessment of DNL in response to oral fructose. Healthy subjects were Nonalcoholic Fatty Liver Disease via Mitochondrial Permeability studied at baseline (n=30) and a repeat visit (subset of n=8). A 13C-acetate Transition Opening infusion and Mass Isotopomer Distribution Analysis were used to assess XUEYAO YIN, FENPING ZHENG, QIANQIAN PAN, SAIFEI ZHANG, DAN YU, HONG DNL (fraction of new palmitate in plasma very low density lipoprotein LI, Hangzhou, China triglycerides). DNL was measured after an overnight fast and during 10-hr Diabetes appeared to be a signifi cant risk factor for the development fructose consumption. Peak DNL during fructose stimulation was adjusted of NAFLD; however, the mechanisms are far from clarifi ed. Hepatocytes for fasting DNL (max DNL). At baseline (n=30), fasting DNL, peak DNL and apoptosis, usually resulting from mitochondrial dysfunction and oxidative max DNL were 8 ± 5%, 36 ± 7% and 28 ± 8%, respectively. There was stress promoting the progression of simple steatosis to nonalcoholic considerable intersubject variability in fasting DNL (range: 1 - 21% and CV steatohepatitis (NASH). Previous studies have found that intermittent of 61%) but less for peak DNL (range: 22 - 47% and CV of 19%) and max high glucose (IHG) plays a more signifi cant role in the oxidative stress DNL (range: 15 - 42% and CV of 28%). The fructose-stimulated DNL time than sustained high glucose (SHG). The present study was to clarify the course was similar for the 2 visits (Fig). Fasting DNL, peak DNL and max contribution of glucose variability to the development of the NAFLD, and the DNL were 7 ± 6% vs. 11 ± 6%; 37 ± 6% vs. 40 ± 4% and 30 ± 8% vs. 29 ± potential mechanisms. Hepatic L02 cells were incubated with palmitic acid 8%, respectively at baseline vs. repeat visit (n=8). Correlations between (PA) to induce steatosis. L02 cells were exposed to control (NG, 5.5mmol/L), the 2 visits for fasting DNL, peak DNL and max DNL were r=0.52, r=0.62 SHG (33.3 mmol/L) or IHG (5.5 and 33.3 mmol/L) alternating every 12 h for 3 and r=0.87, respectively. In contrast to fasting DNL, fructose stimulated days with or without PA. C57BL/6J mice maintained on high fat diet (HFD) DNL represents a reproducible method to study the impact of disease or were injected with glucose (3g/kg) or saline twice a day to induce IHG. therapy on hepatic lipogenic activity. In the L02 cells with PA, compared with NG, SHG induced mitochondrial dysfunction, ROS production and cell apoptosis. However, these effects were more pronounced in IHG with PA. Although glucose homeostasis, hepatic enzyme levels, liver lipid deposition and fi brosis were comparable between HFD mice received glucose and saline injection, HFD mice with glucose injection displayed a marked increase in hepatocytes apoptosis accompanied by increased lipid peroxidation content in liver and cytochrome c release from mitochondria, and decreased ATP production. Co-treatment with the mitochondrial permeability transition (MPT) inhibitor, cyclosporin A (1.5µmol/L), prevented both IHG and SHG with PA induced mitochondrial Integrated

dysfuntion and ROS production, as well as hepatocytes apoptosis. In POSTERS conclusion, glucose fl uctuation together with lipotoxity, might contribute to the development of NAFLD by increasing hepatocytes apoptosis, and MPT Physiology/Obesity may act as a “central executioner.” Supported By: NSFC (81000364)

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A461 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1789-P 1791-P Liraglutide Prevents Steatohepatitis, Liver Fibrosis, and Accom- Liver-specifi c Expression of Dominant Negative Transcription panying Carcinogenesis in a Diabetes and Nonalcoholic Steato - Factor 7-like 2 Causes Impaired Glucose Homeostasis in Mice hepatitis Mouse Model Treated with STZ-HFD WILFRED IP, WEIJUAN SHAO, TIANRU JIN, Toronto, ON, Canada MOTOYASU KOJIMA, TAKUYA KUWASHIRO, MICHIAKI OKADA, YAYOI MAT- Certain common single nucleotide polymorphisms in the transcription SUDA, YOICHIROU KITAJIMA, HIROKAZU TAKAHASHI, KENJI ASHIDA, IWATA factor 7-like 2 (TCF7L2) gene are strongly associated with type 2 diabetes OZAKI, YUICHIROU EGUCHI, KEIZO ANZAI, Saga, Japan, Boston, MA, Fukuoka, risk. Although extensive research has been focused on the role of TCF7L2 in Japan pancreatic beta-cells, recent studies have revealed that TCF7L2 may exert Introduction: Nonalcoholic fatty liver disease (NAFLD) possibly leads important functions elsewhere, including the liver. Using various in vitro to nonalcoholic steatohepatitis (NASH), liver cirrhosis, and hepatocellular approaches, we and others have shown previously that TCF7L2 and Wnt carcinoma. Liraglutide is a glucagon-like peptide-1 (GLP-1) agonist developed signaling are negative regulators of hepatic gluconeogenesis. In the current for the treatment of type 2 diabetes, and has been reported to improve liver study, we aimed to determine the in vivo role of TCF7L2 in hepatic glucose enzymes, oxidative stress and hepatic steatosis in vivo and in vitro. We metabolism. We generated a liver-specifi c dominant negative TCF7L2 hypothesized that liraglutide or linagliptin, known as dipeptidyl peptidase-4 (TCF7L2DN) transgenic mouse model and confi rmed exclusive expression inhibitor (DPP-4I), could prevent steatohepatitis, fi brosis and carcinogenesis of TCF7L2DN in the liver. Transgenic animals exhibited impaired glucose as in NASH. well as pyruvate tolerance, suggesting that TCF7L2DN expression causes Methods: Male STAM mice, a NASH mouse model that shows short- increased hepatic glucose production. Insulin tolerance was comparable term progression to liver cirrhosis and cancer, were used. STAM mice between the transgenic and wild-type control mice, while hepatic glycogen was developed by administration of streptozotocin in C57BL/6J mice at 2 and triglyceride content were increased in transgenic animals. To further days old, and fed a high-fat diet from 4 weeks. STAM mice generally show examine the mechanism by which TCF7L2 regulates hepatic metabolism, hyperglycemia and fatty liver at 5 weeks, hepatic fi brosis and steatohepatitis we expressed TCF7L2DN in primary hepatocytes. Adenoviral delivery of at 9 weeks, and hepatocellular carcinoma at 16 weeks. STAM mice were TCF7L2DN expression in hepatocytes caused higher production of glucose treated with liraglutide (GLP-1 group), linagliptin (DPP-4I group) or saline from gluconeogenic precursors. The mRNA levels of gluconeogenic genes (Vehicle group) from 6 weeks old. Blood glucose, insulin, and histopathology Pck1, G6pc, Fbp1, and Ppargc1a were signifi cantly increased in hepatocytes of the liver were analyzed at 6, 12, and 20 weeks old. expressing TCF7L2DN. In addition, TCF7L2DN expression caused increased Results: The GLP-1 group showed signifi cantly lower plasma glucose levels mRNA expression of Gys2, which encodes the enzyme that catalyzes than the Vehicle group (161.8 ± 45.4 mg/dL vs. 457.7 ± 116.9 mg/dL, P<0.01), glycogen synthesis, as well as the lipogenic gene Fasn. Together, the higher insulin levels (0.18 ± 0.06 ng/mL vs. 0.09 ± 0.03 ng/mL, P<0.05), utilization of TCF7L2DN as a unique tool allows us to suggest that TCF7L2 and a lower NAFLD activity score (NAS) (12 weeks: 2.50 ± 1.73 vs. 5.25 ± serves a benefi cial role in the liver by suppressing gluconeogenic gene 0.95, P<0.05; 20 weeks: 1.00 ± 0.82 vs. 5.75 ± 1.26, P<0.01). Hepatocellular expression. carcinoma was observed in all Vehicle group mice at 20 weeks and not in Supported By: CIHR any GLP-1 group mice (tumor number/mouse: 5.50 ± 3.87 vs. 0, P=0.00). Tumor number was signifi cantly fewer in the DPP-4I group compared with 1792-P the Vehicle group (1.25 ± 0.50, P<0.05). β-Cryptoxanthin Improves Hepatic Insulin Resistance and Conclusion: Liraglutide improved the pathogenesis of NASH, and lira- Infl ammation Through M2 Dominant Shift of Macrophages in Diet- glutide and linagliptin could suppress hepatocarcinogenesis in this NASH induced Nonalcoholic Fatty Liver Disease mouse model. TSUGUHITO OTA, YINHUA NI, MAYUMI NAGASHIMADA, FEN ZHUGE, NAOTO NAGATA, SHUICHI KANEKO, Kanazawa, Japan 1790-P Excessive hepatic lipid accumulation promotes activation of macrophage/ Dawn Phenomenon in Type 1 Diabetes: Dual Tracer Approach to kupffer cells, resulting in exacerbation of insulin resistance and hepatic Measure Effect Size and Tracer Recycling infl ammation. β-cryptoxanthin is a carotenoid compound that is known to have ASHWINI MALLAD, LING HINSHAW, CHIARA DALLA MAN, CLAUDIO COBELLI, a potent anti-infl ammatory effect by primarily modulating the innate immune RITA BASU, YOGISH C. KUDVA, ANANDA BASU, Rochester, MN, Padova, Italy response. In this study, we examined the effect of β-cryptoxanthin on diet- Dawn phenomenon infl uences fasting glucose concentrations in type 1 induced NAFLD to clarify the signifi cance of lipotoxicity-mediated hepatic diabetes (T1D). To determine it’s effect size and hormonal basis, we studied activation/polarization of macrophages in insulin resistance and infl ammation 14 subjects with C-peptide negative T1D (age 45.5±13.1 yrs., BMI 28.7± 5.9 in the pathogenesis of NAFLD/NASH. C57BL/6 mice were fed on a high- kg/m2, HbA1c 7.6 ± 0.7 %) on their individualized open loop insulin pump cholesterol/high-fat diet (CL) or a CL diet containing β-cryptoxanthin 3 mg/kg therapy for 2 successive nights. Endogenous glucose production (EGP) rates (CL+CX) for 12 weeks. Histological examination revealed hepatic steatosis 2 at 4AM and 7AM were measured with infusion of [6,6- H2] glucose from 12 and infl ammation in mice fed CL diet. They showed hyperinsulinemia (CL AM to 7 AM. To determine extent of tracer recycling, we additionally infused 2.5±0.3 vs. normal chow 0.5±0.1 ng/ml, p<0.01), indicating that mice fed on a [6-3H] glucose from 4 AM to 7 AM in 5 of 14 subjects. Counter-regulatory CL diet developed NASH associated with insulin resistance. β-cryptoxanthin hormones were also measured. Plasma glucose concentrations tended to administration improved glucose intolerance and hyperinsulinemia, and rise from 4 to 7AM (night 1: Δ 1.3±0.8 mM, p=0.13; night 2: Δ 1.0±0.5 mM, also enhanced insulin signal assessed by IRβ and Akt phosphorylation in p=0.09). There were no episodes of nocturnal hypoglycemia. While cortisol the liver. β-cryptoxanthin reduced hepatic TG and TC levels by 36% and concentrations rose (p< 0.05) consistently during both nights from 4 to 7AM 22% respectively (p<0.01), and decreased F4/80+ macrophage infi ltration (night 1: Δ 4.9 ± 0.8 µg/ml; night 2: Δ 2.4± 0.9 µg/ml), glucagon concentrations in liver of CL group. Integrated pathway analysis using cDNA microarray 2 increased (p=0.01) only during night 1. EGP measured with [6,6- H2] glucose showed that β-cryptoxanthin signifi cantly downregulated macrophage did not differ during night 1 (18.7±4.9 µM/kg/min at 4AM vs. 18.4±5.4 µM/ activation signal related genes without affecting most FA or cholesterol kg/min at 7AM: p=0.8) or night 2 (17.3± 6.2 µM/kg/min at 4AM vs. 15.7±5.5 metabolism-related genes. Moreover, FACS analysis revealed that mice µM/kg/min at 7AM: p=0.06). However, EGP measured with [6-3H] glucose at fed CL+CX had 51% fewer CD11c+CD206- (M1)-type macrophages but had 2 - + 7AM was higher (p<0.001) than that measured with [6,6- H2] glucose during 170% more CD11c CD206 (M2)-type macrophages in liver than CL diet fed

Integrated 3 2 POSTERS night 1 (38.7±9.4 µM/kg/min vs. 22.1±6.4 µM/kg/min: [6- H] vs. [6,6- H2] mice, resulting in a predominance of M2 over M1 macrophage population. glucose) and night 2 (23.3±8.7 µM/kg/min vs. 16.3±8.1 µM/kg/min: [6-3H] In conclusion, β-cryptoxanthin improves hepatic insulin resistance and Physiology/Obesity 2 vs. [6,6- H2] glucose) indicating signifi cant tracer recycling underestimating infl ammation through an M2-dominant shift in macrophge/Kupffer cells in 2 EGP calculated at 7AM with [6,6- H2] glucose. Likewise, EGP was higher diet-induced NAFLD. 3 2 (p<0.01) at 7AM (with [6- H] glucose) than 4AM (with [6,6- H2] glucose) Supported By: Ministry of Education, Culture, Sports, Science and Technology; during both nights. The data demonstrate consistent but variable overnight Ministry of Agriculture, Forestry and Fisheries rise in glucose concentrations through increased EGP, mediated primarily by rising cortisol concentrations. Closed loop control algorithms will need to account for these changes with variable effect sizes from night to night, to effectively treat T1D. Supported By: NIDDK (085516, 094331)

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A462 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1793-P demonstrated that glucose output is increased in LKB1 KO hepatocytes, Jinlida Reduces Insulin Resistance by Ameliorating Liver Oxidative which is associated with an increased expression of gluconeogenic Stress via JNK and p38MAPK Pathways genes and dephosphorylation of CRTC2. In contrast, glucose production SONG GUANGYAO, LIU YIXUAN, MA BOQING, ZANG SHASHA, WANG CHAO, and gluconeogenic gene expression are not altered in AMPKα1/2 KO Shijiazhuang, China hepatocytes although the phosphorylation of CRTC2 is lost in response to The Chinese medicine Jinlida (JLD) is traditionally used to treat diabetes AMPK activators. by improving insulin resistance. We aimed to elucidate the mechanism of JLD In an effort to understand the mechanism implicated in the increase treatment, in comparison to metformin treatment, on ameliorating insulin of gluconeogenesis in LKB1 KO hepatocytes, we generated a new mouse sensitivity in insulin resistant rats and to reveal its anti-oxidant properties. model deleted for SIK2. Surprisingly, glycemia of liver-specifi c SIK2 KO mice Rats were fed with standard or high-fat diet for 6 weeks. After 6 weeks, the is indistinguishable from control. Moreover, glucose output, gluconeogenic high-fat fed rats were subdivided into fi ve groups and orally administrated gene expression and phosphorylation of CRTC2 were not altered in SIK2 by JLD or metformin for 8 weeks. Fasting blood glucose (FBG), fasting blood KO hepatocytes. We then assumed that AMPK might compensate for insulin, blood lipid and antioxidant enzymes were measured. Intraperitoneal the loss of SIK2. To investigate this hypothesis, we generated triple KO glucose tolerance test (IPGTT) and hyperinsulinemic euglycemic clamp mice deleted of AMPKα1/2 and SIK2 in the liver. Unexpectedly, glycemia technique were carried out to measure insulin sensitivity. Gene expression and hepatic gluconeogenesis were not affected in the absence of AMPK of the major signaling pathway molecules that regulate glucose uptake, and SIK2 in the liver. These results support the concept that the deletion including insulin receptor (INSR), insulin receptor substrate-1 (IRS-1), of AMPK and SIK2 alone or in combination are not suffi cient to mimic the phosphoinositide-3-kinase (PI3K), protein kinase beta (AKT), and glucose alteration of gluconeogenic program observed in LKB1 KO liver. Finally, we transporter type 2 (GLUT2), were assessed by quantitative RT-PCR. The speculated that all SIK isoforms should be simultaneously inactivated to protein expression of IRS-1, AKT, JNK and p38MAPK were determined observe an increase of glucose production in the liver. This hypothesis was by Western blot. Treatment with JLD effectively ameliorated the high-fat supported by using a highly selective pan SIK inhibitor in hepatocytes, which induced hyperglycemia, hyperinsulinemia and hyperlipidemia. Similar to recapitulated completely the increase of gluconeogenesis observed in LKB1 metformin, the high insulin resistance in high-fat fed rats was signifi cantly KO hepatocytes. decreased by JLD treatment. JLD displayed anti-oxidant effects, coupled In summary, our results demonstrate that the LKB1-SIK pathway inhibits with up-regulation of the insulin signaling pathway. The attenuation of hepatic gluconeogenesis and suggest that SIK could be a new therapeutic hepatic oxidative stress by JLD treatment was associated with reduced target to improve blood glucose in patients with type 2 diabetes. phosphorylation protein levels of JNK and p38MAPK. The results suggested Supported By: Région Ile-de-France (CORDDIM) that treatment with JLD can moderate glucose and lipid metabolism as well as reduce hepatic oxidative stress, most likely through the JNK and 1796-P p38MAPK pathways. Effects of Intranasal Insulin on Fasting Glucose, Lipid and Hepatic Supported By: Yiling Company Energy Homeostasis in Humans With and Without Type 2 Diabetes SOFIYA GANCHEVA, CHRYSI KOLIAKI, ALESSANDRA BIERWAGEN, PETER 1794-P NOWOTNY, ANDREA NAGEL, NICOLE ACHTERATH, KAI TINNES, MYRKO ESSER, Free Fatty Acids Induce FGF21 Resistance in HepG2 Cells MARTIN HENI, ANDREAS FRITSCHE, JULIA SZENDROEDI, MICHAEL RODEN, MOHAMED ASRIH, FRANCOIS R. JORNAYVAZ, Geneva, Switzerland Düsseldorf, Germany, Tübingen, Germany Fibroblast growth factor 21 (FGF21) is a key mediator of glucose and lipid Central delivery of insulin may regulate peripheral glucose and lipid metabolism and has been shown to reverse hepatic steatosis and improve metabolism through indirect cross-talk between brain and peripheral tissues insulin resistance. Surprisingly, however, the benefi cial effects of exogenous in rodents. This study investigates the acute effects of central insulin on FGF21 administration are somewhat attenuated in obese patients with type endogenous glucose production (EGP), lipid and hepatic energy metabolism 2 diabetes (T2DM) and in animal models of obesity and T2DM, suggesting an in humans. Eight lean normoglycemic humans (age 26±2 yrs, BMI 23±1 kg/ FGF21-resistant state. T2DM is associated with increased plasma levels of m2, CON) and 8 patients with type 2 diabetes (age 61±3 yrs, BMI 27±1 kg/ free fatty acids (FFA), which lead to impaired insulin signaling and increased m2, T2D) received intranasal insulin to mimic central insulin delivery (160 lipogenesis. Thus, we investigated how FFA impact FGF21 effects on hepatic IU; Actrapid, Novo-Nordisk, Denmark) or placebo in a randomized, single- lipid metabolism. For this purpose, we exposed human liver-derived HepG2 blinded, cross-over trial. Rates of EGP were assessed with [6,6-2H2] cells to FGF21 either in presence or absence of FFA. glucose and metabolic parameters were monitored continuously for 3 hours. In the absence of FFA, treatment of HepG2 with FGF21 reduced lipogenesis Hepatic fat content (HCL) and adenosine triphosphate (ATP) concentrations and increased lipid oxidation. Inhibition of lipogenesis was associated with were determined at baseline and 3 hours after intervention by magnetic the downregulation of SREBP-1c (FGF21 vs. controls (C): -60%, p<0.05) and resonance spectroscopy (31P/1H-MRS) on a 3T clinical MR scanner (Philips, its downstream target genes (FAS, SCD1) as revealed by RT-qPCR. The lipid- Best). Circulating glucose, glycerol and triglycerides were not affected by lowering effect was mediated via the phosphorylation of AMPK and its either insulin or placebo in both groups. Fasting hepatic insulin sensitivity, downstream target ACC, and the upregulation of CPT-1α (FGF21 vs. C: +50%, as determined from EGP and insulin levels, was lower in T2D for 180 min p<0.01). FGF21 was unable to induce neither basal nor insulin-stimulated (9.7±0.9 vs. 13.5±0.7, p=0.002), but not changed after intranasal insulin in Akt phosphorylation, but similarly to glitazones, FGF21 upregulated PPARγ either group compared to placebo. Free fatty acid concentration decreased (FGF21 vs. C: +75%, p<0.01) and adiponectin gene expression (FGF21 vs. C: by 42±8% (p=0.003 vs. placebo) in CON, but was not altered in T2D at 60 +670%, p<0.001), a known mediator of some FGF21 metabolic effects, while min after intranasal insulin. HCL decreased (Δ=0.2±0.1%, p=0.04), whereas it reduced TNFα gene expression (FGF21 vs. C: -50%, p<0.05). In contrast, the absolute γ-ATP concentrations increased (Δ=0.5±0.2 mmol/l, p=0.04) after addition of FFA abolished the benefi cial effects of FGF21 on lipid metabolism. intranasal insulin in CON, but not in T2D. In conclusion, intranasal insulin Moreover, FGF21 did not affect FFA accumulation as assessed by oil red O does not seem to affect glucose production in humans, but associates with staining. benefi cial effects on lipid and energy metabolism. These effects are blunted In conclusion, in the absence of FFA, FGF21 improves lipid metabolism in in T2D patients, which could be due to their lower insulin sensitivity. HepG2 cells notably through an increase of PPARγ gene expression and a Supported By: Helmholtz Alliance ICEMED Integrated reduction of the infl ammatory cytokine TNFα. However, under high levels POSTERS of FFA, FGF21 has no effect on lipid metabolism. We therefore propose that 1797-P high levels of FFA represent an FGF21-resistant state. Synergy between Metformin and Leucine in Sirtuin Signaling and Physiology/Obesity Fat Oxidation In Vitro, and in Reducing Lipid Accumulation in Diet- 1795-P induced Obese Mice Role of AMPK and SIK in the LKB1-dependent Regulation of Hepatic HANG SHI, LIZHI FU, FENFEN LI, ANTJE BRUCKBAUER, QIANG CAO, XIN CUI, RUI Gluconeogenesis WU, MICHAEL B. ZEMEL, BINGZHONG XUE, Atlanta, GA, Knoxville, TN ALLISON MARION, NADIA BOUDABA, KEI SAKAMOTO, BENOIT VIOLLET, MARC We previously found leucine (Leu) to stimulate Sirt1 and AMPK signaling FORETZ, Paris, France, Lausanne, Switzerland in vitro and in vivo. Since metformin (Met) converges on the same pathway, AMP-activated protein kinases (AMPK) and salt-inducible kinases (SIK) we have tested the ability of Leu to amplify the effects of Met on AMPK, are activated by the LKB1 kinase and have been involved in the regulation Sirt1 and lipid metabolism. Met (10-100 µM) and Leu (0.5 mM) combinations of gluconeogenesis via phosphorylation of a common downstream target, synergistically increased AMPK phosphorylation and activity 50% (p<0.01), the CREB regulated transcription coactivator 2 (CRTC2). Recently, we Sirt1 activity 40-60% (p<0.01), mitochondrial mass 24-51% (p<0.04) and

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A463 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

fatty acid oxidation (FAO) by 50-110% (p<0.01) in adipocytes and myotubes after 10 days of treatment. T2 did not alter fasting plasma glucose or insulin and the effects on AMPK and FAO were augmented by low-dose (200 nM) concentration. Basal endogenous glucose production (EGP) was unchanged. resveratrol (Resv). To evaluate this synergy in vivo, diet-induced obese (DIO; T2 treatment modestly improved hepatic insulin action: insulin suppressed induced by high fat diet) mice were fed high Leu (24 g/kg diet) with or without EGP by 74% ± 10 in T2-treated rats as compared with 46% ± 5 suppression in Resv (12.5 mg/kg diet) with or without subtherapeutic levels of Met (0.05- the vehicle group (p = 0.04). This improvement in hepatic insulin sensitivity 0.50 g/kg diet) or therapeutic levels of Met (1.5 g/kg diet; ~300 mg/kg BW). was associated with improvements in hepatic insulin signaling. The increase Compared to control low-fat diet mice, DIO mice exhibited a 10-fold increase in insulin stimulated Akt phosphorylation was ~2.5 fold greater in T2- in inguinal fat pad weight and 69% increase in liver weight (p<0.0002); treated animals as compared with vehicle-treated rats (p = 0.003). T2 did histology confi rmed the latter to be due to steatosis. Neither Leu nor Resv, not activate classical nuclear THRs, as assessed by mRNA expression of individually or in combination, affected fat pad or liver mass in the absence of genes highly sensitive to nuclear THR activation, dio1 and hrl. There was Met. However, addition of subtherapeutic Met reduced fat pad mass by 31%, also no change in expression of genes regulating hepatic lipid and glucose comparable to the effets of therapeutic Met (p<0.0001). Moreover, addition metabolism including fatty acid synthase, PPARa, and glucose 6-phosphatase. of subtherapeutic levels of Met resulted in a dose-responsive decrease in CONCLUSION: In contrast with previous reports, in rats fed an unsaturated liver mass, with the highest dose exerting signifi cantly greater effects than fat diet T2 administration failed to improve NAFLD or whole body insulin therapeutic levels of Met and fully reversing hepatic steatosis (p<0.0002). sensitivity and only modestly improved hepatic insulin action. T2 is unlikely This was accompanied by substantial reversal of DIO-induced infl ammation to be an effective treatment for NAFLD and dyslipidemia. as measured by serum CRP (p<0.001). Thus, Leu and Met exhibit signifi cant synergy with respect to Sirt1-AMPK signaling, and a low-dose Leu-Met 1800-P combination exerts comparable effects on adiposity to therapeutic doses of High Risk of Nonalcoholic Fatty Liver Disease (NAFLD) and Steato- Met and fully reverses hepatic steatosis in DIO mice, enabling 66-83% Met hepatitis (NASH) in Obese Patients with Type 2 Diabetes Mellitus dose reduction in this model. (T2DM) and Normal Liver Aminotransferases Supported By: NuSirt Sciences, Inc. MARYANN MAXIMOS, FERNANDO BRIL, PAOLA PORTILLO SANCHEZ, ROMINA LOMONACO, DIANE BIERNACKI, SREEVIDYA SUBBARAYAN, AMITABH SUMAN, 1798-P KENNETH CUSI, Gainesville, FL The I148M Variant in PNPLA3 Is Associated with Altered Hepatic In patients (pts) with T2DM and normal liver aminotransferases (LFTs) the Lipid Composition in Humans true prevalence of nonalcoholic fatty liver disease (NAFLD), and its more ANDREAS PETER, MARKETA KOVAROVA, ALFRED KOENIGSRAINER, FAUSTO severe form with steatohepatitis and fi brosis (NASH), is unknown. The MACHICAO, NORBERT STEFAN, HANS-ULRICH HAERING, ERWIN SCHLEICHER, common belief is that pts with normal LFTs do not have NAFLD, but emerging Tübingen, Germany, Pardubice, Czech Republic evidence in non-diabetics suggests the opposite. However, no studies have The common sequence variant I148M in patatin-like phospholipase focused on the prevalence of NAFLD/NASH in T2DM or used the gold- domain 3 (PNPLA3) is associated with increased hepatic triacylglyceride standard magnetic resonance and spectroscopy (MRS) to screen for NAFLD, (TAG) content but not with insulin resistance or diabetes in humans. In but just ultrasound or elevated LFTs. To this end, we screened pts with vitro data and a transgenic mouse model demonstrated that PNPLA3 T2DM and normal LFTs for NAFLD and NASH using a 2-step approach (liver I148M not only increases hepatic lipid content, but also confers qualitative MRS, followed by a liver biopsy if positive) and also performed measures changes in hepatic lipid composition which may contribute to the metabolic of sensitivity. We studied 90 pts with T2DM, normal LFTs, and no prior consequences. Therefore we hypothesised that PNPLA3 I148M impacts diagnosis of NAFLD (age:59±1years; gender:79% male;BMI:33.5±0.5kg/m2; the hepatic lipid composition also in humans and analyzed the fatty acid HbA1c: 7.2±0.1%). Also, 37 pts with elevated liver enzymes served as controls 2 composition of 5 lipid fractions in liver biopsies from 52 human subjects (age:56±2years; gender: 78% male; BMI: 33.2±0.9kg/m ; HbA1c: 7.2±0.2%; all including 19 carriers of the PNPLA3 I148M variant.PNPLA3 I148M was p=NS). The prevalence of NAFLD on MRS was 83% in the cohort with normal associated with a strong increase (1.75-fold) in liver TAG but not other lipid LFTs vs. 97% in pts with elevated LFTs (p=0.03). Surprisingly, 52% of those fractions. The relative fatty acid composition of the TAGs showed a distinct with NAFLD and normal LFTs had NASH on histology (vs. 74% in elevated pattern with signifi cantly increased content of the n-3 polyunsaturated LFTs controls; p<0.05). Fibrosis, mostly mild to moderate, occurred in more fatty acid (PUFA) alpha-linolenic acid (18:3 n-3) by 44% and decreased fatty than half of the pts with NASH and normal LFTs, but was higher (78%) in acids of the n-6 series: eicosatrienoic acid (20:3) by 34%, arachidonic acid those with elevated LFTs. The higher prevalence of NAFLD/NASH in pts with (20:4) by 44%, docosatetraenoic acid (22:4) by 50% and the end product elevated LFTs was associated with more insulin resistance in muscle (Rd: docosapentaenoic acid (22:5) by 38%. These results are in part concordant p<0.01) and adipose tissue (Adipo-IR [fasting FFA x insulin], p<0.01). with the changes detected in mouse livers overexpressing human PNPLA3 Conclusion: The majority (8 out of 10) of obese pts with T2DM and I148M. Noteworthy, most PUFAs also showed an inverse correlation with normal liver enzymes have NAFLD. More importantly, half of these pts have TAG content independent of the PNPLA3 genotype. In a multivariate model biopsy-proven NASH, most with some degree of fi brosis, placing them at including liver fat content, PNPLA3 genotype and fatty acid composition, an increased risk of disease progression and cirrhosis. Thus, physicians two signifi cant changes remained: reduced stearic acid and increased must have a high degree of clinical suspicion for NAFLD and NASH in this alpha-linolenic acid content in hepatic TAG. These two changes in fatty acid population. composition can be exclusively attributed to the PNPLA3 I148M mutation Supported By: ADA (1-08-CR-08); Burroughs Wellcome Fund and not simple steatosis. Since stearic acid can induce insulin resistance and alpha-linolenic acid can protect from it, these changes may be part of the 1801-P mechanism how PNPLA3 I148M increases liver fat content without causing Caffeic Acid Ameliorates Hepatic Steatosis via Decreased ER Stress insulin resistance. and Increased Autophagy in High-Fat Diet-induced Obese Mice HONG-MIN KIM, EUN SOO LEE, EUN YOUNG LEE, CHOON HEE CHUNG, Wonju, 1799-P Republic of Korea, Cheonan, Republic of Korea 3,5-Diiodo-L-Thyronine Improves Hepatic Insulin Sensitivity but Nonalcoholic fatty liver disease (NAFLD) is characterized by an increase Not NAFLD in Fat-Fed Rats in hepatic triglyceride (TG) contents which is associated with endoplasmic Integrated POSTERS DANIEL F. VATNER, JACLYN SNIKERIS, VIOLETA B. POPOV, RACHEL J. PERRY, reticulum (ER) stress and insulin resistance. Caffeic acid (CA) is an organic YASMEEN RAHIMI, VARMAN T. SAMUEL, New Haven, CT compound classifi ed as a hydroxycinnamic acid. It is an antioxidant and Physiology/Obesity Hepatic selective thyroid hormone mimetics remain conceptually alluring also has immunomodulatory and anti-infl ammatory activities. We studied treatments for diseases such as dyslipidemia, nonalcoholic fatty liver disease whether CA ameliorates hepatic steatosis or not and how it does. (NAFLD) and insulin resistance. Though diiodothyronines are thought inactive, We categorized three groups such as low fat diet mice (LFD, n=10), high fat pharmacologic treatment with 3,5- Diiodo-L-Thyronine (T2) reportedly alters diet-induced obese mice (HFD, n=10) and HFD fed with CA (50 mg/kg per day, the expression of genes regulating hepatic lipid and glucose metabolism but n=10) for 10 weeks. Plasma glucose, insulin, triglyceride and total cholesterol without nuclear thyroid hormone receptor (THR) activation, thus decreasing levels were measured by enzyme-linked immunosorbent assay. Hepatic fat hepatic steatosis and improving glucose tolerance in fat-fed rats. To further was measured by histologic examination. Hepatocytes (AML12) were treated study this compound, unsaturated fat-fed male SD rats were treated with with palmitate (250 µM). Palmitate-treated AML12 hepatocytes were cultured T2 (0.25 mg/kg-d) or vehicle. Neither 10 nor 30 days of T2 treatment had with or without CA (50 µM) for 24 hours and observed lipid accumulation and an effect on weight, adiposity, plasma fatty acids, or hepatic steatosis. lipogenesis markers. The levels of ER stress and autophagy markers in liver Insulin action was quantifi ed in vivo by a hyperinsulinemic-euglycemic clamp and AML12 hepatocytes were measured by western blot.

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A464 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

CA signifi cantly lowered body and liver weight. It decreased plasma test was two-fold higher compared to GK activity during the intravenous test glucose, insulin, triglyceride and total cholesterol levels. Lipid accumulation the same subjects (OGTT KGK vs. FSIGT KGK: 8 x 10-3 vs. 4 x 10-3 P=0.003). in the liver was decreased in HFD + CA group than HFD group. Glucose Hepatic Glycolysis (K12) and lactate clearance were only modestly increased intolerance and insulin sensitivity were markedly improved in HFD + CA during the OGTT test (40%, P=0.05, 15%, P=0.04 respectively). Oral glucose group. Moreover, the levels of ER stress markers (e.g., BiP, ATF4, CHOP, administration specifi cally activates liver glucokinase activity. These data p-eIF2α and XBP-1) were decreased in the liver of HFD + CA group. Otherwise, argue that a signal related to glucose absorption from the GI tract activates autophagy markers (e.g., ATG7, ATG5 and LC3) were increased in the liver of hepatic GK to enhance liver glucose uptake. The signal, which could activate HFD + CA group. CA (50 µM) treatment reduced the lipid accumulation and GK directly or through GKRP, may be GLP-1 or other GI hormones. lipogenesis markers, decreased ER stress and increased autophagy markers 1. Stefanovski D, et al. Estimating hepatic glucokinase activity using a in palmitate-treated AML12 hepatocytes. simple model of lactate kinetics. Diabetes Care. 2012 May;35(5):1015-20. We suggest that caffeic acid may ameliorate hepatic steatosis via decreased ER stress and increased autophagy. 1804-P Does Nonalcoholic Fatty Liver Disease (NAFLD) Develop in Insulin- 1802-P Sensitive Patients? Prevalence of Nonalcoholic Fatty Liver Disease (NAFLD) and FERNANDO BRIL, ROMINA LOMONACO, PAOLA PORTILLO SANCHEZ, BEVERLY Disease Severity among African Americans (AA) with Prediabetes ORSAK, MARYANN MAXIMOS, JOAN FINCH, AMY WEBB, KENNETH CUSI, (preDM) or Type 2 Diabetes (T2DM) Gainesville, FL, San Antonio, TX FERNANDO BRIL, SREEVIDYA SUBBARAYAN, MARYANN MAXIMOS, PAOLA It is believed that NAFLD can only develop in an insulin-resistant state PORTILLO SANCHEZ, ROMINA LOMONACO, DIANE BIERNACKI, AMITABH and the disease is often referred to as the “hepatic manifestation” of the SUMAN, MICHELLE WEBER, KENNETH CUSI, Gainesville, FL metabolic syndrome. However, it is puzzling that a minority of patients Compared to other ethnicities, there is a perception that AA are (pts) with NAFLD appear to be insulin-sensitive. This population has not “protected” from NAFLD and from severe liver disease (steatohepatitis or been carefully characterized in prior studies. To this end, we studied pts NASH) as well as fi brosis. However, the true prevalence of NAFLD/NASH in with (n=19) and without (n=15) NAFLD (liver fat by magnetic resonance AA with preDM/T2DM is unknown as they have been underrepresented in spectroscopy [MRS]: 22±3 vs. 2±1%, p<0.001) closely matched for clinical clinical studies. variables and that were insulin-sensitive during an euglycemic insulin clamp To this end, we screened for fatty liver with magnetic resonance spectro- with 3-3H-glucose (defi ned as peripheral [muscle] insulin sensitivity [Rd] >9.0 scopy (MRS) 93 patients with preDM/T2DM: 31 AA, 31 Caucasians (C) and mg/kg . min). Diabetes was ruled out by an OGTT. Both groups were well 31 Hispanics (H) well-matched for clinical characteristics using propensity- matched for age (47-50 years, p=0.47), gender (47-53% M, p=0.74), BMI (30-31 score matching (age: 55-57 years; male: 84-85%; BMI: 32-33kg/m2; T2DM: kg/m2, p=0.40), A1c (5.5-5.7%, p=0.19). However, despite these similarities, 61-68%; A1c: 6.7-6.9%; ALT: 29-37U/L, all NS). We assessed: 1) liver fat pts with NAFLD had higher plasma TG (122±14 vs. 86±11 vs. mg/dl, p=0.03) (LFAT) by MRS; 2) liver histology (biopsy); 3) liver, muscle (Rd) and adipose and lower HDL-C (40±2 vs. 54±3 mg/dl, p<0.001). Even as pts were selected tissue insulin resistance (IR) during a low- and high-dose insulin clamp with based on a “normal” peripheral (muscle) insulin-sensitivity, NAFLD pts had a 3-3H-glucose. lower muscle insulin sensitivity (Rd: 11.5±0.5 vs. 13.2±0.6 mg/kgLBM . min, Prevalence of NAFLD (AA vs. C vs. H: 81% vs. 71% vs. 77%, p=0.67) p=0.03), a 53% increase in plasma FFA (p<0.05) and worse adipose tissue and LFAT were similar across ethnicities (12% vs. 14% vs. 14%; p=0.82). insulin resistance (p<0.05). No differences were observed in hepatic insulin Lower adipose tissue IR in AA (p<0.05 vs. H) was associated with a 2-fold sensitivity. Of note, insulin-sensitive pts with NAFLD were not protected reduction in plasma TG (AA: 105 vs. C: 159 vs. H: 194 mg/dL, p=0.01). AA from developing severe liver disease (9 out 13 biopsies had NASH and 1 out with NAFLD had severe muscle (Rd: 6.1±1.0 mg/kgLBM · min) and hepatic IR 13 advanced fi brosis). Accordingly, plasma aminotransferases were higher (insulin suppression of EGP: -29±5%), but both were similar to that in C and in pts with NAFLD (ALT: 58±7 vs. 23±3 U/L, p<0.001 and AST: 41±4 vs. 25±2 H. There was a trend for the prevalence of NASH in patients with NAFLD to U/L, p=0.003). be lower in AA (AA: 36% vs. C: 64% vs. H: 53%). Once AA developed NASH, Conclusion: Even in apparently insulin-sensitive patients, NAFLD is histological severity of steatohepatitis (p=0.22) and fi brosis (p=0.86) was associated with subtle reductions in muscle and adipose tissue insulin action similar to C and H. and a worse metabolic profi le (i.e. higher TG, lower HDL-C). We speculate Conclusion: Contrary to common belief that AA are “protected” from that even mild chronic insulin resistance, particularly in adipose tissue, may NAFLD, 8 out of 10 middle-aged AA with preDM/T2DM had NAFLD (similar still place genetically predisposed subjects at risk of NAFLD/NASH. to well-matched C and H). NASH appeared to be less frequent, likely due Supported By: ADA (1-08-CR-08); Burroughs Wellcome Fund to their lower adipose tissue IR, but still affecting about 1/3 of AA patients with similar histological severity. If confi rmed, AA are truly at risk of NAFLD/ 1805-P NASH and early screening is as important as for other ethnicities to prevent FTO Inhibits Autophagy Activity Leading to Hepatocellular Lipid progressive liver disease. Deposition Supported By: ADA (1-08-CR-08); U.S. Dept. of Veterans Affairs JIANJIN GUO, WEI REN, XING LI, JIE LIU, WEIPING JIA, Taiyuan, China, Shanghai, China 1803-P Background and Aims: Fat mass and obesity-associated (FTO) gene was Oral Glucose Delivery Specifi cally Activates Liver Glucokinase, the fi rst locus unequivocally associated with adiposity. We had demonstrated Possibly via Glucokinase Regulatory Protein FTO involved in lipid deposition in Nonalcoholic fatty liver disease (NAFLD). DARKO STEFANOVSKI, RICHARD N. BERGMAN, STEPHANIE T. CHUNG, ANNE E. Autophagy is now linked with several events in the development of NAFLD. SUMNER, Los Angeles, CA, Bethesda, MD Decreased autophagic function drives the initial development of hepatic It has become evident that glucokinase (GK) activity, which regulates the steatosis and its progression to liver injury. Here we analyze the functions of rate of glucose phosphorylation in the liver, is a major determinant of glucose FTO in the progress of hepatocellular lipid deposition. tolerance. In fact variants for GKRP which regulate GK activity have been Materials and Methods: Establish a stable L02 cell line expressing human identifi ed in those at risk for diabetes, and compounds regulating GK and FTO. Control L02 cell and L02/FTO cell cultures were incubated in RPMI- Integrated GKRP have been introduced. It is therefore important to assess GK activity 1640 medium and oleic acid with fi nal concentrations 20µg/ml. After 48 h POSTERS in vivo to establish diabetes risk and compound effi cacy. We introduced of incubation with OA, monodansylcadaverine (MDC) staining was used to a novel model of lactate kinetics (1), based upon the frequently-sampled evaluate the abundance of autophagic vacuoles in cells. Lipid droplets in Physiology/Obesity intravenous glucose tolerance test (FSIGT) to estimate in vivo hepatic GK cytoplasm were observed by oil red O staining. Autophagy-related genes activity, glycolysis and whole body lactate clearance. In the clinical setting Beclin-1, Microtuble-associated protein light chain 3 (MAPLC3), lysosomal the OGTT is a preferred approach for assessing glucose homeostasis. Our membrane proteins (LAMP), autophagy-associated gene 7 (ATG7) expression objective was to estimate the lactate model indices from the oral glucose were investigated using RT-PCR and Western blot analysis. tolerance, and compare them with the FSIGT data identifi ed indices in the Results: Oil Red O staining showed lipid accumulation was substantially same individuals. We studied a cohort of 21 African immigrants 57% male, increased in the FTO overexpressed group, and little number of auto phagosomes age: 38.9 ± 3.4 y (mean± SE BMI: 28.4 ± 1.2 kg/m2) in whom the OGTT and was detected compared with control cells. LAMP mRNA level increased 1.5-fold, FSIGT were performed ~1 week apart. Parameter values for GK activity (KGK, whereas the corresponding protein increased 1.3-fold. The MAPLC3 mRNA and glycolysis K01, and lactate clearance K12, were all uniquely identifi ed for protein levels were obviously reduced by 30% in the FTO group. No differences both tests in all 21 subjects. Interestingly, GK activity (KGK) during the oral of Beclin-1 and ATG level were found among two groups.

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A465 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

Conclusions: FTO decreases the autophagy activity induced by high-fat 1808-P exposure, and its mechanism was associated with the regulation of several Involvement of Resistin-like Molecule β in the Development of Non- functional autophagy gene and protein. alcoholic Steatohepatitis Supported By: China Postdoctoral Science Foundation (2012M510850) AKIFUMI KUSHIYAMA, HIROFUMI OKUBO, YUSUKE NAKATSU, HIDEYUKI SAKODA, MIDORI FUJISHIRO, TOSHIAKI FUKUSHIMA, SHIN-ICIRO TAKAHASHI, 1806-P TOMOICHIRO ASANO, Tokyo, Japan, Hiroshima, Japan Exposure to a Maternal High Fat Milieu Alters Hepatic and White Resistin-like molecule β (RELMβ) reportedly has multiple functions Adipose Epigenome in Mice including local immune responses in the gut. In this study, it was revealed YOSHINORI SEKI, LYDA WILLIAMS, ALEX CHENG, XINGYI GUO, DEYOU ZHENG, that Kupffer cells in the liver, besides epithelial cells of the gut, express MASAKO SUZUKI, JOHN M. GREALLY, MAUREEN J. CHARRON, Bronx, NY RELMβ. Thus, the role of RELMβ in non-alcoholic steatohepatitis (NASH) Exposure to a maternal high fat (HF) diet is associated with increased was investigated. In mice, RELMβ expression levels in the colon and the incidence of metabolic syndrome later in life. However, the molecular basis numbers of RELMβ-positive Kupffer cells were both markedly increased by of this enhanced susceptibility is poorly understood. The HELP tagging methionine-choline defi cient (MCD) diet feeding. Increased RELMβ positive assay was used to measure genome-wide DNA methylation patterns in Kupffer cells were also observed in the livers of human patients with NASH. two tissues that display dysfunction in metabolic syndrome (liver and white In addition, importantly, RELMβ knock-out (KO) mice were highly resistant to adipose tissue (WAT)) of 5wk old male offspring exposed to a Control (C) or NASH development, based on liver histochemical fi ndings, serum parameters HF maternal diet. WT mice were fed a C (9.5% fat) or HF (35.5% fat) diet for and various mRNA and/or protein expression levels. Interestingly, the 2 wk before mating, throughout pregnancy and lactation. Offspring were proportion of lactic acid bacteria was higher in the gut microbiota of RELMβ weaned to a low fat (5.6% fat) diet (n=4/group). In liver, 10,633 differentially KO mice than in that of wild-type mice. Subsequently, to distinguish the methylated loci (DML) were identifi ed, the majority (65.2%) of which were roles of RELMβ secreted from the gut versus by Kupffer cells, bone marrow hypermethylated in HF. In WAT, 51.5% of the DMLs were hypermethylated transplantation was carried out between RELMβ KO and wild-type mice. in HF. Interestingly, only 10% of the DMLs found in both tissues mapped to These experiments revealed the requirement of both non-hematopoietic and promoters. Although the majority of the epigenetic changes associated with hematopoietic cell-derived RELMβ for full manifestation of NASH, while exposure to maternal HF diet are tissue specifi c, 300 loci displayed the same deletion of each one alone signifi cantly attenuated the development of methylation pattern in liver and WAT of HF offspring. Cardiovascular disease NASH with reduced serum LPS levels. was identifi ed as the top disorder with 33 genes having DMLs associated with These data strongly suggest the contribution of increases in RELMβ in the exposure to maternal HF. Next, eight 500kb-sized hot spots common to both gut and Kupffer cells to NASH development, raising the possibility of RELMβ tissues were identifi ed that displayed a high-density of DMRs correlating being a novel therapeutic target for NASH. with exposure to a maternal HF diet. Curiously, Quantitative Trait Loci associated with diabetes (chr18), atherosclerosis and bone mineral density 1809-P (chr4, chr1) mapped to these hot spots suggesting common chromosomal In Vivo Roles of Three Akt Substrates in the Liver on Insulin Sensi- regions may be vulnerable to epigenomic modifi cation upon exposure to a tivity and Plasma Glucose Level maternal HF diet. In summary, exposure to a maternal HF diet signifi cantly GOTA SAKAI, HIRAKU ONO, TAKASHI SUMITA, SHIGEHIRO KATAYAMA, TAKUYA altered DNA methylation patterns in liver and WAT of HF offspring. Although AWATA, Moroyama, Japan, Saitama, Japan most changes were tissue specifi c, DMLs in common to liver and WAT Insulin is reported to suppress endogenous glucose production (GP) via associated with metabolic syndrome suggest that epigenomic changes an activation of hepatic Akt. Three substrates of Akt, i.e. Foxo1, PGC1α and seen with exposure to a maternal HF diet may contribute to programmed GSK3β, have been reported to mediate this effect. All of them are inactivated development of metabolic disease later in life. when they are phosphorylated by Akt at their specifi c Ser/Thr sites. Foxo1 Supported By: ADA (1-13-CE-06); NIDDK and PGC1α upregulate gluconeogenic genes, so when they are inactivated by Akt, gluconeogenesis are suppressed. GSK3β inactivates glycogen 1807-P synthase, so when it is inactivated by Akt, glycogen synthesis is enhanced. Regular Exercise Corrects Fatty Liver Independent of Hepatic AMPK However, differential roles of these Akt substrates on suppression of HGP in High Fat Fed Mice and on plasma glucose level is not well clarifi ed. To elucidate these points, CLINTON M. HASENOUR, DERRICK E. RIDLEY, LARRY L. SWIFT, BENOIT VIOLLET, we made adenoviral vectors expressing mutants of these Akt substrates MARC FORETZ, DAVID H. WASSERMAN, Nashville, TN, Paris, France whose phosphorylation sites are mutated to alanine to make them Akt- Fatty liver affects ~30% of adults in developed countries. This is unresponsive constitutively-active forms (Foxo1-T24A-S256A-S319A: F, of concern as it associates with the sequalae of metabolic syndrome. PGC1α-S570A: P, and GSK3β-S9A: G), and intravenously injected them in Exercise is remarkably effective in reducing fatty liver. These studies test C57BL6 mice implanted with catheters in jugular vein and carotid artery. whether regular exercise (EX) requires hepatic AMPK to reduce fatty liver After a week of recovery we performed euglycemic-hyperinsulinemic clamp and correct associated metabolism. Male, 6wk old AMPKα1α2lox/lox (WT) study under unanesthetized and non-restrained condition. and AMPKα1α2lox/lox+Albcre (KO) C57Bl6 mice were given a HF diet; 6wks Hepatic expression of the mutants were 2-5 fold of endogenous levels. later, HF fed WT and KO mice were housed with operable or inoperable Glucose infusion rates (GIR) to maintain euglycemia were 5.5±2.0 mg/kg.min running wheels for 10wks. HF diet causes equal changes in body weight, in F, 25.3±8.8 in P, 24.5±9.4 in G, and 26.0±5.5 in control LacZ (Z) groups. GIR body composition, blood glucose, insulin, and leptin in WT and KO mice. was 8.2±1.8 when all three mutants were expressed altogether (T group). The EX response of these variables is unaffected by genotype. Livers were Signifi cant differences in GIR were observed between Z and F groups, characterized by metabolomic and standard biochemical analyses. HF diet and between Z and T groups. GP were 5.0±1.3 mg/kg.min in F, 3.8±5.5 in increases liver triglycerides (TGs) equally in WT and KO mice. EX ameliorates P, 0.5±2.6 in G, 10.2±2.3 in T, and 1.3±4.2 in Z groups. Trend of a difference fatty liver independent of hepatic AMPK, as liver TGs fall by >50% in WT in GP was observed between Z and T groups. Fasting plasma glucose were and KO mice. EX also increases liver TG polyunsaturation and decreases not signifi cantly different among groups. These results indicate that hepatic liver cholesterol esters, independent of AMPK. Liver phospholipids reduce Foxo1 is more important than other two Akt substrates for regulating insulin with AMPK deletion but are unaffected by EX. In contrast to TGs, EX sensitivity, and that there would be other pathways to maintain euglycemia Integrated POSTERS improvement of liver energy state requires AMPK. Acylcarnitines (≤C6) are in the fasted state. also elevated in KO mice, suggesting abnormal oxidative metabolism. This Physiology/Obesity effect of AMPK deletion is not corrected by EX. HF-diet increases the TCA 1810-P cycle intermediates citrate/cis-aconitate and reduces fumarate/malate in Metformin Alleviates Hepatosteatosis by Restoring SIRT1-mediated KO relative to WT mice. These changes are normalized by EX. These studies Autophagy Induction via an AMP-activated Protein Kinase-independent establish that 1) the hepatic TG lowering effect of EX is independent of Pathway AMPK, 2) the improved energy state with EX requires AMPK, and 3) changes YOUNG MI SONG, YONG-HO LEE, JI-WON KIM, HYE JIN YOON, EUN SEOK KANG, in TCA cycle intermediates, but not acylcarnitines, created by AMPK deletion BONG SOO CHA, HYUN CHUL LEE, BYUNG-WAN LEE, Seoul, Republic of Korea are normalized by EX in HF fed mice. The integrated control of metabolism Metformin is established to activate both AMPK and SIRT1. Furthermore, by regular exercise potently reduces fatty liver even in the absence of autophagy is induced by either AMPK-mTOR-ULK1 or SIRT1-FoxO pathways. hepatic AMPK. However, nucleotides and key metabolites related to energy We aimed to elucidate the mechanism by which metformin alleviates metabolism may be critically reliant on exercise-AMPK interactions. hepatosteatosis by examining the molecular interplay among SIRT1, AMPK, Supported By: U24DK59637, R37DK50277 and autophagy.

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A466 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

Ob/ob mice were divided into three groups: one with ad libitum feeding 1812-P of a standard chow diet, one with metformin injection (300 mg/kg, IP) and Fasting Hyperglucagonemia in Patients with Nonalcoholic Fatty one with 3 g/day caloric restriction (CR) for 4 weeks. Primary hepatocytes or Liver Disease With and Without Type 2 Diabetes HepG2 cells were treated with oleic acid (OA) plus high glucose (HG) in the ANDERS E. JUNKER, LISE L. GLUUD, JENS J. HOLST, FILIP K. KNOP, TINA VILS- absence or presence of metformin. BØLL, Hellerup, Denmark, Copenhagen, Denmark Both CR and metformin signifi cantly improved body weight, glucose homeo- Non-alcoholic fatty liver disease (NAFLD) and hyperglucagonemia seems stasis and hepatic steatosis, in ob/ob mice. Furthermore, CR and metformin to infl uence glucose metabolism and both features are associated with both up-regulated SIRT1 expression and stimulated autophagy induction in type 2 diabetes (T2D). We investigated glucagon responses during oral and vivo. Metformin also prevented OA with HG-induced suppression of both SIRT1 intravenous glucose administration in biopsy-verifi ed NAFLD patients with expression and SIRT1-dependent activation of autophagy machinery, thereby and without T2D. alleviating intracellular lipid accumulation in vitro. Interestingly, metformin We performed a 50 gram-oral glucose tolerance test (OGTT), and iso- treatment up-regulated SIRT1 expression and activated AMPK even after glycemic intravenous glucose infusion (IIGI) in participants with I) normal siRNA-mediated knockdown of AMPKα1/2 and SIRT1, respectively. glucose tolerance (NGT) and no liver disease (N=10; age (median±interquartile Taken together, these suggest that metformin alleviates hepatosteatosis 2 range): 58±17 years; BMI: 29±1 kg/m : HbA1c: 34±6 mmol/mol (5.5±0.2%)), II) through AMPK-independent, SIRT1-mediated effects on autophagy machinery. 2 NGT+NAFLD (N=9; age: 54±22 years; BMI: 28±7 kg/m : HbA1c: 33±4 mmol/ mol (5.2±0.3%)), III) T2D and no liver disease (N=8; age: 59±18 years: BMI: 2 27±3 kg/m : HbA1c: 45±9 mmol/mol (6.2±1.1%)), or IV) T2D+NAFLD (N=8; age: 2 65±3 years; BMI: 30±4 kg/m ; HbA1c: 53±19 mmol/mol (7.0±1.4%)). In all groups, isoglycemia was achieved during oral and intravenous glucose administrations. Fasting plasma glucagon was signifi cantly elevated in participants with NAFLD and NGT (7.0±2.7 pM) and T2D (6.7±2.3 pM), compared to participants without NAFLD and NGT (2.8±4.0 pM, P<0.001) or T2D (4.5±2.9 pM, P<0.05). Larger glucagon responses (ΔAUC0-50 min) during OGTT vs. IIGI were seen in both groups with T2D (no liver disease: 40±78 (OGTT) vs. -63±43 pM×50min (IIGI), P<0.05; NAFLD: 30±99 (OGTT) vs. -38±44 pM×50min (IIGI), P<0.05) whereas no differences in glucagon responses to OGTT and IIGI were seen in NGT subjects (no liver disease: -20±26 (OGTT) vs. -88±60 pM×50min (IIGI), P=NS; NAFLD: -40 ± 75 (OGTT) vs. -90±70 pM×50min (IIGI), P=NS). Fasting hyperglucagonemia seems to be a pathophysiological trait of NAFLD independently of concomitant T2D. In contrast, patients with T2D have an inappropriate glucagon response to oral glucose independently of NAFLD.

1813-P Expression of Sfrp5 and Wnt5a in Liver Associates with Infl amma- tion and Hyperlipidemia in Humans MAREN CARSTENSEN, CHRYSI KOLIAKI, PETER NOWOTNY, KARIN ROEHRIG, FRANK JANKOWIAK, MARKUS KRAUSCH, WOLFRAM T. KNOEFEL, MATTHIAS Supported By: Brain Korea 21 PLUS Project for Medical Science, Yonsei University SCHLEN SAK, JULIA SZENDROEDI, CHRISTIAN HERDER, MICHAEL RODEN, Düssel - dorf, Germany 1811-P Secreted frizzled related-protein 5 (Sfrp5) is a new anti-infl ammatory Liver Specifi c G0/G1 Switch Gene 2 (G0S2) Overexpression Exacer- protein, which decreases insulin sensitivity in human adipocytes, but has no bates Hepatic Insulin Resistance by Aggravating Hepatic Steatosis effect on infl ammation and insulin signaling in skeletal muscle. The relevance but Ameliorates Hepatic Fibrosis of the hepatic expression of Sfrp5 for cardiometabolic risk factors is yet YOSHIYUKI SUGAYA, HIROAKI SATOH, TSUYOSHI WATANABE, Fukushima, unknown. Here, we analyzed the association of liver mRNA expression of Japan Sfrp5 and its antagonist Wnt5a with metabolic factors in well-characterized Hepatic steatosis is strongly associated with insulin resistance. Recently patients undergoing abdominal surgery (NCT01477957). it has been reported that G0/G1 switch gene 2 (G0S2) inhibited the lipolysis Eighteen obese normoglycemic humans undergoing bariatric surgery activity of adipose triglyceride lipase. Moreover, we confi rmed that G0S2 (OB; age 42.5±9.1 y, BMI 49.6±9.1 kg/m2) and 8 lean humans undergoing protein content was increased in the livers of high fat diet (HFD)-fed rats. surgery for non-malignant abdominal disease (CON; age 44.1±10.6 y, BMI However, the precise physiological role of hepatic G0S2 is still unknown. In 25.4±2.7 kg/m2) underwent euglycemic-hyperinsulinemic clamps with [6,6- the current studies, we investigated the effect of hepatic G0S2 on insulin 2H2]glucose and intra-operative liver biopsies. Relative mRNA expression of sensitivity in HFD-fed male Wistar rats by overexpressing G0S2 proteins Sfrp5 and Wnt5a were quantifi ed by real-time PCR. using an adenovirus (Ad) encoding mouse G0S2. Male Wistar rats were OB were more insulin-resistant in peripheral tissues (M-value 3.6±1.4 vs. fed with 60% HFD for a total of 4 weeks. After 3-week feeding, the rats 7.0±2.1 mg.kg-1.min-1, p<0.05), but had similar hepatic insulin sensitivity as were injected with control Ad-GFP or Ad-G0S2. On day 7 post injection, CON. Liver Sfrp5 expression tended to be lower, whereas Wnt5a expression glucose tolerance test and euglycemic-hyperinsulinemic clamp studies were was higher in OB than CON (0.13±0.02 vs. 0.05±0.01 a.u., p=0.03). Sfrp5 performed after an 8-hour fast. There were no signifi cant changes in the associated negatively with C-reactive protein (hs-CRP; r=-0.46, p=0.03) and body weight and fasting glucose levels between Ad-GFP and Ad-G0S2 rats. total cholesterol (r=-0.52, p=0.01), while Wnt5a associated positively with However, after the glucose load, the glucose levels were signifi cantly higher hs-CRP (r=0.45, p=0.03) and negatively with adiponectin (r=-0.43, p=0.03). Integrated in the Ad-G0S2 rats at 15 and 30 min. During the clamp studies, the glucose The ratio Sfrp5/Wnt5a was lower in OB than CON (43±11 vs. 110±37, POSTERS infusion rate required to maintain euglycemia was signifi cantly decreased p=0.04) and correlated negatively with hs-CRP (r=-0.64, p=0.0009) and by 16% in Ad-G0S2 rats. Clamp hepatic glucose output was signifi cantly LDL-cholesterol (r=-0.43, p=0.04), and positively with adiponectin (r=0.44, Physiology/Obesity increased by 31% in Ad-G0S2 rats, but there were no signifi cant changes in p=0.03). Sfrp5, Wnt5a and their ratio were not associated with BMI, liver fat insulin-stimulated glucose disposal rate between two groups. Furthermore, content, hepatic or peripheral insulin sensitivity. the Oil Red O staining revealed that overexpression of G0S2 signifi cantly In conclusion, the ratio of Sfrp5 to Wnt5a mRNA levels is lower in the increased lipid accumulation in the liver. However, Masson trichrome liver of obese humans and associates with increased infl ammation and staining revealed that overexpression of G0S2 signifi cantly ameliorated hyperlipidemia, but not with insulin resistance or hepatic steatosis. fi brosis in the liver. Consistent with histological data, expression of TGF-β Supported By: German Center for Diabetes Research; Helmholtz Alliance ICEMED; and Smad2 were signifi cantly decreased in the livers of Ad-G0S2 rats. Schmutzler-Skröder Foundation In conclusion, overexpression of hepatic G0S2 protein exacerbates hepatic insulin resistance by the aggravation of hepatic steatosis but ameliorates hepatic fi brosis.

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A467 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1814-P 1816-P Inhibition of FoxO1 by AS1842856 Decreases Hepatic Glucose ACOT1 Regulates Hepatic Lipid Traffi cking and Signaling Production and Lowers Hepatic and Circulating Lipids MALLORY P. FRANKLIN, AISHWARYA SATHYANARAYAN, MARA T. MASHEK, GREGORY TESZ, MATTHEW F. GORGOGLIONE, PHILIP A. CARPINO, NICHOLAS DOUGLAS G. MASHEK, St. Paul, MN B. VERA, DEREK M. ERION, JAMES F. SMITH, PAUL A. AMOR, ELSIA YOO, Acyl-CoA thioesterases (ACOTs) catalyze the hydrolysis of acyl-CoAs to WILLIAM P. ESLER, TIMOTHY P. ROLPH, JEFFREY A. PFEFFERKORN, Cambridge, free fatty acids and CoA. Surprisingly, little is known about the physiological MA, Greensburg, PA role of this family of proteins. In the liver, ACOT1 is the major cytosolic ACOT Elevated hepatic glucose production contributes to hyperglycemia in type isoform and is highly induced by peroxisome proliferator-activated receptor 2 diabetes mellitus (T2DM). While both glycogenolysis and gluconeogenesis alpha (PPAR-α), an important regulator of hepatic energy metabolism that drive hepatic glucose production, the latter process is increased in people mediates the metabolic adaptation to fasting. ACOTs infl uence long-chain with T2DM. The Forkhead Box class O1 (FoxO1) transcription factor regulates free fatty acids and acyl-CoAs pools, both of which infl uence lipid metabolism the inducible gluconeogenic enzymes, glucose 6 phosphatase (G6Pase) and and fatty acid-mediated cell signaling. Since the physiological role of ACOT1 phosphoenolpyruvate carboxykinase (PEPCK). In T2DM mouse models, has not been studied, the objective of this research was to test the effects both hepatic deletion of FoxO1 and antisense oligonucleotide suppression of hepatic ACOT1 on lipid metabolism and signaling. To test our objectives, reduced gluconeogenesis and improved glycemia. However, FoxO1 deletion we injected male C57/Bl6 mice with adenovirus harboring shRNA targeted in some of these studies, but not all, increased hepatic lipids; while antisense to ACOT1 or a scrambled control. One week post-injection, mice were fasted suppression reduced hepatic lipids. To clarify the role of FoxO1 in lipid overnight and sacrifi ced. There were no differences in food intake, body metabolism and to better understand the consequences of pharmacological weight or tissue weights between treatment groups. Compared to control FoxO1 inhibition, we utilized the recently published specifi c FoxO1 inhibitor mice, ACOT1 knockdown increased serum β-hydroxybutyrate, a marker of AS1842856, a compound which binds only dephosphorylated, active, FoxO1. fatty acid oxidation. Surprisingly, genes involved in mitochondrial biogenesis Treatment of primary human hepatocytes with AS1842856 decreased FoxO1 and mitochondrial DNA copy number were decreased in response to ACOT1 target gene expression (G6Pase, PEPCK, IRS-1 and IGFBP1) and glucose knockdown. Consistent with the in vivo data, radioisotope labeling studies production. To determine if the mechanism of inhibition was specifi c for using primary hepatocytes showed that ACOT1 knockdown signifi cantly FoxO1, HepG2 cells were transfected with constitutively active FoxO1 (ADA- increased oleate oxidation. Additionally, ACOT1 knockdown increased FoxO1) and treated +/- AS1842856. AS1842856 specifi cally suppressed only the turnover of intracellular triglyceride and the subsequent channeling increased FoxO1 target gene expression by ADA-FoxO1, and not non-FoxO1 of hydrolyzed fatty acids towards oxidation. ACOT1 had no effect on the target genes. To better understand the effects of acute FoxO1 inhibition in metabolism of de novo synthesized fatty acids or the incorporation of vivo, diabetic db/db mice were treated with AS1842856. Treatment with exogenous oleate into complex lipids. In summary, these data suggest that AS1842856 lowered fasting glucose, reduced pyruvate stimulated glucose ACOT1 plays an important role in hepatic triglyceride turnover, in partitioning excursion, decreased hepatic gluconeogenic gene expression and reduced long-chain fatty acids/acyl-CoAs to specifi c metabolic pathways and in hepatic and plasma lipids. Reductions in hepatic gluconeogenesis and infl uencing cell signaling pathways that govern mitochondrial biogenesis. dyslipidemia warrant further evaluation of FoxO1 as a target for improving metabolic homeostasis. 1817-P Sugary Water Consumption Leads to Adipose, Liver, and Metabolic 1815-P Dysfunction in a Western Diet-induced Model of NAFLD Benefi ts of Docosahexanoic Acid Tissue Enrichment in Nonalcoholic YUWEN LUO, CHRISTINE M. BURRINGTON, JIAN ZHANG, EMILY C. GRAFF, Fatty Liver Disease: Results from a Randomised, Double-Blind, ROBERT L. JUDD, MICHAEL W. GREENE, Auburn, AL, Cooperstown, NY Placebo-Controlled Study Testing the Effects of Omega-3 Fatty Acid Nonalcoholic steatohepatitis (NASH), a more severe form of nonalcoholic Treatment fatty liver disease (NAFLD) that is associated with obesity, insulin resistance, CHRISTOPHER D. BYRNE, ELEONORA SCORLETTI, LOKE BHATIA, KEITH MC- and diabetes, is characterized by steatosis, infl ammation, oxidative stress, CORMICK, GERALDINE CLOUGH, KATE NASH, LEANNE HODSON, PHILIP CALDER, apoptosis, and fi brosis. It has been postulated that excessive sugary drink ON BEHALF OF THE WELCOME STUDY INVESTIGATORS, Southampton, United consumption may contribute to the development and severity of NAFLD. Kingdom, Oxford, United Kingdom However, its role in adipose tissue expansion and dysfunction is not known. Non-alcoholic fatty liver disease (NAFLD) is a risk factor for type 2 To better understand the role of consumption of liquid sugar in high fat diabetes, cardiovascular and chronic liver disease, but to date there is no diet-induced NAFLD progression, we investigated metabolic and histologic licensed treatment. In a randomised double blind placebo controlled trial parameters and gene expression from hepatic and adipose tissues in mice we tested if 15-18 months Omacor (4 g/day) [containing eicosapentaenoic fed a low or high fat western diet (HFWD) without or with liquid sugar acid (EPA) and docosahexaenoic acid (DHA)] increased EPA and DHA tissue [fructose and sucrose (F/S)] at two time points (2 and 12 week). HFWD+F/S enrichment, decreased liver fat (LF%), and improved two validated liver impaired the storage capacity of epididymal white adipose tissue (eWAT) fi brosis biomarker scores. and exacerbated HFWD-induced glucose intolerance and insulin resistance. 103 patients with NAFLD were randomised to Omacor (n=51) or placebo Consistent with these results, HWFD+F/S fed mice developed more profound (n=52). At baseline and end of study, we measured erythrocyte percentage eWAT infl ammation characterized by macrophage infi ltration, a dramatic DHA and EPA enrichment (a validated proxy for hepatic enrichment), by increase in crown-like structures, and upregulated proinfl ammatory gene gas chromatography; mean LF% (three liver zones) by magnetic resonance expression. Hepatic triglyceride, plasma alanine aminotransferase, and spectroscopy (MRS); and liver fi brosis scores. In intention to treat (ITT) normalized liver weight were signifi cantly increased only in HFWD+F/S and per protocol (PP) analyses, the decrease in LF%, and improvement in fed mice. Hepatic oxidative stress, assessed by superoxide dismutase liver fi brosis scores with Omacor treatment (ITT analyses) and tissue DHA activity, 4-hydroxynonenal blotting, NADPH oxidase activity, and gene and EPA enrichment (PP analyses), were tested, by multivariable linear expression was highest in HFWD+F/S fed mice. HFWD+F/S also resulted in regression modelling. increased hepatic fi brosis and elevated collagen I, collagen III and TGFβ gene Mean, (95%CIs) baseline and end of study LF% was 21.7 (13.7,32.3) and expression. Our results highlight the effect of sugary water consumption in 19.7 (11.3,28.0) respectively for the placebo group; and 23.0 (12.0,47.5) and adipose tissue dysfunction and NAFLD progression. Integrated

POSTERS 16.3 (9.0,30.7) respectively for the Omacor group, but the adjusted difference in change from baseline LF% between groups was -1.7 (-6.3, 3.0), p=0.5

Physiology/Obesity 1818-P (ITT analyses). In PP analyses, percentage erythrocyte DHA enrichment Branched-Chain Amino Acids Alter Mitochondrial Fat Metabolism was independently associated with decrease in LF% (Beta -0.47; SE 0.74; in Mice with Nonalcoholic Fatty Liver Disease p<0.0001), adjusting for age, sex, baseline LF%, a biomarker of apoptosis, NISHANTH E. SUNNY, ADRIANE K. HINES, SRILAXMI KALAVALAPALLI, TIMOTHY and change in weight during the study. Further adjustment for changes J. GARRETT, JUSTIN T. MATHEW, KENNETH CUSI, Gainesville, FL in physical activity and diet did not affect the results. No changes in the Elevated levels of branched chain amino acids (BCAAs; leucine, isoleucine fi brosis biomarker scores were observed in ITT or PP analyses. and valine) are implicated in the etiology of insulin resistance, obesity and These data suggest that high tissue DHA enrichment is required for onset of type II diabetes. Over 70% of obese humans have nonalcoholic achieving a decrease in LF% with Omacor treatment. fatty liver disease (NAFLD), characterized by high liver triglycerides (> 5%). Supported By: NIHR; Diabetes UK Mitochondrial dysfunction is a central feature of NAFLD. Our hypothesis was that BCAAs impact mitochondria by activating fat oxidation through hepatic tri-carboxylic acid (TCA) cycle during fasting. Mice (C57/BL6) were

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A468 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE fed a high fructose- high trans-fat diet (TFD) for 8 weeks to induce NAFLD. and reversed markers of ER dysfunction and lipotoxicity. To determine if Following an overnight fast, either saline or a mixture of BCAAs were infused calcium effl ux from the ER was required to induce the downstream effects into the control and TFD fed animals (n= 6/ group) intravenously for 4 hours. of palmitate, we co-treated hepatic cells with both palmitate and the Infusion of BCAAs increased plasma concentrations twofold. Following this intracellular calcium chelator BAPTA. BAPTA suppressed ROS accumulation acute BCAA challenge we determined a) degradation of BCAAs in the liver and caspase activation in the presence of palmitate. Next, we applied 13C 13 13 from the incorporation of [ C6]leucine into [ C5]isovalerylcarnitine by mass metabolic fl ux analysis (MFA) using 13C-glutamine as an isotopic tracer to spectrometry (MS) and b) fasting hepatic TCA cycle activity using nuclear identify metabolic pathway alterations associated with palmitate treatment magnetic resonance based 13C isotopomer analysis. Degradation of BCAAs and BAPTA rescue. We found that palmitate enhanced citric acid cycle was impaired during NAFLD by 1.5 fold compared to the controls. Surprisingly, (CAC) fl ux, mitochondrial oxygen consumption, and glutamine anaplerosis in long chain acylcarnitines (nmoles g liver protein-1) were elevated in the liver H4IIEC3 cells. Co-treating with BAPTA restored most mitochondrial fl uxes to of control animals following BCAA challenge (3.9± 0.75 vs. 5.9± 0.93 for basal levels and suppressed both ROS and apoptosis. C14; 21± 3.1 vs. 29± 4.2 for C16). Hepatic TCA cycle activity measured as Supported By: NSF (CBET-0955251) anaplerotic fl ux and pyruvate cycling in relation to citrate synthase was also induced 1.5 fold (P < 0.05) by BCAAs in the control animals. However 1821-P BCAAs failed to induce hepatic TCA cycle in animals with NAFLD. Our data Antioxidant Activates AMPK and Ameliorates Nonalcoholic Fatty suggest that BCAAs play a signifi cant role in up-regulation of mitochondrial Liver Disease in OLETF Rats fat metabolism during fasting. Impairment of BCAAs mediated induction YONGSOO PARK, HYUNOK KIM, Seoul, Republic of Korea of hepatic TCA cycle during NAFLD could contribute to mitochondrial Although the pathogenesis of non-alcoholic fatty liver disease (NAFLD) dysfunction and triglyceride accumulation in the liver. has been obscure, oxidative stress might be one of the crucial causes for progression. In this study, we tried to investigate whether it is possible to 1819-P ameliorate NAFLD by regulation of oxidative stress in OLETF rats. Applying ATF4 Defi ciency Protects against High Fructose-induced Hypertri- protein transduction technology, we made Tat-fusion antioxidants, Tat-MT glyceridemia in Mice (metallothionein) and Tat-SOD (superoxide dismutase) and found that Tat- TING ZHANG, GUOZHI XIAO, SHIBING YU, SOJIN LEE, VIRTU CALABUIG- MT and Tat-SOD in combination are effi ciently transduced into HepG2 cells NAVARRO, HENRY DONG, Pittsburgh, PA, Chicago, IL across the lipid membrane. The protective effect of antioxidant in progression Hypertriglyceridemia is the most common lipid disorder in obesity and type of NAFLD was studied in vivo in OLETF rats. A single intraperitoneal injection 2 diabetes. It results from increased production and/or decreased clearance of Tat-fusion proteins resulted in the delivery of these antioxidants to the of triglyceride-rich lipoproteins. To better understand the pathophysiology liver. Decreased hepatic steatosis, decreased apoptosis and reduced of hypertriglyceridemia, we studied hepatic regulation of triglyceride infl ammation resulted from ER stress amelioration along with reduction of metabolism by the activating transcription factor 4 (ATF4), a member of the ROS in the liver were induced by the prolonged treatment of Tat-MT and Tat- basic leucine zipper-containing protein subfamily, also known as cAMP- SOD in combination for 16 weeks, which ultimately delayed the development response element-binding protein 2. ATF4, characterized as a key factor of of NASH. Antioxidant signifi cantly reduced glucolipotoxic injuries, and bone homeostasis, contributes to the regulation of energy homeostasis. apoptosis and infl ammtory marker expression along with the decrease in the Nevertheless it remains unknown as to how ATF4 impacts lipid metabolism level of ROS in HepG2 cells. Moreover, antioxidants decreased lipogenesis and whether ATF4 contributes to the pathogenesis of hypertriglyceridemia. gene and increased β-oxidation gene transcription. Antioxidant treatment We determined the effect of ATF4 on hepatic lipid metabolism in Atf4-/- increased AMPK phosphorylation and Sirt-1 expression along with the mice fed regular chow or provided with free access to fructose drinking lipogenic transcription factor, SREBP-1c. Antioxidants reduced hepatic water. ATF4 depletion preferentially attenuated hepatic lipogenesis without steatosis in OLETF rats and activated AMPK and Sirt-1 in the liver. These affecting hepatic triglyceride production and fatty acid oxidation. This effect results suggest the role of AMPK activation in the protective infl uence of prevented excessive fat accumulation in the liver of Atf4-/- mice, when antioxidant on NAFLD progression. The ameliorative effects of Tat-MT and compared with wild-type littermates. To gain insight into the underlying Tat-SOD in combination on NAFLD progression might be associated with this mechanism, we showed that ATF4 depletion resulted in a signifi cant reduction inhibition effect on ROS signaling pathway. in hepatic expression of peroxisome proliferator-activated receptor-gamma, a nuclear receptor that acts to promote lipogenesis in the liver. This effect was accompanied by a signifi cant reduction in hepatic expression of sterol INTEGRATED PHYSIOLOGY—MACRONUTRIENT regulatory element binding protein 1c (SREBP-1c), acetyl-CoA carboxylase, METABOLISM AND FOOD INTAKE and fatty-acid synthase, three key functions in the lipogenic pathway in Atf4-/- mice. Of particular signifi cance, we found that Atf4-/- mice, as opposed to wild-type littermates, were protected against the development Guided Audio Tour: Food Intake and Fuel Metabolism (Posters: 1822-P to of steatosis and hypertriglyceridemia in response to high fructose feeding. 1827-P), see page 15. These data demonstrate that ATF4 plays a critical role in regulating hepatic lipid metabolism in response to nutritional cues. & 1822-P Acute Effects of the Glucokinase Activator (GKA), AZD1656, on the 1820-P Turnover and Metabolic Fate of Plasma Glucose, and Free Fatty Acid ER Calcium Effl ux Promotes Mitochondrial Dysfunction and Hepatic (FFA) in Obese Zucker Rats Lipotoxicity in Response to Palmitate Overload NICHOLAS D. OAKES, THERESE HAGSTEDT, ANN KJELLSTEDT, KRISTINA WALLEN- ROBERT A. EGNATCHIK, ALEXANDRA K. LEAMY, DAVID A. JACOBSON, MASA- IUS, ANDREW D. CHARLES, MASAKAZU SHIOTA, Mölndal, Sweden, Nashville, TN KAZU SHIOTA, JAMEY D. YOUNG, Nashville, TN Effects of AZD1656 on in vivo glucose and free fatty acid (FFA) metabolism Lipotoxicity resulting from free fatty acid overload has been implicated were investigated in obese insulin resistant Zucker rats. Fasted animals in a variety of disorders including type-2 diabetes and non-alcoholic fatty were intravenously infused with either AZD1656 (0.02, 0.2 or 2.0 µmol/kg/ liver disease (NAFLD). Incubation of hepatic cells with saturated fatty acids min) or vehicle. Blood glucose was maintained at pre-treatment (basal) levels Integrated (SFAs) induces aberrant mitochondrial metabolism, accumulation of reactive by intravenous infusion of glucose. Initial studies using [3-3H]glucose and POSTERS oxygen species (ROS), and apoptosis. However, a consensus mechanism [U-14C]glucose (14C-G) tracers confi rmed that AZD1656 engaged the target linking SFA-induced metabolic alterations to hepatocyte dysfunction and mechanism (immuno-histochemistry showed translocation of glucokinase Physiology/Obesity cell death has been diffi cult to establish. Our central hypothesis is that SFA from the nuclear to cytosolic compartments in hepatocytes) and dose overexposure leads to increased saturation of ER phospholipids, which alters dependently: increased the glucose infusion rate (GIR) needed to maintain membrane stability and induces ER calcium effl ux. Re-uptake of calcium by basal glycemia; increased the whole body glucose disposal rate (Rd); mitochondria subsequently causes metabolic dysfunction, ROS accumulation, increased incorporation of glucose into glycogen in both liver and skeletal and apoptosis. To test this, we fi rst measured 3H-palmitate incorporation muscle; and suppressed hepatic glucose output (HGO). Effects of AZD1656 into various intracellular lipid pools in both H4IIEC3 rat hepatoma cells (given at a dose of 2µmol/kg/min) on the metabolic fate of glucose was 14 14 and primary rat hepatocytes. We found that exogenous palmitate was further investigated using plasma C-G and exhaled CO2 kinetics using preferentially incorporated into phospholipid (PL) pools, and increases in a simple compartmental model. FFA turnover and metabolic fate were PL saturation correlated with ER membrane dilatation and decreases in simultaneously evaluated using [9,10-3H]palmitate and its conversion to 3 ER calcium stores. Co-treatment with oleate prevented PL oversaturation H2O. Compared to vehicle controls the GKA: increased plasma lactate,

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A469 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE

increased the rate of non-oxidative glucose disposal (by 62%, P<0.001), (P<0.05) that was also abolished in HFS rats (P<0.05). High fat feeding that increased whole body glucose oxidation (by 67%, P<0. 01) and stimulated induces IR abolishes the daily peak in circadian DA neuronal activity at the hepatic de novo lipogenesis. AZD1656 also increased hepatic malonyl- SCN area, a neuronal function previously demonstrated to be integral to CoA levels, an important physiologic inhibitor of β-oxidation of fatty acids maintenance of insulin sensitivity. (by 100%, P<0.05), increased incorporation of FFA into lipid and tended to lower FFA oxidation. In conclusion, these data suggest that AZ1656 & 1825-P acutely improves glucose control via increased fl ux of glucose into all Effect of High Protein (HP) vs. High Carbohydrate (HC) Diets on major metabolic pathways of glucose disposal including glycolysis, lactate Satiety and Cardiovascular Risk Factors (CVRF) in Obese Subjects production, oxidation and de novo lipogenesis. At the same time FFA tends FRANKIE B. STENTZ, ABBAS E. KITABCHI, Memphis, TN to be diverted away from oxidation to storage. Our study of effects of dietary macronutrients on metabolic parameters show that a HP diet(30% protein,30% fat, 40%CHO) compared to a HC diet & 1823-P (15%protein,30%fat, 55%CHO) results in greater reduction in oxidative Intermittent, but Not Continuous, Nicotinic Acid (NiAc) Infusion stress, CVRF and insulin resistance in the HP group than the HC group after 6 Preserves Plasma Free Fatty Acid (FFA) Lowering and Ameliorates months(mo). Since the incretin GLP-1 has an important role in insulin sensitivity Insulin Resistance in Obese Zucker Rats we studied the effect of the HP and HC diets on GLP-1. Studies have shown TOBIAS KROON, PIA THALÉN, ANN KJELLSTEDT, JOHAN GABRIELSSON, that HP diets can potentially induce satiety, therefore, we studied HP and HC NICHOLAS D. OAKES, Uppsala, Sweden, Mölndal, Sweden diet effects on ghrelin levels. Additionally, since CVRF decreased more in the Acute NiAc administration results in rapid reduction of FFA concentrations. HP than the HC diet, we determined if the B-Type Natriuretic Peptide(BNP) However, chronic treatment is associated with development of tachyphylaxis released from the heart was affected by either diet. resulting in complete return of FFA to pre treatment levels. The aim of this 24 obese women were randomized to HP or HC diets. All food was study was to determine whether an intermittent dosing regimen could avoid provided for the 6 mo study. Ghrelin, GLP-1 and NT-proBNP(BNP) levels tachyphylaxis and result in reduction of average FFA concentrations and were determined with MTT at baseline and after 6 mo on HP and HC diets. thereby reverse lipid-overload induced insulin resistance. Results are shown in the Table. HP ghrelin results demonstrate that HP diet NiAc-induced FFA lowering was assessed in male Sprague Dawley (Lean) can induce satiety and is more effective than the HC diet. HP diet had a and obese Zucker rats (Obese) during an acute experiment following either greater increase in GLP-1 than the HC diet. The BNP decrease in both groups continuous (24 h/day) or intermittent (12 h/day) NiAc treatment for 5 days, demonstrates the improvement in heart tissue with the HP diet having a achieved using implantable, programmable mini-pumps. We found that in greater effect than the HC diet. Although weight loss was similar (9.8% in the intermittent NiAc groups, lowering of average FFA levels achieved by HP group and 9.3% in HC group) this study demonstrates that the HP diet has plasma NiAc concentrations ~1 µM, was preserved in Lean (-0.30±0.064 vs. additional health benefi ts compared to the HC diet. -0.26±0.019 mM in NiAc naïve) and Obese (-0.41±0.057 vs. -0.55±0.021 mM in Effect of HP and HC Diets on Ghrelin GLP-1 and BNP. NiAc naïve). Extending the intermittent protocol to 11 days in Obese resulted in no signifi cant loss of FFA lowering effi cacy (-0.34±0.086 mM) vs. day 5. HP Diet (n=12) HC Diet (n=12) By contrast, FFA lowering was completely lost in both groups when NiAc Parameters Baseline 6 months P* Baseline 6 months P* P** was continuously infused. In separate experiments, effects of continuous GLP-1 MTT (pmol/l/min) 7860 ± 31 8970 ±37 0.001 6570 ± 29 7006 ±35 0.05 0.001 vs. intermittent NiAc infusion on insulin sensitivity in obese Zucker rats Ghrelin MTT (ng/ml/min) 342 ± 26 88.5 ± 12 0.005 516 ± 22 480 ± 17 0.3 0.001 was assessed using hyperinsulinemic-isoglycemic clamps. We found that, whole body insulin sensitivity (steady state glucose infusion rate needed to NT-proBNP (pmol/l) 492 ± 23 359 ± 18 0.01 486 ± 27 390 ± 21 0.04 0.03 maintain basal glucose levels) was dramatically increased in intermittently * Indicates Wilcoxon Signed Rank Test. vs. continuously NiAc infused animals (by 340%, P<0.01). ** Indicates Wilcoxon Rank Sum Test for 6 mo HP vs. HC. In conclusion, an intermittent NiAc dosing strategy succeeded in retaining Supported By: ADA (1-09-CR-32) FFA lowering and most importantly substantially improved insulin sensitivity in obese Zucker rats. These data suggest that optimized anti-lipolytic & 1826-P treatment regimes might provide a means to reverse lipid overload and its Metabolic Adaptation in Indians Emigrated to USA—Changes in harmful metabolic consequences. DNA Methylation and Amino Acid Metabolites in Comparison with Northern-European Americans & 1824-P DWAIPAYAN BHARADWAJ, TUMPA DUTTA, ANIL GIRI, MOHD NAZIR KHAN, High-Fat Feeding Abolishes the Insulin-Sensitizing Peak in NIKHIL TANDON, K. SREEKUMARAN NAIR, Rochester, MN Circadian Dopamine Activity at the Biological Clock We determined the impact of environmental factors on high incidence of SHUQIN LUO, YAHONG ZHANG, MICHAEL EZROKHI, YELENA TRUBITSYNA, T2D among migrant Indians.To determine the impact of environmental factors ANTHONY H. CINCOTTA, Tiverton, RI on DNA methylation of individuals having same genetic background but The peak in circadian dopaminergic (DA) activity at the area of the different environmental infl uence and with same environment but different biological clock, suprachiasmatic nucleus (SCN), at daily locomotor activity genetic background we performed whole genome DNA methylation study (LA) onset is a major stimulus for maintenance of insulin sensitivity. Loss of in 15 non-diabetic (ND) Indians residing in India, 13 T2D and 13 matched ND this peak either by natural seasonal change in seasonal animals or by SCN of migrant Indians living in USA, 12 T2D and 15 matched ND of Americans DA neurotoxic lesion to insulin sensitive animals induces insulin resistance of Northern-European (NE) origin. Data exhibited hyper-methylation in 287 (IR). Central dosing of dopamine agonist to re-elevate this diminished of 442 CpGs and hypo-methylation in 155 CpGs in ND Indians to migrant endogenous peak in DA activity ameliorates the IR. However, the impact of Indians to NE American. Pathway analysis showed enrichment of signaling a “westernized” IR-inducing, high fat (HF) diet upon this circadian system, pathways of phosphatidyl inositol phosphate, interleukin, cytokine including a major DA projection to this SCN area from the supramammillary mediated and cancer pathways. The methylation profi ling of ND migrant nucleus (SuMN), has never been investigated and was the focus of this Indians were similar to NE and different from resident Indians highlighting study. Rats were fed ad lib either a HF (60% saturated fat) or regular chow the importance of environmental factors in regulating the DNA methylation. Integrated POSTERS (RC) diet for 6 weeks. High fat sensitive (HFS) rats (those gaining 25% more Hierarchical clustering analysis showed a clear segregation of Indians from weight than RC rats, N=10) and RC rats (N=10) were then subjected to a migrant Indians, whereas NE Americans and migrants Indians clustered Physiology/Obesity glucose tolerance test (GTT) followed in 14 days by in vivo SCN microdialysis together. These results demonstrate that DNA methylation changes due for 24 hours. Dialysate samples were analyzed by HPLC for dopamine to environmental factors. Principal Component Analysis of targeted plasma metabolites: homovanillic acid and 3,4-Dihydroxyphenylacetic acid. Brains amino acid metabolites measurements by LC MS/MS approach showed were subsequently collected at 4 and 16 hours after light onset to quantify distinct intra and inter group separation between Indians and NE Americans c-Fos and tyrosine hydroxylase (TH) double immunopositive neurons in and between T2D and ND. The metabolites a-amino adipic acid, cysteine SuMN. HFS rats had elevated GTT plasma glucose (30%, P<0.05) and insulin and glutamine were signifi cantly altered in Indian and NE American T2D AUC (53%, P=0.005) and reduced Insulin Sensitivity Index (Matsuda by 30%; compared to their matched ND. The levels of taurine, alanine, glysine, Belfi ore by 45%; P<0.004) vs. RC rats. In RC rats circadian DA activity at the histidine, sarcosine and glutamic acid were higher in Indians whereas valine, SCN area peaked at LA onset (> 2-fold increase, p<0.0001) and this was leucine, lysine, cysteine, -amino adipic acid, amino butyric acid were higher in abolished in HFS rats (P<0.05). There was a marked (50%) increase in the NE Americans. An inherent metabolic differences and epigenetic adaptation number of c-Fos/TH co-expression neurons in SuMN of RC rats at LA onset to changing environment are shown among Indians and NE Americans.

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A470 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE

1829-P & 1827-P Circulating Oleate, but Not Other Free Fatty Acids, Suppresses Food Higher Intake of Palmitic Acid (PA) Leads to Higher Whole Body PA Intake in Wistar Rats Balance, PA Accumulation in Skeletal Muscle, but Not Incomplete YOUNG TAEK OH, JANG H. YOUN, Los Angeles, CA β-Oxidation of PA The brain plays an essential role in the regulation of energy balance C. LAWRENCE KIEN, JANICE Y. BUNN, DWIGHT E. MATTHEWS, NAOMI K. and metabolism. A crucial component of this regulation is fuel (or nutrient) FUKAGAWA, MATTHEW E. POYNTER, KAREN I. CRAIN, DAVID B. EBENSTEIN, sensing. Dietary fat is sensed in the gut to control food intake. However, Colchester, VT, Burlington, VT it is unclear whether fat is also sensed in circulation after its absorption. Higher intake of PA has been linked to increased ceramide in muscle, with Previous studies showed that, when injected into the brain, oleate, but not infl ammatory stress, and with increased blood concentrations of medium other free fatty acids, reduced food intake in rodents. These data suggest chain acylcarnitines in women in the fed-state, the latter implying less that circulating oleate may be a major signal for the brain control of food complete β-oxidation of fatty acids (Diabetes 62:1054, 2013). We assessed intake. The present study was carried out to directly test this idea. We PA oxidation (PAOX) to determine if a higher PA intake induces a higher conducted two series of experiments in normal male Wistar rats. In the fi rst retention (“balance”) of PA in the whole body and skeletal muscle and/or experiment, rats received a constant tail-vein infusion of olive, saffl ower, or 13 results in relative, incomplete β-oxidation (as evidenced by a 13- C-PAOX/1- coconut oil (100 mg/h; n = 4 each), together with heparin, to selectively raise 13 C-PAOX ratio <1). Using a crossover trial and a repeated dose, oral tracer + circulating oleate, linoleate, or saturated fatty acids (including palmitate), formula diet model, we studied 17 young adults (9 women), during 3 wk. respectively, and their impacts on overnight food intake was evaluated. Food 13 periods of high PA (HPA) and low PA/high oleic acid (HOA) diets. A [1- C]- intake decreased similarly with saffl ower and coconut oil infusions (27% and acetate breath test was used to correct for isotope exchange in the 29%, respectively), which might be responses to the energy intake due to 13 tricarboxylic acid cycle. During HPA, whole body PAOX (1- C) was 226% the oils infused overnight. In contrast, olive oil infusion had a more profound higher than during HOA (P < 0.0001). Net difference between total PA intake effect to reduce food intake (71%; P < 0.01 vs. other oils), suggesting that and PAOX (“PA balance”) also was higher (mean ± SEM) (219.5 ± 12.4 vs. 1.3 ± circulating oleate may have a strong inhibitory effect on food intake. To 3.9 µmol/min, P < 0.0001). Forearm muscle PAOX did not differ from zero directly test this idea, in the second experiment, rats received a constant on HPA and HOA (0.0016 ± 0.006 and 0.0424 ± 0.0433 µmol/min), but the infusion of oleate, linoleate, or octanoate (0.5 mg/h; n = 5 or 6), bound to 13 rate of muscle uptake of the [1- C]-PA tracer (µmol/min) was signifi cantly albumin, and their impacts on overnight food intake was evaluated. Oleate different from zero on the HPA (0.039 ± 0.021, P = 0.035) and HOA diets infusion decreased overnight food intake by 26% (P < 0.001), but no signifi cant 13 13 (0.012 ± 0.004, P = 0.008)(NS, for diet differences). The 13- C/1- C ratio of effect was seen with linoleate, octanoate, or vehicle (0.1% albumin) infusion whole body PAOX was 1.09 ± 0.10 and 1.28 ± 0.14 respectively during HPA (P > 0.05). We also found that the oleate effect was dose-dependent. Taken and HOA (men and women, NS), and in women only, 1.00 ± 0.10 and 1.54 ± together, our data indicate that circulating oleate, but not other free fatty 0.22 (P = 0.04). In conclusion, whole body PA balance was increased during acids, inhibits food intake in rats. Oleate, an essential FFA that originates the HPA diet, and there was net retention of PA in forearm muscle, during from food, may be involved in feedback control of food intake. 13 13 both diets. However, since the 13- C/1- C ratio of whole body PAOX was not Supported By: ADA (7-12-BS-214) less than one during the HPA diet, we speculate that incomplete β-oxidation of OA, not traced in this study, may account for the increased medium chain acylcarnitine production previously observed during a high PA diet. 1830-P Supported By: NIDDK (DK082803) The Role of Starvation-induced Autophagy in Gluconeogenesis and Ketogenesis AYANO TAKAGI, SHINJI KUME, YUKI TANAKA, HISAZUMI ARAKI, KEIJI ISSHIKI, 1828-P SHIN-ICHI ARAKI, TAKASHI UZU, HIROSHI MAEGAWA, Otsu, Japan RYGB Induces Segment-specifi c Changes in Small Intestinal In starvation, fatty acids released from adipose tissue are mainly used Glucose and Lipid Metabolism—A Transcriptomics Analysis for gluconeogenesis and ketogenesis in the liver to maintain systemic LIHONG CHEN, JOHANNES FREUDENBERG, NEETU RAJPAL, HINA BHUTTA, energy homeostasis in animals. Kidney is also known to contribute to gluco- JAMES WAY, DEEPAK RAJPAL, ANDREW A. YOUNG, ALI TAVAKKOLIZADEH, neogenesis. Autophagy, a major intracellular degradation system, is induced Durham, NC, Research Triangle Park, NC, Boston, MA by starvation. Systemic autophagy-defi cient mice die in neonatal starvation Bariatric operations such as Roux-en-Y gastric bypass (RYGB) improve period, suggesting that autophagy is a critical survival response against glucose control and weight via several proposed mechanisms including starvation. However, the exact role of starvation-induced autophagy remains intestinal glucose metabolism. One proposal, stemming from observations unclear. To determine the importance of autophagy, we examined starvation- that the Roux limb (Rx) of RYGB-treated rats that is exposed to undigested induced gluconeogenesis and ketogenesis in liver specifi c or kidney proximal foods, have enhanced basolateral glucose uptake and augmented aerobic tubular epithelial cells specifi c Atg5 knockout mice (L-Atg5-/- and K-Atg5-/-), glycolysis that could promote glycemic control. Similarly, increased and control mice (Atg5lox/lox). Although there were no differences under intestinal gluconeogenesis was proposed as a mechanism in surgery- ad-libitum feeding among 3 groups, after 36-h fast, increase in plasma mediated reduction of food intake and weight loss. Regional differences in ketone levels was impaired in L-Atg5-/- mice but not in K-Atg5-/- mice. The intestinal nutrient exposure after RYGB suggested to us that there are likely impairment of ketogenesis in L-Atg5-/- mice was not fully. In addition, the oil- regional differences in intestinal glucose handling. To test this hypothesis red-O-stained lipid droplets as a main source of ketogenesis and elevation in a obese diabetic rodent model, we analyzed mRNA expression of the of expression of HMG-CoA synthase, a rate-limiting enzyme of ketogenesis Rx, Bilio-Pancreatic (BP), and common (Cm) limbs as well as the terminal were observed in the kidney likely in the liver. We then hypothesized that ileum (TI) of non-fasted ZDF rats four weeks after RYGB, and compared the kidney compensated for the absence of ketone in L-Atg5-/- mice. To to equivalent intestinal sections after sham surgery (n=5 per group) using examine this hypothesis, we generated the mice lacking Atg5 in both the microarray. Although there were no major changes in the expression of liver and kidney. During a 36-h fast, these mice showed further decline in glycolytic enzymes, the expression of Pdk4 (phosphorylates and inhibits plasma ketone concentrations compared with L-Atg5-/- mice. Starvation- pyruvate dehydrogenase) was signifi cantly reduced in the BP and Rx but induced lipid droplets formation was impaired in Atg5-defi cient organs. No not Cm and TI limbs of RYGB-treated rats, suggesting enhanced entry of signifi cant differences in blood glucose levels and the expression of enzymes pyruvate into the TCA cycle. In BP and Rx but not Cm and TI limbs, there Integrated for gluconeogenesis in the liver and kidney were observed among 3 groups, POSTERS were signifi cant reductions in expression of Slc22a5, Slc27a2, Cpt1a, Cpt1b, even instarvation. These results suggest that starvation-induced autophagy Slc25a2 and Cpt2, which are involved in mitochondrial entry of long-chain is essential for ketogenesis relating with lipid accumulation, and that the Physiology/Obesity fatty acids for beta-oxidation, as well as Hmgcs2, the mitochondrial form kidney potentially has a function to generate ketone bodies. responsible for the biosynthesis of ketone bodies. On the contrary, RYGB increased expression of Pepck and decreased Pklr mRNA in Cm and TI, consistent with enhanced gluconeogenesis. In summary, our transcriptomic data indicate that RYGB surgery induces segment-specifi c changes of small intestinal glucose and lipid metabolism.

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A471 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE

1831-P 1833-P Chronic Administration of Apple Procyanidins Ameliorate Insulin Concomitant Intracarotid Infusion of a Lipid Emulsion and Glucose Resistance through the Suppression of Infl ammation in Diabetic ob/ob towards the Brain Increases Endogenous Glucose Production with- Mice out a Change in Neuropeptide Y Expression KASANE OGURA, KAZUAKI NAGASHIMA, TOSHIHIKO SHOJI, YUICHI SATOH, MEREL RIJNSBURGER, LESLIE EGGELS, JULIEN CASTEL, CELINE CRUCIANI- YUMIKO TAHARA, GEN YAMANO, HIROKI SATOH, KAZU SUGIZAKI, NAOTAKA GUGLIELMACCI, CHRISTOPHE MAGNAN, ERIC FLIERS, ANDRIES KALSBEEK, FUJITA, MASAHITO OGURA, ERI MUKAI, SHIMPEI FUJIMOTO, NOBUYA INAGAKI, MIREILLE SERLIE, SERGE LUQUET, SUSANNE LA FLEUR, Amsterdam, Netherlands, Kyoto, Japan, Ibaraki, Japan Paris, France Procyanidins, the main ingredient of apple polyphenols, are known to Providing rats the choice of saturated fat and liquid sugar in addition to possess anti-oxidative and anti-infl ammatory effects. Several studies chow induces glucose intolerance and increases hypothalamic neuropeptide have reported the association of oxidative stress and infl ammation with Y (NPY) expression, whereas providing either fat or sugar only in addition to insulin resistance in obese and diabetic subjects. The aim of this study is to chow does not. Although NPY can modulate glucose metabolism, at present investigate the effect of apple procyanidins (APs) on glucose metabolism. it is unknown whether direct effects of fat and sugar on NPY expression APs were administrated to diabetic ob/ob mice (8 weeks of age) via cause the observed diet-induced effect on glucose. drinking water for 3 weeks. There is no difference in body weight and food We hypothesized that direct infusion of lipids and sugar towards the brain intake between APs-treated and non-treated ob/ob mice. Oral glucose increases glucose concentration through an increase in endogenous glucose tolerance test (OGTT, 1g/kg glucose), insulin tolerance test (ITT, 2U/kg production (EGP) induced by an increase in NPY expression. To study this insulin), and pyruvate tolerance test (PTT, 1g/kg pyruvate) were performed. we used a 2-hours intracarotid infusion of either heparinized Intralipid 20% In OGTT, blood glucose levels were signifi cantly lower at 15 min and 30 min (IL), IL and glucose 1% (IL + G) or control saline (NaCl) at a rate of 5µl/min in in APs-treated ob/ob mice compared with those in non-treated ob/ob mice. male Wistar rats fasted overnight. We assessed EGP using a stable isotope In ITT, blood glucose levels were signifi cantly lower at 15, 30, 45, 60 min in dilution method. APs-treated mice. In PTT, APs-treated mice exhibited lower blood glucose NaCl infusion had no effect on glucose concentrations while IL signifi cantly levels at 15 min and 30 min. Administration of APs suppressed the increase decreased glucose. IL + G did not affect glucose concentrations, but when of pancreatic beta-cell area in ob/ob mice. There is no difference in lean comparing to IL, glucose was signifi cantly higher (P < 0.01). EGP increased mass or fat compositions evaluated by computerized tomography between upon IL + G infusion only. In situ hybridization showed no differences in NPY APs-treated and non-treated mice. Total mRNA and protein were extracted mRNA expression between groups. Surprisingly, IL signifi cantly reduced from various organs including liver from APs-treated and non-treated mice. POMC mRNA expression compared to rats infused with NaCl (P < 0.05). The expression levels of oxidative stress-induced or infl ammation-related Taken together, these data show that intracarotid IL infusion towards the gene were examined by quantitative RT-PCR. The mRNA expression levels brain decreases glucose concentrations and POMC mRNA. Infusion of IL + G of TNF-α and IL-6 were signifi cantly decreased in liver of APs-treated mice. increases EGP which may counteract the glucose lowering effect of IL only. Western blot analyses showed that insulin-stimulated Akt phosphorylation The increase in EGP was independent of changes in NPY or POMC. was signifi cantly enhanced and JNK and ERK phosphorylation were Supported By: ZonMw downregulated in liver of APs-treated mice. These results suggest that APs ameliorates insulin resistance by the improvement of hepatic insulin 1834-P signaling through the suppression of infl ammation in ob/ob mice. Determination of Human Serum Acylcarnitine Profi les in Different Supported By: Ministry of Agriculture, Forestry and Fisheries Glucose Intolerance Statuses XIUYING ZHANG, CHUNFANG ZHANG, LING CHEN, XUEYAO HAN, LINONG JI, 1832-P Beijing, China Metabolic Effects of an Amino Acid Infusion during LPS Exposure Aims: To understand the relationship between serum acylcarnitine profi les Mimicking Acute Infection in Humans and the development of diabetes. NIKOLAJ RITTIG, ERMINA BOSNJAK, HENRIK H. THOMSEN, BJØRN RICHELSEN, Methods: We analyzed 61 subjects who were divided into three groups JENS O. JØRGENSEN, NIELS MØLLER, Aarhus, Denmark based on their glucose intolerance status: normal glucose tolerance (NGT; n = Acute infl ammation is catabolic and increases protein loss. This study was 20, M/F = 9/11, mean age 48 years), pre-diabetes (Pre-DM; n = 20, M/F = designed to test whether a continuous amino acid infusion could counteract 11/9, mean age 51 years), or newly diagnosed type 2 diabetes mellitus these metabolic effects. (T2DM; n = 21, M/F = 8/13, mean age 49 years). Fasting serum free carnitine Eight healthy males randomly underwent 3 different interventions: Control/ and acylcarnitine concentrations were determined using isotope dilution placebo, LPS injection (E. Coli lipopolysaccharide, 1 ng/kg), and LPS + amino electrospray ionization mass spectrometry coupled with high performance acid infusion (2.5 g/kg/day). Metabolic parameters were measured after 4 liquid chromatography. hour in and after a 2 hour hyperinsulinemic euglycaemic clamp. Glucose, fat, Results: In comparison with NGT subjects, Pre-DM and type 2 diabetes and protein metabolism were quantifi ed by isotope tracer dilution, forearm subjects showed clear serum metabonomic changes highlighted by dys- arterio-venous differences and by indirect calorimetry. Muscle and fat regulation of mitochondrial fatty acid combustion. Of the long-chain carni- biopsies were obtained for signaling analysis. tine esters, signifi cantly higher palmitoylcarnitine (C16), 3-OH-hexa deca- On both days LPS caused a similar degree of infl ammation with a signifi cant noyl carnitine (C16-OH), carnitine C20, carnitine C22, and carnitine C24 increase in plasma cortisol levels (185.9±50.0 ng/ml), temperature (2.6±1.6 concentrations (all P < 0.05) were noted in the newly diagnosed type 2 °C), and pulse (37.3±6.0 bpm) compared to placebo. diabetes group, and even the pre-diabetes group. Energy expenditure increased with ≈ 500 kcal/day (CI 368.5;608.0) on Conclusions: This research provides further evidence of alterations in the days with LPS compared to placebo. When adding amino acids to LPS serum acylcarnitine profi les being associated with the progression of glucose there was a signifi cant increase in protein oxidation of 176.4 kcal/day (CI intolerance. The fi ndings may suggest different degrees of involvement of 61.6;291.1) compared to placebo. The glucose clamp caused an increase in dysregulated mitochondrial function and incomplete long-chain fatty acid glucose oxidation and a decrease in fat oxidation in all groups, the insulin oxidation pathways in the natural course of diabetes. levels being lower when comparing LPS alone with placebo (p<0.001) Integrated POSTERS and tending to be lower between LPS + amino acids compared to placebo 1835-P (p=0.062). The glucose infusion rate (GIR) did not differ between the two

Physiology/Obesity Pioglitazone Inhibits Pyruvate Transport and Oxidation In Vitro days of LPS (0.94 mg/kg/min, CI -0.22;2.10). Independent of the Mitochondria Pyruvate Carrier-1 and -2 In conclusion, LPS caused a signifi cant degree of infl ammation and a LUKE NORTON, GIUSEPPE DANIELE, CYNTHIA GALINDO, NARGES MOHAMMAD- catabolic response with a 28 % increase in energy expenditure and amino TAGHVAEI, JERRY R. COLCA, RALPH A. DEFRONZO, MUHAMMAD ABDUL-GHANI, acid supplementation further increased protein oxidation by 38 %. Thus San Antonio, TX, Ahvaz, Islamic Republic of Iran, Kalamazoo, MI i.v administration of amino acids during acute infl ammation has distinct Pioglitazone (PIO) lowers plasma glucose concentration through enhanced metabolic effects on substrate utilization and may have a future clinical role peripheral insulin-stimulated glucose uptake and suppression of hepatic as an anabolic agent. glucose output. While these actions are thought to occur through PPAR-γ mediated adipose tissue expansion, reduced plasma FFA concentration and depletion of ectopic fat from muscle and liver, existing evidence also supports a PPAR-γ-independent action of PIO. The aim of this study was to investigate the effect of PIO on pyruvate transport and oxidation and examine whether

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A472 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE the newly identifi ed mitochondrial pyruvate carriers (MPC1 and MPC2) delivery and uptake. Here, we studied whether perivascular adipose were required for the action of PIO on pyruvate metabolism. We measured tissue (PVAT) controls insulin-induced microvascular recruitment in human pyruvate oxidation/transport and oxygen consumption rate (OCR) in rat H4IIE skeletal muscle and insulin sensitivity in 18 obese and 15 lean women. hepatocytes and L6 myoblasts before and after MPC1 and MPC2 siRNA To this end, we measured insulin-induced microvascular recruitment with silencing. First, we demonstrate that silencing of MPC1 (P < 0.05) or MPC2 contrast-enhanced ultrasound (CEU) before and during a hyperinsulinemic (P < 0.05) signifi cantly reduced pyruvate oxidation in hepatocytes, confi rming euglycemic clamp. Obese women had impaired microvascular recruitment an important role for these transporters in regulating pyruvate metabolism. compared with lean women (10% (-57 to +62) vs. 26% (-44 to +210 (p<0.05)), Second, in control hepatocytes, 10 µM PIO inhibited pyruvate oxidation (P as well as a lower metabolic insulin sensitivity (M-value 6.0 (2.2-15.4) mg/ = 0.001), as did 5 µM of the specifi c pyruvate transport inhibitor UK5099 (P kg/min vs. 10.9 (4.4-14.9) mg/kg/min, p<0.001). Microvascular recruitment < 0.001). These results were confi rmed independently in L6 myoblasts. In signifi cantly attenuated the association between obesity and metabolic isolated mouse muscle mitochondria, PIO (P < 0.05) and UK5099 (P < 0.05) insulin sensitivity from B=-0.76, p<0.001 to B=-0.60, p<0.05. Perivascular dose-dependently inhibited pyruvate transport, and pyruvate-driven basal and adipocytes were smaller in lean than in obese women (1637 (195-3751) um FCCP-stimulated OCR was reduced by PIO (P < 0.05) and UK5099 (P < 0.05). vs. 530 (152-1002) um, p<0.01) and these sizes statistically explained the Third, and most importantly, the ability of PIO or UK5099 to inhibit pyruvate difference in microvascular recruitment between obese and lean women; oxidation was unaffected by the silencing of MPC1 or MPC2. Lastly, inhibition change in beta from B=-0.40, p<0.05 to B=0.01, p=NS. Ex vivo, in resistance of pyruvate metabolism by PIO was associated with signifi cant reductions in microvessels isolated from the skeletal muscle biopsies, PVAT from obese hepatic glucose production (HGP) in hepatocytes, independent of changes in women revealed insulin-induced vasoconstriction (-25±3%, p<0.05), while the expression of HGP enzymes (P < 0.001). These data suggest that PIO is an PVAT from lean women enhanced insulin-induced vasodilation (20±8%, important regulator of pyruvate metabolism, but that its mechanism of action p<0.05). These results strongly suggest that PVAT enhances insulin-induced is independent of the MPC pyruvate carrier proteins. microvascular perfusion and, thereby, insulin-induced glucose uptake in skeletal muscle of lean women, this effect being abolished in obesity. Perivascular adipose tissue may therefore be an important factor in insulin INTEGRATED PHYSIOLOGY—MUSCLE resistance and cardiovascular disease. Supported By: Netherlands Heart Foundation (2009B098)

Guided Audio Tour: Muscle Insulin Sensitivity—Pathophysiological & 1838-P Mechanisms (Posters: 1836-P to 1842-P), see page 15. Glucagon-like Peptide-1 (GLP-1) and Insulin Recruit Muscle Microvasculature and Dilates Conduit Artery Independently but Not & 1836-P Synergistically in Healthy Humans Muscle-specifi c Integrin-linked Kinase Deletion Rescues Muscle SHARMILA C. SUBARAN, MATTHEW A. SAUDER, WEIDONG CHAI, LINDA JAHN, Insulin Resistance in High Fat-Fed C57BL/6J Mice DALE E. FOWLER, KEVIN AYLOR, ANANDA BASU, ZHENQI LIU, Charlottesville, VA, LI KANG, SHILPA MOKSHAGUNDAM, BRADLEY REUTER, DEANNA P. BRACY, Rochester, MN FREYJA D. JAMES, ROY ZENT, DAVID H. WASSERMAN, Nashville, TN Muscle microvasculature regulates oxygen, nutrient and hormone Expansion of extracellular matrix (ECM) components such as collagen and exchanges between plasma and muscle interstitium. Both insulin and GLP-1 its interaction with integrin α2β1 is associated with diet-induced muscle are able to recruit muscle microvasculature and dilate conduit arteries via a insulin resistance (IR). Transduction of ECM signals through integrins requires nitric oxide-dependent mechanism. interaction of integrin cytoplasmic domains with cellular proteins. Integrin- To examine whether GLP-1 and insulin at physiological concentrations act linked kinase (ILK), a downstream scaffold protein of the integrin signaling synergistically, 18 overnight fasted, healthy young adults were studied twice interacts with the cytoplasmic domain of β1 integrin. We hypothesized that and randomly received either a GLP-1 (1.2 pmol/kg/min) or saline infusion for ILK is key in linking ECM-integrin signaling to muscle IR. Wild type (WT) 150 mins with a euglycemic insulin clamp (3 mU/kg/min) superimposed on and muscle-specifi c ILK defi cient (MILK) mice were chow fed (CF) or high the last 120 mins. Microvascular blood volume (MBV), microvascular fl ow fat fed (HFF) for 16 wks. The hyperinsulinemic (4 mU/kg/min) euglycemic velocity (MFV) and microvascular blood fl ow (MBF) were determined using clamp (IC) was used to assess insulin action in conscious, unstressed mice contrast-enhanced ultrasound at baseline, 30 min and 150 min. Brachial with indwelling arterial (sampling) and venous (infusion) catheters. A muscle artery diameter (d), fl ow velocity (v) and blood fl ow (Q) were quantifi ed at glucose metabolic index (Rg) was assessed using 2[14C]deoxyglucose. Body baseline and 150 min. weight and composition did not differ between CF WT and MILK mice. HFF Insulin infusion signifi cantly increased muscle MBV (~57%) and MBF MILK weighed slightly less than HFF WT due to less lean mass (24.1±0.3 vs. (~45%)(p<0.001 for both). GLP-1 infusion alone increased MBV and MBF 26.5±0.4 g). MILK deletion decreased fasting glucose (123±5 vs. 150±8 mg/ by ~30% at 30 min (p<0.001 for both). Superimposition of insulin on GLP-1 dL) and insulin (12±2 vs. 18±1 ng/mL) in HFF mice, with no effects in CF mice. infusion raised MBV and MBF by ~20% (p<0.001 for both) but the values Glucose infusion rate (GIR), IC glucose disappearance rate (Rd), and muscle were not signifi cantly different from insulin alone (p>0.05). This was Rg were the same in CF WT and MILK mice. In contrast, HFF MILK increased accompanied by a slight (~5%) decrease in MFV at 150 min in both groups GIR (30±2 vs. 15±2 mg/kg/min), IC Rd (37±4 vs. 24±2 mg/kg/min) and muscle (p<0.05). Similarly both insulin alone and GLP-1 and insulin combination Rg (17±3 vs. 8±1 µmol/100gtissue/min). MILK deletion did not affect IC signifi cantly increased d (p<0.005 and p<0.001 respectively) and Q (p<0.005 suppression of hepatic glucose production. Despite improved muscle IR, and p<0.05, respectively) but the increases were compatible between the collagen IV deposition in HFF MILK muscle remained elevated as in HFF two studies (p>0.05). WT mice when compared to CF WT mice. Activation of Akt, a downstream We conclude that GLP-1 and insulin at physiological concentrations each component of insulin signaling was increased in muscle of HFF MILK mice. recruits muscle microvasculature and dilates conduit vessels in healthy These results show that MILK deletion uncouples the association between humans but these effects are not synergistic. These indicate that in the muscle IR and increased collagen-integrin interaction, confi rming that ECM- postabsorptive state a defi ned amount of muscle microvascular recruitment integrin-ILK signaling is a key determinant of muscle IR in mice. is suffi cient for physiological interaction between GLP-1 and insulin in the Supported By: DK054902, DK059637 muscle microcirculation. Integrated

Supported By: ADA (7-09-NOVO-11) POSTERS & 1837-P Perivascular Adipose Tissue Mediates the Difference in Insulin & 1839-P Physiology/Obesity Induced Microvascular Recruitment in Muscle between Lean and A Novel Role of Interleukin-10 to Regulate Skeletal Muscle Growth Obese Women in Obesity RICK I. MEIJER, ERIK H. SERNÉ, IBRAHIM H. KORKMAZ, DONALD L. VAN DER SEZIN DAGDEVIREN, RANDALL H. FRIEDLINE, SOCHEATA LY, MATTHEW GODIN, PEET, MICHIEL P. DE BOER, HANS W.M. NIESSEN, VICTOR W.M. VAN HINSBERGH, KEVIN HSU, EMILY RADEMACHER, AVNEET SOIN, JASON K. KIM, Worcester, MA JOHN S. YUDKIN, YVO M. SMULDERS, ETTO C. ERINGA, Amsterdam, Netherlands, Obesity exerts pathophysiological effects on multiple organs including London, United Kingdom musculoskeletal system. We have shown that mice with muscle-specifi c Obesity increases risk of cardiovascular diseases and type 2 diabetes, overexpression of IL-10 (MCK-IL10) are protected from obesity-mediated at least partly through its association with microvascular dysfunction and infl ammation and insulin resistance in muscle. Serendipitously, we found insulin resistance. In the skeletal muscle microcirculation of obese subjects, that MCK-IL10 mice showed an increased lean mass as compared to WT insulin’s vasodilator effects are impaired, reducing insulin-induced glucose mice during chronic high-fat diet (HFD) (Fig. 1; *P<0.05, n=10/group). IL-10

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A473 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

expressing leptin-defi cient ob/ob mice (MCK-IL10ob/ob) also developed higher lean mass than ob/ob mice after 8 wks of age (Fig. 2; n=4/group). & 1841-P To determine the mechanism, we performed histological and molecular Predictors of Whole-Body Insulin Sensitivity analyses in skeletal muscle samples collected from WT (n=2) and MCK-IL10 IAN LANZA, MATTHEW M. ROBINSON, KLAUS DISTELMAIER, MATTHEW mice (n=3) after 16 wks of high-fat feeding. HFD-fed MCK-IL10 mice showed JOHNSON, ADAM KONOPKA, MARIA GLAVIN, JOHN PORT, K. SREEKUMARAN a 50% increase in muscle fi ber number as compared to HFD-fed WT mice NAIR, Rochester, MN (Fig. 3). qRT-PCR was performed to measure muscle expression of growth- Numerous factors are believed to contribute to the decline in insulin associated genes, and muscle expression of Myf5, MyoD, Myogenin, and sensitivity with obesity and aging. Leading hypotheses implicate ectopic MyHC was increased by 2 to 5-fold in HFD-fed MCK-IL10 mice (Fig. 4). Thus, lipids, dysfunctional mitochondria, and accumulation of abdominal fat, IL-10 expression induced muscle growth in obesity. Overall, our fi ndings particularly visceral fat, in the development of insulin resistance. Here, suggest that physiological growth of skeletal muscle to support increased we evaluated a large cohort (127 cases) of men and women with a wide body weight may be affected by obesity-induced infl ammation and insulin range of age (19-75yrs) and BMI (20-44kg/m2) to identify robust predictors resistance, and that IL-10 mediated suppression of infl ammation and insulin of insulin sensitivity. Insulin sensitivity was measured by hyperinsulinemic- resistance maintains normal muscle growth in obesity. euglycemic clamp. Intramyocellular and intrahepatic lipid levels were measured by proton magnetic resonance spectroscopy. Body composition was measured by DEXA. Abdominal subcutaneous and visceral fat were measured by magnetic resonance imaging. Skeletal muscle mitochondrial capacity was measured by high-resolution respirometry of mitochondria isolated from vastus lateralis muscle biopsies (JO2). In vivo mitochondrial capacity was measured by phosphorous magnetic resonance spectroscopy (kPCr). Stepwise multiple linear regression was performed using a backward elimination approach. Initial candidate variables were age, BMI, body fat (%), abdominal subcutaneous fat, visceral fat, liver lipid, muscle lipid, and mitochondrial capacity (JO2 and kPCr). After adjusting for other variables in the model, % body fat, muscle lipid, liver lipid, and muscle oxidative capacity were not signifi cant predictors of insulin sensitivity. Together, age, BMI, visceral fat, and abdominal subcutaneous fat were statistically signifi cant predictors of insulin sensitivity in the adjusted model. Chronological age remained a signifi cant predictor of insulin sensitivity after adjusting for BMI, visceral fat, and abdominal fat, These data indicate that age and body fat distribution are more important predictors of insulin sensitivity than mitochondrial capacity or ectopic lipid levels. Supported By: NIH (R01DK41973, R01AG09531, UL1TR000135)

Supported By: NIH (DK080756) & 1842-P Hyperinsulinemia Augments Endothelin-1 Release and Impairs & 1840-P Vasodilation of Human Skeletal Muscle Arterioles DPP-4 Inhibitor Anagliptin Increases Capillary Recruitment and ABEER M. MAHMOUD, JACOB T. MEY, BRIAN K. BLACKBURN, KARIA COLEMAN, Glucose Uptake in Skeletal Muscle via Endothelial NO-dependent VIKRAM S. SOMAL, JING-TAN BIAN, MARY R. SZCZUREK, AUSTIN T. ROBINSON, Pathway SHANE A. PHILLIPS, JACOB M. HAUS, Chicago, IL HIROYUKI SATO, NAOTO KUBOTA, TETSUYA KUBOTA, ISEKI TAKAMOTO, KUMPEI Increased endothelin 1 (ET-1), a potent vasoconstrictor, is implicated in TOKUYAMA, KEIZO NAKAYA, SHINJI HASHIMOTO, MORITAKA GOTO, TAKAHITO diabetes associated cardiovascular disease (CVD). Balance between ET-1 JOMORI, KOHJIRO UEKI, TAKASHI KADOWAKI, Tokyo, Japan, Bunkyo-ku, Japan, and nitric oxide (NO) is maintained by normal levels of insulin, however Ibaraki, Japan, Inabe, Japan this balance may be disrupted with compensatory hyperinsulinemia One of the pathogenic mechanisms of the skeletal muscle insulin (HI) as a consequence of insulin resistance. The effect of HI on ET-1 is resistance is considered to be attenuation of insulin-induced capillary controversial and has not been evaluated in human skeletal muscle (SkM) recruitment of skeletal muscle as a consequence of impaired endothelial microvasculature. We tested the hypothesis that HI augments ET-1 release insulin signaling in obese patients with type 2 diabetes. Recently, it has and subsequently impairs endothelium dependent fl ow induced dilation (FID) been reported that GLP-1 may affect vascular endothelial cells (ECs), thereby in isolated SkM arterioles. SkM biopsies were performed on 8 lean healthy improving endothelial dysfunction. In this study, we investigated the effect controls (LHC: Age 37±4 y, BMI 21±1 kg/m2, M: 7.1±0.5 mg/kg/min) and 6 of DPP-4 inhibitor on ECs of skeletal muscle using endothelium specifi c IRS-2 obese, type 2 diabetics (OB: Age 65±0.8, BMI 34.1±0.7 kg/m2, M: 2.7±0.3 knockout (ETIrs2KO) mice, a skeletal muscle insulin resistance animal lacking mg/kg/min) before and after a 40 mU/m2/min insulin clamp. SkM protein insulin signaling in ECs. from LHC (30µg) was probed for ET-1 and ET converting enzyme (ECE). In Hyperinsulinemic-euglycemic clamp was performed in ETIrs2KO mice OB (n=6) and a subset of LHC (n=5), the internal diameter of isolated SkM at 8 weeks after administration of DPP-4 inhibitor anagliptin mixed in the arterioles was measured before and during intraluminal fl ow corresponding normal chow at 0.3%. To evaluate the capillary recruitment, capillary blood to increasing pressure gradient, with or without NO synthase inhibitor volume of skeletal muscle after insulin injection was measured. Anagliptin (L-NAME; 10-4 M). LHC subjects showed increased ET-1 and ECE protein signifi cantly increased the insulin-induced glucose uptake in skeletal muscle expression (AU) during HI (ET-1: 0.63±0.04, ECE: 1.08±0.17) compared to (18% increase compared to ETIrs2KO mice not receiving anagliptin) without basal conditions (ET-1: 0.44±0.05, p= 0.01, ECE: 0.80±0.06, p=0.02). FID (% affecting the body weight. The effect of anagliptin on glucose uptake was of dilation at Δ60 pressure gradient) was impaired with HI in LHC (basal: signifi cantly but partially inhibited by a GLP-1 receptor antagonist exendin 74.2±2.0; HI: 57.2±3.3, p=0.003) and in OB (basal: 62.1±3.6; HI: 48.5±5.6,

Integrated p=0.04). Compared to baseline, FID was also reduced in the presence of POSTERS (9-39). In addition, capillary blood volume of skeletal muscle in response to insulin was increased by 36% by treatment with anagliptin in ETIrs2KO L-NAME (LHC: -36.8%, p=0.04; OB: -45.9%, p<0.001). Interestingly, this Physiology/Obesity mice. The increment of capillary blood volume by anagliptin was completely L-NAME impairment was greater with HI (versus basal L-NAME; LHC: inhibited by administration of L-NAME, an inhibitor of NO synthesis. -64.2%, p=0.02; OB: -37.1%, p=0.046), suggesting increased vasodilation Furthermore, in vitro study using human coronary artery endothelial cells dependence on NO. In conclusion, these data support the hypothesis that (HCAEC), GLP-1 but not anagliptin up-regulated the eNOS phosphorylation, HI may disturb the vasoactive substance balance favoring ET-1 release and and this effect was completely inhibited by PKA inhibitor H89. vasoconstriction which may modify risk factors for CVD. These data imply that anagliptin increases the insulin-induced capillary recruitment and glucose uptake in skeletal muscle of ETIrs2KO mice via enhanced endothelial NO production, in part, by increased GLP-1 responses.

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A474 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

1843-P 1845-P Hyperglycemia-induced Deterioration of Insulin Sensitivity in Human Glucocorticoids Stimulate Intramuscular Adipose Derived from Skeletal Muscle Is Dependent on Endoplasmic Reticulum Stress Fibro/Adipogenic Progenitors by an Il-4 Mechanism THOMAS P.J. SOLOMON, SINE H. KNUDSEN, HELEN BRADLEY, KRISTIAN YANJUN DONG, KLEITON SILVA, YANLAN DONG, WILLIAM E. MITCH, LIPING KARSTOFT, MATTHEW J. LAYE, CHARLOTTE J. GREEN, ANTON J.M. WAGEN- ZHANG, Houston, TX MAKERS, BENTE K. PEDERSEN, CHRISTOPHER S. SHAW, Copenhagen, Denmark, Excess body fat associated with developing diabetes and the metabolic Birmingham, United Kingdom, Liverpool, United Kingdom, Melbourne, Australia syndrome via the infl uence of excess glucocorticoids (GCs). In injured muscles To test the hypothesis that hyperglycemia reduces insulin sensitivity of mice treated with dexmethasone (Dex), satellite cell activation and via ER stress, 10 healthy overweight subjects underwent a 24-hour muscle regeneration was impaired but adipocytes increased. To determine hyperglycemic clamp and changes in insulin sensitivity and signaling in the origin of these adipocytes, satellite cells and fi bro/adipogenic progenitor skeletal muscle were determined after 2 and 24 hours. The same variables cells (FAPs) were isolated by FACS cell sorting. We found satellite cells do not were measured in 10 age- and BMI-matched type 2 diabetic subjects during transform into tissues except muscle: Satellite cells only differentiate into a 2-hour hyperglycemic clamp. An in vitro study of human primary skeletal myofi bers even cultured in an adipogenic differentiation media; transplanting muscle (HSkM) cells derived from 10 healthy subjects was also conducted. satellite cells isolated from EGFP transgenic mice into muscles of C57/BL6 Differentiated HSkM cells were incubated for 24-hours in 5 mM or 25 mM mice, they only differentiate into myofi bers even in muslce injury and Dex glucose ± 5 mM taurodeoxycholic acid (TUDCA), an ER stress inhibitor. treatment condition. But, Dex induced FAPs differentiate into adipocytes in Infl ammation (plasma interleukin [IL]-6; muscle tyrosine-185 phosphoryla- vivo and in vitro: Dex increase FAPs differentiate into perilipin positive cells tion of c-Jun N-terminal kinase [p-JNK]) and ER stress (muscle serine-51 in cell culture; transplanting FAPs isolated from EGFP mice into muscles of phosphorylation of eukaryotic translation initiation factor 2 alpha [p-eIF2α]) C57/BL6 mice, they differentiate into adipocyte only in injured muscles of were greater in subjects with type 2 diabetes, who had lower insulin Dex-treated mice. Since Dex increases myostatin expression, we explored sensitivity than healthy subjects. Hyperglycemia for 24 hours in healthy whether myostatin has similar function as GCs. Unexpectedly, myostatin subjects reduced insulin sensitivity, while increasing infl ammation (plasma inhibited FAPs adipogenic while stimulated FAP fi brotic differentiation in IL-6; muscle p-JNK) and ER stress (muscle p-eIF-2α), but without impairment vitro and in vivo. In fact, the mechanism involves Dex-mediated inhibition in canonical insulin signaling. Exposure of HSkMs to 25 mM glucose reduced of IL-4 expression. IL-4 (2g/25g mice) blocked Dex induced adipocytes in insulin-stimulated 2-deoxy-D-[3H]glucose uptake. TUDCA partially but injured muscles. IL-4 also inhibited Dex induced adipogenic differentiation signifi cantly attenuated this impairment, confi rming a role for ER stress in of FAPs in vitro. Our results suggest that glucocorticoid induce intramuscular hyperglycemia-induced insulin resistance. adipocyte from FAPs, a process could be blocked by IL-4, which providing a Hyperglycemia per se reduces insulin sensitivity in humans via a mecha- therapeutical strategy. nism involving the unfolded protein response in skeletal muscle (indicated Supported By: ADA (1-11-BS-194); NIH (R37DK-37175); Norman S. Coplon (to L.Z.) by ER stress). Reducing ER stress in diseases characterized by chronic hyperglycemia may improve insulin sensitivity. 1846-P Supported By: EFSD Role of PI3-Kinase p110 Subunit in Muscle Growth and Insulin Re- sponse 1844-P MENGYAO LI, C. RONALD KAHN, Boston, MA PPARD SNP-dependent Effect of the Human Myotube Secretome on The class IAphosphatidylinsositol 3-kinases (PI3Ks) regulate the metabolic GSIS and the growth-promoting effects of insulin/IGF-1 pathways, however, the ANNA-MARIA ORDELHEIDE, MANUEL CAVADA, FELICIA GERST, SUSANNE precise role of the many different isoforms of this enzyme is still unclear. To ULLRICH, HANS-ULRICH HAERING, HARALD STAIGER, Tübingen, Germany determine the relative roles of the p110α and p110b catalytic subunits of PI3K It is recognized that single nucleotide polymorphisms (SNP) together in skeletal muscle we stably overexpressed each isoform in skeletal C2C12 with lifestyle and environmental factors determine the metabolic fate of muscle cells in vitro. Both resulted in increased rates of cell proliferation (15- human individuals. SNPs in the peroxisome proliferator-activated receptor δ 30%) and almost doubled MTT activity, with p110a being more effective than (PPARD) have been shown to play a role in the change of body composition, p110b. To further explore the role of p110a, we created mice in which this aerobic physical fi tness and insulin sensitivity during lifestyle intervention. gene was specifi cally deleted in skeletal muscle by Cre-lox recombination Additionally, we have shown that several myokines, like ANGPTL4 and (M-p110αKO). By 6 weeks of age, these mice exhibited an ~30% reduction G-CSF, are secreted by human myotubes in a PPARδ-dependent manner. in muscle fi ber size with no changes in muscle, fat or body weight (p < Therefore, we were interested in possible PPARD SNP-dependent effects 0.05). This was accompanied by 40-50% increases in expression of muscle of the myotube secretome on other organs, e.g., the pancreatic β-cell. We growth/differentiation markers MyoD1, Myf5 and myogenin, and decreased cultured myotubes differentiated from primary human myoblasts from 9 expression of ubiquitin ligase 3 Atrogin1 and MurF1 (both ~ 45%) (all p < 0.05). PPARD non-risk allele and 9 risk allele carriers (compound heterozygous for These occurred with no difference in the expression of muscle fi ber type rs6902123, rs2267668 and rs1053049). Conditioned media were generated markers (myosin 1, myosin 2a, myosin 2x and myosin 2b). M-p110αKO mice in the presence of the PPARδ agonist GW501516. These supernatants were also displayed a trend of impaired glucose tolerance with 14% of increase of used to incubate INS-1E cells prior to determination of glucose-stimulated area under curve. Together, these data indicate that both p110α and p110b insulin secretion (GSIS). Surprisingly, supernatants of risk allele carriers isoforms of PI3K play important roles in skeletal muscle with p110a being showed a positive effect on GSIS compared to non-risk allele supernatants. more important in growth and control of glucose tolerance. To fi nd differently expressed myokines in these supernatants, we carried out a cytokine array measuring 274 cytokines. We found several cytokines 1847-P to be differently regulated, such as insulin-like growth factor-binding Pin1 Reduces AMPK Phosphorylation and Thereby Contributes to protein 2 and platelet-derived growth factor homodimer BB (p<0.05). The the Impaired Mitochondrial Activity and Lipid Accumulation in the difference in secretion was mirrored on the mRNA level (p<0.05). We also Muscle examined whether SNPs in PPARD infl uence the expression of adjacent YUSUKE NAKATSU, MISAKI IWASHITA, YASUKA MATSUNAGA, TOSHIAKI genes. We found the transcription factor TEAD3 to be differently regulated FUKU SHIMA, HIDEYUKI SAKODA, MIDORI FUJISHIRO, AKIFUMI KUSHIYAMA, Integrated by the PPARD SNPs. This was also visible on the protein level. Additionally, TAKAFUMI UCHIDA, TOMOICHIRO ASANO, Hiroshima, Japan, Tokyo, Japan, Sendai, POSTERS we found two miRNAs (MIR5690, AL138721.1) to be differently regulated

Japan Physiology/Obesity (p<0.05). These data show that SNPs in PPARD infl uence the secretome of Prolyl isomerase (Pin1) associates with the pSer/Pro or pThr/Pro-con- human skeletal muscle cells and, thus, might have an impact on interorgan taining motif and thereby changes the conformations of target proteins. communication. In particular, we show here the effect on GSIS. High-fat diet feeding for 7 days upregulates Pin1 expression by several Supported By: German Center for Diabetes Research fold and Pin1 KO mice show resistance to high-fat diet induced obesity. These observations strongly suggest the importance of Pin1 in energy metabolism. In this study, it was revealed that Pin1 binds to γ1, γ2 or γ3 but not the α or β subunit of AMPK. The analysis using deletion- and point- mutated Pin1 and AMPKγ1 constructs revealed the N-terminal WW domain of Pin1 and the T221-cintaining motif located in CBS3 of AMPKγ1 to be involved in their association. Overexpression of wild-type Pin1 markedly attenuated phosphorylation of the α subunit of AMPK induced by AICAR

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A475 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

or 2-deoxyglucose treatment in HepG2 cells. In contrast, phosphorylation kinase inactive IGF1R in muscle induced glucose intolerance and elevated of the α subunit of AMPK was enhanced in the HepG2 cells treated with triglycerides in both normal and MIGIRKO mice. Thus, insulin and/or IGF-1 Pin1 siRNA as well as in the muscles of Pin 1 KO mice. The experiments receptor signaling is required for maintenance of muscle mass, but not for using recombinant Pin1, AMPK and LKB1 revealed no signifi cant effects of normal glucose tolerance. Nonetheless, presence of a dominant negative Pin1 on LKB1-induced phosphorylation of the α subunit. On the other hand, receptor in muscle, even in the absence of functional IR and IGF1R, induces dephosphorylation of the α subunit, reportedly by PP2C, was markedly delayed dysglycemia and derangements in metabolism, indicating that interactions in the cells overexpressing Pin1, as compared with control cells. Taking the between these receptors and other proteins in muscle is required for normal Pin1 binding site in the CBS domain into consideration, the protective effect regulation of glucose homeostasis. of AMP against dephosphorylation by PP2C appears to be contradicted by Supported By: NIH (K08DK100543 to B.T.O.) the binding of Pin1 to the CBS domain. In good agreement with the data on the AMPK phosphorylation level, expressions of mitochondria-related 1850-P genes, such as PGC-1α, were markedly higher and triglyceride contents were Differential Effects of Infectious and Metabolic Infl ammation on lower in the muscles of Pin1 KO mice, as compared with control mice. Myokine Secretion by Myotubes from Non-Diabetic and Type 2 In conclusion, high-fat induced up-regulation of Pin1 expression reduces Diabetic Subjects AMPK activity, and this mechanism may be involved in high-fat diet induced THEODORE P. CIARALDI, ALEXANDER J. RYAN, SUSAN A. PHILLIPS, SUNITA mitochondrial activity reduction and lipid accumulation in muscle. BAXI, SUNDER R. MUDALIAR, ROBERT R. HENRY, La Jolla, CA, San Diego, CA Chronic low-grade infl ammation, including elevated infl ammatory cell 1848-P infi ltration in adipose tissue and skeletal muscle, is a common feature Salicylate Ameliorates High-Fat Diet-induced Microvascular and of obesity and Type 2 Diabetes (T2D). To compare the direct effects of Metabolic Insulin Resistance microenvironments representing infection (II) and disordered metabolism LINA ZHAO, ZHUO FU, KEVIN AYLOR, EUGENE J. BARRETT, ZHENQI LIU, Char- (MI) induced infl ammation, cultured myotubes (MT) prepared from non- lottes ville, VA diabetic (ND) and T2D subjects (n=4 each) were treated with LPS (1 µg/ Microvascular insulin resistance contributes to metabolic insulin mL), or combined hyperglycemic (11 mM)/hyperinsulinemic (150 pM)/ resistance and chronic infl ammation plays an important role in both. hyperlipidemic (0.4 mM palmitate) conditions to represent infectious and Salicylate has a potent anti-infl ammatory action and been shown to improve T2D microenvironments, respectively. Conditioned media was collected after vascular function and insulin sensitivity. 1 and 2 d for analysis of myokine secretion and cells extracted to evaluate To examine whether salicylate alters muscle microvascular and metabolic signaling pathways involved in infl ammation. LPS treatment induced large insulin actions, adult male SD rats were fed a high fat diet (HFD, 60% (all p<0.05) increases in the secretion of GROα (1832 ± 382% of control, d1 saturated fat) for 3 days, 1 week, 2 weeks or 4 weeks with or without sodium values), IL6 (647 ± 132%) and IL8 (4020 ± 1392%). The T2D condition (MI) had salicylate oral gavage (120 mg/kg/day). Additional rats were fed a low fat much smaller effects on the secretion of GROα (188 ± 32%), IL6 (112 ± 22%) diet (LFD, 10% saturated fat) as controls. Overnight fasted rats then received or IL8 (203 ± 52%), all signifi cantly less than the paired response to LPS. a 2-hr euglycemic insulin clamp (3 mU/kg/min). Muscle microvascular Changes in MCP1 and VEGF in response to either condition were modest. blood volume, velocity and fl ow were quantifi ed using contrast-enhanced Relative myokine responses to LPS treatment were similar in ND and T2D ultrasound before and after insulin infusion. Insulin-stimulated steady-state MT. However, T2D MT displayed augmented responses to MI treatment for whole body glucose disposal (ISGD) rates were calculated. GROα (138 vs. 255%, p=0.063) and IL8 (108 vs. 329%, p=0.014) secretion, Neither HFD nor salicylate treatment altered body weights. Compared with ND vs. T2D. LPS treatment led to increases in the phosphorylation of both LFD controls, plasma levels of triglyceride, cholesterol and insulin signifi cantly p38 and p44/42 MAPK, while MI had no effect. IkBα protein expression was increased in HFD groups at week 4 (p<0.05 for all). HFD feeding decreased unaltered by either treatment in both ND and T2D MT. In summary: 1) In insulin-mediated microvascular recruitment at day 3 (by ~30%, p<0.05) the absence of infl ammatory cells, LPS can directly stimulate the release of and abolished it at week 1-4 (p<0.05 for all). On the contrary, ISGD did not pro-infl ammatory myokines, 2) A T2D-like metabolic environment induces a signifi cantly decrease at day 3 and then began to progressively decline (by more modest pro-infl ammatory response, 3) T2D MT exhibit an exacerbated ~20% at week 1, ~40% at week 2, and ~60% at week 4, p<0.05 for all) in infl ammatory response to T2D-like conditions. We conclude that infectious HFD fed rats. Salicylate treatment restored microvascular insulin responses at and metabolic infl ammation exert quantitatively different effects on myokine week 1 and this effect persisted at weeks 2 and 4. Salicylate treatment did not secretion. improve ISGD until week 2 (by ~50%) and 4 (by ~70%)(p<0.05 for both). Supported By: U.S. Dept. of Veterans Affairs In conclusion, microvascular insulin resistance develops earlier than metabolic insulin resistance during HFD feeding and salicylate treatment 1851-P ameliorates both. Our data suggest that prevention of microvascular insulin Muscle Phospholipid Synthesis Regulates Insulin Sensitivity and resistance may contribute to the prevention of metabolic insulin resistance Contractile Function and diabetes. KATSUHIKO FUNAI, IRFAN J. LODHI, LARRY D. SPEARS, LI YIN, HAOWEI SONG, Supported By: ADA (1-11-CT-30) SAMUEL KLEIN, CLAY F. SEMENKOVICH, St. Louis, MO Skeletal muscle insulin resistance is an early and perhaps essential defect 1849-P in the development of type 2 diabetes. Molecular mechanisms responsible for Insulin and IGF-1 Signaling in Muscle Is Required for Normal Muscle the reduction in skeletal muscle insulin-stimulated glucose disposal remain Growth but Not for Normal Glucose Tolerance elusive. Recent fi ndings implicate lipogenesis in this process through effects BRIAN T. O’NEILL, MICHAEL F. HIRSHMAN, JIA LI, GRAHAM SMYTH, C. RONALD on sarcoplasmic reticulum (SR) phospholipid composition. We perturbed KAHN, Boston, MA choline/ethanolamine phosphotransferase-1 (CEPT1), the terminal enzyme Skeletal muscle insulin resistance is an early and dominant feature of the in the Kennedy pathway of phospholipid synthesis, to test the hypothesis pathogenesis of type 2 diabetes, yet deletion of muscle insulin receptors (IR) that SR phospholipid composition regulates muscle insulin sensitivity and in mice does not cause diabetes. To determine to what extent IGF-1 receptor contractile function. In C2C12 cells, lentivirus-mediated knockdown of (IGF1R) signaling compensates for the loss of IR in muscle to maintain CEPT1 altered SR phospholipid composition and calcium fl ux. In mice, diet- Integrated POSTERS glucose homeostasis and muscle mass, we generated mice with combined induced obesity increased muscle CEPT1 expression and decreased insulin muscle-specifi c knockout of IGF1R and IR. These MIGIRKO mice showed a sensitivity. In obese humans, surgery-induced weight loss was associated Physiology/Obesity >60% decrease in muscle mass involving all skeletal muscles, an effect not with decreased skeletal muscle CEPT1 expression. In humans spanning a obser ved in mice lacking either IR or IGF1R alone. MIGIRKO mice also displayed spectrum of metabolic health, muscle CEPT1 expression was inversely fasting hypoglycemia due to increased basal glucose uptake in muscle. This correlated with glucose disposal. Mice with skeletal muscle-specifi c was secondary to increased protein levels of glucose transporters 1 and 4 knockout of CEPT1 exhibited improved insulin sensitivity as measured by and increased glucose transporter translocation, as well as increased total glucose and insulin tolerance tests, hyperinsulinemic-euglycemic clamps, and activated Akt levels due to altered rates of Akt degradation. By contrast, and 2-deoxyglucose uptake in isolated muscles. In CEPT1-defi cient muscles, despite the complete lack of insulin/IGF-1 signaling in muscle, MIGIRKO an altered SR phospholipid milieu decreased sarco/endoplasmic reticulum mice displayed normal glucose tolerance and insulin tolerance. Indeed, even Ca2+ ATPase (SERCA)-dependent calcium uptake. This led to the activation after high fat diet challenge, MIGIRKO mice did not develop diabetes and of calcium signaling pathways, known to improve insulin sensitivity. had only moderate glucose intolerance similar to control mice on high fat However, dysregulated muscle SR calcium handling made these mice weak diet. On the other hand, transgenic overexpression of a dominant negative, and exercise intolerant. These results suggest that high fat feeding and

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A476 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE obesity induce CEPT1 to ensure SR compositional remodeling that preserves muscle vascular sensitivity to AII and ii) local muscle AII infusion does not contractile function at the expense of insulin sensitivity. directly cause microvascular or metabolic insulin resistance in muscle, but worsens insulin resistance during HS intake. These data suggest that insulin 1852-P resistance that is specifi cally associated with increased dietary sodium may Muscle Remodeling Is Necessary for Exercise Training to Decrease be ameliorated by AII blockade. Insulin Resistance in Prediabetic Metabolic Syndrome Supported By: University of Tasmania CHARLES A. STUART, MICHELLE L. LEE, BRIAN M. CARTWRIGHT, MARK A. SOUTH, MARY E. HOWELL, MICHAEL W. RAMSEY, MICHAEL H. STONE, Johnson City, TN 1854-P In the absence of weight loss, the impact of endurance or strength training Ceramide Mediates Cigarette Smoke-induced Skeletal Muscle on insulin resistance has not been consistently benefi cial. To determine the Metabolic Disruption subject characteristics that identify those who would benefi t from exercise MIKAYLA THATCHER, TREVOR TIPPETTS, IAN JOHNSON, ZACHARY HOLUB, training, we analyzed data from thirty metabolic syndrome men and women MICHAEL NELSON, DUANE WINDEN, PAUL REYNOLDS, BENJAMIN T. BIKMAN, who had undergone eight weeks of either strength or endurance training with Provo, UT pre- and post- euglycemic insulin clamps and muscle biopsies. No weight Cigarette smokers are known to be insulin resistant and smoke exposure loss was permitted. Fourteen subjects improved their insulin responsiveness increases lung infl ammation and ceramide biosynthesis. We hypothesized (Group A) whereas sixteen were unchanged or worsened (Group B). Pre- that ceramides are released from the lung with cigarette smoke exposure training muscle from Group A had more (44±5% vs. 26±4%) mixed slow- and and result in elevated skeletal muscle ceramide levels, resulting in insulin fast-twitch myosin heavy chain containing fi bers (type IIa) and fi ber types I, resistance and altered mitochondrial respiration. Employing cell and animal IIa, and IIx had 15 to 26% smaller cross-sectional areas. Group A subjects models, we explored the effect of cigarette smoke on muscle cell insulin had less insulin receptors and activated AMPK in pre-training muscle. Group signaling and mitochondrial respiration. Muscle cells were treated with A post-training muscle had decreased the type IIa and increased the IIx fi ber conditioned media from cigarette smoke extract (CSE)-exposed lung cells, portion, suggesting training caused a shift from IIa to IIx. In Group A, the followed by analysis of ceramides and assessment of insulin signaling size of all fi ber types increased during training, but Group B muscle fi ber and mitochondrial function. Muscle cells treated with CSE-exposed composition and fi ber size did not change. The same changes were seen with conditioned medium were completely unresponsive to insulin stimulation either type of training. Change in muscle fi ber composition and increase in and mitochondrial respiration was severely blunted. These effects were fi ber size was necessary for insulin responsiveness to improve in metabolic mitigated when lung cells were treated with the ceramide inhibitor syndrome subjects after exercise training. myriocin prior to and during CSE exposure. Mice were exposed to daily cigarette smoke for 12 wk and received either PBS or myriocin injections. At the conclusion of the period, insulin sensitivity was determined, as well as muscle mitochondrial and ceramides analysis. Daily cigarette smoke exposure in PBS-, but not myriocin-treated animals, resulted in elevated muscle ceramide levels, reduced ceramide mitochondrial respiration, and elevated HOMA scores. Thus, ceramide inhibition appears to prevent whole body and skeletal muscle metabolic disruption with smoke exposure.

1855-P Ceramide Is Necessary for Cigarette Smoke-induced Reduced Heart Mitochondrial Function TREVOR TIPPETTS, DUANE WINDEN, TY CONDIE, PAUL REYNOLDS, BENJAMIN T. BIKMAN, Provo, UT Cigarette smoke exposure increases risk of multiple cardiovascular complications, including heart failure, heart attack, and atherosclerosis. Given that ceramides are both produced in the lung with smoke exposure and are known to alter cardiovascular function, we hypothesized that ceramides may mediate the deleterious effect of smoking on heart mitochondrial function. Employing cell and animal models, we explored the effect of cigarette smoke on myocardial cell mitochondrial function. Muscle 1853-P cells were treated with conditioned media from cigarette smoke extract Local Hindleg Angiotensin II Infusion Impairs Insulin-mediated Micro - (CSE)-exposed lung cells, followed by analysis of ceramides and assessment vascular and Metabolic Actions in High- but Not Normal-Sodium Fed of mitochondrial function. H9C2 cardiomyocytes cells treated with CSE- Rats exposed conditioned medium suffered a dramatic reduction in mitochondrial DINO PREMILOVAC, STEPHEN M. RICHARDS, STEPHEN RATTIGAN, MICHELLE A. respiration. These effects were mitigated when lung cells were treated KESKE, Hobart, Australia with the ceramide inhibitor myriocin prior to and during CSE exposure. Mice Impairments in microvascular actions of insulin in muscle can directly were exposed to daily cigarette smoke for 12 wk and received either PBS cause insulin resistance. Enhanced angiotensin II (AII) signaling has been or myriocin injections. At the conclusion of the period, heart ceramide and implicated in the development of insulin resistance, however the local muscle mitochondrial function was determined. Daily cigarette smoke exposure in effects of AII on insulin-mediated microvascular and metabolic actions are PBS-, but not myriocin-treated animals, resulted in elevated heart ceramide unknown. To study this, we fed male Sprague Dawley rats either normal levels and reduced ceramide mitochondrial respiration. Thus, ceramide (NS, 0.3% NaCl wt/wt) or high (8% NaCl wt/wt) sodium diets for 4 weeks. inhibition appears to prevent heart mitochondrial dysfunction with smoke Overnight fasted anesthetized rats were subjected to either constant-fl ow exposure. perfused rat hindlimb ex vivo or hyperinsulinemic euglycemic clamp in vivo (10 Integrated mU/min/kg x 2hrs) studies with concomitant local hindleg AII infusion. HS- 1856-P POSTERS fed rats had signifi cantly (p<0.05) augmented AII-mediated vasoconstriction Muscle Lipoprotein Lipase Activity Does Not Predict VLDL-TG Fatty Physiology/Obesity (2.5-fold, 2.2-fold and1.8-fold at 1, 3 and 10nM AII, respectively) during Acid Oxidation during Exercise hindlimb perfusion ex vivo compared to NS. Infusion of 5nM AII into one ESBEN SØNDERGAARD, IBEN RAHBEK, LARS P. SØRENSEN, LARS C. GORMSEN, hindleg (retrograde via epigastric artery) of NS-fed rats started 15 min SØREN NIELSEN, Aarhus, Denmark prior to hyperinsulinemic euglycemic clamps did not alter insulin-mediated Lipoprotein lipase (LPL) activity is considered the rate-limiting step in changes in femoral artery blood fl ow, microvascular perfusion or muscle facilitating fatty acid (FA) uptake from lipoproteins. However, previous glucose uptake when compared to the contralateral leg. HS-fed rats had studies have failed to demonstrate an association between LPL activity and impaired insulin-mediated microvascular and metabolic actions in muscle. VLDL-TG FA uptake in muscle and adipose tissue. The aim of the present Local muscle AII infusion in HS-fed rats did not alter femoral artery responses study was to determine whether muscle LPL activity predicts VLDL-TG FA to insulin, but further impaired insulin-stimulated muscle microvascular oxidation during exercise. perfusion (1.3-fold, p=0.022) and muscle glucose uptake (1.5-fold, p=0.001) Sixteen healthy, lean subjects (8 men and 8 women) were examined after compared to the contralateral leg. We conclude that i) HS feeding augments an overnight fast during rest and exercise (90 min at 50% of VO2-max).

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A477 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

Heparin-releasable LPL activity was measured in muscle and adipose tissue results show that lack of SIRT3 in HF, but not chow fed mice, profoundly biopsies. Breath 14CO2 was measured after a primed-constant infusion of impairs insulin-stimulated muscle glucose uptake and respiration creating ex-vivo labelled [14C]-triolein VLDL-TG. Fractional VLDL-TG storage during an increased reliance on fatty acids. We conclude that SIRT3 protects rest was measured in adipose tissue biopsies after 4½ hours infusion of against severe dietary insulin resistance by facilitating glucose disposal and VLDL-TG tracer. mitochondrial function. Muscle LPL activity was not affected by exercise (rest: 0.44 (0.27-0.94) Supported By: NIH (DK54902) µmol FFA/hr/g; exercise: 0.40 (0.17-1.11) µmol FFA/hr/g). No association was observed between muscle LPL activity and VLDL-TG oxidation, neither in the 1859-P basal state (r=-0.31, NS) nor during exercise (r=0.06, NS). In addition, exercise Evaluation and Comparison of Epicatechin Epimer Effects on Skeletal did not affect abdominal (rest: 0.36 (0.25-0.49) µmol FFA/hr/g; exercise: 0.37 Muscle Structure, Function, and Regulators of Metabolism (0.24-0.58) µmol FFA/hr/g) or femoral (rest: 0.35 (0.25-0.50) µmol FFA/hr/g; ISRAEL RAMIREZ-SANCHEZ, GUILLERMO CEBALLOS, ALDO MORENO-ULLOA, exercise: 0.34 (0.20-0.88) µmol FFA/hr/g) adipose tissue LPL activity. The THEODORE P. CIARALDI, ROBERT R. HENRY, FRANCISCO VILLARREAL, Mexico basal adipose tissue fractional VLDL-TG storage (abdominal.13±9%; femoral City, Mexico, San Diego, CA, La Jolla, CA 17%±10%) was not different between adipose tissue regions or associated We reported on the benefi cial effects of fl avanol rich cocoa on skeletal with adipose tissue LPL activity. No sex-specifi c differences was observed muscle (SkM) structure and function of type 2 diabetic and heart failure in LPL activity, VLDL-TG oxidation or VLDL-TG storage. patients. We ascribe such effects to the presence of the fl avanol epicatechin Muscle LPL activity does not predict VLDL-TG oxidation during rest or (Epi) which can be found in its (-) and (+) epimer confi gurations. Whereas (-)- exercise. In addition, no association was observed between adipose tissue Epi is abundantly present in natural products such as cocoa, (+)-Epi is rare. LPL activity and VLDL-TG storage during rest. This suggests that LPL is We previously reported that oral supplementation of 1 mg/kg/day BID of present in excess of what is needed to facilitate lipid uptake, even during (-)-Epi leads to signifi cant increases in exercise capacity, SkM and cardiac exercise where lipid oxidation is stimulated. mitochondria volume and cristae abundance as well as capillarity. The effects of (-)-Epi can be replicated in cultured C2C12 cells evidencing increases in 1857-P protein levels of known regulators of mitochondrial biogenesis (MiB) such as GLP-1(32-36)amide Pentapeptide Regulates Oxidative Phosphoryla- AMPK and PGC1α. However, nothing is known about the relative potencies tion by Activating AMPK Signaling and Raising the NAD+/NADH of the two epimer forms of Epi. To address this issue studies were performed Ratio in C2C12 Myotubes in C2C12 cells and in 6 month old male mice. C2C12 cells were treated with JOEL F. HABENER, EVA TOMAS, Boston, MA incremental doses of (-)- and (+)-Epi for 48 h and effects on AMPK and Infusions of a pentapeptide-derived from GLP-1, GLP-1(32-36)amide, PGC1α protein levels determined. Results indicate signifi cant increases in into diet-induced obese mice for 16 weeks attenuated weight gain and the both proteins that reached maximal effects with (-)-Epi at 100 nM whereas development of manifestations of the metabolic syndrome by increasing the those of (+)-Epi at 10 nm (i.e. 10 fold increase in potency). Male mice (n=3-5/ rates of oxygen consumption (determined by indirect calorimetry). Skeletal group) were treated for 2 weeks via oral gavage with either water (control), muscle is an important contributor to the regulation of whole body energy (-)-Epi (1 mg/kg/day), (+)-Epi (0.1 mg/kg/day), (+)-Epi (0.3 mg/kg/day) or (+)- metabolism. Therefore, we examined the actions of GLP-1(32-36)amide on Epi (1 mg/kg/day). Results evidenced dose dependent increases in treadmill muscle oxidative phosphorylation. Incubation of GLP-1(32-36)amide in C2C12 performance with (+)-Epi with similar effects noted with the 0.3 mg/kg/day myotubes activated AMPK signaling by increasing the phosphorylation of dose as with 1 mg/kg/day of (-)-Epi. Signifi cant dose dependent increases of AMPK and its downstream target ACC in a dose-dependent manner. These 20-100% were observed in SkM, p-LKB1, AMPK and PGC1α protein levels. changes were also accompanied by increases in the intracellular ratio of The comparison of effects evidenced ~3 fold greater potency of (+)-Epi NAD+/NADH. Likewise, in 16 hour palmitate-induced impaired oxidative effects vs. (-)-Epi. In conclusion, the (+)- epimer form of Epi evidences greater metabolism GLP-1(32-36)amide incubation restored both the decreased potency vs. its (-)- confi guration which translates into greater organ function AMPK and ACC phosphorylation levels as well as NAD+/NADH ratio. These and likely mitochondrial levels. effects of GLP-1(32-36)amide were blunted when cells were pre-incubated Supported By: NIDDK with the fatty acid inhibitor Etomoxir. In summary, in C2C12 myotubes GLP- 1(32-36)amide regulates oxidative phosphorylation by increasing NAD+/ 1860-P NADH ratio, AMPK activity and fatty oxidation as seen by the increased Inducible Opa-1 Deletion in Skeletal Muscle Results in Mitochondrial phosphorylation of ACC. Dysfunction and Reduced Muscle Mass in Mice KAREN DE JESUS OLIVEIRA, RENATA O. PEREIRA, KENSUKE TSUSHIMA, 1858-P GABRIELA S. MONTEIRO, KARLA M. PIRES, E. DALE ABEL, Salt Lake City, UT, Iowa SIRT3 Is Crucial for Maintaining Skeletal Muscle Insulin Action and City, IA Mitochondrial Function in High Fat Fed Mice Opa1 an inner membrane mitochondrial protein plays a fundamental LOUISE LANTIER, ASHLEY S. WILLIAMS, FREYJA D. JAMES, DEANNA P. BRACY, role in mitochondrial fusion, quality control and energetics. Germline DAVID R. GIUS, DAVID H. WASSERMAN, Nashville, TN, Chicago, IL haploinsuffi ciency of Opa1 impairs mitochondrial biogenesis in skeletal Sirtuins, a family of NAD+ dependent deacetylases, are involved in muscle, but paradoxically improves endurance exercise capacity. Given the balance between acetylated and deacetylated proteins. Protein potential limitations of germline Opa1 defi ciency we generated mice with hyperacetylation is associated with glucose intolerance, raising the inducible deletion of the Opa1 gene in skeletal muscle of adult C57Bl6 question of the role of Sirtuins in the pathogenesis of insulin resistance mice (Opa1SMKO) by crossing Opa1 fl oxed mice with HSA-Cre (ERT2) mice. (IR). In particular, SIRT3 is a mitochondrial deacetylase that has been Four-week-old mice were treated with tamoxifen for 5 days to induce linked to energy homeostasis and mitochondrial function. We hypothesized recombination, resulting in a 60% reduction in Opa1 protein content, 8 that mitochondrial acetylation state is key to the deleterious effects of a weeks after treatment (13-week-old mice). Body weight was decreased in high fat diet (HF) on IR, and that SIRT3 knockout (KO) mice have increased 12-week-old Opa1SMKO mice, which was accompanied by reduced lean diet-induced IR. One week following catheterization of the jugular vein mass and increased fat mass in females. Changes in body composition were and carotid artery, hyperinsulinemic (4 mU.kg-1.min-1) euglycemic clamps corroborated by a reduction in soleus weight in Opa1SMKO females at 18 Integrated POSTERS combined with isotopic ([3-3H]glucose, 2[14C]deoxyglucose) methods were weeks and in males at 20 weeks of age. In both genders basal and maximally performed. Results show for the fi rst time that while chow fed SIRT3 KO stimulated mitochondria respirations in soleus muscle were preserved 8 Physiology/Obesity mice have no apparent metabolic phenotype, HF KO mice exhibit increased weeks after Opa1 reduction (13-wk-old mice), although ATP synthesis rates IR compared to their WT littermates, as evidenced by a 30% decrease in were reduced. In 20-week old mice, both mitochondrial respiration and ATP glucose infusion rate. This is due to skeletal muscle IR, as Rg, an index of synthesis were reduced in Opa1 SMKO mice. Western blots in gastrocnemius muscle glucose uptake, is decreased by 50% in KO mice, while suppression showed a 2-fold reduction in Complex I (NDUFA9) content in 13-week old of liver glucose production is unaffected. High resolution respirometry on Opa1SMKO mice, relative to controls. Autophagy fl ux was increased in permeabilized fi bers shows that TCA cycle substrate based respiration is 13-week old Opa1SMKO females, as evidenced by a 25% increase in LC3II decreased by 30% in fi bers from HF SIRT3 KO mice while fatty acid based and 25% reduction in p62 content. Mitophagy-related proteins Bnip3 and respiration is increased by 40%, refl ecting a fuel switch from glucose to fatty Parkin were also induced in Opa1SMKO females. In contrast, autophagy acids. Consistent with reduced muscle glucose uptake, hexokinase II (HKII) proteins were unchanged in males at this time point. Thus females are binding to the mitochondria is decreased in muscle fi bers from HF SIRT3 more susceptible to the consequences of OPA1 defi ciency on mitophagy/ KO mice, suggesting decreased overall HKII activity. Taken together, these autophagy. In conclusion, Opa1 is critical for maintaining skeletal muscle

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A478 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE mitochondrial function and regulates muscle mass by modulating skeletal 1863-P muscle autophagy. Dipeptidyl Peptidase-4 Inhibitor Improved Exercise Capacity in Supported By: NIH; AHA Association with Switching to Oxidative Skeletal Muscle Fiber Type in Mice with Post-infarct Heart Failure 1861-P YOSHIHIRO MASAKI, SHINTARO KINUGAWA, SHINGO TAKADA, TOMOYASU Distinct Effects of Eicosapentaenoic Acid (EPA) and Docosa hexae- KADOGUCHI, ARATA FUKUSHIMA, TSUNEAKI HOMMA, TAKAAKI FURIHATA, noic Acid (DHA) on Muscle and Liver RYOICHI KATSUYAMA, TADASHI SUGA, MASASHIGE TAKAHASHI, TAKASHI IAN LANZA, SURENDRA DASARI, CARRIE-JO HEPPELMANN, DANIEL JAKAITIS, YOKOTA, SHOUJI MATSUSHIMA, KOICHI OKITA, HIROYUKI TSUTSUI, Sapporo, JILL SCHIMKE, KATHERINE KLAUS, DAWN MORSE, MATTHEW L. JOHNSON, Japan, Ebetsu, Japan Rochester, MN Exercise capacity is reduced in heart failure (HF) largely due to the Insulin resistance, mitochondrial dysfunction, and sarcopenia are impairment of skeletal muscle function. Glucagon-like peptide-1 (GLP-1) hallmarks of aging, and omega-3 fatty acids (n-3 PUFAs) have been studied has been shown to improve exercise tolerance in HF patients. However, the as potential countermeasures. However, few studies have compared the effects of dipeptidyl peptidase (DPP)-4 inhibitor on the exercise capacity biological effects of eicosapentaenoic acid (EPA) and docosahexaenoic and skeletal muscle characteristics have not been elucidated. Myocardial acid (DHA). Young (6 mo) and old (24 mo) mice were fed control chow or infarction (MI) was created in male C57BL/6J mice (n=20) by ligating the chow containing purifi ed EPA or DHA (3.4% kcals) for 10 weeks. RNA deep left coronary artery, and sham operation was performed (n=20). They were sequencing revealed that EPA and DHA exhibited distinct transcriptional divided into 2 groups of the treatment with or without DPP-4 inhibitor, MK- patterns. EPA infl uenced genes related to energy production, lipid 0626 (sitagliptin analog, 1 mg/kg/day) for 4 weeks. MK-0626 signifi cantly metabolism, and amino acid metabolism in muscle but not liver. In contrast, inhibited plasma DPP-4 activity and increased plasma active GLP-1 levels by the top networks infl uenced by DHA were related to cellular maintenance 1.6 folds. MK-0626 did not affect infarct size from MI and MI+MK-0626 mice in muscle and infl ammation in liver. We then focused our attention on and left ventricular function evaluated by echocardiography. In contrast, mitochondrial biology. Mitochondrial function, measured by high-resolution exercise capacity, measured by treadmill test with expired gas analysis, respirometry, was reduced in old compared to young mice. EPA increased was reduced in MI mice compared to sham mice (work 17±1 vs. 29±1 J and mitochondrial function in skeletal muscle of old mice, whereas DHA increased peak VO2 140±5 vs. 161±3 mL/kg/min) and was signifi cantly ameliorated in mitochondrial function in liver only. Mitochondrial DNA copy number, a MI+MK-0626 (work 22±1 J and peak VO2 158±3 mL/kg/min). A shift toward marker of mitochondrial abundance, was lower in old mice and unaffected fast twitch myosin fi ber type was observed in the skeletal muscle from MI by EPA or DHA. Furthermore, mass spectrometry-based large-scale shotgun mice, and this change was reversed by MK-0626, expressed as the increase proteomics revealed that the abundance of mitochondrial proteins was in type I fi ber and the decrease in type IIb fi ber. Citrate synthase activity lower in mice that received EPA or DHA compared to control fed mice. As a and mitochondrial quantity were reduced in the skeletal muscle from MI measure of mitochondrial biogenesis, mitochondrial protein synthesis rates mice and this decrease was normalized in MI+MK-0626, in association with were measured by injecting a bolus of 13C6 phenylalanine and monitoring the improvement of mitochondrial biogenesis. These effects of MK-0626 on the rate of tracer incorporation into the mitochondrial proteome by mass skeletal muscle fi ber type and mitochondrial function were not observed in spectrometry. Neither EPA nor DHA signifi cantly changed mitochondrial sham mice. The administration of MK-0626 into MI mice improved exercise protein synthesis in muscle or liver. Taken together, these data indicate that capacity and normalized the skeletal muscle fi ber type switch. DPP-4 EPA and DHA enhance mitochondrial function in a tissue-specifi c manner, inhibitor may be a novel therapeutic agent against exercise intolerance seen but do so in the absence of any clear changes in mitochondrial abundance or in HF by improving mitochondrial biogenesis. biogenesis. These data demonstrate that EPA and DHA have distinct effects on tissue biology, the transcriptome, and the proteome. 1864-P AMPK Regulates In Vivo Exercise-induced Fatty Acid Oxidation in 1862-P Skeletal Muscle Twelve Weeks of Pioglitazone Treatment Improves Insulin Sensi- JOACHIM FENTZ, RASMUS KJØBSTED, JESPER BIRK, ANDREAS BØRSTING tivity, Lipid Utilization, and Metabolic Flexibility, but Not Maximal JORDY, JACOB JEPPESEN, ANDERS GUDIKSEN, HENRIETTE PILEGAARD, BENTE ATP Synthesis Capacity KIENS, NIELS F.P. JESSEN, MARC FORETZ, BENOIT VIOLLET, JØRGEN WOJTA- SUDIP BAJPEYI, KEVIN CONLEY, BRADLEY R. NEWCOMER, SHARON JUBRIAS, SZEWSKI, Copenhagen, Denmark, Aarhus, Denmark, Paris, France CECIIA GAMBOA, KORI MURRAY, LAUREN M. SPARKS, STEVEN R. SMITH, El Studies investigating the importance of adenosine monophosphate-activated Paso, TX, Seattle, WA, Birmingham, AL, Baton Rouge, LA, Orlando, FL, Winter Park, FL protein kinase (AMPK) in carbohydrate and fatty acid (FA) oxidation with Pioglitazone (Pio) is known to improve insulin sensitivity in skeletal contraction/exercise in skeletal muscle have utilized a wide range of genetic muscle. However, the role of Pio on skeletal muscle lipid metabolism and mouse models and methodologies. These studies have yielded confl icting results skeletal muscle oxidative capacity is not clear. The purpose of this study which to some extent may pertain to redundant AMPKα1 signaling. was to determine the effects of 12 weeks of pioglitazone treatment Using a novel model lacking both AMPKα1 and -α2 in skeletal muscle on insulin sensitivity measured by hyperinsulinemic euglycemic clamp, specifi cally (mdKO) we approached the question of the signifi cance of the intramyocellular lipid content (IMCL) measured by proton magnetic resonance AMPKα catalytic subunits in regulation of glucose and FA utilization by spectroscopy, metabolic fl exibility measured by calculating delta RQ during contraction/exercise by performing a mouse expression array analysis the steady state of the clamp and muscle maximal ATP synthesis capacity on extracted mRNA from WT and mdKO muscles. This analysis proposed (ATPmax) measured by 31P Magnetic Resonance Spectroscopy. Twenty-four a major role for AMPK in FA oxidation. Performing an ex vivo contraction participants with type 2 diabetes (13M/11F 53.38 ± 2.1 years; BMI 36.47 ± protocol in skeletal muscles of WT and mdKO mice results revealed that 1.1) were randomized to either a placebo (n=8) or a Pio (n=16) group. After 12 basal and contraction-induced FA oxidation were signifi cantly reduced in weeks of Pio treatment, there was an increase in insulin sensitivity (Placebo mdKO mice. Further the mdKO had a markedly higher RER than WT (mdKO 2.2% vs. Pio 90.1% increase; p=<0.05) and an increase in metabolic fl exibility 0.87 ±0.02, WT 0.81 ±0.01; p<0.001) during in vivo exercise performed at the (delta RQ; Placebo 0.063±0.01 to 0.033 ± 0.01; p=0.02 vs. Pio 0.061 ± 0.01 same relative exercise intensity, supporting a key role of AMPK in exercise- to 0.088 ± 0.02; p=0.03). Pio treatment decreased plasma free fatty acids induced FA oxidation in vivo. Skeletal muscle glucose uptake during in vivo Integrated (Placebo vs. Pio 16% increase vs. -7.7% decrease; p=0.04) and IMCL content exercise was signifi cantly increased (or trended to) in mdKO mice. POSTERS in gastrocnemius (Placebo -1.1% vs. Pio -56.5% decrease, p=0.005), Soleus These results are interesting as another mouse model lacking both the (Placebo -0.4% vs. Pio -21.8% decrease, p<0.05) and Tibialis Anterior muscle AMPKβ1 and -β2 in skeletal muscles displays a lower in vivo exercise- Physiology/Obesity (Placebo -1.1% vs. Pio -39.5% decrease, p=0.003). There was no change induced glucose clearance and RER compared to WT, suggesting highly in ATPmax after Pio treatment (Placebo 15.8% vs. Pio 6.1%, p=0.6). These distinct roles of the different AMPK subunits. Moreover we have recently results suggest that 12 weeks of pioglitazone treatment improves insulin published that the AMPKα2 KO mouse does not show alterations in RER sensitivity, metabolic fl exibility and lipid utilization independent of any during in vivo exercise. Together with our present results this suggests that improvement in maximal ATP synthesis capacity in skeletal muscle. AMPKα1 may be suffi cient to maintain a normal skeletal muscle FA oxidation with ex vivo contractions and RER during in vivo exercise. In summary our data show that the catalytically active AMPKα subunits play an important role in FA utilization both with ex vivo contractions and during in vivo exercise. Supported By: Danish National Research Council; Novo Nordisk Foundation

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A479 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

1865-P 1867-P Impaired Cytosolic NADH Shuttling and Elevated UCP3 Contribute Heat Shock Protein 90 Beta (Hsp90 Beta) Isoform Represents a to Ineffi cient Citric Acid Cycle Flux Support of Cardiac Work in Novel 1st in Class Target Regulating Systemic Energy Metabolism Diabetic Hearts Following Ischemia/Reperfusion for Treatment of Metabolic Diseases NATASHA H. BANKE, XUERONG WANG, E. DOUGLAS LEWANDOWSKI, Chicago, IL ENXUAN JING, PRAGALATH SUNDARARAJAN, SAMANTHA FOWLER, VIVEK Diabetic myocardium has increased susceptibility to ischemia/reperfusion VISHNUDAS, RANGAPRASAD SARANGARAJAN, NIVEN R. NARAIN, Boston, (IR) damage. This study examined the effi ciency of citric acid cycle (CAC) fl ux MA and the transfer of cytosolic reducing equivalents into the mitochondria for Hsp90beta was identifi ed as a critical node in Bayesian metabolic oxidative support of cardiac work following IR in hearts of c57bl/6 (NORM) disease network derived by the interrogation of the biology underlying and type 2 diabetic, db/db mouse (DM2) hearts. Flux through the CAC and diabetes in a data driven manner using a proprietary platform integrating malate-aspartate shuttle (MA) was monitored in isolated hearts perfused pan-omic data, mitochondrial-centric metabolic fi ngerprint and phenotypic with 0.4 mM 13C palmitate + 10 mM glucose in a 14.1 T NMR magnet with assays capturing metabolic dysregulation reminiscent of obesity/diabetes end point metabolite quantifi cation. The lactate:alanine ratio provided an sequale. Hsp90beta is an ATPase targeting multiple clients including vital index of cystolic NADH/NAD+ (NADHc) which was limited in DM2 hearts by components of insulin signaling and mitochondrial membrane proteins. glucose availability compared to NORM (DM2: 1.0 +/-0.1*, I/R 1.2 +/- 0.1*; However the role of Hsp90beta in metabolism remains unknown. PoC NORM: 2.4 0.3, I/R 2.2 +/- 0.2, *P<0.05 vs. c57bl6). Consequently, MA fl ux studies on Hsp90beta expression in primary human skeletal muscle cells was reduced in DM2 but unchanged by I/R in both DM2 and NORM (DM2: revealed that it is regulated by various metabolic stimuli. C57B/6 HFD 0.79 ± 0.14 µmol/min/dgw, I/R: 0.61 ± 0.10; NORM: 1.3±0.08, I/R: 1.45±0.23). mice were used as a model to knock-down Hsp90beta using Anti-Sense Expression of oxoglutarate malate carrier protein (OMC) of the MA was Oligonucleotide (ASO) technology. The in vivo results demonstrated that elevated in DM2, consistent with a compensatory response to low NADHc. with 4 weeks of IP ASO administration targeting Hsp90beta in key insulin Baseline CAC fl ux per unit work (Vcac/rate pressure product) was similar sensitive tissues signifi cantly improved glucose tolerance compared between NORM and DM2 (Vtca/RPP: NORM= 0.49±0.05; DM2= 0.45±0.08 with control group. Hsp90beta knock-down signifi cantly suppressed fed nmol/mmHg/gdw). With IR, Vcac/RPP was elevated in DM2 compared to glucose levels accompanied by a trend of decreased fed insulin, suggesting NORM, indicating a 20% ineffi ciency in CAC fl ux for IR cardiac function in improved insulin sensitivity. The whole body heat production was also DM2 (NORM= 0.58±0.03 nmol/mmHg/gdw; DM2= 0.7±0.06). The ineffi cient signifi cantly increased in Hsp90beta ASO mice indicating that Hsp90beta support of workload by oxidative metabolism was consistent with elevated knockdown induced systemic metabolic change. Further in vitro study using UCP3 content in DM2 hearts, which responded to IR by increasing UCP3 primary human skeletal muscle myotubes with siRNA mediated Hsp90beta transcript levels. Therefore, DM2 heart exhibits limited glycolytic NADH knockdown revealed that Hsp90beta knockdown induced signifi cantly contributions to oxidative energy production, with compensatory elevation of elevated glycolysis, beta-oxidation, mito-respiration, and insulin stimulated MA protein. Importantly, IR exacerbates pre-existing mitochondrial defects p-Akt, indicating that the effects of Hsp90beta in skeletal muscle is a critical rendering DM2 hearts more susceptible to mitochondrial ineffi ciencies in regulator of substrate metabolism. Taken together, our data provides novel CAC fl ux, including uncoupling and ROS production. evidence that Hsp90beta isoform is a key regulator of skeletal muscle cell Supported By: R37HL49244 metabolism and whole body metabolism and represents a viable target for potential treatment of diabetes. 1866-P Optimal Muscle Group and Reproducibility of Intramyocellular 1868-P Lipid by Magnetic Resonance Spectroscopy in a Pilot Study of Non- Skeletal Muscle Malonyl-CoA Levels as a Biomarker for Pharma- Diabetic and T2DM Subjects cologic Compounds That Modulate the ACC/MCD Pathway GABRIELE E. SONNENBERG, THERESA A. TUTHILL, SANTOS CARVAJAL-GON- WILLIAM P. ESLER, PAUL A. AMOR, R. KIRK MCPHERSON, SANTOS CARVAJAL- ZALEZ, LINDA MORROW, GAVIN HAMILTON, MICHAEL MIDDLETON, WILLIAM GONZALEZ, DAVID A. BEEBE, UDENI YAPA, VINCENT BERNARDO, ERIC RAVUS- P. ESLER, Cambridge, MA, Chula Vista, CA, San Diego, CA SIN, GABRIELE E. SONNENBERG, Cambridge, MA, Groton, CT, Baton Rouge, LA Alterations in lipid metabolism contribute to net ectopic lipid accumulation Malonyl-CoA regulates lipid metabolism by functioning as the rate-limiting in liver and skeletal muscle which are hypothesized to play a causative role substrate for de novo lipogenesis (DNL), and by controlling fatty acid oxidation in the development of insulin resistance. Pharmacologic agents aimed at (FAOX) through CPT-1 inhibition. Dysregulation of malonyl-CoA pathway rebalancing these alterations in T2DM are presently being investigated. is hypothesized to contribute to the pathogenesis of insulin resistance. A reproducible measure of intramyocellular lipid (IMCL) is important to Pharmacologic agents which modulate malonyl-CoA are presently being evaluate these agents and to understand better the pathogenesis of insulin investigated. The ability to reproducibly measure tissue malonyl-CoA level resistance. The main purpose of the study was to validate IMCL quantifi cation is essential to evaluate such agents and to better understand the biology in lower leg muscles, assess measurement reproducibility, and identify of insulin resistance. Tissue malonyl-CoA levels were measured using LC- muscle groups most suitable for future studies. IMCL was assessed twice MS-MS. Skeletal muscle and liver malonyl-CoA levels were suppressed in a by proton magnetic resonance spectroscopy (1H-MRS) in each of ten lean dose-dependent manner with up to 80% in rodents (n=8) by oral treatment or obese non-diabetic subjects without repositioning to optimize spectral with an acetyl-CoA carboxylase (ACC) inhibitor. The levels of ACC inhibition fi tting parameters. Ten T2DM subjects each had three examinations: two on were highly related to the increase in whole-body fat oxidation (r2=0.92; the same day with repositioning and one on a second day. Using a 3 Tesla MR decrease in respiratory quotient) and to the inhibition in hepatic DNL (r2=0.94) scanner (GE Signal EXCITE HD, GE Medical Systems), voxels of at least 15 indicating malonyl-CoA is a good target engagement biomarker. However, x 15 x 15 mm3 with minimal visible extramyocellular lipid were chosen from reproducible measurement of human malonyl-CoA levels is hampered by (1) each of three muscle groups (soleus, tibialis anterior and gastrocnemius). low concentrations in human muscle, (2) rapid metabolic turnover and (3) IMCL measurements in all three muscle groups were reproducible with no dependence on nutrient status. Sequential muscle biopsies were collected bias between the two same-day scanning sessions. Variance component 2 hours apart from the vastus lateralis of human volunteers (n=11) following analysis in T2DM subjects showed soleus and tibialis anterior had lower standard meal challenge and immediately frozen in liquid nitrogen. Malonyl- total variability (0.09 and 0.16) compared to gastrocnemius (0.90). Using a CoA levels were quantifi able from all samples (mean 0.21±0.05 nmol/g). Integrated

POSTERS maximum likelihood variance component model and constraining negative While levels varied between subjects, within subject assessment of estimates to zero, the between-day variability (% of total variability) for sequential samples was highly reproducible (intra class correlation 0.76). As Physiology/Obesity IMCL were 0.000 (0%) in soleus), and 0.054 (33.8%) in tibialis anterior, and a result, a 20% change in skeletal muscle malonyl-CoA would be detectable 0.544 (61.5%) in gastrocnemius. These observations demonstrate that IMCL in 5 subjects with 80% power. These observations demonstrate that tissue measurement in soleus and tibialis anterior muscles could serve as a reliable levels of malonyl-CoA could be used to assess target engagement by agents biomarker for detection of treatment-induced IMCL change in patients with aimed at tissue fat balance and thus insulin sensitivity. T2DM. Supported By: Pfi zer, Inc. Supported By: Pfi zer, Inc.

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A480 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

1869-P demonstrated that Fyn phosphorylates LKB1, the main upstream AMPK Short-term Insulin Deprivation Activates Skeletal Muscle Autophagy kinase in insulin sensitive target tissues, on Y261 and Y365 residues and but Not Proteasome Activity in Streptozotocin Diabetic Mice regulate its nuclear/cytosolic distribution, leading to modulation of AMPK MATTHEW M. ROBINSON, ADAM R. KONOPKA, DANIEL JAKAITIS, K. SREEKU- activity. However, it was not excluded that Fyn could also directly act on MARAN NAIR, Rochester, MN AMPK itself. Insulin is a key regulator of muscle protein degradation and may affect To investigate this, phospho-proteomic analysis and phospho-specifi c autophagy-lysosome and ubiquitin-proteasome pathways, although the immunoblotting were performed in vivo and demonstrated that Y436 of the contribution of these pathways during insulin deprivation in-vivo remains AMPK alpha subunit is a substrate target of Fyn activity. This fi nding was unknown. We induced diabetes in C57/BL6J mice (n=14) with streptozotocin confi rmed as mutation of Y436 to phenylalanine (AMPK-Y436F) abolished the causing insulin defi ciency then used subcutaneous insulin implants to maintain Fyn-dependent phosphorylation of AMPK. glucose concentrations (4.4±2.1 mmol) for 14 days. Then, implants were removed To evaluate whether phosphorylation on Y436 has an effect on AMPK to induce insulin deprivation (n=7, INS-) for 48-hours compared to insulin activity, we fi rst silenced endogenous AMPK α1 and α2 subunits in HEK293T treatment (n=7, INS+) resulting in elevation of glucose (INS+: 5.7±2.1; INS-: cells using specifi c siRNA technology. Cells were further transfected either 29.1±7.6, p<0.05). Skeletal muscle autophagy was assessed by western blotting with GST-AMPK-WT or GST-AMPK-Y436F. The GST-AMPK-Y436F mutant and confocal microscopy for microtubule-associated protein 1A/1B-light chain 3 displayed increased AMPK activity directly measured in vitro with enhanced (LC3). Proteasome activation was measured using quantitative polymerase chain S79 ACC phosphorylation in vivo. reaction (qPCR) and enzymatic activity using fl uorogenic substrates. Intracellular Taken together, these data suggest that Fyn not only regulates AMPK essential amino acids (EAA) content was measured using mass spectrometry. activity indirectly via its action on LKB1, but also by direct modulation of Insulin deprivation increased (p<0.05) autophagy marker LC3 accumulation AMPK activity by through Y426 phosphorylation of the alpha subunit. (~33%) and LC3-II:I ratio (~73%). Gastrocnemius from INS- mice had higher Supported By: Japan Society for the Promotion of Science mRNA content for the ubiquitin ligase muscle RING-fi nger protein-1 (MuRF1) but no difference in proteasome activity. Intracellular concentrations of valine (~51%) 1872-P and leucine (~61%) were greater (p<0.05) in quadriceps of INS-. Cell culture NFκB Inhibition Improves Glucose Metabolism, but Not Muscle experiments with human myotubes revealed decreased proteasome activity Atrophy, in Aged Mice (~20%, p<0.05) with EAA treatment during high or low insulin concentrations. NING ZHANG, MENGYAO LI, YIQIANG ZHANG, HOLLY VAN REMMEN, KEYT We conclude that insulin deprivation leads to increased autophagy while the E. FISHER, STEVEN N. AUSTAD, STEVEN E. SHOELSON, SOPHIE E. HUSSEY, proteasome pathway is activated at the mRNA level but not enzyme activity, NICOLAS MUSI, San Antonio, TX, Boston, MA, Tampa, FL potentially due to feedback inhibition of the proteasome from EAA. Aging is characterized by impairments in glucose metabolism and muscle Supported By: 5R01DK041973 mass/strength. NFκB is a ubiquitous transcription factor that plays a central role in regulating infl ammatory processes. Because aging is a state of 1870-P low-grade infl ammation, it has been hypothesized that NFκB could play Pharmacological TLR4 Inhibition Protects Against Acute and an important role on glucose metabolism abnormalities and sarcopenia of Chronic Fat-induced Insulin Resistance in Rats aging, partly through proteasome activation. To test this hypothesis, we SOPHIE E. HUSSEY, NING ZHANG, JI LI, NICOLAS MUSI, Tampa, FL, San Antonio, TX studied the effect of NFκB inhibition on aging-induced glucose intolerance, Several lines of evidence suggest that TLR4 may play a role in the pathogenesis insulin resistance, and sarcopenia, in young (3-5 m), mature (12-15 m) and of insulin resistance. Yet, it is not known whether blocking TLR4 action improves very old (33-35 m) transgenic mice with muscle-specifi c inactivation of glucose metabolism. TAK-242 is a small molecule inhibitor of TLR4 studied for NFκB (MISR). In wild type (WT) mice, glucose tolerance (measured with the treatment of sepsis. The goal of the present study was to evaluate whether I.P. GTT) and insulin sensitivity (measured with I.P. ITT and euglycemic blocking TLR4 with TAK-242 prevents acute and chronic fat-induced insulin clamp) worsened with age, whereas MISR mice were protected from age- resistance in vivo. In the acute experiment, rats received TAK-242 (10 mg/ related alterations in glucose metabolism. Muscle mass and strength also kg) or vehicle, and an 8 h infusion of Intralipid 20% (8.5 mg/kg/min) or saline, progressively declined with age in WT mice, however, MISR mice were followed by a 2-step (0.4 and 4 mU/kg/min) hyperinsulinemic-euglycemic clamp. not protected from sarcopenia. To the contrary, muscle mass (14%, P<0.05) The glucose infusion rate (GIR) during step-2 of the clamp was reduced 2-fold in and strength (22%, P<0.05) were lower in very old MISR compared with rats infused with lipid. This effect was partially restored by TAK-242, suggesting WT mice. Consistent with this fi nding, muscle proteasome activity (26S) improved peripheral (muscle) insulin sensitivity. Accordingly, the lipid-induced was elevated in MISR mice (by 7-13%, P<0.05), regardless of the age. The reduction (18%) in insulin-stimulated glucose disposal (Rd) was attenuated by improved glucose metabolism in aged MISR mice indicates that NFκB may TAK-242. Insulin suppressed hepatic glucose production (HGP) in saline- but not play a role in the glucose intolerance and insulin resistance of aging. The lipid-treated rats. TAK-242 partially restored this effect, suggesting improved fi nding that inhibition of NFκB accelerates age-induced sarcopenia suggests hepatic insulin action. In the chronic experiment, rats were implanted with a that infl ammatory pathways have dual roles and may help preserve muscle slow-release pellet of TAK-242 (1.5 mg/d) or placebo. Rats received a high fat diet as age advances. (HFD) or a low fat control diet (LFD) for 10 wk, followed by a 2-step (2 and 8 mU/kg/ Supported By: NIH (R01DK080157, R01DK089229, K23AG030979, POAG013319) min) insulin clamp. The HFD reduced the GIR during step-2 of the clamp by 3-fold, and TAK-242 partially ameliorated this reduction (p<0.05), suggesting improved 1873-P peripheral insulin sensitivity. Accordingly, Rd was reduced 30% by the HFD, but Increase in Specifi c Skeletal Muscle Ceramide May Contribute to completely restored by TAK-242 (p<0.05). The insulin-induced suppression of the Overfeeding-induced Decline in Insulin Sensitivity HGP observed in rats fed a LFD was not evident in rats fed a HFD and TAK-242 DARCY L. JOHANNSEN, DIANA OBANDA, WILLIAM T. CEFALU, JEFFREY D. had no effect on hepatic insulin sensitivity. In summary, TAK-242 provides partial COVINGTON, ERIC RAVUSSIN, Baton Rouge, LA protection against acute and chronic fat-induced insulin resistance in vivo. TLR4 Accumulation of lipid species in skeletal muscle is thought to trigger insulin may represent a novel target to improve insulin action in patients with obesity resistance (IR) by interfering with cellular insulin signaling. Whether skeletal and type 2 diabetes. muscle ceramide accumulation is a major culprit of IR remains controversial. Supported By: NIH (1F32DK095565-01A1, R01DK080157, R01DK089229, K23AG Here we sought to determine in a longitudinal design whether overfeeding Integrated 030979, POAG013319) promotes an accumulation of skeletal muscle ceramides, contributing to a POSTERS decline in insulin sensitivity. 1871-P We overfed 29 males (27±5 y, BMI 25.5±2.3; mean±SD) 40% above Physiology/Obesity Fyn-Dependent Tyrosine Phosphorylation of AMPK on Y436 Regu- baseline energy requirements for 8 weeks (15% protein, 44% fat, 41% CHO). lates Its Intrinsic Activity All meals were consumed under supervision. Whole-body insulin sensitivity EIJIRO YAMADA, SHUICHI OKADA, TSUGUMICHI SAITO, YOKO SHIMODA, YUKO was determined by a hyperinsulinemic-euglycemic clamp (50 mU/m2·min 2 TAGAYA, CLAIRE C. BASTIE, JEFFREY E. PESSIN, Maebashi, Japan, Bronx, NY insulin for 2.5 h with 6,6- H2 glucose tracer) and skeletal muscle tissue was AMP-dependent protein kinase (AMPK) is a central regulator of energy collected by needle biopsy. Post-testing was performed after re-establishing utilization and plays an important role in maintaining the balance of glucose energy balance at participants’ new body weight. and fatty acid oxidation. Dysregulation of this process is associated with Participants gained 7.6±2.1 kg (9.3±2.8%) comprised of 4.2±1.4 kg of states of insulin resistance and Type 2 Diabetes. Previously we reported that fat. Glucose infusion rate decreased from 11.5±2.5 to 10.9±2.5 mg/kg Fyn null mice display increased energy expenditure and fatty acid oxidation FFM·min (p=0.05). Total ceramide content in muscle increased by 123% due to increased AMPK activity in peripheral tissues. More recently, we after overfeeding (842±471 to 1879±673 nM/mg protein, p<0.0001) and

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A481 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

was evident in all species measured (C16:0, C18:1, C18:0, C20:0, C22:0, osteocalcin levels in adults were signifi cantly correlated with BMI (r = -0.20, C24:1, C24:0, all p<0.0001). The change in total ceramide content was not p<0.001), HbA1c (r = -0.20, p<0.001), fasting glucose (r = -0.16, p<0.001), signifi cantly associated with the change in insulin sensitivity (r = -0.28, and 2-hour glucose (r = -0.21, p<0.001). Moreover, a stepwise decrease in p=0.15); however, larger increases in species C24:1 (r = -0.40, p=0.04) and osteocalcin levels was noted across the spectrum of glycemic status from C20:0 (r = -0.37, p=0.05) were related to larger declines in insulin sensitivity. normal, prediabetic, and type 2 diabetic, respectively (43.9±1.3, 39.3±1.3, In conclusion, overfeeding inducing a 10% weight gain caused a marked and 31.9±1.2, p<0.001). increase in total skeletal muscle ceramide content in parallel to a modest These data suggest that the associations between osteocalcin and T2D reduction in insulin sensitivity. Although the increase in total ceramides did risk are age dependent. In youth where osteocalcin levels may be naturally not correlate with the decrease in insulin sensitivity, our data suggests that increased due to bone development and growth, the effects on obesity- specifi c ceramide species may play a role in the pathogenesis of weight-gain related health may be minimal. As fi nal growth is reached in adulthood a induced muscle insulin resistance. more distinct and protective effect of osteocalcin on T2D risk emerges. Supported By: R01DK060412, K01DK089005, 2P30DK072476 & 1876-P Testosterone Restores Insulin Sensitivity and Improves Insulin INTEGRATED PHYSIOLOGY—OTHER HORMONES Signal Transduction in Patients with Diabetes and Hypogonadotropic Hypogonadism MANAV BATRA, SANDEEP DHINDSA, SANAA ABUAYSHEH, NITESH D. KUHA- Guided Audio Tour: Novel Axis for Regulation of Glucose Metabolism DIYA, KELLY GREEN, ANTOINE MAKDISSI, JEANNE HEJNA, HUSAM GHANIM, (Posters: 1874-P to 1881-P), see page 17. AJAY CHAUDHURI, PARESH DANDONA, Buffalo, NY Hypogonadotropic hypogonadism (HH) is associated with increased & 1874-P insulin resistance compared to eugonadal type 2 diabetics (T2DM), based Glycemic Control Is Impaired in the Evening in Prediabetes Through on HOMA-IR. We have now investigated whether insulin resistance is Multiple Circadian Rhythms increased in the HH and the effects of testosterone supplementation on COURTNEY M. PETERSON, TANCE SONNIER, JENNIFER ROOD, JEFFREY M. insulin sensitivity and on mediators of insulin signaling. Twenty six patients GIMBLE, Baton Rouge, LA, New Orleans, LA with HH and T2DM were compared with 24 eugonadal patients with T2DM Recent studies suggest that circadian rhythms regulate glucose at baseline. From the HH patients, 12 were treated with testosterone metabolism, weight loss, and even drug effi cacy. Moreover, molecules 200mg every 2 weeks injected intramuscularly for 6 months. Fasting blood targeted at the circadian clock show promise in treating metabolic disease. samples and fat biopsies were obtained at baseline and at 6 months. Using Therefore, this study set out to better characterize the diurnal rhythms hyperinsulinemic euglycemic clamps, we demonstrated that patients with in glucose metabolism in prediabetes. Ten subjects with prediabetes HH have 28% lower glucose infusion rates (GIR) for a given rate of insulin completed oral glucose tolerance tests at 0700 h and 1900 h on the same infusion compared to eugonadal patients. Following testosterone treatment, day. Lipids and hormones were also measured. Two-hour and three-hour there was an increase in GIR by 30%, consistent with a reversal of insulin glucose tolerances were worse in the evening by 40±52 mg/dl (p=0.02) and resistance. Basal expression of mediators of Insulin signaling including IR, 62±46 mg/dl (p=0.001), respectively. These impairments were explained by IRS-1 and GLUT-4 was lower by 32%, 35% and 27% (p<0.05), respectively, in lower insulin sensitivity (OGIS; 5.14±1.02 vs. 4.74±0.77 mg/kg/min; p=0.03) HH adipose tissue (AT) compared to eugonadal. Following testosterone, the and 2-hour AUC insulin levels (87.4±37.6 vs. 69.8±40.2 mU hr/l; p=0.02) in AT expression of IR, IRS-1 and GLUT-4 increased signifi cantly by 63±15%, the evening. Cortisol and leptin also exhibited marked circadian oscillations, 54±17% and 59±14%, respectively, while there was a decrease in the while lipids, cholesterol, and triglycerides did not. Intriguingly, more insulin expression of PTP-1B by 23±8% and TLR-4 by 21±11%, both of which interfere resistant people had weaker rhythms in insulin sensitivity (r=-0.66; p=0.04), with insulin signaling. There was a signifi cant fall in circulating mononuclear which were offset by stronger rhythms in insulin (r=-0.67; p=0.03) and cortisol cells expression of SOCS-3 by 27±8% and IKKβ by 23±11%, both known to (r=-0.78; p=0.008) levels; this is presumably a compensatory response interfere with insulin signaling. Additionally, plasma concentrations of FFA, intended to preserve better glycemic control in the morning. Interestingly, TNFα and CRP, all of which interfere with insulin signaling, fell by 35%, 18% the difference between glucose tolerance in the morning versus evening and 26%, (p<0.05) respectively. Therefore, testosterone supplementation was strongly determined by the circadian rhythms in cortisol and insulin has an insulin sensitizing effect in HH patients at various levels involving an sensitivity (r=0.86; p=0.002). Surprisingly, the cortisol circadian rhythm increase in the expression of mediators of insulin signaling and a reduction was the most important predictor of large declines in glucose tolerance in factors that interfere with insulin signaling. in the evening. In conclusion, glycemic control is dramatically impaired in Supported By: NIH the evening in people with prediabetes, with those who have the weakest circadian rhythms in cortisol being the most at-risk for large declines. Given & 1877-P the size of the circadian drop in glycemic control, food intake may need to be Gastrointestinal-mediated Glucose Disposal in Total Pancreatecto- curbed at dinnertime in prediabetes. mized Patients Supported By: Jenny Craig; Pennington Biomedical Research Foundation ASGER LUND, JONATAN I. BAGGER, MIKKEL CHRISTENSEN, MAGNUS GRØN- DAHL, ELISABETH R. MATHIESEN, CARSTEN P. HANSEN, JAN STORKHOLM, & 1875-P STEEN LARSEN, JENS J. HOLST, TINA VILSBØLL, FILIP K. KNOP, Copenhagen, Osteocalcin and Type 2 Diabetes Risk across the Lifespan Denmark GABRIEL Q. SHAIBI, IFTIKHAR J. KULLO, ANGELA K. DALENBERG, DAWN K. Gastrointestinal-mediated glucose-disposal (GIGD) after oral glucose COLETTA, LAWRENCE J. MANDARINO, Phoenix, AZ, Rochester, MN, Tempe, AZ tolerance test (OGTT) refl ects the percentage of glucose disposal caused by The bone-derived protein osteocalcin is inversely associated with obesity the oral route of glucose administration. It accounts for as much as 70% in and type 2 diabetes (T2D) in adults, but it is not known whether these healthy subjects. Mediators of GIGD may include the gut incretin hormones, associations are present in youth. Therefore, the purpose of this study was gut microbiota, fi rst-pass hepatic uptake of glucose and at present unknown to examine the associations between osteocalcin and T2D risk in youth and factors. It is likely that incretin-mediated potentiation of pancreatic insulin Integrated POSTERS explore whether differences exist across the lifespan. secretion constitutes a major contributor to GIGD, but so far it has not Data from 121 youth (14.0±0.2yrs) and 508 adults (33.6±0.5yrs) who been possible to discriminate between pancreatic and extrapancreatic Physiology/Obesity participated in the Arizona Insulin Resistance Registry were used for the mechanisms underlying GIGD. We aimed to evaluate the impact of current analysis. Fasting serum osteocalcin was measured by solid-phase extrapancreatic effects on GIGD. Data from 7 total pancreatectomized sandwich immunoassay and T2D risk factors included BMI (percentile for patients (age: 60±4 years; body mass index (BMI): 22.3±1.4 kg/m2; HbA1c: youth and BMI for adults), HbA1c, fasting glucose, and 2-hour glucose after 63±4 mmol/mol (mean±SEM)) and 6 healthy control subjects (age: 57±3 a standard oral glucose tolerance test. years; BMI 22.9±0.8 kg/m2; HbA1c: 32±1 mmol/mol) were included in Osteocalcin (nmol/L) levels were >5-fold higher in youth compared to the present preliminary analysis. Participants were examined over two adults (208.5±11.6 vs. 40.9±0.9, p<0.0001). In youth, osteocalcin levels were experimental days: a 75g-OGTT and a corresponding isoglycemic iv glucose not correlated with BMI percentile (r = -0.03, p=0.8), HbA1c (r = 0.04, p=0.7), infusion (IIGI). Differences between administered glucose during OGTT and fasting glucose (r = 0.12, p=0.2), or 2-hour glucose (r = 0.12, p=0.2). Further, no IIGI within the group of pancreatectomized patients were used to evaluate differences in osteocalcin levels were noted between normoglycemic youth the impact of extrapancreatic mechanisms on GIGD. In healthy control and those with prediabetes (193.4±15.7 vs. 196.1±24.0, p=0.9). In contrast, subjects 28±2 g of glucose was infused intravenously to copy the plasma

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A482 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES glucose profi le from the 75 g-OGTT, resulting in a GIGD of 63±2%. In the total Since R59W is the loss of function mutation of BTH, the subjects with pancreatectomized patients we used 76±3 g of glucose to copy the plasma R59W are supposed to be hereditarily prone to DM or IGT. However, no glucose profi le from the 75 g-OGTT, resulting in a GIGD of -2±4% (i.e. that tendency was observed in the general population, only in those subjects the pancreatectomized subjects disposed of iv and oral glucose similarly). with elevated TG. TG overproduction plays a signifi cant role in subjects with The low GIGD in pancreatectomized patients suggests that extrapancreatic R59W and TG elevation. In subjects with elevated TG, fatty acids derived factors do not play a major role in GIGD. The need for a larger glucose load on from TG would damage beta cells through oxidative stress. Therefore, the IIGI day could however point to the existence of gut-derived factors (e.g. decreased induction potency of beta cells with R59W might be prominent gut derived glucagon) contributing to a worsening of oral glucose tolerance in these subjects. in pancreatectomized patients. Thus, BTH would play a signifi cant role in the progression of diabetes in Supported By: EFSD subjects with hyper TG.

& 1878-P & 1880-P Differential Regulation of Glucocorticoids and Insulin on Hepatic Enteroendocrine Cell Response to Roux-en-Y Gastric Bypass 11β-Hydroxysteroid Dehydrogenase Type 1 Enzyme Activity NICOLAI A. RHEE, CAMILLA D. WAHLGREN, JENS PEDERSEN, EBBE LANGHOLZ, SIMMI DUBE, ISABEL ERRAZURIZ, BARBARA NORBY, CHERYL SHONKWILER, ERIK P. WANDALL, STEFFEN U. FRIIS, PETER VILMANN, SARAH J. PAULSEN, ANANDA BASU, RITA BASU, Rochester, MN VIGGO B. KRISTIANSEN, JACOB JELSING, NIELS VRANG, JENS J. HOLST, TINA Animal studies indicate that insulin decreases and glucocorticoids VILSBØLL, FILIP K. KNOP, Hellerup, Denmark, Copenhagen, Denmark, Hørsholm, increase hepatic 11 β-HSD-1 expression and activity. The current studies Denmark , Hvidovre, Denmark were undertaken to evaluate whether hyperinsulinemia suppress and The impact of Roux-en-Y gastric bypass (RYGB) on glycemic control is glucocorticoids stimulate hepatic cortisone to cortisol conversion via thought to be partly conveyed by an altered anatomy and physiology of the 11β-HSD-1 in humans. 45 nondiabetic subjects (ND) were randomized to enteroendocrine cells of the gut. We aimed to investigate RYGB-induced receive either 50 mg twice a day of hydrocortisone (Group I, n=15) for one alterations in the prevalence of enteroendocrine cells and their hormonal week followed by a six hour saline infusion on study day; or placebo twice a products in 12 patients with type 2 diabetes (T2D) and 11 non-diabetic day (Group II, n= 15) for one week and a six hour hyperinsulinemic (2mU/kg/ individuals. Gut mucosa samples were taken during the RYGB procedure (from min) euglycemic clamp ( ~5mM) on study day; or placebo twice a day (Group the site of the jejuno-jejunal anastomosis) and compared to mucosa samples 2 III, n= 15) for one week and a six hour saline infusion on study day. [6,6 H2] obtained endoscopically 10.4 months postoperatively at 3 different sites glucose was infused to calculate glucose fl uxes and insulin/saline infusion around the jejuno-jejunal anastomosis (alimentary, secretory and common started at time zero along with ingestion of 1 mg of 13C cortisone to measure limb). Biopsies were analyzed using quantitative polymerase chain reaction hepatic C13 cortisol production. (qPCR) for ghrelin, cholecystokinin (CCK), secretin, glucose-dependent Baseline cold cortisol concentrations were higher (p =0.0001) in Group I insulinotropic polypeptide (GIP), glucagon-like peptide-1 (GLP-1), peptide YY vs. III and not different (p = 0.21) in group II vs. III. Rate of appearance (Ra) (PYY), neurotensin and FXR mRNA, and immunohistochemical (IHC) staining of C13 cortisol (derived from ingested 13C cortisone) was signifi cantly higher for CCK, GIP, GLP-1 and PYY. Ghrelin and CCK mRNA was downregulated (p= 0.005) in group I vs. III while not different (p = 0.74) between group II vs. following RYGB in both groups whereas CCK positive cells increased III implying that 11β- HSD-1 activity is augmented by glucocorticoids and numerically after RYGB in the control group. In both groups secretin mRNA unaffected by insulin. Basal rates of endogenous glucose production (EGP) was downregulated in the secretory limb vs. perioperative samples. GIP were signifi cantly higher (17.5 ± 2.4 vs. 16.0 ± 2.4 µmol/kg/min; p =0.0001) mRNA was downregulated in all 3 limbs in both groups vs. perioperatively. and suppression of EGP signifi cantly lower (11.7 ± 2.6 vs. 7.8 ± 3.3 µmol/ Also, decreased densities of GIP positive cells were found in the alimentary kg/min; p = 0.0001) in group I vs. III suggesting that glucocorticoids impair limb vs. perioperative biopsies. After RYGB GLP-1 mRNA expression was hepatic insulin action. Glucose disappearance was signifi cantly lower during upregulated in the secretory limb of the T2D group, whereas no changes study (13.0 ± 2.5 vs. 16.6 ± 4.1 µmol/kg/min; p =0.0001) in group I vs. III. We in the density of GLP-1 positive cells were seen in either of the 2 groups. conclude a) glucocorticoids stimulate hepatic cortisone to cortisol conversion PYY positive cells were less abundant in the common limb vs. perioperative via 11β-HSD-1 causing hepatic and extra hepatic insulin resistance b) biopsies. No signifi cant changes in neurotensin or FXR mRNA were found. insulin has no effect on hepatic 11β-HSD-1 enzyme activity under present In conclusion, a number of alterations in the distribution of enteroendocrine experimental conditions. cell and the expression of their hormonal products occur after RYGB. These Supported By: NIDDK (29953) alterations may constitute important mechanisms underlying RYGB-induced improvements in glycemic outcomes in patients with T2D prior to RYGB. & 1879-P Supported By: Højteknologifonden High Frequency of Diabetes and Glucose Intolerance in the Hypertriglyceridemic Subjects with Betatrophin (ANGPTL8) R59W & 1881-P Mutation Wnt1 Inducible Signaling Pathway Protein 1 (WISP1) Is a Novel JIANHUI LIU, KUNIMASA YAGI, ATSUSHI NOHARA, KENGO WADA, HIDEKO Adipokine Linked to Infl ammation and Insulin Resistance in Visceral KITAMOTO, AZUSA OBATAKE, YUKIKO MORI, SATOKO OKAZAKI, YOSHIYU Fat TAKEDA, MASAKAZU YAMAGISHI, Kanazawa, Japan NATALIA N. RUDOVICH, VERONICA MURAHOVSCHI, OLGA PIVOVAROVA, IRYNA Betatrophin (BTH), which is translated from the gene ANGPTL8, has been ILKAVETS, RENATA M. DMITRIEVA, STEPHANIE DÖCKE, ÖZLEM GÖGEBAKAN, recently recognized as the hormone that regulates pancreatic beta cell MARTIN OSTERHOFF, NORA KLÖTING, ALEXANDRA CONRADI, STEVEN DOOLEY, mass and proliferation under insulin resistance. BTH also acts as a lipase MARTIN STOCKMANN, PETER NEUHAUS, CHRISTIAN VON LOEFFELHOLZ, inhibitor cooperating with ANGPTL3 and ANGPTL4 resulting in triglyceride MATTHIAS BLÜHER, ANDREAS F.H. PFEIFFER, Nuthetal, Germany, Mannheim, (TG) increase. Germany, Saint Petersburg, Russian Federation, Leipzig, Germany, Berlin, Germany, Since we have found that the BTH mutation R59W is not rare in the Jena, Germany Japanese population, we examined the relationship between the BTH WISP1 is a member of the secreted extracellular matrix associated proteins mutation and lipoprotein and glucose metabolism. of the CCN family and target gene of canonical Wnt signaling pathway. Integrated Written informed consent for gene analyses was obtained from all 532 We aimed to validate WISP1 as a novel adipokine and to characterize POSTERS people (M431/F101, DM 228 cases, IGT 61, non-DM 243 cases, age 54±14 the association of WISP1 with parameters of the metabolic syndrome. years old, BMI 24.0±3.7, TC 207±44 mg/dl, TG 157±117 mg/dl, HDL-C 49±14 Methods: WISP1 expression was studied 1) in paired samples of visceral Physiology/Obesity mg/dl, FBS 130±46 mg/dl, HbA1c (NGSP) 6.90±1.64%). Genomic DNA was (VAT) and subcutaneous adipose tissue (SAT) from healthy subjects (n=75); extracted from peripheral white blood cells. The BTH R59W mutation was 2) in paired VAT, SAT and liver tissue from subjects with/without NAFLD identifi ed with PCR-RFLP and all subjects were genetically divided into three (n=47); 3) in human weight reduction study (n=49); 4) in SAT of overweight groups, homozygote (WW), heterozygote (WR) and wild type (RR). subjects (n=14) in the euglycemic-hyperinsulinemic and hyperglycemic- No difference in the frequency of DM among WW, WR and RR was hyperinsulinemic clamp test; 5) in human stem cells derived adipocytes and observed. However, when dividing subjects by TG levels, the frequency of in human monocytes and M-and GM-macrophages. Results: WISP1 protein DM+IGT correlated with the existence of R59W in subjects with TG greater was identifi ed in cell culture medium of human stem cells derived adipocytes. than 150 mg/dl; WW(73.68%), WR(62.03%), RR(50.62%) (p=0.0458). On the Gene expression of WISP1 was not detectable in monocytes as well as in other hand, the frequency of DM+IGT was not signifi cantly correlated in M-and GM-macrophages. A strong and signifi cant increase of IL-6, NFkB subjects with TG less than or equal to 150. gene expression in human stem cells derived adipocytes and of TNFα gene

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A483 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

expression in human monocytes after 4-h stimulation with 1µg/ml WISP1 1884-P was observed. In human studies: 1) WISP1 gene expression correlates with insulin sensitivity, macrophages infi ltration and adiponectin levels in samples WITHDRAWN of VAT and SAT; 2) WISP1 expression is substantially elevated in VAT rather than in SAT; 3) hepatic WISP1 expression has no association with ectopic fat accumulation in obesity; 4) weight reduction decreases WISP1 expression in SAT as well as circulating WISP1 levels in plasma. Discussion: WISP1 is a novel adipokine but no hepatokine that is substantially overexpressed in visceral fat from obese subjects and refl ects insulin resistance and adipose tissue infl ammation. Weight changes regulate circulating WISP1 levels and WISP1 expression in adipose tissue. Therefore, we propose that WISP1 is a novel biomarker and a potential link between obesity and the metabolic syndrome. Supported By: German Research Foundation; German Federal Ministry of Education and Research

1882-P Leptin Reverses Diabetes by Hypothalamic-Pituitary-Adrenal Axis- mediated Reductions in SubstrateWITHDRAWN Delivery to Liver RACHEL J. PERRY, XIAN-MAN ZHANG, DONGYAN ZHANG, NAOKI KUMASHIRO, JOÃO PAULO G. CAMPOREZ, GARY W. CLINE, DOUGLAS L. ROTHMAN, GERALD I. SHULMAN, New Haven, CT Leptin treatment reverses hyperglycemia in animal models of poorly- controlled type 1 diabetes (T1D) yet the mechanism of this remains poorly understood. To this end, we examined liver-specifi c rates of hepatic gluconeogenesis in conjunction with rates of whole-body glycerol, fatty acid and acetate turnover in a rat model of poorly controlled T1D. Rates of hepatic gluconeogenesis were increased by 120% (P<0.0001) in the T1D rats and could be attributed to hypoleptinemia-induced increase in plasma corticosterone (42±12 to 187±30 ng/mL, P=0.001) resulting in three- to four-fold increases in glycerol (P<0.001), fatty acid (P<0.0001) and acetate (P<0.0001) turnover. Hepatic acetyl CoA concentrations were a strong predictor of fasting plasma glucose concentrations (R^2=0.86) and pyruvate carboxylase fl ux (R^2=0.75), independent of changes in gluconeogenic gene or protein expression. All of the altered metabolic fl uxes were mimicked by infusing rats with Intralipid and heparin or corticosterone, and corrected by six hours of leptin infusion, without any difference in plasma insulin or 1885-P glucagon concentrations. Leptin treatment had no effect on plasma glucose Testosterone Defi ciency Exacerbates High-Fat Diet-induced concentrations or rates of hepatic gluconeogenesis when Intralipid and Glucose Intolerance through Inhibition of GLUT4-mediated Glucose heparin were infused to increase fatty acid and glycerol delivery to the liver. Uptake in Skeletal Muscle In conclusion, these data demonstrate a critical role for increased lipolysis KAZUTERU MITSUHASHI, MICHIAKI FUKUI, TAKAFUMI SENMARU, MUNEO secondary to hypoleptinemia-induced hypercorticosteronemia in promoting TSUJIKAWA, HIROSHI OBAYASHI, TAKUYA FUKUDA, HIROYA IWASE, HIROSHI increased hepatic gluconeogenesis in poorly controlled T1D independent of OKADA, MUHEI TANAKA, MASAHIRO YAMAZAKI, GOJI HASEGAWA, NAOTO changes in hepatic gluconeogenic protein expression. NAKAMURA, Kyoto, Japan Testosterone defi ciency is associated with impaired glucose metabolism. 1883-P However the precise mechanism of this association is not apparent. A Trace Gluconeogenesis and Glucose Uptake Under in Diabetic Rats high-fat diet (HFD) induces glucose intolerance in part through inhibition of with GLP-1 Analogues glucose uptake in skeletal muscle. The aim of this study was to investigate HUI WU, CHUNHUA SUI, HUI XU, YINGLI LU, Shanghai, China the combine effects of testosterone defi ciency and HFD on skeletal muscle Glucagon-like peptide-1 (GLP-1) analogues effectively reduce blood glucose metabolism in mice. Male C57BL/6J mice aged 7 weeks were either glucose in diabetes. However, there are few detailed studies about GLP- orchiectomized (ORX) or sham-operated (SHAM), and then were fed a HFD or 1 analogues in the regulation of gluconeogenesis (GNG), hepatic glucose a standard diet for 12 weeks starting at 8 weeks of age. In mice fed standard production (HGP), glucose uptake in peripheral tissue through by trace in diet, orchiectomy did not affect all metabolic parameters. HFD increased homeostasis. To explore the GLP-1 analogues as exenatide, this study had fasting blood glucose concentrations in both SHAM and ORX mice. In the four groups of SD rats: non-diabetic (control, C); non-diabetic + exenatide IPGTT, blood glucose concentrations at 30, 60 and 120 min after glucose (C+E); diabetic (D); and diabetic + exenatide (D+E). After eight weeks, isotope load were signifi cantly higher in ORX mice than SHAM mice (120 min: ORX tracer technology was used to check the rate of appearance (Ra) of glucose, 30.20 ± 4.54 vs. SHAM 25.63 ± 3.95 mmol/L, p < 0.05, n =7). Gene analysis glycerol and GNG following infusion of 3-3H-glucose and U-13C-glycerol. revealed that the expression of GLUT4 was decreased to 1.4-fold by HFD, Hepatic glucose production and the rate of glucose disappearance (Rd) and further decreased to 2.3-fold by testosterone defi ciency. Similarly, the were assessed using a hyperinsulinemic-euglycemic clamp with infusion of expression of myocyte enhancer factor 2A (MEF2A), Lipin1 and p38 mitogen- 3-3H-glucose. Glucose uptake was determined in gastrocnemius muscles activated protein kinase (MAPK) was down-regulated both by HFD and Integrated POSTERS with 2-deoxy-D-14C-glucose. In the D+E group, body weight, fasting blood by testosterone defi ciency. In addition, the expression of Rho family GTP- glucose, triglyceride, total cholesterol, lower density lipoprotein, insulin and binding protein TC10, which is a positive regulator of GLUT4 translocation Physiology/Obesity HOMA-IR were signifi cantly decreased compared with the D group. The Ra of from intracellular site to plasma membrane, was down-regulated in ORX glucose (94.70 ± 13.46 vs. 121.07 ± 16.55 µmol/kg/min, p<0.01), Ra of glycerol mice fed HFD. These perturbations in ORX mice fed HFD were normalized to (15.75 ± 2.04 vs. 24.65 ± 5.39 µmol/kg/min, p<0.01); Gluconeogenesis (48.23 ± the levels of SHAM mice fed HFD by testosterone supplementation. These 11.20% vs. 61.53 ± 7.44%µmol/kg/min) were also decreased p<0.001; Uptake results suggest that the effect of testosterone on glucose uptake is mediated glucose in muscle were (0.24 ± 0.02 vs. 0.17 ± 0.02 µmol/g/min, p<0.01) by the p38 MAPK/MEF2 axis, which is a strong inducer of GLUT4 expression. markedly increased. These trace results show that in addition to decreasing In conclusion, testosterone defi ciency may exacerbate glucose intolerance gastrointestinal glucose absorption, the GLP-1 analogue inhibits hepatic through inhibition of GLUT4-mediated glucose uptake in the skeletal muscle. gluconeogenesis and improves glucose uptake in muscle signifi cantly, and A high-fat diet could be an initiating factor for those changes. improves others parameter of glucose metabolism.

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A484 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1886-P antihypertensive treatment, we examined the associations between the Low-Salt Diet in Subjects with Hypertension: Impact on Insulin GLP-1 and GIP release during a 3-point 75 g OGTT (0, 30 and 120 minutes) Resistance and Its Modulation by Salt Sensitivity and markers of vascular function: brachial and central blood pressure, and RAJESH GARG, BEI SUN, JONATHAN WILLIAMS, Boston, MA aortic pulse wave velocity (PWV)-an assessment of aortic stiffness. Total Salt sensitivity is associated with increased insulin resistance (IR) and area under the curve (AUC) and incremental AUC (iAUC) for GLP-1 and GIP metabolic syndrome. Some experts have suggested salt restriction as a were calculated and transformed by the base 2 logarithm. Associations method of reversing or controlling the metabolic syndrome in these subjects. were analyzed by linear regression adjusting for age and sex. The analyses However, we have shown an increase in IR in relation to low salt (LS) intake of PWV were additionally adjusted for heart rate and mean blood pressure. in healthy subjects. In this study, we evaluated the impact of salt restriction Mean age was 55.1 years (SD: 7.4) and 52% were men. There was a tendency on IR in hypertensive subjects and its modulation by salt sensitivity. This towards a lower blood pressure for a higher GLP-1 release (Figures 1a and study includes 389 hypertensive subjects (44% Females, 16% Blacks, mean 1b), and a doubling in iAUCGLP-1 was associated with a difference in PWV of BMI 28.5±4.2 Kg/m2) studied after one week of high salt (HS) (200 mmol/ -0.04 m/s (95% CI: -0.12;0.04). For GIP, there was no clear tendency, and only day Na) and one week of LS (20 mmol/day Na) diet. On day 6 of each diet, brachial pulse pressure was associated with GIP, showing that a doubling participants remained fasting and supine overnight for 8-10 hours and all of iAUCGIP corresponded to a higher pulse pressure of 1.11 mmHg (95% CI: hemodynamic and laboratory assessments were made the following morning. 0.06;2.16). Although cross-sectional, these results indicate that GLP-1 but The homeostasis model assessment (HOMA) was used as an index of IR. Salt not GIP, has a benefi cial impact on the vascular function. sensitivity was defi ned as the fall in systolic blood pressure (SBP) >15 mmHg on LS diet as compared to HS diet. In the overall group, as expected, BP was lower on LS diet (129±16 / 78±9 mmHg) compared to HS diet (145±18 / 86±10 mmHg) and renin-angiotensin-aldosterone system was activated. Fasting plasma glucose, insulin and HOMA were higher on LS diet (95.4±19.4 mg/dl, 10.8±7.3 mIU/L and 2.6±1.9) as compared to HS diet (90.6±10.8 mg/ dl, 9.4±5.8 mIU/L and 2.1±1.4) (p <0.0001 for all). Based on SBP criterion, 193 subjects were classifi ed as salt sensitive (SS) and 196 as salt resistant (SR). There was no difference in HOMA between SS versus SR subjects on either diet. Increase in HOMA on LS diet was 0.5±1.4 in SS subjects and 0.4±1.5 in SR subjects (p=NS). On multivariate regression analysis, change in SBP was not signifi cantly associated with change in HOMA. The relation did not change after controlling for age, BMI, sex, change in serum and urine aldosterone and change in serum and urine cortisol. We conclude that salt restriction increases IR in hypertensive subjects and this effect is not modulated by salt sensitivity.

1887-P Supported By: Danish Diabetes Academy; EFSD/Pfi zer, Inc. (74550801) Nrf2 Positively Regulates FGF21 in Diabetic Mice: A Potential Effi cacy Biomarker of Nrf2 Induction 1889-P YUKI FURUSAWA, AKIRA URUNO, YOKO YAGISHITA, MASAYUKI YAMAMOTO, Neuroendocrine Regulatory Peptide-4 (NERP-4): A VGF Derived Pep- Gotemba, Japan, Sendai, Japan tide That Expresses in Pancreatic β-Cells and Stimulates Glucose- A transcription factor Nuclear factor E2-related factor 2 (Nrf2) is a key induced Insulin Secretion regulator for the protection from oxidative stresses, which is negatively KOICHIRO SHIMIZU, HIROAKI UENO, EMI EBIHARA, HIROE SHIBATA, CHIKAKO regulated by an adaptor protein Kelch-like ECH-associated protein 1 (Keap1). TSUBOUCHI, YUUTA MORINAGA, WAKABA TSUCHIMOCHI, ABU SALEH M.D. Several studies revealed that Nrf2 also plays a pivotal role in metabolic MOIN, TADATO YONEKAWA, HIDEKI YAMAGUCHI, MASAMITSU NAKAZATO, regulations such as lipid metabolism and energy expenditure as well Miyazaki, Japan as redox homeostasis. Our previous study showed that Nrf2 activation Neuroendocrine regulatory peptide-4 (NERP-4), a VGF derived (VGF489-507) by hypomorphic knockdown of Keap1 or administration of Nrf2 inducer peptide was identifi ed through an ex vivo Ca2+ assay using tissue pieces from synthetic oleanolic triterpenoid 1-[2-cyano-3,12-dioxooleane-1,9(11)-dien- transgenic mice that systemically express the calcium indicator apoaequorin. 28-oyl] imidazole (CDDO-Im) markedly suppresses diabetes in mice. To In this study we identifi ed that NERP-4 raised transient intracellular Ca2+ in explore the mechanisms how Nrf2 regulates glucose metabolism, we pancreas of apoaequorin transgenic mouse. To explore the physiological conducted microarray analysis using Keap1 knockdown mouse livers and roles of NERP-4 in pancreas, we examined its cellular localizations in found that fi broblast growth factor 21 (FGF21), a mediator of glucose and pancreatic islets and functions on insulin secretion. NERP-4 was expressed lipid metabolism, is upregulated in the livers. We have examined roles that in pancreatic islet cells of human and mouse and co-expressed abundantly Nrf2 plays in diabetic model mice and characterized involvement of Nrf2 in with insulin but moderately with glucagon and somatostatin. Immunoelectron the FGF21 regulation. We found that Nrf2 induction by genetic knockdown of microscopy demonstrated that NERP-4 also subcellularly localized in a dense Keap1 increased plasma FGF21 and hepatic Fgf21 expression levels in diabetic core granule, colocalized with insulin granule in the secretory vesicles of db/db mice and high-calorie-induced obesity model mice. Administration of pancreatic β-cell. NERP-4 potentiated glucose-stimulated insulin secretion CDDO-Im upregulated plasma FGF21 and hepatic Fgf21 expression levels in from isolated mouse islets or insulinoma (MIN6-K8) cells. Insulin secretion db/db mice, while this was canceled in Nrf2 knockout db/db mice. Our data in response to different doses of NERP-4 in MIN6-K8 cells exhibited a demonstrate that Nrf2 positively regulates FGF21 in diabetic mice and FGF21 bell-shaped pattern where 10-10 M of NERP-4 showed the highest activity. is a potential effi cacy biomarker that possibly mediates metabolic regulation Peripheral administration of NERP-4 in presence of glucose also increased by the Keap1-Nrf2 system. plasma insulin level in mice and rats. Ca2+ imaging analyses in MIN6-K8 Supported By: Ministry of Education, Culture, Sports, Science and Technology; cells demonstrated that NERP-4 accelerated intracellular Ca2+ infl ux. Taken Integrated

Japan Society for the Promotion of Science; JST; Takeda Foundation; Naito together our fi ndings revealed that NERP-4 is a newly identifi ed bioactive POSTERS Foundation peptide targeting pancreatic β-cells to modulate insulin secretion. Physiology/Obesity 1888-P 1890-P The Relationships between Plasma Incretin Hormone Release Diverse Signaling Systems Activated by the Sweet Taste-sensing and Peripheral and Central Hemodynamics in Individuals without Receptor in Human GLP-1-secreting Cells Known Diabetes: The ADDITION-PRO Study YOSHIAKI OHTSU, YUKO NAKAGAWA, MASAHIRO NAGASAWA, HIROKAZU NANNA B. JOHANSEN, KRISTINE FÆRCH, SIGNE S. TOREKOV, DANIEL R. WITTE, ARAKAWA, ITARU KOJIMA, Maebashi, Japan ANNA JONSSON, OLUF PEDERSEN, TORBEN HANSEN, TORSTEN LAURITZEN, The sweet taste receptor regulates secretion of glucagon-like peptide-1 ANNELLI SANDBÆK, JENS J. HOLST, DORTE VISTISEN, MARIT E. JØRGENSEN, (GLP-1) in enteroendocrine cells. We investigated the intracellular signaling Gentofte, Denmark, Copenhagen, Denmark, Strassen, Luxembourg, Aarhus, Denmark system activated by this receptor using a human enteroendocrine cell line, The role of incretin hormone release on vascular function remains unclear. Hutu-80. We stimulated these cells with four sweet molecules: sucralose, Hence, in 836 Danish individuals without know diabetes and without saccharin, acesulfame K and glycyrrhizin. These four sweeteners stimulated

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A485 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

secretion of GLP-1, which was attenuated by lactisole, an inhibitor of the HepG2 cells. These data suggest that anti-proliferative effect of Ex-4 in sweet taste receptor. All these sweeteners induced rapid and sustained cancer cells might depend on GLP-1R expression level. Further, we transplant elevation of cytoplasmic cyclic AMP concentration ([cAMP]c). Among four HT-29, MCF-7 and HepG2 cells into nude mice subcutaneously and treated sweeteners, sucralose and saccharin induced a monophasic increase in them with Ex-4 for 6 weeks. We observed decreased tumor size of HT-29 cytoplasmic Ca2+ concentration ([Ca2+]c). Removal of extracellular calcium and MCF-7 in Ex-4-treated mice compared with control, but not HepG2. Their or sodium nearly completely blocked the [Ca2+]c response to sucralose and body weight and blood glucose levels were not changed by Ex-4 treatment. saccharin. YM254890, an inhibitor of Gq/11, nearly completely blocked [Ca2+] These data suggest that Ex-4 might be able to inhibit cancer growth, c responses to sucralose and saccharin. In contrast, acesulfame K induced depending on their GLP-1R expression level. a rapid and sustained reduction of [Ca2+]c. In addition, glycyrrhizin fi rst reduced [Ca2+]c which was followed by an elevation of [Ca2+]c. Reductions 1893-P of [Ca2+]c induced by acesulfame K and glycyrrhizin were attenuated by a The Triglyceride to High-Density Lipoprotein Cholesterol (TG/ calmodulin inhibitor W-7 or by knock down of the plasma membrane calcium HDL-C) Ratio as a Predictor of β-Cell Function in African American pump. These results indicate that various sweet molecules act as biased Women agonists, activate the sweet taste receptor differently and evoke strikingly AMITA MATURU, PETER DEWITT, PHILLIP A. KERN, NEDA RASOULI, Aurora, CO, different patterns of intracellular signals. Lexington, KY, Denver, CO The TG/HDL-C ratio is used as a marker of insulin resistance (IR) in 1891-P Caucasians. However, there is confl icting data on TG/HDL-C ratio as a Secretion of GLP-1 and PC1/3-PC2 Expression on Pancreatic α-Cell predictor of IR in African Americans. Compared to Caucasians, African VERONICA SANCHO BORNEZ, ROBERTO LUPI, VALENTINA GARAU, DANIELA Americans have lower TG levels and an increased β-cell response to IR LUCCHESI, LAURA GIUSTI, SILVIA DEL GUERRA, GIUSEPPE PENNO, STEFANO despite a greater risk for diabetes. It has been proposed that low TG levels DEL PRATO, Pisa, Italy in African American (AA) women may be due to higher insulin concentrations GLP-1 has been recently suggested to be released by pancreatic α-cells causing reduced free fatty acid levels. We hypothesized that the TG/HDL-C through modulation of the expression and activity of proconvertase 2 and ratio is predictive of β-cell function and/or IR in AA women. Non-diabetic 1/3 (PC1/3, PC2). Little is known, whether this secretion can be affected by AA women (n=41) with a BMI > 25 kg/m2 underwent an intravenous glucose the metabolic alterations occurring in type 2 diabetes. tolerance test. Insulin sensitivity (SI) was determined using Bergman’s Alpha-TC1/6 (TC1/6) cells from a mice pancreatic cell line were incubated minimal model. Insulin secretion was measured as the acute insulin response in the presence of glucose (G) (5.5 and 16.7 mM) for 16 h with or without free to glucose (AIRg) and disposition index (DI) was computed as SI*AIRg. IR was -4 -1 fatty acid (FA, 2:1 palmitate:oleate, 0.5 mM), IL6 (200 ng/ml) or glucagon defi ned as the lowest tertile of SI (<1.8 x 10 min /µU/ml) and inadequate (GLU: 250 pg/ml). We then measured GLP-1 secretion (ELISA), PC1/3 and β cell compensation was defi ned as the lowest tertile of DI (< 900). Data PC2 expression (RT-PCR and western blot), cell viability and apoptosis were analyzed using logistic regression models and area under the receiver (Fluorescein Diacetate and Propidium Iodine fl uorescence). operating characteristic curve (AUC-ROC). An AUC-ROC > 0.70 was defi ned Upon 16.7G incubation, GLP-1 secretion (total and active) was signifi cantly as signifi cant discrimination. The mean (± SD) age was 38.5 ± 11.3 years, increased (130±9% and 158±19%, respectively, p<0.04 vs. 5.5G) along with with BMI of 33.5 ± 6.7 kg/m2 and fasting glucose of 86.5 ± 10.5 mg/dL. The increased PC1/3 expression (mRNA: 1.37±0.06 folds and protein: 138±10%, AUC-ROC for the prediction of DI < 900 was 0.74 indicating that a higher TG/ both p<0.02) whereas PC2 was decreased (mRNA: 0.51±0.06, p<0.05). An HDL-C ratio was associated with decreased DI. However, the AUC-ROC for increment in GLP-1 secretion (173±21%, p<0.02) and PC1/3 expression was prediction of IR (SI<1.8) by TG/HDL-C ratio was 0.45. AUC-ROC for prediction also observed with 5.5G + FA (mRNA: 2.07±0.24 folds, p<0.006). With 16.7G of AIRg <335 µU/ml by TG/HDL-C ratio was 0.60. Alternate cut-offs to + FA, GLP-1 secretion was similar to basal, but under this condition apoptosis defi ne IR as a SI < 2.0 or < 2.3 demonstrated an AUC-ROC of 0.58 and 0.62 was markedly increased (187±38%, p<0.05). On the contrary, apoptosis was respectively. These data suggest that unlike Caucasians, TG/HDL-C ratio in not affected by neither 16.7G (106±9%) nor 5.5G + FA (112±9%; both p=NS). AA women does not predict IR but it is associated with β-cell function. This IL6 also elicited increased GLP-1 release (156±12%) and PC1/3 expression may be due to race differences in lipoprotein lipase activity and lipolysis (2.78±1.04). No effect was apparent when TC1/6 were incubated in the regulation by insulin and high insulin concentrations may contribute to low presence of GLU. TG levels in this group. These data confi rm that the pancreatic α-cell releases GLP-1 and that this secretion can be modulated by metabolic and humoral environment 1894-P similar to the one occurring in type 2 diabetes. More studies are required to Ramadan as a Model of Intermittent Fasting: Effects on Gut Hor- defi ne the potential role of the α-cell secreted GLP-1 in the alterations of the mones, Appetite, and Body Composition in Subjects With and pancreatic islet in diabetes. Without Type 2 Diabetes Mellitus TURKI ALHARBI, DENNIS K. YUE, JENCIA WONG, TANIA MARKOVIC, TED 1892-P WU, BELINDA A. BROOKS, RADHIKA SEIMON, ALICE GIBSON, STEPHANIE L. Exendin-4, Glucagon-like Peptide-1 Receptor Agonist, Inhibits SILVIERA, AMANDA SAINSBURY, TANYA J. LITTLE, Sydney, Australia Cancer Cell Proliferation Depending on GLP-1R Expression Perturbation of circadian rhythms results in adverse metabolic effects. TAKASHI NOMIYAMA, TAKAKO KAWANAMI, YUICHI TERAWAKI, TAKASHI Recent evidence suggests that Intermittent Fasting (IF) can protect against FUKUDA, YURIKO HAMAGUCHI, TOMOKO TANAKA, TOSHIHIKO YANASE, Fukuo- metabolic abnormalities. Ramadan, requiring abstinence from food/drink ka, Japan during daylight hours, presents a unique opportunity to study the metabolic Incretin therapy has emerged as one of the most popular treatment for effects of IF, on a background of circadian rhythm disturbance that would type 2 diabetes. Exendin-4 (Ex-4), has received much attention, because of otherwise predict an adverse metabolic outcome. its’ tissue protective effects beyond glycemic control. We have previously In 5 subjects with type 2 diabetes (T2DM) and 7 healthy controls (C) at reported that Ex-4 attenuates atheroma formation in apoE−/− mice (Arakawa baseline (pre), and in the last week of Ramadan (post), we assessed fasting M, Diabetes, 2010) and also reduces intimal thickening after vascular injury glucose, HbA1c, lipids, body composition (with DXA) and resting energy (Goto H, BBRC, 2011). On the other hand, the most important cause of death expenditure (REE). Plasma gut hormone and appetite responses to a mixed Integrated POSTERS of type 2 diabetes in Japan is cancer. We have previously reported that Ex-4 meal were also measured. Data are means±SEM. inhibit prostate cancer growth both in vitro and in vivo through ERK-MAPK Ramadan resulted in decreased total fat mass (-907±92 g, p=0.001) and Physiology/Obesity inhibition (ADA 2013), suggesting anti-cancer effect of incretin therapy. trunk fat (-778±190 g, p=0.014) in T2DM but not in controls, without any However, prostate cancer is not major cancer in patients in diabetes. Then, reductions in lean mass or REE. There was a trend towards a decline in we next examined anti-cancer effect of Ex-4 in other cancers, such as colon plasma FFA in both groups. Ramadan had no effect on body weight, glycemia, cancer, breast cancer and hepatic cancer. blood pressure, or plasma lipids in either group. First, we examined GLP-1R expression in human cancer cell lines, HT-29 In T2DM only, the area under the curve for post-meal plasma ghrelin cells (colon cancer), MCF-7 cells (breast cancer) and HepG2 cells (hepatic concentrations increased after Ramadan (pre: 6632±1737 vs. post: cancer), using quantitative RT-PCR. We observed abundant GLP-1R 9025±2518 pg/ml.min-1, p=0.045). Despite this increase in orexigenic ghrelin, expression in HT-29 cells and MCF-7 cells similar amount to our previous subjective appetite scores were not altered by Ramadan. Meal-induced report using prostate cancer cells, but not in HepG2 cells (p<0.01). 0.1~10nM plasma concentrations of the satiety hormone pancreatic polypeptide did Ex-4 signifi cantly decreased cell proliferation of HT-29 cells and MCF-7 cells, not change during Ramadan, but were higher in T2DM compared to controls in a dose-dependent manner (p<0.01), while no reduction was observed in (post: C: 23486±6677 vs. T2DM: 62193±6880 pg/ml.min-1, p=0.003).

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A486 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

In conclusion, Ramadan, as a model for IF in the context of circadian rhythm important clinical implications, as severe refractory hyperandrogenism in disturbance, appears to have more favourable effects on body composition in women with extreme insulin resistance could be treated with suppression T2DM, without adverse effects on metabolic control or subjective appetite. of gonadotropins using leuprolide. These data suggest that IF may be particularly benefi cial in T2DM as a nutritional intervention. Larger studies of longer duration are warranted. 1897-P The Novel SIRT1-FGF21 Axis: A Hepatocyte-derived Endocrine 1895-P Signaling that Protects against Weight Gain by Promoting the Browning Adipokines: The Missing Link between Alzheimer’s Disease and of White Adipose Tissue and Increasing Energy Expenditure Type 2 Diabetes? YU LI, KIMBERLY WONG, JONG WOO LEE, MENGWEI ZANG, Boston, MA ADINA MITREA, SARA FALLUCCA, SILVIA ANGELETTI, ANDREEA SOARE, Fibroblast growth factor 21 (FGF21), the hepatocyte-derived hormone, FABRIZIO VERNIERI, FRANCESCA URSINI, LAURA TROTTA, MARIA MOTA, PAOLO is rapidly gaining interest as a promising therapeutic target in diabetes, POZZILLI, Craiova, Romania, Rome, Italy although the upstream regulators of FGF21 remain elusive. The NAD- Recent studies have focused on the importance of adipokines in the dependent deacetylase SIRT1 and its activator, resveratrol, have the development of both Alzheimer’s disease (AD) and type 2 diabetes (T2D), the therapeutic potential on diabetes. However, whether SIRT1 mediates the most studied were adiponectin, leptin and progranulin (PGRN). benefi ts of resveratrol is the subject of debate. We have demonstrated We carried out a pilot study evaluating serum levels of PGRN and that mice with adenoviral overexpression of SIRT1 in the liver are resistant adiponectin and the contribution of these proteins to the association to diet-induced insulin resistance. Here we determined whether loss of between AD and T2D. The study was conducted in 40 subjects: Group 1: 10 hepatic SIRT1 might affect the benefi cial effects of resveratrol. Liver- subjects with both AD and T2D; Group 2: 10 subjects with AD only; Group 3: specifi c SIRT1 knockout mice (SIRT1 LKO) when challenged with a high fat, 10 subjects with T2D only; Group 4: 10 age and sex matched healthy controls. high sucrose (HFHS) diet were prone to developing more severe obesity PGRN and adiponectin were evaluated by ELISA assays. Recorded data were and glucose intolerance, compared to wild-type (WT) mice. Total body analyzed using SPSS 17.00 software and presented as mean±SD. weight and fat mass were increased in SIRT1 LKO mice without affecting Serum levels of PGRN (ng/ml) in the 4 study groups were: 87.2±19.1 in lean mass. Oxygen consumption rate and carbon dioxide production were Group 1; 105.6±33 in Group 2; 132.9±36.7 in Group 3; 99±22.9 in Group 4. reduced, and total energy expenditure was also decreased, suggesting PGRN was signifi cantly lower in Group 1 vs. Group 3 (p=0.006). Although that decreased energy expenditure may be a primary cause of adiposity in not statistically signifi cant (p=0.078), we also noted that PGRN levels in SIRT1 LKO mice. Strikingly, administration of resveratrol (130 mg/kg/day) to T2D were higher than in the control group. Regarding the PGRN/kg, we WT mice protected against diet-induced obesity and insulin resistance, but found statistically signifi cant lower values in Group 1 than in both Group 2 the improvements were diminished in SIRT1 LKO mice. To identify a novel (p=0.023) and Group 3 (p=0.049). downstream of SIRT1, gene chip expression profi ling of the liver of SIRT1 Serum levels of adiponectin (µg/ml), in the 4 study groups were: 18.1±11.6 LKO mice revealed that FGF21 was the most markedly downregulated gene. in Group 1; 25.05±12.9 in Group 2; 16.1±5.3 in Group 3; 25.6±8 in Group 4. Adenoviral delivery of FGF21 into SIRT1 LKO mice, as evidenced by increased We found higher serum adiponectin in females (p=0.01). Furthermore, hepatic and circulating FGF21 levels, restored energy balance through an the gender specifi c statistics showed that adiponectin/kg was signifi cantly increase of energy expenditure and induction of browning-related genes higher in females of Group 1 compared to those in Group 3 (p=0.045), while in white adipose tissue. Thus, activation of the SIRT1-FGF21 axis provide there was no statistically signifi cant difference between adiponectin/kg insight into a previously unrecognized liver-adipose tissue crosstalk that between females with AD and females with both AD and T2D. potentially combats obesity. PGRN and adiponectin serum levels were not correlated in our study Supported By: NIH/NIDDK population. Our fi ndings suggest that adipokines play a role in the association AD with 1898-P T2D, but further studies are needed to elucidate how these two cytokines Effect of Chenodeoxycholic Acid and Colesevelam on Glucose contribute to the association of these diseases. Metabolism in Patients with Type 2 Diabetes and Healthy Control Subjects 1896-P MORTEN HANSEN, MATTHIJS SCHELTEMA, DAVID P. SONNE, JENS F. REHFELD, Pubertal Gonadotropins Are Required for Insulin-induced Hyper- JENS J. HOLST, TINA VILSBØLL, FILIP K. KNOP, Hellerup, Denmark, Copenhagen, androgenism Denmark JALAJA JOSEPH, ELAINE K. COCHRAN, PHILLIP GORDEN, REBECCA J. BROWN, In patients with type 2 diabetes, rectal administration of bile acids, and Bethesda, MD oral bile acid sequestrants lower blood glucose. We evaluated the effect Gonadotropins are physiologic regulators of ovarian growth and of chenodeoxycholic acid (CDCA) and the bile acid sequestrant colesevelam steroidogenesis. In contrast, hyperinsulinemia causes a pathological rise (COL), delivered by intragastric tube, on plasma glucose, insulin, C-peptide, in ovarian volume (OV) and testosterone (T) production. We hypothesized cholecystokinin (CCK), gastric emptying, gallbladder volume, appetite and that hyperinsulinemia could cause ovarian growth in the absence of food intake, in a placebo-controlled, double-blinded study. gonadotropins, but that pubertal levels of gonadotropins are required for On 4 separate days 10 patients with type 2 diabetes (age (mean±SD): 2 hyperinsulinemia-induced ovarian steroidogenesis. 62.37.4 years, BMI: 28.92.3 kg/m ; HbA1c: 7.01.1%) and 10 matched healthy 2 We conducted an analysis of 49 girls (<18y) with insulin resistance due to control subjects (age: 61.39.8 years, BMI: 28.23.2 kg/m ; HbA1c: 5.50.4%) lipodystrophy, insulin receptor mutations, and insulin receptor antibodies, received 1) CDCA (1.25 g); 2) COL (3.75 g), 3) CDCA+COL, or 4) placebo; enrolled in studies at the NIH. Patients were categorized as pre-pubertal suspended in 100 ml of water with 1.5 g acetaminophen (for evaluation of (PRE), mid-pubertal (MID), or post-pubertal (POST) by breast Tanner staging. gastric emptying). During 180 min blood was drawn, gallbladder volume was OV, insulin, and T were measured. evaluated by ultrasound, and appetite was evaluated by visual analogue Pubertal Insulin Ovarian Volume Testosterone Correlation of insulin Correlation of insulin scale. At the end of each day ad libitum food intake was evaluated. stage (mcU/mL) (cc) (ng/dL) and ovarian volume and testosterone None of the interventions changed plasma glucose in either group. In the patients with type 2 diabetes, CDCA, COL and CDCA+COL elicited a

PRE 51.1± 48 3.8±2.2 10.9±7 p=0.28, R2=0.18, m=0.01 p=0.08, R2= 0.08, m= 0.04 Integrated (n=12) (normal < 15) (normal < 2.3) (normal < 20) small but signifi cant (P<0.05, for all) increase in C-peptide/glucose ratio vs. POSTERS placebo, with CDCA eliciting the most pronounced effect. Also in the healthy MID 79.9± 83 9.7±7.8 49.8±81 p=0.02, R2=0.09, m=0.03 p=0.005, R2=0.09, m= 0.3 subjects, CDCA elicited the highest C-peptide/glucose ratio. In the healthy Physiology/Obesity (n=22) (normal < 30) (normal < 6) (normal < 50) subjects, COL increased CCK signifi cantly vs. CDCA, CDCA+COL and placebo, POST 190.7±454 30.5±91.8 62.8±90 p<0.0001, R2=0.5, m=0.17 p=0.02, R2=0.06, m= 0.05 but not in patients with type 2 diabetes. CDCA slowed gastric emptying in (n=33) (normal < 30) (normal < 6.5) (normal < 50) both groups. COL tended to decrease gallbladder volume, whereas CDCA Insulin and OV were increased in all pubertal stages. In MID and POST tended to increase gallbladder volume. The interventions did not affect children, there was a signifi cant positive correlation between insulin and appetite or food intake. OV. A similar but non-signifi cant relationship was observed in PRE children. In conclusion, CDCA, COL and CDCA+COL increased insulin secretion In PRE patients there was no relationship between insulin and T, whereas in (measured as C-peptide/glucose ratio), and CDCA slowed gastric emptying. MID and POST patients there was a signifi cant positive correlation. We speculate that these effects may be mediated by bile acid-induced TGR5 Our fi ndings are consistent with the hypothesis that gonadotropins play activation on L cells and subsequent GLP-1 secretion. a permissive role in hyperinsulinemia-induced hyperandrogenism. This has Supported By: Novo Nordisk Foundation

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A487 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1899-P 1901-P Acute Cardiovascular and Renal Effects of Glucagon-like Peptide-1 Leptin Signaling in Neurons of the Ventromedial Hypothalamic in Healthy Men Nuc leus Is Not Required for Intact Responses to Fasting or Neuro- ALI ASMAR, LENE SIMONSEN, MEENA ASMAR, STEN MADSBAD, JENS J. glucopenia HOLST, ERIK FRANDSEN, CEDRIC MORO, JENS BÜLOW, Copenhagen, Denmark, THOMAS H. MEEK, VINCENT DAMIAN, MICHAEL W. SCHWARTZ, GREGORY J. Hvidovre, Denmark, Toulouse, France MORTON, Seattle, WA Studies have shown confl icting results with regard to cardiovascular and Glutamatergic neurons in the ventral medial hypothalamic nucleus (VMN) renal effects of glucagon-like peptide-1 (GLP-1). The present experiments that express steroidogenic factor 1 (SF1) are implicated in the counter- were performed in order to elucidate the effects of GLP-1 on cardiac and regulatory response (CRR) to hypoglycemia. Since leptin receptors (LepR) renal hemodynamics in healthy men. are expressed by glutamatergic SF1 neurons in the VMN that are depolarized On a controlled diet (2.822 cal/day, 2 mmol Na/kg/day) eight healthy by leptin, and since deletion of LepR from these neurons impairs glucose middle-aged men were examined on two different occacions. During a 3-h homeostasis, we hypothesized that leptin signaling in SF1 neurons is also infusion of either GLP-1 (1.5 pmol/kg/min) or saline, renal plasma fl ow was required to mount an intact CRR to neuroglucopenia. To test this hypothesis, estimated by continuous 51Cr-EDTA infusion and arterial and renal venous we bred SF1-Cre mice with LepRlox/lox mice to delete the LepR within the catheterization. Cardiac output was measured by a Finometer® device, and SF1 neurons (LepR-VMN KO). Consistent with prior reports, on a standard intra-arterial blood pressure was measured in a radial artery. During the chow diet, adult male LepR-VMN KO animals exhibit slightly increased body whole experiment the subjects remained supine. adiposity (p<0.05) despite a similar body weight relative to wild-type (WT) During GLP-1 infusion, arterial blood glucose decreased by 1.3 mmol/L littermates, and are hyperinsulinemic (p<0.05) while remaining euglycemic. with maximum after 40 min (p<0.05), while it remained constant during Relative to WT mice, the effect of a 24-h fast to lower levels of both blood saline infusion. During GLP-1 infusion the systolic blood pressure and arterial glucose and plasma insulin was intact in LepR-VMN KO mice, as was the pulse pressure increased 6 ± 1 mm Hg (p<0.05), whereas blood pressure fasting-induced increase of plasma corticosterone levels (44% and 37% remained unchanged during saline infusion. Heart rate increased 7 ± 2 bpm decrease in glucose; 75% and 70% decrease in insulin; 2.3-fold and 2.4- (p<0.05) and remained unchanged during saline infusion. Cardiac output fold increase in CORT; for WT and KO respectively; p=ns for all genotype increased with 14 % (p<0.05), while no signifi cant changes occurred during comparisons). Similarly, the effect of an intraperitoneal injection of the saline infusion. The renal plasma fl ow and glomerular fi ltration rate remained glucoprivic agent (2-deoxyglucose) to raise blood glucose as well as plasma constant in both experiments. corticosterone and plasma glucagon levels did not differ between LepR- Further plasma noradrenaline concentrations remained constant in both VMN KO and WT mice (40% and 50% increase in CORT; 9-fold and 11-fold experiments, whereas plasma renin concentrations decreased during GLP-1 increase in glucagon for WT and KO respectively; p=ns for all genotype infusion. comparison). Collectively, these data suggest that leptin signaling in SF1 Atrial natriuretic peptide and the urinary sodium excretion remained neurons is not required to mount intact responses either to fasting or to unchanged in both experiments. neuroglucopenia. Thus, leptin-independent mechanisms appear to underlie An acute administration of GLP-1 leads to an increased cardiac output the contribution to glucose counter-regulation made by glutamatergic SF-1 due to a simultaneous increase in stroke volume and heart rate compared to neurons in the VMN. placebo, while having no effect on renal plasma fl ow. The renin-angiotensin- Supported By: NIH (DK089053 to G.J.M); (F32-DK097859 to T.H.M.) aldosterone system is inhibited by GLP-1, whereas the urinary sodium excretion is unaffected. 1902-P AgRP Signaling Is Not Required for Diabetic Hyperglycemia 1900-P THOMAS H. MEEK, MILES E. MATSEN, VINCENT DAMIAN, STREAMSON C. The Association between Incretin Hormones and Bone Metabolism CHUA, MICHAEL W. SCHWARTZ, GREGORY J. MORTON, Seattle, WA, New York, in Patients with Long-term Type 1 Diabetes Mellitus NY BARBARA KATRA, DANUTA FEDAK, ANNA WEDRYCHOWICZ-NIEDZWIECKA, Recent evidence suggests that in rodents with uncontrolled diabetes MACIEJ T. MALECKI, Krakow, Poland (uDM) the anti-diabetic effects of leptin require intact neuronal melanocortin There is growing evidence that incretin hormones may infl uence bone signaling, but that increased melanocortin signaling is not suffi cient to mimic status. However, little is known about the impact of incretins on bone leptin’s glucose-lowering effect. One potential explanation for these fi ndings health in patients with type 1 diabetes (T1DM) and their association with is that in uDM, the melanocortin 3/4 receptor (Mc3/4r) antagonist, AgRP, is osteocalcin (OC) which is considered as a link between glucose and bone expressed at high levels in the hypothalamus. We therefore hypothesized metabolism. that mice lacking AgRP would be 1) protected against hyperglycemia in We aimed to determine the relationships between incretin hormones and uDM, and 2) susceptible to the anti-diabetic effect of increased Mc3/4r bone density as well as undercarboxylated OC (ucOC) and carboxylated OC signaling. To test this hypothesis, we induced uDM using the β-cell toxin, (cOC) in T1DM patients with a long duration of diabetes. streptozotocin (STZ; 150 mg/kg x 2) in AgRP-defi cient mice and their wild- We included 82 T1DM patients. The mean DM duration was 18.87±5.99 type (WT) littermate controls. Both diabetic and nondiabetic control groups yrs, mean HbA1c level was 8,09±1,18. The control group consisted of 53 received chronic icv infusion of either the Mc3/4r agonist MTII (10ug/d) or its apparently healthy individuals matched by age. Blood samples were drawn vehicle. As expected, blood glucose levels were markedly elevated following for the assessment of incretin hormones (GLP-1, GLP-2, GIP), selected STZ administration in WT mice and, although levels were slightly lower in markers of bone metabolism (ucOC, cOC) additional parameters (Ca, P, AgRP-defi cient mice, the difference was not statistically signifi cant (249 25(OH)D3, PTH) and HbA1c. HbA1c was assessed by HPLC, while all the other +/-22 vs. 226 +/-27mg/dl; p=ns), suggesting that AgRP is not required for biochemical parameters were evaluated with ELISA. Bone mineral density diabetic hyperglycemia. As expected, icv MTII did not attenuate diabetic (BMD) was measured in the lumbar spine and femoral neck by DXA scan. hyperglycemia or hyperphagia in WT mice with uDM, whereas the drug T Student test, ANOVA with post hoc Tukey test and multiple regression effectively reduced body weight in non-diabetic WT mice. In AgRP-defi cient analysis were used for statistical analysis. mice with uDM, icv MTII also failed to lower either food intake (7.7 +/-1 for Differences in serum GIP levels were found (p=0.0002) between T1DM WT vs. 7.5 +/-1g/day for AgRP-KO) or blood glucose levels (348 +/-34 for WT Integrated

POSTERS patients (56.3±26.2 pg/ml) and controls (35.8±14 pg/ml); the levels of vs. 310 +/-28mg/dl for AgRP-KO). We conclude that while previous evidence GLP-1 and GLP-2 were not different between the groups. GLP-1 correlated indicates that melanocortin pathway activity is required for leptin’s anti- Physiology/Obesity with neck BMD (g/cm2) (r=0.265, p=0.016) in T1DM patients in univariate diabetic effects, increased melanocortin signaling per se is insuffi cient regression analysis; however, this association was not signifi cant in multiple to ameliorate diabetic hyperglycemia, even when AgRP is absent. These regression. Moreover GLP-1 and GLP-2 levels correlated positively with cOC data suggest that AgRP is not a major driver of diabetic hyperglycemia and level in univariate (r=0.546, p<0.001 and r=0.272, p=0.013) and multivariate that leptin’s glucose-lowering actions involve mechanisms additional to regression (r=0.643, p=0.0005 and r=0.7857, p=0.019, respectively). These increased melanocortin signaling. correlations were not present in the control group. Supported By: NIH (DK089053 to G.J.M); (F32-DK097859 to T.H.M.) In conclusion, we identifi ed a strong association between incretin hormones and some markers of bone metabolism in T1DM patients. Supported By: Polish Diabetes Association

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A488 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1903-P Eight T1DM [7 male 1 female, age 38±6 years, HbA1C 71.3±10.6 mmol/mol] The Effect of Premeal Chewing Gum on Blood Levels of GLP-1 in attended the laboratory fasted and on two separate mornings, having taken Healthy Nonobese Men their usual basal insulin. Participants completed a REST trial or 3SET of RE (8 JIANPING XU, Beijing, China exercises x 10 repetitions) at 67±3%1RM followed by 60 min recovery. Venous Reports varied about of the effect of chewing on blood GLP-1 level. In blood samples were taken before and at 0, 5, 15, 30 and 60 min after exercise. this study, we evaluated the effect of chewing by chewing sugarless Data (mean±SEM) were analyzed using ANOVA (p≤0.05). Resting plasma chewing gum, intended to explore the neural mechanisms of GLP-1 secretion N-arachidonoylethanolamine (AEA) (3SET 0.11±0.01 vs. REST 0.09±0.02 regulation. pmol/ml, p=0.381), 2-arachidonoylglycerol (2-AG) (3SET 1.61±0.44 vs. REST After fasting 12 hours, 12 healthy male non-obese volunteers (18 hunger were evaluated from 0 to 100 of time or condition (p>0.05). Growth hormone (GH) concentrations were at each time point. highest following exercise (3SET 5.1±0.7 vs. CON 0.6±0.3 ng/mL, p<0.001) Compared with the control group, by chewing sugarless gum 80 times but for cortisol there was no effect of time or condition (p>0.05). Under 3SET, every 2 minutes, the satiety of the test group increased at 15, 25 and 30 peak AEA correlated with peak GH concentrations (r=-0.733, p=0.039); peak minutes after the beginning (p = 0.043, p = 0.014 and p = 0.018, respectively), cortisol correlated with immediate post-exercise AEA (r=0.812, p=0.014) and blood GLP-1 level of the test group at 30 minutes was 49.6 ± 20.3pmol /l, peak OEA (r=0.712, p=0.048) and PEA (r=0.719, p=0.044) concentrations, signifi cantly higher than the control group(38.9 ± 20.9 pmol /l, p = 0.031). but OEA and PEA did not correlate with GH. 3SET responses for 2-AG did There was no signifi cant difference in the test group at each time point in not correlate with cortisol, but the rise in concentration from rest to peak GLP-1 levels. In the control group, compared to baseline, levels of GLP-1 at 2-AG correlated with GH values at 60 min post-exercise (r=0.840, p=0.009). 15, 25 and 30 minutes were signifi cantly decreased (p = 0.042, p = 0.0214 and Although RE did not alter resting EC concentrations in T1DM, the strong p = 0.012, respectively). No signifi cant differences in blood levels of glucose, relationships between the glucocorticoid response to RE and the EC system insulin and GIP, and hunger were shown between groups. warrants further investigation. Our study suggests, fasting chewing sugarless gum can increase satiety, Supported By: ERDF and decrease the decline of GLP-1 levels. 1906-P 1904-P Duodenojejunal Bypass Liner Increases Bile Acids Levels in Effects of Acute Manipulation of Serum FFA During Prolonged Patients with Severe Obesity and Type 2 Diabetes Mellitus Fasting on Serum GLP1 Concentrations in Non-Diabetic Patients R. MUÑOZ, J.F. MIQUEL, M. ARRESE, V. IRRIBARRA, F. PIMENTEL, A. SHARP, GUIDO LATTUADA, ANNA SALERNO, GIUSEPPINA MANZONI, FEDERICO D. TURIEL, P. PIZARRO, A. YOUNG, K. BROWN, D. MAGGS, L.M. KAPLAN, A. MARTUCCI, ALICE OLTOLINI, GABRIELE FRAGASSO, ALBERTO MARGONATO, ESCALONA, Santiago, Chile, Research Triangle Park, NC, Lexington, MA, Boston, MA GIANLUCA PERSEGHIN, Monza, Italy, Milan, Italy Endoscopic placement of a duodenojejunal bypass liner (DJBL) has Serum FFA concentration is a metabolic endogenous factor potentially been shown to improve glycemic control in patients with type 2 diabetes able to infl uence incretin response and may play a role in the impairment of mellitus (T2DM). Because DJBL implantation induces nutrient bypass of the GLP1 secretion and/or action in type 2 diabetes. To assess whether acute duodenum, its mechanisms have been proposed to mimic those of Roux-en-Y changes in serum FFA concentrations performed during prolonged fasting gastric bypass (RYGB). Elevated circulating bile acid (BA) concentrations had an effect on incretin hormones level, serum glucose, insulin, c-peptide, appear to mediate some of the metabolic benefi ts of RYGB and other bariatric glucagon, GLP1 and GIP concentrations were assessed in 9 non diabetic procedures, including improved glucose homeostasis. We therefore sought men with stable, chronic heart failure during two experimental conditions to determine whether DJBL implantation increases circulating BA levels in performed 7 days apart. Study 1: before and during a 4-hours i.v. bolus + patients with T2DM. continuous heparin infusion titrated to achieve a serum FFA concentration Seven patients (mean 34.8±9.5 years) with T2DM and obesity (mean body of 1.2 mM. Study 2: before and for 4 hours after the oral administration of mass index 42.6±5.2 kg/m2) underwent frequent blood sampling after a acipimox titrated to achieve a serum FFA concentration of 0.2 mM. Serum standard liquid test meal before and after 52 weeks of DJBL implantation. hormones were measured using a multiplex protein array system enabling Primary BA (PBA) and secondary BA (SBA) levels and their glycine and taurine their simultaneous detection and quantifi cation in a single sample volume. conjugates were measured by HPLC/mass spectroscopy. Serum FFA increased in Study 1 and dropped in Study 2, as experimentally The baseline fasting level of total BAs (TBA) was 0.7±0.3 µmol/L. Fasting planned. Serum glucose, insulin, c-peptide, GLP1 and GIP concentrations all PBA and SBA levels were 0.04±0.01 and 0.07±0.02 µmol/L, respectively. showed small but signifi cant reductions during the prolonged fasting with After 52 weeks of DJBL treatment, fasting TBA levels increased to 4.3±0.8 respect to baseline but only serum GLP1 concentrations were different µmol/L (p<0.05 vs. baseline). At this time, fasting PBA and SBA levels between the two procedures with an early and pronounced reduction during increased to 2.1±0.4 and 1.5±0.4 µmol/L, respectively (p<0.05 vs. baseline Study 1 (- 50±32%) when compared to Study 2 (- 24±23%; p=0.03). Serum for both). Total glycine-and taurine-conjugated BA concentrations exhibited glucagon, in contrast, showed a trend for higher concentrations during the no signifi cant change 52 weeks after device implantation. last part of Study 1 (+4±9%) when compared to Study 2 (-11±12%; p=0.05) Similar to the effects of RYGB, DJBL treatment is associated with increased with respect to baseline. circulating levels of unconjugated PBAs and SBAs. Given the proposed In conclusion, during prolonged fasting conditions an acute, non effects of BAs on glucose homeostasis, these elevated BA concentrations nutritionally mediated increase in serum FFA levels is associated with after DJBL may contribute to the robust therapeutic effects of this device on lower serum GLP1 concentrations in comparison with acute FFA deprivation. T2DM. Further studies of the mechanisms by which DJBL alters circulating This effect seems not to be temporally related to the slightly higher serum BA levels should provide insight into the signaling pathways infl uenced by glucagon concentrations and may underline a direct effect of the circulating this therapeutic device, allowing the identifi cation of novel targets for the FFA on GLP1 homeostasis. effective treatment of T2DM. Integrated

Supported By: Family of Angela Musazzi and Mario Stellato POSTERS 1907-P 1905-P Short-term Effects of NPH Insulin, Insulin Detemir, and Insulin Glar- Physiology/Obesity Intense Resistance Exercise Does Not Alter Circulating Endo can- gine on the IGF-IGFBP-GH Axis in Patients with Type 1 Diabetes nabinoids Despite Signifi cant Elevations in Growth Hormone in JAN FRYSTYK, ZHULIN MA, TORBEN LAURSEN, TORSTEN LAURITZEN, JENS S. Type 1 Diabetes Individuals CHRISTIANSEN, Aarhus, Denmark DANIEL TURNER, STEPHEN D. LUZIO, BENJAMIN J. GRAY, STEPHEN C. BAIN, Insulin-like growth factor I (IGF-I) and growth hormone (GH) regulate insulin MATTHEW D. CAMPBELL, LIAM P. KILDUFF, DANIEL J. WEST, DAVE BARRETT, sensitivity. Insulin regulates the hepatic synthesis of IGF-I and IGF-binding RICHARD M. BRACKEN, Swansea, United Kingdom, Newcastle upon Tyne, United protein 1 (IGFBP-1). In this way insulin indirectly controls the secretion of GH. Kingdom, Nottingham, United Kingdom Insulin and insulin analogues have different receptor affi nities and they may To determine the infl uence of resistance exercise (RE) on endocannabinoid also possess different liver accessibility. Therefore, we found it of interest to (EC) concentrations in type 1 diabetes individuals (T1DM), and explore compare responses of GH and the circulating IGF-system to human NPH insulin, relationships between the EC system and physiological markers of stress. insulin detemir and insulin glargine in patients with type 1 diabetes (T1D).

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A489 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

Seventeen patients (seven women) with T1D (41.9 (24-63) yr (mean and index by homeostasis model assessment ratio (HOMA-R) (r=-0.238, range), BMI 24.7 (19.5-28.3) kg/m2, HbA1c 7.2% (6.3-8.0), T1D duration 26.2 p=0.019), CD16- monocytes (r=-0.252, p=0.003) and CD16+/CD16- monocyte (8-45) yr) were studied using a randomized, three-period crossover design. ratio (r=0.193, p=0.022). GLP-1R levels on CD16+ monocyte were signifi cantly Patients received subcutaneous injections of equal individual doses of NPH, higher than those on CD16- monocytes. (mean fl uorescent intensity 303±127 determir and glargine at 18:00 h. Plasma glucose, serum total IGF-I, bioactive vs. 208±111 (mean±SD), p=0.022). Multiple regression analysis indicated that IGF-I, IGFBPs and GH were measured hourly for 14 hours after injections. systolic blood pressure (β=-0.280, p=0.011), IRI (β=-0.232, p=0.029), CD16+/ As compared to NPH and glargine, detemir resulted in lowest 6-14 hrs CD16- monocyte ratio (β=0.243, p=0.024) and CD16- monocytes (β=-0.243, AUC (mean and range) of IGFBP-1 (1518 [1280-1800]) vs. 1621 [1367-1922] p=0.033) were the signifi cant independent contributors to monocyte GLP- vs. 1020 [860-1210] µg/l x h) and GH (17.1 [14.1-20.6] vs. 15.4 [12.7-18.6] vs. 1R levels. (R2=0.149, p<0.001). These fi nding may suggest atherogenic risk 10.2 [8.5-12.3] µg/l x h), but highest AUC of bioactive IGF-I (3.8 [3.5-4.2] vs. factor and immune status of monocyte are potential modulator of GLP-1R. In 3.7 [3.4-4.0] vs. 4.4 [4.1-4.8] µg/l x h) (all p-values <0.01). Interestingly, these conclusion, blood pressure, IRI and monocyte subset are associated with the differences were unrelated to plasma glucose. By contrast, the profi les of monocyte GLP-1R levels in type 2 diabetes. total IGF-I, IGFBP-2 and IGFBP-3 were comparable between the three insulin Supported By: Japan Society for the Promotion of Science (23791041) preparations. Independent of plasma glucose, a single dose of detemir caused larger 1910-P suppression in serum IGFBP-1 than NPH and glargine, whereas levels of Identifi cation and Characterization of Glucagon-like Peptide-1 bioactive IGF-I were higher, hereby explaining the lower GH levels. Thus, Receptor Expressing Neurons and Thyroid C-Cells Using a Trans- detemir appears to be more liver specifi c than NPH insulin and glargine. genic Mouse Model The physiological signifi cances of this observation remains, however, to be PAUL RICHARDS, FRANK REIMANN, FIONA M. GRIBBLE, Cambridge, United determined. Kingdom Supported By: Novo Nordisk A/S Glucagon-like peptide-1 (GLP-1) is an intestinal hormone with widespread actions on metabolism. Therapies based on GLP-1 are highly effective in 1908-P treating diabetes, but reports suggest that GLP-1 has additional benefi cial, Subtle Defects in Glucose Metabolism Prior to Development of Dia- or in some cases potentially dangerous, actions on other tissues. Long term betes in Patients with Cystic Fibrosis: The Role of Gut Hor mones treatment of mice with GLP-1 mimetics, for example, causes thyroid C-cell LIMOR MARKO, KAREN J. HERSHKOP, NOA ESHEL, HILA ELIASHAR IRON, hyperplasia, and concerns have been raised that GLP-1-based treatments MELANA COHEN, EITAN KEREM, RAM WEISS, Jerusalem, Israel may be linked to pancreatic cancer in humans. The lack of specifi c antibodies Cystic fi brosis related diabetes (CFRD) is a common co-morbidity in people to the GLP-1 receptor (GLP1R) has hindered reliable identifi cation of GLP-1 with CF. The underlying pathophysiology of CFRD has not been revealed. target cells. To counter this we generated transgenic mice in which Cre- We hypothesized that it might be associated with an abnormal gut derived recombinase is driven by the glp1r promoter. Our objective was to identify hormonal profi le, specifi cally of lower incretin hormone responses, prior to and characterise cellular targets of GLP-1 in mice. development of CFRD. Glp1r-cre mice were crossed with reporter strains expressing red Ten non-diabetic CF patients (5F/5M) and nine healthy controls underwent fl uorescent protein (tdRFP) or the Ca2+ sensor GCaMP3, to label cells an oral glucose tolerance test (OGTT) for evaluation of insulin secretion, expressing glp1r. Initial work identifi ed glp1r-fl uorescence in vascular smooth sensitivity and incretin dynamics. muscle, pancreatic β and δ-cells, enteric neurones, vagal ganglia, brainstem, Fasting and 2-hour glucose levels of patients with CF were signifi cantly hypothalamus and rare pancreatic ductal cells. In follow up studies, we found higher compared to healthy controls. In the patients with CF the peak glucose no glp1r-fl uorescence in white or brown adipose cells or skeletal muscle. By occurred later (60 min vs. 30 min in controls) and was higher (205.5±12.8 mg/ immunomicroscopy, ~50% of thyroidal C-cells (identifi ed by calcitonin/CGRP dL vs. 117.2±9.88 mg/dL compared to normal controls. Insulin levels were staining) exhibited RFP-fl uorescence. signifi cantly lower in the CF group at 30-min, compared to control group. Ca2+ concentrations were monitored in glp1r-fl uorescent cells using C-peptide levels were signifi cantly lower in the CF group at fasting and all GCaMP3 or fura2. Primary cultured thyroidal C-cells showed Ca2+ responses time points and they had a lower and delayed peak compared to controls. to KCl (n=12), and subpopulations responded to cholecystokinin (CCK) and Gut hormone response to OGTT was lower in the CF patients yet shared GLP-1. Glp1r-RFP fl uorescent myenteric neurones showed Ca2+ responses the same magnitude as the normal reference group. AUC for GLP-1and GIP to KCl (n=111) and acetylcholine, and some responded to ATP, CCK and was signifi cantly lower for CF patients compared to control (p=0.02, p=0.04 serotonin. respectively). A similar trend was seen for PYY levels yet did not reach Our data suggest that GLP-1 directly targets a range of tissues. Afferent statistical signifi cance. CF patients had lower insulin sensitivity (p=0.022), neurones and pancreatic β-cells may underlie effects on food intake and a lower acute insulin response (p<0.0001) and the oral disposition index was insulin secretion. In the mouse, GLP-1 directly targets thyroid C-cells. signifi cantly lower in CF patients compared to controls (p<0.0001) as well as Supported By: Wellcome Trust (WT084210\Z\07|Z, WT088357\Z\09\Z) the insulin clearance (p=0.002). We propose that impaired incretin hormone response is an additional 1911-P contributor to the pathophysiology of CFRD. This may be due to lower Nutrient-induced Suppression of Thyroid Stimulating Hormone in secretion responses or due to reduced effects at the cellular level. Patients with Type 2 Diabetes—The Role of Gallbladder Emptying and Gut Incretin Hormones 1909-P DAVID P. SONNE, ASGER LUND, JENS FABER, JENS J. HOLST, TINA VILSBØLL, Risk Factors of Atherosclerosis and Immune Status of Monocytes FILIP K. KNOP, Copenhagen, Denmark, Hellerup, Denmark, Herlev, Denmark Are Associated with Circulating Monocyte GLP-1R Levels in Type Bile acids (BAs) are possible candidate agents in newly identifi ed 2 Diabetes pathways through which carbohydrate metabolism, lipid metabolism and KOKA MOTOYAMA, MASANORI EMOTO, TAKASHI MISHIMA, YUKO YAMAZAKI, energy expenditure are regulated. Preclinical studies suggest that BAs TOMOAKI MORIOKA, KATSUHITO MORI, SHINYA FUKUMOTO, TETSUO SHOJI, activate the enzyme type 2 iodothyronine deiodinase, which deiodinates MASAAKI INABA, Osaka, Japan thyroxine (T4) to the biologically active triiodothyronine (T3). We aimed to Integrated POSTERS GLP-1 receptors (GLP-1R) are expressed not only on islet cells or intestine evaluate the infl uence of various postprandial factors on thyroid function but also on brain, heart and leukocytes. GLP-1R mediates the effects of parameters in patients with type 2 diabetes. Physiology/Obesity incretin axis and GLP-1R expression levels may modulate incretin signals. Thyroid stimulating hormone (TSH) and thyroid hormones (total T3 and free T4) However, the regulation of GLP-1R in type2 diabetes is unknown. In this study, were measured in plasma from two human studies: I) 75g-OGTT and 3 isocaloric we focused on the expression levels of GLP-1R on circulating monocytes and (500 kcal) and isovolemic (350 ml) liquid meals of varying fat and carbohydrate aimed to investigate the factors associate with GLP-1R levels on monocytes content with concomitant ultrasonographic evaluation of gallbladder emptying in type2 diabetes. 141 type 2 diabetic patients (age; 62 ± 14 (SD) years, in 15 patients with type 2 diabetes and 15 healthy age, gender and BMI-matched duration; 14 ± 12 (SD) years) were enrolled in this cross-sectional study. Cell controls (meal-study); and II) 50g-OGTT and isoglycemic iv glucose infusions surface GLP-1R on circulating monocyte was fl uorescently labeled and GLP- (IIGI) alone or in combination with glucose-dependent insulinotropic polypeptide 1R levels were quantifi ed by fl ow cytometry. In simple regression analysis, (GIP), glucagon-like peptide-1 (GLP-1), and glucagon-like peptide-2 (GLP-2), in 10 monocyte GLP-1R levels are signifi cantly associated with smoking index (r=- patients with type 2 diabetes (IIGI-study). 0.165, p=0.050), triglyceride (r=-0.163, p=0.050), serum creatinine (r=-0.176, In the meal-study, gallbladder emptying increased with increasing meal p=0.036), fasting serum insulin (IRI) (r=-0.225, p=0.026), insulin resistance fat content in both type 2 diabetes patients and controls (P<0.01), but no

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A490 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES intergroup differences were demonstrated (P=NS). In both studies, TSH evaluated as a preventive and inhibitory agent for HCC. Our study, for the levels declined (P<0.01) to the same extent after all meal and infusion fi rst time, suggests the possible clinical use of Ex-4 as therapies that could stimuli, respectively. T3 and T4 concentrations did not change in response to reduce HCC risk in type 2 diabetic patients. any of the applied stimuli. TSH levels declined independently of the degree Supported By: NSFC (81170722, 81270438); Chinese University of Hong Kong of gallbladder emptying, route of nutrient administration and infusion of gut hormones. 1914-P In conclusion, intestinal bile fl ow and iv infusions of GIP, GLP-1 or GLP-2 Effect of Rosiglitazone on Acid Labile Subunit Expression in do not seem to affect thyroid function parameters. Thus, the presence of a Adipocytes and Patients with Type 2 Diabetes Mellitus ‘gut-thyroid-pituitary’ axis seems questionable. YING-CHUEN LAI, LEE-MING CHUANG, Yunlin, Taiwan, Taipei, Taiwan Supported By: Novo Nordisk Foundation Acid-labile subunit (ALS) is an important protein for maintaining insulin- like growth factor system. Recently, ALS was found to control animal growth 1912-P as well as carbohydrate and fat metabolism. The aim of the study was to Effect of Known Disruptors of Peripheral Clocks, High-Fat Diet, and evaluate the change of ALS expression in adipocytes and patients with type Constant Glucocorticoid Levels on the Daily Rhythm of Glucagon- 2 diabetes mellitus after rosiglitazone treatment. like Peptide-1 (GLP-1) Secretory Responses Mouse 3T3-L1 preadipocytes were inducted into adipocytes in adipogenic MANUEL GIL-LOZANO, W. KELLY WU, PATRICIA BRUBAKER, Toronto, ON, media. ALS expression was analyzed by Western blot and reverse Canada transcription-polymerase chain reaction during adipocytes differentiation We have previously shown the existence of a diurnal rhythm in GLP-1 and after rosiglitazone treatment (4.5 µM). To evaluate the effect of secretory responses to oral glucose in rats that correlates with the pattern rosiglitazone on circulating ALS in vivo, we measured serum ALS levels at in insulin secretion. Since a circadian pattern of GLP-1 responses was also three time points over 24 weeks in 61 diabetic subjects randomized to either found in vitro, this rhythm may be driven by an independent clock in the rosiglitazone (2mg/day) or placebo group. L-cell. Herein, we investigated the effects of two well-established disruptors In vitro, ALS expression was up-regulated during adipocyte differentiation of peripheral clocks, high fat diet and constant glucocorticoid levels, on the and reduced by rosiglitazone. In clinical studies, serum ALS levels signifi cantly diurnal rhythm of GLP-1 responses in rats. correlated with age, gender, body mass index (BMI), and triglyceride. At the Male Wistar rats were fed regular chow or high-fat high-sucrose (HFHS) end of study, HbA1c and fasting plasma glucose decreased in rosiglitazone diet for 5wk (n=7 each). Other rats were implanted with placebo or 50 mg group as compared to placebo group. BMI, total cholesterol, low-density corticosterone (B) pellets for 2wk (n=6-8 each). GLP-1, insulin and glucose lipoprotein cholesterol, and homeostatic model assessment version 2 β-cell responses to oral glucose were examined at ZT10 vs. ZT22, previously shown function increased in subjected treated with rosiglitazone over time. Overall, as peak vs. trough, resp. of both GLP-1 and insulin responses. rosiglitazone did not decrease ALS serum levels (p=0.474). In subgroup HFHS-fed rats gained more weight than chow animals (237±9 g vs. 180±9 analysis, rosiglitazone decreased ALS in the group with middle tertile of g, resp. p<.001) and had a disrupted pattern of food intake, consuming more baseline ALS levels (adjusted p=0.047). Multivariate linear regression calories during the light hours (p<.05). Control animals showed the expected analysis showed change in serum ALS levels correlated with rosiglitazone rhythm in GLP-1 and insulin responses, with higher responses at ZT10 (p<.05- treatment, age, and change in body weight, total cholesterol, HbA1c and .01). In contrast, this rhythm was abolished in the HFHS-fed rats, showing fasting plasma insulin. augmented responses at ZT22 and, thus, comparable responses at both time In conclusion, ALS expression is up-regulated during adipocyte points. No circadian variation in B levels was observed in the animals with differentiation. Rosiglitazone suppresses ALS expression in vitro and B pellets, showing constant levels around the daily average. Surprisingly, in vivo. Change in ALS may associate with age, change in glycaemia, although these animals showed the same diurnal variation in GLP-1 and hyperinsulinemia, and adiposity. insulin responses as controls, the magnitude of these responses was increased in the rats with constant B levels (p<.05). 1915-P Our data demonstrate that known disruptors of peripheral circadian Glucose Stimulates GLP-1 Secretion from Isolated Perfused Rat clocks exert differential effects on the diurnal variation in GLP-1 and insulin Small Intestine by SGLT1 and GLUT2 Mediated Uptake, Causing secretory responses to causealterations in the timing (HFHS diet) and V-gated Calcium Channel Activation magnitude (constant B) of the responses to nutrient ingestion. RUNE E. KUHRE, CHARLOTTE RASMUSSEN, BERIT SVENDSEN, JENS J. HOLST, Copenhagen, Denmark 1913-P We characterized the mechanisms of glucose-stimulated glucagon-like Activation of Receptor of GLP-1, but Not GIP, Inhibits Tumorigenesis peptide-1 (GLP-1) secretion from isolated rat small intestine perfused at in Animal Models of Hepatocellular Carcinoma constant rate. Luminal glucose (5 and 20 % (w/v)) stimulated secretion dose MINGYAN ZHOU, ALFRED S.L. CHENG, YU HUANG, KAI WING TSE, GANG XU, dependently (ΔGLP-15% (w/v): 4.77 ± 0.8 vs. GLP-120% (w/v): 18.5 ± 2.8 pM, P< 2 Sha Tin, China, Kowloon Tong, Hong Kong 0.0001, n = 6) at rates correlating with glucose absorption (R 5% (w/v) = 0.68, 2 Recent epidemiological studies revealed the increasing frequency of R 20% (w/v) = 0.87, P < 0.001, n = 6), independent of the osmotic load. Vascular hepatocellular carcinoma (HCC) in patients with type 2 diabetes. The glucose stimulated secretion at 15 mM (21.7 ± 0.5 vs. 25.2 ± 0.6 pmol/l, P < incretin-based therapies, including GLP-1 receptor activation and DPPIV 0.001, n = 6), but not at 5 and 10 mM (P > 0.05). Luminal glucose activated inhibitors, are widely used for the treatment of diabetes. Both GLP-1 voltage-gated calcium channels in the L-cells, as inhibition with nifedipine (NI; receptor agonist exendin-4 (Ex-4) and the GIP active form (D-GIP) are 10 µM) or hyperpolarization with diazoxide completely abolished secretory extremely potent and effective anti-diabetic drugs. However, there are responses (Glu. 9.19 ± 0.6 vs. 25.1 ± 0.56 pM, P < 0.01, Glu.+NI: 7.90 ± reports on the risk of pancreatic and thyroid cancers after the treatment. 0.56 vs. 10.57 ± 1.92 pM, P > 0.05, n = 6) (Glu.: 12.0 ± 0.6 vs. 31.8 ± 1.9 pM, P While diabetes treatment may reduce risk for HCC, chronic administration < 0.05, Glu. + DZ: 20.1 ± 1.0 vs. 20.9 ± 0.7 pM, P > 0.05, n = 6). Nifedipine also of Ex-4 raises concern of undesired induction of hepatocellular malignant eliminated depolarization driven GLP-1 responses to vascular KCl (50 mM) transformation. To test whether incretin hormone has any tumor-promotion (KCl: 8.60 ± 0.4 vs. 47.8 ± 13.0 pM, P < 0.05, KCl+NI: 7.83 ± 0.49 vs. 15.23 ± potential, we applied chronic daily Ex-4 and D-GIP treatment in two HCC 1.32 pM, P < 0.05, n = 6). The non-metabolizable SGLT1 substrate, α-MGP, Integrated models, namely spontaneously HCC development model (HBx transgenic stimulated release (15.7 ± 0.5 vs. 29.4 ± 0.7 pM, P < 0.0001, n = 6), while both POSTERS mice) and carcinogen (Diethylnitrosamine)-induced HCC model. The in vivo α-MGP and glucose failed to stimulate GLP-1 secretion in presence of the study demonstrated Ex-4 did not increase, but reduced HCC incidence SGTL1 inhibitor phloridzin (10 µM) (P > 0.05). However, glucose-stimulated Physiology/Obesity by 58% in HBx mice. In contrast, D-GIP did not exert any effect on HCC GLP-1 secretion was also sensitive to luminal GLUT2 inhibition (1 mM incidence. In carcinogen-induced HCC model, Ex-4 treatment decreased phloretin) (Glu. 13.94 ± 2.5 vs. 30.90 ± 10.9 pM, Glu.+PT: 12.44 ± 1.3 vs. 17.18 ± tumor multiplicity by 70%. The amount of proliferating cells determined by 3.4 pM, P > 0.0001, n = 6), and KATP-channel closure by two sulfonylurea Ki67 positive staining was reduced by Ex-4 in carcinogen-induced HCC mice. drugs stimulated GLP-1 release (tolbutamide: 14.7 ± 0.2 vs. 28.6 ± 5.2 pM; Moreover, in vitro studies showed that Ex-4 reduced cell number in both gliclazide: 15.2 ± 0.3 vs. 24.6 ± 2.4 pM, P < 0.05, n = 8). Our data indicates dose and time dependent manners in HepG2 hepatoma cells. Furthermore, that SGLT1 activity is the driving force for glucose induced GLP-1 secretion, Ex-4 inactivated the proliferation-related epidermal growth factor receptor though secretory responses may be potentiated by GLUT2 activity and KATP- signaling pathway and this effect was largely attenuated by PKA inhibitor channel closure. H-89. Taken together, these results show that Ex-4 could decrease the HCC Supported By: Novo Nordisk Foundation incidence at least by inhibition of cancer cell proliferation, which should be

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A491 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1916-P 1918-P DNA Methylation, Gene Expression and Plasma Levels of Leptin in Regulation of Glucose Tolerance via the Proopiomelanocortin Young Low Birth Weight Men and Controls, Before and After a 36- (Pomc) Glucagon-like Peptide-1 (Glp1) Receptor (Glp1r) Hour Fasting Intervention MELISSA A. BURMEISTER, ADRIANA LANDIVAR-ROCHA, HANNAH SMOUSE, LINE HJORT, SINE WANDA JØRGENSEN, CHARLOTTE BRØNS, LINN GILLBERG, JENNIFER E. AYALA, DORIS A. STOFFERS, RANDY J. SEELEY, JULIO E. AYALA, ELIN HALL, CHARLOTTE A. LING, ALLAN A. VAAG, Copenhagen, Denmark, Malmö, Orlando, FL, Philadelphia, PA, Cincinnati, OH Sweden Glp1 regulates glucose tolerance via pancreatic Glp1r-mediated stimulation Low birth weight (LBW) is associated with increased risk of Type 2 of insulin secretion. Pharmacological Glp1r activation in the hypothalamus is Diabetes (T2D), increased fat mass and differential expression of adipokines, also glucoregulatory. Here we test the hypothesis that the hypothalamic Glp1r including leptin (LEP). Knowing that plasma leptin levels decline with fasting, regulates glucose tolerance in response to endogenous Glp1. Hypothalamic we aimed to study if this is associated with transcriptional regulation of Glp1r knockout mice (HypoKO) were generated (Nkx2-1 Cre bred to Glp1rfl ox/ LEP by changes in DNA methylation degree and whether leptin expression fl ox mice) and compared to controls (C) at 12 wks on chow. Glucose area under is regulated differently in LBW and normal birth weight (NBW) individuals the curve (AUC) was measured following oral and intraperitoneal (ip) glucose before and after a fasting challenge. tolerance tests (GTT). AUC was not different between HypoKO and C during 20 LBW men and 17 matched NBW controls were studied during a 36 hour oral (8059±684 vs. 7593±973 mg·min·dL-1) or ip (16483±1421 vs. 17187±2628 fasting intervention. After 8-12 weeks, a subset of both groups participated mg·min·dL-1) GTT. Since the Glp1r is expressed in multiple hypothalamic in an overnight fasting control study. After the overnight or 36 hours fast nuclei, we generated arcuate (ARC) Pomc-specifi c Glp1r knockout mice subcutaneous adipose tissue biopsies were excised. P-leptin levels were (PomcKO) using Pomc-Cre mice. Pharmacological doses of Glp1 targeted to measured after 12, 36, 39 and 42 hours. Adipose tissue DNA methylation the ARC exert glucoregulatory effects, and the Glp1r is primarily expressed degree was studied at 39 CpG sites in the proximal LEP promoter region and in Pomc neurons. Oral glucose tolerance was unaffected in male PomcKO. LEP mRNA expression levels were measured. Interestingly, female PomcKO showed lower AUC compared to C (5637±201 Average DNA methylation degree of the LEP promoter was after the vs. 9511±749 mg·min·dL-1). Ip glucose tolerance was unaffected in PomcKO overnight fast higher (~3%) in LBW compared to NBW subjects (p<0.05). of either sex. Improved glucose tolerance in female PomcKO was surprising After 36 hours fasting the average methylation degree of the LEP promoter since we show that targeted delivery of the Glp1r agonist Exendin-4 was increased (~5%) compared to the control study in NBW subjects only. (Ex4) to the ARC also lowers AUC compared to vehicle during an ip GTT No differences were found in LEP mRNA expression between the birth (16692±2425 vs. 25845±2795 mg·min·dL-1). In sum, loss of Glp1r in the whole weight groups, neither did LEP mRNA expression change with fasting. hypothalamus does not affect glucose tolerance, but deletion of the Glp1r in P-leptin levels were higher among LBW subjects after 12 hours of fasting Pomc neurons improves glucose tolerance in female mice. Pharmacological (p=0.04) and decreased more than 3-fold in both groups after 36 hours activation of ARC Glp1r also improves glucose tolerance. Thus, the ARC Glp1r fasting (p<0.0001). is a regulator of glucose tolerance, and this is dependent on sex and on the LBW subjects were characterized by higher baseline DNA methylation targeting of pharmacological Glp1r agonists to this site. This proposes novel degree at the proximal LEP promoter in adipose tissue and higher baseline incretin-independent glucoregulatory mechanisms for Glp1r agonists that p-leptin levels. A short-time fasting challenge induced small but signifi cant merit further exploration as therapeutic targets for diabetes. changes in LEP promoter DNA methylation in NBW subjects only. No evidence was found to support the hypothesis that DNA methylation changes of the 1919-P LEP promoter regulate the drastic decrease in p-leptin during fasting. Hypothalamic Lipid Sensing Mechanism Is Not Required for Gluca- gon to Regulate Glucose Production 1917-P MARY P. LAPIERRE, MONA A. ABRAHAM, JESSICA T.Y. YUE, GUY A. RUTTER, Decrease Glyoxalase 1 Activity Increases Dicarbonyl Stress in PETER E. LIGHT, BEATRICE M. FILIPPI, TONY K.T. LAM, Toronto, ON, Canada, Periodontal Ligament Fibroblasts in Model Hyperglycaemia London, United Kingdom, Edmonton, ON, Canada AMAL ASHOUR, PAUL J. THORNALLEY, NAILA RABBANI, Coventry, United Direct administration of glucagon into the mediobasal hypothalamus Kingdom (MBH) of rodents under a pancreatic (basal-insulin) euglycemic clamp inhibits Periodontal ligament infl ammation (periodontitis) is a common disease hepatic glucose production (GP), in contrast to the hormone’s systemic characterized by gradual destruction of connective tissue fi bres that attach effect on GP. The downstream signaling of this MBH glucagon action remain a tooth to the alveolar bone within which it sits. Diabetes and infl ammation unknown. In parallel, a lipid-sensing pathway involving MBH AMPK -> enhances periodontal bone loss through enhanced resorption and diminished malonyl-CoA -> CPT-1 -> LCFA-CoA -> PKC-δ leading to the activation of bone formation. It has been shown that human PDL fi broblast (hPDLF) KATP channels lowers GP. Given that glucagon signals through the MBH attachment and function to type 1 collagen was impaired by methylglyoxal PKA to lower GP, and that PKA inhibits AMPK in hypothalamic cell lines, (MG) modifi cation in vitro. We hypothesise that increased hPDLF detachment we tested whether MBH glucagon-PKA inhibits AMPK, elevates LCFA-CoA and dysfunction may be mediated by increased exposure to high glucose levels to activate PKC-δ, and activates KATP channels to lower GP. Here, we concentration with increased formation of MG, impaired MG metabolism fi rst confi rmed that direct administration of glucagon, but not saline, into the by glyoxalase 1 (Glo1) and increased formation of MG-derived advanced MBH of rats during a pancreatic clamp increased the glucose infusion rate glycation endproducts (AGEs). The aim of this study was to evaluate the required to maintain euglycemia (5.8 ± 0.8 vs. 1.1 ± 0.5 mg/kg/min, p= 0.002) effects of high and low glucose concentrations on MG metabolism in due to a suppression of GP (4.3 ± 1.3 vs. 8.7 ± 0.6 mg/kg/min, p= 0.02) with no hPDLFs. changes in glucose uptake (10.2 ± 0.7 vs. 9.3 ± 0.4 mg/kg/min). Co-infusion hPDLFs were cultured in low and high glucose media (8 mM and 25 mM, of glucagon and AMPK activator AICAR failed to negate the effect of MBH respectively) to model hyperglycaemic conditions for three days; n = 3. The glucagon on GP suppression, compared to saline (3.8 ± 0.9 vs. 8.8 ± 0.7, p= activity of Glo1, concentration of MG, MG-derived AGE, MG-H1, and fl ux of 0.0007). MBH glucagon was equally potent to lower GP in rats expressing D-lactate (surrogate of MG fl ux) were measured. There was a signifi cant MBH CA AMPK compared to Ad-GFP (3.9 ± 0.8 vs. 3.4 ± 0.9). PKC-δ inhibitor decrease in Glo1 activity (42.5%, p<0.001), increase in fl ux of D-lactate (42%, rottlerin also failed to abolish the MBH glucagon-induced GP suppression, p<0.012) and increased cellular concentration of MG (41%, <0.002) in hPDLFs compared to saline (5.8 ± 1.0 vs. 8.7 ± 0.6, p= 0.03). MBH glucagon lowered Integrated POSTERS incubated with high glucose concentration. Relatedly, there was increased GP in rats injected with MBH DN PKC-δ and control LacZ virus (4.9 ± 1.3 vs. MG-H1 content of cell protein (85%, p<0.05) and fl ux of MG-H1 free adduct 5.4 ± 1.2). In contrast, KATP channel inhibitor glibenclamide negated MBH Physiology/Obesity (107%, p<0.0001). We conclude that hPDLFs suffer dicarbonyl stress and glucagon’s effect to lower GP compared to saline (9.7 ± 1.4 vs. 9.1 ± 0.7). The MG-derived AGE accumulation in high glucose concentration. Similar GP-lowering effect was also abolished in rats expressing MBH DN Kir6.2 changes induced by hyperglycemia in vivo may contribute to increased AAA compared to Ad-GFP injected rats (9.6 ± 0.8 vs. 3.4 ± 0.9, p= 0.001). hPDLF dysfunction and development of periodontitis in diabetes. Thus, MBH glucagon signals through a lipid-sensing (i.e., AMPK and PKC-δ) Supported By: Saudi Arabia Ministry of Education independent but KATP channel-dependent pathway to regulate GP. Supported By: CDA

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A492 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1920-P 1922-P Effect of Lowering Plasma Glucose and FFA Concentrations on Expression of Products from D, K, and L Enteroendocrine Cells along Insulin Sensitivity and Beta-Cell Function in T2DM Individuals the Entire Intestinal Tract in Patients with Type 2 Diabetes and Non- AURORA MEROVCI, CAROLINA SOLIS-HERRERA, JUAN XIONG, GIUSEPPE Diabetic Controls Reveals Altered Glucagon and Somatostatin DANIELE, ALBERTO CHAVEZ-VELAZQUEZ, DEVJIT TRIPATHY, RALPH A. DE- Expression in Type 2 Diabetes FRONZO, MUHAMMAD A. ABDUL-GHANI, San Antonio, TX JENS PEDERSEN, NICOLAI A. RHEE, SARA L. JEPSEN, CAMILLA D. WAHLGREN, Aim: Chronic elevation in plasma FFA (lipotoxicity) and plasma glucose TINA JORSAL, BRYNJULF MORTENSEN, PETER VILMANN, HAZEM HASSAN, (gluco-toxicity) concentrations have been shown to have detrimental JAKOB W. HENDEL, STEEN S. POULSEN, JENS J. HOLST, TINA VILSBØLL, FILIP K. effect on both core defects (insulin resistance and beta cell dysfunction) KNOP, Copenhagen, Denmark, Hellerup, Denmark, Herlev, Denmark in T2DM. We have shown that lowering plasma FFA and plasma glucose Little is known about the organization of the enteroendocrine cells along concentrations separately in T2DM individuals improves beta cell function the intestinal tract. We used double-balloon enteroscopy (DBE) to collect and ameliorate insulin resistance. In the present study, we examined the mucosal gut biopsies with the aim to evaluate the expression of products effect of lowering both the plasma FFA and glucose concentrations on beta from enteroendocrine D, K and L cells along the entire intestinal tract in cell function and insulin sensitivity in T2DM individuals. subjects with type 2 diabetes (T2D) and in healthy controls. Methods: 12 T2DM subjects (age=49 ±3, BMI=31.1± 1.8, HbA1c = 8.5 ± 0.3, The study involved 12 subjects diagnosed with T2D and 12 age and body diabetes duration = 8.5 ± 0.3 years) received treatment with dapaglifl ozin (10 mass index matched non-diabetic controls. All subjects underwent antero- mg/day) for 3 weeks and during the third week they also received acipimox and retrograde DBE. Biopsies were collected from every 30 cm from pylorus (250 mg four times daily). 75 gram OGTT and euglycemic insulin clamp were to the ileocecal valve and from 6 specifi c sites in colon. RNA was isolated performed at baseline and at weeks 2 and 3. and analyzed for expression of glucagon (GCG), peptide YY (PYY), glucose- Results: Compared to baseline, dapaglifl ozin lowered the fasting plasma dependent insulinotropic polypeptide (GIP) and somatostatin (SST). glucose concentration at 2 weeks (181±9 to 146±9, p<0.01); the decrease Expression of markers for L cells (GCG, PYY) and K cells (GIP) showed in (FPG) was accompanied by signifi cant increase in insulin-stimulated signifi cantly higher expression of L cell genes in the distal versus the total glucose disposal (TGD) (4.16 ±0.71 to 5.15±0.73, p<0.05), and beta proximal parts of the small intestine and signifi cantly higher expression of K cell function. ΔC-pep/ΔG[0-120]÷IR increased by 105% (p=0.003). At week cell marker (GIP) in the proximal part of the small intestine. Expression of the 3, plasma FFA concentration was decreased by 28% compared to week 2 L cell marker (GCG) was signifi cantly higher in the proximal small intestine and this decrease was accompanied by a small, non-signifi cant increase in subjects with T2D compared to healthy controls (P<0.05). Expression of (by 9%) in insulin-mediated TGD, beta cell function (ΔC-pep/ΔG[0-120]÷IR)÷IR the D cell marker (SST) was signifi cantly lower in the distal part of the small increased further by 53% (0.15±0.03 to 0.23 ±0.04, p=0.02). intestine in T2DM subjects compared with healthy controls P<0.05). Conclusion: Simultaneously lowering the plasma glucose and FFA con- Our study in patients with type 2 diabetes and non-diabetic controls centrations has an additive effect to improve beta cell function, but not to confi rmed the notion of higher expression of the L cell marker (GCG) distally, improve insulin sensitivity. and higher expression of the K cell product (GIP) proximally in the small Supported By: NIH intestine. Importantly, we show that intestinal expression of GCG and SST is altered in patients with type 2 diabetes. This supports the concept of 1921-P the enteroendocrine system as highly dynamic system which is modifi ed by High-Density Lipoproteins Mediate Intercellular Communication in internal and external factors. Type 2 Diabetes Through tRNA-derived Small RNAs Supported By: Novo Nordisk Foundation LESLIE A. ROTETA, QUANHU SHENG, YAN GUO, CARINE BEYSEN, SCOTT TURNER, KASEY C. VICKERS, Nashville, TN, Emeryville, CA 1923-P Small RNAs, namely microRNAs (miRNA), have proven to be powerful Serum Concentrations and Adipose Tissue Expression of the regulators of glucose metabolism and contribute to the pathogenesis of type Macrophage Activation Marker CD163 in Subjects with Type 2 2 diabetes (T2D). Recently, we reported that high-density lipoproteins (HDL) Diabetes Mellitus and Obesity: The Infl uence of VLCD and Gastric transport and deliver functional miRNAs to recipient cells. However, miRNAs Plication are only a small fraction (16%) and small RNAs derived from tRNAs (tDRs) MILOS MRAZ, JANA DRAPALOVA, ZDENKA LACINOVA, PETRA KAVALKOVA, account for over 60% of HDL-RNAs. Here we report that pancreatic beta PAVEL TRACHTA, MIKULAS KOSAK, ANNA CINKAJZLOVA, MARTIN MATOULEK, cells (INS-1) export both miRNAs and tDRs to HDL in vitro and this process STEPAN SVACINA, MARTIN HALUZIK, Prague, Czech Republic is altered in hyperglycemic conditions. In addition, tDRs, namely tRNA- CD163 is a novel marker of macrophage activation suggested to link low- derived halves (tRHs) were found to be signifi cantly altered (4 down, 9 up) on grade infl ammation to obesity and type 2 diabetes mellitus (T2DM). The HDL from Zucker Diabetic Fatty (ZDF) rats compared to lean controls using aim of our study was to assess the infl uence of selected weight-reducing smRNA sequencing. tRH4128, a tRH originating from the 5’ half of tRNA4128- interventions on serum levels and adipose tissue mRNA expression of CD163 HisGTG, was identifi ed to be signifi cantly reduced in both HDL and islets in subjects with obesity and T2DM. (-3.69-fold) from ZDF rats, yet signifi cantly increased 2.75-fold in ZDF livers. Systemic concentrations of soluble CD163 (sCD163) and its mRNA Strikingly, 104 of the 224 tDRs found to be signifi cantly reduced in the ZDF expression in subcutaneous adipose tissue (SCAT) were measured in 22 islets were found to be signifi cantly increased in ZDF rat livers. Colesevelam, subjects with T2DM and obesity (T2DM group), 22 obese patients without a bile acid sequestrant, was found to correct (increase in islets, decrease in T2DM (OB group) and 15 healthy lean subjects (C group) at baseline, after 2 livers) many of the observed changes to tDRs in both tissues. In addition, weeks of very low calorie diet (VLCD, 15 T2DM and 15 OB subjects) and 6 angiogenin, an enzyme reported to cleave parent tRNAs into tRHs, was months after gastric plication (7 T2DM subjects). found to be decreased in plasma from T2D subjects suggesting a possible At baseline sCD163 levels were signifi cantly increased in T2DM group as mechanism for tDR biogenesis in diabetic pathology. Collectively, our data compared to OB and C subjects (275±26 vs. 197±29 vs. 155±17 ng/ml, p<0.05) suggest that tDRs in plasma, livers, and pancreatic islets are highly sensitive and correlated positively with markers of low-grade infl ammation. mRNA to hyperglycemia and that HDL-tDR transfer may represent a novel form of expression of CD163 in SCAT of T2DM subjects was higher compared to C endocrine-like communication originating from pancreatic beta cells that is group, while no signifi cant difference was present between T2DM and OB Integrated altered in T2D. Although the functional impact of these novel RNAs are not group. Both VLCD and gastric plication markedly reduced body weight and POSTERS currently understood, they have great potential as diabetes biomarkers and improved metabolic and infl ammatory parameters. sCD163 was signifi cantly likely regulate gene expression and phenotypes in ways that are both similar decreased in both T2DM and OB subjects after VLCD (308±31 vs. 191±20 Physiology/Obesity and distinct from other small RNAs, including miRNAs. ng/ml, p<0.001 for T2DM and 230±39 vs. 188±35 ng/ml, p<0.01 for OB group), while no effect on mRNA expression in SCAT was observed in either group. Gastric plication tended to decrease sCD163 (178±35 vs. 107±7 ng/ ml, p=0.147 at month 6 after plication) and reduced the number of CD163+ macrophages in stromal vascular fraction of SCAT (p<0.05). We conclude that serum concentrations and SCAT mRNA expression of the macrophage marker CD163 are elevated in subjects with T2DM and obesity. The decrease of sCD163 might contribute to favourable metabolic effects of VLCD and gastric plication. Supported By: IGANT13299-4, SVV264503, RVO-VFN64165

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A493 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1924-P live in normal health for over 18 month with no immunosuppression nor Effects of Niemann-Pick C1-like 1 on Glycemic Control by Modu- additional treatment. BAT transplants in diabetic mouse models result in lating Glucagon-like Peptide-1 decreased infl ammatory cytokines, robust replenishment of white adipose CHEOL-YOUNG PARK, EUGENE CHANG, LISA KIM, JUNG-MOOK CHOI, SUNG tissue (WAT), followed by euglycemia and reversal of diabetes. These WOO PARK, Seoul, Republic of Korea effects are independent of insulin, and require the animal’s fat and muscle Many clinical and animal studies show that the NPC1L1 inhibitor, ezetimibe, tissue to remain in a healthy un-infl amed state. We have now expanded ameliorates several metabolic disorders including obesity, diabetes, and this work in two ways. We have shown that embryonic BAT transplants heart disease. In our previous study, we found that ezetimibe treatment can reverse T1D in a spontaneous model of the disease (NOD mouse, which improves glycemic control and pancreatic beta cell mass by the increase mimics many features of human type 1 diabetes). Of NOD mice that survive of GLP-1 (glucagon like peptide-1), an incretin hormone having antidiabetic the BAT transplant surgery, over 75% achieve euglycemia within 1-2 months, properties. The aim of this study was to determine how NPC1L1 regulates and remain euglycemic for over 6 months (studies on-going). We have also intestinal GLP-1 secretion and improves glycemic control. Male KK/H1J mice utilized tissue cultured BAT cells (Cell 150:366 (2012)) to achieve a similar were fed either normal chow or high fat (HF) diet (45% fat) supplemented effect, albeit with <50% success rate. Towards elucidating the mechanism(s) with or without ezetimibe (10 mg/kg day). 6 wk ezetimibe attenuated HF- underlying this effect, we have measured the levels of many hormones and reduced insulin sensitivity during insulin tolerance test. Serum and intestinal adipokines beginning within the fi rst week after transplant. Early changes GLP-1 levels were signifi cantly higher in ezetimibe supplemented HF diet include increases in adiponectin, IGF-1, and leptin along with a decrease group than HF group. The direct effects of ezetimibe on GLP-1 secretion and in TNFα, Il-6, and plasma glucagon levels. After six months, TNFα, Il-6, L cell secretory mechanism were examined in human NCI-H716 intestinal leptin, and glucagon levels return to near their normal non-diabetic levels, cells. Ezetimibe signifi cantly increased active GLP-1 concentration, con- while IGF-1 and adiponectin remain elevated signifi cantly over normal. This comitant with the activation of ERK1/2 phosphorylation. Incubation of suggests a model where the initial effects leading to euglycemia differ from ERK inhibitor (PD98059) in NCI-716 cells prevented ezetimibe-induced the long-term maintenance of glucose homeostasis. GLP-1 secretion. Moreover, any concentration of ezetimibe did not inhibit Supported By: Iacocca Family Foundation dipeptidyl peptidase-4 (DPP-4) activity, indicating ezetimibe exerts direct DPP-4 independent effects on intestinal GLP-1 secretion. These fi ndings 1927-P indicate, for the fi rst time, NPC1L1 inhibition enhances intestinal GLP-1 Increasing GABAa/AMPA Receptor Ratio Stimulation at the secretion by ERK activation and a novel biological function of ezetimibe in Supramammillary Nucleus (sum) Induces Insulin Resistance (IR) glycemic control. without Weight Gain in Rats YAHONG ZHANG, SHUQIN LUO, MICHAEL EZROKHI, YELENA TRUBITSYNA, 1925-P ANTHONY H. CINCOTTA, Tiverton, RI Novel Glucagon-like Peptide-1(9-37) Fusion Proteins Suppress High fat feeding that induces IR is coupled to loss of the daily circadian Hepatic Gluconeogenesis in Zebrafi sh Larva and Mice peak in dopaminergic (DA) activity at the area of the biological clock, XIAOMING LI, WANPING SUN, YOUDONG WANG, JOEY ZHOU, JIN XING, XIAO- suprachiasmatic nuclei (SCN). Furthermore, loss of this circadian peak by DA YAN WEN, QINGHUA WANG, Toronto, ON, Canada neurotoxin lesion at the SCN induces IR in insulin sensitive animals. A major Glucagon-like peptide 1 (GLP-1)(7-37, or 7-36amide) is an insulinotrophic neuronal origin of this circadian DA activity at the SCN area is the SuM. We hormone and is rapidly metabolized by dipeptidyl peptidase-IV to form GLP- therefore investigated the possibility that the SuM may be a focal center 1(9-37) which is a primary GLP-1 metabolite in the body. Recent evidence within a hypothalamic circuitry to modulate peripheral glucose metabolism suggested that GLP-1(9-37) previously considered inactive has insulinomimetic in response to its afferent communication that a) inhibits its clock directed effects in a subject. This is supported by clinical studies demonstrating DA activity output and b) potentially also modulates its communication that administration of GLP-1(9-37) in human reduced postprandial glycemia with other metabolic homeostatic centers. High fat diet resistant (HFDr) independent of insulin secretion and insulin responsiveness. We hypothesized rats that were selected based on their resistance to weight gain response that the extrapancreatic glucose lowering effects of GLP-1(9-37) is achieved to HFD and rats on regular chow (RC) of similar age were studied. RC and through the suppression of hepatic gluconeogenesis. We constructed the HFDr rats were each randomized to direct SuM inhibition (SuMi) with either GLP-1 fusion proteins by the fusion of GLP-1(9-37 or 7-37) to a human IgG2 vehicle or a GABAa receptor agonist/AMPA receptor antagonist cocktail Fc segment (9-37/Fc, 7-37/Fc). Micro-injection of fl uorescein-labelled fusion (muscimol/CNQX : 3.6/1.8 pmol/day, respectively via osmotic pump) to proteins in zebrafi sh larva revealed that 7-37/Fc were concentrated in the silence supramammillary DA activity for 3 weeks. Subsequently, all animals pancreas and islet, whereas, 9-37/Fc were mainly localized in the liver. This were subjected to a glucose tolerance test (GTT) and HFDr animals were suggests that GLP-1(9-37) has a distinct pharmacokinetic profi le as that of also subjected to a 180 minute pyruvate tolerance test (PTT). In both RC GLP-1(7-37). Real-Time qRT-PCR was conducted to quantify the transcript fed and HFD fed rats, SuMi increased the GTT glucose area under curve expression of PEPCK, the enzyme of gluconeogenesis, in the zebrafi sh (AUC) (by 11%, P=NS and 28%, P<0.05), insulin AUC (by 122% and 150%, injected with the fusion peptides and glucagon (control). The results showed P<0.05) and decreased Belfi ore insulin sensitivity index (by 46%, P=0.03 and that 9-37/Fc drastically and signifi cantly suppressed PEPCK expression in 69%, P=0.002) relative to vehicle groups. SuMi also increased the plasma a time-dependent fashion, while glucagon enhanced and 7-37/Fc reduced glucose level by 34% during the last 90 minutes of the PTT (P=0.01). This these gene expression to certain levels (9-37/Fc vs. Glucagon p<0.001 n=5; induction of IR was not accompanied by an increase in body weight or fat. 9-37/Fc vs. 7-37/Fc p<0.01, n=5). The qRT-PCR was also performed in total These data suggest that the SuM, a hypothalamic site heretofore unknown RNA extracted from the liver tissues in CD1 mice injected with equal mole as a regulator of peripheral metabolism, is in fact so via regulation of clock GLP-1 fusion proteins and glucagon. We found that 9-37/Fc signifi cantly directed dopaminergic signaling and other to be determined pathways. suppressed hepatic PEPCK expression in mice (9-37/Fc vs. Glucagon p<0.001 n=5; 9-37/Fc vs. 7-37/Fc p<0.05, n=5). These fi ndings suggest that the 1928-P glucose lowering effects of GLP-1(9-37) may be achieved at least in part via Angiopoietin-like 4 Fibrinogen-like Domain as an Antiobesity and suppressing gluconeogenesis in the liver. Antidiabetic Agent Supported By: CDA ALLISON MCQUEEN, DEEPTHI KANAMALURU, NORA GRAY, JEN-CHYWAN Integrated POSTERS WANG, Berkeley, CA Angiopoietin-like 4 (Angptl4) is a secreted protein from adipose tissue

Physiology/Obesity 1926-P Reversal of Type 1 Diabetes Phenotypes by Brown Adipose Tissue and liver that acts in an endocrine, paracrine and/or autocrine fashion. Transplant Angptl4 exists either in full-length, or truncated forms, as N-terminal coiled- DAVID W. PISTON, SUBHADRA C. GUNAWARDANA, TROY J. HUTCHENS, Nash- coil domain (CCD) or C-terminal fi brinogen-like domain (FLD) in plasma or ville, TN tissues. Extensive studies have been focused on the lipoprotein lipase (LPL) Type 1 diabetes (T1D) is a serious disease affecting over 2 million inhibitory function of CCD. In contrast, we recently discovered that FLD Americans, with fatal consequences if untreated. Current therapies, involving increases intracellular cAMP levels to promote lipolysis in adipocytes. We direct insulin replacement or islet/pancreas transplantation, have numerous overexpressed FLD, full length ANGPTL4 or LacZ (control) with adenoviral limitations and complications. The ultimate goal in treating T1D is to restore delivery system in mice fed with high fat diet. Overexpressing FLD or glucose homeostasis. We previously demonstrated that euglycemia in mice ANGPTL4 did not affect food intake. Elevating plasma FLD levels, however, lacking insulin can be achieved, using subcutaneous embryonic brown protected mice from diet-induced obesity and glucose intolerance, and adipose tissue (BAT) transplants (Diabetes 61:674(2102)). These animals reduced triglyceride (TG) accumulation in skeletal muscle and liver. Gene

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A494 OBESITY—ANIMALCATEGORY expression analysis showed that the expression of fatty acid oxidative tissues including liver and heart despite elevated levels of ER stress markers. genes was increased in white adipose tissues, skeletal muscle and liver Our expression analysis ruled out involvement of known ER stress activated of FLD-overexpressing mice. Elevating full length ANGPTL4 levels in transcription factors including XBP1, ATF4, ATF6, C/EBPb, CREB3 and CREB3L3. plasma protected mice from diet-induced obesity but did not improve Hence, our data suggest that ER stress-induced TRIP-Br2 expression in visceral glucose tolerance, and markedly increased plasma TG levels. Notably, FLD fat is mediated by a previously unknown ER stress inducible-transcription overexpression did not affect plasma TG levels. We also overexpressed FLD regulator, which is absent in subcutaneous fat and other metabolic tissues. Taken using adenoviruses in mice fed with chow diet. These FLD-overexpressing together, our study suggests that identifi cation of the visceral fat-specifi c TRIP- mice had improved glucose and pyruvate tolerance. The expression of Br2 regulator could potentially reveal novel mechanism regulating differential gluconeogenic genes, such as phosphoenolpyruvate carboxykinase (Pepck) metabolic risks in different white fat depots in obesity. and glucose-6-phosphatase (G6Pase), was signifi cantly lower in liver of FLD- Supported By: R00DK090210 overexpressing mice than that of control mice. Overall, our results suggest that FLD is a potential anti-obesity and anti-diabetic agent that can modulate & 1931-P both lipid and glucose metabolism. Infl ammation and ER Stress Downregulate Branch Chain Amino Supported By: NIH (R01DK83591) Acid Uptake and Metabolism in Adipose Tissue JOEL S. BURRILL, ERIC K. LONG, BRIAN REILLY, YINGFENG DENG, PHILIPP E. 1929-P SCHERER, IAN M. ARMITAGE, DAVID A. BERNLOHR, Minneapolis, MN, Dallas, TX WITHDRAWN Infl ammation and ER stress play critical roles in the pathology of obesity- linked insulin resistance and type-2 diabetes and are linked to increased reactive oxygen species (ROS). Elevated ROS levels lead to potentiated lipid peroxidation and protein carbonylation of mitochondrial proteins including those of the branch chain amino acid (BCAA) metabolism pathway. To address BCAA metabolism regulation in adipocyte biology we utilized high fat fed mice, evaluated enzyme expression and identifi ed coordinate down regulation of BCAA metabolism genes in visceral but not subcutaneous adipose tissue or other tissues. Furthermore, TNFα reduced BCAA pathway gene expression, reduced [3H]-leucine internalization and conversion to triglycerides and metabolomic analysis revealed intracellular accumulation of BCAA in 3T3-L1 adipocytes. Surprisingly, induction of ER stress using either pharmacologic treatment of 3T3-L1 adipocytes or genetic mouse models similarly down-regulated genes linked to BCAA metabolism. Consistent with an adipose-derived source of serum BCAA, high fat fed Fabp4 null mice that exhibit reduced adipose tissue infl ammation, exhibit decreased serum leucine levels. These results demonstrate that infl ammation and ER stress attenuate lipogenesis in visceral adipose depots in part by down regulating the BCAA pathway and suggest that the accumulation of BCAA in the serum as a biomarker of the obese insulin-resistant state is the result of adipocyte mitochondrial dysfunction. Supported By: NIH (R01DK084669, P30DK050456)

& 1932-P Lycopene Attenuates the Infl ammatory Adipose Tissue Response to High Fat Feeding by Regulating Both Macrophage Recruitment and M1/M2 Status NAOTO NAGATA, YINHUA NI, MAYUMI NAGASHIMADA, LIANG XU, FEN ZHUGE, SHUICHI KANEKO, TSUGUHITO OTA, Kanazawa, Japan Adipose tissue macrophages (ATM) recruitment and polarization are considered pivotal in the development of insulin resistance. However, OBESITY—ANIMAL promising treatment modalities targeting ATM remain limited. In the present study, we show that lycopene, an antioxidant carotenoid compound, ameliorates adipose tissue infl ammation and whole-body insulin resistance Guided Audio Tour: Endoplasmic Reticulum Stress and Infl ammation in in high-fat diet (HFD)-induced obese (DIO) mice. C57BL/6J mice were fed the Pathogenesis of Obesity (Posters: 1930-P to 1936-P), see page 17. on a HFD or a HFD containing lycopene (HFD+LY; 12 mg/kg body weight). After 8 weeks of feeding, lycopene improved HFD-induced glucose & 1930-P intolerance, hyperinsulinemia (DIO 4.0±0.2 vs. HFD+LY 2.0±0.1 ng/ml, p < Endoplasmic Reticulum Stress Selectively Upregulates TRIP-Br2 in 0.01; fed state), and also enhanced insulin signaling assessed by IRβ and Visceral Fat, a Novel Transcriptional Co-regulator for Adiposity and Akt phosphorylation in epididymal white adipose tissue (eWAT) of DIO Energy Metabolism, During Obesity mice. HFD+LY mice had decreased macrophage infi ltration and crown-like GUIFEN QIANG, Chicago, IL structure formation in eWAT compared with DIO mice even though weight Obesity results from storage of excess nutritional calories as fat in adipose and adiposity were similar. To further assess the impact of lycopene on tissue. Excess nutritional infl ux has been shown to promote endoplasmic adipose tissue infl ammation, fl ow cytometry was performed on stromal reticulum (ER) stress leading to insulin resistance, infl ammation and metabolic vascular cells isolated from eWAT. ATMs identifi ed as CD45+CD11b+F4/80+ Integrated disorders. We have recently shown that a novel transcriptional co-regulator, cells were increased in DIO mice by 11.4-fold compared with WT mice. In POSTERS TRIP-Br2, is selectively up-regulated in visceral fat of obese humans and mice addition to reduction of total ATM content, HFD+LY mice had 35% fewer and TRIP-Br2 ablation in mice exerts a protective effect on obesity as well as CD11c+CD206- (M1) ATMs whereas 60% more CD11c-CD206+ (M2) ATMs Physiology/Obesity associated metabolic dysfunctions. Accumulation of excess fat in visceral but than DIO mice, resulting in predominance of M2 over M1 ATM population. not subcutaneous fat depot has been associated with increased metabolic However, the predominance of the Ly6C- over Ly6Chigh monocyte population risks during obesity. However, the regulation of functional heterogeneity was not observed in both peripheral blood and bone marrow of HFD+LY mice. among white fat depots is poorly understood. In this study, we observed that In parallel, lycopene (10-50 nM) suppressed LPS-induced M1 markers mRNA TRIP-Br2 in adipocytes is induced by high-fat fed-mice mouse serum and expression (TNFα, MCP-1, and RANTES) in Raw264.7 macrophage cells lipopolysaccharides (LPS)-treated macrophages conditioned media compared to whereas it augmented IL-4-induced M2 markers mRNA expression (IL-10 and control via the activation of ER stress pathway. This induction is abolished by Arg1) in a dose-dependent manner. In conclusion, lycopene causes dynamic the ER stress inhibitor (tauroursodeoxycholic acid, TUDCA) treatment. Further, M2 dominant phenotypic shift of ATM and thereby attenuates obesity- in vivo induction of ER stress in mice using tunicamycin selectively up-regulate induced infl ammation and insulin resistance. TRIP-Br2 in visceral but not subcutaneous fat depots as well as other metabolic Supported By: Ministry of Education, Culture, Sports, Science and Technology

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A495 OBESITY—ANIMALCATEGORY

& 1933-P & 1935-P Selective Inactivation of c-Jun NH2-Terminal Kinase in Visceral Perinatal Nutritional Programming of Adipose Tissue Infl ammation Fat Alleviates Atherosclerosis in Apo-E Defi cient Mice with Diet- and Metabolic Dysfunction in Diet-induced Obesity induced Obesity BRANDON D. KAYSER, SEBASTIEN G. BOURET, MICHAEL I. GORAN, Los Angeles, KELVIN H.M. KWOK, FRANCOIS Y.L. LI, YUK C. CHAN, RUBY L.C. HOO, DEWEI YE, CA AIMIN XU, KAREN S.L. LAM, Hong Kong, China Obesity causes adipose tissue (AT) infl ammation and metabolic diseases It has been increasingly recognised that infl ammation plays an important in some but not all individuals. We tested whether perinatal overnutrition role in the development and progression of atherosclerotic diseases. In programs vulnerability to obesity-induced infl ammation and metabolic obesity, c-Jun N-terminal kinase (JNK) mediates the infl ammation and dysfunction. Litters of C57BL/6J mice were reduced to 3 pups (SL) or aberrant production of pro-infl ammatory adipokines in adipose tissues. maintained at 7 pups (NL) from postnatal day 4 until weaning. Male NL and Our aim is to investigate the role of adipose tissue infl ammation in obesity- SL mice were then fed normal chow (NC) or 60% high fat diet (HF) from 5 related atherosclerosis and whether inactivation of JNK in visceral fat can to 17 weeks. Liver and fasting serum were collected and AT harvested for protect against atherosclerosis in diet-induced obese mice. weight, histology, and fl ow cytometry. Data were analyzed by 2x2 mixed Transgenic mice with adipose-specifi c expression of a dominant negative effects ANOVA to test for diet by litter-size interactions and are reported (dn) JNK transgene were crossbred with ApoE-/- mice to generate an as mean±SE with interaction effects and post-hoc comparisons considered ApoE-/-/dnJNK (ADJ) mouse strain . Both ADJ and ApoE-/- mice were fed signifi cant at p<0.05. Compared to NL, SL mice grew 10% heavier on NC with a high fat high cholesterol diet (0.21% cholesterol) for 20 weeks and and 18% heavier on HF. HF caused at least a 5-fold increase in epididymal, examined for the development of atherosclerosis. En face Oil Red O staining inguinal, and retroperitoneal depot weights (main effect) with SL rearing revealed a marked reduction in atherosclerotic lesion area in ADJ mice further increasing only inguinal weight by 33%. Findings for adipocyte areas compared to ApoE-/- mice. Macrophage infi ltration, as indicated by F4/80 paralleled those of fat mass. Yet despite comparable adiposity, AT from staining, was signifi cantly reduced in adipose tissue from ADJ mice when SL-HF had more crown-like structures per 3 fi elds than NL-HF (epididymal: compared to that from ApoE-/- mice. Expression levels of mRNA for pro- 14.1±3.2 vs. 2.3±2.2; inguinal: 2.2±1.0 vs. 0.3±0.7; retroperitoneal: 23.7±5.2 infl ammatory adipokines including tumour necrosis factor α, monocyte vs. 9.6±3.7) and epididymal fat from SL had more total macrophages chemotactic protein-1, F4/80 and CD68 were signifi cantly lower in adipose (393±78 x103 vs. 195 x103±32cells/g) and M1 macrophages (249x103±58 tissue from ADJ than from ApoE-/- mice. Transplantation of visceral fat vs. 66x103±41cells/g). Furthermore, SL-HF relative to NL-HF had greater from ADJ mice, but not from ApoE-/- mice, alleviated the progression of diet-induced hepatic steatosis (173±23 vs. 110±16mgTG/g), hyperglycemia atherosclerosis in recipient ApoE-/- mice, as evidenced by reduced lesion (195±17 vs. 140±10 mg/dl) and hyperinsulinemia (1.02±0.3 vs. 0.28±0.01ng/ area on the aortic tree, despite similar body weight, serum glucose levels ml), and worse glucose and insulin intolerances. In conclusion, SL rearing/ and glucose tolerance in the two types of recipient mice. perinatal overnutrition amplifi es adipose tissue infl ammation and metabolic Alleviation of atherosclerosis in ADJ mice could be attributed to the impairment with high fat feeding, and this effect is independent of adiposity. reduced production and release of pro-infl ammatory adipokines from adipose Perinatal nutrition may therefore be a critical determinant of diabetes risk in tissues, especially from visceral fat depots. Adipose-specifi c inhibition of obese adults by directly programming the infl ammatory capacity of adipose JNK may represent a potential therapeutic strategy for atherosclerotic tissue. diseases in obesity. Supported By: Hong Kong Research Grants Council (GRF769410) & 1936-P TLR4 Expression in Bone Marrow-derived Cells Modulate Insulin & 1934-P Resistance and Infl ammation in Diet-induced Obesity Meta-analysis of Genome-Wide Gene Expression Data Identifi es DANIELA S. RAZOLLI, JULIANA C. MORAES, JOSEANE MORARI, RODRIGO F. the Receptor CD44 as a Therapeutic Target for Type 2 Diabetes MOURA, MARCO A. VINOLO, LICIO A. VELLOSO, Campinas, Brazil KEIICHI KODAMA, KYOKO TODA, JUNICHIRO IRIE, SATORU YAMADA, ATUL J. Saturated fatty acids induce the activation of TLR4-dependent innate BUTTE, Stanford, CA, Tokyo, Japan immune response. It is currently unknown if the TLR4-dependent impairment Type 2 diabetes (T2D) is characterized by obesity-induced insulin of insulin signaling is due to the activation of TLR4 receptors in immune cells resistance, and growing evidence has indicated that the causative link and/or in insulin-sensitive tissues. The aim of the present study was to between obesity and insulin resistance is associated with visceral adipose investigate if a defective TLR4 signaling in bone-marrow (BM) derived cells is tissue infl ammation and liver steatosis. To fi nd causative genes implicated in suffi cient to modulate the whole-body insulin action. TLR4-mutant (TLR4-/-) the pathogenesis of insulin resistance, we performed a meta-analysis of 130 and wild-type (WT) mice were irradiated in a cobalt 60 source and submitted independent microarray experiments (totally 1,175 T2D case-control subjects), to bone marrow transplantation (BMT). TLR4-/- received BM from WT and and found that the immune-cell receptor CD44 is highly and frequently up- vice-versa. After recovery, mice were fed on a high fat diet (HFD) or standard regulated in liver and adipose tissue in T2D subjects. We demonstrated that chow for 8 weeks. We evaluated body mass gain, food intake, energy CD44 defi ciency ameliorates insulin resistance, adipose tissue infl ammation expenditure and thermogenesis markers in brown adipose tissue (BAT). and hepatic steatosis in diet-induced obese mice. We also found, in humans, Insulin sensitivity was analyzed by GTT, ITT and AKT phosphorylation in liver CD44-positive infl ammatory cells are infi ltrated into obese adipose tissue. and adipose tissue. In addition, markers of infl ammation were evaluated In addition, to assess if this receptor can be useful as a therapeutic target in both tissues by real-time PCR. WT submitted to BMT from TLR4-/- for T2D, we performed daily injections of anti-CD44 monoclonal antibody donors were protected from HFD-induced obesity. In TLR4-/- submitted (CD44 mAb) in a high-fat diet mouse model. Four weeks of therapy with to BMT from WT donors this effect was blunted and mice gained weight. CD44 mAb suppressed visceral adipose tissue infl ammation compared to Energy expenditure was higher in TLR4-/- compared to WT mice, which was controls and reduced fasting blood glucose levels, body weight gain, liver accompanied by increased UCP1 mRNA expression in brown adipose tissue. steatosis, and insulin resistance to levels comparable with or better than TLR4-/- mice fed on HFD and WT submitted to BMT from TLR4-/- donors other common anti-insulin resistance drugs (metformin and pioglitazone). presented increased IL10 mRNA expression and reduced IL6, TNFα and IL1β These fi ndings suggest that CD44 mAb may be useful as a prototype drug mRNA levels in liver and adipose tissue. The opposite occurred when WT

Integrated for fundamental therapy of T2D by breaking the links between obesity and and TLR4-/- mice were submitted to BMT from WT donors. TLR4-/- mice POSTERS insulin resistance. showed improved tolerance to glucose, insulin sensitivity and increased AKT

Physiology/Obesity phosphorylation in liver and adipose tissue. Thus, we show that the absence of a functional TLR4 in bone marrow derived cells is suffi cient to protect mice from diet-induced insulin resistance and defective regulation of whole body energy homeostasis. Supported By: FAPESP

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A496 OBESITY—ANIMALCATEGORY

Guided Audio Tour: Role of the Gut in Metabolic Regulation (Posters: 1937-P to 1943-P), see page 17. & 1939-P Antibiotic Treatment Alters the Gut Microbiome, Adipose Tissue & 1937-P Infl ammation, and Energy Metabolism SHIHO FUJISAKA, SIEGFRIED USSAR, C. RONALD KAHN, Boston, MA Gut Microbiota Confer a Protective Role of Fibersol-2 Against Meta- We have previously shown that different strains of mice and mice from bolic Syndrome different vendors exhibit different rates of obesity following high fat diet KAZUHIRO SHIMOMURA, ZHENG CHEN, Wilmette, IL, Houston, TX (HFD) challenge. C57BL/6J (B6J) mice from Jax Labs are obesity prone, Dietary fi bers are well recognized as a benefi cial nutrient component; while 129 mice from Jax (129J) are obesity resistant. 129 mice from Taconic however, average daily fi ber intake in the U.S. is markedly lower than the (129T), on the other hand, are similar to B6J mice and develop HFD-induced recommended amount. In-depth research on their preventive/therapeutic obesity. This difference between strains of 129 mice correlates with effi cacy and cognate mechanisms will serve to raise awareness and promote differences in the gut microbiome. In the present study, we have treated fi ber intake for better health. Fibersol-2 is a soluble, digestion-resistant these 3 strains with either vancomycin, which kills gram positive bacteria, maltodextrin widely incorporated and consumed in various food products. In or metronidazole, which kills anaerobic bacteria, and challenged them the current study, Fibersol-2 intake was found to mitigate body weight gain with HFD to further explore the role of the microbiome in obesity. Both in both diet-induced obesity (DIO) and type 2 diabetic db/db mouse models; antibiotics greatly decreased the number of gut bacteria and changed their concomitantly, glucose tolerance tests revealed improved glucose tolerance composition. Vancomycin treatment resulted in increased lean mass in both and insulin sensitivity in Fibersol-2 treated mice. Real-time qPCR analysis of B6J and 129T mice with no change in food intake or activity. In B6J mice, white adipose tissues showed diminished expression of pro-infl ammatory this was associated with a signifi cant decrease in basal metabolic rate as cytokine genes including TNF-α and IL-6 from Fibersol-2 treated mice, and assessed in a CLAMS apparatus. Vancomycin and metronidazole treatment H&E staining also found reduced numbers of crown-like structures, indicating also signifi cantly improved glucose metabolism, decreased serum TNF attenuated adipose tissue infi ltration of macrophages. Consistent with a α levels, decreased infl ammatory markers in both adipose tissue and colon prebiotic function of Fibersol-2, 16S rRNA sequencing of gut microbiome and improved insulin signaling in adipose tissue of B6J mice. On the other revealed phylum- and genus-level changes in Fibersol-2 treated db/db mice hand, in 129J mice, both antibiotics increased infl ammatory markers in compared with control db/db mice. Remarkably, transplantation of microbiota adipose tissue and colon and impaired insulin sensitivity. Thus, antibiotic from Fibersol-2 treated mice to untreated mice resulted in pronounced and treatment modifi es the gut microbiome, and this impacts on energy balance progressive improvement in glucose tolerance and insulin sensitivity. In and infl ammation induced by HFD. These effects are strain dependent, agreement with previous results from T2D patients, 16S sequencing of gut indicating an important interaction of the gut microbiome with host in microbiome from recipient mice elucidated a positive correlation between determining metabolic state. abundance of Firmicutes and improved insulin response. In particular, Supported By: Sunstar Foundation Lactobacillus and Ruminococus were enriched in Fibersol-2 recipient mice relative to control recipient mice. Taken together, these studies establish a robust protective role of Fibersol-2 against metabolic syndrome, and indicate & 1940-P a predominant contribution from its prebiotic action on gut microbiota. Roux-en-Y Gastric Bypass in a Severely Type 2 Diabetic Rodent Supported By: Matsutani America, Inc. Model RAYMOND G. LAU, LOUIS RAGOLIA, COLLIN BRATHWAITE, DREW RIDEOUT, & 1938-P MICHAEL S. RADIN, KENETH HALL, Mineola, NY, St. Petersburg, FL Background: Roux-en-Y gastric bypass (RYGB) is a weight loss surgery Gut Microbial Metabolomic Predictors of Dietary-induced Obesity that causes weight-independent improvement of type 2 diabetes (T2DM). and Diabetes Growing evidence also supports that RYGB has benefi cial effects on MARC E. DUMAS, LESLEY HOYLES, JULIEN CHILLOUX, SOPHIE CALDERARI, cardiovascular dysfunction, although the mechanism is unknown. Over ALICE ROTHWELL, CLAIRE L. BOULANGE, QUAN GU, CHRISTOPHE HUE, JANE the last several years, RYGB studies in rodents have attempted to better F. FEARNSIDE, JAMES SCOTT, JEREMY K. NICHOLSON, DOMINIQUE GAUGUIER, understand the physiologic changes that occur after the surgery. To date, London, United Kingdom, Paris, France, Randwick, Australia, Oxford, United few studies have performed RYGB in the Zucker Diabetic Fatty (ZDF) rodent, Kingdom a model of diabetes and obesity. Hypothesis: We hypothesized that RYGB Phenotype heterogeneity is classically infl uenced by genetic, epigenetic will improve cardiometabolic parameters, even in a severely diabetic rodent and environmental factors but the gut microbiome is also thought to model. Methods: Five ZDF rodents underwent RYGB and were compared to contribute to this phenomenon. Here we studied a population of 50 isogenic four pair-fed ZDF rodents that underwent sham surgery. Fasting glucose, C57BL/6J mice developing a range of dietary-induced diabetes, obesity and insulin, glucagon-like peptide-1, leptin, and blood pressure were measured anxiety sub-phenotypes after 20 days of high-fat feeding. We then profi led prior to surgery, and then four and 14 weeks post surgery. Tissue samples the urinary metabolome of these mice by untargeted 1H Nuclear Magnetic of aorta, pancreas, liver, and kidney were obtained at euthanasia. Results: Resonance spectroscopy before high-fat-diet feeding and sequentially Rodents that underwent RYGB lost 20% of their body weight by 2 weeks. afterwards. Multivariate statistical models were constructed to predict Fasting glucose levels between RYGB and pair-fed at four weeks was the phenotype heterogeneity and disease sub-phenotypes (lean non diabetics, same (202 +/- 14.7 mg/dL, 194 +/- 19 mg/dL). However, by 14 weeks RYGB lean diabetics, obese diabetics) from using baseline metabolic profi les. rodents fasting glucose was 332.6 +/- 45 g/dL as compared with the pair- Among the markers, we found that excretion of gut microbial detoxifi cation fed >500 mg/dL. Fasting total cholesterol in RYGB was lower (140.6 +/- 16 metabolites such as phenylacetylglycine (PAG), hippurate, dimethylamine mg/dL) as compared to pair-fed group at 14 weeks (282.5 +/- 27 mg/dL). and trimethylamine-N-oxide (TMAO) was strongly predictive of obesity and Systolic and diastolic blood pressure increased in the pair-fed group by disease sub-phenotypes, insulin resistance, body/organ weights as well as nearly 30% post-operatively. For the RYGB group, the systolic and diastolic anxiety/activity outcomes. Genome-wide adipose tissue gene expression blood pressures were unchanged as compared to the pre-operative values. profi ling tissue suggests that TMAO excretion correlates with reduced Aortic wall thickness in RYGB was 9% less as compared to the pair-fed lipogenesis and insulin action. We then set up cellular investigations group. Conclusions: RYGB attenuates the progression of T2DM in a weight- to further document the causative role of these microbial metabolites.

independent manner in a severely diabetic rodent model. The surgery also Integrated For instance, we tested whether TMAO, PAG, hippurate affect cellular POSTERS has a weight-independent effect on hyperlipidemia and cardiovascular phenotypes associated with obesity and diabetes, through inhibition parameters. Physiology/Obesity of in vitro cell differentiation, glucose uptake and lipid accumulation in adipocytes. This work describes new mechanistic detail that underpins the role of the microbiome in cardiovascular disease and also suggests future possibilities for disease prognosis based on microbial signatures and new therapeutic interventional approaches. Supported By: MetaCardis (HEALTH-F4-2012-305312)

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A497 OBESITY—ANIMALCATEGORY

& 1941-P & 1943-P Characterization of Hypothalamic Gene Expression in a Rat Model GLP1 Decreases Adiposity and Food Intake Partially via Improvement of Roux-en-Y Gastric Bypass of Insulin and Leptin Action/Signaling in the Hypothalamus and PERNILLE BARKHOLT, PHILIP J. PEDERSEN, SARAH PAULSEN, ANDERS HAY- Amygdala of Dio Mice SCHMIDT, NIELS VRANG, JACOB JELSING, HENRIK B. HANSEN, Hørsholm, GIOVANNA T. RICARDO, LAIS WEISSMANN, PAULA GABRIELE FERNANDES Denmark, Copenhagen, Denmark QUARESMA, THAYANA MICHELETTI, GISELE CASTRO, PATRICIA O. PRADA, Roux-en-Y gastric bypass (RYGB) surgery promotes robust weight loss and Campinas, Brazil resolution of type II diabetes. However, relatively little is known about how Glucagon-like-peptide-1 (GLP1) and its analogs such as liraglutide are the metabolic effects of RYGB involve central signaling pathways. Here, used to treat type 2 diabetes (T2D) and are also critical regulators of energy we aimed at identifying adaptations in the hypothalamic gene expression balance. GLP1 receptors are expressed in the hypothalamus and amygdala pattern of neuropeptides known to be involved in the homeostatic regulation that is part of mesolimbic system which contribute to control food intake and of energy balance in an RYGB model in rats. the hedonic value of the food. However, it is not known whether liraglutide 18 lean male Sprague-Dawley rats were included in this study; nine treatment may improve leptin and insulin signaling in the hypothalamus and underwent RYGB surgery and nine underwent sham surgery. Body weight amygdala. Thus, the aims of the present study are: 1) to investigate whether was monitored daily and the study was terminated at day 60. Semi- liraglutide treatment alters adiposity, insulin sensitivity and 2) insulin and quantitative in situ hybridization using 33p-labelled probes against AgRP, leptin action/signaling in the hypothalamus and amygdala, in vivo, of diet- NPY and CART mRNA was applied to systematic uniform randomly sampled induced obesity (DIO) mice. DIO mice were treated for 7 days with liraglutide sections of the hypothalamus. intraperitoneal (200 mg/kg/day) or vehicle. Liraglutide treatment decreased RYGB led to a sustained and statistically signifi cant 35% weight loss adiposity, fasting glycemia and improved insulin sensitivity. Decreased food when compared with sham-operated controls. Furthermore, the expression intake and increased IR, AKT and FoxO1 phosphorylation were observed of the orexigenic peptides AgRP and NPY was signifi cantly upregulated in in response to intracerebroventricular (ICV) insulin in the hypothalamus RYGB operated rats (by 83% and 95% respectively) in the arcuate nucleus of liraglutide treated DIO mice. Also, liraglutide treatment decreased (ARC) compared with sham-operated controls, whereas no changes were food intake, increased leptin receptor, JAK2, STAT3 and decreased AMPK seen in the CART mRNA expression. phosphorylation in response to ICV leptin in the hypothalamus. Insulin injected Our data indicates that the effect of RYGB surgery on body weight loss in amygdala reduced food intake and increased IR and AKT phosphorylation cannot be explained by reduced orexigenic or increased anorexigenic signaling in response to insulin in the amygdala of DIO mice treated with liraglutide. from the AgRP/NPY and CART/POMC cell pool of the ARC, suggesting that Liraglutide and vehicle pair-fed mice had improved insulin/leptin signaling in other central mechanisms are triggered to overrule hunger-associated those brain regions, suggesting that the improvement was due to GLP1. In hypothalamic signaling upon RYGB surgery. These may tentatively involve summary, our results suggest that liraglutide treatment improves insulin and forebrain-associated dopaminergic and neuropeptidergic pathways. leptin action/signaling in the hypothalamus and amygdala of DIO mice, which may contribute to the reduction of food intake, adiposity and the hedonic & 1942-P value of the food. Furthermore, we suggest that GLP1 potentiates insulin and Duodenal Linoleic Acid Sensing Lowers Glucose Production in Rats leptin action in different brain regions that regulate energy balance. and Mice via a CCK-independent Neuronal Network Supported By: FAPESP BRITTANY A. RASMUSSEN, CLEMENCE D. COTE, MELIKA ZADEH TAHMASEBI, FRANK A. DUCA, TONY K.T. LAM, Toronto, ON, Canada 1944-P The disruption in glucose homeostasis characteristic of diabetes and Possible Involvement of Anorexigenic Protein, Nesfatin/Nucleo- obesity is due partly to an increase in glucose production (GP). However, the bindin-2 (NUCB2) in Blood Pressure Regulation by Regulating Water mechanisms underlying the regulation of GP in healthy and obese/diabetic Reabsorption in Renal Collecting Duct settings remain to be elucidated. A duodenal Intralipid infusion activates a HIROYUKI SHIMIZU, AYA OSAKI, Nasushiobara, Japan, Maebashi, Japan duodenal PKC-δ → cholecystokinin (CCK) → CCK1 receptor → PKA neuronal Nesfatin/nucleobindin-2 (NUCB2), a precursor protein of anorexigenic signaling cascade to lower GP in normal but not high-fat fed rodents. Intralipid protein, nesfatin-1, is ubiquitously expressed in the body. Since circulating is an emulsion of fatty acids containing the highest percentage of linoleic concnetrations of nesfatin/NUCB is correlated with body mass index, acid (LA; polyunsaturated fatty acid) and oleic acid (OA; monounsaturated nesfatin/NUCB2 is supposed to involve in the development of metabolic fatty acid). The relative ability of individual duodenal fatty acids to regulate syndrome. Recently, we have demonstrated an involvement of this protein GP in normal and obese conditions is currently unknown, as are the potential in the regulation of blood pressure in vivo, since peripheral administration mechanisms. Therefore, we investigated whether LA and OA regulate GP of nesfatin-1 increases blood pressure mediated by beta-adrenergic when infused into the duodenum and whether they utilize similar mechanisms mechanism in mice (Hypertens Res 2013). The present studies were as Intralipid in rats and mice in vivo. First, an intraduodenal infusion of OA undertaken to investigate a possible involvement of nesfatin/NUCB2 in the or LA lowered GP during the pancreatic (basal insulin) euglycemic clamps in regulation of blood pressure. The immunoreactivity against nesfatin/NUCB2 regular-chow fed rats and mice. Second, co-infusion of duodenal OA with was selectively found in vascular endothelial cells of aorta, pulmonary artery the CCK1 receptor inhibitor, MK-329, abolished the ability of OA to lower and renal artery, cardiac muscle and skeletal striated muscle cells, but not GP. Interestingly, co-infusion of duodenal LA with MK-329 did not block the in vascular smooth muscle cells at all. Furthermore, the immunoreactivity ability of LA to lower GP. Third, an intraduodenal OA or LA infusion failed against nesfatin/NUCB2 was selectively found in renal collecting duct cells, to lower GP when co-infused with the anesthetic tetracaine. Fourth and which contain aquaporin (AQP)-2 and/or epithelial sodium channel (ENaC). In most importantly, high fat feeding disrupted the ability of duodenal OA or medullary collecting ducts, cells expressing nesfatin/NUCB2 co-expressed LA infusion to lower GP. In summary, duodenal LA and OA sensing lower GP AQP-2, but did not co-expressed AQP-2 in renal cortical collecting ducts. via a CCK-independent and dependent neuronal network in rats and mice. On the other hand, collecting ductal cells expressing ENaC were totally Interestingly, despite different signalling mechanisms, both OA and LA fail to compatible with those expressing nesfatin/NUCB2 in both renal medullary lower GP in rodents fed a high fat diet. Thus, dissecting duodenal OA and LA and cortical collecting ducts. Thus, there is a possibility that nesfatin/

Integrated signaling will unveil dietary and/or pharmacological strategies to activate NUCB2 is involved in the development of hypertension in obese patients POSTERS duodenal signaling that lower GP in diabetes and obesity. through vasoconstriction and increase of plasma volume by increased water

Physiology/Obesity Supported By: CIHR reabsorption in the kidney. Supported By: Ministry of Education, Culture, Sports, Science and Technology

1945-P 3-D Imaging and Illustration of Perivascular Remodeling of Islet Pericytes and Parasympathetic Innervation in Obese db/db Mice SHIUE-CHENG TANG, HUNG-JEN CHIEN, SHIH-JUNG PENG, TZU-EN HUA, Hsinchu, Taiwan Obesity increases the islet β-cell mass to cope with the elevated insulin demand. Obesity also induces diabetes and hypertension which create disturbances in circulation and in turn infl uence the islet vasculature. Despite

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A498 OBESITY—ANIMALCATEGORY the expected changes of the islet microstructure, vasculature, and innervation glucagon-like peptide-1(GLP-1) receptor agonist on the differentiation and in response to the metabolic challenge, visualization of the tissue remodeling function of brown adipocytes in mice through certain pathways. Primary in an integrated fashion has been diffi cult. Here, we prepared transparent brown and white adipocytes in mice were cultivated in vitro, and were islets from lean control and obese (db/db, 8-10 weeks, diabetic) mice to further divided into four subgroups receiving: (1) GLP-1(10-8M),(2) GLP-1(10- characterize the perivascular remodeling of pericytes and parasympathetic 9M),(3) STAT3 inhibitor(5*10-5M),(4) Control. We examine specifi c genes of innervation via 3-D histology with optical clearing (Gastroenterology 137: brown adipocytes and white adipocytes. Treatment with GLP-1 receptor p453-465, 2009). Islet pericytes were identifi ed by their NG2+ cell bodies agonist increased the expression of specifi c genes(UCP1,Cidea,PGC1- and processes contacting the capillaries. Parasympathetic innervation was α),differentiation genes(PRDM16,PPAR-γ),STAT3 pathway genes(STAT3) in confi rmed by the peri-islet VAChT+ ganglion and the scattered intra-islet brown adipocytes signifi cantly compared with the control group (P<0.01), varicosities. In the db/db mice, we observed the lateral spreading of the while the effects on BAT specifi c genes were eliminated by the treatment pericyte processes in comparison with the slender, longitudinal extension with STAT3 inhibitor partly. Furthermore, GLP-1 receptor agonist attenuated of the normal processes on the capillary walls. Meanwhile, in the islet core, the expression of brown adipocytes specifi c genes in the white adipocytes. the VAChT+ varicosities were found mainly associated with the capillaries Thus, Glucagon-like peptide-1(GLP-1) receptor agonist may regulate the rather than randomly distributed in space. These new histological features differentiation and function of brown adipocytes as well as white adipocytes identify the perivascular tissue remodeling of the diseased islets in the obese in mice, thereby enhance the browning of adipose, partly through STAT3 db/db mice. activation, which provides a theoretical approach to the therapy of GLP-1 in diabetes and obesity.

1947-P Adipose Tissue (AT) Immune Responses Distinguish the Differential Insulin Resistance (IR) of Obesity Associated with Aging or Diet KANTHI BANGALORE KRISHNA, MAJA STEFANOVIC-RACIC, GABRIELE SCHOISWOHL, NIKOLAOS DEDOUSIS, IAN SIPULA, ROBERT M. O’DOHERTY, Pittsburgh, PA AT of obese humans and rodents is heavily infi ltrated with Th-1 skewed immune cells, which play an important role in the pathogenesis of IR. Aging is also associated with IR, but the role of immunophenotypic changes with aging is unknown. Here, we evaluated IR and AT immunophenotypic changes in young diet-induced obese mice (41% fat diet, YO, see Table 1), young lean mice (11% fat diet, YL), aged lean (AL) and aged obese (AO) mice (11% fat diet throughout their lives). Despite similar body weight and fat mass, YO mice were ~50% more IR than AO mice. Both AL and AO mice were somewhat more IR compared to YL (~25%), and AO were more IR than AL, based on a greater insulin response (~150%). The outstanding pathophysiological characteristic that distinguished YO from AO mice was a substantially greater (at least 200%) AT accumulation of CD11b+ and CD11c+ cell sub-populations i.e. macrophages (MAC) and dendritic cells (DC), with no differences in the lymphoid populations (data not shown). There were no immunophenotypic differences between YL and AL. Together, these data suggest that differences in the severity of IR in diet-induced and aging associated obesity are related to the degree of MAC and DC infi ltration of AT.

Supported By: Taiwan NHRI (EX102-10044EI); Taiwan NSC (102-2628-B-007-002- Integrated MY2 to S.C.T.) POSTERS Physiology/Obesity 1946-P Regulation of Brown Adipose Tissue’s Function and Differentiation by Glucagon-like Peptide-1 Receptor Agonist in Mice HUAN SHI, SHAN-MEI SHEN, DA-LONG ZHU, YAN BI, RAN MENG, YA-LI JING, Nanjing, China Besides its evident effect of lowering glucose levels, GLP-1 and its receptor agonist can reduce weight and regulate energy metabolism through multiple mechanisms, which was the similar to the function of brown adipose tissue. Recent study illustrated some potential effect of GLP-1 on brown adipose tissue. In this study, we investigate the effects of

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A499 OBESITY—ANIMALCATEGORY

1948-P 1950-P Combinations of Adenosine A2a Agonism with Inhibition of Active Antihyperglycemic Effect of Triterpene from Panax Notoginseng: Bile Acid or Glucose Uptake from the Gut Lumen Are Synergistic for Panaxatriol, but Not the Panaxatriol-type Saponin, Promotes Weight Loss in DIO Mice Glucose Uptake via PI3 Kinase-akt Signaling Pathway in Skeletal KAMAL A. AL-BARAZANJI, STEVE M. SPARKS, WILLIAM G. BENSON, THOMAS Muscle Cells J. RIMELE, ANDREW YOUNG, LIHONG Z. CHEN, Research Triangle Park, NC, TAKU IWAMOTO, KUMIKO KITAMURA, YUSUKE TAKAMURA, MITSURU NOMURA, Durham, NC TOMOJI ONO, MICHIAKI MURAKOSHI, YASUKO MANABE, NOBUHARU L. FUJII, Previously, we and others have demonstrated antidiabetic effects Odawara, Japan, Hachioji, Japan of inhibitors of sodium-coupled glucose transporter 1 (SGLT1) and the The root of Panax notoginseng has been used as an herbal treatment apical sodium-dependent bile tranporter (ASBT). We further explored the in traditional Chinese medicine for centuries, and a group of saponins effects of combining GR163819, a potent agonist for the G-protein coupled (ginsenosides) is considered as the major active ingredients. Since ginseno- adenosine A2a receptor (A2aR) with GSK2594928, an SGLT1 inhibitor or with sides are converted into dammarane-type triterpenes as aglycones during the GSK2331583, an ASBT inhibitor, in a diet-induced obese (DIO) mouse model. digestion process, we produced a Panax notoginseng root extract containing Relative to vehicle controls, 15-day treatment with GR163819 (30 mg/kg, bid) high concentrations of dammarane-type triterpenes (DTE) in which all caused 5.2%±1.1 (p<0.05) weight loss, while GSK2594298 (-0.4%±0.8) and sugars were fully removed from ginsenosides. Previously we reported that GSK2331583 (0%±1) singly caused no weight loss. Combination of GR163819 DTE has hyperglycemia inhibitory effect in mouse obese/diabetic models with GSK2594298 and GSK2331583 caused 9.1%±1.1 and 10.8%±1.7 weight and pre-diabetic people (ADA, 109-LB, 2011 and, 1140-P, 2013). DTE had loss, respectively. The weight loss was predominantly from fat. Cumulative a greater anti-hyperglycemic effect than Panax notoginseng root extract food intake was reduced by GR163819 (17.1% relative to vehicle), with containing high concentrations of ginsenosides. Panaxatriol (PT), one of minimal effect from the other two agents dosed singly (1.7% increase, 2.3% the dammarane-type triterpenes in DTE, has been found to improve whole reduction, respectively). Combination of GR163819 with GSK2594298 and body insulin resistance and enhance glucose uptake in skeletal muscle (ADA, GSK2331583 caused a reduction of 25.4% and 24.9% in cumulative food 1933-P, 2012). intake (p<0.05), respectively. The enhanced effects of GR163819 combined In this study, we aimed to elucidate the molecular mechanisms of glucose with GSK2594298 or GSK2331583 were associated with increased neuronal uptake in skeletal muscle elicited by PT. We found that treatment of skeletal activation in the brainstem (AP, NTS) and hypothalamic PVN regions and muscle cells with PT signifi cantly increased glucose uptake. In contrast, increased plasma GLP-1 concentrations. In conclusion, activation of the ginsenoside Rg1, precursor of PT, did not promote glucose uptake. Treatment A2aR combined with inhibition of SGLT1 or ASBT transporters enhanced with PT signifi cantly enhanced Akt, but not AMPK, phosphorylation. The anorectic and weight loss effects in DIO mice. effect of PT on glucose uptake was completely abolished by treatment with a PI3 kinase and Akt1/2 kinase inhibitor. Treatment with PT did not 1949-P affect on the protein expression of glucose transporter GLUT1, GLUT4 and Antihyperglycemic Effect of Triterpene from Panax Notoginseng: insulin receptor IR-β. These results indicate that PT, but not ginsenoside Chronic Exercise Combined with Panaxatriol Effectively Suppresses Rg1, promotes glucose uptake via the PI3 kinase-Akt signaling pathway in Hyperglycemia and Increases Skeletal Muscle Mass in KKAy Mice skeletal muscle cells. Our results also suggest that the anti-hyperglycemic KUMIKO KITAMURA, TAKU IWAMOTO, YUSUKE TAKAMURA, MITSURU mechanism of PT intake due to the enhancement of glucose transport via the NOMURA, TOMOJI ONO, MICHIAKI MURAKOSHI, YASUKO MANABE, NOBUHARU insulin signaling pathway in skeletal muscle. L. FUJII, TOHRU FUSHIKI, Odawara, Japan, Hachioji, Japan, Kyoto, Japan Previously we reported that Panax notoginseng extract containing 1951-P dammarane-type triterpenes (DTE) elicits an anti-diabetic effect (ADA, 109- Testosterone Upregulates Mitochondrial Biogenesis in Skeletal LB, 2011) and panaxatriol (PT, one of the dammarane-type triterpene contained Muscle in DTE) improved whole body and skeletal muscle insulin sensitivity. In this YOSHIHIKO KITADA, KOICHIRO TAGUCHI, TARO USUI, HIDEYUKI OKADA, ICHIRO study, we explored the combined effect of exercise and ingestion of PT on MORI, TAKAHIDE IKEDA, KAZUO KAJITA, TADAHIRO KITAMURA, TSUTOMU SASAKI, hyperglycemia. Type2 diabetic KKAy mice were divided into four groups; TAKASHI SATO, TATSUO ISHIZUKA, Gifu, Japan, Gero, Japan, Maebashi, Japan sedentary (control group), 0.24 % PT fed (PT group), exercise (Ex group) and Androgen reduces fat mass, although the underlying mechanisms are exercise + 0.24 % PT fed (Ex + PT group). Mice assigned to the exercise group unknown. Here, we examined the effect of testosterone on heat production performed treadmill exercise 5 day/week and were made to run at a running and mitochondrial biogenesis. Testosterone-treated mice exhibited weight intensity of 15 m/min for 45 min. At the end of the exercise training, skeletal reduction and elevated heat production. Treatment with testosterone muscles were removed and weighted. Compared with control group, blood increased the expression level of PGC1α, ATP5B and Cox4 in skeletal glucose levels of Ex group were signifi cantly lower on day31 and thereafter, muscle, but not those in brown adipose tissue and liver. Expression levels of moreover, blood glucose levels of Ex + PT group were signifi cantly lower these mitochondrial proteins were reduced in skeletal muscle isolated from from day10 until end of the test period (Fig.1). Only in Ex + PT group, muscle androgen receptor defi cient mice (ARKO) showing late-onset obesity caused weight was signifi cantly increased compared to control group. Both GLUT4 by decreased energy expenditure. Mitochondrial DNA and mRNA levels of and PGC-1α mRNA expression in Ex + PT group were signifi cantly higher genes involved in mitochondrial biogenesis including PGC1α, NRF-1, NRF-2 than those in control group. Chronic exercise combined with ingestion of PT and Tfam were elevated in skelet al muscle isolated from testosterone-treated effectively suppresses hyperglycemia and increases skeletal muscle mass. mice, but those were down-regulated in ARKO. The expression levels of myoglobin and troponin I1 mRNA were increased compared with troponin C2 and troponin T3 in skeletal muscle isolated from testosterone-treated mice, which imply testosterone promotes conversion of muscle fi bers from type 2 (fast twitch) to type 1 (slow twitch). Treatment with 10 nM testosterone increased mRNA levels of genes involved mitochondrial biogenesis in C2C12 myotubes. These results demonstrated that the testosterone-induced increase in energy expenditure is derived from elevated expression level Integrated POSTERS of PGC1α and subsequent mitochondrial biogenesis in skeletal muscle. Since PGC1α is known to regulate muscle fi ber type, we speculate that Physiology/Obesity testosterone-induced increasing in expression level of PGC1α elevates type 1 fi ber inskeletal muscle.

1952-P Increased Akt Phosphorylation and Nrf2 Activation by Low Dose Radiation Was Associated with Reduction of Testicular Apoptotic Cell Death in Type 2 Diabetic Rats YUGUANG ZHAO, YANG BAI, FANGCHAO GONG, ZHIQIANG WAN, WEI LI, LU CAI, Changchun, China, Louisville, KY Infertility is a common complication in diabetic men due to the loss of germ cells. Our previous study had demonstrated that repetitive exposures

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A500 OBESITY—ANIMALCATEGORY to low dose X-ray’s radiation (LDR) can attenuate type 1 diabetes-induced also markedly increased in casein group. Decreased insulin sensitivity as testicular apoptotic cell death. To defi ne whether repetitive exposures well as increased triglyceride in liver were found in casein mice. The mRNA to LDR can also attenuate type 2 diabetes (T2DM)-induced testicular and protein expressions of CD36, a critical protein to facilitate the fatty acid apoptotic cell death, T2DM rats were established by high fat diet feeding, uptake in the liver, were signifi cantly increased in the casein group. While, followed by a small dose of streptozotocin. After diabetes onset, diabetic the mRNA expressions of genes accelerate fatty acid oxidation such as and age-matched control rats were treated with or without LDR at 25 mGy microsomal triglyceride transfer protein, carnitine palmitoyltransferase 1a, for 4 weeks every other day. Western blotting assay revealed that T2DM and acyl-coenzyme A oxidase 1, a key factor in hepatic lipid secretion were increased testicular apoptosis (i.e.: increased AIF expression), oxidative obviously decreased in the casein group. Meanwhile, chronic infl ammation and nitrosative damage (3-NT and 4-HNE), and endoplasmic reticular stress not only minished the size of adipocytes, but also down-regulated the (CHOP and caspase 12). All these effects were attenuated by repetitive expressions of PPARγ and C/EBPα, indicating an impaired adipogenesis. The exposures to LDR. Mechanistic studies showed that Akt-mediated GSK-3β conclusion is that in addition to disturbed lipid metabolism in the liver per was down-regulated, but Akt negative regulators PTP1B and TRB3 were up- se, impaired adipogenesis in the adipose tissue might also be associated regulated in T2DM group. T2DM-induced the above testicular Akt related with hepatic lipid deposition induced by chronic infl ammation in mice with changes were partially prevented by repetitive exposures to LDR. Expression chow diet. of Nrf2 and its downstream gene NQO1 was up-regulated in T2DM group, Supported By: NSFC (81170751, 81200294, 81300486, 81370954) and further up-regulated by exposures to LDR. Nrf2-related downstream anti-oxidants, such as SOD activity and catalase content, were decreased 1955-P in T2DM group, which were signifi cantly attenuated by exposures to LDR. Proteomic Analysis Reveals the Role of SIRT1 in Regulating Diverse Therefore, the attenuation of T2DM-induced testicular apoptotic cell death Substrates in Mature Adipocytes by repetitive exposures to LDR is likely mediated by up-regulation of Akt/ SUN-YEE KIM, FENG XU, Singapore, Singapore Nrf2-mediated anti-oxidative pathway. Mammalian SIRT1 deacetylates target proteins that are responsible Supported By: NSFC (81300724) for apoptosis, cancer, mitochondrial function, and metabolism. In spite of important functions for numerous cellular processes, not much is known 1953-P about the specifi c role for SIRT1 in deacetylating proteins in fat cells. Impairment of Primary Cilia Contributes to High-Fat-Diet–Induced Metabolic pathways in fat cells are unique compare to those of normal cells Visceral Adiposity WITHDRAWN because of their high lipid content. To explore a possible role of SIRT1 in NI QIU, CONG LI, WEIJIN FANG, YAN XIONG, Guangzhou, China regulating fat cell metabolism, we aimed to identify SIRT1 target proteins in Obesity, especially visceral adiposity, is thought to a major cause for mature adipocytes by a proteomic analysis. We performed two-dimensional insulin resistance and metabolic syndrome. Our previous data showed polyacrylamide gel electrophoresis in 3T3-L1 mature adipocytes upon that disruption of primary cilia related gene Kif3a in osteoblasts displayed addition of a pharmacological inhibitor or activator of SIRT1. By comparing impairment of osteogenesis and enhanced makers of adipogenic gene acetylated protein spots from mature adipocytes to those of from samples expression. Whether high fat diet-induced adiposity is due to impairment of treated with SIRT1 inhibitor or activator, we found seven spots out of 264 primary cilia in adipose tissues needs further investigation. Results showed spots which showed a dynamic change of acetylation levels. Identifi cation that the levels of primary cilia related genes Kif3a and IFT88 as well as of these proteins was assessed by mass spectrometry analysis. Five of the a-acetylated tubulin, a primary cilia special protein, were two-fold higher in candidates were found to be mitochondrial proteins that are involved in epididymal adipose tissue and stromalvascular fractions (SVF) than that in fatty acid metabolism and the electron transport system. SIRT1 is commonly subcutaneous adipose tissue and SVFs from lean mice. However, both levels known to play a role in the nucleus, but is not known whether SIRT1 is of Kif3a and IFT88 genes and a-acetylated tubulin protein were reduced active in the mitochondria, or whether it is directly involved in deacetylating accompanying with expansion of adipocyte cell size in epididymal adipose mitochondrial proteins. To elucidate how SIRT1 can regulate mitochondrial tissues and SVFs from high fat diet (60% fat chow) induced obese mice, proteins deacetylation, we performed western blot analysis and SIRT1 which suggested that visceral adiposity may associated with dysfunction of activity assay. Consequently, we demonstrated that full length SIRT1 is primary cilia. Further investigation revealed that either disrupted the number mainly localized in the nucleus, whereas the N-terminal short version of of primary cilia by 2mM choral hydrate or impaired the function of primary SIRT1 (N-SIRT1) is translocated into the mitochondria, and deacetylates the cilia via lentivirus mediated Kif3a and IFT88 shRNA, lipid accumulations candidate proteins. These data suggest that SIRT1 is directly involved in were both enhanced in epididymal SVFs from lean mice on normal chows vital cellular processes in fat cells by regulating the acetylation levels of in vitro. In addition, insulin sensitivity were attenuated, markers of lipo/ mitochondrial metabolic proteins in the mitochondria. adipogenesis such as Pparγ and Srebp-1c were upregulated while the Supported By: Agency for Science Technology and Research (A*STAR) marker of lipolysis such as HSL was downregulated in both epididymal adipose tissue and SVFs from HFD-induced obese mice and SVFs treated 1956-P with 2mM choral hydrate or shRNA lentivirus. In conclusion, primary cilia act Unique and Overlapping Transcriptomic Landscapes Defi ne Tissue- as an insulin signaling sensor to maintain the balance of lipo/adipogenesis specifi c Responses to Long-term Dietary Calorie and Methionine and lipolysis in adipose tissue, once disrupted by extracellular environments, Restriction in Rats it will affect insulin sensitive and lead to adiposity either by enhance of SUJOY GHOSH, DESIREE WANDERS, KIRSTEN P. STONE, NANCY VAN, CORY lipogenesis or impairment of lipolysis. CORTEZ, THOMAS GETTYS, Singapore, Singapore, Baton Rouge, LA Supported By: Postdoctoral Science Foundation of China (2012M521590, Dietary methionine and calorie restriction (MR/CR) without malnutrition 2013T60792) produce an integrated series of biochemical and physiological responses that improve biomarkers of metabolic health, limit fat accretion, enhance 1954-P insulin sensitivity, and extend lifespan. Using comprehensive transcriptional Impaired Adipogenesis in Adipose Tissue Associated with Hepatic profi ling to evaluate the molecular responses to these dietary regimens, we Lipid Deposition Induced by Chronic Infl ammation in Mice with identifi ed liver and adipose tissue as the primary targets of transcriptional Chow Diet programs that were generally more pronounced in MR than CR. The Integrated SHUMIN YANG, WENLONG ZHANG, RUFEI GAO, QIFU LI, Chongqing, China responses to chronic CR include up-regulation of genes involved in fatty acid POSTERS Impaired adipogenesis in adipose tissue has rarely been discussed in an biosynthesis and down-regulation of cytoskeletal matrix related genes in animal model with hepatic lipid deposition induced by chronic infl ammation adipose tissue. Chronic MR also increased expression of genes associated Physiology/Obesity in mice with chow diet. This study aims to explore impaired adipogenesis with fatty acid synthesis in adipose tissue along with a down-regulation in the adipose tissue as well as hepatic lipid disturbance in the liver of the of immune-infl ammation signaling genes in both adipose tissue and liver. C57BL/6J mice with hepatic lipid deposition induced by chronic infl ammation. Additionally, a coordinate program associated with gene translation was C57BL/6J mice was injected with casein for 18 weeks to establish an animal evident in liver of the MR group, driven by up-regulation of a cluster of genes model of chronic infl ammation. Serum levels of pro-infl ammatory cytokines encoding ribosomal proteins. The transcriptional responses to the two diets were analyzed by ELISA. Hepatic triglyceride was analyzed quantitatively and in brown adipose tissue were similar and less encompassing, although both expressions of molecules involved in hepatic lipid metabolism and adipose diets produced a notable down-regulation of genes linked to PPAR-signaling. adipogenesis were examined. Serum levels of amyloid A protein, IL-6, TNF-α Under similar statistical thresholds, no signifi cant pathway changes were and monocyte chemoattractant protein-1 were signifi cantly increased in identifi ed in skeletal muscle from either dietary regimen. Additionally, casein mice. Concentrations of fasting serum insulin and free fatty acid were topological relationships among identifi ed pathways were examined and

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A501 OBESITY—ANIMALCATEGORY

showed signifi cant evidence of expression co-regulation and the formation 1959-P of functionally interacting networks. These results provide an integrated Interesterifi ed Fats Induce Nonalcoholic Steatohepatitis or Enhance picture of the breadth of transcriptional responses to MR and CR among key the Adipose Tissue and Adipocyte Hypertrophy metabolic tissues. As such, they have the potential to serve as an analytic MARIA SILVIA F. LAVRADOR, MILESSA S. AFONSO, MARCIA KOIKE, RENATA P. platform for further hypothesis generation and testing regarding overlapping BOMBO, DENNYS E. CINTRA, VALÉRIA S. NUNES, SÉRGIO CATANOZI, EDNA R. and unique cellular response signatures produced by the respective diets. NAKANDAKARE, ANA MARIA LOTTENBERG, São Paulo, Brazil Supported By: ADA (1-12-BS-58); NIH (DK-096311, U24DK076169, P20GM103528, The interesterifi ed fats, rich in saturated fatty acids and trans free P30DK072476) have been used in the food industry. However, their impact on metabolic alterations is not yet elucidated. 1957-P Weaning LDLr-KO male mice were randomly distributed into fi ve groups Magnolia Extract and Its Bioactive Constituent 4-O-methylhonokiol fed with a high fat diet (40% of energy as fat) containing polyunsaturated Prevent Cardiac Hypertrophy via Suppression of Lipid Accumulation, (PUFA), palmitic (PALM), palmitic interesterifi ed (PALM INTER), stearic Oxidative Stress, and Infl ammation in Obesity Mice Induced by (STEAR) or stearic interesterifi ed (STEAR INTER) fats during 16 weeks. High-Fat Diet Dietary intake, body composition, liver fat and weight, total cholesterol, ZHIGGUO ZHANG, XIAOQING YAN, YANG ZHENG, YI TAN, LU CAI, YOUNG HEUI triglycerides, glucose, insulin concentrations and adipose tissue histology KIM, Changchun, China, Louisville, KY, Shanghai, China were determined. Histological markers of NAFLD were also assessed. Obesity increase cardiac hypertrophy associated with cardiac lipid There was no difference in dietary intake among the groups. The inter- accumulation, infl ammation, and oxidative stress. Magnolia extract (BL153) as esteri fi cation process did not alter plasma lipid concentrations (mg/dL) as an herbal obtained from Magnolia offi cinalis was found to play an important observed between PALM vs. PALM INTER (499.2 ± 22.5 vs. 526.8 ± 20.7) and role in anti-infl ammation and -oxidative stress. Here we investigated the STEAR vs. STEAR INTER (342.6 ± 21.8 vs. 363.9 ± 22.5). However, PALM and effects of BL153 and its bioactive constituent 4-O-methylhonokiol (MH) PALM INTER presented higher total plasma cholesterol, insulin and glucose on cardiac damage in obese mice induced with high fat diet (HFD) feeding. as compared to the others. Regarding triglycerides, all the groups were higher C57BL/6 mice were treated with vehicle, BL153 (5 mg/kg), MH (0.5 mg/kg or than PUFA (129.9 ±46.3; P<0.05). PALM INTER presented higher epididimal 1.0 mg/kg) by gavage daily with normal diet or HFD feeding for 24 weeks. (4.04 ±0.45; p<0.05) and subcutaneous (2.17 ± 0.39; p<0.05) adipose tissue MH treatment at 0.5 mg/kg signifi cantly ameliorated insulin resistance with than other groups (values expressed as g/100gbody weight) and additionally slightly decreased body weight gain induced by HFD feeding. MH signifi cantly more hypertrophy adipocytes (4356 µm2 ±1190; p<0.01). The liver weight, attenuated HFD-induced hypertriglyceridemia and hypercholesteremia. Both cholesterol and TG content did not differ among the groups. Nevertheless, BL153 and MH treatments signifi cantly ameliorated cardiac hypertrophy along STEAR INTER developed nonalcoholic steatohepatitis characterized by a with the reduction of cardiac lipid accumulation, fatty acid translocase/CD36 diffuse macrosteatosis, hepatocellular ballooning, infl ammatory infi ltration expression, and infl ammation (tumor necrosis factor-α; and plasminogen and fi brosis. activator inhibitor-1) and elevation of peroxisome proliferator-activated In conclusion, interesterifi ed fats bring on an increase in adipose tissue receptor γ coactivator 1 α (PGC1α) expression, all of which was associated and induce adipocyte hypertrophy (PALM INTER) and lead to NASH (STEAR with the suppression of oxidative damage (malondialdehyde) by increasing INTER). nuclear-factor-E2-related-factor-2 (Nrf2). Our study indicated that BL153 Supported By: FAPESP (2011/24083-7, 2012/50249-2) and MH ameliorated HFD-induced cardiac hypertrophy, lipid accumulation, infl ammation, oxidative stress via downregulation of CD36 and upregulation 1960-P of PGC1α and Nrf2. Reciprocal Effects of 14-3-3ξ Knockout and Overexpression on Murine Obesity 1958-P GARETH E. LIM, NANCY N. FANG, SUSANNE M. CLEE, AMPARO ACKER-PALMER, CD47 Defi ciency Protects Mice from High-Fat Diet-induced Obesity THIBAULT MAYOR, JAMES D. JOHNSON, Vancouver, BC, Canada, Frankfurt, and Infl ammation Germany HEATHER L. NORMAN-BURGDOLF, HASIYETI MAIMAITIYIMING, SHUXIA WANG, Obesity stems from the expansion of fat mass by hypertrophy of adipocytes Lexington, KY and the differentiation of adipose precursor cells. A complex transcriptional Obesity is becoming a global epidemic, which is associated with chronic network regulates adipogenesis, and molecular adaptor proteins, such as infl ammation and the development of insulin resistance and other metabolic those belonging to the 14-3-3 protein family, are required to ensure the proper complications. Although there are signifi cant advances in the studies relating degradation rates and spatial localization of transcription factors. We found to obesity, the exact mechanism still remains unclear of how obesity- essential roles for the 14-3-3ζ isoform in adipogenesis through the analysis associated chronic infl ammation initially develops within adipose tissue. of 3T3L1 cells treated with 14-3-3 siRNA or a small molecule inhibitor, as Previous research in our lab has established that thrombospondin-1 (TSP1), well as the study of 14-3-3ζ knockout mice, which have signifi cantly reduced a stress protein secreted and associated with various cellular responses, adiposity and poorly differentiated adipocytes. Conversely, transgenic contributes to obesity-induced infl ammation and impaired insulin signaling. mice with 14-3-3ζ over-expression were signifi cantly heavier than WT CD47, a key receptor for TSP1 expressed on a variety of cells, once activated mice at 1 year of age, but did not exhibit impaired glucose tolerance or inhibits the Nitric Oxide (NO)/cGMP/PKG signaling cascade and impairs anti- insulin sensitivity. Over-expression of 14-3-3ζ potentiated weight gain infl ammatory mechanisms within the vasculature. However, it is unknown in response to high-fat diet feeding. DEXA analysis revealed signifi cantly whether this proinfl ammatory effect that was elicited by TSP1 under obese increased fat mass in 14-3-3ζ over-expressing animals, but this increase in conditions is mediated by CD47. This hypothesis was tested by using CD47 adiposity had no effects on glucose tolerance or insulin sensitivity. To gain defi cient mice. Interestingly, we found that CD47-/- mice were resistant mechanistic insight into how 14-3-3ζ facilitates adipocyte differentiation, to high fat diet-induced obesity, which was associated with reduced we used proteomics to determine the 14-3-3ζ interactome from MEFs macrophage infi ltration into adipose tissue and decreased expression of derived from TAP-14-3-3ζ transgenic mice. Of the 350 proteins detected several proinfl ammatory cytokines (TNF-a, IL-6, IL-1B). Knowing that both by mass spectrometry, 180 proteins were unique to differentiating cells. TSP1-/- and CD47-/- mice display similar phenotypes when challenged with When grouped by their biological functions, these proteins were involved Integrated POSTERS high fat diets, we believe that their ligation could be responsible for the in RNA processing, actin cytoskeleton remodeling, nucleic acid transport, increased adipokine and cytokine production seen in both adipocytes and translation, and macromolecular protein complex assembly, all functions that Physiology/Obesity MI macrophages within adipose tissue depots. Further research is needed to are essential for adipogenesis. Collectively, we report an obesogenic effect understand how activation of CD47 plays a role in regulating adipose function of elevated 14-3-3ζ levels and suggest targeting 14-3-3ζ or constituents of and macrophage interactions in obesity-associated chronic infl ammation. its interactome as novel therapeutic approaches for obesity. Supported By: 5T32DK007778-14 Supported By: JDRF; CDA

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A502 OBESITY—ANIMALCATEGORY

1961-P (p<0.001) higher iron concentration. To further phenotype and characterize Physiologic and Genetic Evidence Links Hemopexin to Adipocytes the adipose tissue iron overload and evaluate the associated systemic in Obesity effects, 27 male KK mice aged 47-79 wks were evaluated. Fourteen mice had HEATHER A. LAWSON, LARRY D. SPEARS, IRFAN J. LODHI, CLAY F. SEMENKO- discolored eAT (DC) and 13 had normal colored eAT (NC). Fasting serum was VICH, St. Louis, MO collected and tissues were harvested for iron content, gene expression, and Iron impacts diabetes risk through poorly understood mechanisms that histology. Robust elevations of iron were confi rmed in the eAT of DC versus extend beyond those involved in classic disorders of iron storage such NC mice. Surprisingly, iron levels in subcutaneous and brown adipose as hemochromatosis. The adipocyte appears to possess iron-sensing depots were not different between groups (p>0.05). However, tissue iron properties that may be related to the secretion of adipokines, control of levels were also increased in liver (26%, p<0.01), pancreas (44%, p<0.01) lipolysis, and modulation of infl ammation, all potential contributors to and heart (29%, p<0.01) of the DC group. The eAT histology with H&E and obesity-related morbidities such as type-2 diabetes and cardiovascular F4/80 staining revealed a robust macrophage clustering, while trichrome disease. To test the hypothesis that a discrete iron-related protein is and caspase-3 staining revealed more fi brosis and cell death in the DC eAT. regulated by the nutritional state of adipocytes, we studied the adipokine The eAT rtPCR showed notably decreased Lep (leptin), Adipoq (adiponectin) hemopexin, a heme-binding protein that circulates in plasma. We found that and Vegf (vascular endothelial growth factor) (p<0.05), whereas Tnfα during the adipocyte differentiation program in 3T3-L1 cells, hemopexin (tumor necrosis factor) and the iron exporter, Slc40a1 (ferroportin), were up expression increases strikingly with peak levels observed around the time regulated in DC group (p<0.05, DC, n=4; NC, n=6). No difference of fasting of substantial development of lipid droplets at 5 days following induction. serum glucose, serum insulin and liver TG was observed between groups To confi rm this observation in an animal model, we studied hemopexin (p>0.05). Our data suggest that the deposition of iron is site (eAT) specifi c expression in chow fed and high-fat (42%) fed mice. Circulating hemopexin in male KK/HIJ mice. The increased macrophages, collagen and cell death levels were increased in high-fat fed relative to chow fed animals. Further, indicate a robust adipose tissue remodeling in the high iron eAT. However, hemopexin expression was signifi cantly increased in adipose tissue but not no association was evident between eAT iron deposition and glucose liver of high-fat fed as compared to chow fed mice. Since hemopexin may homeostasis. control the functional status of circulating HDL particles, we also pursued genetic evidence of a relationship between hemopexin and mediators of 1964-P obesity-related disease. We identifi ed hemopexin as a top candidate gene New Diagnostic Criteria for Type 2 Diabetes Determined for Non- associated with three independently mapped quantitative trait loci (QTL) for human Primates variation in serum lipids. For cholesterol, Chldq4, was identifi ed in a MRL/ BARBARA C. HANSEN, JENNIFER D. NEWCOMB, ELLEN H. LINDEN, Tampa, FL MpJ x SJL/J cross and Hdlcl1 was identifi ed in a C57BL/6J x PERA/EiJ cross. Many metabolic processes change dynamically during the progression For triglycerides, Dserum7b was identifi ed in a LG/J x SM/J F16 advanced from obesity to overt type 2 diabetes (T2DM), and differ with disease intercross. These fi ndings suggest that factors promoting fat cell mass, such progression rates. Because nonhuman primates (NHPs) develop obesity- as dietary fat, increase hemopexin in adipose cells to alter the infl ammatory associated T2DM extraordinarily like that of humans, the standard ADA milieu in obesity. diagnostic criteria have been used to date. NHPs, however, have distinct differences which we now show warrant an independent diabetes diagnostic 1962-P threshold. Most notably, the fasting plasma glucose (FPG) in normal NHPs is Antioxidant α-Tocopherol Ameliorates Glucose Intolerance, Lipid 57.3±0.4 mg/dl compared to ~85 mg/dl in humans, a difference of > 25 mg/dl. Metabolism, and Liver Steatosis More Effi ciently Than γ-Tocopherol To determine an appropriate point in the disease progression for a monkey in High-Fat Diet-Fed Mice to be diagnosed with overt T2DM we studied longitudinally 67 overweight KYOKO TODA, KEIICHI KODAMA, YOSHIFUMI YAMADA, SHOJIRO MORINAGA, rhesus macaques (Macaca mulatta, 62 males) who became diabetic while SATORU YAMADA, ATUL J. BUTTE, Tokyo, Japan, Stanford, CA under study. Upon reaching an FPG of 126 mg/dl they ranged in age from 10.3 An excess of reactive oxygen species that overcomes the capacity of to 30.4 yrs and weight 6.3 to 26.1 kg. FPG, triglycerides (TGs), fasting Insulin, the antioxidant system induce oxidative stress in cells. Oxidative stress Acute Insulin Response (AIR) to IV glucose, weights and ages were compared is associated with metabolic disorder condition such as nonalcoholic within and across subjects at three different potential diagnostic thresholds steatohepatitis, insulin resistance and type 2 diabetes. Antioxidant (FPG levels of 80, 100 and 126 mg/dl) which occurred at ages 17.2±0.6, vitamin E consists of two major forms: α- and γ-tocopherol. α-Tocopherol 18.9±0.6 and 19.1±0.6 yrs, respectively. (Note the brief time between the is quantitatively the major form of vitamin E in humans and animals and 100 and 126 thresholds.) TGs were elevated at all three thresholds, with the is a more potent antioxidant in vitro than γ-tocopherol. To determine if 80 mg/dl level signifi cantly below those of the 100 and 126 levels which did antioxidant vitamin E (α-/γ-tocopherol) has benefi cial effects on the glucose not differ from each other. Prior to T2DM onset in both humans and NHPs, and lipid metabolism, eight-week-old male C57BL/6J (B6J) mice were fed a insulin rises signifi cantly over baseline, peaks and then drops. At the 80 mg/ high-fat diet containing 0 (control) or 0.1% (wt/wt) α- or γ-tocopherol for 16 dl threshold, both insulin and AIR were at or near longitudinal peak levels weeks. Intraperitoneal glucose tolerance test (IPGTT) revealed a signifi cant and signifi cantly higher than at both the 100 and 126 mg/dl FPG thresholds decrease in blood glucose levels at 30, 60, 90 and 120 min and a signifi cant (all p’s <0.001) indicating that the NHPs were progressing toward diabetes. increase in insulin secretion at 2 to 120 min after glucose injection into These extensive data, together with the naturally occurring ~25 mg/dl lower the B6J mice fed with α-tocopherol diet. The serum triglyceride (TG) and FPG levels in normal NHPs, lead us to conclude that for NHPs T2DM should LDL-C levels were reduced in the α-tocopherol diet group. Examination of be diagnosed at an FPG level of 100 mg/dl (repeated 2X), which is most histological sections revealed an obvious decrease in lipid accumulation equivalent to the current human T2DM diagnostic level of 126 mg/dl. in the livers of α-tocopherol diet mice compared to controls. Hepatic TG Supported By: NIH (HHSN2632008000022C) (mg/g liver) analysis indicated that hepatic TG levels in the α-tocopherol group were decreased compared to controls. The metabolic measurements 1965-P indicated above were also ameliorated by γ-tocopherol, but not statistically Astaxanthin, a Strong Antioxidant, Ameliorated Glucose Metabolism signifi cantly, compared to control. These fi ndings demonstrated that in Obese Mouse Through Various Mechanisms α-tocopherol supplementation ameliorated glucose and lipid metabolism MANABU ISHIKI, YASUHIRO NISHIDA, AKIKO TAKIKAWA, YUKIKO KOSHIMIZU, Integrated and fatty liver of diet-induced obese mice, suggesting the importance of MINORU IWATA, ISAO USUI, KAZUYUKI TOBE, Toyama, Japan POSTERS antioxidative intervention as a supportive therapy of type 2 diabetes. Oxidative stress is one of the major causes of impairment of glucose metabolism in obesity. We have reported that astaxanthin, an antioxidant Physiology/Obesity 1963-P belonging to the carotenoid, restored insulin signaling impaired by oxidative Robust Tissue-specifi c Iron Elevation and Adipose Tissue Remodel- stress generated by cytokine or free fatty acid in L6 cells, although its effect ing in a Polygenic Obese Mouse Model in vivo remains to be elucidated. Here we explored if it improves glucose XIAOYA MA, VINH T. PHAM, YATRIK M. SHAH, PETER F. BODARY, Ann Arbor, MI metabolism in high fat -diet mouse by carrying out glucose tolerance Iron overload is frequently observed in type 2 diabetes. Iron dysregulation test, insulin tolerance test, real time PCR and western blotting in various is a potential contributor to the pathology of obesity-related metabolic tissues. We found that astaxanthin improved insulin resistance and glucose complications. KK/HIJ (KK) mice, a polygenic mouse model of obesity and metabolism. Although it restored PGC1α, PPARα, ATP5G1, Tfam, MCH1, insulin resistance, have elevated serum iron levels and a propensity for MCHIIa, CD36, Cpt1b, MCD in their mRNA level in skeletal muscle and MCP1, tissue iron deposition. In subsets of older KK males, we have observed a CD11C in adipose tissue in early period, it had little infl uence on liver. On discoloration of epididymal adipose tissue (eAT) associated with >100-fold the contrary, it restored IL-1β, MCP1, CD11C, FASN, G6PC in liver during late

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A503 OBESITY—ANIMALCATEGORY

period. Phosphorylation of signaling molecules such as insulin -induced Akt, or AMPkinase were restored by astaxanthin in skeletal muscle. Furthermore, the results obtained by hyperinsulinemic euglycemic clamp indicated the improvement of insulin resistance by this antioxidant in early period seems to be occurred mainly in skeletal muscle. We speculate that astaxanthin improved βoxidation in skeletal muscle during the process of accumulation of fat tissue in early period through acceleration of formation of slow twitch fi ber and modulation of several genes. And then, in late period, infl ammation in the fat tissue, which followed by insulin resistance in liver and skeletal muscle, appears to be restored. Moreover, insulin dependent or independent signaling pathways in skeletal muscle seem to involve in modulation of glucose metabolism by astaxanthin. These results indicate that the very unique character of astaxanthin and its variable actions, which could make this antioxidant be a very effective tool which improves oxidative stress- induced abnormal glucose metabolism in vivo.

1966-P Medica Analog Ameliorates Obesity-induced Diabetes MICHAEL VALITSKY, YAEL KUPERMAN, AMNON HOFFMAN, JACOB BAR-TANA, Supported By: Shanghai Natural Science Foundation Jerusalem, Israel, Rehovot, Israel Background: MEDICA analogs consist of substituted long-chain, α, ω dioic 1968-P fatty acids that are not esterifi ed into lipids, nor β oxidized, but excreted Resolvin D1 Reduces Body Mass in Diet-induced Obesity in bile as respective glucuronides. MEDICA analogs proved anti-diabetic LIVIA APARECIDA BITENCOURT PASCOAL, LETÍCIA MARTINS IGNÁCIO DE effi cacy in a series of genetic diabetic animal models, being partly accounted SOUZA, ANDRESSA COOPE DOS SANTOS, RODRIGO FERREIRA DE MOURA, for by activating AMPK and by promoting low-conductance gating of the BRUNA BOMBASSARO, DANIELA SOARES RAZOLLI, RODRIGO RAMOS CATHA- mitochondrial permeability transition pore. RINO, LÍCIO AUGUSTO VELLOSO, Campinas, Brazil, Cuiabá, Brazil, Ann Arbor, MI Objectives: Current study aimed at probing MEDICA effi cacy in ameliorating Hypothalamic infl ammation is a common feature of diet-induced obesity. high-fat diet (HFD)-induced diabesity. It is installed right after the introduction of a high-fat diet and results from Methods: MEDICA effi cacy and its mode-of-action were studied in 10- saturated fatty acid activation of TLR4 signaling and endoplasmic reticulum week old C57Bl male mice fed HFD for 120 days, with MEDICA analog added stress. It was recently shown that ω-3 unsaturated fatty acids can reverse in diet (0.04% W/W). the obese phenotype and reduce hypothalamic infl ammation in rodents fed Results: MEDICA treatment resulted in decrease in body weight gain on a saturated fatty acid rich diet (Cintra PLoS One 7: e30571, 2012). Here (7.6±0.3 gr vs. 19±1 gr) and body fat (25.1% vs. 40.4%), despite of increase we tested the hypothesis that, resolvins play a role in the anti-infl ammatory in food consumption. Decrease in body fat was accompanied by increase effect of ω-3 unsaturated fatty acids in the hypothalamus of obese mice. Our in lean body mass. The decrease in metabolic effi ciency was accounted for results show that, in mice, the enzymes involved in the synthesis of resolvins, by increase in total body energy expenditure, without an apparent change lipoxigenase -5 and -15 are present in the hypothalamus and their expressions in locomotion activity. MEDICA treatment resulted in normalizing glucose are modulated upon high-fat feeding, as determined by real-time PCR and intolerance, insulin resistance and hepatic glucose production as verifi ed by immunoblot. This is accompanied by decreased hypothalamic concentration glucose, insulin and pyruvate tolerance tests. of resolvin D1 (RvD1) in mice fed on a high-fat diet and increased in mice fed Conclusions: MEDICA analogs ameliorate diet-induced diabesity by on a ω-3 unsaturated fatty acids diet as determined by MALDI. The receptor increase in energy expenditure and decrease in metabolic effi ciency. for RvD1, GPR32 is expressed in the hypothalamus, both in NPY and POMC neurons and in CD11c expressing microglia cells. The intracerebroventricular 1967-P (icv) treatment of obese mice with 3.0 ng/day of RvD1 for 11 days promoted CTSK Inhibitor Exerts Its Antiobesity Effects through Regulating reduced body mass, reduced visceral adipose tissue mass, increased Adipocyte Differentiation in High-Fat Diet-induced Obese Mice hypothalamic expression of the anti-infl ammatory cytokines IL10 and IL6, JUNFENG JAN, Shanghai, China increased O2 consumption and reduced RQ, suggesting increased energy Objective: Cathepsin k (CTSK) is highly expressed in adipose tissues of expenditure and higher lipids oxidation. All these effects of the icv treatment obesity patients and obese animal models. While it had demonstrated that with RvD1 were accompanied by increased expressions of UCP1 and PGC1α CTSK promote the process of adipocyte differentiation in 3T3-L1 cells. The in the brown adipose tissue (BAT) and reduction of glucose intolerance as actions of CTSK inhibitor in the prevention of mouse body weight gain and determined by the glucose tolerance test. Thus, icv treatment with RvD1 insulin resistance in vivo are not well understood. reverses the obese and glucose intolerance phenotypes. Activation of anti- Methods: High-fat diet (HFD)-induced obese Male C57BL/6 mice were infl ammatory activity in the hypothalamus and induction of thermogenic fed with or without CTSK inhibitor (CKSI, 0.1-10umol/kg, every other day) genes in the BAT are potentially involved in the benefi cial effects of this ω-3 for 8 weeks. The mechanism was further studied using western-blot and derived anti-infl ammatory substance. immunohistology methods. Results: The HFD induced adipose tissue weight gain, increase of HOMA 1969-P index and an accumulation of large adipocytes. After CKSI treatment, all Small Molecule Kaempferol Prevents Type 2 Diabetes by Promoting these effects were blunted compared with the HFD control group. Mechanism Insulin Sensitivity and Reducing Hepatic Gluconeogenesis study demonstrated a role for CTSK inhibitor in signifi cantly down-regulation WILLIAM MOORE, HANA ALKHALIDY, RYAN MCMILLAN, WEI ZHEN, AIHUA the expression of two key transcription factors, peroxisome proliferator- WANG, DONGMIN LIU, Blacksburg, VA activated receptor-γ (PPARγ) and CCAAT/Enhancer-Binding Protein α (C/

Integrated Insulin resistance is a primary risk factor for developing prediabetes and POSTERS EBPα), which are markers of adipogenic differentiation. type 2 diabetes (T2D). Thus, searching for natural, low-cost compounds to Conclusions: These results indicated that CKSI possess anti-obesity Physiology/Obesity prevent insulin resistance or promote its sensitivity could be a promising effect, possibly involving the inhibition of adipocyte differentiation. CTSK strategy to prevent the pathogenesis of T2D. is likely to be a new target of therapeutic strategy in the management of In the present study, we show that dietary intake of kaempferol (0.5 g/ obesity. kg diet), a plant-derived fl avonol, signifi cantly ameliorated hyperglycemia and hyperinsulinemia, which were associated with the improved peripheral insulin sensitivity in high-fat diet-induced obese mice. Kaempferol treatment reversed high-fat diet impaired glucose transport-4 (Glut4) and AMP- dependent protein kinase (AMPK) expression in both muscle and adipose tissues from obese mice. Oral administration of kaempferol to obese mice inhibited glucose production in vivo. We further found that kaempferol treatment signifi cantly increased basal glucose uptake in C2C12 myotubes. In addition, pre-treatment of C2C12 cells with kaempferol (1-10 µM) pre-

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A504 OBESITY—ANIMALCATEGORY vented the harmful effect of saturated fatty acids on lowering glucose uptake. Further, kaempferol increased PGC-1α expression and AMPK activity in skeletal muscle cells and suppressed basal and insulin-regulated gluconeogenesis in hepatocytes. These data demonstrate that kaempferol may be a low-cost natural compound that can be used to prevent T2D by improving insulin sensitivity and reducing hepatic gluconeogenesis. Supported By: ADA (1-14-BS-018); NCCAM/NIH (1R01AT007077)

1970-P High-Fat Diet Impairs the Myocardial Inotropic Response to β-Adrenergic Stimulation by Inducing Phosphodiesterase (PDE4) YUAN ZHANG, TREVOR P. FIDLER, BHARAT P. JAISHY, YANG K. XIANG, E. DALE ABEL, Iowa City, IA, Salt Lake City, UT, Davis, CA The global epidemic of obesity, insulin resistance and diabetes mellitus has increased cardiovascular disease and heart failure via complex mechanisms that may involve impaired beta-adrenergic signaling. We therefore placed C57BL6/J mice on a high fat diet (HFD- 45% fat) for 12 weeks and hearts were subjected to Langendorff perfusion without pacing in KHB buffer. In control hearts, LV developed pressure (LVDP) and the maximum and minimum rates of change in LV pressure (±dP/dt) increased by 2.7, 2.3 and 2.8 fold respectively in response to increasing concentrations of isoproterenol (ISO) from 10-18M to 10-7M. Although the heart rate response to ISO was similar in both groups, the increase in LVDP and ±dP/dt was blunted in HFD hearts (1.3 - 1.5 fold, P<0.05 vs. control). Protein levels of PDE4 increased by 3.07±0.77 fold in HFD mouse hearts relative to chow- fed controls (P<0.05), and were associated with a 1.39±0.12 fold change of PDE4 mRNA. Consistent with increased PDE4-mediated hydrolysis of cAMP, Supported By: NSFC (81070667, 81270925) PKA signaling was impaired by HFD. Specifi cally, levels of phosphorylated phospholamban were decreased by 78 % in HFD vs. controls (p<0.005). 1972-P Additionally, the β-AR mediated increase in Akt and mTOR phosphorylation Evidence That a Subset of Beige Cells Arise from Smooth Muscle was blunted in the HFD hearts. PDE4 expression was induced following 12hr. Precursors incubation with either insulin (100nM) or the β-adrenergic receptor agonists JONATHAN Z. LONG, KATRIN J. SVENSSON, XING ZENG, LINUS TSAI, HYUN C. (Dobutamine and Clenbuterol, 10µM) in H9C2 cardiomyocytes. Interestingly, ROH, XINGXING KONG, RAJESH R. RAO, JESSE LOU, ISHA A. LOKURKAR, EVAN this increase was blunted by the combined treatments of insulin+Dobutamine D. ROSEN, BRUCE M. SPIEGELMAN, Boston, MA or insulin+Clenbuterol, suggesting inhibitory crosstalk between insulin and Thermogenic Ucp1-positive adipocytes, which include brown and beige β-adrenergic signaling in the regulation of PDE4 expression. Thus insulin adipocytes, transform chemical energy into heat and increase whole body resistance and obesity blunt the inotropic response to β-adrenergic receptor energy expenditure. While classical brown fat arises from a skeletal muscle agonists by a mechanism involving PDE4-mediated degradation of cAMP and lineage, the origins and molecular characteristics of beige adipocytes in vivo PKA signaling, representing a novel mechanism linking insulin resistance, are less well understood. Here, we adopt a translational profi ling approach hyperinsulinemia, obesity and cardiac dysfunction. to directly and selectively characterize in vivo Ucp1-positive adipocytes Supported By: NIH in mice. We show that the majority of the gene expression between stimulated brown and beige fat is equivalent and represents a common 1971-P core thermogenesis program. Of the differentially expressed genes, beige Wnt/β-catenin Signaling Pathway and Lipolysis Enzymes Partici- adipocytes express a signature classically associated with smooth muscle pate in Methylprednisolone Induced Fat Differential Distribution cells. In vivo fat mapping shows that at least a subset of beige cells arise between Subcutaneous and Visceral Adipose Tissue from a smooth muscle lineage. Finally, in vitro, ectopic expression of XINHUA XIAO, HAN LI, JIAOJIAO YANG, XUYU ZU, MINGYAN GAO, JIANGHUA PRDM16, a transcription factor required for the development of beige cells, LIU, GEBO WEN, Hengyang, China in smooth muscle cells promotes their differentiation into Ucp1+ adipocytes. Different adipogenic and lipolytic effects between subcutaneous Taken together, these results establish the fi rst global portrait of brown and adipose tissue (SAT) and visceral adipose tissue (VAT) is likely to play a beige adipocyte gene expression in vivo and identify a hitherto unrecognized role in glucocorticoids (GCs) induced fat differential distribution. Sprague- origin for at least a subset of beige cells. Dawley rats were randomly divided into two groups, control (n=15) and Supported By: ADA/CDA (7-12-CDA-01) methylprednisolone (MPL) (n=16). The MPL group was intramuscularly injected with 4 mg/kg/d MPL for 12 weeks. The body weight and food 1973-P consumption were measured respectively. The SAT and VAT were stained Myeloid Cell-specifi c Deletion of Hypoxia-Inducible Factor 1-alpha with H&E. QPCR and Western blotting were used to detect mRNA and protein Gene Protected against Diet-induced Insulin Resistance in Mice expression. MPL attenuated body weight gain and increased VAT mass but AKIKO TAKIKAWA, ARSHAD MAHMOOD, SHIHO FUJISAKA, TAKASHI not SAT mass. MPL impaired glucose tolerance and lipid profi le by increasing NAKAGAWA, AMINUDDIN AMINUDDIN, SATOKO SENDA, ISAO USUI, KAZUYUKI TC, TG, and LDL level. Subcutaneous adipocyte size was signifi cantly smaller TOBE, Toyama, Japan and the average number of adipocytes exhibited an increase in MPL treated Adipose tissue becomes hypoxic as obesity progresses because oxygen rats. The expression of β-catenin and Wnt-10b were lower and DKK-1 was supply cannot meet the demand of expanding adipose tissue. Although Integrated higher in MPL rats than those in control rats. MPL showed an obviously adipose tissue hypoxia has been implicated in the obesity-induced POSTERS suppressive effect on protein expression of ATGL and HSL in VAT but little infl ammation and insulin resistance, the causal relationship has not Physiology/Obesity change in SAT. Our data suggest that differential expression of lipolysis precisely elucidated. We investigated the role of hypoxia-inducible factor enzymes induced by MPL between SAT and VAT might play crucial role in fat (HIF)-1α gene in myeloid cells in the development of high-fat diet (HFD)- distribution. Those fi ndings would offer novel insights into the mechanisms induced systemic insulin resistance in mice model. Mice with myeloid- of GCs induced fat distribution. specifi c targeted disruption of the gene encoding Hif-1α (Mye-HIF-1α KO) were generated using a LysM-Cre transgene with the Cre/LoxP system. The mice were fed an HFD for 16 weeks and their metabolic phenotypes were determined. Gene expression patterns in adipose tissue, liver, and skeletal muscle were also determined by quantitative PCR. Mye-HIF-1α KO mice showed improved glucose tolerance with lower serum insulin concentrations, indicating improved insulin sensitivity in KO mice. KO mice have less crown- like-structures in adipose tissue along with the reduced expression of CD11c,

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A505 OBESITY—ANIMALCATEGORY

an M1 macrophage marker, hypoxia-related genes including VEGF, Glut 1 and pharmacokinetic properties. Accordingly, these data demonstrate that osteopontin and infl ammatory genes including TNF-α and MCP-1. KO mice lipidated NMU analogues may represent promising drug candidates for the also demonstrated to express increased expression of metabolic genes treatment of obesity and diabetes. including adiponectin, PGC-1α, Glut4 and PPARγ. In the liver, expression of gluconeogenic genes as well as a pan-macrophage marker F4/80, and an M1 1976-P macrophage marker, CD11c but not an M2 macrophage marker CD206, was Proteomic Analysis of Serum Proteins in LEA Rat, a New Rat Model markedly decreased in KO mice. KO mice also exhibited increased expression of Nonobese Type 2 Diabetes Mellitus of PGC-1α and several mitochondrial genes in muscle compared to that in ERI TAKAHASHI, HIROYUKI UNOKI-KUBOTA, YUKIKO SHIMIZU, AKINORI OKU- wild- type mice. In conclusion, Myeloid-specifi c deletion of HIF-1α gene has MURA, TADASHI OKAMURA, MITSUHIKO NODA, YASUSHI KABURAGI, Tokyo, a benefi cial role against obesity-associated infl ammation, thus contributing Japan to the improvement of the whole body resistance. Recent advances in proteomics have shed light to discover serum proteins or peptides as biomarkers for tracking the disease progression as well as 1974-P understanding their molecular mechanisms. To identify candidate serum Pro-infl ammatory State of Bone Marrow Monocyte Might Contribute molecules associated with the progression of type 2 diabetes mellitus to Adipose Tissue Infl ammation in Diabetic Obese State (T2DM), we carried out a differential serum proteomic analysis using BATTSETSEG BATCHULUUN, TOYOSHI INOGUCHI, NORIYUKI SONODA, SHUJI a novel, non-obese rat strain with spontaneous diabetes, Long-Evans SASAKI, YUKA SAKAKI, BAIGALMAA BATCHULUUN, Fukuoka, Japan Agouti (LEA) rat. We employed two-dimensional fl uorescent difference The obesity epidemic caused an increased prevalence of obesity- gel electrophoresis-based proteomic approach to analyze the proteomic induced insulin resistance. In obese state, macrophage accumulation and changes in the sera collected from a pair of 8 week-old LEA versus control polarization in adipose tissue contribute to chronic infl ammation, leading BN rats (n=4/each group). Among a total of 2348 protein spots detected, to the development of insulin resistance. However, the role of monocyte 209 protein spots displayed signifi cant fold-change alterations in expression is not understood. Thus, we investigated whether monocyte is affected in between LEA and BN rats. We established the identity of 64 spots out of the diabetic obese state. Monocyte was isolated from bone marrow, peripheral total of 209, resulting in the identifi cation of 15 distinct proteins. In addition, blood and epididymal adipose tissue of db/db and db/+ mice, using monocyte the 15 candidates were validated by multiple reaction monitoring mass enrichment kit and subsequently stained with monocyte/macrophage spectrometry in the sera collected from another pair of LEA and BN rats (n=4/ markers for fl owcytometry analysis. Bone marrow monocyte was shifted each group), where 6 proteins were confi rmed to be differentially expressed into pro-infl ammatory state in db/db mice. Ly6C+ monocytes migrated into between the two groups. Among them, serine protease inhibitor (SERPIN) epididymal adipose tissue twice more than Ly6C- cells did. The ratio of M1 to A3K levels were signifi cantly increased in the sera of LEA rats compared M2 macrophage was signifi cantly increased in epididymal adipose tissue of with those in BN rats. We also investigated a possible association between db/db mice compared to that of db/+ mice. To determine the characteristics human homologue SERPINA3 levels and T2DM in 25 T2DM patients and 33 of bone marrow monocyte subtypes, Ly6C+ or Ly6C- monocytes were sorted healthy control subjects. The serum levels of SERPINA3 were signifi cantly out for in vivo monocyte tracking and total RNA microarray analysis. In vivo higher in T2DM group (195.9±38.1 µg/mL) compared with those in healthy tracking, fi rstly Ly6C+ and Ly6C- monocytes were stained with red (PKH26) group (176.3±26.7 µg/mL), and was signifi cantly associated with fasting and green (PKH67) dyes separately and then mixed prior administration of tail plasma glucose levels (β=0.29, p=0.036). These fi ndings suggest a possible vein injection. Labeled Ly6C+ monocyte were higher in adipose tissue of db/ role of SERPINA3 in the progression of T2DM although further replication db mice compared to Ly6C- monocyte. It also migrated into the kidneys more studies and functional investigation regarding the role of SERPINA3 are than Ly6C- monocytes did. To further determine whether Ly6C+ monocyte is needed. preprogrammed, Ly6C+ monocytes from db/db and db/+ mice were tracked Supported By: Grants-in-Aid for Scientifi c Research; Ministry of Health, Labour at the same time. Db/db Ly6C+ monocyte migrated into adipose tissue more and Welfare than db/+ Ly6C monocyte did. We also found distinct characteristics of bone marrow monocyte subpopulations in microarray analysis, including toll-like 1977-P receptors and cell adhesion molecules. In conclusion, monocyte is affected Effects of Sucrose or Fructose-and-Glucose Solutions on Gut Flora as early as in bone marrow level, contributing to adipose tissue infl ammation Composition in Sprague-Dawley Rats in diabetic obese state. GRZEGORZ WYSTRYCHOWSKI, EWA ZUKOWSKA-SZCZECHOWSKA, EWA OBUCHOWICZ, WLADYSLAW GRZESZCZAK, ANTONI WYSTRYCHOWSKI, Zabrze, 1975-P Poland, Katowice, Poland Lipidated NMU Analogues Decrease Acute Food Intake in Mice An increase of obesity in the U.S. coincided with use of high-fructose corn LOUISE S. DALBØGE, SØREN L. PEDERSEN, SØREN BLOK VAN WITTELOOSTUIJN, syrups (HFCS: HFCS55, blend of 55% fructose (F) and 42% glucose (G) and BIRGITTE HOLST, NIELS VRANG, JACOB JELSING, Hørsholm, Denmark, Copen- HFCS42, blend of 42%F and 53%G) as sweeteners. Although higher F content hagen, Denmark in HFCS55 than in sucrose (S) was suspected guilty, this is undermined by a Neuromedin U (NMU) is a 25 amino acid peptide expressed in the relatively stable ratio of F to G in the American diet. brain and gastrointestinal tract. In-house data has shown that peripheral Gut fl ora participates in the pathogenesis of obesity. S is not utilized by all administration of NMU decreases food intake and body weight and improves gut bacteria, which implicates that gut fl ora may differ with ingestion of free glucose tolerance in rodents. Therefore NMU receptors constitute a possible or bound F and G. In S-D rats we aimed to compare effects of S vs. F and G anti-diabetic and anti-obesity drug target. The clinical use of native NMU is on the gut contents of bacteria reported to affect body mass. hampered by the very short half-life. Consequently, we aimed to create a 60 male new-born rats were assigned to 18 weeks of unlimited access to series of NMU analogues with a longer half-life. The analogues were tested standard chow containing ~60% carbohydrates with no F and 5% solutions for their ability to activate both the peripheral and central NMU receptors of S (19 rats, S group), 55%F and 45%G (21 rats, 55F/45G) or tap water (20 (NMUR1 and NMUR2) and to decrease food intake in vivo. rats, W). Gut fl ora composition was assessed with quantitative analysis of A library of NMU analogues were established using solid-phase peptide DNA encoding bacterial 16S rRNA extracted from feces (Illumina HiSeq). synthesis in combination with an acylation scan using Nε-Lys lipidation. The F accounted for ~10% caloric intake in S and 55F/45G alike. Certain lean Integrated POSTERS functional activity of the analogues was next measured using an inositol phenotype-associated bacteria were more prevalent in S than in 55F/45G phosphate accumulation assay on HEK293 cells transfected with NMUR1 or W (table). Physiology/Obesity or NMUR2. Finally, in vivo activity was assessed in lean mice as reduction Unrestricted intake of S solution in rats, as opposed to solution of free F in food intake after acute subcutaneously administration of 1, 0.3, 0.1, 0.03 and G or just regular chow, increases the gut contents of bacteria reportedly µmol/kg, respectively. associated with lean phenotypes. If present in humans, this may account for All NMU analogues were potent activators of the receptors. The acylation obesity increase following sucrose replacement by HFCS. scan demonstrated that the potency of the analogues increased as the lipidation site was positioned towards the N-terminal and away from the receptor binding C-terminal octapeptide segment. All analogues signifi cantly and dose- dependently suppressed food intake with more than 50% over 18 hours as compared to vehicle whereas native NMU only resulted in a 17% reduction. The potent reduction in food intake observed after administration with the lipidated NMU analogues is considered an effect of improved

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A506 OBESITY—ANIMALCATEGORY

to reduce APP phosphorylation at T668. We also observed similar changes S 55F/45G W in tau phosphorylation by insulin and IGF-I treatment. Our results suggest Body weight [g] 441.2±42.2 438.1±40.4 456.3±61.7 InsR-induced increases in T668 phosphorylation of APP as a possible link Firmicutes phylum/Bacteroidetes phylum ratio 0.47±0.20 0.46±0.32 0.40±0.17 between obesity and cognitive impairment. Furthermore our data reveal a potential and benefi cial effect of IGF-I signaling as a therapeutic target. Bifi dobacterium genus [%×10-3] 0.52±0.6* 0.12±0.13 0.31±0.6 Supported By: University of Michigan Bacteroides uniformis species [%×10-6] 6.06±6.5*^ 0.14±0.66 0.31±0.98 Bacteroides cellulosilyticus species [%×10-3] 0.15±0.08*^ 0.09±0.1 0.1±0.12 1980-P Parabacteroides merdae species [%×10-3] 0.15±0.09*^ 0.09±0.04^ 0.05±0.03 Collinsella aerofaciens species [%×10-3] 0.04±0.12* 0 0.01±0.03 Improving Hepatic Insulin Resistance in a High-Fat-Diet-Induced Model of Type 2 Diabetes Using a Manganese Porphyrin Catalytic -6 Lactobacillus reuteri species [%×10 ] 0.39±1.2 1.7±4.2 0.19±0.9 Antioxidant Mann-Whitney U-test: *P<0.05 vs. 55F/45G; ^P<0.05 vs. W GINA M. COUDRIET, MEGHAN M. DELMASTRO-GREENWOOD, SOJIN LEE, Supported By: Polish Diabetes Association HENRY H. DONG, JON D. PIGANELLI, Pittsburgh, PA Insulin resistance is exaggerated through excessive consumption of a 1978-P fatty diet resulting in an obese state, including hepatic steatosis, oxidative P7435, a Novel, Selective, and Potent DGAT1 Inhibitor as a New stress, and chronic infl ammation. In type 2 diabetes (T2D), impaired insulin Therapeutic Option for Hyperlipidemia and Obesity signaling and resultant hepatic insulin resistance leads to hyperglycemia and AMOL DIXIT, NITIN DESHMUKH, PARIKSHIT GAIKWAD, MOHAN PATIL, JESSY long-term complications. These fi ndings led to the hypothesis that targeting ANTHONY, RAGHURAM ANUPINDI, HARENDRA JHA, MAHESH REDDY, ANAGHA secondary infl ammation resulting from oxidative stress would assuage DAMRE, AMOL GUPTE, KISHORKUMAR KADAM, SHIVAJI KANDRE, RAVI hepatic infl ammation and enhance insulin sensitivity and glucose tolerance JADHAV, TANDRA GUHA, KUMAR NEMMANI, UMAKANT BAHIRAT, MANOJA in a high fat diet (HFD) mouse model of T2D. To address this hypothesis, BRAHMA, LALIT DOSHI, PRAMOD KOLHE, YOGESH AHIRE, SOMESH SHARMA, we treated high fat-induced obese mice with subcutaneous injection of a RAJIV SHARMA, VEENA AGARWAL, Mumbai, India manganese porphyrin (MnP) (5 mg/kg) every 3 days. We showed that mice Diacylglycerol acyltransferase (DGAT) enzymes catalyze the fi nal step in treated with MnP exhibited signifi cantly decreased weight gain (p<0.5) and triglyceride synthesis pathway. DGAT1 knock-out mice have been shown whole body adiposity (p<0.5), accompanied by diminished hepatic steatosis to be resistant to diet-induced obesity with increased insulin sensitivity, characterized by decreased liver weight (p<0.01), total liver triglyceride decreased absorption of triglycerides when fed a high fat diet, resulting levels, and LDL cholesterol levels (p<0.001), when compared to mock-treated in reduced post-absorptive chylomicronemia. P7435, a novel, potent and dietary obese mice. MnP also decreased liver infl ammation by reducing selective DGAT1 inhibitor, is being developed as a potential treatment for expression of TNF. Treatment with MnP improved glucose tolerance and dyslipidemia/hyperlipidemia and obesity. insulin sensitivity as well as decreasing serum adipokine levels, including P7435 inhibits hDGAT1 with an IC50 of 22.7 nM (enzyme inhibition leptin (p<0.01), resistin, and insulin (p<0.5). Our studies show that MnP assay) having >10,000-fold selectivity over hDGAT2 (IC50 >300 µM). P7435 was capable of scavenging reactive oxygen species and suppressing showed good pharmacokinetic profi le with half life of 4-5 hours in rodents. proinfl ammatory cytokine production, thus reducing the progression of Treatment with P7435 resulted in a signifi cant reduction in body weight HFD induced insulin resistance and steatosis. Overall, MnP is a potential gain, food intake, plasma triglycerides, cholesterol and glucose in rodent therapeutic agent for impeding infl ammatory-driven T2D pathogenesis, models of obesity and dyslipidemia without causing steatorrhea. In high which may decrease complications and provide a window for lifestyle fat high cholesterol-fed hamsters, P7435 signifi cantly (p <0.01) decreased intervention between transitions from metabolic syndrome to overt T2D. plasma triglyceride (50%) and cholesterol (58%) associated with a reduction Supported By: ADA (1-12-BS-161); JDRF of 35% in food intake. P7435 also elicited a dose-related reduction in body weight gain of 7% (p <0.05) along with fat mass reduction. In high fat diet 1981-P (HFD)-fed ob/ob mice, P7435 elicited dose related reduction in food intake, Collagen VI Knockout Abrogates Obesity-related Adipose Tissue body weight gain and plasma glucose by 21, 21 and 57% (p <0.01) along Stiffening with fat mass reduction. In HFD-fed SD rat model, P7435 reduced plasma LIKANG CHIN, BOBBY MONKS, MORRIS J. BIRNBAUM, PAUL A. JANMEY, triglycerides, food intake and body weight gain by 57, 26 and 16% (p <0.01) MAKOTO FUNAKI, Philadelphia, PA, Tokushima, Japan respectively. Tissue stiffness is tightly controlled under normal conditions. Numerous In conclusion, P7435 has signifi cant effi cacy in rodent models of hyper- fi ndings - in liver fi brosis, kidney disease, and cancer - strongly suggest that lipidemia and obesity. P7435 is currently in phase I trial in USA (Clintrials.gov altered tissue mechanics is not only a symptom but also a causative factor in number - NCT01910571). disease. We previously demonstrated that adipose tissue stiffness (storage modulus) increases 2-fold in ob/ob mice compared to lean littermates. 1979-P Obesity-related adipose tissue stiffening is presumably a result of increased Insulin Resistance (InsR) Increases the Phosphorylation of APP and extracellular matrix proteins, such as collagen VI (col6). Prior work has shown Tau: Possible Connection of InsR and Cognitive Defi cit that knockout (KO) of col6 from ob/ob mice yields improved metabolism and BHUMSOO KIM, CATRINA SIMS-ROBINSON, CAREY BACKUS, SANGSU OH, EVA decreased infl ammation, despite adipocyte hypertrophy. We hypothesize L. FELDMAN, Ann Arbor, MI that col6 contributes to the adipocyte mechanical environment and thereby Diet-induced obesity is a risk factor for Alzheimer’s disease (AD). Insulin changes to col6-dependent mechanical properties may affect infl ammation. resistance (InsR) is a major feature of obesity. Impaired insulin/insulin- To investigate this hypothesis, the viscoelastic properties of adipose tissue like growth factor (IGF-I) signaling cascades due to InsR may provide a from col6-KO and -expressing mice, both lean and ob/ob, were measured link between obesity and cognitive impairment/AD. In addition to the using a multi-step stress-relaxation test over a range of physiologically- well-known pathological features of increased tau phosphorylation and relevant compressional strains. Stiffness of adipose tissues for all genotypes Abeta accumulation in AD, recent studies also highlight the critical role of increases with increasing compression. KO of col6 not only decreases the phosphorylation of amyloid precursor protein (APP) at threonine 668 (T668). stiffness of obese adipose tissue (signifi cant up to 10% compression), but Integrated In this study we examined the effects of insulin/IGF-I signaling on it does so to levels comparable to lean, abrogating obesity-related tissue POSTERS APP and tau phosphorylation and the possible contribution of T668-APP stiffening. Col6 KO decreases the loss (viscous) modulus of obese adipose Physiology/Obesity phosphorylation on obesity-induced cognitive impairment. We induced to levels similar to lean, but there are no differences amongst genotypes obesity in C57Bl/6 mice using a high fat diet (HFD) for 24 wk. Obese mice for the viscoelastic properties of stress-relaxation ratio or rate. Thus, col6 displayed signifi cant cognitive impairment at 24 wk in parallel with the contributes to obesity-related tissue stiffening and more specifi cally, to the increased tau and T668-APP phosphorylation in the cortex. We previously elastic, not viscous, properties of adipose tissue. Moreover, the difference reported that embryonic cortical neurons (eCN) develop neuronal InsR with in stiffness between adipose tissues from ob/ob and ob/ob col6 KO mice decreased insulin and IGF-I signaling following chronic insulin treatment. suggests that adipocytes are stiffness-sensitive, and this gives insight into IGF-I treatment of eCN resulted in a time-dependent decrease in T668-APP a possible mechanism by which adipocytes from ob/ob col6 KO mice have phosphorylation. Insulin also decreased APP phosphorylation but the effect improved metabolism despite exhibiting uninhibited hypertrophy. was much weaker compared to IGF-I. Chronic treatment of eCN with insulin increased Thr668-APP phosphorylation. IGF-I was able to reduce T668 phosphorylation after chronic insulin treatment; insulin itself was unable

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A507 OBESITY—ANIMALCATEGORY

1982-P 1984-P High-Fat-Diet-Induced Paternal Obesity Effects on Offspring Pheno - Role of Developmental Genes in Heterogeneity of White Adipocytes type KEVIN Y. LEE, SIEGFRIED USSAR, ANDRE KLEINRIDDERS, C. RONALD KAHN, YURIY SLYVKA, YIZHU ZHANG, ALEXIS ZONTINI, FELICIA V. NOWAK, Athens, OH Boston, MA Recent studies have shown that parental diet affects offspring phenotype. White adipose tissue (WAT) in from depots differentially infl uences whole- However, few studies have concentrated on the effects of paternal diet- body physiology. Previously we showed that these differences among WAT induced obesity on metabolic parameters in offspring. Such environmentally depots can be related to differences in expression of developmental genes, induced changes may refl ect altered gene expression due to transmitted such as T-box 15 (Tbx15), HoxC8/C9, and Shox2. In the present study, we epigenetic marks. have explored heterogeneity of WAT adipocytes within a single depot by To induce obesity 8 male C57BL/6 mice were fed a high fat diet (HFD, preparing and characterizing 94 clonal preadipocyte cell lines derived from 45% kcal fat) for 12 weeks, starting at 4 weeks of age, and mated with 8 fat pads of the Immortomouse, which expresses an interferon inducible, females fed for 12 weeks with low fat diet (LFD, 10% kcal fat) starting at the thermolabile large T antigen. Of the clonal cell lines, 51 of the 94 lines were same age. Control group offspring were obtained from 8 parental pairs both capable of differentiation into mature adipocytes. Interestingly, despite a on LFD. After weaning, all offspring were maintained on regular lab chow. high level of differentiation, ~50% of the adipocyte cell lines were resistant At the age of 20 days, 6 weeks, 6 and 12 months, body weight and length to insulin induced uptake of [3H] 2-deoxyglucose with <2-fold stimulation, were taken. Body fat was detected by nuclear magnetic resonance, serum while the remainder showed high insulin sensitivity (~9-fold stimulation). leptin measured by ELISA, insulin sensitivity tests performed and voluntary This was paralleled by differences in insulin stimulated lipogenesis as physical activity measured with metered running wheels. measured by incorporation of [14C] D-glucose into lipid. These differences Paternal HFD results in the following changes in offspring compared to were independent of fat depot of origin, but were correlated with higher the control group. Body weight is greater at 6 weeks, and 6 and 12 months in levels of HoxC8 and HoxC9 in the insulin resistant adipocytes. Expression males. Female offspring are heavier only at 12 months and this is associated of Tbx15 was also inversely correlated with oxidative metabolism of the with longer body length. Percent body fat is higher in male offspring at 6 adipocytes as assessed in a Seahorse apparatus (p<0.05). Furthermore, months. Male offspring at 6 weeks engage in more voluntary physical while Tbx15(+) preadipocytes obtained by FACS sorting from Tbx15-LacZ activity Both males and females at 6 weeks are more sensitive to insulin. knock-in mice differentiated into adipocytes that were morphologically Insulin sensitivity decreases with age in all groups of offspring. Leptin indistinguishable from Tbx15(-) adipocytes, these adipocytes exhibited increases with age in all groups of offspring but no difference is observed ~40% lower levels of expression of markers of oxidative metabolism (PGC- with type of paternal diet. 1α and cytochrome c oxidase subunit 8b), while having 30-100% increases Paternal HFD induced obesity results in phenotypic differences in fi rst in markers of glycolytic metabolism (hexokinase 2 and GAPDH). Thus, generation offspring, both males and females, although greater differences preadipocytes even within a single fat depot are heterogeneous with respect in the above parameters are observed in males. It is hypothesized that to expression of developmental genes, and these differences are linked to paternal HFD induces epigenetic changes in sperm some of which may functional differences between adipocytes, including major differences in protect against diet-induced energy imbalance in offspring. insulin sensitivity and oxidative metabolism.

1983-P 1985-P Dynamic Regulation of DNA Methylomes in Fat, Liver, and Muscle Lack of CEP19 Gene Causes Insulin Resistance with Enhanced in Obesity Insulin Signaling and Mitochondrial Function in Skeletal Muscle RUI WU, JEONG-HYEON CHOI, HANG SHI, HUIDONG SHI, BINGZHONG XUE, JUAN XIONG, FENG DONG, CYNTHIA GALINDO, MARCEL FOURCAUDOT, JEFFREY Atlanta, GA, Augusta, GA L. BARNES, LUKE NORTON, MUHAMMAD MAHROUM, RALPH A. DEFRONZO, ZVI Obesity and its associated metabolic disorders are consequences of BOROCHOWITZ, ADEL SHALATA, MUHAMMAD A. ABDUL-GHANI, San Antonio, gene and environment interactions, and one of the mechanisms whereby TX, Haifa, Israel environmental factors (eg, diets) affect gene expression is through epigenetic Both genetic background and lifestyle contribute to the pathogenesis modifi cations. Here, we have characterized the detailed DNA methylomes in of insulin resistance. Insulin resistance is closely associated with obesity fat, liver and muscle of mice fed a low fat (LF) or a high fat (HF) diet using and insulin resistant individuals manifest impaired insulin signaling and reduced representation bisulfi te sequencing (RRBS) analysis. Out of all the decreased mitochondrial ATP synthesis rate. We previously have shown that methyl-cytosines (mCG, mCHG and mCHH) analyzed, we fi nd that >90% lack of CEP19 gene results in an obese phenotype in both mice and man. The are mCG, with the remaining (<10%) being mCHG and mCHH, indicating aim of the present study is to characterize the metabolic phenotype of mice that most of the methyl-cytosines in adult fat, liver and muscle are mCGs. lacking the CEP19 gene. From mapped methyl-cytosine sites, we have identifi ed 2119 Differentially Compared to WT mice (n=12, age=4-6 months), mice lacking the CEP19 gene Methylated Regions (DMRs) in fat tissues between LF- and HF-fed mice, had 18% increase in body weight. CEP19 KO mice had normal fasting plasma with fewer DMR identifi ed in liver and muscle (686 and 233, respectively). glucose concentration (126±4 vs. 118±7 mg/dl) but 64% increase in the fasting Most of the DMRs in fat and liver (70%) and about 30% DMRs in muscle plasma insulin concentration. The 2-hour plasma glucose concentration was show increased methylation rate in HF-fed mice compared to LF-fed mice. markedly elevated in CEP19 KO mice compared to WT (405±34 vs. 219±11 Consistently, treating diet-induced obese mice with 5-aza-2’-deoxycitydine mg/dl, p=0.0002). Whole body insulin-stimulated glucose disposal measured to inhibit DNA methylation protects mice from developing obesity-induced with the euglycemic hyperinsulinemic clamp was decreased by 34% in mice insulin resistance and diabetes. DNA hypermethylation of estrogen receptor lacking CEP19 gene compared to WT. Surprisingly, despite insulin resistance, α (Esr1) is frequently associated with cancer and atherosclerosis. We fi nd insulin-stimulated muscle Akt and AS160 phosphorylation were enhanced that DNA methylation at the promoter/5’-untranslated region of Esr1 is in CEP19 KO mice compared to WT littermates. Moreover, CEP19 KO mice signifi cantly up-regulated in fat tissues of HF-fed mice compared to LF-fed manifested increased mitochondrial ATP synthesis rate (by 45% and 39% for mice. This is confi rmed by pyrosequencing. Consistent with DNA methylation complex 1 and 2 substrates, respectively, p<0.01), oxygen consumption rate status, Esr1 expression is signifi cantly down-regulated in fat tissues of HF- (by 83%, p<0.01) and muscle protein content of mitochondrial complexes 1, 3 fed mice. In summary, our data suggest that DNA methylomes are under and 4 compared to WT animals. Integrated

POSTERS dynamic regulations in fat, liver and muscle during the development of obesity Conclusion: (i) lack of CEP19 gene results in increased body weight and and its associated metabolic disorders, and inhibiting DNA methylation skeletal muscle insulin resistance; (ii) the defect in insulin-stimulated muscle Physiology/Obesity protects against obesity-induced insulin resistance. In addition, Esr1 may be glucose disposal is downstream of the insulin signaling cascade; and (iii) an epigenetically regulated target in fat tissues during the development of skeletal muscle insulin resistance in CEP19 defi cient mice is not associated obesity and its associated metabolic disorders. with impaired mitochondrial function. Supported By: ADA (7-13-BS-159); NIH (1R01DK084172, 1R01HL107500); AHA (10SDG Supported By: AHA (to M.A.G.) 3900046, 11GRNT7370080)

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A508 OBESITY—ANIMALCATEGORY

1986-P protein (GFAP), whereas expression of phospho glycogen synthase kinase 3 CDP138 Knockout Mice Display Altered Glucose Metabolism (GSK3β), synaptophysin and insulin degrading enzyme (IDE) were decreased QIONG L. ZHOU, JUN-YUAN HUANG, ZHENWEI GONG, XIANGYANG XIE, KARLA compared to brain of lean mice. In ob/ob mice, amyloid beta concentration CANDELARIA, ZHEN Y. JIANG, Boston, MA, Orlando, FL in brain was increased. Resveratrol treatment improved the expression of CDP138 is a newly identifi ed C2 Domain containing Phosphoprotein IDE, synaptophysin and GSK3β level and decreased tau phosphorylation that has been indicated as an Akt downstream target and is involved in and amyloid-beta concentration. Because tau hyperphosphorylation and insulin-regulated GLUT4 translocation and glucose transport in cultured increased amyloid beta content are associated with memory impairment, 3T3 L1 adipocytes. Here we report that mice with whole-body deletion of these fi ndings demonstrate a potential neuroprotective effect of resveratrol CDP138 have been generated and that these mice exhibited no signifi cant in brain of obese diabetic mice. difference in steady-state glucose infusion rates, tissue glucose uptakes Supported By: Auburn University (to R.J.); MWU-ORSP (to T.L.B.) or glucose appearance/disappearance compared to WT mice during 120 minutes hyperinsulinemic-euglycemic clamps under the normal chow diet 1989-P condition. However, after challenged with a high fat diet (HFD, 60% fat) for 6 weeks, CDP138-/- mice gained signifi cantly more weight than WT mice. HFD decreased the steady-state glucose infusion rate in CDP138- WITHDRAWN /- mice to an average rate of 27mg/kg/min, while WT mice have the average rate of 36mg/kg/min. Glucose uptakes in skeletal muscles, white adipose tissues and heart were signifi cantly slower in CDP138-/- mice than in WT mice. Under HFD condition, both basal and clamped glucose turn over rates were lower in CDP138-/- mice than in WT mice. Glucose and insulin tolerance tests also indicated that HFD fed CDP138-/- mice were less insulin sensitive and more glucose intolerant. In addition, deletion of CDP138 signifi cantly inhibited insulin-induced GLUT4 translocation in differentiated mouse embryonic fi broblast (MEF) cells. Taken together, our data shows that CDP138 plays an important role in glucose metabolism in mice. Supported By: ADA (7-11-BS-72); NIDDK

1987-P A Novel Carbonyl Scavenging Compound Mitigates Mitochondrial Dys- function and Cardiac Remodeling Caused by Diet-induced Obesity: Preclinical Studies using Wild-type and GPx4-defi cient Mice LALAGE A. KATUNGA, HANNAH E. MITCHELL, GIANCARLO ALDINI, ETHAN J. ANDER- SON, Greenville, NC, Milan, Italy Oxidation of polyunsaturated fatty acids (PUFAs) yields lipid peroxidation end products (LPPs), including α,β-unsaturated aldehydes, which are thought to con- tribute to a number of chronic diseases associated with obesity. However, the precise role of these LPPs and the mechanisms involved remain unknown. Gluta- thione Peroxidase 4 (GPx4) is an antioxidant enzyme with both cytosolic and mito- chondrial isoforms, and is unique in that it specifi cally neutralizes lipid peroxides. Here, we fed mice defi cient in GPx4 (GPx4+/-) and their wild type (WT) littermates a high fat high sucrose (HFHS) diet for 16 weeks in order to examine the role of LPPs in cardiac remodeling that occurs with obesity. To determine whether scav- enging LPPs can mitigate this remodeling we treated separate GPX4+/- and WT mice with FL-926-16, a novel carbonyl scavenging compound. FL-926-16 interven- tion (40mg/kg) began after 8 weeks on the HFHS diet. At end of the study, cardiac structure/function was assessed, along with cardiac mitochondrial function in permeabilized ventricular myofi bers. A number of whole body metabolic param- eters were also examined. GPx4 – HFHS mice were substantially more insulin resistant compared to WT- HFHS. FL-926-16 increased glucose tolerance by 60% 1990-P and 22% in WT and GPx4+/- respectively. FL-926-16 mitigated HFHS-diet induced Diet Quality, Not Time of Ingestion, Is Associated with Early cardiac hypertrophy in WT only. FL-926-16 improved mitochondrial effi ciency – the Development of Increased Cardiometabolic Risk in Rats ratio of ATP production to O2 consumption by ~2 fold, and restored mitochondrial ISRAEL ORTA, NATASHA SOARES, LASHANNI BUTLER, JOYCE M. RICHEY, Los fatty acid oxidation rates to levels comparable to control diet in both WT-HFHS Angeles, CA and GPx4-HFHS groups. These fi ndings illustrate a critical role of LPPs in the pro- A great deal of data demonstrate that clock disturbances lead to disease, gression of cardiomyopathy caused by HFHS diet-induced obesity, and provide consistent with increased metabolic risk in shift workers, persons with sleep evidence that novel therapies directed at scavenging LPPs have the potential to deprivation and night eating syndrome. However, few studies in animals or mitigate the progression of this disease. humans have examined the role of feeding times on the development of obesity and subsequent cardiometabolic (CMet) risk. Thus, we undertook 1988-P the present study to determine the effects of fat feeding during the inactive Resveratrol Protects Proteins Associated with Memory Loss in (Day-D) vs. active (Night-N) phases on CMet risk. Towards this end, male Integrated Obese Diabetic Mice Wistar rats were restricted to 60% high fat feeding (HF) or control chow (CC) POSTERS

SHRADDHA D. REGE, GEETHA THANGIAH, TOM L. BRODERICK, RAMESH JEGA- during either Active (lights off: 6 pm to 6am) or Inactive phases (lights on: 6am Physiology/Obesity NATHAN, Auburn, AL, Montgomery, AL, Glendale, AZ to 6pm) for a 3 week period. A total of 4 groups were analyzed (n=7 for each Resveratrol is a polyphenolic phytoalexin and known to exert anti- group); (1) HF-N; (2) HF-D; (3) CC-N and (4) CC-D. Body weight (BW) and food diabetic and anti-infl ammatory actions. Several studies have demonstrated intake were measured bi-weekly and daily, respectively. Vascular function a link between the obese diabetic state and Alzheimer’s disease. The was assessed in thoracic aortic ring preparations and glucose metabolism present study evaluated the neuroprotective action of resveratrol in the was measured using an intravenous glucose tolerance test (IVGTT, n=3 for ob/ob mouse, a model of severe obesity and diabetes. Resveratrol was each group). BW increased signifi cantly in all 4 groups, with the greatest administered orally at the dose of 25 mg kg-1 body weight daily for 3 weeks energy intake and weight gain in CC-N (p<.05), despite epididymal (EPI) and to male lean and ob/ob mice. Resveratrol did not show any effect on body retroperitoneal (RET) fat depots being substantially greater in HF-N and HF-D weight or blood glucose levels in ob/ob mice. Formic acid fractions in brain of groups. EPI and RET fat were 65% and 100%, respectively, greater in HF ob/ob mice were found to have increased expression of tau, phosphorylated compared to CC groups (p<.01). Vascular dysfunction, evidenced by reduced forms of tau (CP-13, S202/205; PHF1, S396/404), and glial fi brillary acidic responsiveness to the vasoconstrictor, phenylephrine was observed only in

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A509 OBESITY—HUMANCATEGORY

HF-D and HF-N groups (p<.01). Similarly, there was a strong tendency (p<.09) 1993-P for both HF groups to exhibit decreased glucose tolerance compared to CC Validation of Nonhuman Primate Adiposity Indices with Body Com- groups. However, differences were not detected between active versus po sition in Obesity With and Without Naturally-occurring Type 2 inactive groups with similar diets. These data support the notion that quality Diabetes of diet and not timing of food consumption is the predominant contributor STACEY L. CONARELLO, ELLEN H. LINDEN, JENNIFER D. NEWCOMB, BARBARA of increased cardiometabolic risk, particularly during the early stages of C. HANSEN, EFFIE TOZZO, Rahway, NJ, Tampa, FL development. Overweight/obese nonhuman primates (NHPs) are considered the ideal animal model for studying the underlying mechanisms of type 2 diabetes 1991-P under constant environmental conditions, and for studying potential therapies Metformin Alters Energy Balance by Reducing Both Food Intake and for their likely applicability to humans. NHPs progress through the same Energy Expenditure phases as humans, and develop varying degrees of obesity prior to the onset THAÏS ROUQUET, CLÉMENT PIERRE, MICHEL DALLAPORTA, STÉPHANIE GAIGÉ, of overt T2DM. Body composition, determined by DXA, is the gold standard JULIEN ROUX, JEAN-DENIS TROADEC, BRUNO BARIOHAY, BRUNO LEBRUN, for assessment of adiposity (% body fat), and, as such, is useful in examining Marseille, France the interactions of increasing obesity in the prediabetes (PreDM) phase. DXA Since the late 1950s, metformin, a biguanide derivative, is widely used is, however, not available to most primate colonies and is not applicable to to treat hyperglycemia in individuals with type 2 diabetes. It has been screening of large colonies for risk of diabetes. In order to validate potential shown that metformin acts primarily on hepatocytes to reduce glucose indices of adiposity in NHPs, we have therefore studied 236 adult rhesus output, and secondarily in peripheral tissues to increase glucose uptake. monkeys (167 males) (ages 10 to 25 yrs) ranging in body weights from 4 to Recent evidences indicate that metformin also exerts an anorectic effect. 30 kg. Overt type 2 diabetes had spontaneously developed in 58 of these To investigate the mechanisms of action of metformin on food intake and monkeys, and metabolic syndrome or preDM was evident in 73 animals, energy metabolism, we submitted C57BL/6J and db/db adult mice to an with males and females similarly distributed across these groups. In males, acute treatment with either metformin (per os, 300mg/kg) or vehicle and % body fat by DXA, abdominal circumference, body mass index (BMI), and measured food intake and energy expenditure for the subsequent 48h in Body Adiposity Index (BAI; abdominal circumference/height) all were similarly metabolic cages (Physiocages system, Panlab). We showed that metformin elevated in both preDM and overtly DM monkeys compared to normal animals. induced a signifi cant decrease in cumulative food intake in both C57BL/6J In males only, there was also an increase in % body fat with age, separate and db/db mice. This anorectic effect lasted 24h and was more pronounced from metabolic status. Females and males show signifi cant sexual dimorphism in leptin-receptor defi cient db/db mice compared to C57BL/6J mice. Meal- and differences in body conformity and composition with age and metabolic pattern analysis revealed that metformin caused a signifi cant decrease in status. The females showed the increase in abdominal circumference between meal size without affecting meal frequency, suggesting that its anorectic normal and preDMs, but did not show this increase in overt DM, and unlike effects do not result from a non-specifi c adverse effect. However, indirect the males, the females showed no relationship of % body fat or BMI to age calorimetry analysis revealed that metformin transiently decreased VO2 or metabolic status. In both males and females, BAI was highly correlated to and energy expenditure in C57BL/6J mice but not in db/db mice. We also both % body fat and to BMI, and thus represents the optimal and most readily investigated the impact of metformin on cerebral structures involved in available new method for estimating NHP body adiposity. food intake control, using c-Fos protein as a marker of neuronal activation. Supported By: NIH (HHSN2632008000022C); Merck & Co., Inc. Metformin treatment resulted in a strong neuronal activation in the Nucleus Tractus solitarii of the Brainstem in both genotypes, whereas no activation was detected in the Arcuate and Paraventricular Nuclei of the Hypothalamus. OBESITY—HUMAN A more thorough c-Fos cartography and phenotyping of activated neurons is underway. Taken together, our results support the view that in addition to its peripheral effects on glucose metabolism, metformin also interacts with Guided Audio Tour: New Insights in Human Obesity Biology (Posters: the central control of food intake and energy expenditure in both leptin- 1994-P to 2000-P), see page 17. dependent and leptin-independent manners. & 1994-P 1992-P Genetic Effects on the Distribution of Abdominal Adipose Tissue TM4(UBAC2) Improves db/db Mouse Status by Regulating Nur77- Compartments and the Presence of Nonalcoholic Fatty Liver IKKβ-NF-κB Pathway Disease: A Twin Study REMING HU, REMING HU, YINTAO LI, LILI CHEN, XIAOCHENG FENG, KUANPING ADAM L. JERMENDY, ZSOFIA DROBNI, CSILLA CELENG, TAMAS HORVATH, YE, YING HUANG, QIN LI, Shanghai, China FERENC I. SUHAI, ADAM D. TARNOKI, BELA MERKELY, PAL MAUROVICH- TM4 (UBAC2) protein is a membrane protein containing four transmembrane HORVAT, GYORGY JERMENDY, Budapest, Hungary domin, which contains a PKC phosphorylation site of serine residue in the In patients with diabetes and/or obesity, accumulation of abdominal 89th amimo acid and a ubiquitin associated domain (UBA). We screen the adipose tissue and non-alcoholic fatty liver disease (NAFLD) are linked candidate interacting partners of TM4 protein using yeast two-hybrid to increased cardiometabolic risk. Little is known about the genetic and system, immunoprecipitation and pull down experiment, we found NR4A1 environmental effects on the distribution of the abdominal adipose tissue protein was probably a candidate partner of TM4 protein. Sliencing the TM4 compartments and those of NAFLD. The aim of the study was to assess the by siRNA and lentivirus shRNA leads to upregulate the protein levels of magnitude of genetic impact on the size of various abdominal adipose tissue Nur77 (45%) and its down stream molecule IKKβ (20%) and NF-κB p65. compartments and the presence of NAFLD within a sample of twin pairs. To better explore the physiologic function of TM4, we generated the In this classical twin study, 82 twin subjects (22 monozygotic [MZ] pairs with mouse model of TM4 gene knockout by gene trapping. Mice were fed 60% age of 59±9 years and 19 dizygotic [DZ] pairs with age of 55±9 years) were kcal% HFD for 4 months, the TM4-/- mice became morbidly obese. The weight, involved. The twin pairs were investigated with a 256-slice CT-scanner (Brilliance blood pressure, fasting and 2 hours’ glucose of TM4 knockout mouse were iCT, Philips Healthcare, Best, The Netherlands). For each patient CT-based waist signifi cantly higher than those in the wild type. circumference, abdominal subcutaneous (SAT) and visceral adipose tissue (VAT) Integrated POSTERS To study the impact of TM4 over-expression in db/db mouse, thirty db/ quantifi cation was performed. Liver and spleen attenuation was determined db mice aged two weeks were treated with saline, empty lentivirus vector by calculating the average of three 300 mm2 ROIs. NAFLD was characterized Physiology/Obesity and TM4 (m) lentivirus vector respectively. After 10 weeks of TM4 lentivirus by attenuation ratios (CTL/S) and liver-to-spleen attenuation differences (CTL-S). infection, the weight (5%, P<0.01), fasting plasma glucose (7%, P<0.01), Concordance between MZ and DZ pairs were assessed by Pearson correlations. HOMA-IR (3%, P<0.05), systolic BP (4%, P<0.05), TG (3%, P<0.05), and LDL-c Rough heritability was calculated according to the Falconer-method. (4%, P<0.01) were lower than those in the control group. On IPGTT, the The median SAT was 193.0 [IQR:144.1-250.3] cm2, whereas median VAT glucose of TM4 lentivirus treated mice in 60 minutes was lower by 18% than was 128.5 [IQR:77.2-180.5] cm2. Relatively high heritability was found in SAT the control. Nur77, IKKβ, and phosphorylated NF-κB protein expression in (0.79) and VAT (0.74), while the derived parameters regarding NAFLD (CTL/S the skeletal muscle, liver, as well as epididymis fat tissues were signifi cantly and CTL-S) showed lower genetic dependency (both 0.12). decreased. In this classical twin study we were able to show that the distribution In conclusion, TM4 may decrease the body weight, blood pressure and of abdominal fat compartments has relatively strong heritability, while the fasting glucose of diabetic mice by regulating Nur77-IKKβ-NF-κB Pathway. presence of NAFLD is less infl uenced by genetic effects. Supported By: NSFC (81030014) Supported By: FESD

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A510 OBESITY—HUMANCATEGORY

& 1995-P & 1997-P Appetite-Regulating Effect of Gut-Derived Clusterin/Apolipoprotein J Plasma Free Fatty Acids Modulate Chemokine Ligand 10 Gene JONG HAN CHOI, JI HEE YU, CHURL NAMKOONG, SO YOUNG GIL, KYONG HAN Expression and Plasma Concentration in Humans PARK, MIN-SEON KIM, Seoul, Republic of Korea, Chuncheon, Republic of Korea HANYU LIANG, HELEN LUM, JOSE DE JESUS GARDUNO GARCIA, NICOLAS Clusterin (also called apolipoprotein J) is a sulfated glycoprotein implicated MUSI, San Antonio, TX in neurodegeneration, lipid transport, immune modulation, and cancer Evidence derived primarily from cell culture and animal studies suggest biology. Our recent work has shown that clusterin exerts anorexigenic effect that free fatty acids (FFA) are involved in the infl ammatory state and insulin in the hypothalamus by activating hypothalamic leptin signalling pathway resistance of obesity and T2DM. However, the infl ammatory mediators and acts as appetite regulating hormone actions. of FFA-induced insulin resistance in human subjects have not been fully Immunohistochemistry of clusterin and double immunofl uorescence characterized. To identify chemokines regulated by FFA in humans, 14 normal staining of clusterin and enteroendocrine cell marker chromogranin and glucose-tolerant subjects (age=39±3 y, BMI=23.5±0.7 kg/m2, FFA=355±33 PGP9.5 were performed in mouse stomach and rat nodosum ganglion. umol/l) received a 48 h intralipid or saline infusion (30 ml/h) on two different Clusterin peptide was administered intraperitonally alone or together with occasions, followed by peripheral monocyte qPCR with an “infl ammation” cholecystokinin (CCK) to examine the effect on food intake and body weight. panel array of 84 genes. Of all the chemokine genes evaluated, the mRNA Clusterin peptide were in vagotomised mice or sham-operated mice. level of CXCL10, a chemokine that attracts monocytes/macrophages, T Clusterin was also abundantly expressed in the epithelial cells in human cells, NK cells, and dendritic cells, was upregulated the most by FFA (1.7- and mouse stomach mucosa. Moreover, clusterin also expressed in mucosa fold, P<0.05). In line with this fi nding, lipid infusion, but not saline, increased endocrine cells expressing chromogranin and coexpressed with ghrelin in plasma CXCL10 concentration by 63% (137 ± 31 vs. 223 ± 47 pg/ml; P<0.05). submucosal ganglionic neurons. Single intraperitoneal (IP) administration After observing that lipid infusion upregulates CXCL10 gene expression of clusterin signifi cantly reduced fasting- and ghrelin-induced hyperphagia. and plasma concentration in lean subjects, we tested whether lowering Conversely, coinjection of clusterin at subeffective dose prolonged the plasma FFA with the antilipolytic agent acipimox (250 mg four times/day anorexigenic effect of CCK. Clusterin was also highly expressed in the for 7 days) would reduce CXCL10 in obese nondiabetic (n=9, age=48±3 nodosa ganglion, a sensory ganglion of the vagus nerve. Furthermore, the y, BMI=31±1, FFA=673±70 µmol/L,) and obese T2DM (n=11, age=48±3, feeding effect of clusterin was almost abrogated in vagotomized mice. BMI=34±1, FFA=812±135, HbA1c=7.7±0.6) subjects. Both obese nondiabetic Consistently, IP administration of clusterin induced c-fos expression in the (133%; P<0.05) and T2DM (79%; P=0.1) subjects had higher plasma CXCL10 solitary tract nucleus and the hypothalamic paraventricular nucleus, which concentration compared to the lean group. Moreover, acipimox, which is known for the nerve conduction route of gut-derived appetite regulating lowered plasma FFA by ~50%, also decreased plasma CXCL10 levels in hormones. both obese nondiabetic (16%; P=0.05) and T2DM (46%; P<0.05) subjects. These fi ndings collectively suggest that clusterin may function as a gut- Summary/Conclusions: (i) Insulin resistant subjects (obese and T2DM) have derived hormone that may transfer satiety signals from the gut to the brain increased plasma CXCL10 concentrations; (ii) CXC10 expression and plasma through the vagus nerve. level is modulated by systemic FFA availability; (iii) CXCL-10 may be involved Supported By: NRF in the infl ammatory state and insulin resistance of obesity and T2DM. Supported By: R01DK080157, R01DK089229, K23AG030979, POAG013319 & 1996-P Insulin Sensitisation Normalises the Human Brain Response to High & 1998-P Calorie Visual Food Cues After Meal Ingestion in Insulin Resistant Gremlin-1, a Secreted BMP Antagonist by Human Preadipocytes, Men Modifi es Beige Adipogenesis YEE S. CHEAH, SARAH LEE, BULA M. WILSON, ANDREW PERNET, LUCY BIRGIT GUSTAFSON, HAMID SALARI, JOSEPH GRIMSBY, CRISTINA RONDINONE, DIAMOND, CHRIS CHEYETTE, ANITA BECKWITH, LAURENCE J. REED, MICHAEL ULF SMITH, Gothenburg, Sweden, Gaithersburg, MD J. BRAMMER, FERNANDO O. ZELAYA, STEPHANIE A. AMIEL, London, United BMP4 and 7 are important regulators of mesenchymal stem cell Kingdom commitment, including white and brown adipogenesis. However, BMP Altered brain responses to meal ingestion and food cues have been signaling and effects are under the regulation of a number of endogenous demonstrated in obesity and diabetes, conditions associated with insulin BMP antagonists. We here focus on Gremlin 1 (GREM1), which we found resistance. We investigated the impact of systemic insulin sensitization to inhibit BMP4 and 7 pSMAD signaling and to be highly expressed in on central responses to food cues in non-obese insulin resistant (IR) men human adipose tissue. GREM1 mRNA levels were positively correlated with using Blood Oxygenation Level Dependent (BOLD) MRI. Fourteen healthy subcutaneous adipose cell size and BMI of the donors. Thus, GREM1 may right-handed IR (HOMA2-IR 1.7±0.3, BMI 25.9±2.5 kg/m2) and 16 insulin be an important contributor to the development of hypertrophic obesity sensitive (IS) men (HOMA2-IR 0.5±0.1, BMI 24.2±1.9 kg/m2) matched and insulin resistance. GREM1 mRNA levels in human subcutaneous for age (33.9±10.6 vs. 32.1±5.8 years p = 0.57) and HbA1c (5.5±0.3% vs. preadipocytes undergoing differentiation were rapidly reduced (around 4h 5.3±0.2% p = 0.06) were studied twice in random order after an overnight following induction of differentiation) and with a time-course similar to the fast. Regional brain responses to foods and non-food objects compared to PPARγ activation. Furthermore, GREM1 protein was secreted by the cells blurred images presented in a block design were measured in a 1.5T scanner and secretion reached steady-state after 6 days. Secretion of GREM1 by 14 mins after subjects consumed 50 ml water (fasted) or 630 kcal mixed fully differentiated adipocytes was negatively correlated with PPARγ meal (fed). IR then received 3 months insulin sensitization therapy (2g/ activation and positively correlated with the adipose cell size of the donors. day metformin and lifestyle advice), which improved HbA1c (5.3±0.3% p = We then examined the effect of silencing GREM1 during white adipogenesis. 0.01) but not BMI (25.5±2.5 kg/m2 p = 0.2) before studies were repeated, Silencing GREM1 did not enhance overall adipogenesis but had a marked during which fullness in fasted and fed states increased (main effect p = positive effect (4-5-fold, p<0.01) on the induction of the well-established 0.02). IS showed a reduction in food image evoked deactivation after the beige adipogenesis marker TMEM26 while no effect was seen on the brown meal (whole brain cluster wise p < 0.05, voxel wise p < 0.001 uncorrected) adipose cell markers UCP1 or PRDM16. Addition of BMP4 or 7 to the cells in precuneus, superior temporal gyrus (STG) and anterior cingulate-medial exerted no further effect following GREM1 silencing. Furthermore, silencing

frontal gyrus (ACC/MFG), the latter driven by responses to high calorie food GREM1 in preadipocytes without induction of white adipogenesis also Integrated POSTERS images. This was not observed in IR at baseline but after treatment there increased TMEM26 mRNA levels and this was further enhanced (2-fold) by was a trend towards meal-induced diminution of precuneus (p = 0.09) and the addition of BMP4. GREM1 is a highly expressed and secreted inhibitor Physiology/Obesity STG (p = 0.09) deactivation which was signifi cant in ACC/MFG (p = 0.01). of BMP4 and 7 signaling and related to both hypertrophic obesity and BMI. These data suggest that in systemic insulin resistance the diminished Silencing GREM1 promotes a beige phenotype of human preadipocytes. modulatory effects of meal ingestion on responses to further food stimuli Thus, GREM1 is a novel and interesting target in human adipocytes in order in brain networks involved in reward and appetite regulation may normalize to promote a beige phenotype, which may help prevent obesity and insulin with insulin sensitization therapy and may be a potential target for obesity resistance. and diabetes prevention strategies. Supported By: King’s College; Diabetes UK; GSTT

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A511 OBESITY—HUMANCATEGORY

Guided Audio Tour: In Vivo Metabolism in Human Obesity (Posters: 2001-P & 1999-P to 2007-P), see page 17. Cerebrospinal Fluid Fructose Levels Are Signifi cantly Elevated Compared to Plasma Fructose Levels in Pregnant Women & 2001-P JANICE J. HWANG, ANDREA JOHNSON, RENATA BELFORT DE AGUIAR, DENIS Different Contributions of Human Brown Adipose Tissue to Drug SNEGOVSKIKH, GARY CLINE, CHRISTINA HAN, ROBERT S. SHERWIN, New and Cold Induced Thermogenesis Haven, CT AARON M. CYPESS, LAUREN S. WEINER, GERALD M. KOLODNY, Boston, MA In humans, fructose ingestion has differential effects on brain activity Acute mild cold exposure and β3-adrenergic receptor (AR) agonists can compared to glucose. Furthermore, unlike glucose, central infusion of each stimulate brown adipose tissue (BAT) activity and increase energy fructose is known to promote feeding in rodents. Despite evidence that expenditure. However, it is not known what human BAT’s proportional increased fructose consumption plays a role in the rising incidence of contribution is to whole-body thermogenesis. We studied 12 healthy obesity, type 2 and gestational diabetes, little is known about the central male volunteers (age 22.2 ± 0.6 y, BMI 22.7 ± 0.5 kg/m2) three times each: nervous system (CNS) metabolism of fructose, particularly in pregnancy they were given the oral β3-AR agonist mirabegron (200 mg), placebo, or where there are dramatic shifts in maternal energy homeostasis to meet the treated for 1 hour with cold exposure at 14°C using a cooling vest. Blood increased energy demands of the fetus. was drawn to measure plasma metabolites, and then subjects were In this study, we assessed CNS fructose metabolism during pregnancy administered [18F]-fl uorodeoxyglucose (FDG). One hour later, BAT activity in 16 healthy non-diabetic women (6 lean: pre-pregnancy BMI 22.7±0.6 kg/ was determined via PET/CT. Vital signs were measured throughout the m2, age 33.5±2.2 years; 10 overweight/obese: pre-pregnancy BMI 40.4±5.6 study, and resting metabolic rate (RMR) was assessed before and after each kg/m2, age 31.6±2.0 years) with uncomplicated singleton pregnancies intervention. Compared to placebo, BAT activity was higher in response to undergoing elective cesarean section at term. Fasting maternal plasma cold exposure (P<0.001) and treatment with the β3-AR agonist (P<0.002). and cerebrospinal fl uid (CSF) were collected for measurement of glucose, RMR increased in response to cold by 128 kcal/d (+8%; P<0.005) and to fructose and sorbitol (using gas chromatography-liquid mass spectrometry) the β3-AR agonist by 203 kcal/d (+13%; P<0.002), and these changes were in the absence of labor and prior to spinal anesthesia. Strikingly, CSF fructose correlated among the subjects (r=0.71). However, the physiological effects levels were 20-25 fold greater than plasma levels (mean CSF 3.27±0.31 mg/dl of the two interventions were different. Heart rate decreased in response vs. plasma 0.14±0.03 mg/dl). Thus, while fructose was negligible compared to mild cold (-4 bpm; P=0.05), but increased with β3-AR agonist (+14 bpm; to glucose in plasma, it was ~7% of the CSF glucose level. CSF sorbitol P<0.001). Univariate regression analysis showed that the predictors of cold- levels were also >6-fold greater than plasma levels (mean CSF 2.92±0.3 vs. induced thermogenesis were change in core temperature (P<0.001), total T3 plasma 0.45±0.1 mg/dl). There were no differences in CSF or plasma levels (P<0.03), and white fat glucose uptake (P<0.05). In contrast, the predictors of fructose or sorbitol between lean and obese women. of the β3-AR agonist-induced thermogenesis were BAT glucose uptake This study shows, for the fi rst time, that CSF fructose and sorbitol levels (P<0.007), body fat percentage (P<0.003), and insulin secretion (P<0.05). are markedly higher than plasma levels amongst pregnant women. As both These data emphasize that an accurate determination of BAT thermogenesis fructose and sorbitol were elevated, endogenous CNS production of fructose should incorporate several functional measures, including tissue substrate likely occurred via the polyol pathway. These data indicate the presence of utilization and oxygen consumption. Accordingly, although cold exposure much higher fructose levels in the CNS than previously thought which may and β3-AR agonists both increase BAT glucose uptake and RMR, their play a role in pregnancy-related weight gain. mechanisms of thermogenesis are markedly different. Supported By: EFF Supported By: NIDDK

& 2000-P & 2002-P Metagenomic Analysis of the Human Gut Microbiome Pre- and The Healthy Obese Are Alive and Well: Reduced Risk of Diabetes, Post-surgical Bariatric Intervention in Type 2 Diabetics with Morbid CHD, and Mortality Obesity: Signifi cant Postoperative Microbiome Modifi cation and FANGJIAN GUO, W. TIMOTHY GARVEY, Birmingham, AL Correlation with Infl ammatory and Metabolic Parameters Obesity is a chronic disease that affects 35% of U.S. adults. There has VIRGINIA KAMVISSI, YOUW QIN, HUANZI ZHONG, JULIO LICINIO, MA-LI been an escalating debate as to whether a proportion of obese individuals WONG, AM XU, TRIANTAFYLLOS CHAVAKIS, TOBIAS D. BORNSTEIN, MONIKA are free of cardiometabolic disease, termed the ‘metabolically healthy obese’ EHRHART-BORNSTEIN, VALERIA LAMOUNIER-ZEPTER, TOBIAS LOHMANN, (HO). Here, we assessed prevalence of HO in U.S. adults, and its association TILLMANN WOLF, STEFAN R. BORNSTEIN, JUERGEN GRAESSLER, Dresden, with incident diabetes, CHD, stroke, and mortality using data from 3 Germany, Shenzhen, China, Canberra, Australia, Hong Kong, China large cohorts, CARDIA, ARIC, and NHANES. HO met all 3 indices: (i) blood Roux-en-Y gastric bypass (RYGB) is increasingly becoming one of the pressure (untreated systolic <130 mmHg and diastolic <85); (ii) blood glucose leading therapies for long-term treatment of morbid obesity and type 2 (untreated fasting <100 mg/dl or HbA1c <5.7%); (iii) blood lipids (untreated diabetes mellitus (T2D). The precise effects and pathogenetic consequences triglycerides <150 mg/dl, total cholesterol <240 mg/dl, and HDL ≥40 mg/dl in of RYGB on metabolism and gut microbiome are poorly understood. The aim men and ≥50 in women). Unhealthy obese (UHO) crossed the risk threshold of this study was to characterise those effects before and 3 months after for all 3 criteria. First, subjects who were obese at baseline remained obese; RYGB with metagenomic sequencing. Subjects suffered from morbid obesity in CARDIA young adults (mean 25y) only 1.5% became lean at year 20, and (BMI>40 kg/m²) and T2D. We identifi ed 1,061 species, 729 genera, 44 phyla in older adults in ARIC (mean 54 y) only 0.4% became lean after 10 years. and 5127 KOs (KEGG Orthology). The overall metagenomic RYGB-induced Secondly, the percentage of HO adults was substantial; in NHANES the % shift was characterised by a reduction of Firmicutes and Bacteroidetes HO remained fairly constant averaging 16% from 1988-2012. Thirdly, risk and an increase of Proteobacteria. Species were then gathered into 2 factors were more predictive of outcomes than obesity. During 18.7 years common components. Component 1 consisted of species which were mainly follow-up in ARIC, compared with HO, the multivariable adjusted hazard associated with BMI and C-reactive protein and was characterised by an ratio in UHO was increased for diabetes (5.6, 95% confi dence interval, CI increase of Proteobacterium Enterobacter cancerogenus and a decrease 4.1-7.5), CHD (6.6, CI 3.4-12.8), stroke (5.2, CI 2.1-12.8), and mortality (2.6, CI of Firmicutes Faecalibacterium prausnitzii (FP) and Coprococcus comes. 1.9-3.6). Among healthy subjects, HRs were similar among HO, overweight,

Integrated Functional analysis of carbohydrate metabolism by KO revealed signifi cant POSTERS and lean for CHD, stroke, and mortality, but were decreased for diabetes effects in 13 KOs assigned to phosphotransferase system. Spearmen’s Rank in lean (0.5) compared with HO. Among unhealthy, HRs for diabetes, CHD, Physiology/Obesity correlation indicated an association of 10 species with plasma total- or stroke, and mortality were comparable among UHO, overweight, and lean. LDL-cholesterol, 5 species with triglycerides. FP was directly correlated to In conclusion, (i) obese people rarely become lean; (ii) percentage of HO has fasting blood glucose. These results provide the fi rst clinical demonstration been stable in the U.S. at ~16%; (iii) HO have dramatically lower risks for of a profound and highly specifi c intra-individual alteration of gut microbial diabetes, CHD, stroke, and mortality compared with unhealthy subjects composition. A correlation of microbiome composition and gene function with whether obese, overweight, or lean. Metabolic syndrome risk factors confer an improvement in metabolic and infl ammatory parameters is clearly shown. much higher risk of diabetes and CVD than obesity per se. This provides a better understanding of the effects of bariatric surgery and Supported By: U.S. Dept. of Veterans Affairs; University of Alabama, Birmingham allows for future exploration of diagnostic and therapeutic strategies based (P30-DK079626); NIH (DK-038765, DK-083562) on metagenomic sequencing of the human gut microbiome.

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A512 OBESITY—HUMANCATEGORY

up at 12 and 24 months (m). T2D remission was defi ned as HbA1c <6.5% & 2003-P without anti-diabetic medication. Indices of insulin secretion and sensitivity Receiver Operating Characteristic (ROC) Curve Analysis of Sex were calculated from plasma glucose, insulin and C-peptide during a 2 h Hormone Binding Globulin as a Marker of Categories of Glucose mixed-meal tolerance test. Body fat (DXA), incretins, and adipokines were Intolerance and Undiagnosed Diabetes in First-Degree Relatives also assessed. Bariatric surgery-induced 40% and 27% T2D remission (FDR) of Type 2 Diabetic Patients rates at 12 and 24m, with RYGB yielding higher remission than SG at 12m NABILLA ABDELLA, OLUSEGUN MOJIMINIYI, Safat, Kuwait, Kuwait, Kuwait (P<0.05). At baseline, non-remitters were older, had longer T2D duration, This study explores the associations of sex hormone binding globulin and higher leptin than remitters (P≤0.05), despite similar BMIs. After 12 (SHBG) with categories of glucose intolerance and undiagnosed diabetes and 24m, weight/fat loss, insulin secretion (C-peptide0-120/glucose0-120), in fi rst degree relatives (FDR) of T2DM patients. BMI, waist (WC), Hip insulin sensitivity (Matsuda Index), and β-cell function (C-peptide0-120/ circumference and waist:hip ratio (WHR)), fasting lipids, glucose, C-peptide, glucose0-120 x Matsuda index) improved more in remitters than non- insulin, adiponectin, SHBG, estradiol (E2), testosterone (T), androstenedione remitters (P<0.05). Incretins were unrelated to remission, but compared (AND), DHEAS, high-sensitivity CRP (hsCRP) and alanine aminotransferase to non-remitters, adiponectin levels rose more in remitters at 12 (0.73[- (ALT) were measured in 141FDR. Homeostasis model assessment-estimated 0.08,37.1] vs. 3.2[2.5,5.5] ng/ml, P<0.05) and 24m (-0.1[-2.2,0.9] vs. 2.9[1.7,4.0] insulin resistance (HOMA-IR), beta cell function (%B), insulin sensitivity ng/ml, P<0.01). Baseline adiponectin predicted lower HbA1c at 12 and 24m (%S) and Free androgen index (FAI) were calculated. Categories of glucose (R=0.34, P=0.04 and R=0.33, P=0.04). Elevated adiponectin correlated with intolerance and diagnosis of diabetes were defi ned based on fasting glucose decreased android body fat (R=-0.34, P<0.04) and weight (R=-0.32, P=0.05) and/or HbA1c (ADA criteria). 82 subjects were normoglycemic; 40 had at 12 and 24 m. Higher adiponectin was also linked to enhanced β-cell impaired fasting glucose and 19 had undiagnosed diabetes. SHBG showed function at 12 and 24 m (R=0.41, P<0.01 and R=0.47, P<0.01). Thus, smaller signifi cant positive correlations with adiponectin (r=0.35), %S (0.33) and rises in adiponectin depict T2D non-remission 2 years after bariatric surgery. HDL-C (r = 0.45) and signifi cant negative correlations with BMI (r=-0.39), WC Therapies targeting greater weight loss and/or raising adiponectin may be (r = -0.35), WHR (r = -0.62), T (r = - 0.35), FAI (r = -0.72), DHEAS (r= -0.26), needed after bariatric surgery for higher T2D remission rates. C-Peptide (r = -0.30), insulin (r = -0.), %B (r=-0.38), HOMA-IR (-0.39), ALT (r=- Supported By: ADA (1-11-26 CT); Ethicon Endo-Surgery, Inc.; 1UL1RR024989; NIH 0.39), Triglycerides (r = - 0.32) and HbA1c (r=-0.22). After partial correlation (R01DK089547) analysis correcting for BMI, only correlations with WHR, FAI, TG and HDL-C remained signifi cant. SHBG decreased stepwise with worsening categories & 2006-P of glucose intolerance in females but not in males whereas FAI decreased Bariatric Surgery Induced Weight Loss and Systemic Glucose stepwise with worsening categories in males only. The area under the Regulates Intracellular Unfolded Protein Response within Human Receiver Operating Characteristic Curve for detection of diabetes for FAI Adipose Tissue from Type 2 Diabetes Mellitus Patients and SHBG were 0.711 and 0.386 respectively for males and 0.430 and PHILIP D. VOYIAS, ANTONYSUNIL ADAIKALAKOTESWARI, WARUNEE KUMA- 0.660 respectively for females. Lower levels of SHBG contribute to the risk SAIYAI, IOANNIS KYROU, JANA VRBIKOVA, VOJTECH HAINER, MARTIN FRIED, of T2DM through obesity dependent metabolic pathways but low FAI is a PETRA SRAMKOVA, PONNUSAMY SARAVANAN, SUDHESH KUMAR, PHILIP better marker of diabetic state in males. MCTERNAN, GYANENDRA TRIPATHI, Coventry, United Kingdom, Prague, Czech Supported By: KFAS (2011-1302-01) Republic In obesity unfolded proteins accumulate in the endoplasmic reticulum and & 2004-P activate the unfolded protein response (UPR). UPR in adipose tissue (AT) is Metabolic Surgery in Obese Patients with Type 1 Diabetes Mellitus critical to the initiation and integration of infl ammation and insulin signalling CHRISTINE K. STIER, Frankfurt, Germany pathways in obese and type 2 diabetes mellitus (T2DM) patients. The aim Introduction: The prevalence of obesity and type 2 diabetes mellitus is of this study was to examine the effects of novel malabsorptive and bypass increasing world wide. Obesity surgery is an effective method for treating bariatric surgery in obese women with T2DM on AT UPR and systemic obesity and diabetes type 2. But little is known about bariatric surgery in insulin and glucose homeostasis. Abdominal subcutaneous AT biopsies: type 1 diabetes. 30 Caucasian obese T2DM women (age 54.1±1.3(mean±SE)years, BMI Method: We report of ten female patients with diabetes type 1. Five 41.2±1.0kg/m2) who underwent malabsorptive; gastric band n=9, or gastric patients underwent Roux-en Y gastric bypass. They were 33-50 years old plication n=13, or bypass; biliopancreatic diversion n=8, bariatric surgery. with diabetes since 2-34 years. They controlled it with CSII (continuous Data was collected at surgery and 6 months post-surgery. mRNA and protein subcutaneous insulin infusion), respectively three of them with an intensive expression for UPR, infl ammatory and macrophage markers were measured insulin-therapy (ICT). Two patients had sleeve gastrectomy. They were and correlation analysis performed. Six months post-surgery all subjects 35 and 38 years old, had had diabetes since 23 respectively 19 years and improved HbA1c, glucose, insulin, and body weight (p<0.001) with mean both controlled it with CSII. Three patients had biliopancreatic diversion/ excess BMI lost 33.4±2.4%. Irrespective of operation type, UPR markers: duodenal switch. At surgery they were 42-52 years old, with diabetes since ATF6, IRE1α, XBP1s, ATF4, and CHOP mRNAs and ATF6, pIRE1α, XBP1s, 8-25 years. They were treated with an ICT. Calnexin and Bip proteins, and pro-infl ammatory marker IL-6 mRNA were Results: Our results showed a remarkable weight reduction as well as all signifi cantly reduced (p<0.05). UPR signalling pathway correlations were an improvement in blood glucose control and the insulin requirement. Pre- strengthened post-surgery between ATF4 and CHOP (p=0.041 to p<0.001) surgery BMI ranged between 37,3-55,3 kg/m2 and improved to 25,3-29,0 kg/ and IRE1α and ATF6 (p=0.853 to p<0.001). Post-surgery plasma glucose m2 one year after surgery. HbA1c decreased from 5,7-10,9% prior to 5,7-8,5 correlated signifi cantly with XBP1s mRNA controlled for BMI (p=0.034). No % one year post-surgery. The insulin requirement was reduced from 0,48- signifi cant change in macrophage CD14 or CD68 mRNA expressions were 1,13 IU/kg body weight pre-surgery to 0,14-0,62 IU/kg after one year. detected. In summary malabsorptive and bypass bariatric surgery reduced Conclusion: The results are impressive and show an improvement in UPR and IL-6 expression in AT. Weight loss surgery modifi ed UPR signalling insulin sensitivity following obesity surgery. However, an optimal blood coupled with systemic glucose highlighting the fi nding that systemic factors glucose control still remains very important in the therapy of diabetes type 1 can have direct impact on AT UPR signalling. to avoid long-term complications. Obesity surgery is an effective method not Supported By: EFSD; BBSRC & AZ Studentship

only for type 2 diabetes but also for obese type 1 diabetes patients. Integrated POSTERS & 2007-P & 2005-P Berberine vs. Pioglitazone for Treatment of Nonalcoholic Fatty Liver Physiology/Obesity Attenuated Rises in Adiponectin and Fat Loss Characterize Type 2 Disease and Its Associated Impaired Glucose Metabolism Diabetes Non-remission Status 2 Years Post-Bariatric Surgery HONGMEI YAN, MINGFENG XIA, XINXIA CHANG, HUA BIAN, HUANDONG STEVEN K. MALIN, JAMES BENA, BETH ABOOD, CLAIRE E. POTHIER, DEEPAK L. LIN, LINSHAN ZHANG, XIUZHONG YAO, SHENGXIANG RAO, MENGSU ZENG, BHATT, STEVEN E. NISSEN, STACY A. BRETHAUER, PHILIP R. SCHAUER, JOHN P. YINFANG TU, WEIPING JIA, WEI DENG, XIN GAO, Shanghai, China KIRWAN, SANGEETA R. KASHYAP, Cleveland, OH, Boston, MA To evaluate the effi cacy and safety of berberine (BBR) or pioglitazone Although bariatric surgery improves glycemic control, not all patients (PGZ) in treatment of non-alcoholic fatty liver disease (NAFLD) patients with achieve type 2 diabetes (T2D) remission. This study sought to identify impaired glucose regulation (IGR) or type 2 diabetes mellitus (DM). This is metabolic determinants of T2D non-remission after bariatric surgery. 37 an open label RCT study. We randomly assigned 184 adults with NAFLD adults (48±9y, BMI: 36±3kg/m2, HbA1c: 9.7±2%) randomized to Roux-en-Y and impaired glucose metabolism to 16 weeks of treatment with lifestyle gastric bypass (RYGB, n=18) or sleeve gastrectomy (SG, n=19) were followed- intervention (LI), LI plus BBR (1.5g daily), or LI plus PGZ (15mg daily). Before

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A513 OBESITY—HUMANCATEGORY

and after treatment, we assessed hepatic fat content (HFC) by 1H-MRS, knockout mice exhibited reduced adipose tissue mass and infl ammation liver enzymes, glucose and lipid metabolism, serum insulin, osteocalcin together with enhanced glucose tolerance and insulin sensitivity. and C reactive protein (CRP) levels. BBR plus LI treatment, as compared To assess the role of miR-150 in humans, we studied 10 overweight or obese with LI control, was associated with a signifi cantly higher reduction in adults (ages 26-57 years) placed on a very low calorie diet (VLCD), which HFC (by 51.30% vs. 27.98%, p=0.035), total cholesterol, triglyceride, we have shown increases insulin sensitivity and reduces intramyocellular alanine aminotransferase (ALT), body weight and waist circumference, a lipid. Subjects were fi rst equilibrated for 1 week on an isocaloric diet (50% signifi cantly higher increasing in early phase insulin secretion (ΔI30/ΔG30) carbohydrate, 30% fat, 20% protein), then studied before and after a 1-week and serum osteocalcin levels (all p<0.05). PGZ plus LI had a signifi cantly VLCD (800 kcal/day for females, 1000 for males) of the same macronutrient higher reduction than LI group in 2-hour glucose, ALT (all p<0.05), but the distribution. Body composition was assessed by DEXA, insulin sensitivity by difference of HFC between PGZ and control was not signifi cant (by 41.02% OGTT and Matsuda index, and circulating miR-150 was measured by RT-PCR vs. 27.98%, p>0.05). Although these data had higher improvement in LI plus in fasting blood. After the 1-week VLCD, patients experienced signifi cant BBR or PGZ than in LI group, such as fasting glucose, HbA1c, HDL-c, LDL-c, weight loss (p<.001), decreased waist circumference (p=.039), and reduced aspartate aminotransferase, glutamyltransferase and serum CRP levels, fat mass (p<.001). There was also an increase in insulin sensitivity (p=.012) the differences of them were insignifi cant (all p>0.05). The incidence of that was accompanied by a marked 58.4% reduction in miR-150 from baseline drug-related adverse event between groups were signifi cantly different (p<.001). We also measured plasma miR-33, which has been implicated as (p<0.001), with no serious adverse events occurred. For NAFLD patients with a marker for atherogenesis, and found that the VLCD also reduced fasting abnormal glucose metabolism, LI and LI plus BBR or plus PGZ are all effective miR-33 levels by 42.9% (p<.001). treatments in improving glucose metabolism, reducing HFC and liver In conclusion, a short-term VLCD increases insulin sensitivity and leads to enzymes. BBR was better than the other two treatments in improving early pronounced reductions in circulating concentrations of miR-150 and miR-33. phase insulin secretion, lipid profi le, reducing weight, waist circumference, When considered in light of our miR-150 data in cultured macrophages and and serum osteocalcin levels. BBR and PGZ were all well tolerated. knockout mice, the data indicate that microRNAs could play a signifi cant role Supported By: NSFC (81200627, 81270933); National Basic Research Program of in the regulation of human metabolism, and in the pathophysiology of insulin China (2012CB524906) resistance and cardiometabolic disease. Supported By: ADA (1-13-IN-19); U.S. Dept. of Veterans Affairs (P30-DK079626); NIH (DK-03876, DK-083562) Guided Audio Tour: Pathophysiology of Human Obesity (Posters: 2008-P to 2014-P), see page 17. & 2010-P Next Generation Sequencing Methylation Profi ling and Whole & 2008-P Human Genome Microarray Analysis in Skeletal Muscle from Lean Magnetic Resonance Spectroscopy for Analysis of Fatty Acid and Obese Subjects Composition in Humans—Intra- and Interindividual Differences in SAMANTHA E. TANGEN, LATOYA E. CAMPBELL, MARGARET K. LINAN, LORI the Amount of Unsaturated Fatty Acids R. ROUST, VALENTIN DINU, LAWRENCE J. MANDARINO, DAWN K. COLETTA, JÜRGEN MACHANN, ERWIN SCHLEICHER, NORBERT STEFAN, HANS-ULRICH Tempe, AZ, Scottsdale, AZ HAERING, ANDREAS FRITSCHE, FRITZ SCHICK, Tübingen, Germany Obesity leads to alterations in metabolic function, and the role of epigenetic Proton magnetic resonance spectroscopy (1H-MRS) offers a non- factors is not well understood. The aim of our study was to determine invasive tool for in-vivo analysis of the composition of fatty acids (FA) in whether skeletal muscle from obese patients has different patterns of human adipose tissue (AT). Especially the amount of mono- (MUFA) and DNA methylation that could explain molecular changes related to metabolic polyunsaturated (PUFA) FA can be quantifi ed. This study was performed to differences. Obese (n=4; BMI=33.0±1.1 kg/m2) and lean (n=6; BMI=24.7±0.9 assess intra- and interindividual differences in composition of various AT kg/m2) subjects had vastus lateralis muscle biopsies taken basally, and next- compartments. generation sequencing reduced representation bisulfi te treatment and 4x44K Fifty overweight/obese volunteers (36m, 14f) participated in this study. Whole Human Genome Microarray analyses were performed on DNA and RNA 1H-MRS was performed on a 3 T whole-body imager in four subcutaneous isolated from the biopsies. From sequencing analysis, 1,198,747 methylation (SCAT) compartments: neck (SCATneck), calf (SCATcalf), superfi cial (SSCAT) sites were captured, and of these 1,935 were differentially methylated and deep (DSCAT) abdominal SCAT. Two internal fat storages were (P<0.05). A methylation site in SH3PXD2A was the most hypermethylated in under investigation: bone marrow in the tibia (BM) and visceral AT (VAT). the obese vs. lean (0.86±0.08 vs. 0.13±0.08) and a site in KCNK4 was the most Unsaturation indices are calculated as ratio of vinyl-resonance (MUFA+PUFA) hypomethylated (0.32±0.15 vs. 0.97±0.03). FANK1 had 9 unique methylation to methyl (CH3) (=vinyl/CH3) and diallylic (PUFA) to methyl (=diallylic/CH3). sites (8 located upstream of the gene) that were signifi cantly altered in the Regarding the entire cohort, vinyl/CH3 is highest for SCATcalf (0.622) obese subjects. The microarray analysis resulted in 605 signifi cant probes and lowest in BM (0.527). The highest diallylic/CH3 was observed for representing 422 unique genes (P≤0.05 and Fold Change ≥1.2). Between the SSCAT (0.108), the lowest in BM (0.093). Highly signifi cant intraindividual signifi cant methylation and microarray gene lists there were 25 genes in differences for vinyl/CH3 are present for almost all compartments. There is common; 2 were hypomethylated and increased in mRNA expression; 12 were no association of unsaturation indices with BMI or age. Except for SCATcalf hypermethylated and decreased in mRNA expression and the remaining 11 (lower vinyl/CH3 for females, p<0.05), there are no signifi cant gender related genes methylation status did not correlate with mRNA expression. Functional differences in unsaturated FA. However, females are characterized by clustering analysis identifi ed 4 transcription factors (ZNF717, CRTC1, ARID1A, slightly lower diallylic/CH3 in all compartments (n.s.). GTF3C2) that were all hypermethylated and decreased in mRNA expression Signifi cant differences in the composition of FA in several AT compartments in obesity. Interestingly, CREB regulated transcription coactivator 1 (CRTC1) are shown in this cross-sectional study. There are only moderate gender- is a known coactivator of PPARGC1A and induces mitochondrial biogenesis in related differences with a vague trend towards less unsaturated FA (mainly skeletal muscle. Our results provide evidence that the manifestation of specifi c PUFA) in females. Further examinations are mandatory to evaluate a genes depend upon methylation levels and suggests that these patterns could probable relation of unsaturation indices with respect to metabolic disorders lead to obesity. (as insulin resistance) or to show up short term changes in FA composition Supported By: NIH/NIDDK Integrated

POSTERS during dietary lifestyle intervention. Supported By: German Center for Diabetes Research (DZD e.V. 01GI0925) & 2011-P Physiology/Obesity Impaired Insulin-mediated VLDL-Triglyceride Kinetics in Obese & 2009-P Men with Nonalcoholic Fatty Liver Disease Role of MicroRNAs -150 and -33 in Human Insulin Resistance MARIANNE K. POULSEN, BIRGITTE NELLEMANN, STEEN B. PEDERSEN, HENNING ELIZABETH MA, NANLAN LUO, LING TIAN, WEI ZHANG, DENNIS STEVERSON, GRØNBÆK, SØREN NIELSEN, Aarhus, Denmark YUCHANG FU, W. TIMOTHY GARVEY, Birmingham, AL Non-alcoholic fatty liver disease (NAFLD) is an obesity-associated Insulin resistance plays a key role in Metabolic Syndrome, as well as condition with increased fasting VLDL-TG secretion as well as insulin the risk for future Type 2 Diabetes and cardiovascular disease. However, resistant glucose and fatty acid metabolism. The purpose of this study was the mechanism linking insulin resistance with cardiometabolic disease to compare basal and insulin-mediated VLDL-TG and glucose kinetics in pathophysiology is unclear. Based on miRNA profi ling, we previously obese men with (NAFLD+) and without NAFLD (NAFLD-). discovered that microRNA-150 (miR-150) was upregulated during the Twenty-seven obese (BMI > 28 kg/m2) men, 18 NAFLD+ and 9 NAFLD- transformation of THP-1 macrophages to foam cells, and that miR-150 determined by MR spectroscopy, were included. 14C-labeled VLDL-TG and

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A514 OBESITY—HUMANCATEGORY

3H-labeled glucose tracers were used in a hyperinsulinemic-euglycaemic subjects whereas no difference was found in portal vein and hepatic arterial clamp in combination with indirect calorimetry and blood and breath blood fl ow (NS). After surgery, notable decreases were observed in weight samples, to assess VLDL-TG and glucose kinetics. DEXA scan assessed body (by 24%, P < 0.0001), insulin resistance (HOMAIR, by 51%, P < 0.0001), fasting composition. glucose levels and HbA1c (both by -12%, P < 0.0001, respectively). Moreover, Liver fat content was 19.8% in NAFLD+ and 3.7% in NAFLD- men, in former diabetic subjects β-cell glucose sensitivity and insulinogenic respectively. However, body composition did not differ between groups. index were signifi cantly improved (by 44%, P < 0.02, and by 70%, P < 0.002, VLDL-TG secretion, oxidation and plasma concentration were signifi cantly respectively), and clinical remission of diabetes was observed in nine of greater in NAFLD+ men compared with NAFLD- men during hyperinsulinemia these 11 subjects. Compared to preoperative rates, blood fl ow was further (secretion: 45.5[21.8-143.5] vs. 11.1[1.8-87.0]µmol/min, P=0.009, oxidation: decreased by 17% in the pancreas (P = 0.02), by 76% in the portal vein and 37.4[15.4-102.8] vs. 10.0[1.6-50.9]µmol/min, P=0.008, concentration: 0.56 ± by 56% in the hepatic artery (both P < 0.0001, respectively). The amount of 0.08 vs. 0.23 ± 0.07 µmol/L, P=0.01). Likewise, the percent decrease in these weight loss showed a trend with the decrease in portal vein blood fl ow (r = parameters was signifi cantly less in NAFLD+ compared with NAFLD- men 0.36, P = 0.10), whereas increased insulin sensitivity - as characterized by 2h (secretion: -33.1 vs. -63.7%, P<0.001, oxidation: +7.1 vs. -44.0%, p<0.001, OGIS index - associated with the decrease in pancreatic blood fl ow (r = -0.48, concentration: -12.3 vs. -55.7%, p<0.001). Insulin-mediated glucose disposal P < 0.04) after surgery. The outcome was similar in patients irrespective rate was signifi cantly decreased in NAFLD+ compared with NAFLD- men of the type of bariatric procedure. In conclusion, obesity is characterized (3.9±0.3 vs. 5.5±0.8 mg/kg/min, P=0.027). Insulin was comparable in the two by hyperemic splanchnic vasculature which is normalized after bariatric groups in both periods. However, FFA was signifi cantly greater in NAFLD+ as surgery and weight loss. Decrease in pancreatic blood fl ow is associated compared to NAFLD- men during hyperinsulinemia (0.04±0.01 vs. 0.03±0.01 with the increase in insulin sensitivity. mmol/L, P=0.038). Supported By: Academy of Finland In conclusion, the inability of NAFLD+ men to suppress VLDL-TG kinetics during hyperinsulinemia seems to be an early pathophysiological & 2014-P manifestation preceding impaired glucose kinetics. Macrophage Coculture Inhibits PGC1-alpha and UCP-1 Expression in Human Adipocytes & 2012-P BRIAN S. FINLIN, BEIBEI ZHU, CHARLOTTE A. PETERSON, PHILIP A. KERN, Lexing- Role of Fat Storage Inducing Transmembrane Protein 2 (FIT2) in ton, KY Regulating Human Adipocyte Function PGC1-α is a transcriptional co-activator of PPARγ that regulates MADHUR AGRAWAL, CHIA ROU YEO, ASIM SHABBIR, JIMMY SO BOK YAN, mitochondrial biogenesis and uncoupling protein-1 (UCP-1) expression in CHIN MENG KHOO, E. SHYONG TAI, SUE-ANNE E.E. SHIOW TOH, Singapore, brown adipocytes. Recent studies suggest that PGC-1α plays a benefi cial Singapore role in white adipose tissue (WAT). In humans, PGC-1α is induced in WAT Adipose tissue is increasingly appreciated as an important regulator of by pioglitazone, an insulin sensitizer, and PGC-1α is decreased in WAT with whole body energy metabolism. Poor function of fat, rather than amount of increasing obesity. Since macrophages are increased in WAT with obesity, fat, has been suggested as an important determinant for insulin resistance we sought to determine whether macrophages inhibit PGC-1α expression (IR). Failure to partition and store excess triglycerides (TG) and free fatty acids in human adipocytes. Furthermore, recent studies suggest that PGC-1α may be detrimental to healthy adipocyte function, triggering aberrations is inhibited by TGF-β, which is increased in adipose tissue with obesity; in downstream pathways leading to IR. We aimed to validate the role of we previously reported that adipocytes increase TGF-β signaling when a recently discovered lipid droplet protein, FIT2, in regulating adipocyte cocultured with macrophages. We treated differentiated adult-derived function. Primary human adipocytes were obtained from (1) subcutaneous human adipocyte stem cells (ADHASC) with a TGF-β receptor blocker adipose depot from non-diabetic (SC_NDM) and diabetic (SC_DM) subjects, (SB505124) and found that the adipocytes expressed higher levels of PGC- (2) subcutaneous (SC_BA) and omental (OM_BA) adipose tissue from obese 1α and UCP-1 (n=3; P<0.05). Next, we cocultured differentiated ADHASC diabetic and obese non-diabetic patients undergoing bariatric surgery. FIT2 cells with polarized macrophages, which were derived by treating THP-1 mRNA and protein expression in adipocytes derived from individuals within monocytes with LPS/γ-interferon (M1), IL-4 (M2a), or IL-10 (M2c). Coculture the respective groups were assessed by real-time qPCR and western blot. with M1, M2a, or M2c macrophages potently (>90 %) inhibited adipocyte Up-regulation of FIT2 (p<0.05) with increasing days of adipogenesis was PGC-1α mRNA and protein expression (n=3; P<0.05) and UCP-1 mRNA observed at both mRNA and protein levels. However, in fully differentiated expression (n=3; P<0.05). However, inhibition of TGF-β receptor signaling adipocytes, FIT2 mRNA expression was signifi cantly lower (p<0.05, n=3 with SB505124 failed to protect against the reduction in PGC-1α expression per group) in SC_DM when compared to SC_NDM, suggesting reduced by macrophage coculture. We next evaluated other mechanisms of regulating lipid storage capacity in the diabetic (DM) state. FIT2 mRNA and protein PGC1-α expression and found that macrophage coculture induced known expression were signifi cantly lower (p<0.05, n= 6) in OM _ BA compared to SC_ inhibitors of PGC-1α such as the pocket protein Rb and RIP140 2-fold (n=3; BA derived from the same individuals, demonstrating metabolically distinct P<0.05). These data suggest that PGC-1α and its targets such as UCP-1 are features between the depots. Amount of intracellular TG accumulation in negatively regulated by macrophage infi ltration into adipose that occurs response to palmitate correlated signifi cantly with FIT2 protein expression with obesity. This may contribute to adipose dysfunction and inhibit the in SC_DM and SC_NDM (p<0.001, n=6) and was higher in SC_NDM when beiging process that may occur in WAT. compared to SC_DM (p<0.05, n=3). In conclusion FIT2 expression correlates Supported By: GM103527, DK71349 with lipid storage capacity, is lower in OM as compared to SC and is lower in DM as compared to non-DM adipocytes. These fi ndings indicate the 2015-P importance of FIT2 protein in effective TG storage in human adipocytes to Heat Treatment with Mild Electrical Stimulation Attenuates maintain metabolic homeostasis. Systemic Infl ammation and Reduces Cytokine Expression in Mono- cytes from Subjects with Metabolic Syndrome & 2013-P TATSUYA KONDO, RINA MATSUYAMA, RIEKO GOTO, KAORU ONO, SAYAKA The Effects of Bariatric Surgery on Splanchnic Blood Flow and KITANO, JUNJI KAWASHIMA, HIROYUKI MOTOSHIMA, TAKESHI MATSUMURA, Metabolism in Morbidly Obese Subjects HIROFUMI KAI, EIICHI ARAKI, Kumamoto, Japan Integrated HENRI HONKA, JARNA C. HANNUKAINEN, HEIDI IMMONEN, JETRO J. TUULARI, Insulin resistance and chronic infl ammation represent key pathophysio- POSTERS VESA OIKONEN, NOBU KUDOMI, PAULINA SALMINEN, RIITTA PARKKOLA, logical characters in metabolic syndrome (MS). Physiology/Obesity ANDREA MARI, PATRICIA IOZZO, PIRJO NUUTILA, Turku, Finland, Kagawa, Japan, We have identifi ed that induction of heat shock protein (HSP) 72 using Tampere, Finland, Padua, Italy, Pisa, Italy mild electrical stimulation with hyperthermia (MET) improves glucose The underlying mechanisms of improved glucose tolerance seen after homeostasis, insulin resistance, chronic infl ammation and reduces visceral bariatric surgery are not fully elucidated. In the present study our aim was adiposity in diabetic mice as well as in human with MS or type 2 diabetes. to investigate the changes in splanchnic blood fl ow and metabolism after The present study investigated whether MET treatment has anti- bariatric surgery and concomitant weight loss. Thus, a total of 27 morbidly infl ammatory effects in circulating monocytes. Ten male subjects with MS 15 obese subjects and 15 age-matched controls were recruited to [ O]H2O-PET/ were received MET treatment (1.4 ± 0.1 V/cm: distance between pads, 0.1 CT study. For obese subjects imaging studies were performed before and millisecond duration. 55 pulses/sec, 60 min at 42oC, 4 times a week) for 4 six months after the bariatric procedure (either Roux-en-Y gastric bypass or weeks. Circulating peripheral blood mononuclear cells (PBMC) were isolated sleeve gastrectomy). Compared to controls, preoperative rate of pancreatic at fasted state before and after the MET treatment. Then, monocytes were blood fl ow was lower (1.1 ± 0.2 versus 1.5 ± 0.5 ml/ml*min, P < 0.02) in obese further isolated from PBMC using CD14 antibody with magnetic beads.

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A515 OBESITY—HUMANCATEGORY

After MET treatment, HSP72 protein levels in monocytes signifi cantly Within the SG group, signifi cant improvements were observed at 1 and increased by 34.5 ± 6.5% compared to baseline. Phosphorylation of AMP- 6 months respectively in measures of insulin sensitivity (fasting insulin: kinase also increased by 79.9 ± 35.7%. Stress signal-activated phospho- -4.8mU/L, -8.5mU/L; fasting C-peptide: -0.6pmol/L, -1.1pmol/L; Homeostasis c-jun N-terminal kinase was attenuated by 40.0 ± 18.6 %. LPS-stimulated Model Assessment [HOMA-IR]: -0.144, -0.174; HOMA insulin sensitivity nuclear factor (NF)-κB p65 subunit nuclear translocation, visualized by [%S]: +29.6, +92.4) and hepatic insulin clearance (+0.07, +0.13). Signifi cant immunofl uorescence, was attenuated by MET treatment. Quantitative increases were observed in AUC0-30mins, AUC0-60mins and AUC0-120mins measurements of mRNA expression after LSP stimulation for C-reactive for GLP-1 at 1 and 6 months postoperatively (AUC0-30 pmol h L-1: +300, +331, protein, Interleukin-6, NF-κB and tumor necrosis factor-α showed a AUC0-60: +300, +294, AUC0-120: +316, +295). There were no signifi cant reduction by 28.0 ± 3.7% (P = 0.006), 7.5 ± 5.1% (P = 0.36), 22.7 ± 4.8% (P = changes in GIP. These results were comparable to the BPD group. 0.044) and 23.3 ± 7.3% compared to baseline (P = 0.03), respectively. SG is associated with early improvements in glucose homeostasis and We confi rmed that MET treatment improved systemic chronic infl ammation remission of diabetes. Our results suggest that these improvements occur as in subjects with MS. As MS is clustering risk of cardio-vascular diseases, a result of improved hepatic insulin sensitivity, enhanced postprandial GLP-1 this anti-infl ammatory effect of MET may provide novel anti-atherogenic response and subsequent restoration of the early phase insulin release. The therapy in life-style related diseases such as MS and type 2 diabetes. rates of remission of T2DM are comparable to those associated with BPD. Supported By: Ministry of Education, Culture, Sports, Science and Technology; Japan Society for the Promotion of Science (2259098701) 2018-P WNT Signaling May Disassociate Obesity from Insulin Resistance 2016-P in Polycystic Ovary Syndrome and WNT-Inhibition Appears to Be Changes in Cardiovascular Risk Factors at 1 and 6 Months Following Anti-infl ammatory Laparoscopic Sleeve Gastrectomy SIDIKA E. KARAKAS, ROGELIO U. ALMARIO, Sacramento, CA AKHILA MALLIPEDHI, SARAH PRIOR, GARETH DUNSEATH, RICHARD BRACKEN, Proteins that transmit or inhibit WNT signaling regulate adiposity, insulin KATHIE WAREHAM, JANE GRIFFITHS, NIA AYRE, JONATHAN D. BARRY, SCOTT resistance and infl ammation. We investigated serum WNT5 and WNT- CAPLIN, JOHN BAXTER, IMRAN ALAM, JAMES MORGAN, SAM RICE, STEVE inducible secreted protein 2 (WISP2) and WNT-inhibitors SFRP4 and SFRP as LUZIO, STEPHEN BAIN, JEFFREY STEPHENS, Swansea, United Kingdom, Llanelli, they relate to obesity and metabolic disorders seen in PCOS. United Kingdom Fasting samples were obtained from 32 obese-PCOS, 25 non-obese PCOS The aim of the study was to examine the early effects of Sleeve gastrecto- and 27 reference women; 14 reference men who were otherwise healthy my (SG) on cardiovascular risk factors and glycaemic control in patients with and on no medication. type 2 diabetes (T2DM). The effects were also examined in participants Results: All subjects had measurable WNT5, WISP2 and SFRP4. SFRP5 undergoing Bilio-pancreatic Diversion (BPD). was undetectable in 33% of reference women, 86% of reference men and A non-randomized prospective study comprising of 22 participants only in 10.5% of PCOS women. As shown in the Table, reference men had undergoing SG (body mass index [BMI] 50.1kg/m2, glycated hemoglobin lower WNT5 than women; reference women and men had lower SFRP5 than [HbA1c] 53 mmol/mol) and 15 participants undergoing BPD (BMI 62.1kg/m2, PCOS women. In PCOS, WNT5 tended to correlate inversely with weight HbA1c 58 mmol/mol). Clinical and biochemical measurements were carried (r= -0.228, p = 0.087), fat-mass (r = -0.236, p = 0.078) and leptin (r = -0.235, out at baseline and 1 and 6 months post-operatively in addition to oral p = 0.081) and correlated directly with insulin during OGTT (90 min r = 0.364, glucose tolerance testing (OGTT). p = 0.005; 2 h: r=0.257, p = 0.054) and IL-6 (r = 0.277, p = 0.056). In reference In both groups, signifi cant reductions were observed at 1 and 6 months subjects WNT5 correlated inversely with weight but not with insulin in weight, BMI and waist circumference at 1 and 6 months with a mean resistance. Serum SFRP5 correlated inversely with IL-1β (r = -0.433, p = reduction of 31.1kg after SG and 37.5kg after BPD at 6 months. There was 0.002), TNFα (r = -0.396, p = 0.005), cholesterol (r = -0.297, p = 0.025) and a signifi cant reduction in the systolic blood pressure at 1 month following apoprotein B (r = -0 317, p = 0.016). SG (p=0.04). Signifi cant changes were observed in fasting glucose, 2-hour Conclusion: WNT signaling and WNT inhibitors may play important roles glucose and HbA1c in both groups at 1 and 6 months. Within the SG group, in insulin resistance and infl ammation in PCOS. 33.2% of participants had normal glucose concentrations during the OGTT at Table 1. Clinical and Biochemical Variables of Women with Polycystic Ovary 1 month and 82.4% at 6 months. Within the BPD group the respective values Syndrome (PCOS), Reference Women and Reference Men (Mean SEM). were 23.8% and 53.2%. In the SG group, 87.5% of subjects had discontinued Obese-PCOS Non-Obese-PCOS Reference Reference all treatments within 6 months (71.4% in the BPD group). There was also a (n = 32 ) (n = 25) Women Men corresponding 50% reduction in the number of participants receiving oral (n = 27 ) (n = 14 ) therapies for T2DM in both the groups. Age (years) 31.7±6.4 32.8±6.8 44.5±9 a,b 37.8±113.5 SG is recognized as a stand-alone bariatric procedure with a superior safety profi le. Our results suggest that SG is effective in managing IGT and Weight (kg) 114.1±14.0 73.9±10.5 73.7±8.3 89.1±10.1 a,b T2DM in subjects with morbid obesity, with comparable remission rates of BMI (kg/m2) 40.6±5.6 28.7±4.4 a 27.6±2.3 a 27.0±1.9 a T2DM associated with BPD. WNT5 (ng/ml) 1.63±0.12 1.72±0.13 1.41±0.12 1.01±0.11 a,b WISP2 (ng/ml) 209.6±79.3 305.6±70.1 250.0±51.5 168.7±25.2 2017-P SFRP5 (ng/ml) 2.84±2.39 4.01±4.52 0.52±0.98 a,b 0.20±0.64 a,b Temporal Changes in Glucose Homeostasis and Incretin Hormone Response at 1 and 6 Months Following Laparoscopic Sleeve Gas- SFRP4 (ng/ml) 5.29±5.49 7.43±9.72 5.57±6.73 5.70±6.84 trectomy a: p< 0.05 vs. obese-PCOS; b: p< 0.05 vs. non-obese-PCOS. AKHILA MALLIPEDHI, SARAH PRIOR, GARETH J. DUNSEATH, RICHARD M. Supported By: NCCAM (AT002280, AT00340 to S.E.K.) BRACKEN, KATHIE WAREHAM, JANE GRIFFITHS, NIA EYRE, JONATHAN D. BARRY, JOHN N. BAXTER, SCOTT CAPLIN, IMRAN ALAM, JAMES MORGAN, SAM RICE, STEPHEN D. LUZIO, STEPHEN C. BAIN, JEFFREY W. STEPHENS, 2019-P Swansea, United Kingdom, Llanelli, United Kingdom Ectopic Colorectal Fat Accumulation in Colorectal Polyp Patients Integrated POSTERS Our aim was to examine the detailed metabolic effects of laparoscopic with Insulin Resistance sleeve gastrectomy (SG) on the temporal changes in insulin and glucose SUMIO KAWATA, SERIKO IKEZOE, YUICHI YASUNAGA, KAZUMASA OKA, NAOKI Physiology/Obesity homeostasis, incretin hormones and hepatic insulin clearance in patients DAN, HITOSHI MATSUMOTO, YOSHIAKI INUI, Nishinomiya, Japan with type 2 diabetes mellitus (T2DM). These effects were also examined in Obesity is known to be associated with ectopic fat accumulation in the participants undergoing Bilio-pancreatic Diversion (BPD). liver and skeletal muscle. However, obesity-related accumulation of fat A non-randomized prospective study comprising of 22 participants in the colorectal mucosal and submucosal regions has not been reported. undergoing SG (body mass index [BMI] 50.1kg/m2, glycated hemoglobin Several lines of evidence have suggested that obesity and insulin resistance [HbA1c] 53mmol/mol) and 15 participants undergoing BPD (BMI 62.1kg/m2, are independent risk factors for the development of colorectal cancer and HbA1c 58 mmol/mol). Serial measurements of markers of insulin sensitivity, adenoma. C-peptide, glucagon like peptide-1 (GLP-1) and glucose-dependent We conducted the present study to seek evidence of obesity-related insulinotropic hormone (GIP) were performed during oral glucose tolerance colorectal accumulation of fat in patients with colorectal polyps, and its testing pre-operatively and 1 and 6 months post-operatively. Areas under relation with insulin resistance and metabolic disorders. For the 27 patients the curve (AUC) were examined at 30, 60 and 120 minutes. (15 males and 12 females) with colorectal polyp enrolled in this study, we

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A516 OBESITY—HUMANCATEGORY measured the triglyceride (TG), total cholesterol (TC) and phospholipid (PL) 2022-P contents of non-tumorous tissues surrounding the polyps which were obtained Impaired Lipid Accumulation Capacity in Induced Pluripotent Stem by endoscopic mucosal and submucosal resection. Immunohistochemistry Cells from Lipodystrophic Patients with BSCL2 Gene Mutations examination using perilipin (PLIN1) antibody (Progen Biotechnik, Gemany) EISAKU MORI, JUNJI FUJIKURA, KAZUHIRO NAKAO, MICHIO NOGUCHI, MASA- was done to examine the localization of lipid droplets. KATSU SONE, DAISUKE TAURA, TORU KUSAKABE, KEN EBIHARA, TAKAYUKI Of the 27 patients (median age, 65 yrs (34 to 84), 17 were overweight TANAKA, KIMINORI HOSODA, ISAO ASAKA, NOBUYA INAGAKI, KAZUWA NA- or obese (BMI > 25). Tertile analysis of TG/PL value to assess the factors KAO, Kyoto, Japan associated with higher TG/PL value showed that serum TG (P = 0.025), Congenital generalized lipodystrophy (CGL) is an autosomal recessive fasting plasma insulin (P = 0.041), and HOMA-IR (P = 0.013) were signifi cantly disorder characterized by marked scarcity of adipose tissue, extreme insulin higher in the highest tertile than in the lowest tertile. Serum adiponectin resistance, hypertriglyceridemia, hepatic steatosis and early onset of level (P = 0.046) was also signifi cantly lower in the highest tertile than in diabetes. Mutation of the BSCL2/Seipin gene causes the most severe form of the lowest tertile. Lipid droplets were observed in the submucosal region of CGL. We generated induced pluripotent stem (iPS) cells from two Japanese colorectal tissue, associated with a high TG/PL value. CGL patients with different BSCL2 gene nonsense mutations (E189X and Ectopic colorectal fat accumulation was observed in colorectal polyp R275X). Each of the patient-derived iPS (BSCL2-iPS) clones showed all the patients with higher fasting plasma insulin and HOMA-IR and lower hallmarks of pluripotency and could differentiate into derivatives of all three adiponectin. In addition, lipid droplets were localized in the colorectal germ layers in vitro and in vivo. Furthermore, BSCL2-iPS cells maintained submucosal region in patients with higher TG/PL values. These results the BSCL2 gene mutation and lacked the SEIPIN protein expression. Upon suggest an association of ectopic colorectal fat accumulation with insulin adipogenic differentiation, BSCL2-iPS cells exhibited marked reduction of resistance. both lipid content and PPARγ2 gene expression compared with those of iPS cells from healthy individuals. Forced expression of SEIPIN rescued the lipid 2020-P accumulation in differentiated BSCL2-iPS cells. BSCL2-iPS cells that could Reduced Striatal D2/3R Binding in Obesity Is Partially Reversible by recapitulate the lipodystrophic phenotypes in vitro have the potential to Long-term Weight Loss After Roux-en-Y Gastric Bypass Surgery provide valuable models to study the pathology of human lipodystrophy and ESTHER M. VAN DER ZWAAL, BARBARA A. DE WEIJER, MICHELLE C. PANTON, to develop drug screening assays. IGNACE JANSEN, FRITS J. BERENDS, ARNOLD VAN DE LAAR, MARIETTE T. ACKERMANS, ERIC FLIERS, SUSANNE E. LA FLEUR, JAN BOOIJ, MIREILLE J. 2023-P SERLIE, Amsterdam, Netherlands, Arnhem, Netherlands Human Abdominal Subcutaneous Adipocytes as an Active Source of Previous studies repeatedly reported reduced striatal D2/3 receptor (D2/3R) LpPLA2 Infl uenced by Metabolic State, with Adipocytes Converting availability in obese subjects compared to lean controls. However, it remains LDL, through LpPLA2, into the More Potent Atherogenic OxLDL unknown whether this refl ects a cause or a consequence of becoming obese. WARUNEE KUMSAIYAI, PHILIP VOYIAS, SARAVANAN PONNUSAMY, To examine the reversibility of reduced striatal D2/3R binding in obesity, we IOANNIS KYROU, NASSER AL-DAGHRI, THOMAS BARBER, SUDHESH KUMAR, previously determined D2/3R availability in 19 morbidly obese women 2 GYANENDRA TRIPATHI, PHILIP MCTERNAN, Coventry, United Kingdom, Riyadh, weeks before and 6 weeks after Roux-en-Y gastric bypass surgery (RYGB) Saudi Arabia using [123I]IBZM SPECT. Although baseline scans replicated a reduction in Obesity, infl ammation, lipotoxicity appear heavily intertwined as causes D2/3R binding compared to lean controls of ± 20 %, D2/3R binding did not of cardiovascular (CVD) risk in type 2 diabetes mellitus (T2DM). CVD is change signifi cantly despite signifi cant weight loss 6 weeks after surgery. correlated with high oxidized LDL and lipoprotein-associated phospholipase However, as body weight had not yet stabilized at that time, it remained A2 (LpPLA2) activity in serum. However, how important the adipocyte is in uncertain whether D2/3R binding might normalize after long-term weight contributing to systemic oxidation of LDL in obese T2DM through LpPLA2 loss. We therefore invited subjects of the short-term study to participate in activation is unknown. These studies characterised LpPLA2 and PLA2 a long-term study to repeat striatal D2/3R measurements. Repeated imaging isoforms in lean, obese, T2DM abdominal subcutaneous (AbdSc) and omental was performed in 12 subjects, with time since RYGB ranging from 2.5 to 3.6 (Om) AT by microarray, RT-PCR and Western Blot; the in vivo role of lipids and yrs. Mean body weight had declined from 129 ± 20 kg to 89 ± 16 kg (p<0.001), infl ammatory markers on circulating LpPLA2 levels were evaluated; coupled BMI from 46 ± 7 kg/m2 to 32 ± 6 kg/m2 (p<0.001) and the quantitative insulin with the in vitro regulation of LDL oxidation by LpPLA2 in human adipocytes sensitivity check index (QUICKI) increased from 0.34 ± 0.03 to 0.37 ± 0.03 (Chub S7). LpPLA2 mRNA levels were higher in AbdSc AT than Om AT in (p<0.01). Although an increase in D2/3R binding was observed in most obesity by 2 fold (P<0.05). The cPLA2 protein expression was increased subjects, this effect was not signifi cant for the entire group (specifi c binding with obesity in AbdSc AT (P<0.01). T2DM showed increased LpPLA2 mRNA ratios: 0.83 ± 0.13 to 0.88 ± 0.14, p=0.23; one-sided t-test). However, when levels in AbdSc (P<0.001) and OmAT (P<0.01). Serum LpPLA2 showed positive one outlier (showing a marked, unexplainable decrease in D2/3R binding correlations with cholesterol, TG, LDL, endotoxin and oxidized LDL (OxLDL) from 1.15 to 0.69) was excluded, the effect did reach signifi cance (0.80 ± (P<0.001) in non-diabetic subjects and with OxLDL (P<0.001), LDL (P<0.01) 0.09 to 0.90 ± 0.13, p= 0.032). Changes in D2/3R binding did not correlate and cholesterol (P<0.05) in T2DM. In human adipocytes, LpPLA2 protein with changes in BMI, QUICKI or weight loss. These fi ndings suggest that expression increased in response to LDL and OxLDL (P<0.001). Use of a striatal D2/3R availability increases with long term weight loss after RYGB, co-treated LpPLA2 inhibitor with LDL, in vitro signifi cantly reduced OxLDL but remains lower compared to lean controls. The reduction in striatal D2/3R production (P<0.05). The adipocyte appears to be an active source of LpPLA2, availability therefore appears only partially reversible. increased in T2DM, with LpPLA2 protein expression being induced by LDL and OxLDL in vitro. As such, increased LpPLA2 protein from the adipocyte in 2021-P obesity and/or T2DM appears to contribute to raise circulating OxLDL, which promotes further infl ammation and CVD risk. WITHDRAWN Supported By: Royal Thai Government Integrated POSTERS Physiology/Obesity

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A517 OBESITY—HUMANCATEGORY

2024-P 2026-P The Association of Dihomo-Gamma-Linolenic Acid Levels with Correction of Intermittent Hypoxia with CPAP Improves Cardio- Obesity and Nonalcoholic Fatty Liver Disease (NAFLD) in Type 2 metabolic Abnormalities and Systemic and Adipose Tissue Infl am- Diabetic Patients ma tion in Obese Subjects with Obstructive Sleep Apnea MARIKO HIGA, KAORU YAMASHITA, AYANO DOI, KEN KANAZAWA, MAI TASAKI, SEBASTIO PERRINI, MARIA BARBARO, ALESSANDRO CIAVARELLA, STELLA RIEKO KUNISHITA, TAKAMASA ICHIJO, HIROMI OUCHI, Yokohama, Japan KOUNAKI, ANGELO CIGNARELLI, VITALIANO QUARANTA, ROMINA FICARELLA, A polyunsaturated fatty acid (PUFA) is called nutritionally an essential ANTONIO DI TRANI, PASQUALE NIGRO, ANNALISA NATALICCHIO, LUIGI fatty acid, being an important nutrient for the body, but the excessive intake LAVIOLA, ONOFRIO RESTA, FRANCESCO GIORGINO, Bari, Italy may cause obesity and fatty liver. A dihomo-gamman-linolenic acid (DGLA) It is unclear whether in obesity chronic intermittent hypoxia (CIH) is n-6 PUFA, and is produced by desaturation and carbon chain elongation resulting from Obstructive Sleep Apnea (OSA) fosters the development reaction from linoleic acid, then to be further converted into arachidonic of cardiometabolic abnormalities. We hypothesized that treatment of CIH acid (AA). We investigated whether the blood DGLA levels associate with with continuous positive airway pressure (CPAP) in obese individuals with obesity and non-alcoholic fatty liver disease (NAFLD) in type 2 diabetic OSA would modify cardiometabolic and infl ammatory outcomes. 82 obese patients. The subjects were 62 type2 diabetic patients with no history patients with OSA were studied at baseline (T0) and after 3 months (T1) of of alcohol drinking and with negative type B and type C hepatitis viruses therapeutic CPAP, and compared with 25 BMI- and age-matched subjects (Age: 63.3±9.3 years, Male: 35 cases and Female 27 cases). They did not who had a negative single-night polysomnogram. Subcutaneous (Sc) adipose take eicosapentanoic acid (EPA) agents. They were divided into 2 groups, tissue (AT) biopsies were taken at T0 and T1 to assess mRNA expression i.e., Obese Group (n=29) having BMI of more than 25 kg/m2 and non-Obese of ER stress-related genes, CD36 and SIRT1. At T0, neck circumference Group (n=33) having BMI below 25 Kg/m2. The measurement of blood (+3.0 cm; p=0.01), systolic (+8.6 mmHg; p=0.03) and diastolic (+8.2 mmHg; PUFA was performed by a gas chromatography. The level of blood DGLA p=0.017) blood pressure, HOMA-IR (+4.5; p=0.005), A1c (+0.6%; p=0.01), was found to be signifi cantly (p<0.01) higher in Obese Group (39.2±12.0 µg/ serum triglyceride (+52.9 mg/dL; p=0.03) and white blood cells (+1,694 ml) than those in non-Obese Group (32.1±9.5 µg/ml). The ratio of DGLA/AA per µL; p=0.001) were higher in OSA than control obese subjects. At T1, were signifi cantly (p<0.01) higher in Obese Group (0.24±0.09) than in non- CPAP treatment resulted in decreases in BMI (-2.0 kg/m2; p=0.026), neck Obese Group (0.19±0.06). In the patients with ultrasound diagnosed NAFLD, circumference (-2.3 cm; p=0.005), systolic blood pressure (-15.4 mmHg; blood DGLA showed signifi cant high level in comparison with the patients p=0.015), heart rate (-8.7 bpm; p=0.032), and serum total cholesterol (-15.8 without NAFLD. Blood DGLA level showed a signifi cant positive correlation mg/dL; p=0.04) in OSA subjects, whereas no signifi cant changes in these with BMI, waist circumference, AST, ALT, blood triglyceride level and leptin, parameters were observed in the control obese or OSA obese subjects respectively.These results suggest that high levels of blood DGLA in type 2 exposed to non-therapeutic CPAP. Serum concentrations of IL-1ra, IL-2, IL-4, diabetic patients with obesity and NAFLD were attributable to an excessive IL-6, MCP-1 and PDGF were also reduced by CPAP in OSA subjects (p<0.05), intake of n-6 PUFA and the decrease in desaturase activity converting DGLA remaining unaltered in controls. Furthermore, in the Sc AT of OSA subjects, to AA. Insulin resistance might contribute to cause for higher blood levels of CPAP resulted in decreased CHOP, ATF4, ERO-1 and CD36 mRNA levels DGLA through a decrease in an activation of desatulation and elogation of and increased SIRT1 gene expression (p=0.02). Thus, correction of CIH by carbon chain in fatty acid. CPAP therapy in obese subjects with moderate-to-severe OSA improves anthropometric variables, cardiometabolic abnormalities, systemic infl am- 2025-P ma tion, and adipose tissue-specifi c ER stress/infl ammation. Metabolically Healthy Obese and Metabolically Unhealthy Lean Phenotypes 2027-P SIMONA GEORGIANA G. POPA, ADINA MITREA, ANDREEA ROTARU, EUGEN MOTA, Effects of Liraglutide 3.0 mg Cessation on Effi cacy and Safety/ MARIA MOTA, Craiova, Romania Tolerability Following a 56-Week Randomized Treatment Period in Metabolically healthy obese and metabolically unhealthy lean are two Obese/Overweight Adults with Type 2 Diabetes (SCALE™ Diabetes) atypical cardiometabolic phenotypes whose characteristics and prognosis RALPH A. DEFRONZO, BRUCE W. BODE, ROBERT F. KUSHNER, ANDREW J. are a subject of extensive scientifi c debate. LEWIN, TRINE V. SKJØTH, CHRISTINE B. JENSEN, MELANIE DAVIES, San The aims were to assess the prevalence of different metabolic phenotype Antonio, TX, Atlanta, GA, Chicago, IL, Los Angeles, CA, Bagsværd, Denmark, Søborg, and to analyze the characteristics of metabolically healthy obese and Denmark, Leicester, United Kingdom metabolically unhealthy lean phenotypes. Effects of liraglutide cessation on effi cacy (body weight, FPG) and safety We performed a cross-sectional analysis in a random sample of 966 (AEs, safety lab, mental health evaluation, binge eating) were investigated individuals, aged 20-79 years, derived from PREDATORR study. Subjects in a 12-week off-treatment period after 56 weeks of treatment in obese/ were classifi ed as metabolically unhealthy overweight/obese (MUHO) or overweight adults with type 2 diabetes (T2D). lean (MUHL) and metabolically healthy overweight/obese (MHO) or lean 846 individuals (age 54.9 yr, weight 105.9 kg, BMI 37.1 kg/m2 [27.0−67.6], (MHL), based on body mass index (BMI) and metabolic health defi ned as HbA1c 7.9%, FPG 158.2 mg/dL,7.3 yr of T2D) were randomized (2:1:1) to HOMA-IR<2,5. liraglutide 3.0 mg, 1.8 mg or placebo, as adjunct to diet and exercise, which The prevalence of MHO, MUHL and MUHO phenotypes was 38.4%, 2.9% continued in the off-treatment period. and, respectively 35.7% among all participants. More individuals with liraglutide (3.0 mg: 77%, 1.8 mg: 78%) than placebo The MUHL subjects had more impaired glycemic control (FPG: 126.2±76.4 (66%) completed the 56-week period. For the completers, liraglutide 3.0 mg vs. 78.3±20.8 mg/dl; HbA1c: 6.4±2.3 vs. 5.3±0.5%; p<0.001 for all), but higher and 1.8 mg induced greater reductions in body weight and FPG than placebo insulinemia (15.5±9.9 vs. 6.3±2.7 µUI/ml; p<0.001) than MHL subjects. (Table). After treatment cessation, weight regain occurred with liraglutide The MHO subjects had a better glycemic control (FPG: 82±14.3 vs. 3.0 mg (2.3%) and 1.8 mg (2.0%) but mean weight loss remained greater 106.9±39.4 mg/dl; HbA1c: 5.5±0.6 vs. 6.1±1.2%; p<0.001 for all), signifi cantly with 3.0 mg (-4.7%) than placebo (-2.7%) at week 68. FPG reverted to lower HOMA%B (226.1±218.8 vs. 245.5±213.9, p=0.005), insulin levels placebo levels within 2 weeks of treatment cessation. No adverse effects of (7.8±2.8 vs. 19.5±18.7 µUI/ml, p<0.001), uric acid levels (5±1.3 vs. 5,6±1.6 treatment cessation on safety or binge eating were noted. mg/dl, p<0.001), triglycerides (129.4±73.1 vs. 167.9±101.4 mg/dl, p<0.001), In conclusion, 12 weeks after liraglutide cessation, the benefi cial treatment Integrated

POSTERS waist (99.5±11.3 vs. 106±13 cm, p<0.001) and higher HDL-Cholesterol levels effects on body weight were reduced and the effects on FPG were reversed, (56.4±14.7 vs. 50.8±15.3 mg/dl, p<0.001) than MUHO individuals. In contrast, emphasizing the need for continued treatment. Physiology/Obesity all the analyzed cardiometabolic parameters were signifi cantly higher, Clinicaltrials.gov ID: NCT01272232. except HDL-Cholesterol which was lower, in MHO subjects compared to MHL subjects. Overall the MHO and MUHL subjects had more impaired cardiometabolic profi le than MHL subjects, but MHO subjects had a better cardiometabolic profi le than MUHO subjects.

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A518 OBESITY—HUMANCATEGORY

(133 mg/dl vs. 102 mg/dl, p=0.001), triglyceride (140 mg/dl vs. 117 mg/dl, Effect of Liraglutide Cessation on Body Weight and FPG (Completers). *Sub- p=0.045) and uric acid (5.8 mg/dl vs. 4.9 mg/dl, p=0.035) levels than obese jects with Data in Analysis. subjects without OSA. A1c levels and PPG120 were found to be signifi cantly Liraglutide 3.0 mg Liraglutide 1.8 mg Placebo correlated with raised apnea-hypopnea index (AHI) (p=0.007 and p=0.004, (Randomized (Randomized (Randomized respectively), oxygen desaturation index (p=0.002 and p=0.04, respectively), individuals, n=423) individuals, n=211) individuals, n=212) and percent of sleep time with oxyhaemoglobin saturation at <90% (ST90) n* Mean n* Mean n* Mean (p=0.002 and p=0.004, respectively). Furthermore, increasing quartiles of Weight change from baseline 317 -6.7 158 -5.0 116 -2.8 ST90 and AHI were associated with increasing levels of A1c and PPG120 to week 56 (%) (p=0.015 and p=0.003, respectively, for ST90; and p=0.035 and p=0.014, Weight change from week 56 303 2.3 146 2.0 113 -0.1 respectively, for AHI). Multiple regression analysis showed that ST90 was to 68 (%) the strongest independent determinant of A1c, after controlling for sex, age, Weight change from baseline 306 -4.7 148 -3.6 114 -2.7 BMI, waist circumference, CRP, HOMA-IR (β=0.442, p=0.007). Similarly, AHI to week 68 (%) persisted as an independent determinant of A1c (β=0.414, p=0.005). Both ST90 and AHI persisted as determinants of PPG120, albeit not signifi cantly FPG change from baseline to 310 -39.1 155 -26.3 115 -7.6 week 56 (mg/dL) (β=0.377, p=0.095; and β=0.358, p=0.119, respectively). In conclusion, OSA acts as an independent factor exacerbating the metabolic risk attributed to FPG change from week 56 to 298 33.6 145 28.0 112 7.5 obesity. Recognition and treatment of OSA may decrease the progression week 68 (mg/dL) toward type 2 diabetes in obese subjects. FPG change from baseline to 300 -5.2 146 -1.7 114 0.8 week 68 (mg/dL) 2030-P Supported By: Novo Nordisk A/S Risk of Development of Type 2 Diabetes by Overall Obesity, Abdom- inal Obesity, and Metabolic Abnormalities—Does Metabolically 2028-P Healthy Obesity Really Exist? Striatal Dopamine Transporter Binding Correlates with Body YORIKO HEIANZA, KIMINORI KATO, AKIKO SUZUKI, CHIKA HORIKAWA, SATOSHI Composition, Energy Expenditure, and Visual Attention Bias for MATSUNAGA, SAKIKO YOSHIZAWA, OSAMU HANYU, SATORU KODAMA, KOJI Food-related Cues SATO, HIROHITO SONE, Niigata, Japan, Mito, Japan KARIN E.M. KOOPMAN, ANNE ROEFS, DANNE C.E. ELBERS, ERIC FLIERS, JAN To clarify whether the concept of healthy obesity would really exist BOOIJ, MIREILLE J. SERLIE, SUSANNE E. LA FLEUR, Amsterdam, Netherlands, in terms of the development of diabetes, we investigated the effect of Maastricht, Netherlands the degree of overall obesity, abdominal adiposity and the number of It is currently believed that a downregulated dopaminergic system in metabolic factors on the development of diabetes. This prospective study obesity stimulates food-motivated behavior in pursuit of reward, but the included 29564 individuals without diabetes. Cut-off points of BMI 23.0, physiological role of dopamine (DA) in energy balance regulation is not 25.0 and 27.5 kg/m2 were used to assess overall obesity; cut-off points of well established. Since motivation for food is an essential evolutionary waist circumference (WC) of 90 cm for men or 80 cm for women defi ned mechanism for survival and lean body mass is a well-known determinant abdominal obesity. Metabolic health was assessed by blood pressure, of survival, we hypothesized that the striatal dopaminergic system might be triglycerides, HDL-cholesterol and fasting glucose concentrations. The involved in maintaining energy balance and lean body mass. presence of metabolic syndrome (Mets) and the number of abnormalities We therefore included 36 lean, healthy, male subjects and measured striatal were assessed by Adult Treatment Panel-III criteria. During a 5-year follow- DA transporter (DAT) binding using 123I-FP-CIT SPECT, body composition using up, 1188 individuals developed diabetes. Even in the absence of Mets, obese bioimpedance analysis, resting energy expenditure (REE), visual attention bias individuals with BMI of 23.0-<25.0, 25.0-<27.5 and ≥27.5 had a 1.40 (95% CI for food and degree of impulsivity using response-latency based computer 1.17, 1.69), 1.78 (1.43, 2.21) or 2.65 (1.95, 3.59) times, respectively, increased tasks. Ad libitum food intake was self-reported online. risk of diabetes than those with BMI <23.0 and with the absence of Mets. Striatal DAT binding correlated with lean body mass (p=0.009, r=-0.47), fat Further, individuals with BMI ≥25.0 even with none of the Mets components mass (p=0.003, r=-0.53) and lean:fat mass ratio (p=0.02, r=0.40) but not with except elevated WC had a 2.46 (1.57, 3.87) times increased risk of diabetes body mass index (BMI; p=0.07). Moreover, striatal DAT binding positively compared with healthy normal weight individuals. However, only 1.1% of correlated with REE expressed as kcal per kg (p=0.02, r=0.42) and with visual total study participants had BMI ≥25.0 but with no component of Mets attention bias for food (p=0.004, r=0.54), but not with impulsivity (p=0.71). including elevated WC. This metabolically healthy obese phenotype did Furthermore, visual attention bias for food positively correlated with ad not show a signifi cantly increased risk of diabetes (odds ratio, 1.53 (0.61, libitum total caloric intake (p=0.002, r=0.55), protein intake (p=0.04, r=0.37) 3.82)). Among overweight or obese individuals, having even ≥1 components and carbohydrate intake (p=0.04, r=0.37). of Mets was not harmless as to the risk for future diabetes. The concept of Based on our results we propose a physiological role for the striatal DA healthy obesity does not appear to be a myth if a metabolically healthy state system in maintaining energy balance by responding to changes in body was defi ned by the absence of any of the components of Mets including composition. When fat and/or lean mass changes, the striatal DA system abdominal obesity. Consideration should be made to defi ne healthy obesity acts to adjust energy expenditure and/or energy intake through modulation with a detailed assessment of the number of metabolic abnormalities. of visual attention to food. As obese subjects have a reduced lean: fat Supported By: Japan Society for the Promotion of Science mass ratio, our proposed mechanism could explain the downregulated dopaminergic system as shown in the striatum of obese subjects. 2031-P PET-MRI Is a Valuable Tool to Detect Brown Adipose Tissue 2029-P KARIM GARIANI, JOANNA GARIANI, GAËL AMZALAG, BÉNÉDICTE DELATTRE, Presence and Severity of Obstructive Sleep Apnea Is Independently OSMAN RATIB, VALENTINA GARIBOTTO, Geneva, Switzerland Associated with Glycometabolic Abnormalities in Obese Non- The purpose of this study was to assess the performance of 18F-FDG PET- Diabetic Subjects MRI to detect the presence of brown adipose tissue (BAT). Integrated ANGELO CIGNARELLI, SEBASTIO PERRINI, ALESSANDRO CIAVARELLA, STELLA We analyzed 197 consecutive 18 F-fl urodeoxyglucose (18F-FDG) positron- POSTERS KOUNAKI, VITALIANO QUARANTA, MARIA BARBARO, ANTONIO DI TRANI, emission tomographic and magnetic resonance imaging (PET-MRI) exams Physiology/Obesity VITO A. FALCONE, ANNALISA NATALICCHIO, LUIGI LAVIOLA, ONOFRIO RESTA, performed for various diagnostic reasons for the presence of brown adipose FRANCESCO GIORGINO, Bari, Italy tissue (BAT). BAT was defi ned as a tissue collection that was more than 4 mm Obstructive sleep apnea (OSA) is a common underdiagnosed condition in diameter, had the characteristics of fat tissue according to MRI and had a in the obese population, and has been associated with worse glycemic maximal standardized uptake value of 18F-FDG of at least 2.0 g per militer, control in individuals with type 2 diabetes. The relationship between OSA indicating high metabolic activity. and glycometabolic parameters was investigated in obese non-diabetic A total of 197 PET/MRI were performed in 192 patients during a period individuals. Ninety-one obese subjects (57% male, mean age 45.3 ± 12 yrs, of 36 months. 105 (54.7%) of these patients were female and 87 (45.3%) mean BMI 42.1 ± 9 kg/m2) underwent polysomnography and a 2-h oral glucose were male. tolerance test (OGTT). OSA was identifi ed in 64% of subjects (73% in male, Brown fat tissue was observed in 5 of the 192 patients (2.6%). Amongst 41% in female, p=0.032 χ2). Obese subjects with OSA showed higher A1c the patients with BAT 5 female (4.8%) and no male (0.0%) with a mean age (5.8% vs. 5.5%, p=0.009), plasma glucose at 120 min during OGTT (PPG120) of 42.4 years range, 23.2-58.5 years.

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A519 OBESITY—HUMANCATEGORY

187 patients did not have BAT on the PET/MRI exam, 100 (53.5%) were In conclusion, hypothyroid patients regularly undergoing LT4 replacement female and 87 (46.5%) were male. The mean age and the mean BMI were therapy showed higher BMI regardless of age, sex and TSH levels in respectively 56.2 years and 24.3 kg/m2. Body weight was signifi cantly lower comparison with MNG patients. In contrast, no difference in the prevalence in the brown fat group than in the group of patients without brown fat (52.4. of the metabolic syndrome and in surrogate biomarkers of fatty liver and kg ± 6.91vs 67.67 kg ± 14.21, p<0.05) as BMI (19.49 kg ± 2.06 vs. 24.34 kg/m2 ± visceral adiposity was observed when the analysis was adjusted for the 4.21, p < 0.05) and age (42.4 ± 13.1 vs. 56.2 ± 16.8). different age and BMI. The isolated fi nding of higher BMI may be due to an In conclusion, we showed that PET/MRI is a radiological tool that can initial weight gain associated to the diagnostic-related delay in beginning detect BAT. To our knowledge this is the fi rst study showing that PET/ LT4 therapy but also to persistent subtle defects of energy metabolism not MRI allows to visualize BAT in humans. Further studies are needed to fully reverted by the LT4 therapy. better understand the possibilities of PET/MRI in detecting BAT and the Supported By: Family of Angela Musazzi and Mario Stellato combination of PET with new sequence of MRI such as Dixon may be an interesting way to improve the evaluation of BAT presence or activity. PET/ 2034-P MRI could be used in a protocol evaluating new pharmaceutical therapies Chromosome 3 Locus Associated with Fetal Adiposity Is Not that activate BAT with the advantage of avoiding irradiation. Associated with Childhood Adiposity WILLIAM L. LOWE, DAVID R. MCCANCE, IAN S. YOUNG, CHRISTOPHER C. 2032-P PATTERSON, JANANI RANGARAJAN, LOREN L. ARMSTRONG, LYNN P. LOWE, Whole Body Energy Metabolism in Metabolically Healthy Obese DENISE M. SCHOLTENS, GEOFF HAYES, BOYD E. METZGER, Chicago, IL, Belfast, (MHO) Subjects Ireland ALICE OLTOLINI, GIUSEPPINA MANZONI, FEDERICO MARTUCCI, SIMONA VILLA, High birth weight and newborn adiposity are associated with adverse GUIDO LATTUADA, GIANLUCA PERSEGHIN, Monza, Italy metabolic outcomes from childhood onwards. The intrauterine environment Metabolically healthy obese (MHO) individuals do not manifest the and fetal and maternal genes contribute to fetal birth weight and adiposity. obesity-related metabolic complications associated with insulin resistance. Through genome wide association studies (GWAS), we and others identifi ed To test whether MHO subjects are characterized by adaptation of whole a locus between CCNL1 and LEKR1 on chromosome 3 (3q25.31) associated body energy metabolism 71 obese individuals consecutively admitted with birth weight which, in our study, was strongly associated with newborn to measure resting energy expenditure (REE) and body composition were sum of skinfolds. These data suggest this locus impacts fetal adiposity, but studied using indirect calorimetry and bioimpedentiometry (BIA). They were whether it is also associated with childhood adiposity is unknown. To address defi ned as MHO (n=30) or unhealthy obese (MUHO: n=41) subjects based this, we used phenotype data collected during childhood in a European on the presence of the metabolic syndrome (ATP III defi nition). MHO were ancestry cohort (Belfast, UK) who participated in the Hyperglycemia younger and had lower blood pressure, transaminases, γGT, uric acid, and Adverse Pregnancy Outcome (HAPO) GWAS study described above. triglycerides and creatinine and higher HDL-cholesterol and platelet count, Genotype and sum of skinfolds (fl ank, subscapular and triceps) data were when compared to MUHO. BIA-derived parameters of body composition did available for 293 children at birth and age 2y and for 350 children at birth not differ between groups meanwhile HOMA-IR was lower in MHO when and age 6y. Linear regression analyses with SNPTEST under an additive compared to MUHO (3.63±2.91 vs. 6.24±4.58; p<0.05) because of both model adjusting for known confounders (ancestry, parity, smoking, drinking, lower fasting serum glucose (92±12 vs. 115±18 mg/dL; p<0.001) and insulin maternal age, BMI, height and MAP at ~28 wks gestation) were performed. (16±8 vs. 22±10 µU/m; p<0.05) concentrations. Respiratory quotient was We examined 4 SNPs at 3q25.31 strongly associated with newborn sum of not different between groups (0.89±0.06 vs. 0.91±0.07; p=0.58) meanwhile skinfolds (lead SNP T allele of rs17451107; p = 1.9 x 10-13) in the HAPO GWAS the ratio between the measured REE and predicted REE (based on Harris study. Among the 293 children in the Belfast cohort with data both at birth Benedict equation) was lower in MHO when compared to MUHO (89±9 vs. and 2y and 350 children with data at birth and 6y, a positive association was 94±11%; p=0.03). The reduced energy metabolism in MHO was more evident observed between the T allele of rs17451107 and newborn sum of skinfolds when the analysis was restricted to the subgroups of those with BMI ≥ 40 (p = 3.1 x 10-4 and p = 1.1 x 10-4, respectively). In contrast, at 2y, a negative kg/m2 (n=13 in MHO e n=21 in MUHO) in which whole body REE (1712±293 association of nominal signifi cance (p = 0.16) was observed between sum of vs. 2022±353 Kcal/day; p=0.01), the ratio between the measured REE and skinfolds and the T allele of rs17451107, while at 6y, there was no evidence predicted REE (85±11% vs. 98±12%; p<0.001) and the REE adjusted for kg of of association (p = 0.68). These data demonstrate that an intergenic locus free fat mass (28.5±3.2 vs. 33.4±6.7 Kcal/kg of FFM/day; p<0.05) were lower on chromosome 3 previously shown to be strongly associated with newborn in MHO when compared to MUHO. adiposity is no longer associated with adiposity by age 2y, suggesting that In conclusion, MHO subjects are characterized by lower whole body REE its impact may be largely limited to fetal fat accretion. when compared to MUHO subjects and this feature is more pronounced in Supported By: NIH (R01DK099820-01) patients with a more severe degree of obesity. Supported By: Family of Angela Musazzi and Mario Stellato 2035-P Elevated Levels of GROα Impair Capillary Formation via Regulation 2033-P of MMP-9 in Adipose Tissue from Type-2 Diabetic Subjects Obesity and Metabolic Syndrome in Patients with Primary Acquired YIFAT AMIR LEVY, THEODORE P. CIARALDI, SUSAN A. PHILLIPS, SUNDER R. Hypothyroidism Undergoing Chronic LT4 Replacement Therapy MUDALIAR, ROBERT R. HENRY, La Jolla, CA, San Diego, CA GIUSEPPINA MANZONI, FEDERICO MARTUCCI, ALICE OLTOLINI, SIMONA Obesity and Type 2 Diabetes (T2D) are associated with not only insulin VILLA, DARIO ZIMBALATTI, GUIDO LATTUADA, EMANUELA ORSI, GIANLUCA resistance, but impaired capillary density of adipose tissue (AT), as well. PERSEGHIN, Monza, Italy, Milan, Italy Impaired vascularization limits the availability of insulin and glucose, amongst Hypothyroidism is a risk factor for obesity, central adiposity and ectopic many other factors, and exacerbates T2D. IL8, GROα and IL15 are myokines fat accumulation. To assess whether this risk could be detected also in that are known to modulate angiogenesis; our group recently reported that stable patients taking regularly the LT4 replacement therapy we assessed T2D myotubes (MT) secrete higher levels of all 3 factors when compared to retrospectively the prevalence of metabolic syndrome (ATP III defi nition) non-diabetic (ND) MT. Studies were conducted to determine how the secretory and calculated the Fatty Liver Index (FLI) and the Visceral Adiposity Index performance of myokines are linked to vascularization of AT. Integrated POSTERS (VAI) in patients attending our Outpatient Endocrinology Clinic because We hypothesized a direct impact of myokines secreted from skeletal of known primary acquired hypothyroidism undergoing LT4 replacement muscle on AT vascularization with potentially negative effects exerted by Physiology/Obesity therapy on a regular basis (n=997) and used as control group patients with the higher levels produced by diabetic muscle. To test our hypothesis, a stable euthyroid multi-nodular goitre (MNG; n=502). Hypothyroid patients mixture of exogenous recombinant (r) IL8, GROα and IL15 was added to AT were younger (56±15 vs. 60±14 years; p<0.001), and showed higher BMI explants embedded in matrigel in concentrations equal to the averages of (28.7±6.4 vs. 27.2±5.3 kg/m2; p<0.001) and FLI (58±31 vs. 49±32; p=0.02) but those found in conditioned media from T2D vs. ND MT (T2D-IL8=3280 pg/ml no difference in VAI (4.6±3.1 vs. 4.6±3.5) and prevalence of the metabolic vs. ND-IL8=2071 pg/ml, Groα-4620 pg/ml vs, 1859 pg/ml and IL15-1.07 pg/ syndrome (47% vs. 47%), neither in the prevalence of type 2 diabetes (11% ml vs. 0.62 pg/ml), for 24h-168h and evaluated for capillary sprout formation vs. 11%) and known CVD (11% vs. 12%) when compared to MNG patients. and MMP-9 content. When AT explants were treated with the mixture of The actual BMI was directly associated with the duration of LT4 replacement r IL8, GROα and IL15 [ND] vs. [T2D], no consistent effects were observed therapy also when adjusted for age, sex and average TSH levels (R=0.30; in T2D AT (n=4). Next, individual myokines were added to T2D AT explants ρ=0.03). No difference in metabolic syndrome, FLI and VAI was detected at ND and T2D concentrations. No consistent effects were found with IL8 when the comparison between groups was adjusted for age, sex and BMI. (n=5) or IL15 (n=2) alone, but T2D levels of GROα impaired sprout formation

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A520 OBESITY—HUMANCATEGORY in T2D AT explants when compared to ND levels (n=5, p<0.01 from 72 h). Western blot analysis showed higher expression levels of MMP-9 secreted Table. Annualized Percent Increase or Decrease in Lower Leg Skeletal Muscle Composition by Race/Ethnicity*. from T2D AT explants treated with ND levels of GROα (n=2, p<0.01), that could support elevated vascularization. Taken together, the data indicate Lower leg skeletal Annualized increase or decrease % P-value for differences that skeletal muscle has the capacity to communicate directly with AT to muscle composition (95% CI) by race/ethnicity regulate vascularization, possibly via MMP-9. Diabetic muscle appears to Caucasian men Afro-Caribbean men impair these properties. (N = 229) (N = 228) Supported By: U.S. Dept. of Veterans Affairs TAT (mm2) 1.46 (-0.21, 3.13) 2.4 (0.87, 3.95) 0.027 SAT (mm2) 1.80 (0.09, 3.51) 1.92 (0.33, 3.51) 0.89 2036-P IMAT (mm2) 8.80 (0.80, 16.80) 7.53 (0.05, 15.0) 0.76 Is Defi nite Metabolically Healthy Obesity Really Healthy? A Cross- Muscle Attenuation (mg/cm3) -0.05 (-0.46, 0.36) -0.88 (-1.29, -0.47) 0.0002 Sectional Study EMMANUEL COSSON, ELIANE HAMO-TCHATCHOUANG, CAMILLE CUSSAC- Muscle Area (mm2) -1.99 (-2.71, -1.28) -0.27 (-0.93, 0.39) <0.0001 PILLEGAND, ISABELA BANU, MINH TUAN NGUYEN, SABRINA CHIHEB, PAUL *Values in italics indicate that changes are signifi cantly different from zero (P < 0.05). VALENSI, Bondy, France Supported By: NIH Patients displaying the metabolically healthy but obese (MHO) phenotype have an intermediate cardio-metabolic prognosis as compared with normal weight healthy individuals and metabolically unhealthy obese (MUO) 2038-P patients. We aimed to evaluate the prevalence and characteristics of defi nite Change in A1C in Overweight (OW) Patients with Type 2 Diabetes MHO phenotype in a large sample of obese patients. with the Healthy Lifestyle Program (HLP) Used in the Lorcaserin Defi nite MHO phenotype was defi ned as having one or more Inter- (lor) Phase 3 Trials national Diabetes Federation metabolic syndrome criteria in addition to KENNETH FUJIOKA, RANDI FAIN, WILLIAM SOLIMAN, YUHAN LI, SCOTT waist circumference criterion. We included 1159 patients (943 women) STUBBE, JOANNE QUAN, DONNA RYAN, La Jolla, CA, Woodcliff Lake, NJ, San consecutively admitted in stable conditions for obesity (body mass index Diego, CA, Baton Rouge, LA (BMI) 38.4±6.3 kg/m2). All of them were free of known diabetes. Patients A1C and comorbidity risk in patients (pts) with type 2 diabetes (T2D) were characterized for cardio-metabolic disorders. improves with 5%-7% weight loss (WL), supporting a weight-centric The 202 (17.4%) MHO patients were younger and had lower BMI than the management of T2D as the standard of care to improve glycemic outcomes. 957 MUO patients. The MHO patients were matched for gender, age and BMI The AACE recommends lifestyle intervention targeting WL for OW T2D pts, with 404 MUO patients. In addition to the features of metabolic syndrome, but this can be challenging in outpatient settings. T2D pts enrolled in a phase the MHO patients as compared with the MUO ones had lower prevalences of 3 trial of lor participated in the HLP, a program designed for pts with diabetes insulin resistance (HOMA-IR ≥ 3: 23.6 versus 38.9%, p<0.001), of abnormal like that described by AACE. This report focuses on outcomes for the HLP + oral glucose tolerance test (13.9 vs. 23.9%, p<0.001), of HbA1c ≥ 5.7% (43.9 placebo (pbo) group in BLOOM-DM, the study specifi c to OW pts with T2D, to vs. 54.2%, p<0.05) and pulse pressure ≥ 60 mmHg (0 vs. 59.4%, p<0.001) but assess the impact of the HLP, which may serve as a support model applicable non signifi cant difference for the prevalences of microalbuminuria (11.1 vs. to outpatient T2D clinical care. 12.3%), cardiac autonomic dysfunction (20.9 vs. 10.8%) and fatty liver index T2D pts aged 18-65 yrs (N=603) with a BMI of 27-45 participated in ≥60 (5.6 vs. 10.2%); and even a higher prevalence of LDL cholesterol ≥ 4.9 the HLP and received pbo or lor, a selective 5HT2C agonist approved for mmol/L (4.0 vs. 0.6%, p<0.01). chronic weight management in adults as adjunct to a reduced-calorie diet Our data do not support the characterization of MHO, even defi nite, as and increased physical activity. The HLP was delivered to outpatients via really healthy, as this phenotype does not exclude abnormal cardio-vascular one-to-one 15-min consultations with a trained counselor twice monthly in markers and glucose and liver abnormalities. the fi rst month, and then monthly thereafter for 1 year. Pts received support materials and were counseled to exercise 30 min/day and to reduce intake to 600 kcal/day below their estimated energy requirements (using WHO 2037-P criteria). WL over time and glycemic parameters were monitored. Analysis is Longitudinal Assessment of Skeletal Muscle Composition in White for the MITT population (LOCF) treated with HLP alone (n=248). vs. Black Elderly Men HLP + pbo resulted in adjusted mean percent changes in weight of -1.5%, IVA MILJKOVIC, TANUSHREE PRASAD, JANE A. CAULEY, YAHTYNG SHEU, adjusted mean change in A1C of -0.4(%), and in FPG of -11.9(mg/dL) for CHRISTINE G. LEE, PEGGY M. CAWTHON, ERIC S. ORWOLL, TIEN DAM, CLARE- patients with T2D. For pts in this group, 16.1% had 5% and 4.4% had 10% ANN H. BUNKER, ALAN L. PATRICK, JOSEPH M. ZMUDA, Pittsburgh, PA, Portland, WL. For those with 7% WL (10.1%) adjusted mean A1C and FPG reductions OR, San Francisco, CA, New York, NY, Scarborough, Trinidad and Tobago were 1.0(%) and 33.4(mg/dL), respectively, after 52 wks. Adipose tissue (AT) infi ltration in skeletal muscle (myosteatosis) is greater The HLP program is adaptable to outpatient clinical settings and is in black compared to white men in cross-sectional studies and may play an associated with reductions in WL, A1C, and FPG in OW pts with T2D. This important role in the development of type 2 diabetes (T2D). Longitudinal behavior-modifi cation program may be useful for practitioners who provide studies examining myosteatosis with aging are lacking. We examined care to OW and obese patients with T2D. longitudinal changes in computed tomography measures of the lower leg skeletal muscle composition including total AT (TAT); subcutaneous AT (SAT); inter-muscular AT (IMAT); muscle attenuation (lower muscle attenuation 2039-P is associated greater intra-muscular AT); and muscle area. Data are from Lorcaserin (lor) Free Plasma Levels at Recommended Dose Are the Osteoporotic Fractures in Men Study (2 years follow-up in white men Suffi cient to Activate 5-ht2c but Not 2a or 2b Receptors aged 65+) and black men from Tobago Bone Health Study (6 years follow-up ERIC RAVUSSIN, YUHAN LI, DAVID UNETT, MICHAEL MORGAN, WILLIAM in black men aged 65+). TAT increased with aging among black men only, SHANAHAN, WILLIAM SOLIMAN, Baton Rouge, LA, Woodcliff Lake, NJ, San whereas muscle area decreased with aging in white men only. Men of both Diego, CA race groups gained SAT and IMAT to a similar degree. In contrast, skeletal Appetite is controlled by multiple signaling pathways, including 5-HT Integrated muscle attenuation decreased to a greater extent in black men. These (serotonin) through the 5-HT2CR which is involved in food intake regulation POSTERS changes were independent of baseline age, physical activity, smoking, and may be linked to glucose homeostasis. Lor, a selective 5-HT2CR agonist Physiology/Obesity alcohol intake, diabetes status, and annualized change in total body fat and (EC50=39nM, 7.6ng/mL), is approved for chronic weight management as an muscle area. Our results illuminate potential differences in skeletal muscle adjunct to lifestyle modifi cation in obese (O) and overweight (OW) patients with composition with aging between black and white men. Additional studies a weight-related comorbidity. Lor is believed to decrease weight in humans via are needed to test if aging-related changes in skeletal muscle attenuation food intake reduction, not increased energy metabolism. In vitro, in studies contribute to ethnic/race differences in T2D risk. controlled for receptor reserve, lor is 14-and 61-fold more selective for the 5-HT2CR vs. 5-HT2AR (associated with mood, perception, and vascular/platelet effects) or the 5-HT2BR, (associated with cardiac valvular insuffi ciency). Here we compare in vivo plasma free lor and estimated brain concentrations and in vitro EC50 values needed for activation of 5-HT2 receptors. Lor plasma concentrations were measured pre-dose and in 2 time windows post-dose (1.5-2.5h and 3.5-6h) at week 12 at a subset of phase 3 study sites.

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A521 OBESITY—HUMANCATEGORY

Free plasma and estimated brain concentrations at the approved dose of multidisciplinary glycemic management (GMP) pathway that recommends 10mg bid in O/OW subjects (N=512) were overlaid with in vitro EC50 (nM) documentation of HbA1C 3 months before and 6-12 months after BS, standard potency data for 5-HT2A, 2B and 2C receptors - calculated by inositol phosphate correction insulin orders immediately post-BS for BG>140 mg/dl, basal insulin and calcium release assays. for persistent hyperglycemia (BG>180 mg/dl x 3), a prescription for metformin In vivo free lor concentrations achieved EC50 for 5-HT2C, but did not achieve at discharge, referral to endocrinology or PCP for post-BS A1C>6.5%, and EC50 for 5-HT2A and 5-HT2B receptors. education of surgical staff. We report the results investigating feasibility and effi cacy of the pre-BS and inpatient aspects of this GMP. Glycemic data was compared among patients undergoing BS before (median age 55 yrs, 84% female, pre-BS A1C 7.1 ± 1.5%) and following (median age 50 yrs, 81% female, pre-BS A1C 7.1 ± 1.1%, 29% on insulin) implementation of the protocol. Improvements were observed in frequency of documentation of pre-BS A1C (38% vs. 98%, p <0.001), and the % of hyperglycemic patients discharged with basal insulin and/or metformin (8.7% (2/23) vs. 73.7% (14/19), p<0.001). In summary, an inpatient BS glycemic management pathway (GMP) increases the % of patients who have an A1C measured and the % with post- BS hyperglycemia discharged home on scheduled diabetes medications. We demonstrate that a BS GMP is feasible and effi cacious for improving the attention to GM of these patients. We plan to investigate safety (i.e. frequency of hypoglycemia) and long-term effi cacy. Although the benefi t of our protocol has to be proven, a system wide multidisciplinary protocol can change the patterns of care in targeted areas to ultimately improve long-term diabetes outcomes.

2042-P Association of Metabolically Healthy Obesity (MHO) with Sub- clinical Coronary Atherosclerosis in a Korean Population: Is MHO 2040-P A Really Benign Condition? Early, Rapid Weight Loss and Very Low Calorie Diet After Bariatric MIN JUNG LEE, YU MI KANG, CHANG HEE JUNG, KEE-HO SONG, SUNG RAE KIM, Surgery Uncouples Changes in Total and HMW Adiponectin CHEOL-YOUNG PARK, SUNG WOO PARK, JOONG-YEOL PARK, Seoul, Republic of LAUREN K. MEYER, ALAN C. WITTGROVE, TRACY MARTINEZ, THEODORE P. Korea, Bucheon, Republic of Korea CIARALDI, ROBERT R. HENRY, SUSAN A. PHILLIPS, San Francisco, CA, La Jolla, The long-term outcome of metabolically healthy obesity (MHO) has been CA, San Diego, CA investigated by several studies, however, the results were confl icting. Bariatric surgery reduces body fat mass and increases total and HMW In this study, we aimed to compare the degree of subclinical coronary adiponectin (Ad). Information is limited regarding the impact of diet and acute atherosclerosis detected in coronary MDCT among groups defi ned by state weight loss (WL) on total and multimeric Ad and leptin. We prospectively of metabolic health and obesity in an asymptomatic Korean population. We enrolled 6 obese subjects (BMI 38±3.4) undergoing elective Roux-en-Y collected the data of asymptomatic 4,009 subjects (mean age 53.2 yr) who gastric bypass surgery for WL. Following surgery subjects adhered to a very participated in a routine health screening examination of Asan Medical low calorie diet (VLCD, <500 kcal/day) for week 1 then progressed to an Center (Seoul, Korea). Signifi cant coronary artery stenosis defi ned as >50% ad lib diet at week 16. Anthropometric and biochemical assessments were stenosis, coronary artery plaque and coronary artery calcium scores (CACS) made at 0, 1, 2, 4, 8, 16, 24, and 52 weeks to study acute versus chronic were assessed by MDCT. Participants were stratifi ed by BMI categories effects of WL. All subjects reached a BMI<30 by 24 weeks. By 52 weeks (25 mg/m2 as the cut off value), and metabolic health state defi ned by the subjects achieved a 37.5±5% WL (P=0.001), reduced waist circumference Wildman criteria. The prevalence and risk of coronary artery atherosclerosis (34.4±3%, P=0.0003) and leptin (92±3%, P<0.0001), and increased total and were analyzed according to the metabolic health and obesity, i.e., meta- HMW Ad (232±54%, P=0.01; 230±27%, P=0.001). Levels of leptin, total and bolically healthy non-obese (MHNO), metabolically unhealthy non-obese HMW Ad correlated with WL (P=0.002, r=-0.90; P=0.002, r=0.91; P=0.001, (MUNO), MHO, and metabolically unhealthy obese (MUO). Subjects with r=0.92), as did %HMW and %LMW Ad (P=0.03, r=0.75; P=0.02, r=0.79). MHO had signifi cantly higher prevalence of signifi cant subclinical coronary Acute effects of WL and VLCD were assessed at 1 week. The most rapid atherosclerotic burden compared with MHNO. Overall, MHO subject WL occurred acutely with VLCD (P<0.001 vs. all other diet intervals). Acute showed intermediate coronary MDCT fi ndings between MHNO and MUNO WL was 5.9±0.1% (P<0.0001) and predictive of WL at 24 weeks (P=0.05, or MUO. The odds ratios of MHO group for various coronary MDCT fi ndings r=0.80). Leptin decreased most rapidly (P=0.005) during the acute 5.9% (MHNO group as the reference) such as coronary artery stenosis, any plaque, WL, with a 55.7±10% reduction (P=0.003). In contrast, a chronic 31.5% WL calcifi ed plaque, mixed plaque, CACS>0, and CACS>100 (CACA=0 as the was associated with only a 36.4% reduction in leptin and a reduced rate of reference) were 1.92 (95% CI 1.18-3.11), 1.33 (1.02-1.72), 1.39 (1.05-1.85), decline (P=0.01). Signifi cantly, during acute WL and VLCD, HMW increased 1.61 (1.02-2.53), 1.38 (1.05-2.53) and 1.66 (1.01-2.73), respectively, even after (P=0.02) despite a reduction in total Ad (P=0.1), increasing the SA ratio adjustment for conventional cardiovascular risk factors. Our data suggested (HMW/total, P=0.05). Chronic WL, which occurred at a signifi cantly slower that MHO might not be a benign disease in terms of coronary atherosclerotic rate (P<0.006), on the ad lib diet was associated with increases in total Ad burden. Thus, it is important to consider obesity and metabolically health (P=0.01). Although surgically-induced WL improved total and HMW Ad over state in evaluating cardiovascular risk. 52 weeks, acute rapid WL achieved with VLCD resulted in selective elevation Supported By: NRF (2013R1A1A1004798) of HMW. Early improvements in metabolic function following gastric bypass surgery may relate to early and selective increases in HMW Ad. 2043-P Integrated POSTERS Supported By: U.S. Dept. of Veterans Affairs Loss of Sympathetic Drive May Explain Loss of Brown Adipose Tissue Activity in Elderly but Not in Obese Males Physiology/Obesity 2041-P LONNEKE BAHLER, WANDA ADMIRAAL, HEIN J. VERBERNE, MAARTEN R. Implementation of Glycemic Management Pathway after Bariatric SOETERS, JOOST HOEKSTRA, FRITS HOLLEMAN, Amsterdam, Netherlands Surgery: A Feasible and Effi cacious Protocol Metabolically active brown adipose tissue (BAT) could facilitate weight SANN YU MON, MARY T. KORYTKOWSKI, DAVID ROMETO, Pittsburgh, PA loss by increasing energy expenditure. Cold is a potent stimulator of BAT, Bariatric surgery (BS) is associated with remission of type 2 diabetes activating BAT primarily through the sympathetic nervous system (SNS). (T2D); however, long-term remission rates may occur in only 24% of Old or obese individuals have less metabolic BAT activity than lean and patients. Early attention to post-BS glycemic management with glargine and young, but the role of the SNS in this decrease is unknown. We determined metformin has been reported to increase the % of patients experiencing whether the lower metabolic BAT activity can be explained by a lower SNS long-term remission. response to cold. To address this, we conducted a retrospective analysis of patients who We studied 10 young obese (26 [21-31] years, BMI 32 [31-39] kg/m2), underwent BS prior to (n = 50) and following (n = 59) implementation of a 10 old lean (55 [51-60] years, BMI 23 [22-25] kg/m2) and 14 young lean (26

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A522 OBESITY—HUMANCATEGORY

[21-28] years, BMI 22 [21-23] kg/m2) males. Metabolic BAT activity was Group with ALT <30 IU/L: South Asians had signifi cantly lower mean measured as maximal standardised uptake value and volume (SUVmax, brachial and ankle pressures [R PT, 146 vs. 156 mmHg]. ALT ≥ 30: South SUVvol) on 18F-Fluorodeoxyglucose positron emission tomography CT Asians had signifi cantly lower brachial pressures with similar use of (FDG). SNS activity was measured as semiquantative uptake values of antihypertensive medications and cholesterol lowering medications, but 123I-metaiodobenzylguanidine (MIBG) on single photon emission computed their ankle pressures were not signifi cantly different to those of nine years tomography scans, with the mediastinum as reference region. Scans were older Europeans [R PT, 151 vs. 159 mmHg]. made after an overnight fast and 2 hours of cold exposure. Increase in ankle pressures and cardiovascular diseases is signifi cantly Metabolic BAT activity and volume were different between young vs. greater with increased metabolic obesity in South Asians than Europeans, old (median [IQR] SUVmax (g/L) 8.0[4.7-17.4] vs. 3.4[2.8-4.2] p<0.01, SUVvol therefore prospective studies to determine predictive value of increased (cm3) 104.8[15.1-303.9] vs. 8.9[2.7-21.3] p<0.05) but unexpectedly, not for absolute ankle blood pressures independent of brachial pressures along lean vs. obese (SUVmax (g/L) 8.0[4.7-17.4] vs. 6.3[3.5-18.2] p=0.7, SUVvol with WHtR as non-invasive screening tools for cardio metabolic diseases in (cm3) 104.8[15.1-303.9] vs. 145.6[3.9-296.0] p=0.9). young apparently healthy South Asians are necessary. HOMA-IR differed between lean and obese (p<0.01) but not for young vs. old. There was no correlation between HOMA-IR and either SUVmax or SUVvol. There were no differences in the SNS activity in BAT between lean and obese males (p=0.2) but for young and old males this was borderline signifi cant (p=0.058). We confi rmed the strong positive correlation between BAT activity measured with FDG and MIBG in the whole group (spearman correlation ρ=0.71, p<0.01), which we previously showed. We conclude that loss of sympathetic drive may explain the loss of BAT activity in elderly but not in obese males. Supported By: University of Leeds

2044-P 2046-P Six-Year Follow-up after Nonsurgical and Surgical Antiobesity GLP-1 Analogues Treatment Was Not Associated with Additional Treatments Weight Loss Over Intensive Lifestyle Intervention in Severely Obese MAJA J. EKMAN, FREDRIK NORSTRÖM, ANNE-MARIE RÄIHÄ, MARIA KLINTEN- Patients with Type 2 Diabetes BERG, MARTIN RIDDERSTRÅLE, Malmö, Sweden ALICE WRIGHT, PUNITH KEMPEGOWDA, GILLIAN ABERNETHY, HAMED SADIQI, Maintenance of weight loss and reduction of co-morbidities such as type CHARLOTTE R. PHILIPS, JANE MCLEASE, REBECCA GOLICS, ELIZABETH 2 diabetes (T2D) are major goals for anti-obesity care. In light of the current MAXWELL, REGGIE JOHN, JAYDAVE SHAKHER, MARTIN J. STEVENS, SRIKANTH obesity epidemic there is a need for affordable, and scalable intervention BELLARY, ABD A. TAHRANI, Birmingham, United Kingdom strategies. Clinical trials of GLP-1 analogues in patients with type 2 diabetes (T2D) We performed a questionnaire-based observational long-term follow- typically exclude patients with a BMI > 40 kg/m2 and do not include intensive up study after different weight loss treatments carried out at an obesity lifestyle intervention (ILI). We aimed to assess the impact of GLP-1 analogues outpatient unit. 1,130 subjects were included; response rate was 67%. on weight in severely obese patients with T2D undergoing ILI. The proposed treatment alternative was based on patient preference and We conducted an observational cohort study of severely obese adults with medical history, and was carried out either as therapy in groups (GT; n=285) T2D attending ILI at a tertiary bariatric center for 12 months. All patients or individually (IT; n=156). When suitable, GT could be initiated by three who received GLP-1 analogues for at least 6 months (GLP+) were included month on low calorie diet (GT-LCD; n=137). Eligible patients could be referred and were age, gender and BMI matched to patients with T2D attending the to bariatric surgery (gastric bypass; GBP; n=151) at any time during or after same ILI who did not receive GLP-1 (GLP-). completing a program. Follow-up time was 6.1±0.1 years (mean±SEM). 155 patients were included (121 GLP+ and 34 GLP-) with a median BMI 45.0 At follow-up, the GBP group had lost most weight; BMI 46.1±0.6 to (IQR 42.0-50.6) kg/m2. GLP-1 analogues treatment duration was 10.3 ± 2.8 32.2±0.6 kg/m2, vs. BMI 40.9±0.3 to 38.5±0.3 kg/m2 for the non-surgical months. Patient characteristics are summarised in Table 1. treatment (NST) subgroups (p<0.001). Both the largest NST loss and regain There was no difference in weight loss between groups (GLP- vs. GLP+) of weight was seen in the GT-LCD subgroup; a maintained loss of 3.6, 4.9, (-7.8 ± 8.2 vs. -7.6 ± 8.7 kg, p=0.9; as a % of baseline weight -6.1 ± 6.2% and 7.6% for IT, GT, and GT-LCD, respectively (p=0.006); 57% maintaining a vs. -5.9 ± 6.8%, p=0.9). After adjusting for baseline weight, and the use of >5% loss of body weight at follow-up in the GT-LCD group. Incident T2D was insulin, orlistat, sulphonylureas, DPP-4 inhibitors and thiazolidinediones, lower (2.6% [-3.5-5.6] vs. 9.2% [6.6-11.8], [95% CI]; p=0.02), and remission GLP-1 analogues treatment was not associated with weight change (B= -1.4, of T2D was greater (52.8% [35.6-69.9] vs. 9.2% [6.5-11.1], [95% CI]; p<0.001) p=0.5). for GBP vs. NST. Nonetheless, T2D remission in the GT-LCD and GT groups In conclusion, GLP-1 analogues did not offer additional weight loss over specifi cally were 17.2% (2.6-31.9 [95% CI]; p<0.001.) and 7.0% (4.7-10.0 [95% and above that achieved by ILI in severely obese patients with T2D. Further CI]; p=0.04) .The increase in T2D prevalence in the NST group was only 5.6% studies are needed. (1.2-10.0 [95% CI]; p=0.01). Table 1. Baseline Characteristics of Patients in the GLP+ and GLP- Groups. Bariatric surgery fulfi ls promises from academic trials in clinical practice. However, only a minority of patients will or should be offered surgery. GLP- GLP+ P value For those, non-surgical treatments may offer long-term effectiveness, Age (years) 53.7 ± 10.8 52.6 ± 10.0 0.6 particularly concerning the risk of developing, and the chance of curing, BMI (kg/m2) 46.4 (43.0-50.6) 45.0 (41.7-50.8) 0.4 diabetes. HbA1c (%) 8.0 ± 2.0 8.5 ± 1.7 0.3 Weight (kg) 132.4 ± 25.2 127.9 ± 22.6 0.3 2045-P

Women 70.6% 62.8% 0.4 Integrated Metabolic Obesity and Ankle Pressures POSTERS KIRTI KAIN, THOMAS STEWART, TASNEEM ISHFAQ, SETOR KUNUTSOR, NAKUL Smoking (current) 13.8% 16.1% 0.4 Physiology/Obesity KAIN, JOHN WALLEY, Leeds, United Kingdom, Norwich, United Kingdom Metformin 79.4% 86.8% 0.3 Obesity per se is not equivalent to poor cardio metabolic status. The Sulphonylureas 35.3% 14.9% 0.01 increase in ankle pressures and cardiovascular diseases with type 2 diabetes is greater in South Asians than Europeans. Outcome measures in Thiazolidinediones 11.8% 0.8% 0.008 case control study were bilateral systolic pressures of posterior tibial (PT) DPP-4 inhibitors 17.6% 0% <0.001 and dorsalis pedis arteries based on waist height ratio (WHtR)

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A523 OBESITY—HUMANCATEGORY

2047-P 2049-P Subclinical Vascular Infl ammation in Subjects with Normal Weight Lactate/Pyruvate Production and/or Utilization Are Altered by Obesity and Its Association with Body Fat: An 18F-FDG-PET/CT Study Gastric Bypass Surgery SHINAE KANG, CHANHEE KYUNG, JONG SUK PARK, SOHEE KIM, SEUNG-PYO TERRY E. JONES, MORGAN M. PEARCE, MELISSA A. REED, CHARLES J. TANNER, LEE, MIN KYUNG KIM, HYE KYOUNG KIM, HAERI BAEK, SEOHUI LEE, JI SUN JOSEPH A. HOUMARD, ROBERT C. HICKNER, WALTER J. PORIES, G. LYNIS NAM, SEHEE JO, JIWOON KIM, JAEYOUNG CHEON, KYUNG RAE KIM, TAE JOO DOHM, Greenville, NC, West Chester, PA JEON, CHUL WOO AHN, Seoul, Republic of Korea Results of a large longitudinal study demonstrated that fasting blood lactate Although body mass index (BMI) is the most widely accepted parameter was the best predictor of subsequent development of diabetes. Previously, we defi ning obesity, recent studies have reported the existence of subjects with studied diabetic patients before and after gastric bypass surgery and found normal BMI and excess body fat (BF), termed normal weight obesity (NWO). that fasting lactate and glucose were high before surgery but both dropped Increased BF is an established risk factor for atherosclerosis. However, it is after surgery. From this we proposed that in type 2 diabetes there is a “vicious” unclear whether NWO subjects have a higher risk of vascular infl ammation, Cori cycle of increased lactate/pyruvate production in muscle and increased and the association of BF with vascular infl ammation in NWO subjects is conversion of lactate/pyruvate to glucose in liver. We further proposed that largely unknown. this cycle is disrupted after gastric bypass surgery by increased lactate/ Equal numbers of NWO and normal weight lean (NWL) subjects (n=82 per pyruvate oxidation in muscle and liver which may play a role in remission of the group) were consecutively identifi ed from the database of a self-referred diabetes. To investigate this hypothesis we measured lactate in blood samples Healthcare Promotion Program. The mean/maximal target-to-background taken from non-diabetic patients during an intravenous glucose tolerance test ratios (TBRmean/max) of the carotid artery were evaluated using 18F-FDG- (IVGTT; injection of glucose followed by injection of insulin at 20 min). Patients PET/CT. NWO subjects exhibited a higher TBR than NWL subjects (TBRmean: underwent IVGTT 1 week prior to gastric bypass or gastric band surgery and 1.33±0.16 versus 1.45±0.19, p<0.001; TBRmax: 1.52±0.23 versus 1.67±0.25, again at 1 week post-surgery. The area under the lactate curve (AUC) starting p<0.001). TBRmax was signifi cantly associated with age (r=0.165, p=0.035), at fasting and ending at minute 100 was calculated. The AUC was 64% lower BMI (r=0.206, p=0.008), fasting glucose level (r=0.182, p=0.020), and in gastric bypass patients post-surgery compared to pre-surgery (43.07±10.12 total BF (r=0.289, p<0.001). In multiple linear regression analysis, BF mM·min pre vs. 15.51±3.81 mM·min post, p=0.03). Gastric band surgery, a was an independent determinant for TBRmean and TBRmax (regression purely restrictive operation, did not alter lactate/pyruvate production and/or coeffi cient=0.020/0.028, p=0.008/0.005 for TBRmean/TBRmax). NWO was utilization. Insulin sensitivity index did not change as a result of either gastric associated with elevated TBRmax after adjustment for confounding factors bypass or gastric band surgery. Both gastric bypass patients and gastric band (odds ratio=2.887, 95% confi dence interval=1.206-6.914, p=0.017). patients followed identical prescribed caloric intake diets prior to the pre- NWO carries a higher risk of subclinical vascular infl ammation. BF is a surgery IVGTT through the post-surgery IVGTT. These data support the notion major contributing factor of vascular infl ammation. These results warrant that changes in lactate/pyruvate production and/or utilization is an outcome investigations for subclinical atherosclerosis in NWO patients. from gastric bypass surgery and this may contribute to the remission of the diabetes. 2048-P Relationship between Visceral Fat and All-Cause Mortality in a 2050-P Lean Elderly Asian Population EUN SHIL HONG, AH REUM KHANG, EUN ROH, EU JEONG KU, YE AN KIM, WITHDRAWN KYOUNG MIN KIM, JAE HOON MOON, SUNG HEE CHOI, KYONG SOO PARK, KI WOONG KIM, HAK CHUL JANG, SOO LIM, Seongnam, Republic of Korea, Seoul, Republic of Korea Obesity, especially abdominal obesity, is considered to be a signifi cant risk factor for coronary heart disease and mortality. However, recent studies indicate that overweight may be protective against mortality, a concept that is termed the “obesity paradox”. Therefore, we evaluated the relationships between mortality and various obesity markers in an elderly Asian cohort. We analyzed data from the Korean Longitudinal Study on Health and Aging (KLoSHA) (n = 1 000, age ≥ 65 years, mean age = 76.0 years, 43.9% men). Anthropometric and clinical data were examined using standard methods. The visceral fat area (VFA) and subcutaneous fat area (SFA) were measured using computed tomography. Total fat mass was obtained using bioelectric impedance analysis. Prospective associations between obesity markers and mortality were assessed using Cox proportional hazards models. A total of 222 deaths occurred during the 7-year follow-up (median = 5.2 [range 0.1-6.3] years). Higher body mass index, VFA, SFA, and total fat mass were negatively associated with all-cause mortality in the univariate analyses (p < 0.001). VFA remained the only variable that was signifi cantly associated with all-cause mortality, following adjustment for age, sex, presence of diabetes or hypertension, smoking status, alcohol consumption, exercise habits, serum low-density lipoprotein-cholesterol level, liver enzyme activity, and creatinine level (hazard ratio 0.80, 95% confi dence interval 0.67-0.96, p = 0.018). When stratifi ed by quartiles, the hazard ratio associated with VFA was the lowest in the third quartile and demonstrated a U-shaped pattern for both men and women. Integrated

POSTERS Higher amounts of visceral fat, a marker for central obesity, were associated with decreased all-cause mortality in this relatively lean elderly Physiology/Obesity Asian sample. This fi nding suggests that visceral fat is a marker for mortality in a U-shaped pattern.

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A524 OBESITY—HUMANCATEGORY

2051-P In a randomized cross-over design 9 obese and 8 lean healthy individuals Effect of Roux-en-Y Gastric Bypass Surgery on Microvascular were examined on 2 separate occations 1) after an overnight fast of 12 h Compli ca tions of Type 2 Diabetes Mellitus (control) and 2) after 72 h of fasting (fast). After a 4 hour basal period a LING L. CHUAH, ALEX D. MIRAS, DIMITRIS PAPAMARGARITIS, AMOOLYA VU- 2 h hyperinsulinemic euglycemic clamp was performed to evaluate insulin SIRI KALA, SABRINA N. JACKSON, NICK OLIVER, TORSTEN OLBERS, CAREL W. sensitivity. Indirect calorimetry was performed to assess substrate oxidation LE ROUX, London, United Kingdom rates, labeled tracers were used to calculate substrate fl uxes and muscle Roux-en-Y gastric bypass (RYGB) surgery is known to rapidly improve and fat biopsies were taken. glycaemic control in obese patients with type 2 diabetes; however, its effect We found that fasting induced insulin resistance in lean subjects (Glucose on microvascular complications, particularly retinopathy and neuropathy, is Infusion Rate (GIR) 5.30±0.76 vs. 2.09±0.25 mg/kg/min, p<0.01), whereas largely unknown. obese individuals were insulin resistant in the control situation and did In this prospective study, we assessed the effect of RYGB on microvascular not become additionally insulin resistant after fasting (GIR 1.94±0.21 vs. complications. Sixty-nine obese type 2 diabetic patients with HbA1c >8.5% or 1.29±0.12 mg/kg/min, p=0.37). In the control situation glucose oxidation was on insulin treatment undergoing best medical care in combination with RYGB lower and lipid oxidation was higher in obese subjects compared to lean had 2 fi eld retinal screening, urine microalbumin and nerve conduction study subjects during the hyperinsulinemic euglycemic clamp. at baseline and at one year post-surgery. This was compared to a comparative Our data show that obese individuals are metabolically infl exible in terms group of obese type 2 diabetic patients who had best medical care but not of a rigid insulin sensitivity response pattern in the transition from the basal surgery in the same hospital. Primary end points were glycaemic control, state to fasting. diabetic retinopathy, nephropathy and neuropathy at 12 months follow up. At 12 months after best medical care combined with surgery, HbA1c 2054-P reduced from 9.6 (8.0- 10.4) % to 6.3 (5.9- 7.0) % (p=0.03); and albuminuria Comparison of Subtotal Gastrectomy with Billoth-I or with Billoth-II reduced from 3.6 (1.7-9.3)µg/mg to 1.7 (1.0-4.9) µg/mg. There was no on Metabolic Effects statistically signifi cant change in retinal eye disease (p=0.59). The proposed KYU CHANG WON, JAE HO CHO, SANG HYUN PARK, JUNSUNG MOON, JI SUNG clinical diabetic retinopathy disease severity scale was 1.0(0.0-1.0) YOON, HYOUNG WOO LEE, Daegu, Republic of Korea preoperative and 1.0(0.0-1.0) post-surgery. There was also no signifi cant Even though bariatric surgery has been widely performed in western change in peripheral neuropathy; as evidenced by sural sensory nerve action countries, but little is known about the effects in Asians for low incidence potential (p= 0.15) and peroneal motor nerve conduction velocity (p=0.20). of extremely obese. We attempted to evaluate metabolic effects of subtotal The comparator group who had best medical care without surgery showed gastrectomy (STG), and differences between with gastroduodenostomy no signifi cant changes in glycaemic control and retinopathy at 12 months (Billoth I operation, B-I) and with gastrojejunostomy (Billoth II, B-II) in Korean follow up, whereas urine albuminuria worsened from 1.8 (1.2-5.3) to 3.6 ( subjects. 2.0-10.8) at follow up (p= 0.0002). This was a single center, retrospective follow-up study. From Jan 2008 RYGB showed an early positive effect on marker of diabetes nephropathy to Dec 2010, patients were included which were performed subtotal at one year. RYGB appears to be safe for diabetes related complications gastrectomy for gastric cancer. Exclusion criteria were renal impairment as supported by the lack of negative consequences on retinopathy and (estimated GFR < 60 ml/min/1.73 m2, n=124) and follow-up loss. We neuropathy in the face of rapid improved glycaemia. compared anthropometric parameters, hemoglobin A1c (HbA1c) and lipid profi les at baseline, 6 and 12 months after surgery. 2052-P 792 patients were analyzed. Of them, 331 patients (mean age 54.5 ± 13.4 Sexual Dimorphism of the Human Milieu Intérieur Proteome in years, M: F = 208: 123) were fi nally included. In subjects without diabetes, 196 Overweight and Obese Men and Women patients were performed by B-I and 52 were patients with type 2 diabetes NASSER AL-DAGHRI, MAJED ALOKAIL, THEODOROS ROUMELIOTIS, OMAR (B-I vs. B-II, n=33 vs. 19). At baseline, mean BMI was 23.8 ± 4.2 kg/m2 and AL-ATTAS, KHALID ALKHARFY, SHERIF ABD-ALRAHMAN, SHAUN SABICO, serum total cholesterol (T-Cho), triglyceride (TG), and HDL-cholesterol (HDL-C) ANTIGONI MANOUSOPOULOU, AKUL SINGHANIA, CHRISTOPHER WOELK, of all subjects were 202.7 ± 41.8 mg/dl, 168.6 ± 82.3 mg/dl, 40.3 ± 9.1 mg/ PAUL TOWNSEND, GEORGE CHROUSOS, SPIROS GARBIS, Riyadh, Saudi Arabia, dl, respectively. All lipid profi les except LDL-cholesterol (LDL-C) signifi cantly Southampton, United Kingdom, Athens, Greece improved at 12 months after operation (172.6 ± 33.8, 108.3 ± 46.8, and 48.1 ± Obesity is a basic component of the metabolic syndrome that has been 10.6 mg/dl, respectively; p<0.05). HbA1c also signifi cantly decreased at 12 associated with insulin resistance, hypertension and an increased risk of months (6.0 ± 1.2 %, vs. baseline 6.9 ± 1.3 %; p<0.05). B-II showed more diabetes mellitus type-2 and cardiovascular disease, as well as several favorable effects on lipid profi les than B-I in patients with type 2 diabetes, cancers. This proof-of-principle study compared the proteomic profi les of whereas no differences between 2 groups in non-diabetics. Regardless of age-matched non-diabetic overweight and obese females (n=28) and males diabetes, B-II was superior to B-I in lowering HbA1c signifi cantly after 1 year. (n=31). This study measured the expression of a total of 2470 serum proteins (ΔHbA1c in B-I vs. B-II, -0.4 vs. -0.5 % non-diabetes; -0.6 vs. -0.8 % diabetes; of which the majority were found for the fi rst time in human serum. Of these p<0.05). ~10%, corresponding to about 250 proteins, exhibited differential expression STG with B-II was more benefi cial on metabolic parameters than B-I, between men and women (p<0.05). The log2-fold change ranged from ± 0.3 especially in diabetic patients, and it is supposed for improvement of incretin to over 5.0 and included both novel and known proteins. The proteins of effects. known function belonged to the following pathways: beta-estradiol function, development, lipid and prostanoid metabolism, vitamin D function, Central 2055-P Nervous System function, immunity/infl ammation and tumorigenesis. Abdominal Obesity and Serum Metabolome: A Twin Study Approach In general, this proteomics evidence shows measurable gender-specifi c SANNA KAYE, LEONIE BOGL, JOEL RÄMÖ, ANTTI J. KANGAS, PASI SOININEN, differences in the serum proteome, explaining the results of many physiologic NINA LUNDBOM, JESPER LUNDBOM, AILA RISSANEN, MIKA ALA-KORPELA, and pharmacologic studies showing marked sexual dimorphism. We suggest JAAKKO KAPRIO, KIRSI PIETILÄINEN, Helsinki, Finland, Oulu, Finland, Kuopio, that the pleiotropic nature of obesity and its metabolic syndrome component Finland also manifest sexual dimorphism in the proteome that should be taken into This study aimed to investigate whether the associations between ab- Integrated account in clinical study design, diagnosis and treatment. dominal obesity and serum metabolomics profi le are explained by genetic or POSTERS environmental factors. 2053-P Body mass index, waist circumference (WC) and serum metabolite Physiology/Obesity Metabolic Infl exibility in Obese Subjects: Unaltered Insulin Sensi- concentrations were measured in a large sample of 1369 twin individuals tivity after 72 hours of Fasting (531 monozygotic (MZ) and 838 dizygotic (DZ) ,age range: 21.0-31.5, 52.5% ANN M. BAK, MIKKEL H. VENDELBO, JENS OTTO L. JØRGENSEN, NIELS JESSEN, female) by proton NMR spectroscopy. In addition, body composition was NIELS MØLLER, Aarhus, Denmark measured by DEXA, abdominal fat distribution by MRI and liver fat by MR- The term metabolic fl exibility has been introduced to denote the spectroscopy in a sub-sample (n=84) of the twins (age range: 22.8-36.2, physiological capability of switching energy substrate metabolism between 52.9% female). Cholesky bivariate decomposition models were performed to glucose and lipid utilization. Insulin resistant subjects are metabolically investigate the genetic and environmental contributions to the covariation infl exible and this defi cit may precipitate the metabolic syndrome and type of WC and serum metabolites. 2 diabetes. In this study we aimed to investigate the metabolic fl exibility in Abdominal obesity, insulin resistance and infl ammation correlated with insulin resistant obese subjects in response to fasting. an unfavorable serum metabolic profi le in twin individuals. Sex- and age-

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A525 OBESITY—HUMANCATEGORY

adjusted bivariate Cholesky model-fi tting analyses in all twins revealed that GBP patients compared to controls, where the prevalence was below 3 % for 55-83% of the phenotypic correlations between WC and serum metabolites vitamin B12 as well as for iron-defi ciency anaemia (P <0.001). In conclusion, were explained by shared genetic factors and the remaining by non-shared the risk of anaemia was markedly higher in patients who had underwent environmental factors. Genetic correlations were strongest for alpha-1- GBP compared to controls, even though a greater proportion reported taking glycoprotein (rg=0.38), essential amino acids (rg=0.32-0.40) and HDL-C (rg=- B12 and iron supplements in the GBP group compared to the control group. 0.30). The analyses within MZ-pairs strongly supported the major role of Our results highlight the importance of careful monitoring of patients who genetic and shared environmental factors. undergo GBP with regard to anaemia and nutritional defi ciencies as well as Abdominal obesity is associated with changes in the serum metabolome prescribing adequate substitution. toward increased cardiometabolic disease risk. Genetic factors are important Supported By: Swedish Research Council; Sahlgrenska University Hospital in explaining these associations. In particular, abdominal obesity may share some common genetic determinants with alpha-1-glycoprotein, essential 2058-P amino acids and HDL-C. However, non-shared environmental factors cannot Transfer of Liraglutide from Blood to Cerebrospinal Fluid Is an be disregarded. Unlikely Contributor to Body Weight Loss in Patients with Type 2 Diabetes 2056-P MIKKEL B. CHRISTENSEN, ALEXANDER H. SPARRE-ULRICH, ULRIK GREVSTAD, Metabolic Flexibility Differs among Asian Ethnic Groups in Lean but BOLETTE HARTMANN, METTE M. ROSENKILDE, JENS J. HOLST, TINA VILSBØLL, Not in Overweight or Obese Individuals FILIP K. KNOP, Hellerup, Denmark, Copenhagen, Denmark TAMMY SONG, KHEE SHING MELVIN LEOW, KAVITA VENKATARAMAN, SURESH The mechanisms by which subcutaneously administered glucagon- ANAND SADANANTHAN, YIN HAO ERIC KHOO, S. SENDHIL VELAN, PETER like peptide-1 (GLP-1) receptor agonists confer weight loss in humans are GLUCKMAN, YUNG SENG LEE, YAP SENG CHONG, E. SHYONG TAI, SUE-ANNE incompletely understood. A centrally mediated effect by passage of the TOH, CHIN MENG KHOO, Singapore, Singapore blood-brain barrier has been suggested from animal studies. There are no Metabolic fl exibility (MF) is defi ned as the change in RQ (ΔRQ), indicating human data on blood-brain transfer of GLP-1 receptor agonists. the ability of the organism to adapt fuel utilization to fuel availability. We measured liraglutide in plasma and cerebrospinal fl uid (CSF) obtained Metabolic infl exibility has been related to insulin resistance and obesity. We by lumbar puncture in 8 patients with type 2 diabetes (male: 4/8, age: 63±2 have previously shown that Chinese are more insulin sensitive than Asian years [mean±SEM], BMI: 30±1 kg/m2, Hba1c 56±4 mmol/mol) treated with Indians when they are lean individuals but not when they are overweight. 1.8 mg liraglutide subcutaneously for at least 1 month and with weight In this study, we examined the association between obesity and MF, and loss 8.4±0.4 kg in response to treatment. Liraglutide was measured with a between ethnicity and MF, using a mixed meal tolerance test (MMTT). radioimmunoassay (antibody no. 98302) specifi c for the intact N-terminus of Body composition (DEXA and MRI) and euglycemic hyperinsulinemic the GLP-1 moiety of liraglutide. For validation of the CSF-assay, we obtained clamp data were obtained from 220 healthy men (91 Chinese, 64 Malays, control CSF from subjects (n=9) not treated with liraglutide. 65 Asian-Indians) who were lean (BMI:18.5-22.9 kg/m2), overweight (BMI Plasma-liraglutide was in the nanomolar range (31±4.4 nmol/L). When 23.0-27.4 kg/m2) and obese (27.5-30.0 kg/m2). Respiratory quotient (RQ) applying the assay to the CSF control samples, we measured a picomolar and macronutrient utilization were evaluated by indirect calorimetry during baseline level of 17±1.2 pmol/L. Recovery of liraglutide added to CSF a 3-hr MMTT (600 kcal/day; 30% fat, 15% protein, 55% carbohydrates) was 100 % and sensitivity < 2 pmol/L. The non-corrected CSF-liraglutide conducted after an overnight fast. ΔRQ was defi ned as difference between concentrations in the liraglutide-treated patients were 23±1.8 pmol/L RQ at 90 min and fasting. ΔRQ was negatively correlated with BMI (r= -0.33), (comparison to untreated controls, Mann-Whitney test: P<0.01). Corrected body fat percentage (BF%) (r= -0.33), abdominal visceral adipose tissue for baseline levels of the controls, the ratio of CSF:plasma-liraglutide (VAT) (r= -0.34), abdominal subcutaneous adipose tissue (SAT) (r=-0.31) concentrations was 0.02% (range: 0.07-0.002%). The CSF-liraglutide even after correction for age, ethnicity and insulin sensitivity. ΔRQ was concentration neither correlated with weight loss nor plasma concentrations positively correlated with insulin sensitivity (r= 0.17) but this relationship (Spearman correlations, P>0.6). disappeared when adjusted for BMI. A larger ΔRQ was observed amongst In conclusion, blood-brain barrier transport of liraglutide appears very lean Asian Indians and Malays compared with Chinese (P<0.05) indicating limited and CSF-liraglutide does not seem to correlate with the plasma greater propensity for carbohydrate oxidation and greater suppression of fat liraglutide concentrations or with clinical weight loss. Therefore, our results oxidation in Asian Indians and Malays than Chinese, even after correction do not support that blood-to-CSF transfer is a signifi cant contributor to the for covariates. In overweight and obese individuals, the differences in anorectic effects of GLP-1 receptor agonists. ΔRQ between ethnic groups were no longer seen. In summary, obesity is Supported By: University of Copenhagen associated with metabolic infl exibility, independent of insulin sensitivity. Ethnic differences in MF are observed in the lean, but not in the overweight 2059-P or obese individuals. Effects on Weight of a Cluster-Randomized, Controlled Trial of a Faith-based Adaption of the Diabetes Prevention Program within 2057-P African-American Churches Nutrient Defi ciency in Gastric Bypass Surgery Patients—Longi- RICHARD W. SATTIN, JAMES K. DIAS, LOVORIA B. WILLIAMS, THOMAS JOSHUA, tudinal Results from the Swedish Obese Subjects (SOS) Study LUCY N. MARION, Augusta, GA KRISTIN LINDBERG, PER-ARNE SVENSSON, LARS V. SJOSTROM, LENA Weight is a major factor for Type 2 DM. More than 75% of African CARLSSON, KAJSA SJÖHOLM, Gothenburg, Sweden Americans (AAs) ages 20 or older are overweight and nearly 50% are obese. Bariatric surgery as treatment for obesity is increasing due to its long- Adaptations of the Diabetes Prevention Program (DPP) lower diabetes risk term effectiveness in reducing weight and decreasing the incidence of type factors; however, such programs in AA populations are scarce and mostly II diabetes, cardiovascular events and overall mortality. However, bariatric clinic-based. Our objective was to determine, in a cohort of overweight AAs surgery is associated with a risk of postoperative adverse events and participating in a single-blinded, cluster randomized, community trial, the nutritional defi ciencies. After gastric bypass (GBP) procedures, vitamin B12- effects on weight of a faith-based adaptation of the DPP called Fit Body and iron-defi ciencies are common side effects and may result in anaemia. The and Soul (FBAS) compared with health education (HE) conducted in 20 AA Integrated POSTERS aim of this longitudinal study was to investigate the prevalence of anaemia churches. AA church parishioners in a Southern U.S. community, aged 20 due to vitamin B12- and iron-defi ciencies among GBP patients and matched to 64 years with a BMI ≥ 25 kg/m2 and without history of diabetes, were Physiology/Obesity controls in the SOS study. In total, 263 GBP patients and 1866 controls were eligible to participate. Demographic data, anthropometric, physiological, included in this report. Data was obtained from medical examinations and physical activity, and health-related quality of life measures, and fasting self-reported surveys at the date of inclusion and after 0.5, 1, 2, 3, 4, 6, 8 plasma glucose were obtained at baseline, 12-14 weeks post-baseline and and 10 years. B12 and iron defi ciency anaemia was diagnosed at primary 12 months post-baseline. Standard descriptive statistics and χ2 tests of health care centres according to standard care principles. Results showed homogeneity were calculated, and general linear mixed model repeated- that mean haemoglobin (Hb) levels, prevalence of vitamin B12 and iron- measures analysis of covariance was used to model and test hypotheses defi ciency anaemia were similar for patients undergoing GBP and controls about weight change. From October 2009 through April 2013, 604 AAs were at baseline. After 10 years, mean Hb (g/l) was lower in the GBP group for consented: 83.4% were female; mean age was 46.5 years (SD = 10.88); both men (GBP 140.2±14.6, control 149.2±11.9, P <0.001) and women (GBP and mean BMI was 35.7 kg/m2 (SD = 7.29). Compared with the HE group, 130.0±11.4, control 139.2±11.5, P <0.001). In addition, the prevalence of the FBAS group had a signifi cant difference in adjusted weight loss from vitamin B12 and iron-defi ciency anaemia was 16 % and 24 % respectively in baseline to 12 months post-baseline (2.39 kg vs. -0.465 kg; F = 8.01, 1 ndf,

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A526 OBESITY—HUMANCATEGORY

505 ddf, p-value = 0.005). From baseline to 12 months post-baseline, FBAS 2062-P subjects (18.8%) were signifi cantly more likely than HE subjects (8.1%) to Elevated Content of Plasma Mitochondrial DNA Correlates with achieve a 7% weight loss (χ2= 12.8, 1df, p-value < 0.001). Our fi ndings show Insulin Resistance and Is Associated with Increased Mitochondrial that weight was signifi cantly reduced and maintained in FBAS compared DNA Damage and Oxidative Stress in Skeletal Muscle from Obese with HE. Successful translation and adaptation of the DPP for AAs could Type 2 Diabetes Patients have the potential of reducing the currently estimated 4.9 million AAs with LARYSA YUZEFOVYCH, VIKTOR PASTUKH, MYKHAILO RUCHKO, KELLEY diabetes. PRUTZMAN, SUSAN LEDOUX, GLENN WILSON, WILLIAM RICHARDS, LYUDMILA Supported By: NIDDK (R18DK082401) RACHEK, Mobile, AL Mitochondrial dysfunction and oxidative stress have been implicated in 2060-P the development of insulin resistance (IR), but the question of what are the mechanisms of oxidative stress-induced mitochondrial dysfunction remains WITHDRAWN unclear. Among the potential mechanisms is damage to mitochondrial DNA (mtDNA), since mtDNA is highly specialized and encodes for proteins essential for energy metabolism and, also, damage to mtDNA heightens mitochondrial oxidative stress, which is very critical for insulin signaling pathways. Recently it has been shown that cells that are damaged during several pathological conditions such as, trauma, injuries, or different infections can release mtDNA into circulation, that trigger infl ammation. As IR and type 2 diabetes (T2D) are characterized by a persistent chronic infl ammation, the aim of this study was to evaluate whether T2D patients have elevated content of plasma mtDNA and to determine whether this elevated content is associated with increased mtDNA damage, mitochondrial dysfunction, and oxidative stress in skeletal muscle from T2D patients. Blood and skeletal muscle samples were collected from healthy non-obese (BMI<30, no history of T2D) and obese T2D diabetic patients. Quantitative Real-time PCR was used to quantify selected sequences of plasma mtDNA. T2D patients have increased plasma mtDNA levels in all sequences examined. Also, T2D patients have signifi cantly decreased mtDNA integrity, increased oxidative stress, and reduced content of mitochondrial proteins in skeletal muscle. Furthermore, levels of plasma mtDNA correlated with the degree of IR in T2D patients. To the best of our knowledge, this is the fi rst evidence that elevated level of plasma mtDNA is associated with mtDNA damage and oxidative stress in skeletal muscle, and correlates with IR in obese T2D patients.

2063-P Body Mass Index Signifi cantly Modulates the Power of C-Reactive Protein to Predict Cardiovascular Event Risk among Angiographied Coronary Patients 2061-P ABDUHARAN SAID, ALEXANDER VONBANK, CHRISTOPH H. SAELY, PHILIPP Validation in Africans of the Simple-to-Use Body Adiposity Index as REIN, DANIELA ZANOLIN, HEINZ DREXEL, Feldkirch, Austria, Triesen, Liechtenstein, a Measure of Percent Body Fat Philadelphia, PA AMBER B. COURVILLE, DILALAT BELLO, ORETHA POTTS, MADIA RICKS, MICHELLE Epidemiological studies in various populations show that obesity is O’CONNOR, CAROLINE THORESON, STEPHANIE CHUNG, ANNE E. SUMNER, associated with infl ammation and with increased cardiovascular risk, and Bethesda, MD that the infl ammatory marker C-reactive protein (CRP) strongly predicts the As the obesity epidemic overtakes Africa, practical methods to measure incidence of cardiovascular events. Whether CRP is equally predictive of percent body fat are essential and validation is required. Therefore, with cardiovascular event risk in obese patients and in non-obese subjects is not dual x-ray absorptiometry (DXA) as the criterion method, we tested the known and is addressed in the present study. Body Adiposity Index (BAI) as well as 3 bioelectric impedance equations Cardiovascular events were recorded over a follow-up period of 10 (NHANES III, Durenberger and Kyle) in 116 Africans (70% male; age 39.8+10.4 years in a large high-risk population of 1731 consecutive patients y, range 22 to 64; BMI 27.9+4.5 kg/m2, range 19.7 to 41.2). These methods undergoing coronary angiography for the evaluation of established or have not been either simultaneously compared or validated in Africans. suspected stable coronary artery disease (CAD). Obesity was defined BAI is the most simple as it is the same equation for men and women and as BMI ≥30kg/m². has only 2 variables, hip circumference and height. The BAI formula is: hip At baseline, CRP surprisingly was signifi cantly higher in non-obese circumference (cm) ÷ (height(m)1.5-18). The equipment required for BAI is a subjects (n=452) than obese individuals (0.6±1.5 vs. 0.5±0.8 mg/dl; p<0.001). tape measure and a wall stadiometer. In contrast, each of the bioelectric Prospectively, 27.8% of our patients suffered vascular events. CRP proved impedance equation is different in men and women and requires knowledge to be a strong and independent predictor of vascular events in non-obese of 5 variables: weight, height, age, reactance and gender. The equipment subjects (HR 1.13 [1.06-1.20]; p<0.001) but not in obese subjects (HR 1.08 required for the bioelectric impedance equations are: scale, wall stadiometer [0.94-1.235]; p=0.262). An interaction term BMI x CRP was signifi cant and an analyzer to calculate reactance. To validate and compare each of (p<0.001), indicating that the body mass index signifi cantly modulated the these techniques to DXA measurement of percent fat, Bland-Altman and power of CRP to predict vascular events. Integrated concordance analyses were performed. Bland-Altman analyses revealed From the results of this large 10-year prospective cohort study we POSTERS that the mean difference between DXA and BAI measures of percent fat conclude that obesity signifi cantly modulates the power of CRP to predict was 1.1% compared to 4.2%, 6.1% and 7.4% for the 3 bioelectric impedance cardiovascular event risk among angiographied coronary patients. Physiology/Obesity equations. Concordance between DXA and BAI measures of percent fat was Supported By: Austrian National Bank 0.79, P=.04. Concordance between DXA and the 3 bioelectrical impedance equations were: 0.86, 0.88 and 0.89 (all P<.001). Each of the bioelectric 2064-P impedance equations overestimated percent fat to a greater degree than Fetuin-A and Ser312phosphofetuin-A Responses to Moderate Body BAI. In short, with DXA as the standard, BAI and the 3 bioelectric impedance Weight Loss in Obese Individuals equations all provided very good estimates of percent fat in Africans. GUANG REN, PETER W. GRANDJEAN, ROBERT L. BOWERS, FELIPE ARAYA- Yet because of its simplicity in both measurement and calculation, BAI is RAMIREZ, LAUREL A. LITTLEFIELD, XIAOMING HE, TEAYOUN KIM, REBECCA a better option than any of the bioelectrical impedance equations for the LUDVIGSEN, A. JACK MAHURIN, SURESH T. MATHEWS, Auburn, AL, Waco, TX determination of percent body fat. Fetuin-A inhibits insulin-stimulated insulin receptor tyrosine kinase and is positively associated with obesity, insulin resistance, and incident

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A527 OBESITY—HUMANCATEGORY

diabetes. Weight-loss has been shown to decrease serum fetuin-A levels, (Bio-impedence method) were also determined. Pearson correlation analysis correlating with an improvement of insulin sensitivity. Fetuin-A, secreted revealed that VFA (r = 0.111; P < 0.001) but not BMI (r = 0.044; P = 0.057) and by the liver, exists in phosphorylated and dephosphorylated forms in BAI (r = 0.031; P = 0.137) signifi cantly correlated with PWV. When considered circulation, and a few reports suggest that phosphorylation status is critical in isolation, BMI (β = 0.052, P = 0.001), VFA (β = 0.004, P = 0.043) and BAI (β = for its inhibitory effect on insulin action. However, there are no reports of the 0.032, P = 0.048) were predictive of PWV after adjusting for age, ethnicity, phosphorylation status of fetuin-A with weight loss. The goal of this study gender, blood pressure, Hba1c, low density lipoprotein, and estimated was to characterize the effects of a moderate weight loss (8-10%) on serum glomerular fi ltration rate (eGFR) using general linear model. In stepwise linear total fetuin-A and Ser312phosphofetuin-A in obese individuals. Sixteen regression analysis, BMI and BAI but not VFA were signifi cant predictors obese men achieved a targeted weight-loss goal of 8-10% of initial body of PWV in the fi nal model together with age, Hba1c, blood pressure, and weight over a 6 to 10-month period. As expected, weight loss signifi cantly eGFR. After inclusion of BMI in the model, global R2 was minimally improved decreased waist circumference, total and regional body fat. Similarly, serum with further inclusion of BAI (from 0.217 to 0.220). Taken together, the data insulin, HOMA, QUICKI, and glucose/insulin ratio signifi cantly improved after suggest that BMI (a measure of global adiposity) is a better predictor of AS weight loss. Fetuin-A and phosphofetuin-A levels were signifi cantly lowered than BAI and VFA in T2DM. with weight loss. Following weight loss, unlike fetuin-A, phosphofetuin-A Supported By: NMRC (PPG/AH (KTPH)/2011) levels were correlated with insulin, HOMA, QUICKI, and glucose/insulin ratio. Further, area under the curve (AUC) for phosphofetuin-A and insulin during 2067-P an oral glucose tolerance test were signifi cantly decreased after weight loss. Next, we investigated the changes in fetuin-A, phosphofetuin-A, and WITHDRAWN markers of insulin sensitivity, in response to incremental weight loss. While serum glucose, insulin, HOMA, QUICKI, and glucose/insulin ratio were signifi cantly altered with 6-8% or 8-10% weight loss, total fetuin-A and phosphofetuin-A were signifi cantly decreased with as little as 2-4% weight loss. Taken together, these results suggest that changes in serum fetuin-A and Ser312phosphofetuin-A with modest weight loss may play a role in the improvement of insulin sensitivity. Supported By: ADA (7-04-JF-36); Alabama Agricultural Experiment Station

2065-P Implication of Circulating Irisin Levels with Brown Adipose Tissue and Sarcopenia in Humans JA YOUNG RYU, HAE YOON CHOI, HO CHEOL HONG, HYE JIN YOO, SEI HYUN BAIK, KYUNG MOOK CHOI, Seoul, Republic of Korea Irisin is an exercise-induced novel myokine that drives brown-fat-like conversion of white adipose tissue and is suggested as a promising target for the treatment of obesity-related metabolic disorders. We examined circulating irisin concentrations in individuals with brown adipose tissue (BAT) detected using 18F-Fluorodeoxyglucose Positron Emission Tomography (18FDG-PET) and individuals with sarcopenia defi ned using dual-energy X-ray absorptiometry (DXA). Forty BAT-positive and 40 BAT-negative subjects were enrolled and matched for age, gender, and season/year of 18FDG- PET scanning. We also examined 401 apparently healthy adults enrolled in the Korean Sarcopenic Obesity Study (KSOS), an ongoing prospective observational cohort study. Sarcopenia was defi ned using skeletal muscle mass index (SMI) compared with sex-specifi c reference values in the young population. Among 6,877 consecutive 18FDG-PET scans in 4,736 subjects, 146 subjects (3.1%) had positive BAT scans. The BAT-detected group and the matched undetected group did not differ signifi cantly in circulating irisin levels measured using two kinds of ELISA kits (P = 0.747 and P = 0.160, respectively). Furthermore, serum irisin levels were not different between individuals with sarcopenia and those without sarcopenia (P = 0.305 and P = 0.569, respectively). Although visceral fat area (VFA) measured using 2068-P abdominal computed tomography (CT) and skeletal muscle mass measured DPP4 Inhibition Prevents Over-Nutrition-induced Diastolic Dys- using DXA showed signifi cant difference according to tertiles of SMI levels, function by Reducing Myocardial Fibrosis irisin concentrations did not differ. In conclusion, circulating irisin levels were BRIAN P. BOSTICK, JAVAD HABIBI, ANNAYYA AROOR, VINCENT DEMARCO, not different in individuals with BAT or those with sarcopenia compared to NATHAN REHMER, RAVI NISTALA, MELVIN R. HAYDEN, ALEX MEUTH, JAMES control subjects and were not correlated with skeletal muscle mass index. R. SOWERS, Columbia, MO Obesity has reached epidemic proportions in the United States with more 2066-P than two thirds of the population overweight or obese. Striking correlations Relationship between Anthropometric Measurements of Adiposity exist between the rise in obesity and consumption of a western diet (WD) and Arterial Stiffness in Subjects with Type 2 Diabetes high in fat and fructose. Over-nutrition and obesity are associated with MEI CHUNG MOH, TAVINTHARAN SUBRAMANIAM, CHEE FANG SUM, LEE YING multiple complications, notably insulin resistance and cardiovascular Integrated POSTERS YEOH, XIAO WEI NG, SIMON B.M. LEE, WERN E.E. TANG, SU CHI LIM, Singapore, disease. Treatment of insulin resistance has recently been enhanced by Singapore inhibitors of dipeptidyl peptidase 4 (DPP4) which lower glucose levels by Physiology/Obesity Obesity and arterial stiffness (AS) are major determinants of cardio- preventing degradation of incretins. Further study of DPP4 has revealed vascular disease. The body adiposity index [BAI = (hip circumference)/ correlations between increased DPP4 activity and cardiac dysfunction. Based ((height)1.5) - 18] is a recently developed method to estimate percentage body on these results, we hypothesized that DPP4 activity would be increased in adiposity. However, the relationship between BAI and AS is not well studied. WD induced heart disease and DPP4 inhibition would ameliorate diastolic We hypothesize that measurements of adiposity (global and central) are dysfunction. To test these hypotheses, we treated C57BL6/J mice with a predictive of AS. Therefore, we aim to explore the predictive value of BAI high fat/high fructose WD with or without the DPP4 inhibitor MK0626 for and other conventional anthropometric measurements of adiposity for AS 16 weeks. HOMA-IR confi rmed insulin resistance with WD. We performed in a cross-sectional cohort of 1,275 Singaporean adults (age 21-89) with detailed anatomic study and cardiac MRI to assess cardiac function. To type 2 diabetes (T2DM) (SMART2D study). Aorto-femoral AS was estimated understand the underlying mechanisms, we studied oxidative stress (ROS), using pulse wave velocity (PWV) measured by the applanation tonometry infl ammation and fi brosis. We found that WD increased plasma DPP4 activity method (SphymoCor®). Body mass index (BMI) and visceral fat area (VFA) 3 fold over that of control mice and MK0626 reduced DPP4 activity by 75%

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A528 OBESITY—HUMANCATEGORY from control levels. WD induced signifi cant obesity and diastolic dysfunction and had lower levels of triglycerides (89.4 vs. 139 mg/dL, p<0.001) and lower by cardiac MRI. MK0626 treatment normalized diastolic function. Histologic triglyceride/HDL ratios, a proxy for metabolic syndrome (1.83 vs. 3.26 mg/ staining revealed increased peri-arterial and interstitial fi brosis in WD, both dL, p<0.001). Additionally, those patients who achieved complete 12-month were reduced by MK0626. 3-nitrotyrosine staining revealed increased ROS remission achieved diabetes resolution sooner than those who achieved with WD. MK0626 blocked the increased buildup of ROS. TGF-β and MCP-1 partial 12-month remission (3-month resolution Complete 82.6% vs. Partial were increased after WD with a trend towards improvement by MK0626. 29.4%, p<0.001 and 6-month resolution Complete 93.8% vs. Partial 67.9%, This study suggests an important role for DPP4 in over-nutrition induced p<0.001). heart disease. DPP4 inhibition may represent a key tool in prevention of Supported By: Stanford Medical Scholars diastolic dysfunction by reducing infl ammation and oxidative stress to prevent myocardial fi brosis. 2071-P Improved Glycemic Control Is Associated with Improvements in 2069-P Excess Daytime Sleepiness After Bariatric Surgery Multiplatform Metabolomics Approach Shows That Evolution of JOHN MORTON, ULYSSES ROSAS, KAFI HEMPHILL, DANIEL ROGAN, HARRISON Prediabetic State Is Different between Lean, Overweight, and HINES, Stanford, CA Obese Humans Morbid obesity and diabetes remain a growing health concern throughout MICHAL CIBOROWSKI, EDYTA ADAMSKA, ANNA CITKO, MAGDALENA the world. Bariatric surgery continues to be the most enduring and effective WASZCZENIUK, JULIUSZ WILK, ANNA GOLONKO, JUSTYNA HRYNIEWICKA, intervention for both morbid obesity and diabetes. However, few studies DANUTA LIPINSKA, JOANNA GOSCIK, MAGDALENA RUSAK, JOANNA GODZIEN, correlate improvements in daytime sleepiness with changes in diabetes CORAL BARBAS, MARIA GORSKA, ADAM KRETOWSKI, Białystok, Poland, Madrid, markers after bariatric surgery. Pre-operative (pre-op), 6-month and Spain 12-month post-operative (post-op) data were collected on 191 bariatric Development of type 2 diabetes mellitus (T2DM) is preceded by insulin patients. The Epworth Sleepiness Scale (ESS) was used to evaluate resistance (IR), which may evolve into pre-diabetic state: impaired fasting excessive daytime sleepiness (EDS) for all patients. EDS was defi ned as glucose (IFG) and/or impaired glucose tolerance (IGT). Up to 70% of ESS ≥10. Values obtained include glucose, insulin, HbA1c, BMI, and percent subjects in pre-diabetic state eventually develop T2DM. The risk of T2DM excess weight loss. Student’s T-tests were used to analyze changes in development is higher in overweight (OW) and obese (OB) people; however daytime sleepiness before and after surgery. Preoperatively, there were lean (L) individuals also suffer from T2DM. The purpose of this study was to no signifi cant differences in age, sex, race, insurance status, and income evaluate metabolic differences between healthy and pre-diabetic individuals between patients with EDS and those without. Post-op data revealed that at in different BMI groups. Two metabolomics platforms (LC-MS and CE-MS) 12-months post-surgery patients with EDS had higher levels of HbA1C (EDS were used to fi ngerprint serum samples obtained from L, OW or OB humans 5.99 vs. No EDS 5.55, p=0.002), fasting glucose (EDS 107.9 vs. No EDS 95.1 whom were healthy, IR or had IFG and/or IGT. mg/dL, p=0.005), and fasting insulin (EDS 9.50 vs. No EDS 6.36, p=0.015). Serum samples obtained from non-diabetic 166 individuals (99 female, 67 At 12-months post-op, patients who were considered diabetic based on male, mean age 50 years; age and sex matched in subgroups with different ADA criteria had higher ESS scores (Diabetic 8.41 vs. Non-Diabetic 6.05, BMI: L, OW or OB) were fi ngerprinted by LC-QTOF-MS (Agilent 6520) and p=0.035). Diabetic patients demonstrated worsening ESS scores from pre-op by CE-TOF-MS (Agilent 6224). Participants were classifi ed according to to 12-months post-op compared to non-diabetics (Diabetic -22.2% vs. Non- health/insulin sensitivity status (controls, IR or pre-diabetic) based on Diabetic 16.0%, p=0.040). Although diabetic and non-diabetic patients both fasting glucose level, HOMA-IR, and 2-h 75-g OGTT. Statistical analysis lost a signifi cant amount of their percent excess body weight at 12-months was performed to compare controls with pre-diabetes in each BMI group. (53.8% vs. 67.6%), patients who remained diabetic after bariatric surgery Identifi cation of signifi cant metabolites was performed based on MS/MS had greater levels of EDS. Improved glycemic control after bariatric surgery fragmentation or analysis of the standards. can improve symptoms of excess daytime sleepiness in these patients. Over 20 metabolites discriminating controls from pre-diabetes were Supported By: Stanford Medical Scholars identifi ed. Fatty acid amides, cortisol, and sphingosine-1-phoshate were signifi cantly higher in pre-diabetes as compared to controls in the L group. 2072-P The differences in level of lysophospholipids between healthy and pre- The Predictive Value of the DiaRem Score: Predicting Diabetes diabetic individuals were higher in L and OW, than in OB. Valine, leucine Remission After Bariatric Surgery and acylcarnitines discriminated controls from pre-diabetes independently JOHN MORTON, HOMERO RIVAS, ULYSSES ROSAS, TRIT GARG, KAFI HEMPHILL, on BMI. Stanford, CA Metabolites discriminating controls from pre-diabetes are different The DiaRem Score has recently emerged as a useful tool in predicting between L, OW and OB individuals. diabetes remission after Roux-en-Y Gastric Bypass surgery (RYGB). However, it’s predictive value has only been verifi ed with patients who have 2070-P undergone RYGB surgery. This study aims to understand the reliability of the Complete and Partial Diabetes Remission After Bariatric Surgery DiaRem score at predicting 12-month diabetes resolution after RYGB, sleeve JOHN MORTON, HOMERO RIVAS, ULYSSES ROSAS, TRIT GARG, KAFI HEMPHILL, gastrectomy (SG), and adjustable gastric band (AGB) surgery. 288 diabetic Stanford, CA patients who underwent RYGB (223), SG (40), or AGB (25) at a single academic Bariatric surgery continues to be the most enduring and effective institution were included in this retrospective analysis of a prospective intervention for both morbid obesity and diabetes. However, the exact database. 12-month diabetes resolution was defi ned based on ADA criteria. mechanism of diabetes remission after bariatric surgery is still not well Patients with lower scores had greater rates of diabetes remission than understood. This study seeks to better characterize differences in patients those with larger scores. At 12-months post-op RYGB patients had greater who achieve complete versus partial diabetes remission after weight loss diabetes resolution rates compared to SG or AGB patients respectively surgery. Pre-operative (pre-op) and 12-month post-operative (post-op) data (79.8% vs. 67.5% vs. 60.0%, p=0.031). When broken down by DiaRem score were collected on 220 diabetic patients who underwent bariatric surgery. category, diabetes resolution rates were signifi cantly different in those Values collected at pre- and post-op include glucose, insulin, HbA1c, BMI, patients with pre-op scores of 3-7 (RYGB 96.2% SG 82.4% Band 76.9%, Integrated and percent excess weight loss (EWL) and various biochemical cardiac risk p=0.009). There were no other signifi cant differences in 12-month diabetes POSTERS factors. Complete and partial diabetes remission was classifi ed based on resolution rates in any other score category. A multivariable hierarchical ADA criteria (complete HbA1C <6.0 or FG<100 and partial HbA1C <6.5 or logistic regression demonstrated that RYGB patients with a score from 0-2 Physiology/Obesity FG<125). Preoperatively, patients who achieved complete diabetes remission were more likely (OR 7.93, 95% CI 1.54-40.74, p=0.013) to achieve 12-month at 12-months were younger (48.6 vs. 52.2 y/o, p=0.043), had shorter operative resolution than those with a score of 18-22. This was also true in patients times (172 vs. 195 minutes, p=0.043), and had lower rates of CAD (11.3% vs. with a score from 3-7 (OR 23.8, 95% CI 6.91-81.94, p<0.001). In SG patients, 29.4%, p=0.005) and hyperlipidemia (59.1% vs. 76.5%, p=0.056) compared lower DiaRem Scores did not show a signifi cantly greater predictive value patients who achieved only partial diabetes remission. Additionally, those than higher scores at determining 12-month diabetes resolution. While the patients with complete remission were on diabetes medications (insulin or DiaRem score has emerged as a useful tool to predict 12-month diabetes oral) for less time before surgery (2.7 vs. 5.55 years, p=0.007) and had lower remission after RYGB, it may not predict diabetes remission as well in SG rates of pre-op insulin use (19.9% vs. 38.2%, p=0.019). At 12-months post- and AGB patients. op, patients who achieved complete diabetes remission lost more weight Supported By: Stanford Medical Scholars (67.9% vs. 58.4% of EWL, p=0.020), had lower BMIs (32.7 vs. 34.8, p=0.053)

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A529 OBESITY—HUMANCATEGORY

2073-P and PGC1α RNA was 1.5-2-fold higher (p<0.05); Western blotting showed a Nesfatin-1, an Anorexigenic Modulator of Food Intake, Is Signifi - 3-fold increase in UCP1 protein in winter. To demonstrate the acute effects cantly Increased after Bariatric Surgery of cold, an ice pack was applied to the thigh of lean subjects (n=16) for 30 JOHANNA M. BRIX, CLEMENS HOEBAUS, ASTRID FEDER, HANS P. KOPP, GUNT- min, followed by a fat biopsy 4 hr later. The acute cold stimulated a 2.7 fold RAM SCHERNTHANER, GERIT SCHERNTHANER, Vienna, Austria, Wien, Austria increase in PGC1α RNA (p<0.05) and a 1.9 fold increase in UCP1 (p=0.07). Nesfatin-1 was introduced as a central novel anorexigenic modulator The effects of winter to increase abdominal SC WAT UCP1 was considerably of food intake and body weight, whereas the peripheral mechanism of diminished in subjects with a BMI>30 kg/m2, and winter WAT UCP1 RNA was Nesfatin-1 is less clear. In addition, Nesfatin-1 might be a new goal to inversely associated with adipose macrophage number (r=-0.53, p<0.05) and infl uence hunger and satiety. Furthermore it might have an impact on glucose positively associated with capillary number (r=0.65, p=0.05), suggesting that homoestasis. Therefore it was of interest to investigate Nesfatin-1 levels in adipose beiging occurs less in dysfunctional adipose of obesity. patients before and after weight loss induced by bariatric surgery (BS). Human adipocytes (from stem cells) were exposed to cold (16C) in culture We included 63 patients (mean age: 42± 11 years; mean BMI: 46.7 ± 7.0 for 30 min, followed by 4 hr at 37C, and this resulted in a 2-3 fold increase in kg/m²) with morbid obesity (MO), who were investigated before and 2 years PGC1α and UCP1 RNA. These cold-mediated changes in vitro were blocked after BS. Apart from weight and CV risk-markers (blood pressure, lipids), by propranolol and H-89 (a PKA inhibitor) and were inhibited by the addition a glucose tolerance test (75g), renal and infl ammation parameters were of macrophage-conditioned medium or TNFα (1-100 nM) to the culture. assessed. Nesfatin-1 levels were assessed by a commercial ELISA. These data demonstrate the ability of human SC WAT to increase Nesfatin-1 levels increased signifi cantly after bariatric surgery (3.49±1.26 mitochondrial genes involved with thermogenesis (beiging properties) under vs. 7.96±2.18 ng/ml; p<0.001). We conducted a correlation analysis, where normal physiologic (seasonal) conditions. The cold mediated changes were post surgery Nesfatin levels correlated signifi cantly with Blood Glucose inhibited by infl ammation. Because SC WAT represents an enormous depot 2h post challenge ( r= -0.289; p=0.049). In another correlation analysis, in humans, the upregulation of thermogenic capacity may be important in this result was augmented as the change pre surgery versus post surgery overall energy regulation, and may become dysfunctional with obesity. (delta) Blood Glucose 2h post challenge correlated signifi cantly with delta Supported By: UL1RR033173 Nesfatin-1 levels ( r= -0.502; p=0.001). This is the fi rst study demonstrating increased Nesfatin-1 levels in patients 2076-P after weight loss induced by BS. Thus Nesfatin-1 might contribute to the Brain Activation in Response to Visual Food Cue: Effect of Gender weight loss in patients after bariatric surgery by improvement of satiety. RENATA BELFORT-DEAGUIAR, DONGJU SEO, SARITA NAIK, CHERYL M. LACADIE, JANICE HWANG, TODD CONSTABLE, RAJITA SINHA, ROBERT S. SHERWIN, New 2074-P Haven, CT, London, United Kingdom Effect of Obesity on β-Cell Function in Subjects with Normal Gender differences in eating behavior, satiety hormones and brain Glucose Tolerance, Prediabetes, and Diabetes responses to food cues have been previously described, however, the DAI ZHE, XU YANCHENG, ZHANG HONGWU, Wuhan, China, Urumqi, China interplay between these factors in individuals of both genders has received Obesity has parallel prevalence with diabetes all over the world. To little attention. We therefore studied 8 male and 12 female healthy subjects, elucidate the effect of obesity on the β cell function in the process of matched by age (31±3 vs. 28±2 years) and BMI (25±1 vs. 23±1 kg/m2) 2 hours diabetes development, a cross sectional study was taken place in Xinjiang after a standardized meal, a time when they were neither hungry nor full. Province, China, which consisted of 3352 subjects aged from 30-80 They underwent functional MRI while viewing high-calorie food (HC), low- years. The questionnaires, physical examination laboratory tests and oral calorie (LC), and non-food (NF) pictures during a hyperinsulinemic-euglycemic glucose tolerance test (OGTT) with 75g glucose were performed in all the clamp to standardize insulin and glucose levels. Prior to study, glucose, participants. We used the HOMA-βindex, insulinogenic index (ΔI30/ΔG30) insulin as well as ghrelin levels were similar in males and females. Leptin and ΔAUCI/G(0-120) as the indicators of β cell function, then reevaluated levels were lower in males (baseline: 3.3±0.5 vs. 10.4±1.2 ng/mL; P<0.001) the results after adjusting the β cell function for insulin sensitivity ([ΔAUCI/ and increased less during the clamp (0.8±0.4 and 2.7±0.7; P<0.05). Hunger G(0-120)]/HOMA-IR index). Altogether 481 prediabetes and 271 diabetes ratings were not different at baseline, but signifi cantly increased in both patients were defi ned. The result showed that β cell function decreased genders (p=0.01) upon looking at food pictures. Hunger ratings, however, gradually from normal glucose tolerance to prediabetes and diabetes rose to 2-fold higher levels in males (P=0.05). Similarly, males rated wanting states. Compared to normal weight, the adjusted β cell function increased for HC foods higher compared to females throughout the study. Baseline in overweight state but moderately decreased in obese state both in leptin levels negatively correlated with wanting for food pictures (want HC: prediabetes and diabetes patients (Table 1). Simple correlation analysis and r=-0.55, P=0.01; LC: r=-0.55, p=0.01). During euglycemic hyperinsulinemia, multiple regression analysis have confi rmed the relationship of body mass gender x task (HC/LC/NF) interaction was signifi cant in the left insula and index and β cell function in normal glucose tolerance and glucose intolerance right inferior parietal lobe (IPL); males showed reduced activity in both the states. The research concluded that obesity have important and complicated insula (HC28kg/m2) in Patients with Type 2 Diabetes: Results from a Large Pooled Integrated POSTERS Database of Post-marketing Observational and Non-interventional Studies Physiology/Obesity 2075-P OLIVER SCHNELL, JIANPING WENG, WAYNE H.H. SHEU, HIROTAKA WATADA, Temperature and Seasonal Changes in Human Subcutaneous (SC) SANJAY KALRA, SIDARTAWAN SOEGONDO, NORIYUKI YAMAMOTO, WLADY- White Adipose Tissue (WAT): Evidence for Browning and Inhibition SLAW GRZESZCZAK, Munich, Germany, Guangzhou, China, Taichung, Taiwan, by Infl ammation Tokyo, Japan, Karnal, India, Jakarta, Indonesia, Beijing, China, Zabrze, Poland PHILIP A. KERN, BRIAN S. FINLIN, BEIBEI ZHU, NEDA RASOULI, ESTHER E. Patients with diabetes often have problems controlling their body weight DUPONT-VERSTEEGDEN, Lexington, KY, Denver, CO and are overweight or obese. Many international and national guidelines Although humans can generate heat through brown adipose tissue, little therefore recommend weight loss as even a modest reduction can meaning- attention has been paid to the potential browning/beiging of WAT, which fully improve glucose control. This study aimed to assess the impact of the clearly occurs in many rodent WAT depots but has not been studied in humans. alpha-glucosidase inhibitor acarbose on body weight alterations in a real- We examined both abdominal and thigh SC WAT from 71 subjects who were life setting. Data were pooled from 10 post-marketing surveillance and biopsied in the summer (June-Aug) or winter (Jan-Mar). In WAT of subjects non-interventional studies from 21 areas around the world. The effect of biopsied in winter (versus summer), UCP1 RNA was 4-10 fold higher (p<0.05),

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A530 OBESITY—HUMANCATEGORY acarbose on body weight was then analysed taking into account baseline Index. Our data suggest that, in obese subjects, low 25(OH)D levels are body weight, glycemic parameters, and other baseline characteristics. independently associated with liver insulin resistance, but not with beta Data were available for 67,682 patients in the safety analysis and 62,905 cell function. Further studies are needed to clarify the relationship between patients in the effi cacy analysis. Mean patient follow up (± standard glucose homeostasis and vitamin D levels. deviation [SD]) was 12.2±4.8 weeks. The mean relative reduction in body weight at the 3-month visit was 1.45±3.24% (n=43,510) and at the last visit was 1.40±3.28% (n=54,760), with both p<0.0001 vs. baseline. Body weight reductions were dependent on baseline body weight, and greatest reductions occurred in patients who were ≥100 kg. In these patients the relative reduction at the 3-month visit was -2.82±3.93% (p<0.0001; n=2,416). When analyzed according to quartiles based on baseline glycemic parameters, the reduction in body weight was independent of fasting plasma glucose (FPG), postprandial plasma glucose (PPG), glycosylated hemoglobin (HbA1c) and glucose excursion (PPG minus FPG). Real-life data from this post-hoc analysis showed that acarbose treatment reduces body weight independent of glycemic control. This response is dependent on baseline body weight and several other factors.

2078-P Circulating Irisin Is Upregulated by Insulin Infusion in Obese, but 2080-P Not in Lean, Humans and Is Inversely Associated with Insulin Contribution of Abdominal Obesity to Cardiovascular Risk in Sensitivity and Respiratory Exchange Ratio Patients with Cerebrovascular Disease: Insights from the IRIS Trial MAREK STRACZKOWSKI, AGNIESZKA NIKOLAJUK, NATALIA MATULEWICZ, SILVIO E. INZUCCHI, KAREN L. FURIE, MARK GORMAN, ANNE M. LOVEJOY, MAGDALENA STEFANOWICZ, MONIKA KARCZEWSKA-KUPCZEWSKA, Białystok, CATHERINE M. VISCOLI, LAWRENCE H. YOUNG, WALTER N. KERNAN, New Poland, Olsztyn, Poland Haven, CT, Providence, RI, Burlington, VT Irisin is a recently discovered myokine, with the potential to induce The importance of general and abdominal obesity (AO) for heart disease brown-fat-like development of white adipose tissue and to increase energy risk is well known in the general population. There are far less data, however, expenditure. In humans, data on the relationships of circulating irisin with on the metabolic implications of excess body weight and weight distribution insulin sensitivity and other metabolic parameters are inconsistent. The aim in stroke populations. We sought to determine the association between AO of the present study was to assess the associations of serum irisin with and cardiovascular (CV) risk factors in patients with TIA/stroke enrolled in an insulin sensitivity and substrate oxidation, as well as the effect of insulin NIH-funded trial. The Insulin Resistance Intervention after Stroke (IRIS) trial infusion on circulating irisin. The study group consisted of 148 healthy is testing the hypothesis that pioglitazone will reduce stroke and myocardial subjects (115 males and 33 females, age between 18 and 35 years), 81 lean infarction in insulin resistant patients with TIA/stroke. We screened 7146 and 67 overweight or obese. Euglycemic hyperinsulinemic clamp and indirect patients for insulin resistance. Mean age was 64±11 yrs, 64% were male, calorimetry were performed. Serum irisin was measured in the baseline and mean BMI was 28.5±5.0 kg/m2; 42% of men and 64% of women had AO and after the clamp. Baseline irisin was not different between lean and (>40” [102 cm] in men; >35” [88 cm] in women.) We examined the relationship overweight/obese group (p=0.44). Insulin infusion resulted in an increase between AO and metabolic features (glucose, insulin, HOMA-IR, lipids). in serum irisin in overweight/obese (p=0.006), but not in the lean group The presence of abdominal obesity was associated with greater insulin (p=0.73). In consequence, post-clamp irisin was higher in the overweight/ resistance, higher levels of triglycerides and lower levels of HDL-C, after obese (p=0.042). Baseline irisin was related to fasting free fatty acids adjusting for gender, age and BMI. (r=0.26, p=0.008). Post-clamp irisin was positively related to BMI, waist Table. Least Squares Means for Biochemical Features. circumference, triglycerides and resting energy expenditure (all p<0.05) and negatively to insulin sensitivity (r=-0.20, p=0.014) and respiratory Feature Abdominal Obesity exchange ratio, both at baseline (r=-0.21, p=0.046) and during the clamp (r=- Yes No P 0.34, p=0.001). We also observed negative association of post-clamp irisin n=3551 n=3595 with glucose oxidation (r=-0.26, p0=0.008) and positive with lipid oxidation Fasting Plasma Glucose (mg/dL) 98 96 <.0001 (r=0.37, p<0.0001) during hyperinsulinemia. Our data show that insulin Insulin (uU/mL) 20 16 <.0001 increases circulating irisin, but this effect is dependent on the presence of HOMA-IR 4.9 4.0 <.0001 overweight/obesity and may represent a compensatory response to increase lipid oxidation in these conditions. Triglycerides (mg/dL) 138 119 <.0001 Supported By: Program Innovative Economy (UDA-POIG.01.03.01-00-128/08) HDL cholesterol (mg/dL) 48 51 <.0001 We conclude that in a large group of non-diabetic patients with recent TIA/ 2079-P stroke, the presence of AO was associated with greater CV risk. Strategies Hypovitaminosis D Is Associated with Liver Insulin Resistance in targeted at reducing obesity, and specifi cally AO, appear warranted in the Obese Subjects setting of cerebrovascular disease. CATERINA CONTE, CHIARA MARIA ASSUNTA CEFALO, CATERINA POLICOLA, Supported By: U01NS44876 SIMONA MOFFA, TERESA MEZZA, GIAN PIO SORICE, ANDREA GIACCARI, Rome, Italy 2081-P Hypovitaminosis D is highly prevalent in obese subjects. Serum Rate of Initial Weight Loss and Low Fat Oxidation After Bariatric 25-hydroxy vitamin D3 [25(OH)D] concentration, the best marker of human Surgery Predisposes Individuals to Weight Regain vitamin D status, has been reported to be associated with glucose status,

ROBERT A. STANDLEY, PAUL M. COEN, CHUCK J. TANNER, NIKKI L. HELBLING, Integrated insulin resistance (IR) and beta cell function. To specifi cally investigate the POSTERS GABRIEL S. DUBIS, HUI XIE, STEVEN R. SMITH, GEORGE M. EID, FREDERICO G.S. relationship between 25(OH)D and liver IR we performed a comprehensive TOLEDO, JOSEPH A. HOUMARD, BRET H. GOODPASTER, Orlando, FL, Greenville, Physiology/Obesity metabolic assessment (2-h OGTT, hyperinsulinemic euglycemic clamp [HEC], NC, Pittsburgh, PA body composition by DXA) in 20 obese non-diabetic subjects (42.9±2.7 The reasons for weight regain following Roux-en-Y gastric bypass (RYGB) yrs; BMI 37.7±0.8 kg/m2) with 25(OH)D insuffi ciency (<30 ng/mL). Liver IR surgery are not clear. The aim of this study was to perform a post-hoc was estimated using the index by Vangipurapu et al. (-0.091 + [log insulin analysis on data from a previously conducted randomized controlled trial AUC 0-120 min x 0.400] + [log fat mass% x 0.346] - [log HDL cholesterol to determine how initial weight loss following surgery infl uences weight x 0.408] + [log BMI x 0.435]). There was a signifi cant inverse correlation regain. between 25(OH)D and the liver IR index (r = -0.514, p = 0.02). This correlation 1-3 months following RYGB, participants (n=128) were randomized to maintained its signifi cance after adjusting for age, gender, total cholesterol, 6-months of either moderate exercise (EX) or lifestyle control (CON). After triglycerides and whole body insulin sensitivity (M value assessed by HEC) the intervention, participants returned for 1 and 2 year follow-up visits. in multiple linear regression analysis. There was no signifi cant correlation Cardiorespiratory fi tness was determined by VO2 peak test, body composition between 25(OH)D and beta cell function estimated by the Disposition by DXA and physical activity levels by wearable monitors.

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A531 OBESITY—HUMANCATEGORY

For the post-hoc analysis, completers (n=117) were divided into two groups Women and men did neither differ in age (40±12 vs. 41±12, p=0.679), based on weight loss after bariatric surgery; those who lost the least (LL; nor in BMI (46.5±7.1 vs. 46.0±6.7 kg/m2; p= 0.603). But men had a slightly n=65) and lost the most (LM; n=52). The LM group weighed more pre-surgery higher but not signifi cant excess BMI loss compared to women (73.7±23.2 (131.7±3.8 vs. 119.0±2.5 kg, P<0.05), while there was no difference in weight vs. 66.4±29.8%; p= 0.064). Before surgery men had signifi cantly more often post intervention. The LM group regained more weight compared to LL group type 2 diabetes (37.7% vs. 15.8%) and equal frequency of patients with at both follow-up visits (Year 1: -1.4±1.5 vs. -8.1±1.3%; Year 2: 5.0±1.9 vs. prediabetes (19.7 vs. 19.5%); Chi-Square 16.941; p<0.001. -2.7±2.0%, P<0.05). Fat mass was greater in the LM group at both follow up After surgery, only 1.6% of the male and 2.6% of the female patients were visits (Year 1: 31.4±2.0 vs. 25.7±1.7 kg; Year 2: 37.0±4.9 vs. 29.8±2.3 kg, P<0.05, still suffering diabetes and 6.3% vs. 6.7% were suffering prediabetes; Chi P=0.07), while fat free mass tended to increase in the LM group (53.8±2.5 vs. Square 0.079; p= 0.961. 49.9±1.3 kg; 57.1±4.6 vs. 50.5±1.7 kg, P=0.06). Respiratory quotient (RQ) was Men had a greater decrease in HOMA-IR (p= 0.013), LDL-cholesterol (p= greater in the LM group post-intervention (0.78±0.01 vs. 0.76±0.01, P<0.05) 0.030), triglycerides (p<0.001) and HbA1c (p=0.002) and an increase in HDL- indicating less reliance on fat substrate oxidation. Weight change was not cholesterol (p=0.002) than women. There were no differences regarding related to initial group randomization (EX vs. CON), cardiorespiratory fi tness total cholesterol. or physical activity levels at follow-up. In conclusion, individuals who initially Our data indicate that, even though there are no differences in excess lose the most weight early after RYGB are predisposed to later weight BMI loss men benefi t more from BS regarding diabetes and lipid profi le than regain, independent of physical activity or fi tness. Moreover, greater weight women. In limited surgery capacity, one might consider to prefer men to regain was associated with a reduced reliance on fat oxidation, suggesting women for bariatric procedures. a link between fuel selection and weight regain. 2084-P 2082-P Visceral Fat Area Is Signifi cantly Related with Hepatic Steatosis The Short-term Improvement in Glucose Homeostasis after Roux- Assessed by Controlled Attenuation Parameter en-Y Gastric Bypass in Type 2 Diabetes Is Explained by Acute KWANG JOON KIM, KYU SIK JUNG, YOUNG EUN CHON, JAE HYUK LEE, EUN Calorie Restriction Despite Enhanced Incretin Effects SEOK KANG, BYUNG-WAN LEE, BONG SOO CHA, HYUN CHUL LEE, JAE BOK SARAH STEVEN, PETER K. SMALL, SEAN WOODCOCK, KIEREN HOLLINGSWORTH, CHUNG, DONG YEOB SHIN, JUN YONG PARK, Seoul, Republic of Korea, Goyang- ANDREA PUCCI, RACHEL L. BATTERHAM, ROY TAYLOR, Newcastle upon Tyne, si, Republic of Korea United Kingdom, Sunderland, United Kingdom, North Shields, United Kingdom, Controlled attenuation parameter (CAP) is noninvasive method of London, United Kingdom measuring hepatic steatosis with high accuracy. The aim of this study is The mechanism of improved glucose control following acute calorie to investigate the relationship between hepatic steatosis assessed by CAP restriction and Roux-en-Y gastric bypass surgery (RYGB) in type 2 diabetes and clinical factors including regional fat distribution measured by computed remains controversial. Individuals with type 2 diabetes listed for RYGB were tomography (CT). randomised to very low calorie group (VLCD) group (studied before and after Clinical and metabolic measurement, laboratory data, regional fat a 7 day VLCD) or Surgery group (studied just prior to and 7 days after RYGB). distribution, and CAP value were evaluated in 304 patients (165 male, 139 At baseline, the VLCD (n=9) and Surgery (n=9) groups were well matched: female) who underwent CAP and abdominal fat CT. In this study, signifi cant 52.2±1.6 vs. 45.9±2.5 yr; 121.4±3.7 vs. 120.8±5.0 kg; HbA1c 7.8±0.6 vs. steatosis is defi ned when CAP value is ≥ 250 dB/m. 7.3±0.5 %. Figure 1 showed visceral fat area (VFA) was signifi cantly related with CAP Weight loss was superior in the Surgery vs. VLCD group: 5.1±0.5 vs. value, together with triglycerides (TG), and alanine aminotransferase. Figure 3.5±0.4 %. However, fasting plasma glucose fell from 10.0 (5.9-16.1) to 7.2 2 showed severity of NAFLD according to the glucose tolerance status. In (5.1-16.2) mmol/L in the VLCD group (p=0.009) and 7.5 (6.0-15.9) to 7.1 (4.5- the multiple logistic regression analysis, VFA (odd ratio [OR], 1.010; 95% 9.9) mmol/L in the Surgery group (p=0.171). confi dence interval [CI], 1.001-1.019; P=0.028) and TG (OR, 1.006; 95% CI, Incremental AUC insulin after a 100kcal semi-solid meal did not change 1.001-1.011; P=0.022) were selected as independent risk factor for signifi cant after VLCD: 950±373 to 994±314 mU/L min-1 but tended to increase after hepatic steatosis. In sub analysis of 110 patients with BMI <23 kg/m2, only Surgery: 783±149 to 1083±187 mU/L min-1. The post meal rise in plasma VFA was signifi cantly related with hepatic steatosis (OR, 1.006; 95% CI, glucose was lower after VLCD than after Surgery (at 30min: 0.6±0.2 vs. 1.001-1.011; P=0.022). 1.7±0.2, p=0.003; at 60min: 1.1±0.2 vs. 1.6±0.2, p=0.096). Our data demonstrated that VFA was signifi cantly related with signifi cant These changes occurred despite a lesser meal related change in the hepatic steatosis assessed by CAP.even in non-obese patients. VLCD group in total GLP-1 (0-20 min: +0.7±0.3 to -0.1±0.1 pmol/L) compared with the Surgery group (+0.7±0.3 to +6.6±0.9 pmol/L). Similarly, total GIP did not increase after VLCD (+93.4±15.7 to +41.5±18.5 pg/ml) but did after Surgery (+62.8±21.2 to +168.4±37.5 pg/ml). Liver triacylglycerol measured by magnetic resonance decreased signifi cantly in both groups; VLCD: 11.8±2.2 to 9.8±1.9 % (p=0.003) and Surgery: 6.5±1.6 to 4.5±1.0 % (p=0.027). Greater improvement in fasting and post-meal glucose occurred following VLCD compared to RYGB despite greater weight loss after surgery. The acute effect on glucose control following RYGB is explained by the associated sudden calorie restriction.

2083-P Diabetes Remission Is Higher in Men Compared to Women After Bariatric Surgery JOHANNA M. BRIX, HANS P. KOPP, EVA C.H. KRZIZEK, GERIT H. SCHERNTHANER, GUNTRAM SCHERNTHANER, Vienna, Austria, Wien, Austria Integrated POSTERS The majority of patients undergoing bariatric surgery(BS) are women. Many studies demonstrated that women and men have a different risk for Physiology/Obesity gaining cardiovascular (CV) disease. It is still not clear if morbidly obese men have the same risk profi le as women. However, due to increasing numbers of morbid obese patients, a patient selection for BS becomes more and more important. Therefore we investigated gender differences regarding diabetes and weight loss in patients before and after bariatric surgery. We included 438 patients with MO (mean age was 40±12 years, 347 (84%) females, 66 males) before and two years after BS. In patients without diabetes a 75g oGTT was performed. Diabetes Mellitus was defi ned by ADA criteria. Insulin levels were assessed and HOMA-Insulin resistance (IR) was calculated. Apart from demographic and CV risk factors renal and infl ammation parameters were assessed.

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A532 OBESITY—HUMANCATEGORY

2085-P obese postmenopausal women (60±6 yr; mean±SD) to MSyn. MSyn and non- Shifts on Microbiota Composition and Clinical Parameters Are MSyn were divided into overweight and obese groups; MSyn had abnormal Associated to Diabetic Status in Morbidly Obese Subjects: Impact metabolism and higher waist (*P<0.05 vs. non-MSyn) (Table). In non-MSyn of Bariatric Surgery women, increasing ABD fat cell weight (FCW) was associated with higher MARIA CARMEN COLLADO, JARNA C. HANNUKAINEN, PAULINA SALMINEN, LPLA (r=0.29, P<0.05), suggesting greater adipogenic capacity. In contrast, SEPPO SALMINEN, PIRJO NUUTILA, Valencia, Spain, Turku, Finland FCW and LPLA were not related in MSyn (r=0.14, P=NS), independent of BMI Recent research has shown that microbiota play an important role in (r=0.13-0.15, P=NS). This suggests that similar to obese MSyn, overweight metabolic disorders such as obesity and diabetes. Our objective was to MSyn have lost the ability to expand their ABD FCWs, contributing to their determine the impact of diabetes in microbiota and evaluate the effects of metabolic dysfunction. Thus, despite weighing 18 kg less (13 kg lower bariatric surgery. Thus, gut microbiota of 12 morbidly obese subjects (T2DM fat mass, P<0.01) than obese, overweight MSyn women have reduced with metformin, IGT and healthy, n=4 in each subgroup) and 12 age-matched capacity to store fat in ABD tissue and comparable visceral to total ABD controls was characterized by 16S gene pyrosequencing and specifi c qPCR. fat ratio (overweight: 0.29±0.07 vs. obese: 0.28±0.07), which is higher than For obese subjects analyses were performed prior to and six months after non-MSyn women (0.24±0.07; P<0.01). This suggests that the protective bariatric surgery. Clinical and biochemical data including HOMA-IR and adipogenic capacity of ABD fat may be a protective mechanism against Matsuda index were also assessed. visceral adiposity and MSyn after menopause. A total of 211,010 high-quality pyrosequencing reads were analysed. The dominant phylum belonged to Firmicutes, Proteobacteria, Actinobacteria and BMI Body Fat FCW TG HDL Fasting Glucose HOMA-IR (kg/m2) (%) (µg TG/cell) (mg/dL) (mg/dL) (mg/dL) Bacteroides. We found that intestinal concentrations of Bacteroidetes were lower and of Firmicutes higher in obese than in controls. Obese patients MSyn: Overweight (N=21) 29±1 43±3 0.67±0.15 167±56* 47±11* 99±7* 3.2±1.1* with T2DM had lower microbial alpha-diversity compared to non-diabetic MSyn: Obese (N=55) 35±4* 49±4 0.79±0.17* 159±62* 45±10* 102±11* 4.5±2.0* obese and controls and also, they had higher abundance of Bacteroides and Non-MSyn: Overweight (N=39) 28±1 43±4 0.63±0.18 118±34 57±14 93±7 2.2±0.6 Streptococcus genus as compared to those without diabetes. They also Non-MSyn: Obese (N=40) 33±3 49±4 0.69±0.17 108±25 55±13 93±6 2.7±1.1 higher levels of Akkermansia spp, which has been associated with the use of metformin. In obese subjects, Fusobacterium genus, Eubacteriaceae and Enterobacteriaceae family levels were associated positively with HOMA- 2088-P IR and negatively with insulin sensitivity evaluated with Matsuda Index. Serum Chemerin Levels Are Positively Correlated with Abdominal Six months after bariatric surgery T2DM was in remission in all patients. Visceral Fat in Type 2 Diabetes The Firmicutes-to-Bacteroidetes ratio was decreased and the microbiota MOONSUK NAM, JU-YOUNG HAN, SO HUN KIM, SEONG BIN HONG, YONG composition was similar between obese and lean healthy controls. SEONG KIM, Incheon, Republic of Korea In conclusion, morbidly obese subjects with T2DM have signifi cantly Chemerin is a recently identifi ed adipokine suggested to play a role in lower diversity in microbiota than obese with IGT or healthy subjects. The obesity and its metabolic complications. Although a positive correlation microbiota shifts are associated with indexes of insulin sensitivity and beta between visceral fat accumulation and serum chemerin levels in non-diabetic cell function and normalized after bariatric surgery. subjects has been shown, the relationship in type 2 diabetic subjects is Supported By: Academy of Finland unknown and may differ with non-diabetic subjects. We evaluated whether serum chemerin was associated with visceral abdominal fat accumulation in 2086-P type 2 diabetic patients. Brain Iron Overload Correlates with Metabolic Homeostasis and A total of 218 Korean type 2 diabetic patients were enrolled. Abdominal Cognitive Dysfunction in Obese Subjects visceral and subcutaneous fat areas were measured by computed GERARD BLASCO, JOSEP PUIG, JOSEP DAUNIS-I-ESTADELLA, ROSER LLAGOS- tomography and serum chemerin levels were measured by ELISA. TERA, XAVIER MOLINA, SALVADOR PEDRAZA, WIFREDO RICART, JOSÉ MANUEL Mean age was 52.2±7.5 years, BMI 25.3±2.9 kg/m2, HbA1c 7.5±1.3%, FERNÁNDEZ-REAL, Girona, Spain visceral abdominal fat area 112.0±48.9 cm2, subcutaneous abdominal Brain iron overload may run in parallel to changes on metabolic fat area 153.1±66.6 cm2, and serum chemerin 80.3±1.5 ng/ml. Serum homeostasis and is linked to body composition in obese subjects. We aimed chemerin level was higher in women (78.6±22.0 vs. 82.9±22.6, p=0.159) and to identify the components related to obesity that contribute to an increased positively correlated with basal insulin(r=0.250, p<0.001), HOMA-IR(r=0.189, brain iron overload and its impact on cognitive function. p=0.006), triglyceride (r=0.358, p<0.001), serum creatinine(r=0.208, p= We prospectively recruited 23 middle-aged obese subjects (13 women; 0.002), creatinine clearance(r=-0.178, p=0.009) Albumin/Cr Ratio(r=0.249, 50.4±7.7 years; body mass index [BMI] 43.7±4.4Kg/m2) and 20 healthy lean p<0.001), hsCRP(r=0.309, p<0.001), fi brinogen(r=0.320, p<0.001), total volunteers (10 women; 48.8±9.5 years; BMI 24.3±3.54Kg/m2) in whom abdominal fat (r=0.156, p<=0.022), abdominal visceral fat area(r=0.279, brain iron load (measured by R2* values in white matter and grey nuclei) p<0.001) and V/S ratio(r=0.3, p<0.001), and negatively correlated with and hepatic iron concentration (HIC) were evaluated by magnetic resonance HDL-cholesterol(r=-0.189, p=0.005) after adjustment for age, gender and imaging (MRI). A neuropsychological test-battery was used to evaluate BMI. Multiple regression analysis showed that abdominal visceral fat potential cognitive dysfunction. Multivariate regression analysis was used area, serum triglyceride, creatinine clearance, hs-CRP and fi brinogen to identify independent predictors of brain iron load and cognitive function. independently affected serum chemerin levels. The serum chemerin was Iron load was increased at the caudate nucleus (p<0.001), lenticular probably predictors of visceral adiposity and may play an important role in nucleus (p=0.004), hypothalamus (p=0.002) as well as at liver (p<0.001) in infl ammation, diabetic nephropathy and cardiovascular disease that cause obese subjects. After adjusting for age and gender, waist circumference, mortality in type 2 diabetes. Clinical utility of chemerin as a biomarker of BMI and fat mass were independent predictors of brain iron load in caudate metabolic syndrome in type 2 diabetes could be useful in early detection of and lenticular nuclei, as well as for HIC. Iron load in the hypothalamus and these pathological states. caudate nucleus were independently associated to cognitive dysfunction Supported By: Korea Healthcare Technology R&D Project (p=0.048 and p=0.027, respectively). Our preliminary data show that brain iron load in the hypothalamus 2089-P Integrated and caudate nucleus are increase in obese subjects resulting in cognitive Metabolic Response to Overfeeding in Insulin Resistant vs. Sensi- POSTERS dysfunction. Brain iron load could be a potential MRI biomarker in obesity-

tive Humans Physiology/Obesity associated cognitive dysfunction. TRACEY MCLAUGHLIN, COLLEEN CRAIG, CINDY LAMENDOLA, LI-FEN LIU, DALIA PERELMAN, DANIEL SPIELMAN, SAMUEL W. CUSHMAN, Stanford, CA, Bethesda, 2087-P MD Adipogenic Capacity, Not Obesity, Is Associated with Metabolic Weight gain has been used in mouse models to evaluate causal mecha- Syndrome (MSyn) in Postmenopausal Women nisms of obesity-induced insulin resistance and type 2 diabetes. We evaluated MONICA C. SERRA, ALICE S. RYAN, ANDREW P. GOLDBERG, Baltimore, MD whether metabolic responses to weight gain differed in insulin-resistant (IR) Fat cells with high adipogenic capacity protect against visceral obesity versus insulin-sensitive (IS) humans and whether expansion of regional fat and MSyn. Obesity predisposes to MSyn; yet, many normal and overweight depots correlated with development of insulin resistance. To do this, we enrolled women develop MSyn. We hypothesized that the inability to expand fat cells 26 healthy overweight/moderately-obese individuals and subjected them to across a range of obesity and abdominal (ABD) lipoprotein lipase activity controlled short-term overfeeding with 900 kcal/day added to usual diet. Insulin- (LPLA) (regulates lipid storage, adipogenesis), predisposes overweight and mediated glucose uptake was quantifi ed by the modifi ed insulin-suppression test

ADA-Funded Research & Guided Audio Tour poster For author disclosure information, see page A743.

A533 ISLET BIOLOGY—APOPTOSISCATEGORY

and expressed as steady-state plasma glucose (SSPG); regional fat deposition variables. Receiver operator characteristic curves were used to calculate the was quantifi ed by CT abdomen and MRS liver. At baseline, similarly-obese IR vs. DTI-metrics cutoffs to predict obesity-associated hypothalamic damage. IS subjects (BMI 30) demonstrated signifi cantly increased waist circumference, Results: λ1 values in the hypothalamus were signifi cantly lower in diastolic blood pressure, fasting triglyceride concentrations, visceral adipose obese subjects (P<0.0001). The sensitivity, specifi city, and positive and tissue (VAT), and intrahepatic fat. After mean weight gain of 3.25 kg over four negative predictive values for obesity-associated hypothalamic damage weeks, followed by one week of weight maintenance, tests were repeated. by λ1<1.072 were 75%, 87.5%, 83.3%, and 80.7%, respectively. Patients The group as a whole, experienced signifi cant increases in SSPG (38%), total with hypothalamic λ1<1.072 presented signifi cantly increased values of cholesterol (15 mgdL), triglycerides (45%), waist circumference (3.2 cm). IS BMI, and infl ammatory markers. These fi ndings were in parallel to lower but not IR individuals exhibited signifi cant increases in waist circumference, scores on cognitive tests. Combined BMI and alanine aminotransferase VAT, %VAT, VAT/SAT ratio, and intrahepatic fat. Furthermore, multiple linear was the strongest predictor of obesity-associated hypothalamic damage regression demonstrated signifi cant group (IR/IS) interactions in the relationship (AUC=0.89). between change in regional fat mass and worsening of SSPG: specifi cally, in IS Conclusion: DTI detects obesity-associated hypothalamic damage but not IR individulas, increase in intrahepatic fat correlated with increase in associated with infl ammatory markers and impaired cognitive function. This SSPG (r=0.77, p=0.027, and as did increase in VAT (r=0.63, p=0.06). These results study highlights the potential utility of λ1 as a surrogate marker of obesity- show that short-term weight gain induces insulin resistance in both IR and IS associated hypothalamic damage. individuals, but that IS are more prone to experience signifi cant expansion of intraabdominal and intrahepatic fat, which in turn correlate with development 2092-P of insulin resistance, pointing to a causal relationship. Infl uence of Adiposity on Insulin Requirements and Glycaemic Supported By: ADA (1-11-CT-35) Control in Children and Adolescents with Type 1 Diabetes THOMAS G. COTTER, PAUL JENNINGS, HELEN BURKE, MARCIA BELL, FIDELMA 2090-P DUNNE, SEAN DINNEEN, TIMOTHY O’BRIEN, FRANCIS M. FINUCANE, Galway, Depot specifi c MicroRNA Expression in Human Subcutaneous Ireland Adipose Tissue While excess body fat is associated with insulin resistance, the ADELINE DIVOUX, HUI XIE, JIAN-LIANG LI, KALYPSO KARASTERGIOU, RANJAN infl uence of adiposity on insulin requirements in young people with type 1 J. PERERA, SUSAN K. FRIED, STEVEN R. SMITH, Orlando, FL, Boston, MA, Winter diabetes (T1DM) is not well established. We sought to determine whether Park, FL standardised body mass index (zBMI) infl uenced total daily-, quick acting- Peripheral fat is associated with lower cardiometabolic risk. Physiological and basal-insulin dosing (TDI, QA and BI, respectively) and HbA1c in a cohort differences in gluteal adipocyte functions are known but the molecular of 136 T1DM patients aged 2-20 years attending our university hospital- basis for depot differences in adipocyte function is poorly understood. based diabetes clinic. MicroRNAs (miRNA) have emerged as key post-transcriptional regulators of Mean age was 13.4±4.2 years. 52.2% were female, 97.8% were Caucasian gene expression. As part of a broader exploration to identify the molecular and 32% were overweight or obese. Mean zBMI was 0.55±1.04, zBP was mechanisms underlying the fundamental differences in gluteal-femoral and 0.43±1.1 and HbA1c was 77±17.8 mmol/mol. Mean TDI, QA and BI doses abdominal subcutaneous adipose tissue, we developed the hypothesis that were 47.5±30, 25.6±17.6 and 22±15.9 units per day, respectively. Among a differential miRNA profi le exists between abdominal and gluteal adipose lean, overweight and obese patients, mean TDI was 44.4±25.6, 46.7±7.6 and tissue depots. Furthermore we explored in silico the potential interactions 73.5±47.6 units, respectively (ANOVA p=0.004), with similar fi ndings for QA with HOX gene expression. Abdominal and gluteal adipose tissue aspirates and BI doses. obtained from 9 premenopausal women (age 28.5±2.32 years, BMI 28.16±2.26 In linear regression modelling, there were strong and statistically 2 2 kg/m ) and 9 men (age 30.9±1.93 years, BMI 26.4±1.08 kg/m ) were used to signifi cant associations between zBMI as the exposure and TDI, QA and BI analyze small RNAs (size<35bp) and miRNAs expression profi les by the next- doses as outcomes. Each unit rise in zBMI was associated with an increase generation DNA sequencing. The raw reads were mapped to miRBase v17, of 8.6±2.4, 5±1.4 and 3.5±1.3 units of TDI (p<0.001), QA (p<0.001) and BI and differentially expressed miRNAs were confi rmed by qRT-PCR. (p=0.007), respectively. These associations were similar after adjusting for Among the 1733 known miRNAs, 640 were detected in adipose tissue; age, sex and HbA1c. There was no association between zBMI and HbA1c. 2.1% were differentially expressed between abdominal and gluteal adipose Increased BMI is associated with higher insulin requirements in young tissue. Twenty fi ve miRNA were down-regulated in gluteal adipose tissue people with T1DM, with obese patients requiring approximately 60% more whereas 12 miRNA were up-regulated. MiR196a2, miR196a1, miR196b and insulin than lean ones. Adiposity appears to affect the QA more than the miR204 showed a higher expression in gluteal (fold change=2.7, 2.3, 1.7 and BI dose, which may refl ect higher carbohydrate intake in heavier patients. 2.3 respectively) independent of the sex. MiR205 was exclusively observed Given its strong association with insulin dose, adiposity appears to be an in male gluteal adipose tissue (fold change between male and female 12.4 important determinant of metabolic health in young T1DM patients. for abdominal depot and 154.5 for gluteal depot). Bioinformatic analysis suggested that the differentially expressed miRNAs are modulators of HOX genes expression. ISLET BIOLOGY—APOPTOSIS A specifi c miRNA signature exists for each adipose tissue depot often in a sex specifi c fashion. miRNA’s may explain fat distribution difference observed between individuals. Guided Audio Tour: Beta Cell Apoptosis (Posters: 2093-P to 2100-P), see Supported By: NIH (DK072476, R24DK087669, P30DK46200) page 15.

2091-P & 2093-P Hypothalamic Damage Is Associated with Infl ammatory Markers Differential Cytokine Effects on β-Cell TXNIP Expression and Cognitive Dysfunction in Obese Subjects KYUNGHEE HONG, GUANLAN XU, ANATH SHALEV, Birmingham, AL JOSEP PUIG, GERARD BLASCO, JOSEP DAUNIS-I-ESTADELLA, XAVIER MOLINA, In type 1 diabetes the autoimmune reaction and the release of ROSER RODRIGUEZ, WIFREDO RICART, SALVADOR PEDRAZA, JOSÉ MANUEL proinfl ammatory cytokines (interleukin (IL)-1β, tumor necrosis factors (TNF) FERNÁNDEZ-REAL, Girona, Spain α, and interferon (IFN) γ) leads to β-cell destruction and apoptosis. Recently, Background: Growing evidence implicates hypothalamic infl ammation in we identifi ed thioredoxin-interacting protein (TXNIP) as an important the pathogenesis of diet-induced obesity and cognitive dysfunction in rodent regulator of β-cell apoptosis and found that its deletion prevented type 1 models. Few studies have addressed the association between obesity and and type 2 diabetes. TXNIP is highly induced by glucose via the carbohydrate hypothalamic damage in humans and its relevance. We sought to evaluate response element binding protein (ChREBP), but the effects of cytokines on obesity-associated hypothalamic damage by magnetic resonance imaging TXNIP expression have remained largely unknown. We therefore cultured diffusion tensor imaging (DTI) and its impact on cognitive function. INS-1 β-cells with a cocktail of IL-1β (1 ng/ml), TNFα (5 ng/ml) and INFγ Methods: We prospectively studied 24 consecutive middle-aged obese (5 ng/ml) under low (5 mM) and high (25 mM) glucose conditions and subjects (13 women; 49.8±8.1 years; BMI 43.9±0.92 Kg/m2) and 20 healthy measured TXNIP by quantitative real-time RT-PCR. Cytokine cocktail

POSTERS volunteers (10 women; 48.8±9.5 years; BMI 24.3±0.79 Kg/m2). We analyzed treatment increased TXNIP expression at 5 mM, but not at 25 mM glucose. Islet Biology/ the following DTI-metrics in the hypothalamus: primary (λ1), secondary (λ2), Insulin Secretion Interestingly, we found that this discrepancy was due to IL-1β, which up- and tertiary (λ3) eigenvalues; fractional anisotropy (FA); and mean diffusivity regulated TXNIP mRNA at 5 mM glucose, but down-regulated TXNIP at (MD). A neuropsychological test-battery was used to evaluate cognitive 25 mM glucose. To investigate the molecular mechanism involved in this

For author disclosure information, see page A743. & Guided Audio Tour poster ADA-Funded Research

A534