INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

1770-P & 1772-P DNA Repair Enzyme OGG1 Regulates Obesity and Insulin Sensitiv- Association of a Type 1 Diabetes Genetic Risk Score with C-Peptide ity with Age and Age of Diagnosis in Type 1 Diabetes LARYSA YUZEFOVYCH, MYKHAILO RUCHKO, MICHELE SCHULER, LYUDMILA DELNAZ ROSHANDEL, RICHARD A. ORAM, ANDREW T. HATTERSLEY, MICHAEL RACHEK, Mobile, AL N. WEEDON, ROSE GUBITOSI-KLUG, SHELLEY B. BULL, ANDREW D. PATERSON, Oxidative stress and mitochondrial dysfunction in skeletal muscle are DCCT/EDIC RESEARCH GROUP, Toronto, ON, Canada, Exeter, United Kingdom, important factors leading to insulin resistance (IR) during the aging process Cleveland, OH but the underlying mechanisms are still unknown. Among the potential tar- A proportion of people with type 1 diabetes (T1D) have detectable C-pep- gets is mitochondrial DNA (mtDNA), since mtDNA is highly specialized and tide years after diagnosis: they maintain better glycemic control and experi- encodes for proteins essential for energy metabolism and, also, damage to ence lower risk for diabetic complications. Multiple loci have been identifi ed mtDNA heightens mitochondrial ROS (mtROS) production that is very criti- for risk of T1D, but it is unknown whether they or a T1D genetic risk score cal for IR. Despite extensive age-related studies on mtDNA mutations, until (GRS) are associated with C-peptide. recently, the integrity of mtDNA and repair mechanisms have received little The study population included 1303 Caucasian subjects from the Diabetes attention in diabetes research. We hypothesized that progressive oxidative Control and Complications Trial (DCCT), recruited in 2 cohorts: primary and mtDNA damage triggers development of age-associated oxidative stress in secondary with 1-5, and 1-15 yrs of diabetes, respectively. We calculated skeletal muscle, and thus, potentiates development of IR with age. To vali- a weighted T1D GRS of 30 SNPs of which 5 were in HLA. T1D GRS, a DR3/ date this hypothesis, we performed studies in vivo using DNA repair enzyme, DR4 GRS (2 SNPs) and individual SNPs (coded additively) were tested for 8-oxoguanine (8-OxoG) DNA glycosylase (Ogg1) knockout and overexpressing association with stimulated C-peptide after a standard meal measured at models: 1) Ogg1 -/- (KO); 2) Tg MTS-hOGG1 (Tg, mice overexpressing human DCCT baseline. Separate analyses of primary cohort (N = 651), secondary OGG1 (hOGG1, subunit 1-α) in mitochondria; 3) wild type (WT, ogg1 +/+). We cohort with duration 1-5 yrs (N = 135), and secondary cohort with duration evaluated obesity and IR phenotypes, oxidative mtDNA damage, oxidative 5-15 yrs (N = 517) were combined through meta-analysis using Tobit models stress and expression of several mitochondrial proteins in skeletal muscle for c-peptide adjusting for sex, age at diagnosis and duration. Similarly, T1D from young (~5 month old) and aging (~15 month old) males from all three OGG1 GRS, DR3/DR4 GRS and individual SNPs were tested for association with groups. Our results showed that mitochondrial hOGG1 overexpression amelio- age of diagnosis using linear regression. rated age-associated obesity and IR phenotypes and protected against oxi- T1D GRS and DR3/DR4 GRS were not associated with stimulated C-pep- dative mtDNA damage and oxidative stress. Additionally, protein content for tide. The risk alleles (T) of SNPs in HLA-A24 region (rs1264813, β (SE) = -0.3 transcriptional coactivator peroxisome proliferator activated receptor alpha (0.1), p = 3E-5) and Insulin gene (rs689, β (SE) = -0.2 (0.1), p = 0.01) were (PGC-1α) and mitochondrial protein porin was signifi cantly increased in skel- associated with lower stimulated C-peptide. T1D GRS (β (SE) = -0.8 (0.1), p = etal muscle from aging Tg mice. Collectively, these proof-of-concept results 3E-8), DR3/DR4 GRS (β (SE) = -0.5 (0.2), p = 6E-4) and the risk allele (T) of provide fi rst direct evidence that oxidative mtDNA damage triggers oxidative the non-HLA SNP rs5753037 (HORMAD2, β (SE) = -1.0 (0.3), p = 4E-4) were stress and development of age-associated IR in mice, thus making mtDNA all negatively associated with age of diagnosis, and exceeded Bonferroni damage and repair a novel target for treatment of IR and type 2 diabetes. correction (< 0.002). At all loci, there was no signifi cant heterogeneity of the effect among the 3 subgroups. We identifi ed risk alleles of 2 T1D susceptibility loci, HLA-A24 and INS, INTEGRATED PHYSIOLOGY—INSULIN SECRETION associated with lower C-peptide levels. T1D GRS, DR3/DR4 GRS and IN VIVO rs5753037 were associated with age of diagnosis. Supported By: National Institutes of Health (U01 DK094157, U01 DK094176, DP3 DK104438) Moderated Poster Discussion: Insulin Secretion in Animals and Humans (Posters: 1771-P to 1777-P), see page 14. & 1773-P Racial Differences in Early GIP Contribute to Postprandial Hyperin- & 1771-P sulinemia in Black Women: The Federal Women Study Insulin Receptor Knockout Using Two Beta-Cell-Specifi c Cre Mouse PAOLA C. ALDANA, AMBER COURVILLE, MARY WALTER, MICHELLE T. DUONG, Lines Transiently Improves Glucose Homeostasis BRIANNA A. BINGHAM, LILIAN S. MABUNDO, MADIA RICKS, JOON HA, ARTHUR SØS SKOVSØ, LYNDA ELGHAZI, DEREK A. DIONNE, MELISSA M. PAGE, HONG S. SHERMAN, ANNE E. SUMNER, STEPHANIE T. CHUNG, Bethesda, MD LI, DARIA HUTCHINSON, XIAOKE HU, FARNAZ TAGHIZADEH, ERNESTO BERNAL- Black women are more hyperinsulinemic than whites after exposure to MIZRACHI, JAMES D. JOHNSON, Vancouver, BC, Canada, Miami, FL intravenous glucose but the post-meal insulin response by race is unclear. Insulin signaling is important for glucose homeostasis and is disrupted in Since incretins mediate 50% of postprandial insulin secretion, we hypothe- type 2 diabetes. The insulin receptor is highly expressed in beta cells, where sized that black women would have higher insulin, active glucagon-like pep- its function remains to be fully established. Previous studies on mice gener- tide-1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP) compared ated to delete insulin receptors (InsR f/f) from beta-cells reported impaired glu- to whites during a 2 h mixed meal test (MMT). To quantify insulin and incretin cose tolerance, reduced insulin secretion, and reduced beta-cell mass. How- response, early (0-30 min) and total (0-120 min) area under the curves were ever, the so-called BIRKO model employed Cre recombinase under the control calculated for the MMT (52% carbohydrate, 15% protein, 33% fat) in 36 fed- of a short fragment of the RIP, which led to deletion of the insulin receptor in erally employed women without diabetes (17 African-American, 6 African both beta-cells and the brain. We generated two new mouse models to re- immigrants, 13 whites; age 43±9 (mean±SD), range 25- 62 y; BMI 30.8±5.8, address the function of insulin receptors specifi cally in beta-cells. First, we range 20.5-45.3 kg/m2). Glucose tolerance status was previously determined crossed InsR f/f mice with a mouse line expressing Cre within the endogenous by OGTT and percent (%) body fat by DXA. Analyses of covariance were Ins1 gene locus (Ins1 Cre), as our group and others have shown little or no Ins1 used to examine insulin and incretin response by race accounting for glucose gene expression in the brain. At 25 weeks of age, female InsR f/f:Ins1 Cre mice and % fat. Glucose tolerance status did not differ by race. During the MMT, exhibited improved glucose tolerance relative to InsR wt/wt:Ins1 Cre littermate glucose concentrations did not differ by race (P≥0.50). After adjusting for % f/f Cre Obesity controls. Interestingly, InsR :Ins1 mice were also signifi cantly heavier fat, early insulin response was 50% higher in black vs. whites (insulin0-30min

POSTERS from 7-16 weeks of age. Second, we employed the tamoxifen-inducible Ins1- 1100±770 vs. 719±371, P=0.01) and total insulin response was 20% higher ERT Cre transgenic mouse model, which has been shown to have virtually no (insulin0-120min 8012±5645 vs. 6598±3305 µU·min/mL, P=0.10) but this did not f/f ERT Integrated Physiology/ recombination in the brain. Similarly, InsR :Ins1-Cre mice had signifi cantly reach signifi cance. Postprandial early and total GIP was higher in blacks vs. improved glucose tolerance 4 weeks after tamoxifen injection relative to both whites (GIP0-30min: 3468±3053 vs.1778±1003, GIP0-120min: 54159±22792 vs. InsR wt/wt:Ins1-CreERT and InsR f/f littermate controls, although this reversed to 39849±15582 pg·min/mL, both P<0.03). There were no differences by race impaired glucose homeostasis later in life. There were no signifi cant dif- in GLP-1 or C-peptide concentrations. Early GIP (P<0.001) and % fat (P<0.001) ferences in body weight in the tamoxifen-inducible model, suggesting that mediated 46% of the racial differences in early insulin response. Greater effects of beta-cell InsR deletion were restricted to developing and/or young early GIP was related to black race (P=0.03), % fat (P=0.04), and glucose0-30min mice. Collectively, these data are consistent with the concept that insulin (P<0.01). Overall, racial differences in postprandial insulin response were inhibits its own secretion, at least initially. Together, these data provide new identifi ed during a meal and may be partially explained by higher early GIP, information about the function of beta-cell insulin receptors and will provide not GLP-1, in black women relative to whites. more insight into the pathogenesis of type 2 diabetes. Supported By: National Institute of Diabetes and Digestive and Kidney Diseases Supported By: Canadian Institutes of Health Research

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A458 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

& 1774-P & 1776-P Greater Increase in Insulin Clearance after Gastric Bypass vs. Gas- Proinsulin as a Marker of Beta-Cell Health tric Banding in Patients with Type 2 Diabetes HILARY R. THOMAS, JACQUELINE HOWELLS, JEFFREY A. BLUESTONE, San Fran- MARLENA HOLTER, SARAH STANO, FATIMAH RIMAWI, MARGARITA SALA, cisco, CA CAROLINA ESPINOSA, JAMES MCGINTY, SCOTT BELSLEY, NINAN KOSHY, ROX- Type 1 diabetes (T1D) is an autoimmune disease thought to result in com- ANNE DUTIA, BLANDINE LAFERRERE, New York, NY plete depletion of the insulin producing β-cells. However, 20-60% of patients About 70% of secreted insulin is cleared by the liver prior to entering the with longstanding T1D produce detectable levels of C-peptide (CP) and systemic circulation. Hepatic insulin clearance is an important regulator of 10-15% produce > 0.6 ng/mL of CP, which is associated with fewer micro- glycemia. Fasting and post-oral insulin clearance rate (ICR) are elevated and macro-vascular complications and more stable glycemic control. Proin- after gastric bypass (GBP). Whether the effect of GBP on ICR is due to a sulin (PI) is produced by the endoplasmic reticulum (ER) and is then cleaved mechanism other than weight loss is not known. We hypothesized that: 1.) in secretory granules to produce insulin and CP. Normally, little PI is secreted ICR will increase more after GBP than after matched weight loss by gastric and when PI is increased relative to insulin and CP, it is thought to refl ect banding (GB); 2.) The difference in ICR after oral vs. IV glucose will be exag- changes in protein folding and processing in the ER or ER stress. Emerging gerated after GBP. We calculated ICR fasted (ICR-f), after oral glucose (ICR- data indicate that ER stress might accelerate β-cell death. We characterized o) and after an iso-glycemic IV glucose infusion (ICR-iv), as the molar ratio of β-cell function in 27 patients (36.5 +/- 14.1 years) at least 4 years following C-peptide to insulin and as the ratio of insulin secretion rate (ISR) to insulin diagnosis (13.4 +/- 11.5 years). We evaluated β-cell function using random levels, in patients with type 2 diabetes (T2D) before and after 10% matched CP and PI and β-cell stress using PI:CP and investigated the relationship weight loss by either GBP (n=26) or GB (n=15). Before surgery: BMI, HbA1c, with clinical parameters. Sixteen of 27 patients had detectable CP levels diabetes duration, HOMA-IR, ICR, glucose infused, GLP-1, GIP, the incretin (0.90 +/- 1.0 ng/mL). All patients with detectable CP values also had detect- effect and ISR were not different between groups. ICR-o tended to be lower able PI values. Importantly, all 11 patients with undetectable CP levels had than ICR-iv with no difference between groups (p=0.559). After weight loss, detectable PI levels (14 +/- 12.6 pM), indicating functionality of some β-cells. HOMA-IR, glucose infused and ISRAUC180 changed similarly in the 2 groups. Patients with detectable CP values had signifi cantly lower PI:CP values com- GLP-1 and GIP increased only after GBP. The incretin effect increased more, pared to patients with no detectable serum CP (111.6 ± 48.72, n=16 vs. 490.6 and glucose (fasting, 120’ and AUC) decreased more after GBP than after ± 194.5, n=11), even after matching for HbA1c, age and duration of diabetes. GB. ICR-f, ICR-iv, and ICR-o increased in both groups, but only signifi cantly This indicates that there may be recoverable β-cells even in patients with after GBP, and the increase in ICR-iv was greater than ICR-o (0.46±0.24 vs. no detectable CP. Targeting ER stress in this population may improve insulin 0.15±0.24, p<0.001). However, ICR-o did not correlate with GIP or GLP-1. ICR- production and clinical outcomes. f, ICR-o and ICR-iv were inversely correlated with HOMA-IR, ISR-o and ISR- Supported By: JDRF; The Leona M. and Harry B. Helmsley Charitable Trust iv, respectively. Conclusion: The increase in ICR after GBP is not evident after GB. The & 1777-P determinants of ICR may differ after GBP and GB. The inverse relationship Reduced Beta-Cell Secretory Capacity in Pancreatic Insuffi cient between ICR and insulin concentrations suggests that insulin itself deter- Cystic Fibrosis despite Normal Glucose Tolerance mines ICR. In addition, the lesser rise in ICR-o vs. ICR-iv suggests an incretin SABA SHEIKH, LALITHA GUDIPATY, DIVA D. DE LEON, DENIS HADJILIADIS, effect in the suppression of ICR-o. CHRISTINA KUBRAK, AMY PELECKIS, NORA ROSENFELD, SARAH NYIRJESY, Supported By: American Diabetes Association (7-08-CR-39 to B.L.); National SALONI MALIK, MARINA CUCHEL, DARKO STEFANOVSKI, RONALD C. RUBEN- Institutes of Health STEIN, ANDREA KELLY, MICHAEL R. RICKELS, Philadelphia, PA, Newark, DE Patients with pancreatic insuffi cient cystic fi brosis (PI-CF) are at increased 1775-P risk for diabetes development, attributed to insulin insuffi ciency. We hypoth- esized that even with normal glucose tolerance (2 hr plasma glucose (PG2) WITHDRAWN < 140 mg/dL and 1 hr PG (PG1) < 200 mg/dL during oral glucose tolerance testing (OGTT)), individuals with PI-CF would manifest insulin secretory defects relative to pancreatic suffi cient CF (PS-CF) and control subjects. We determined β-cell secretory capacity and insulin secretory rates (ISR) from glucose-potentiated arginine (GPA) and mixed meal tolerance tests (MMTT), respectively, in PI-CF (n = 11), PS-CF (n = 9) and matched controls (GPA, n = 11; MMTT, n = 10). Acute insulin secretory responses were determined at base- line (AIRarg) and during hyperglycemic clamps at 230 mg/dL (AIRpot) and 340 mg/dL (AIRmax). Acute C-peptide (ACR), proinsulin (APR), and glucagon (AGR) responses were also calculated. Incremental areas under the curve for ISR, gastric inhibitory peptide (GIP) and glucagon like peptide (GLP-1) were calcu- lated within 30 (AUC30min) and 180 (AUC180min) minutes post meal ingestion. PI-CF had signifi cantly higher OGTT PG1 than PS-CF and controls (p < 0.05). AIRpot, ACRpot, ACRmax, and AGRs during GPA were lower in PI-CF vs. controls and PS-CF (p < 0.05). There was a trend towards higher proinsulin: C-peptide secretory ratios (PCSR) during the hyperglycemic clamps in PI-CF (p<0.1). During the MMTT, the ISR, GLP-1, and GIP AUC30min were lower in PI-CF (p < 0.05). While the AUC180min ISR was similar between PI-CF and controls, the AUC180min glucose remained higher (p = 0.004). Despite “normal” glu- cose tolerance, young adults with PI-CF have greater glucose excursions, impaired early phase insulin and incretin secretion, and decreased β-cell

secretory capacity; likely resulting in clinically relevant hyperglycemia at Obesity

1 hr on OGTT. Furthermore, increased PCSR during hyperglycemia suggest POSTERS PI-CF β-cells are susceptible to glucose stress. Integrated Physiology/ 1778-P The Roles of NOD Receptors in Fat-induced Beta-Cell Dysfunction ALEXANDER IVOVIC, TEJAS DESAI, KHAJAG KOULAJIAN, KAITAI YE, YUSAKU MORI, ADRIA GIACCA, Toronto, ON, Canada Studies suggest intracellular nucleotide-oligomerization domain (NOD) receptors of innate immunity play a signifi cant role in metabolism and diabe- tes. The bacterial cell wall components meso-glutamyl diaminopimelic acid (DAP) and muramyl dipeptide (MDP) are endogenous ligands of NOD1 and NOD2 receptors, respectively. NOD1 activation leads to a proinfl ammatory

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A459 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

response, whereas NOD2 has an immunoregulatory role. Saturated fat can Results: For participants with CKD, correlation of SIclamp with Matsuda also activate and synergize with NOD activators. Finally, recent data sug- (r=0.45) was lower than for participants without CKD (r=0.58). For 1/HOMA-IR, gest NOD1-null mice have better glucose tolerance with high fat feeding, the correlation in CKD was (r=0.35) was lower than those for participants whereas in NOD2-null mice glucose tolerance is worse, than controls. This without CKD (r=0.51). When adjusted for sex, age, race, and SIclamp, partici- suggests a β-cell role of NODs since glucose tolerance refl ects β-cell func- pants with CKD had a mean Matsuda that was 1.0 lower (95% CI -2.0, 0.1; tion. NOD1/2 mRNA was assessed in islets. Wild-type (WT) islets were then p=0.07) and a mean 1/HOMA-IR that was 0.3 (dL·mL)/(µIU·mg) lower (CI -0.5, exposed to the NOD1 ligand FK565 and the NOD2 ligand L18-MDP in the 0.0; p=0.03). Holding sex, age, race and SIclamp constant, obese subjects presence or absence of palmitate (and oleate as negative control) for 48 h, had a mean Matsuda index that was 2.6 lower (95% CI -3.7, -1.5, p<0.0001) followed by glucose-stimulated insulin secretion. NOD1-KO islets were also and a mean 1/HOMA-IR that was 0.5 (dL·mL)/(µIU·mg) lower (95% CI -0.7, - exposed to palmitate. Ex vivo studies were performed in islets of WT and 0.3, p<0.0001). Correlation coeffi cients of BMI with SIclamp, Matsuda, and 1/ NOD2-KO mice following 96 h infusion of saline or Intralipid. Finally, WT mice HOMA-IR were -0.19, -0.56, and -0.52, respectively. were injected with FK565 or MDP or saline 6 h before undergoing hypergly- Conclusions: Matsuda and 1/HOMA-IR underestimate insulin sensitiv- cemic clamps in vivo. Islets express NOD1/2 mRNA. FK565 decreases β-cell ity compared to clamp in CKD and obesity, while CKD also introduces less function in vitro, whereas MDP alone does not have any affect. FK565 has precision. Clamp and surrogate indices derived from fasting or OGTT mea- an additive effect with palmitate in decreasing β-cell function and MDP surements may refl ect different aspects of insulin action that may vary by appears to prevent palmitate- and oleate-induced β-cell dysfunction. NOD1- disease state. KO islets are protected from palmitate-induced β-cell dysfunction. Ex vivo Supported By: National Institute of Diabetes and Digestive and Kidney Diseases data also suggest NOD2 deletion increases susceptibility of β-cells to fat- induced dysfunction. Finally, the NOD1 activator decreased β-cell function 1781-P whereas the NOD2 activator did not have a signifi cant effect on β-cell func- Circulating GLP-1 May Not Be the Main Determinant of Insulin tion in vivo. These data suggest NOD1 plays a detrimental, and NOD2 a ben- Release after Roux-en-Y Gastric Bypass efi cial, role in modulating β-cell function in the presence and absence of fat KARIN STERL, SONGYAN WANG, BRUCE W. PATTERSON, LAUREN ZEITER, and suggest their potential as novel therapeutic targets. DOMINIC N. REEDS, BURTON M. WICE, St. Louis, MO Supported By: Banting and Best Diabetes Centre; Canadian Institutes of Health Xenin-25 (Xen) is a 25-amino acid neurotensin-related peptide secreted by Research a subset of enteroendocrine cells. Xen effects are mediated by neurotensin receptor 1 present on neurons. In our previous study, continuous intravenous 1779-P infusion of Xen during liquid mixed meal tolerance tests in humans with nor- mal glucose tolerance (NGT) potently inhibited glucagon-like peptide-1 (GLP- WITHDRAWN 1) secretion without reducing insulin secretion rates (ISRs). Thus, circula t ing GLP-1 is not required for the incretin response in healthy humans. Roux-en-Y gastric bypass (RYGB) is associated with marked improvements in glucose homeostasis, thought to be related to the rapid and profound increase in postprandial GLP-1 release. The current study was designed to determine if exogenously administered Xen also modulates GLP-1 levels and the incretin response in humans after RYGB. Fasted subjects (n=8) with prior RYGB and no history of type 2 diabetes ingested a liquid mixed meal at time zero. On separate visits, a primed- constant intravenous infusion of albumin or Xen at 4 (Lo-Xen) or 12 (Hi- Xen) pmoles/kg/min was administered from -15 until 300 minutes. Plasma glucose, insulin, C peptide, glucagon, Xen, GIP, GLP-1, pancreatic polypep- tide (PP), and neuropeptide Y (NPY) were measured and ISRs calculated. Repeated measures 2-way ANOVAs and AUCs assessed treatment effects. Responses were compared to those in well-matched healthy humans from our earlier study. Hi-Xen infusion in subjects with RYGB did not affect glucose, glucagon and GLP-1 responses and ISRs but still amplifi ed PP and NPY release (cholin- ergic input to islets and sympathetic tone, respectively). Thus, RYGB disrupts a neural circuit that inhibits GLP-1 release. Early insulin and GIP responses were superimposable and preceded GLP-1 release. Compared to the non- surgical group, the ISR to GLP-1 ratio was reduced 5-fold but the ISR to GIP ratio was increased 2-fold. Hence, GIP may play an underappreciated but important role in regulating ISR after RYBG. Supported By: National Institutes of Health

1782-P The Postprandial Serum C-Peptide Correlate with Beta-Cell Func- tion in Japanese Patients with Type 2 Diabetes Mellitus 1780-P YUKI MATSUHASHI, SHINJI CHIKAZAWA, HIDEYUKI OTAKA, KAZUHISA TAKA- Effect of Chronic Kidney Disease and Obesity on Insulin Sensitivity HASHI, AYA KANBA, YUKI KIMURA, KOKI MATSUMURA, HIROSHI MURAKAMI, Indices KOTA MATSUKI, ERI SATO, JUTARO TANABE, MIYUKI YANAGIMACHI, HIROSHI

Obesity IRAM AHMAD, LEILA ZELNICK, KRISTINA M. UTZSCHNEIDER, STEVEN E. KAHN, MURAKAMI, MAKOTO DAIMON, Hirosaki, Japan

POSTERS IAN H. DE BOER, Seattle, WA The onset of type 2 diabetes mellitus (T2DM) is based on the insulin resis- Introduction: Insulin sensitivity can be measured by hyperinsulinemic- tance and decreasing of insulin secretion (loss of β-cell function). Glucagon

Integrated Physiology/ euglycemic clamp or estimated using surrogate indices. Chronic kidney induces insulin secretion by directly stimulating β-cells, therefore glucagon disease (CKD) and obesity may differentially affect insulin sensitivity esti- stimulaton test (GST) is a reliable marker for β-cell function. Meal also stim- mates, compared with clamp measurements, due to differences in insulin ulates insulin secretion after ingestion. We evaluated the effi cacy of meal kinetics and organ-specifi c insulin responses. tolerance test (MTT) as a tool for evaluating β-cell function in T2DM. Methods: In a cross-sectional study of 59 subjects with nondiabetic CKD Japanese T2DM patients with inadequate glycemic control (HbA1c>7.0) (estimated GFR <60 mL/min/1.73 m2) and 39 healthy controls, we quanti- were enrolled in the study (n=216, insulin therapy =155). Fasting plasma glu- fi ed insulin sensitivity by clamp (SIclamp), OGTT (Matsuda index), and fast- cose, C-peptide (CPR) and HbA1c were measured. For the MTT, postprandial ing glucose and insulin (1/HOMA-IR). We compared the Matsuda index and plasma glucose and CPR were measured. MTTΔCPR (postprandial CPR-fast- 1/HOMA-IR to SIclamp using descriptive statistics, graphical analyses, cor- ing CPR) was calculated. For the GST, CPR was determined before and 6 relation coeffi cients, and linear regression. Multivariable modelling was min after 1mg of glucagon injection. GSTΔCPR (CPR [6 min]-CPR [0 min)] was adjusted for age, sex, and race. calculated. Factors correlated with GSTΔCPR were analyzed using simple

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A460 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER and multiple stepwise regression analysis. P value<0.05 was defi ned as sta- mice, respectively, with expansion of β-cell mass correlated (R2=0.854) with tistically different. HbA1c (beta coeffi cient=0.449, p=0.054) and body weight (beta coeffi cient= Fasting CPR, postprandial CPR and MMTΔCPR were 1.85±1.00, 4.43±2.78 0.562, p=0.023). Before onset of obesity in 4 wk KKAy mice, islet GLUT2 was and 2.58±21.4 ng/ml, respectively. GSTΔCPR was 1.64±1.03 ng/ml. Simple already reduced by 75% in fasted mice, cell-surface GLUT2 expression was regression analysis showed that GSTΔCPR was signifi cantly correlated with 32% more robust in fed mice, and HbA1c was increased by 16%. With con- age, height, body weight (BW), BMI, gender, duration of diabetes, diabetic tinued uncontrolled hyperglycemia, GLUT2 transporters were progressively retinopathy, insulin therapy, fasting CPR, postprandial CPR and MMTΔCPR. internalized to a maximum of 85% and GLUT2 content was reduced by 76%. The postprandial CPR in the patients with DPP-4 inhibitor (DPP-4i) and/or In KK mice, islet GLUT2 transiently decreased by 84% then recovered by 16 biguanide (BG) therapy was signifi cantly higher than that without. Multiple wks, while β-cell GLUT2 remained stable in the membrane. Food intake regu- stepwise regression analysis revealed that independent factors contribut- lated cell-surface expression of GLUT2 independently of glycemic control. At ing to GSTΔCPR were BW (β=0.367, p<0.001), postprandial CPR (β=0.527, 4 wk, food intake reduced membrane GLUT2 by 50% and 20% in KK and KKAy p<0.001), presence of diabetic retinopathy (β= -0.134, p=0.009), DPP-4i (β= mice, respectively. In contrast, at 8 wk, food intake increased membrane -0.124, p=0.012) and BG (β= -0.142, p=0.007). GLUT2 by 10% and 59% in KK and KKAy mice, respectively. In addition, food The present date demonstrated that postprandial CPR is valuable in intake increased islet GLUT2 by 51% in only KKAy mice. Reduced islet GLUT2 assessing β-cell function and can help clinicians in clinical practice. and altered food-induced traffi cking of GLUT2 may be among the earliest defects in β-cell function contributing to persistent hyperglycemia. 1783-P Supported By: Midwestern University Activation of Angiotensin Type 2 Receptors Protects Pancreatic Islet Function in Obese Rats Induced by High-Fat Diet 1785-P MIN LIU, XIAOJING LI, ANPING WANG, YU LIU, DANQING JING, SHINAN YIN, Increase in Urinary N-acetyl-β-D-glucosaminidase Excretion YIMING MU, Beijing, China Might Refl ect Glucose Fluctuation and Decreased Insulin Secretary Activation of Ang II type 2 receptor (AT2R) has been examined as a Capacity in Patients with Type 2 Diabetes potential therapeutic strategy in cardiovascular and central nervous sys- SO RA KIM, JI-YEON LEE, GYURI KIM, EUGENE HAN, YONG-HO LEE, EUN SEOK tems. However, there is few fi ndings in insulin resistance and diabetes mel- KANG, CHUL WOO AHN, BONG-SOO CHA, BYUNG-WAN LEE, Seoul, Republic of litus type 2 pathogenesis. In the current study, we evaluated the effects Korea of Compound 21 (C21), a nonpeptide AT2R agonist, on islets in obese rats Urinary N-acetyl-β-D-glucosaminidase (NAG) present in high concentra- induced by high-fat diet (HFD). Adult male Sprague-Dawley (SD) rats were tion in lysosomes of proximal tubular cells is released into the urine after randomly assigned into fi ve groups: normal (fed with normal diet), HFD and renal proximal tubule injury. By comparing glomerulopathic marker of albu- HFD respectively plus telmisartan (Tel, 1 mg/kg/d), C21 (1 mg/kg/d), Tel+C21. minuria, we investigated the clinical relevance of urinary NAG, a renal tubu- Tel and C21 were continually given by oral administration for four weeks. lopathic marker, regarding demographic, gluco-metabolic parameters (glu- After treatment, the rats received an intraperitoneal glucose tolerance test, cose, glycated hemoglobin (HbA1c), glycated albumin (GA), insulin, C-peptide, and the pancreas were saved to examine islet morphology and biochemi- HOMA-IR, HOMA-β, and postprandial C-peptide-to-glucose ratio (PCGR)) cal parameters of insulin secretion. We found that, compared with control and nephropathic parameters including albumin-to-creatinine ratio (ACR), HFD rats, those HFD rats treated with C21 alone and combined with Tel dis- glucose-to-creatinine ratio (GCR), and eGFR. A total of 563 participants with played lower blood glucose lever, higher serum insulin concentration and type 2 diabetes (T2D) were enrolled. Median age and diabetes duration were improved glucose tolerance. These rats had more integrated islets, larger 62.0 and 8.75 years, respectively. Median HbA1c was 7.30%. In correlation positive insulin-staining islet mass and up-regulated insulin and GLUT2 pro- analysis, urinary NAG was more strongly correlated with BMI (r=-0.222; tein expressions accompanied with slightly higher AT2R expression in the P<0.001 vs. r=-0.015; P=0.741), plasma postprandial glucose (r=0.251; P<0.001 pancreas. These data suggest that C21 protects pancreatic islets function vs. r=0.076; P=0.099), GA (r=0.204; P<0.001 vs. r=0.132; P=0.005), PCGR (r=- against lipotoxicity via activatin of AT2R. 0.171; P=0.001 vs. r=-0.087; P=0.107), and HOMA-β (r=-0.100; P=0.049 vs. Figure. r=-0.015; P=0.791) than urinary ACR. In the multiple regression analysis, age (STD β=0.26), GA (STD β=0.13), plasma postprandial glucose (STD β=0.13), and urinary ACR (STD β=0.27) predicted increased urinary NAG. We classi- fi ed the subjects with normoalbuminuria into two groups according to NAG levels. The participants with increased urinary NAG levels (>4 U/gCr, group II) were older (62.0 vs. 52.0 years) and had signifi cantly higher levels of plasma postprandial glucose (11.5 vs. 9.44 mmol/l), GA (19.5 vs. 16.9%) and urinary GCR (0.12 vs. 0.07 mg/mg) than those with normal urinary NAG levels (group I). From these results and based on previous reports on GA, we postulate that increase in urinary NAG might be related to plasma glucose fl uctuation and decreased insulin secretary capacity in patients with T2D.

INTEGRATED PHYSIOLOGY—LIVER

Moderated Poster Discussion: Integrated Physiology—Liver Supported By: National Natural Science Foundation of China (Posters: 1786-P to 1792-P), see page 15.

1784-P & 1786-P Effect of Hyperglycemia and Food Intake on Beta-Cell Mass and Suppression of Lipolysis Is Not the Major Mechanism by which Obesity Membrane GLUT2 in 4-32 wk Female KK and KKAy Mice a Rise in Insulin Secretion Inhibits Hepatic Glucose Production POSTERS KATHY J. LEPARD, JOSEPH CELLINI, KUNAL NAIK, BAO CHAU TRAN, RODNEY (HGP) FULLMER, VICTORIA ANDROS, STEPHEN GUNNINK, RYAN KING, STACEY TIN- DALE S. EDGERTON, GUILLAUME KRAFT, L. MERKLE MOORE, BEN FARMER, Integrated Physiology/ KOFF, Downers Grove, IL MARTA SMITH, JON HASTINGS, IAN HAJIZADEH, ALAN D. CHERRINGTON, Type 2 diabetes mellitus may be averted by expansion of pancreatic Nashville, TN β-cell mass with suffi cient cell-surface expression of GLUT2. The duration Recent studies in rodents concluded that suppression of lipolysis and and severity of hyperglycemia required to alter β-cell mass and cell-surface reduced FFA fl ux to the liver is the major mechanism by which insulin inhib- GLUT2 expression was determined in fed and fasted female KK and KKAy its HGP (Cell 160 (4):745-758, 2015). In that study insulin was infused into a mice. Fixed pancreata from 4-32 wk mice were sectioned and H and E stained peripheral vein, underexposing the liver to insulin relative to adipose tis- or immunostained for GLUT2 and insulin. Islet size and number were deter- sue. Therefore the present study tested the importance of a decrease in FFA mined, GLUT2 location was blindly scored by three observers using a Likert to the control of HGP during a physiologic increase in insulin. Two weeks scale, and islet section GLUT2 content was estimated by relative intensity prior to study 10 dogs underwent trans-liver vascular catheterization. Dur- of immunofl uoresence. Islet size doubled after 16 or 24 wks in KKAy or KK ing a euglycemic hyperinsulinemic clamp insulin was infused intraportally

ADA-Supported Research & Moderated Poster Discussion For author disclosure information, see page A696.

A461 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

(1.8 mU/kg/min) resulting in a 5-fold increase in arterial and hepatic insulin & 1788-P levels, although liver insulin exposure was 2.5-fold greater than adipose, Liver-specifi c Knockout of DsbA-L Impairs Mitochondrial Function as occurs when insulin is secreted by the pancreas. In the control group and Exacerbates Diet-induced Hepatosteatosis and Insulin Resis- (CON; n=5), hyperinsulinemia quickly suppressed lipolysis and circulating tance FFA, while infusion of intralipid plus heparin in a 2nd group (IL; n=5) pre- HONGZHI CHEN, YUN ZHANG, JULI BAI, LILY Q. DONG, FENG LIU, Changsha, vented that fall. Net hepatic glucose output was rapidly reduced in both China, San Antonio, TX groups (down 75±13 and 85±10% at 1 hr in CON and IL, respectively), even- Mitochondrial dysfunction is implicated in various metabolic diseases tually being completely suppressed, regardless of whether net hepatic FFA including type 2 diabetes, yet the underlying mechanisms remain elusive. uptake was reduced (down 67±16%) or not (3±14%). Therefore under physi- We found that the expression levels of disulfi de-bond-A oxidoreductase-like ologic hyperinsulinemic conditions it is the direct effect of insulin on the liver protein (DsbA-L), a protein that is highly abundant in mitochondria, are signif- which inhibits HGP; the fall in FFA has little to no impact. icantly suppressed in the livers of obese mice and humans compared to their Figure. respective controls. Liver-specifi c disrupting DsbA-L expression exacerbates high fat diet-induced glucose intolerance, insulin resistance, and hepatoste- atosis. Suppressing DsbA-L expression in mouse hepatocytes reduced com- plex I activity, blunted ATP synthesis, and enhanced reactive oxygen spe- cies production. Yeast two-hybrid screening revealed that DsbA-L binds to NADH dehydrogenase (ubiquinone) Fe-S protein 2 (NDUFS2), an essential component of the mitochondrial respiratory chain complex I. Overexpression of wild type but not an NDUFS2-binding-defi cient mutant of DsbA-L reduces DsbA-L defi ciency-induced reactive oxygen species levels in hepatocytes. In summary, our study identifi es DsbA-L as a novel regulator of mitochondrial function and reveals that hepatic DsbA-L down-regulation contributes to obesity-induced hepatic dysfunction and whole body insulin resistance. Supported By: National Basic Research Program of China; National Natural Sci- ence Foundation of China; National Institutes of Health

& 1789-P Deletion of Cannabinoid 1 Receptor Prevents the Aging Phenotype of the Liver in Mice Supported By: National Institutes of Health ISABEL GONZALEZ MARISCAL, ERIN HERCULES, JOSEPHINE M. EGAN, Balti- more, MD & 1787-P The endocannabinoid (EC) system, particularly cannabinoid receptor 1 Physical Cell-Cell Interaction Regulates Hepatic Glucose Metabo- (CB1), plays a substantial role in regulating metabolism in peripheral organs lism in Mice such as liver, gastrointestinal tract, pancreas and adipose tissue. CB1 acti- KYOICHIRO TSUCHIYA, YASUTAKA MIYACHI, CHIKARA KOMIYA, KUMIKO vation, specifi cally in liver, has been linked to progression of liver disease in SHIBA, NORIKO SHIMAZU, SHINOBU YAMAGUCHI, MIZUKO OSAKA, MASAYUKI obesity and alcoholism. In contrast, blockade of CB1 in the periphery leads YOSHIDA, KOJI INOUE, KENJIRO WAKE, YUTA SATO, JUNICHI KIKUTA, MASARU to benefi cial metabolic effects independent of food intake. To determine ISHII, YOSHIHIRO OGAWA, Bunkyo, Japan, Tokyo, Japan, Yokohama, Japan, Suita, the mechanism of EC system regulation in liver and its impact on meta- Japan bolic changes with age, we developed a liver specifi c CB1 knockout mice - - Objective: Obesity-associated leukocyte infi ltration into the liver has (HepCB1 / ) and compared the phenotypical profi le to wild type (Wt) mice as a pivotal role in the pathogenesis of glucose intolerance. Liver sinusoidal well as CB1 global knockout mice (CB1KO). During the span of 3-18 months endothelial cells (LSEC) form the sinusoidal wall between hepatocytes and on standard chow diet, Wt mice had gradual weight gain with age going - - sinusoidal lumen, and regulate cell traffi cking and infi ltration via adhesion from 31g to 46.7±5.4g, unlike HepCB1 / and CB1KO mice who maintained at - - molecules. However, the roles of LSEC in obesity-associated leukocyte infi l- 28.3±0.5g and 27.7±0.4g, respectively. The leaner phenotype of HepCB1 / tration into the liver are unclear. In addition, the regulation of hepatic glu- and CB1KO mice was corroborated with NMR body composition analysis. cose metabolism through physical cell-cell interaction between infi ltrated Plasma leptin levels were found to rise with age in Wt mice, peaking at 24.3 - - leukocytes and hepatocytes remains unknown. ng/mL, whereas HepCB1 / and CB1KO mice had lower levels (7.4 and 2.5 Methods and Results: In LSEC from high-fat diet (HFD)-fed mice, genes ng/mL, respectively). Plasma EC levels, such as AEA, PEA, and OEA, were - - related to adhesion molecules were upregulated compared to those from also lower in the HepCB1 / and CB1KO. Consistent with the physical profi le, standard diet-fed mice. Intravital imaging for Lysozyme M (LysM)-eGFP mice Wt mice progressively accumulated triglycerides in liver with age, which was - - demonstrated that the number of adhered LysM-positive cells to the sinusoi- dramatically reduced in HepCB1 / and CB1KO mice. Wt mice also showed dal wall was enhanced in ob/ob: LysM-eGFP mice compared to those in LysM- pronounced age-related infl ammation of the liver as a result of immune cell - - eGFP mice. In vivo blockade of VLA-4, a counterpart of an adhesion molecule infi ltration and increased MCP-1 levels, which were mitigated in HepCB1 / VCAM-1, by blocking antibodies improved HFD-induced glucose intolerance and CB1KO mice. These results indicate that deletion of CB1 in only the liver associated with decreased leukocyte infi ltration into the liver. In the liver of is just as benefi cial to the aging phenotype of the liver as global CB1 dele- HFD-fed mice, Notch signaling, which is activated through cell-cell contact, tion. For the fi rst time, a relationship between fatty liver, CB1, and aging was attenuated by VLA-4 blockade. Co-culturing with leukocytes upregu- has been demonstrated, suggesting pharmacological blockade of peripheral lated gluconeogenic genes in hepatocytes via Notch-dependent manner. CB1 can protect the liver from a variety of stressors that trigger the onset Conclusion: Our data indicate that LSEC mediate HFD-induced glucose of liver disease. intolerance via VLA-4-mediated leukocyte adhesion. The infi ltrated leuko-

Obesity cytes could increase glucose production in hepatocytes by cell-cell contact- & 1790-P

POSTERS triggered activation of Notch signaling. Our observation suggests a novel A Systems Transcriptomic Approach in Mice Identifi es ACOT8 as a mechanism of hepatic glucose metabolism regulated by physical cell-cell Gene Regulating Hypertriglyceridemia in Humans

Integrated Physiology/ interaction of leukocytes with LSEC and hepatocytes. LUDIVINE DORIDOT, SARAH KRAWCZYK, MISUNG KIM, MAGALI SOUMILLON, Supported By: Japan Diabetes Society; Uehara Memorial Foundation, MSD K.K.; TARJEI MIKKELSEN, MICHELLE LAI, MARK HERMAN, Boston, MA, Cambridge, AstraZeneca; Banyu Life Science Foundation International; Japan Foundation for MA Applied Enzymology; Novartis Pharma LLC; Eli Lilly and Company; Sanofi Excessive fructose consumption contributes to the epidemic of cardiomet- abolic disease. We determined that male C3H/HeJ mice fed a high-fructose diet (HFrD) for 6 weeks develop obesity, hypertriglyceridemia, and hyperin- sulinemia (p<0.05 for each trait compared to chow), whereas HFrD had no effect in C57/Bl6 mice. HFrD fed F2 progeny of a C3HxC57 cross showed large variation in body weight and serum triglycerides and insulin indicative of complex, multigenic traits. As fructose is largely metabolized in the liver, we

For author disclosure information, see page A696. & Moderated Poster Discussion ADA-Supported Research

A462 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER assessed the transcriptome of 48 HFrD fed F2 mice to map hepatic gene pro- reduction in fasting blood glucose (FBG) and ~30% reduction in normalized grams contributing to fructose-induced metabolic disease. Using weighted area under the curve (AUC) of glucose tolerance tests (GTTs) compared to correlation network analysis, we identifi ed a gene expression module con- wild type (WT) littermates (WT-n=30; KO-n=20). Lower (~36%) fasting insu- sisting of 312 genes (out of 8563 detected genes) highly correlated with lin levels and normal glucose-stimulated insulin in LILKO mice, in addition to serum triglycerides (R=0.45, p=0.001). Gene-set analysis revealed that this glucose tolerance data, indicate improved insulin sensitivity. However, we module is enriched for genes involved in lipid metabolism including steroid observed no effect on gene expression of phosphenolpyruvate carboxyki- biosynthesis (p=1.7E-7), PPAR signalling (p=1.4E-6) and fatty acid metabolism nase (PEPCK) or glucose-6 phosphatase catalytic subunit (G6PC) (WT-n=6; (p=1.6E-3). We also observed that this module is enriched for genes in prox- KO-n=7). Interestingly, we found that LILK-KO mice have a ~50% decrease imity to polymorphisms associated with hypertriglyceridemia in humans. in P38 MAPK (P38) protein activation and ~60% decrease in PPARγ gene One such gene was Acot8, a member of the acyl-CoA thioesterase family expression. Previous reports demonstrated that activation of P38 stimulates implicated in the regulation of peroxisomal lipid metabolism. The human hepatic glucose production pathways through activation of PPARγ coacti- ACOT8 gene is located near a SNP (rs4810479) highly associated with hyper- vator 1 alpha Therefore, the activation of integrin-stimulated pathways in triglyceridemia in humans. This SNP was previously “mapped” to another hepatocytes may contribute to hepatic insulin resistance through ILK-depen- gene in this locus, PLTP. In human liver samples, we determined that ACOT8 dent activation of P38 pathways and inappropriate hepatic glucose output. expression is signifi cantly higher (p=0.028) in individuals homozygous for the C (risk) allele (n=18) at rs4810479 compared to those homozygous for the T 1793-P allele (n=55), while the expression of PLTP was not different. By integrating ANGPTL8 (Betatrophin) Relates to Hepatic Steatosis in Two Inde- a systems transcriptomic approach in genetically diverse mice with bioin- pendent Clinical Collectives formatic analysis of human metabolic traits, we identify ACOT8 as a new CHRISTIAN VON LOEFFELHOLZ, ANDREAS F.H. PFEIFFER, STEFFI LIESKE, JENNI- candidate gene to regulate hypertriglyceridemia in humans. FER KRIEBEL, NATALIA RUDIVICH, MARTIN STOCKMANN, JOACHIM SPRANGER, Supported By: National Institutes of Health SVEN HAUFE, JENS JORDAN, STEFAN ENGELI, ANDREAS L. BIRKENFELD, Jena, Germany, Berlin, Germany, Dresden, Germany, Munich, Germany, Potsdam, Germany, & 1791-P Hannover, Germany ATF6 Directly Increases Peroxisome Proliferator-activated Recep- Angiopoietin-like protein 8 (ANGPTL8), known as betatrophin is involved tor Alpha Activity to Reduce Hepatic Steatosis in Mice in lipid homeostasis and perhaps glucose metabolism. However, associa- XUQING CHEN, FEIFEI ZHANG, QI GONG, AOYUAN CUI, SHU ZHUO, ZHIMIN HU, tions of ANGPTL8/betatrophin with liver steatosis have not been assessed YAMEI HAN, JING GAO, YIXUAN SUN, ZHENGSHUAI LIU, ZHONGNAN YANG, in human subjects. We studied 57 overweight to obsese subjects in a pro- YINGYING LE, XIANFU GAO, LILY Q. DONG, XIN GAO, YU LI, Shanghai, China, San spective, randomized dietary weight reduction intervention trial. A cross- Antonio, TX sectional group included 18 subjects with type 2 diabetes (T2D) or liver Aberrant fatty acid metabolism plays important roles in the development steatosis. In the intervention group, oral glucose testing and liver magnetic of hepatic steatosis. Although ATF6α knockout mice developed tunicamy- resonance scanning at baseline (n = 57) and after 6 months (n = 52) were per- cin-induced hepatic steatosis, the role of ATF6 defi ciency in the liver in the formed. Liver and adipose tissue were sampled in the cross-sectional study. regulation of hepatic lipid metabolism in response to dietary overloading is Samples were analysed for ANGPTL8/betatrophin by enzyme-linked immu- unknown. Here, we show that adenovirus-mediated overexpression of domi- nosorbent assay or by real-time PCR and microarray analysis. In the interven- nant negative form of ATF6 (dnATF6) repressed expression of PPARα’s down- tion study, baseline circulating ANGPTL8/betatrophin correlated with age (r stream targets such as CPT-1α and MCAD, leading to exacerbated hepatic = 0.32, P = 0.010) and triglycerides (r = 0.30, P = 0.02) and was increased with steatosis and insulin resistance in the liver of HFHS diet-fed mice. The effi - hepatic steatosis (P = 0.033). Weight loss reduced liver fat by 45% and signif- ciency of dnATF6 was evidenced by reduction of UPR mediators such as icantly decreased plasma ANGPTL8/betatrophin by 11% (288 ± 17 vs. 258 ± GRP78 and CHOP. Strikingly, dnATF6 was suffi cient to abrogate PPARα ago- 17 pg/ml; P = 0.015). In the cross-sectional study, circulating ANGPTL8/beta- nist WY14643-stimulated expression of fatty acid oxidation genes in HepG2 trophin correlated with liver fat (r = 0.60, P = 0.017), age (r = 0.48, P = 0.044) cells, resulting in repressed the mitochondrial oxygen consumption rate. We and hepatic ANGPTL8/betatrophin mRNA (r = 0.49, P = 0.037). ANGPTL8/ showed that the N-terminal active form of ATF6 (nATF6) induced the tran- betatrophin mRNA was associated with liver fat and insulin resistance as scriptional activity of ERSEI and ERESII luciferase reporters in HepG2 cells. assessed by HOMA-IR (P < 0.05, resp.). T2D in addition to steatosis had no In contrast, nATF6 was suffi cient to stimulate expression of genes involv- effect on circulating and hepatic ANGPTL8/betatrophin (P > 0.05). We con- ing fatty acid oxidation in HepG2 cells. Mechanically, we showed that GST- clude that hepatic steatosis is associated with an increased expression and tagged nATF6 physically interacted with endogenous PPARα. Interestingly, circulating concentration of ANGPTL8/betatrophin. We speculate that this hepatic inhibition of ATF6 abrogated the binding of endogenous PPARα on association also links ANGPTL8/betatrophin to insulin resistance. the promoter of its target gene FGF-21, suggesting that ATF6 is required for Supported By: German Diabetes Federation; German Federal Ministry of Educa- the transcriptional activity of PPARα. Moreover, administration of PPARα tion and Research (BMBF-0313868); German Obesity Network of Competence; agonist WY14643 partially rescued hepatic steatosis in adenovirus-medi- ADIPOSETARGET (01Gl0830) ated dnATF6 overexpressed mice fed on HFHS diet. Therefore, our results indicate strategies to activate ATF6 could be used as an alternative avenue 1794-P to improve liver function and treat hepatic steatosis in obesity. Hepatic Mechanisms Contributing to Early Improvement in Glyce- Supported By: National Natural Science Foundation of China (81270930, mic Control following Bariatric Surgery 31471129) WILLIAM C. ROELL, LUCA MEOLI, SIMONE GUPTA, JIANNONG DAI, BOMIE HAN, NICHOLAS STYLOPOULOS, MELISSA K. THOMAS, Indianapolis, IN, Boston, MA & 1792-P Improvements in glucose and lipid metabolism from bariatric surgery are Regulation of Glucose Homeostasis by Integrin-Linked Kinase in attributed to many mechanisms, including weight loss. In humans, resolution Hepatocytes of hyperglycemia is observed earlier than maximal weight loss, associated ELIJAH TREFTS, DAVID H. WASSERMAN, ASHLEY S. WILLIAMS, Nashville, TN, with reduced fasting glucose and insulin levels indicative of improved insulin

Durham, NC sensitivity. To identify weight-loss independent mechanisms that enhance Obesity

In the face of chronic nutrient excess obesity arises with several comor- glucose metabolism, we studied early changes in metabolic and cellular POSTERS bidities. Amongst obesity-linked pathologies, nonalcoholic fatty liver disease signaling in an insulin resistant mouse model of bariatric surgery. Diet-

(NAFLD) and nonalcoholic steatohepatitis (NASH) directly affect the mor- induced obese mice underwent RYGB or sham surgery followed by postop- Integrated Physiology/ phology and functional homeostasis of the liver. These disturbances include erative recovery and resumption of normal feeding behavior. We identifi ed differential cellular signaling responses to endocrine and paracrine stimuli an early window post-RYGB when fasting blood glucose levels were sig- as well as expansion of the extracellular matrix (ECM). Integrin receptors nifi cantly reduced without signifi cant changes in body weight. To identify and their immediate intracellular machinery convert sensory inputs from the associated RYGB-dependent, weight loss-independent changes in hepatic ECM to biochemical processes within the cell. We show that mice fed a high signaling, we conducted liver microarray analyses. We observed substantial fat (HF) diet (60% kcal from fat) for 16 wks have altered expression of sev- modulation of hepatic mRNA expression of insulin signaling, carbohydrate, eral integrin signaling components, most notably a decrease in hepatocyte and lipid metabolism pathways in RYGB- as compared to sham-operated integrin-linked kinase (ILK). ILK acts as a scaffold, linking the intracellular tail mice, despite the absence of weight loss. mRNA expression levels of insulin of integrin receptors to various signaling components. We demonstrate that signaling regulators were signifi cantly changed, including IRS1, AKT1, and 6 wk old chow-fed mice lacking ILK in hepatocytes (LILKO mice) have a ~33% FOX01. Differential expression of key enzymatic regulators of gluconeogen-

ADA-Supported Research & Moderated Poster Discussion For author disclosure information, see page A696.

A463 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

esis, including PEPCK and G6PC, was accompanied by robust transcriptional 1797-P regulatory signatures, including upregulation of HNF4α, PGC1α, and associ- Extrahepatic Effect in Irs1/Irs2 Liver Knockout Mice ated targets. Transcriptional modulation of lipid metabolism was prominent, OLIVER STOHR, Boston, MA including downregulation of PPARα and downstream targets. Our fi ndings LDKO (hepatic Irs1/Irs2 double knockout) have a diabetic phenotype owing suggest that substantial changes in hepatic signaling likely contribute to at least in part to uncontrolled gluconeogenesis. However, other tissues, early improvements in glycemic control and metabolic function that precede including muscles and various adipose tissues, might contribute to the meta- weight loss following bariatric surgery. By modeling bariatric surgery in bolic dysregulation. LDKO mice showed a strong phenotype in the brown adi- mice, we can elucidate mechanisms of improved glycemic function distinct pose tissue (BAT). BAT of LDKO mice was less brown and enlarged compared from and complementary to benefi ts of reductions in body weight. to BAT of wildtype controls. FGF-21 plays an important role in the develop- Supported By: Eli Lilly and Company ment of BAT. FGF-21 serum levels in LDKO mice are signifi cantly decreased compared to wildtypes. On a molecular level, common BAT genetic mark- 1795-P ers showed less expression compared against control mice. In a [14C] 2-DOG KMO Overexpression Contributes to Glucose Overproduction in glucose uptake assay LDKO mice displayed less glucose uptake into BAT in Type 2 Diabetes both basal and insulin stimulated states. An injection of FGF-21 adenovirus VALÉRIE AUTIER, SOPHIE RAYNAL, ANNICK AUDET, CHRISTINE CHARON, improved insulin sensitive and glucose tolerance in LDKO mice, consistent FRANCK LEPIFRE, MICHELINE KERGOAT, Chilly Mazarin, France with improved glucose uptake by BAT. BAT also plays a crucial role in ther- Kynurenine pathway (KP), the main route of Tryptophan metabolism, is mogenesis. Wildtype mice adapted with a moderate drop in body core tem- overactivated in chronic infl ammatory diseases including type 2 diabetes. perature when exposed to a 4°C environment, whereas LDKO mice failed to This activation leads to increased activity of KP key enzymes (Indoleamine maintain their body core temperature in the cold. Wildtype animals showed 2, 3-Dioxygenase (IDO)-Kynurenine Mono Oxygenase (KMO) and higher tis- a strong induction of genes involved in thermogenesis when exposed to 4°C sue concentrations of several intermediate metabolites deleterious for cell whereas LDKO mice induced thermogenic genes to a far lesser degree. After function and survival. an FGF-21 AV injection the expression levels of BAT markers in LDKO were Glucose overproduction is a major defect of human type 2 diabetes. Under nearly equal to the expression levels of these markers in wildtype mice, and diabetic conditions in several animal models, we observed that KMO protein strongly elevated compared against GFP AV injected LDKO mice. In a 4°C was upregulated in liver. Human HepG2 cells overexpressing KMO exhibited environment FGF-21 injected LDKO mice sustained a normal body core tem- a 30 to 64% increase in glucose production. Similar results were obtained perature, whereas GFP AV injected LDKO mice failed to sustain their core with KMO overexpressing hepatocytes isolated from overnight fasted rats. temperature. In summary, dysregulated metabolism of the FGF-21-defi cient Conversely, siRNA KMO transfected HepG2 cells or rat hepatocytes induced LDKO mice can be treated by genetic or medical replacement of FGF-21 to a 25-30% decrease in glucose production induced by lactate/pyruvate. improve BAT function, thermoregulation and glucose homeostasis. The decreased glucose production induced by KMO knockdown was also found using the KMO inhibitor PNU167532. PNU167532 (1 and 10µM) signifi - 1798-P cantly decreased glucose production from overnight fasted rat hepatocytes The Role of Insulin Receptor Substrate in Hepatocellular Carcinoma by 30 to 60% respectively. Moreover, PNU167532 (25 to 200mg/kg) signifi - YOSHITAKA SAKURAI, ISEKI TAKAMOTO, NAOTO KUBOTA, TAKASHI KAD- cantly improved fasting glycemia (17 to 30%) in type 2 diabetic nSTZ rats. OWAKI, Tokyo, Japan These results demonstrated for the fi rst time the involvement of Kynure- Epidemiologic studies have shown that people with diabetes are at nine pathway in the regulation of hepatic glucose production and the benefi - increased risk for various cancers, especially hepatocellular carcinoma (HCC). cial effect of KMO inhibition on glucose overproduction and glycemic control However, the role of insulin signaling during hepatocarcinogenesis has yet to in type 2 diabetic rats. KMO inhibition is a promising therapeutic strategy be elucidated. Insulin receptor substrate1 (IRS1) and 2 (IRS2) mainly convey for type 2 diabetes. the insulin signaling in the liver. In our preceding study, liver specifi c IRS1 KO mice (LIRS1KO) and liver specifi c IRS2 KO mice (LIRS2KO) fed standard chow 1796-P showed normal TG content levels in the liver, which are otherwise confound- Empaglifl ozin (an SGLT2 Inhibitor), Alone or in Combination with ing factors in hepatocarcinogenesis. In this study, we investigated the role Linagliptin (a DPP-4 Inhibitor), Prevents Steatohepatitis in a Novel of IRS in HCC by induction of chemical carcinogenesis. 15-day-old LIRS1KO Mouse Model of Nonalcoholic Steatohepatitis and Diabetes and LIRS2KO mice were injected diethylnitrosamine (DEN) at a dose of 25 TERUO JOJIMA, KAZUNORI YANAGI, TOSHIE IIJIMA, KUNIHIRO SUZUKI, YOSHI- mg/kg and maintained on standard chow. Ten months after DEN adminis- MASA ASO, Mibu, Japan, Mibu, Kazakhstan tration, the development of HCC and histological and biochemical analyses Sodium-glucose co-transporter-2 (SGLT2) inhibitors are new oral antidi- were performed. IRS1 and IRS2 fl ox/fl ox mice administered DEN were used abetic drugs that reduce hyperglycemia by promoting urinary glucose excre- as controls respectively. First of all, we observed upregulation of Akt phos- tion. Glycosuria produced by SGLT2 inhibitors is associated with weight loss, phorylation in HCC. Insulin receptor and IRS1 protein were also upregulated mainly due to reduced fat volume. We investigated the effects of empagli- in HCC although IRS2 showed only slight increase in HCC. LIRS1KO showed fl ozin (an SGLT2 inhibitor) and linagliptin (DPP-4 inhibitor) on steatohepatitis decreased DEN induced HCC development. The number and size of tumors, and fi brosis in a mouse model of nonalcoholic steatohepatitis (NASH) with serum ALT levels were much smaller in LIRS1KO than in controls. LIRS1KO diabetes. A novel NASH model was generated by administration of strepto- was associated with reduced cancer cell proliferation. Akt phosphorylation zotocin to C57BL/6J mice at 2 days old, with a high-fat diet from 4 weeks. levels were signifi cantly low in tumor lesions of LIRS1KO. Gene expression NASH mice aged 6 weeks were divided into four groups of 6 animals: vehicle, analysis revealed tumors in LIRS1KO showed decreased infl ammation, inva- linagliptin (10 mg/kg), empaglifl ozin (10 mg/kg), and linagliptin +empaglifl ozin. sion, Warburg effect compared with tumors in controls. On the other hands, The histological nonalcoholic fatty liver disease activity score was signifi - LIRS2KO showed similar HCC development compared with controls. These cantly lower in the empaglifl ozin and linagliptin+empaglifl ozin groups than in data suggest that IRS1, not IRS2, plays a crucial role in hepatocarcinogen- the vehicle or linagliptin groups. Hepatic expression of infl ammatory genes esis through activation of insulin signaling independent of fatty liver. (tumor necrosis factor-α, interleukin-6, and monocyte chemoattractant

Obesity protein-1) was decreased in the empaglifl ozin and linagliptin+empaglifl ozin 1799-P

POSTERS groups compared with the vehicle group. Immunohistochemistry showed Osteocalcin Improves Nonalcoholic Fatty Liver Disease in Mice that expression of α-smooth muscle actin, a marker of myofi broblasts through Activation of Nrf2 and Inhibition of JNK

Integrated Physiology/ (fi brosis), was reduced in the linagliptin+empaglifl ozin group compared JING DU, MINGLIANG ZHANG, YUQIAN BAO, Shanghai, China with the vehicle group, as was expression of type 1 and 3 collagen mRNA. Recent studies have demonstrated a protective effect of osteocalcin Linagliptin+empaglifl ozin decreased expression of mRNAs for genes related against nonalcoholic fatty liver disease (NAFLD), although the specifi c to fatty acid synthase, but did not increase mRNAs for β-oxidation-related underlying mechanisms remain unclear. The present study aimed to discuss genes. In conclusion, while empaglifl ozin alone attenuates development the molecular mechanism underlying osteocalcin regulation of NAFLD. Male of NASH showing anti-steatotic and anti-infl ammatory effects, combined C57/BL6J mice were fed a high-fat diet for 12 weeks to induce NAFLD and administration of empaglifl ozin and linagliptin synergistically ameliorates were treated with osteocalcin (30 ng/g) or vehicle by daily intraperitoneal NASH with stronger anti-fi brotic and anti-infl ammatory effects. injection during this period. Osteocalcin treatment protected mice from diet-induced hepatic triglyceride accumulation (p<0.001) and liver injury. Increased levels of malondialdehyde and 8-iso-prostaglandin F2α in the liver of mice fed a high-fat diet were decreased by osteocalcin (p<0.01).

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A464 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

Osteocalcin treatment not only activated nuclear factor-E2-related factor-2 hepatic CB1R has been shown to promote both processes. However, lipid (Nrf2) nuclear translocation and up-regulated the expression of antioxidant accumulation in liver can be experimentally dissociated from insulin resis- enzyme genes (catalase, superoxide dismutase, and glutathione peroxidase) tance under certain conditions, suggesting that other mechanisms are also (p<0.05), but also inhibited the activation of c-Jun N-terminal kinase (JNK) involved. Obesity is also associated with proinfl ammatory changes which, in in the liver (p<0.05). G protein coupled receptor family C, group 6, subtype A turn, can promote insulin resistance. Kupffer cells (KCs), the liver’s resident (GPRC6A), the putative receptor of osteocalcin, was found in the liver. These macrophages, are the major source of proinfl ammatory cytokines in the liver, results suggest that osteocalcin improves NAFLD directly by activating the such as TNF-α, which has been shown to inhibit insulin signaling in multiple Nrf2 pathway to alleviate oxidative stress, and inhibiting JNK to mediate cell types, including hepatocytes. Using intravenously administered β-D- obesity and insulin resistance. glucan-encapsulated siRNA to silence gene expression selectively in KCs Figure. in vivo, we demonstrate that an 80% knock-down of the expression of Cnr1, the gene encoding CB1R, results in improved glucose tolerance and insulin sensitivity in diet-induced obese mice, without affecting hepatic lipid con- tent. Moreover, Cnr1 knock-down in KCs was associated with a shift from pro-infl ammatory M1 to anti-infl ammatory M2 cytokine profi le and improved insulin signaling as refl ected by increased insulin-induced Akt phosphoryla- tion. These fi ndings suggest that CB1R expressed in KCs play a critical role in hepatic insulin resistance via a pro-infl ammatory mechanism. Supported By: National Institute on Alcohol Abuse and Alcoholism

1802-P Glucose Lowering and Vascular Protective Mechanisms of Dop- amine Agonist Therapy in Patients with Type 2 Diabetes CHRISTINA AGYIN, MARIAM ALATRACH, JOHN ADAMS, VINUTHA GANAPA- THY, SUGANYA CHANDRAMOHAN, CHRISTINA CAGA-ANAN, PAULA PHILIPS, CURTIS TRIPLITT, RALPH A. DEFRONZO, EUGENIO CERSOSIMO, San Antonio, TX, Galveston, TX

Bromocriptine, a D2-dopamine agonist [DA] improves glycemic control and has benefi cial CV effects in patients with type 2 diabetes mellitus [T2DM]. To assess its glucose lowering and vascular mechanisms of action, we studied 6 T2DM (3M/3F, age 52±4 y, duration ~6 y, BMI 33±5 kg/m2) Supported By: National Natural Science Foundation of China (31571212) before and after 4 months of DA (3.2 mg/day). All T2DM were treated with GLP-1RA+Metformin (none were on Pioglitazone). At baseline and study 1800-P end, HbA1c and Fasting plasma glucose [FPG] were measured and subjects received: i) 6-hour double tracer (i.v. [3H-glu] and oral [14C-glu)] meal tolerance test to quantitate endogenous glucose production [EGP], glucose disappear- WITHDRAWN ance [Rd] and oral [RaO] glucose appearance rates; and mean prandial plasma glucose [PPG] concentrations; ii) systolic, diastolic and mean ambulatory pressure (SBP, DBP and MAP) and pulse pressure (PP) for 3 days [Spacelabs]; and iii) reactive hyperemia index (RHI) and Arterial Stiffness (AS); [EndoPat 2000]. FPG remained unchanged (157±4 vs.156±8 mg/dl) after 4 months of DA, while HbA1c declined from 8.3±0.5 to 7.4±0.4% and PPG from 215±5 to 192±4 mg/dl (p<0.05); EGP did not change (~2.1 vs. ~2.3 mg/kg.min) following DA, % suppression of EGP dropped slightly (46 vs. 40%) and RaO decreased from 55.4±3.8 vs. 45.8±5.1 mg/kg.min (p<0.01). Pulse rate (76±2 vs. 71±3 bpm) declined (p<0.05), despite reductions in MAP (90±1 to 87±1), SBP (120±2 to 16±1); DBP (74±1 to 72±) and PP (47±2 to 44±1 mmHg). Mean RHI (1.57±0.11 to 2.00±0.18 au) improved and AS (19±3 to 14±3 au) decreased (both p<0.05). Conclusion: Adjunct therapy with bromocriptine DA improves vascular indices and glycemic control in T2DM by reducing oral glucose appearance, thus decreasing post-prandial hyperglycemia. Assuming DA has no effect on gastric emptying, our results suggest that bromocriptine enhances hepatic glucose uptake. Improved vascular indices may partly explain the reduction in CV events observed with Cycloset therapy in T2DM patients. Supported By: VeroScience

1803-P FGF-21-Regulated Hepatic Autophagy Activity Contributes to the Protective Effects of SIRT1 on Liver Steatosis and Insulin Resis- tance in the Diet-induced Obese Mice XIANLIANG RUI, TING LUO, ALLISON NOCON, ALEX SHERBAN, MENGWEI ZANG,

Boston, MA Obesity

Fibroblast growth factor-21 (FGF-21) is a liver-derived endocrine hormone POSTERS with pleiotropic metabolic functions on obesity. Our recent studies indicate

that FGF-21 acts as a critical downstream regulator of the NAD-dependent Integrated Physiology/ deacetylase SIRT1 that protects against liver steatosis. However, the mechanism underlying the metabolic actions of FGF-21 remains unknown. 1801-P Here we show that FGF-21 promotes hepatic autophagy activity, maintains Cannabinoid-1 Receptor (CB1R) in Kupffer Cells Regulate Hepatic hepatic homeostasis, and enhances insulin sensitivity in diet-induced obese Insulin Sensitivity in Obese Mice mice. Liver-specifi c SIRT1 knockout mice (SIRT1 LKO), and their wild-type lit- TONY JOURDAN, SARAH M. NICOLORO, RESAT CINAR, ZHOU ZHOU, JIE LIU, termates (WT) were placed on a high fat, high sucrose (HFHS) diet, followed MINGJIANG XU, MYRIAM AOUADI, MICHAEL P. CZECH, GEORGE KUNOS, Rock- by tail vein injection with adenovirus-mediated gene transfer of FGF-21 or ville, MD, Worcester, MA, Huddinge, Sweden control GFP. Our results showed that autophagy activity, as refl ected by LC3- Obesity-induced accumulation of ectopic fat in the liver is thought to con- II accumulation and p62 degradation, was impaired in livers of HFHS-fed WT tribute to the development of insulin resistance, and increased activity of mice. Autophagy defects were exacerbated in SIRT1 LKO mice with more

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A465 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

severe hepatic steatosis and these metabolic dysfunction was effectively liver fat content (R=-0.615, P=0.0332) but not with whole-body insulin sen- reversed by adenoviral delivery of FGF-21 in these mice. Mice with hepatic sitivity. These results show that MRS allows measurement of insulin-stim- overexpression of FGF-21 showed improved glucose and insulin tolerance ulated changes in adipose tissue fat composition, likely due to synthesis of responses and reduced body fat mass and plasma TG levels. Consistent with saturated fatty acids by DNL. These data suggest that impaired DNL in DSAT these fi ndings, quantitative PCR revealed dramatically increased hepatic in type 2 diabetes humans could trigger NAFLD. expression of genes related to fatty acid β-oxidation (Cpt1α and MCAD) Supported By: German Federal Ministry of Health; German Ministry of Innova- and ketogenesis (HMGCS2), and decreased expression of lipogenic genes tion, Science and Research; German Federal Ministry of Education and Research; (SREBP-1c, ACC1, and FAS). Taking together, FGF-21-induced autophagy may German Center for Diabetes Research represent a novel mechanism for benefi cial effects of hepatic SIRT1 on met- abolic disorders. 1806-P Supported By: American Diabetes Association (1-15-BS-216 to M.Z.); National Hepatic, but Not Extra-Hepatic, Insulin Clearance Is Much Lower Institutes of Health in African American Women Compared with European American Women 1804-P DAVID C. POLIDORI, BARBARA A. GOWER, RICHARD N. BERGMAN, San Diego, Polyol Accumulation in Skeletal Muscle and Liver in a Mouse Model CA, Birmingham, AL, Los Angeles, CA of Type 2 Diabetes Is Reversed upon CL-316,243 Treatment African Americans (AA) tend to have higher serum insulin (SI) than Euro- EMILY J. GALLAGHER, JOSEPH SHILOACH, MARILYN STASINOPOULOS, ZARA pean Americans (EA); this been attributed to both increased secretion and ZELENKO, DEREK LEROITH, New York, NY, Bethesda, MD decreased clearance. Whether lower insulin clearance in AA is due to Type 2 diabetes is a complex metabolic disease that leads to complica- lower hepatic and/or extra-hepatic clearance has not been established. A tions in multiple organs. Diabetic myopathy and liver disease are common recently-developed mathematical model utilizing insulin and C-peptide mea- complications of type 2 diabetes, but are incompletely understood. To gain surements during an insulin-modifi ed FSIGT to estimate hepatic and extra- insight into the pathogenesis of these conditions, we aimed to identify hepatic insulin clearance was applied to data from 42 women (mean age = 25 changes in metabolites that occur in the setting of type 2 diabetes using a yr, BW = 72 kg, fasting glucose = 90 mg/dL, insulin = 65 pM). During the fi rst mouse model of type 2 diabetes with skeletal muscle and liver dysfunction 20 min (when all SI came from endogenous secretion), SI was approximately (the MKR mouse). We performed a metabolomic study of the skeletal muscle twice as high in AA vs. EA, whereas SI was more similar in AA and EA fol- and liver in the adult MKR mouse model of type 2 diabetes, compared with lowing the IV insulin infusion (occurring from 20-25 min) (Figure). Hepatic wild type (WT) FVB/n mice. MKR mice were treated with the β-3 adrenergic insulin clearance was markedly lower in AA vs. EA: fi rst-pass hepatic extrac- receptor agonist, CL-316,243 by i.p injection (1mg/kg/day) or vehicle (PBS) for tion in overnight fasted state was 43 (4)% in EA vs. 16 (4)% in AA, p<0.001; 3 weeks to determine metabolite changes after correcting hyperglycemia. similar differences were seen throughout the FSIGT. In sharp contrast, extra- Tissue metabolomics were performed by Metabolon (Durham, NC) by GC/ hepatic insulin clearance was not lower in AA (1.14 (0.20) L/min for AA vs. MS and LC/MS. Statistical analysis was performed using Welch’s two sam- 0.87 (0.11) for EA). In the overnight fasted state, the percentage of whole- ple t-tests, n=5 per group. At the end of the study, blood glucose was higher body insulin clearance occurring in liver was 59 (5)% in EA vs. 29 (6)% in AA, in MKR mice (311.4±37.8mg/dL) compared with WT mice (147.1±8.3mg/ p<0.001. Hepatic, but not extra-hepatic insulin clearance differences can dL), and normalized with CL-316,243 treatment (117.4±4.3mg/dL). The MKR help explain hyperinsulinemic response to glucose in African Americans. mice had 2.5 fold greater concentrations of sorbitol in the skeletal muscle Figure. (p=0.01), 1.7 fold lower concentrations of reduced glutathione (p=0.01), and 7.6 fold lower NAD+ (p=0.002). In liver, MKR mice had 1.95 fold higher con- centrations of the pentitol ribitol (p=0.04). CL-316,243 treatment normalized sorbitol and ribitol concentrations in the MKR muscle and liver, respectively, returning levels to those of the WT mice. These results demonstrate tissue specifi c accumulation of polyols in a mouse model of type 2 diabetes. Our fi ndings provide novel insights into the potential pathogenesis of skeletal muscle myopathy and liver disease in type 2 diabetes and identify ribitol as a potential biomarker of hepatic disease in type 2 diabetes. Supported By: American Diabetes Association (1-13-BS-108 to D.L.); National Institute of Diabetes and Digestive and Kidney Diseases (to D.L.); National Cancer Institute (to D.L., E.J.G.)

1805-P Impaired Insulin-stimulated Fatty Acid Synthesis in Subcutaneous Adipose Tissue Associates with Liver Fat Content in Type 2 Diabetes JESPER LUNDBOM, MARIA APOSTOLOPOULOU, MARTIN RÖHLING, ALES SAN- DRA BIERWAGEN, JULIA SZENDROEDI, MICHAEL RODEN, Düsseldorf, Germany Nonalcoholic fatty liver disease (NAFLD) associates with insulin resis- tance and type 2 diabetes (T2D). Approximately 25% of hepatic triglycerides are derived from de-novo lipogenesis (DNL), while adipose tissue-derived Supported By: National Institutes of Health free fatty acids (FFA) contribute 60%. Insulin suppresses FFA release and stimulates FA synthesis and DNL in adipose tissue, but not in insulin resis- 1807-P tant states. Thus, adipose tissue DNL is considered benefi cial, while hepatic DNL may contribute to NAFLD. We have developed a non-invasive method

Obesity of measuring double bonds and saturated chain length of FA using non-inva- WITHDRAWN POSTERS sive magnetic resonance spectroscopy (MRS) that may refl ect adipose tis- sue DNL which predominantly has saturated FA as end products. This study

Integrated Physiology/ aimed at determining adipose tissue FA composition and liver fat content by non-invasive MRS in 12 patients with type 2 diabetes before and after hyperinsulinemic-euglycemic clamp tests. Spectra were collected from the deep subcutaneous adipose tissue (DSAT) at the height of the umbilicus and from the liver. DSAT spectra were analysed for total unsaturation (=CH/CH2) and saturated chain length (CH2/CH3), while liver spectra were analysed for total fat content. DSAT unsaturation did not change during the clamp (13.7%±0.5% vs. 13.7%±0.5%), while the saturated chain length slightly increased (5.50±0.06 vs. 5.58±0.07, P<0.01). No change was observed in liver fat content (15.2%±2.2% vs. 15.2%±2.2%). The change in saturated chain length (delta CH2/CH3) during the clamp correlated inversely with baseline

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A466 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1808-P 1810-P Kidney-Liver Cross Talk: Bilateral Surgical Denervation of the Exenatide Enhances the Expression of Heat Shock Protein 70 and Kidneys Normalizes Hepatic Glucose Production in Obese Insulin Attenuates Lipid-induced Hepatic ER Stress via Upregulation of Resistant Dogs by Reducing the Expression of Liver Gluconeogenic SIRT1 Genes XIAOBIN ZHENG, FEN XU, HUA LIANG, JIANPING WENG, Guangzhou, China MALINI S. IYER, RICHARD N. BERGMAN, JEREMY E. KORMAN, ORISON O. Recent studies have indicated endoplasmic reticulum (ER) stress to be a WOOLCOTT, MORVARID KABIR, CATHRYN KOLKA, Los Angeles, CA major contributor to the progression of hepatic steatosis. Exenatide (Exen- Obesity-induced insulin resistance has been widely associated with acti- din-4), a glucagon-like peptide-1 receptor agonist, is known to improve vation of the sympathetic nervous system (SNS). Inhibiting SNS by renal hepatic steatosis by increasing silent mating type information regulator 2 denervation (RDN) has been suggested to improve glucose metabolism. homolog 1 (SIRT1). In this study, we investigated whether exenatide attenu- However, attempts to denervate the kidney using radiofrequency (RF) have ated lipid-induced hepatic ER stress via SIRT1 and illustrated the detailed yielded inconsistent results. To determine the unequivocal effect of RDN on molecular mechanisms. Male C57BL/6J mice were fed with a chow diet or insulin sensitivity (SI) and to study the mechanism involved we performed a high-fat diet (HFD) for 12 weeks, and then the mice in the HFD group were direct surgical (as opposed to RF) bilateral RDN in obese normotensive insu- further treated with or without exenatide for 4 weeks by intraperitoneal lin resistant dogs. injection. After 4 weeks of exenatide treatment, the body weight and sys- Insulin resistance (measured by euglycemic clamp) was induced by feeding temic insulin resistance were signifi cantly reduced when compared with the -4 -1 -1 -1 dogs high-fat diet for 6 wks (w6-HFD) (SI (x 10 dl kg min pM ): 11.6±0.9 at HFD only group. Exenatide notably decreased liver weight and ameliorated w0 vs. 7.9±2.8 at w6-HFD, P<0.01). RDN was performed (n=8) by stripping all the hepatic steatosis induced by HFD challenge. HFD feeding induced ER visible nerves along the renal artery and painting the artery with 10% phenol stress marker proteins including GRP78 and phosphorylated forms of PERK solution. RDN was confi rmed by measuring noradrenaline in the renal cortex and eIF2a. Interestingly, these increases were signifi cantly inhibited by (P<0.001). For Sham surgery (SS; n=7) nerves were exposed but not severed. exenatide treatment. Consistently, in human HepG2 cells, exendin-4 also sig- Renal denervation per se completely restored the impaired ability of insulin nifi cantly attenuated the ER stress markers and lipid accumulation induced to suppress endogenous glucose production in the fat-fed dogs (EGP) (SI EGP (x by thapsigargin (TG) or palmitate (PA). Mechanistic analysis by western blot- 10-4 dl kg-1 min-1 pM-1): 0.5±0.3 at w6-HFD vs. 1.8±0.3 at HFD+RDN, P<0.001, ting showed exenatide enhanced the expression of SIRT1 and heat shock 1.6±0.3 at w0). The mechanism of restoration was revealed by measuring protein 70 (HSP70) compared with the HFD group. Importantly, inhibition expression of gluconeogenic (GNG) genes in liver; RDN reduced the expres- of SIRT1 by genetically whole-body heterozygous knock-out and lentivirus- sion of G6Pase (P<0.05), PEPCK (P<0.001) and Fox01 (P<0.05) by at least 3 mediated shRNA, greatly blocked the effect of exenatide (exendin-4) on the fold in the liver but not in the renal cortex (P=ns), SS did not restore SI EGP in expression of HSP70 and lipid-induced ER stress both in vivo and in vitro. In fat-fed dogs (w6-HFD vs. 1.0±0.3 at HFD+SS, P=ns); nor did it restore liver conclusion, our results indicate exenatide enhances the expression of HSP70 GNG enzymes (data not shown). in a SIRT1-dependent manner, which in turn may reduce the lipid-induced ER We for the fi rst time demonstrate that direct bilateral RDN restores fat- stress and hepatic steatosis. These fi ndings further support exenatide as a impaired liver insulin resistance by reducing the expression of GNG genes in promising therapeutics for hepatic steatosis. the liver. Our data demonstrates crosstalk between renal SNS and hepatic glucose regulation. Crosstalk pathway (s) remain to be elucidated. 1811-P Supported By: National Institutes of Health E2F1 Transcription Factor Controls Hepatic Gluconeogenesis and Contributes to the Development of Type 2 Diabetes and Insulin 1809-P Resistance Cross Talk between ChREBP and SHP in Hepatic Glucose and Lipid DENECHAUD PIERRE-DAMIEN, ALBERT GIRALT, ISABEL C. LOPEZ-MEJIA, EMILIE Metabolism BLANCHET, CAROLINE BONNER, FRANÇOIS PATTOU, JEAN-SEBASTIEN ANNI- KATSUMI IIZUKA, HIROYUKI NIWA, WUDELEHU WU, HIROMI TSUCHIDA, TAKE- COTTE, LLUIS FAJAS, Lausanne, Switzerland, Montpellier, France, Lille, France HIRO KATO, YUKIO HORIKAWA, JUN TAKEDA, Gifu, Japan E2F1 transcription factor regulates the expression of genes required for Carbohydrate response element binding protein (ChREBP) and small het- cell cycle progression through G1 into S phase. Signifi cant and specifi c energy erodimer partner (SHP) play important roles in hepatic de novo lipogenesis needs are necessary to support cell cycle checkpoints and we believe that and cholesterol metabolism, respectively. ChREBP and SHP gene deletion cell cycle regulators, like E2F1, could also control metabolic gene expres- in ob/ob mice improved fatty liver. However, whether ChREBP and SHP sion. We think that this metabolic role of cell cycle regulators should not interact remains unclear. Here, we clarifi ed the crosstalk between SHP and only be restricted to a proliferative context. In fact, we recently established ChREBP in glucose and lipid metabolism. First, a mammalian two-hybrid sys- E2F1 as a key metabolic transcription factor controlling liver lipogenesis and tem revealed that ChREBP and SHP interact. Next, we established Chrebp -/- thus contributing to the development of NAFLD (nonalcoholic fatty liver dis- Shp -/- double knockout (DKO) mice. Compared with WT mice, body weights ease). Here, we reveal a new role of E2F1 in liver physiology through the were larger in DKO mice. Increased liver and decreased visceral fat weights regulation of glucose production. Loss of E2F1 in murine liver results in a were observed in Chrebp -/- and DKO mice. Fasted plasma glucose, insulin, reduction of gluconeogenic gene expression, such as pyruvate carboxylase and triglyceride levels were comparable between the groups. In contrast, (PCX), PCK1 and G6Pase, contributing to lower blood glucose. Inversely, E2F1 fasted plasma cholesterol levels were signifi cantly lower in Shp -/-, Chrebp -/-, overexpression in murine hepatocytes contributes to increase glucose pro- and DKO mice. Plasma β-hydroxybutyrate and fi broblast growth factor-21 duction via gluconeogenic program induction. Mechanistically, the upstream (FGF-21) levels were lower in Chrebp -/- and DKO mice. Interestingly, plasma cyclin dependent kinase CDK4 requires E2F1 to control this pathway. We FGF-21 levels were signifi cantly higher in Shp -/- mice. Hepatic Shp mRNA also show that E2F1 binds directly PCX but not PCK1 and G6Pase promoter. levels were higher in Chrebp -/- mice; however, hepatic Chrebp mRNA levels These results suggest different levels of regulation, in part via FOXO1, that were unchanged in Shp -/- mice. Farnesoid X receptor and liver type pyru- E2F1 uses to modulate glucagon action. Importantly, we determine that E2F1 vate kinase mRNA levels were decreased in Chrebp -/- and DKO mice. CyP7a1, and PCK1 gene expression are positively correlated in liver biopsies from Fatty acid synthase, and peroxisome proliferator-activated receptor mRNA lean and obese patients. Finally, in the physiopathological context of insulin -/- -/-

levels were decreased in Shp , Chrebp , and DKO mice. Consistent with resistance, in which CDK4-E2F1 pathway is over-activated, we demonstrate Obesity -/- plasma FGF-21 levels, FGF-21 mRNA levels were lower in Chrebp and DKO that, contrary to E2F1 reduction, E2F1 overexpression in liver contributes to POSTERS mice and higher in Shp -/- mice. Liver receptor homolog-1 mRNA levels were hyperglycaemia and insulin resistance in mouse models of type 2 diabetes. -/-

decreased in Shp and DKO mice. Therefore, SHP and ChREBP appear to These results provide partly an answer to the “selective” insulin resistant Integrated Physiology/ regulate glucose and lipid metabolism independently; however, SHP and liver paradox (high gluconeogenesis and lipogenesis) and highlight potential ChREBP could interact in vitro. therapeutic targets for the treatment of metabolic diseases. Supported By: Japan Society for the Promotion of Science

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A467 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1812-P 1814-P Diminished Impact of Glucagon (G) to Regulate Hepatic Glucose Reduced Circulating Levels of IGFBP2 Associate with Liver Steato- Production (HGP) in Zucker Diabetic Fatty Rats (ZDF) sis and Insulin Resistance in Patients with Type 2 Diabetes SHANEA K. ESTES, TRACY P. O’BRIEN, ERIN C. JENKINS, RICHARD L. PRINTZ, PIA FAHLBUSCH, MARLIES BEKAERT, FREDERIQUE VAN DE VELDE, YVES VAN MASAKAZU SHIOTA, Nashville, TN NIEUWENHOVE, MARLEEN PRAET, WEENA J.Y. CHEN, MICHAELA DIAMANT, G excess with diabetes is thought to promote elevated HGP resulting in ADRIAAN A. LAMMERTSMA, BRUNO LAPAUW, D. MARGRIET OUWENS, Düs- hyperglycemia, yet with liver steatosis, known to occur with obese type 2 seldorf, Germany, Ghent, Belgium, Amsterdam, Netherlands diabetes (O-T2DM), it has been reported that G desensitization occurs in Accumulating evidence implicates insulin-like growth factor binding pro- liver. We investigated the impact of basal G on the determination of HGP teins (IGFBP) in the development of type 2 diabetes (T2D) and nonalcoholic and plasma glucose levels (PG) in 10 week-old ZDF, a model of the early fatty liver disease (NAFLD). This study analyzed the role of IGFBP2 in T2D and stage O-T2DM. Endogenous glucose production (EGP) was measured using NAFLD. Circulating levels of IGFBP2 were determined by ELISA in a cohort [3-3H] glucose in chronically catheterized, 6 h fasted conscious ZDF and lean of patients with uncomplicated T2D (n=65) and age- and BMI-matched con- littermates (ZCL). During a 4 h test period (TP), after 1 h control period (CP), trols (n=13) that underwent an euglycemic-hyperinsulinemic clamp to assess somatostatin was infused at 5 µg·kg-1·min-1 and basal insulin (I) clamps were insulin sensitivity, and proton MRS to quantitate hepatic triglyceride (TG) performed with intraportal infusion (PI) of I at 1 and 7 mU·kg-1·min-1 for ZCL content. Circulating levels of IGFBP2 were lower in men with uncomplicated and ZDF, respectively, without G infusion (LG) or with basal G clamp (BG) T2D (152 ng/ml) vs. healthy controls (234 ng/ml; p<0.01), and associated with PI of G at 1.6 and 1.8 ng·kg-1·min-1 for ZCL and ZDF, respectively. Dur- positively with insulin sensitivity (M-value) (r=0.509), adiponectin (r=0.305), ing the second half of TP, basal glycemic or hyperglycemic clamp (B-PGC or HDL-cholesterol (r=0.423) (all p<0.05). Negative associations were observed H-PGC) was performed. ZDF during CP exhibited mild hyperglycemia (mM: with fasting glucose (r =-0.352), fasting insulin (r=-0.581), TG (r=-0.385) and 9.4±0.9;ZDF vs. 6.1±0.3;ZCL), markedly higher basal I (ng/ml: 6.5±0.7;ZDF hepatic TG content (r=-0.523) (all p<0.05). All associations remained signifi - vs. 0.9±0.1;ZCL), slightly higher basal G (pg/ml: 36±3;ZDF vs. 30±2;ZCL) and cant when adjusting for age, BMI and adiponectin. The association between elevated EGP (µmol·kg-1·min-1: 54±3;ZDF vs. 44±3;ZCL). During LG, G levels fell IGFBP2 and NAFLD was validated in a cohort consisting of morbidly obese by ~90% without (3±2;ZCL and 4±2;ZDF) compared to those with (36±3;ZDF men (n=79), that had liver biopsies collected during surgery. Here, circulat- and 29±2;ZCL) IPI of G. In ZCL, EGP (27±3;LG vs. 46±3;BG) and PG (2.9±0.2;LG ing levels of IGFBP2 were lower in morbidly obese men with biopsy-proven vs. 6.0±0.3;BG) decreased by 40% and by 50%, respectively; but in ZDF, NAFLD or NASH (117 ng/ml) vs. men without liver disease (242 ng/ml; only by 10% and 12% for EGP (50±4;LG vs. 55±4;BG) and PG (8.3 ± 0.6;LG p<0.001). There were no changes in circulating IGFBP2 among patients with vs. 9.4±1.1;BG), respectively. A B-PGC (6.0±0.2;ZCL and 9.0±0.4;ZDF) reduced varying degrees of lobular infl ammation, ballooning or fi brosis. However, EGP (18±1.4;LG vs. 45±2.8;BG) by 60% in ZCL, but only by 20% (43±4;LG vs. IGFBP2 levels were signifi cantly lower in patients with >66% steatosis (71 53±4;BG) in ZDF. Whereas, a H-PGC at PG levels of 12.4 mM completely ng/ml) vs. morbidly obese men with <5% steatosis (199 ng/ml; p<0.01). In suppressed EGP (-7± 5;LG vs. 23±3;BG) in ZCL, it was reduced by only 28% conclusion, these fi ndings suggest a protective role for IGFBP2 in the devel- (31±6;LG vs. 45±5;BG) in ZDF. In conclusion, for determination of HGP and opment of insulin resistance and NAFLD during the pathophysiology of T2D. glycemia, basal G plays a major role in normal rats, but, unexpectedly, has Supported By: German Center for Diabetes Research much lesser impact in rat model of O-T2DM. Supported By: National Institutes of Health (DK060667) 1815-P Hepatic Denervation Improves Glucose Metabolism in a Canine 1813-P Model of Insulin Resistance Impact of Liver Transplantation on Glucose Homeostasis in Cir- GUILLAUME KRAFT, MELANIE SCOTT, ERIC ALLEN, DALE S. EDGERTON, PHILLIP rhotic Patients E. WILLIAMS, SCOTT B. VAFAI, BOBAK R. AZAMIAN, ALAN D. CHERRINGTON, VALERIA GRANCINI, MARIA ELENA LUNATI, EVA PALMIERI, VERONICA RESI, Nashville, TN, Eden Prairie, MN MASSIMILIANA SMIRAGLIA, GIUSEPPE PUGLIESE, EMANUELA ORSI, Milan, Chronic consumption of a high fat high fructose diet (HFFD) induces Italy, Rome, Italy hepatic insulin resistance, abnormal glucose tolerance, and impaired β-cell Abnormalities of glucose regulation are common in cirrhotic patients and function in the dog. Since enhancement of hepatic glucose uptake (HGU) may be affected by liver transplantation (LT). This study aimed at assessing by portal glucose delivery appears to depend in part on the withdrawal the impact of LT on glucose homeostasis in 89 cirrhotic patients who under- of hepatic sympathetic input, we hypothesized that hepatic denervation went an anthropometric and metabolic evaluation, including an OGTT, prior would improve HGU in this model. Dogs were fed a HFFD for 4 weeks and and 3, 6 and 12 months after LT. Before LT, 18 patients had normal glucose an OGTT was performed. The following week surgical hepatic denervation tolerance (NGT), 30 had impaired glucose tolerance (IGT), and 41 had diabe- (HFFD+HDN, n=11; reducing hepatic norepinephrine >98%) or SHAM sur- tes mellitus (DM). HOMA-IR increased from NGT to DM, which was associ- gery (HFFD+SHAM, n=9) was performed. After 3 more weeks of HFFD a 2nd ated with lower total and LDL cholesterol, albumin, pseudocholinesterase OGTT was performed. Data were compared to OGTTs in untreated chow- (CHE) and platelet count than IGT and NGT. After LT, 24 patients recovered fed dogs (CTR, n=8). The abnormal glucose excursion induced by the diet from DM (regressors, R), 17 did not (non-regressors, NR), 3 developed DM, was improved in HDN but not SHAM dogs. Values for CTR, HFFD (wk 4), and 45 remained nondiabetic. After LT, DM prevalence gradually decreased HFFD+SHAM (wk 7), HFFD+HDN (wk 7) were 3215±352, 7581±688, 7136±1241 from 46.0% to 34.8% at 3 months, 33.7% at 6 months and 22.4% at 1 year, and 5104±654 mg·120 min-1·dL-1 respectively (P<0.05 for HFFD+HDN vs. HFFD when 35 patients had NGT and 34 had IGT. At 1 year, patients who had and for HFFD+HDN vs. HFFD+SHAM). This improvement appears to relate DM at baseline showed a signifi cant improvement in insulin, cholesterol, to both a β-cell and liver effect. The HDN group showed partial restoration triglycerides, total proteins, albumin, bilirubin, AST, ALT, CHE, and platelet of β-cell function (Δ AUC Insulin/Δ AUC Glucose = 0.506±.06, 0.308±.05, count, whereas HOMA-IR improved signifi cantly only in R subjects (2.36±1.8 0.310±.07 and 0.452±.07 in CTR, HFFD, HFFD+SHAM and HFFD+HDN respec- vs. 4.22±3.0, P<0.05), though all these parameters remained signifi cantly tively with P<0.05 for CTR vs. HFFD and HFFD+SHAM and P=0.099 for HFFD worse than in IGT and NGT subjects. At multiple regression analysis, inde- vs. HFFD+HDN). A subset of dogs (n=6 HFFD-HDN and n=3 HFFD-SHAM) pendent predictors on NR vs. R were male gender (β=0.7269; P=0.015) and underwent a hyperglycemic hyperinsulinemic clamp a week after OGTT#2.

Obesity baseline HbA1c (β=0.727; P= 0.015) and γ-GT (β=0.0038; P=0.05) levels and Net hepatic glucose balances were -1.5 ±.1 (net uptake), +0.2 ±.1 (net pro- -1 -1 POSTERS inversely CHE (β=-0.0003; P=0.008) and platelet count (β=-0.0089; P=0.05). duction) -0.5 ±.1 (net uptake) mg·kg ·min respectively in CTR, HFFD+SHAM In conclusion, LT is associated with regression of DM in ~50% of patients, and HFFD+HDN (P<0.05 for HFFD+SHAM vs. HFFD+HDN) indicating a partial

Integrated Physiology/ with failure to regress being predicted by male sex and baseline HbA1c and restoration of normal insulin stimulated hepatic glucose uptake in response liver function. to HDN. In summary, HDN improved glucose tolerance in an insulin resistant canine model by improving β-cell function and the liver’s response to glu- cose. In conclusion, HDN has the potential to enhance post-prandial glucose clearance in insulin resistant states.

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A468 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1816-P overnight cortisol concentrations. However the contribution of this physi- Quantifying Divergent Effects of ANGPTL3 Antisense Oligonucle- ologic rise in nocturnal cortisol to endogenous glucose production (EGP) and otide on Hepatic Lipid and Glucose Metabolism in Fructose-Fed the extent to which this is altered in T2D has not been investigated. The cur- Rats rent study was designed to compare EGP in T2D and controls in the presence DANIEL F. VATNER, GABRIELA V. MOREIRA, ABUDUKADIER ABULIZI, MARK GRA- vs. absence of a nocturnal rise in cortisol. 9 C (5M, age 57 ± 7 yr, HbA1c 36 ± HAM, GERALD I. SHULMAN, VARMAN T. SAMUEL, New Haven, CT, Carlsbad, CA 2 mmol/mol) and 9 T2DM (5M, age 60.6 ± 9.6 yr, HbA1c 57 ± 12 mmol/mol) The endogenous circulating lipoprotein lipase inhibitor Angiopoietin-Like matched for BMI were studied on two occasions in random order. Subjects 3 (ANGPTL3) is a promising drug target for hypertriglyceridemia, particu- were admitted to the Clinical Research Trials Unit at Mayo Clinic for two larly for mixed dyslipidemia. Knockdown of ANGPTL3 has been studied in overnight visits lasting 48 hours. Subjects with sleep apnea were excluded genetic models of metabolic disease, but less is known about the basic biol- from participation. Endogenous cortisol was blocked with metyrapone 500 ogy of ANGPTL3 in models of diet-induced obesity. We used a liver-targeted mg given orally every 4 hours from 1800 to 0600 starting on the second day GalNAc-modifi ed antisense oligonucleotide (ASO) to assess the effects of of each visit. At 2200, hydrocortisone infusion was started at 0.15µg/kg/min ANGPTL3 knockdown on hepatic lipid metabolism in male Sprague Dawley and continued until 0700 on one occasion; on the other occasion infusion rats. In high fat fed rats, ANGPTL3 ASO decreased fasting plasma triglyc- rates were adjusted (0.3µg/kg/min from 2200 to 2400, 0.6 µg/kg/min from eride (TG) by 38% (P=0.01). The impact of ANGPTL3 ASO on plasma lipid 2400 to 0400, 1.0 µg/kg/min from 0400 to 0700) to mimic the physiological concentrations was greater in high fructose fed rats, with a 75% reduction nocturnal rise in cortisol. [3-3H] glucose was infused starting at 2200 until in plasma TG (P<0.01), and a 42% reduction in plasma fatty acid concentra- 0700. Blood was sampled from an arterial catheter. Fasting glucose, gluca- tion (P<0.01). Body weight, epidydimal adipose weight, and skeletal muscle gon and insulin were not different between visits in either group. Cortisol TG content were unchanged in both diet groups. Though hepatic TG was concentrations by design (18.3 ± 4.0 vs. 7.7 ± 1.5; 16.4 ± 2.7 vs. 7.1 ± 1.3µg/dl) not altered in fat fed rats, hepatic TG content was surprisingly increased by and rates of EGP at 0700 were slightly higher (11.6 ± 2.6 vs. 12.9 ± 1.7; 17.0 ± 2.7-fold (P < 0.01) in fructose-fed rats and associated with a 64% increase in 4.6 vs. 18.5 ± 4.7 µmol/kgFFM/min) in the presence than absence of a noctur- hepatic de novo lipogenesis (DNL, P < 0.01), and a 23% decrease in fasting nal rise in cortisol in both C and T2D respectively. We conclude that a physi- hepatic VLDL secretion (P < 0.001). There was no change in the expression of ological nocturnal rise in cortisol at least in part contributes to the increased key enzymes of hepatic lipid metabolism, including fasn, scd-1, and mttp. In fasting EGP observed in T2D. Further studies are required to test whether spite of the increased hepatic steatosis, glucose metabolism did not appear cortisol-induced increase in EGP is clinically relevant in T2D during nocturnal to be affected. Plasma glucose and insulin concentrations after a fast and rise in cortisol. oral glucose tolerance test were not different. FGF-21 has been identifi ed as Supported By: National Center for Advancing Translational Science (R01 a potent regulator of hepatic lipid metabolism and ANGPTL3 ASO decreased DK29953, UL1 TR000135) hepatic FGF-21 expression by 57% (P < 0.04). Conclusion: While ANGPTL3 knockdown consistently decreases plasma 1819-P TG concentration, it can also worsen hepatic steatosis by potentiating Human Kallistatin Improves Diet-induced Insulin Resistance in hepatic DNL from dietary fructose associated with a decrease in FGF-21, Mice though without signifi cantly increasing lipid-induced dysglycemia. JULIA REINKE, SEBASTIAN BRACHS, DIANA M. WILLMES, JOSEPH TIO, Supported By: National Institutes of Health JOACHIM SPRANGER, JEFFREY D. MCBRIDE, JIAN-XING MA, STEFAN R. BORN- STEIN, GERALD I. SHULMAN, ANDREAS L. BIRKENFELD, Berlin, Germany, Dres- 1817-P den, Germany, Oklahoma City, OK, New Haven, CT Exenatide Improves Hepatic Insulin Resistance Kallistatin (KST), also known as SERPIN A4, is a circulating, broadly act- AMALIA GASTALDELLI, MELANIA GAGGINI, GIUSEPPE DANIELE, DEMETRIO CIO- ing protein with anti-angiogenic and anti-infl ammatory properties. Clinical CIARO, EUGENIO CERSOSIMO, DEVJIT TRIPATHY, CURTIS TRIPLITT, PETER FOX, studies in patients with diabetes revealed increased levels of KST in poor NICOLAS MUSI, PATRICIA IOZZO, RALPH A. DEFRONZO, San Antonio, TX, Pisa, glycemic control and microvascular complications. Whether or not KST has a Italy direct effect on glucose homeostasis in the setting of insulin resistance and The effect of glucagon like peptide 1 receptor agonists (GLP-1RA) on the T2D is currently unknown. liver is still debated. Recent studies suggest that GLP-1 acts on the liver to To address this, transgenic mice overexpressing human KST systemi- improve hepatic glucose/lipid metabolism and to reduce insulin resistance cally (hKST-TG) and littermate-control wildtype mice (WT) were studied (IR) and hepatic fat. However, it is diffi cult to discern where these effects under chow (ND) and high fat diet (HFD) conditions. Plasma hKST levels are secondary to weight loss or GLP-1 per se. To examine this, we studied were 7.2±0.83 µg/ml on ND and 2.8±0.5 µg/ml on HFD in the hKST-TG mice, 15 subjects (12 IGT and 3 newly diagnosed T2DM, age=56±8 y, BMI=29±1kg/ respectively. As expected, hKST was not detectable in the WT group. Body m2, HbA1c=5.7±0.1%) on two occasions. Subjects received subcutaneous weights were similar between groups up to an age of 24 weeks on both exenatide (EX, 5 mcg) or placebo (PLC) in double blind fashion, 30 min before diets. IPGTT yielded similar glucose and insulin excursion curves in ND ani- a 4 hour oral glucose tolerance test (75g). During OGTT we measured hepatic mals. In the weight matched HFD cohort, an IPGTT revealed an improve- glucose uptake (HGU) by 18FDG-PET; hepatic glucose production (HGP), oral ment in glucose homoestasis in hKST-TG mice. Additionally, the HOMA-IR glucose absorption (RaO) and glucose clearance (Cglu) by simultaneous dou- was lower in hKST-TG on HFD (2.2±0.27, hKST-TG vs. 4.42±0.87, WT, p<0.05), ble tracer. Hepatic IR was calculated as HGP x Mean Plasma-Insulin (I) conc indicating improved insulin sensitivity in hKST-TG mice. To better understand the tissue specifi c contribution to the protective effect of hKST, hyperinsu- during OGTT. RaO was reduced by EX compared to PLC (mean RaO0-240min 8.0±1.0 vs. 12.7±0.6 µmol/min kg). This was explained, in part, by improved linemic euglycemic clamps with labelled glucose were performed. Glucose hepatic uptake of oral glucose by EX (HGU/RaO: 3.9±1.5 vs. 0.38±0.07 [nmol/ infusion rates were higher in hKST-TG mice (31.5±3.7 mg/kg/min, hKST- min ml]. [ µmol/min kg]-1, p=0.03). Despite lower plasma insulin levels with TG vs. 18.1±3.5 mg/kg/min, WT, p<0.05), validating the insulin sensitizing EX vs. PLC during the OGTT (mean-I=23±5 vs. 41±5 mU/l, p<0.02), EX mod- effect of hKST in HFD fed mice. Specifi cally, hKST overexpression protected estly decreased postprandial mean HGP (6.1±0.6 vs. 5.1±0.6 µmol/min.kg, against HFD induced hepatic (clamp hepatic glucose output: 7.7±1.9 mg/kg/ p=ns), signifi cantly reduced Hep-IR (130±37 vs. 197±28, p<0.02) and sig- min, hKST-TG vs. 12.1±0.8 mg/kg/min, WT, p=0.05) but not peripheral insulin -1 resistance. nifi cantly augmented Cglu (0.16±0.04 vs. 0.05±0.01 [µmol/min kg]. [mU/l] Obesity

p<0.05). Glucagon levels were similar in the two studies. These data show that human KST protects against diet induced insulin POSTERS Conclusion: EX acutely improves Hep-IR enhancing HGU and decreasing resistance in mice. We speculate that increased KST levels in the setting of poor glycemic control and microvascular complications are rather protective

EGP. These results demonstrate that GLP-1 exerts its effect on hepatic glu- Integrated Physiology/ cose metabolism independent of weight loss. than harmful. Further studies are needed to uncover the underlying molecu- Supported By: Amylin; Bristol-Myers Squibb; AstraZeneca lar mechanisms.

1818-P 1820-P Effects of Physiological Rise in Nocturnal Cortisol on Endogenous Impact of Loss of Hepatocyte PPARγ on Mechanisms Mediating Glucose Production in Type 2 Diabetes Liver Steatosis in Adult Mice RITA BASU, SONA VEETTIL, LING HINSHAW, BRENT MCCONAHEY, NISHA ABIGAIL WOLF GREENSTEIN, NEENA MAJUMDAR, PENG YANG, PAPASANI V. JOSHI, ROBERT A. RIZZA, ANANDA BASU, Rochester, MN SUBBAIAH, RHONDA D. KINEMAN, JOSE CORDOBA-CHACON, Chicago, IL Studies have shown that nocturnal rise in glucose concentrations Hepatic triacylglycerols (TAG) accumulation “steatosis,” is the leading observed in individuals with type 2 diabetes (T2D) correlate with higher cause of chronic liver disease and it is associated with insulin resistance

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A469 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

and diabetes. Reduced hepatic fatty acid (FA) oxidation and VLDL release respectively, which were also highly correlated with the multiple metabolic OR increased esterifi cation of FA [from diet, adipocyte-lipolysis or de novo disorders such as hyperglycemia, hypertryglycemia and obesity. HR have lipogenesis (DNL)] results in steatosis. In order to develop new strategies positively correlated with FIB4 index, Aspartate aminotransferase to Plate- to prevent steatosis, we need to understand how these mechanisms are let Ratio Index (APRI), and BARD score. The postmortem histopathological regulated. Peroxisome proliferator-activated receptor gamma (PPARγ) is a analysis also revealed that some of the obese and dysmetabolic NHPs had nuclear receptor expressed in multiple tissues, and its hepatic overexpres- liver morphological changes assemble to human NAFLD/NASH. Thus, this sion is associated with steatosis. Although indirect evidence suggests that computer-assisted noninvasive method for analysis of hepatic echogenicity PPARγ increases DNL and FA esterifi cation, the precise PPARγ-regulated can be used as a powerful tool in discovery and development of novel thera- mechanism that promotes liver steatosis has not been fully elucidated. We pies for NAFLD/NASH in both clinical and preclinical research. sought to determine how loss of hepatocyte PPARγ impacts hepatic lipid Figure. The Correlation between H/R Ratio and FIB4 Index, BARD Index and metabolism in a mouse model of adult-onset, hepatocyte (liver)-specifi c APRI Index. knockdown of PPARγ (aLivPPARγkd) as compared to their respective lit- termate controls under various metabolic conditions: fed-fast-refeeding of a chow diet and diet-induced obesity. aLivPPARγkd blunted high-fat diet induced steatosis, but did not impact VLDL release, plasma TAG, or the expression of lipolytic and FA oxidation genes. Expression of DNL genes and DNL or SCD-indexes (16:0/18:2 or 16:1/16:0, respectively, indicative of DNL rate) were not reduced in aLivPPARγkd mice. However, expression of Cd36, Fabp1 and Mogat1 (genes involved in the hepatic FA uptake, transport and esterifi cation) and the hepatic diacylglycerol (DAG)/monoacylglycerol (MAG) ratio were reduced independent of diet, while plasma levels of NEFA were elevated in obese aLivPPARγkd mice. These data suggest that PPARγ may promote TAG accumulation by enhancing FA esterifi cation via MAG path- way [esterifi cation of MAG to DAG by the action of MOGAT (monoacylg- lycerol acyltransferase)], independent of hepatic DNL, FA oxidation or VLDL release. Supported By: U.S. Department of Veterans Affairs (BX001114, BX001090); 1823-P National Institutes of Health (R01DK088133, R21AT008457, S10OD010660); Chi- Altered Metabolic Reprogramming in Male Offspring in a Genetic cago Biomedical Consortium (PDR-033); Endocrine Society Model of Insulin Resistance DARIO F. DE JESUS, ROHIT N. KULKARNI, Boston, MA Despite the identifi cation of several genes associated with diabetes, only 1821-P a small fraction of its hereditability can be explained. This argues for the Role of the Hepatic Mammalian Indy Homolog (mIndy, Slc13a5) in relevance of epigenetics in the etiology of diabetes. We aimed to deter- Energy Homeostasis mine the effects of parental insulin resistance on metabolic programming DIANA M. WILLMES, ANICA KURZBACH, MARTIN A. DANIELS, STEFANIE LIESKE, in the offspring of the liver-specifi c insulin receptor knockout (LIRKO) mice. STEFAN G. KAUSCHKE, GERD LUIPPOLD, STEFAN R. BORNSTEIN, ANDREAS L. At 3 months of age LIRKOs are insulin resistant and glucose intolerant. Male BIRKENFELD, Dresden, Germany, Berlin, Germany, Biberach, Germany control offspring from father LIRKO (COFL), mother LIRKO (COML), or con- INDY (I’m Not Dead Yet) is a transporter of TCA cycle intermediates, trol mothers and fathers (C) were weaned on chow (21%) or high-fat diet mainly handling cellular citrate uptake. Reduced expression of Indy in lower (HFD) (60%) and followed for 3 months. An independent cohort was followed organisms extends lifespan and reduces whole body fat content. In mam- on chow for 12 months. COFL and COML exhibited impaired growth until mals, whole body deletion of the mammalian Indy homolog (mIndy, Slc13a5) 2 months of age on chow. Total fat mass was 1.4 fold higher in COFL and increases energy expenditure and protectes mice from diet induced obesity COML on chow (p<0.01, n=5, 12 months of age) and HFD (p<0.05, n=4-5, 3 and insulin resistance, while reducing mIndy expression in the liver using anti- months of age) compared to C. Indirect colorimetry revealed impaired meta- sense oligonucleotides improves insulin sensitivity but not body weight. To bolic fl exibility in COFL and COML on HFD (p<0.01, n=4). COFL and COML characterize the role of hepatic mIndy on energy homeostasis, we analyzed exhibited a phenotype of insulin resistance on chow and HFD. With aging conditional liver selective KO mice with and without high fat feeding. Body and on HFD, COFL and COML develop hyperglycemia and hyperinsulinemia weight was similar between knockouts and littermate controls up to an age (p<0.05, n=5) and prominent hepatic steatosis compared to C. COFL and of 20 weeks (HFD 39.6g±2.23, Ctr vs. 41.2g±1.75, KO). Calorimetric analysis COML on HFD revealed increased hepatic expression of fatty acid transport (average 24 h O2 consumption: HFD 5941.95mL/h/kg lean mass ±230.27, Ctr and lipogenesis-associated genes. Furthermore, RNA-sequencing of COFL vs. 5910.82mL/h/kg lean mass±241.41, Ko), food intake and punctual physi- and COML livers on chow revealed down-regulation in enriched-pathways cal activity where unchanged. However, glucose tolerance was improved in associated with insulin and MAPK signaling (p<0.001, n=4), while only COML diet induced heterozygous liver-mIndy KO mice and tended to be improved presented increased regulation in enriched-pathways associated with in homozygous liver-mINDY mice (ipGTT AUC: HFD 65102.7±18448.01, Ctr nonalcoholic fatty liver disease (NASH) (p<0.001, n=4) compared to C. These vs. 48106.56±7358.2, Het (P<0.05 vs. Ctr) vs. 51875.6±11186.45, Ko (P=0.08 data suggest that prenatal insulin resistance have detrimental effects on vs. Ctr)). From these data, we conclude that hepatic deletion of mIndy is metabolic adaptation and transcriptional regulation of hepatic metabolism involved in the regulation of glucose metabolism but is not suffi cient to in the offspring that contribute to impaired growth and metabolic response induce changes related to whole body energy homeostasis. We speculate to dietary challenges. that other organs than the liver also contribute to the effect. 1824-P 1822-P The Effect of One-Year Treatment with the Glucagon-Like Peptide-1 Noninvasive Sonographic Quantifi cation of Hepatic Lipidosis in Receptor Agonist Liraglutide on NAFLD in Women with Prior Gesta- Obesity Nonhuman Primate (NHP) Model of Obese, Dysmetabolism, and tional Diabetes Mellitus POSTERS Diabetes LOUISE VEDTOFTE, EMILIE BAHNE, SIGNE FOGHSGAARD, CAMILLA ANDREA- HAIHUA GU, YONGQIANG LIU, HUI WANG, BINGDI WANG, GUOFENG SUN, SEN, CHARLOTTE STRANDBERG, THORA BUHL, LISELOTTE K. CHRISTIANSEN, Integrated Physiology/ XIAOLI WANG, YONG-FU XIAO, KEEFE CHNG, YAO-PING LIN, YIXIN (JIM) WANG, JENS J. HOLST, JENS A. SVARE, TINE D. CLAUSEN, ELISABETH R. MATHIESEN, Kannapolis, NC, Taicang, China, Taipei, Taiwan PETER DAMM, LISE L. GLUUD, FILIP K. KNOP, TINA VILSBØLL, Copenhagen, Den- Liver biopsy is a gold standard in diagnosis of nonalcoholic fatty liver mark, Herlev, Denmark, Hillerød, Denmark disease (NAFLD)/steatohepatitis (NASH) in the clinic. We have developed Prior gestational diabetes mellitus (GDM) is associated with an increased a noninvasive sonographic method to quantify hepatic lipidosis and other risk of nonalcoholic fatty liver disease (NAFLD). This study evaluates the pathologic changes in correlation with multiple metabolic disorders in a NHP effect of one-year treatment with liraglutide on NAFLD in nondiabetic model of obese, dysmetabolic and diabetes, which has been proved to be the women with prior GDM. The study includes a subgroup of patients from a most translatable animal model for human diseases. Both the hepatic/renal randomised, double-blinded, investigator-initiated, placebo-controlled trial echo-intensity ratio (H/R=1.60 ±0.12 vs. 1.36 ± 0.09, p=0.012) and hepatic examining nondiabetic women with prior GDM for whom we have full data echo-intensity attenuation rate (HA=0.41 ± 0.07 vs. 0.17 ± 0.04, p=0.005) set (n=82, age (mean±standard deviation): 38±5 years, body weight: 88±16 signifi cant increase in the obese (n=13) compared to control (n=23) NHPs, kg, BMI: 32±5 kg/m2) allocated to liraglutide 1.8 mg subcutaneously once-

For author disclosure information, see page A696. & Moderated Poster Discussion ADA-Supported Research

A470 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE daily (n=37) or placebo (n=45). All women underwent abdominal ultrasound activity in vitro and in vivo. We measured MGAT2 catalyzed formation of to diagnose NAFLD at baseline and after one year and a subgroup (n=60) also didecanoyl-glycerol from 1-decanoyl-rac-glycerol and decanoyl-CoA, to underwent transient elastography (FibroScan®) with assessment of intra- produce predominantly 1,3-didanoyl-glycerol. Unlike 1, 2-DAG, 1,3-didanoyl- hepatic fat by controlled attenuation parameter (CAP) both at baseline and glycerol is not susceptible to further acylation to triglyceride (TG). This LC/ after one year (CAP: 266±48 dB/m). None had elevated liver blood tests or MS assay allowed the quantifi cation of MGAT activity in tissue lysates. cirrhosis at baseline. Based on the abdominal ultrasound and elastography, To measure MGAT2-driven cellular activity of TG synthesis, we utilized 18 women had NAFLD at baseline. Eleven women (13%) developed NAFLD 1-oleoyl-glycerol-d5 as a substrate to trace its incorporation into TG in during the one-year follow-up. Eight women had NAFLD at baseline, but not MGAT2 expressing cells. The oleoyl-glycerol-d5 incorporated major TG spe- after one year. These women had lower alanine aminotransferase levels at cies were then quantifi ed by LC/MS. We assessed the ability of MGAT2 the end of follow-up compared to baseline (P=0.042). None of the remain- inhibitors to suppress fat absorption in mice by a meal tolerance test with ing patient characteristics were associated with resolution of NAFLD. Lira- U13C-TG oil. The newly absorbed and re-synthesized plasma heavy TGs con- glutide had no effect on the prevention, development or resolution of NAFLD taining three 13C in the glycerol backbone and two U13C-acyl-chains, which (NS), but signifi cantly reduced intrahepatic fat assessed by CAP compared to represented the newly absorbed and resynthesized TGs, was then quan- placebo (-35±52 vs. -5±52 dB/m, P=0.003). Liraglutide reduced bodyweight titated by LC/MS. We showed the inhibition of metabolic fl ux of 1-oleoyl- (4.7±5.2 vs. 1.4±5.5 kg, P=0.007), but this effect was not associated with CAP glycerol-d5 and U13C-TG in the MG pathway by a MGAT2 inhibitor in cells and values or resolution of NAFLD in multivariable analyses. This one-year trial in vivo, Our results indicated that MGAT2 plays a major role in the intestinal found no effect of liraglutide on presence of NAFLD but a signifi cant weight MG pathway for dietary fat absorption. loss and reduced steatosis as measured by CAP was observed. Supported By: Novo Nordisk A/S 1827-P Use of a New, Inducible Genetic Method Reveals Critical Roles of 1825-P GABA in the Control of Metabolic and Mental Functions Integrated Regulation of Hepatic Lipid and Glucose Metabolism by FANTAO MENG, YONG HAN, DOLLADA SRISAI, WENWEN CHEN, VALERY Fox0 Proteins and Adipose Triacylglycerol Lipase BELAKHOV, SICONG DONG, MONICA FARIAS, YONG XU, RICHARD D. PALMITER, WENWEI ZHANG, SO YOUNG BU, MARA MASHEK, INSUG O-SULLIVAN, TIMOR BAASOV, QI WU, Houston, TX, Iowa City, IA, Haifa, Israel, Seattle, WA ZAKARIA SIBAI, SALMAAN A. KHAN, OLGA ILKAYEVA, CHIRISTOPHER NEW- Currently available inducible Cre/loxP systems, despite their considerable GARD, DOUGLAS G. MASHEK, TERRY G. UNTERMAN, Chicago, IL, Minneapolis, utility in gene manipulation, have pitfalls in certain scenarios, such as unsat- MN, Durham, NC, Saint Paul, MN isfactory recombination rates and deleterious effects on physiology and Fox0 proteins are major targets of insulin action and contribute to the behavior. To overcome these limitations, we designed a new, inducible gene- regulation of glycolytic, gluconeogenic and lipogenic metabolism in the targeting system by introducing an in-frame nonsense mutation into the cod- liver (J Biol Chem 281:10105-17, 2006). Adipose triacylglycerol lipase (ATGL) ing sequence of Cre recombinase (nsCre). Mutant mRNAs transcribed from mediates the fi rst step in triacylglycerol (TAG) hydrolysis and Fox01 stimu- nsCre transgene can be effi ciently translated into full-length, functional Cre lates ATGL expression in adipose tissue. Here, we examined the regulation recombinase in the presence of nonsense suppressors such as aminoglyco- of ATGL and its inhibitor, the G0/G1 switch gene protein 2 (G0S2), by Fox0 sides. In a proof-of-concept model, GABA signaling from hypothalamic neu- proteins, and their role in mediating Fox0 effects on gene expression and rons expressing agouti-related peptide (AgRP) was genetically inactivated metabolism in the liver. Studies in Fox0 transgenic and knockout mice and within 4 days after treatment with a synthetic aminoglycoside. Disruption hepatocytes show Fox0 proteins stimulate ATGL and suppress G0S2 mRNA of GABA synthesis in AgRP neurons in young adults led to a dramatic loss levels and mediate effects of insulin on ATGL and G0S2 expression in the of body weight due to reduced food intake and elevated energy expendi- liver. Studies with adenoviral vectors show Fox0 proteins stimulate TAG ture; they also manifested glucose intolerance. In contrast, older mice with turnover and fatty acid oxidation (FAO) in isolated hepatocytes and in the genetic inactivation of GABA signaling by AgRP neurons had only transient liver in an ATGL-dependent fashion. Studies in transgenic mice expressing reduction of feeding and body weight; their energy expenditure and glucose constitutively active Fox01 in the liver show ATGL also mediates effects of tolerance were unaffected. They also had less obsessive compulsive-like Fox01 on serum TAG levels, liver TAG content and on glycolytic, lipogenic and behaviors. These results indicate that GABAergic signaling from AgRP neu- gluconeogenic gene expression and metabolism in the liver. Adenoviral ATGL rons not only plays a key role in the control of feeding and body weight but knockdown or G0S2 expression, and inhibition of FAO with etomoxir restore also modulates affective disorders that are comorbid with eating disorders. glucose tolerance in Fox01 transgenic mice. Western blotting studies indi- This new genetic technique will augment current tools used to elucidate cate that intracellular signaling and energy sensing pathways may mediate mechanisms underlying many physiological and neurological processes. ATGL-dependent effects of Fox0 proteins on gene expression and metabo- Supported By: American Diabetes Association (7-13-JF-61 to Q.W.) lism in the liver. Together, these results reveal ATGL-dependent lipolysis and FAO play an important role in mediating effects of Fox0 proteins and inte- 1828-P grating the regulation glucose and lipid metabolism in the liver. VMH Glucose Sensing Neurons Are Altered by High Fat Diet Sensi- Supported By: U.S. Department of Veterans Affairs; National Institutes of Health tivity to Facilitate Dysglycemia YANG LI, ANTHONY H. CINCOTTA, Tiverton, RI The ventromedial hypothalamus (VMH) of insulin resistant animals exhib- INTEGRATED PHYSIOLOGY—MACRONUTRIENT its elevated noradrenergic input activity potentiating counter-regulatory METABOLISM AND FOOD INTAKE responses even at euglycemia and diminished insulin sensitizing response to physiological postprandial hyperglycemia. As such, glucose (G) (excitatory 1826-P and inhibitory) sensing neurons within the VMH of insulin resistant (IR) ani- Metabolic Tracing of Monoacylglycerol Acyltransferase-2 Activity mals would be expected to be hyper-responsive to local mild glucopenia and In Vitro and In Vivo unresponsive to local physiological hyperglycemia. We therefore studied the JIANSHEN QI, WENSHENG LANG, MARGERY A. CONNELLY, FUYONG DU, YIN in vivo VMH electrophysiological dose response to exogenous G (1-5 mM

LIANG, JOHN MASUCCI, GARY W. CALDWELL, TONYA MARTIN, GEEHONG KUO, [observed in VMH following feeding)] or deoxyglucose (DG) (1-15 mM) admin- Obesity MICHAEL GAUL, WILLIAM MURRAY, MICHAEL K. HANSEN, JAMES LENHARD, istration to the VMH of rats resistant or sensitive to the fattening effects of POSTERS ALESSANDRO POCAI, KEITH T. DEMAREST, SEUNGHUN LEE, Spring House, PA 1 month of a high (60%) saturated fat diet (HFR or HFS respectively) vs. lean

Monoacylglycerol acyltransferase 2 (MGAT2) is highly expressed in the rats held on regular diet (controls [C)] to test this postulate. VMH neuronal Integrated Physiology/ small intestine where it catalyzes the synthesis of diacylglycerol (DAG) from activity was assessed via current recording from a direct wire electrode at free fatty acids (FFA) and sn-monoacylglycerol (MG). MGAT2 knockout (KO) the VMH amplifi ed and surveyed through an AD Instrument Bio Amp ML136. mice have delayed absorption of FFA and MG from the proximal to distal Fasting plasma G was also determined. IR HFS rats were 26% more obese intestine, leading to enhanced incretin release and improvement in glucose and hyperglycemic than HFR rats (P<0.01) while HFR rats did not differ in metabolism. Diet-induced obesity (DIO) and glucose intolerance are pre- these parameters from C rats. VMH neurons of C rats exhibited a marked vented in both congenital and inducible MGAT2 KO mice. A MGAT2 inhibitor 2.8 fold increase in fi ring in response to 5mM G (P<0.05) that was abolished recapitulates in DIO mice the phenotype observed in MGAT2 KO mice with in HFR and HFS rats (P<0.05). VMH neurons of C rats also exhibited a dose improved glucose tolerance and decreased fat weight. Thus, inhibition of dependent 1.9 fold increase in fi ring rate in response to increasing doses of MGAT2 is a promising strategy to treat and prevent obesity and diabetes. DG between 3 to 15 mM. Responsiveness to low (3-5mM) doses of DG was Here, we report novel LC/MS based metabolic fl ux assays to trace the MGAT2 enhanced only in HFS rats vs. C (by 50%, P< 0.05). These fi ndings suggest

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A471 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE

that HF feeding reduces the activity of VMH glucose excitatory neurons and PYY3-36 separately; GLP-1R blockage alone markedly augmented the con- that in HFS rats, there is also an enhanced sensitivity of VMH glucose sens- centration of active PYY3-36, while sita alone greatly increased active GLP- ing neurons to local glucopenia. In HFS rats, HF feeding induces changes 1, likely counteracting effects on food intake on these days. These results in VMH G sensing (hyper-responsive to local mild glucopenia and reduced show that combined actions of GLP-1 and PYY are important for inhibition of response to local hyperglycemia) that may facilitate increases in fasting and appetite and food intake after RYGB. postprandial plasma G levels. 1831-P 1829-P The Composition of Gastrointestinal Bacterial Flora in HNF1A- The Effect of Hyperglycemia on Brain Activity Is Infl uenced by MODY Seems to Be Different from Type 2 Diabetes Weight Loss SANDRA MROZINSKA, PIOTR RADKOWSKI, TOMASZ GOSIEWSKI, MAGDALENA RENATA BELFORT-DEAGUIAR, DONGJU SEO, MARY SAVOYE, JANICE J. HWANG, SZOPA, MALGORZATA BULANDA, AGNIESZKA H. LUDWIG-GALEZOWSKA, CHRISTIAN SCHMIDT, CHERYL LACADIE, ROBERT T. CONSTABLE, RAJITA SINHA, IWONA MORAWSKA, AGNIESZKA SROKA, BARTłOMIEJ MATEJKO, MACIEJ T. ROBERT S. SHERWIN, New Haven, CT MALECKI, PAWEL WOLKOW, TOMASZ KLUPA, Krakow, Poland Obese, but not lean, subjects exhibit increased activity in reward and There have been many reports on the composition of the gastrointestinal decreased activity in executive-control brain areas in response to food cues (GI) bacterial fl ora in patients with type 2 diabetes (T2DM) at various stages measured by functional MRI (fMRI). However, is not well known if these dif- of disease progression. However, no data exists concerning the profi le of GI ferences can be ameliorated by diet-induced weight loss. Thus, to evaluate bacterial fl ora in individuals with monogenic forms of the disease. The aim the effect of a weight loss diet on brain response to visual food cues during of the investigation was to compare the parameters of the colonic fl ora in hyperglycemia, we studied 8 obese subjects (age 46±6, 7F/1M, BMI 32±2) patients with HNF1A-MODY and T2DM. We collected stool samples from before and after an 8-week low calorie diet. They underwent an fMRI scan 10 patients with HNF1A-MODY and 23 patients with T2DM. The bacterial combined with a 2-step hyperglycemic (200 mg/dL)-euglycemic (100 mg/dL) DNA was isolated from specimens and then 16S RNA sequencing using the clamp study. The participants lost ~4% of their weight. Fasting glucose lev- Ilumina MiSeq platform was performed. The mean age in the T2DM group els decreased from 96±7 to 87±12 mg/dL (P=0.02); while glucose levels were was 58, 13 years (SD± 14, 69 years), in the HNF1A-MODY- 42, 5 years (SD± indistinguishable before and after the diet during the 2-step clamp. Leptin 4, 63 years), p<0.05. The duration of diabetes was 4, 46 years (SD± 2, 95 levels decreased from 50±28 to 33±23 (P<0.01). fMRI scans before and after years) vs. 19, 1 (SD± 13, 37 years), p<0.05, the HbA1c level was 8, 15% vs. 6, the diet are seen below (subtraction maps hyper-euglycemia). Before the 95%, p<0.05, respectively. After preprocessing we received an average of diet, there was a signifi cant increase in brain activity (yellow/red) in the ven- 90304, 89 reads per sample (SD±43368, 92). The differential abundance of tral prefrontal cortex (PFC), insula and striatum during hyper vs. euglycemia operational taxonomic unit (OUT) across sample groups was analyzed using (P<0.05 whole brain corrected-WBC). Post-diet, hyperglycemia decreased DESeq2. At the genus level the Megamonas was 18-fold, Slackia 10, 4-fold, brain activity (blue/purple) in reward areas (insula, and striatum) (P<0.05, Lactobacillus 6, 86-fold, Faecalibacterium 4-fold, other genus of Enterobac- WBC). These results suggest that the brain of obese subjects after a weight teriaceae 3, 7-fold, unnamed genus of Ruminococcaceae 2, 2-fold higher in loss diet may respond more appropriately to food cues in the presence of T2DM group than HNF1A-MODY group. In opposite the Weissella was 7-fold, elevated blood glucose levels. Cellulosimicrobium 5-fold, other genus of Promicromonosporaceae 6-fold, Figure. unnamed genus of Clostridiaceae 8, 6-fold, other genus of other family of Actinomycetales 5, 8-fold lower in T2DM patients vs. MODY individuals (p<0, 05 for each comparison). Multivariate analysis (Wilcoxon test, Mul- tivariate Linear Regression including body mass index, age, HbA1c level) revealed that the relative abundance of two unnamed genus of Clostridi- aceae was independently lower in T2DM group, p=0.00019. In conclusion, it seems that there are differences in gut microbiome composition between patients with HNF1A-MODY and type 2 diabetes. Supported By: Ministry of Science and Higher Education; European Union Supported By: National Institutes of Health 1832-P 1830-P Manipulating the Sequence of Nutrient Ingestion Improves Real- Combined Actions of GLP-1 and PYY Reduce Food Intake after Roux- Life Glycemic Control in Type 2 Diabetes en-Y Gastric Bypass DOMENICO TRICO’, EMANUELE FILICE, SILVIA TRIFIRÒ, ANDREA NATALI, Pisa, MARIA S. SVANE, NILS B. JORGENSEN, KIRSTINE N. BOJSEN-MØLLER, SIGNE Italy NIELSEN, CARSTEN DIRKSEN, VIGGO B. KRISTIANSEN, NICOLAI J. WEWER Lipid and protein ingested before carbohydrate reduce post-prandial ALBRECTSEN, SIGNE TORÄNG, BOLETTE HARTMANN, STEN MADSBAD, JENS hyperglycemia. We tested the feasibility, safety and clinical effi cacy of JUUL HOLST, Hvidovre, Denmark, Copenhagen, Denmark manipulating the sequence of nutrient ingestion in outpatients with type 2 Exaggerated secretion of the appetite inhibiting hormones glucagon-like diabetes (T2D). Seventeen overweight, well-controlled (HbA1c 48-58 mmol/ peptide-1 (GLP-1) and peptide YY (PYY) may explain the substantial weight mol) subjects with T2D were randomized to either a hypocaloric control diet loss after Roux-en-Y gastric bypass (RYGB). GLP-1 actions can be blocked (CD) or to a hypocaloric experimental diet (ED) allowing the consumption of by the GLP-1 receptor (GLP-1R) antagonist Exendin 9-39 (Ex9), while PYY high-carbohydrate foods only after high-protein and high-fat foods at lunch actions can be inhibited by a dipeptidyl-peptidase 4 (DPP-4)-inhibitor block- and dinner (not at breakfast to comply with Italian habits). We collected ing conversion of PYY to active PYY . We hypothesized that both GLP-1 blood glucose self-monitoring data, in addition to relevant clinical and meta- 1-36 3-36 bolic parameters, during the 4-week run-in and the 8-week diet periods. Both and PYY3-36 reduce appetite post-RYGB and investigated effects of Ex9 and DPP-4 inhibition on ad libitum food intake. In a placebo-controlled, random- diets were well accepted, accurately followed (meal compliance ≥ 90%), and ized, design, 12 patients with normal glucose tolerance (age: 35±7 years, neutral on arterial blood pressure, plasma lipids and indices of hepatic and Obesity sex (f/m): 8/4, BMI: 33±6 kg/m2) were studied 5±1 month after RYGB with kidney function. After 8 weeks, in spite of a similar reduction of body weight POSTERS mixed meal tests followed 4 hours later by an ad libitum meal. On separate (ED -1.9 95% CI [-3.4/-0.4] kg, p<0.03; CD -2.0 [-3.6/-0.5] kg, p<0.02) and days, patients received: 1.) placebo, 2.) the DPP-4 inhibitor sitagliptin (sita) waist circumference (ED -2.9 [-4.3/-1.5] cm, p<0.002; CD -3.3 [-5.9/-0.7] cm, Integrated Physiology/ to decrease PYY , 3.) Ex9 to block the GLP-1R or 4.) sita and Ex9 to inhibit p<0.02), the ED only was associated with a statistically signifi cant reduction 3-36 of HbA1c (-0.3 [-0.50/-0.02]%, p<0.04), of fasting plasma glucose (-1.0 [-1.8/- both PYY3-36 and GLP-1 simultaneously. Combined administration of sita/Ex9 increased ad libitum food intake ~20% vs. placebo, whereas no effect was 0.3] mmol/L, p<0.01), and of both postprandial glucose excursions (lunch -1.8 seen with sita or Ex9 alone (Ad libitum food intake: placebo: 245±22; Ex9: [-3.2/-0.4] mmol/L, p<0.01; dinner: -1.0 [-1.9/-0.1] mmol/L, p<0.04) and other 264±30, p=0.38; sita: 252±20, p=0.71; sita/Ex9: 290±36 grams, p=0.04). Ex9 indices of glucose variability measured over 4 days (Standard Deviation: -0.5 [-0.7/-0.2] mmol/L, p<0.02; Coeffi cient of Variation: -6.6 [-10.4/-2.7]%, increased AUCPYY3-36 by 124±36% compared with placebo (p<0.01), whereas a 66±6% decrease was seen during concurrent administration of sita/Ex9 p<0.02). When compared to the CD, the ED was associated with lower post- lunch glucose excursions (p<0.02) and lower 4-days glucose coeffi cients of compared with Ex9 alone (p<0.01). AUCActive GLP-1 increased 154±38% during Ex9 and 226±50% during sita (both p<0.01). In conclusion, combined block- variation (p<0.05) by MANOVA for repeated measures over the 8 weeks. ade of GLP-1 and PYY signaling increased food intake by ~20% after RYGB, Manipulating the sequence of nutrient ingestion might reveal a rapid, fea- whereas there was no effect when blocking the GLP-1R or formation of sible, economic and safe strategy for optimizing glucose control in T2D.

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A472 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE

1833-P intermediates to substrates were positively correlated with age in both lean Postprandial VLDL-TG Storage in Type 2 Diabetes and obese individuals (18 of 30 metabolites with p < 0.05) and was more ESBEN SØNDERGAARD, RAKEL F. JOHANSEN, SØREN NIELSEN, Aarhus, Denmark pronounced after weight loss. Following weight loss, medium chain acyl- Ectopic lipid storage is closely linked to insulin resistance and develop- carnitine levels fell more in obese with high VO2max while BCAAs and their ment of type 2 diabetes. Whether this is a consequence of impaired adipose metabolites fell more in obese with lower VO2max. Multiple linear regression tissue storage capacity is unknown. To determine this, we investigated 11 analysis showed that higher VO2max reduced the age-dependent increase in men with type 2 diabetes and 10 weight-matched healthy men using ex-vivo fatty acid metabolite levels. The effect of VO2max remained when obese labeled VLDL-TG tracers before and after an oral fat tolerance test to deter- were assessed alone and was independent of sex, race, degree of obesity mine postprandial VLDL-TG storage and turnover. As expected, we found and diabetes status. Following diet-induced caloric restriction, there was that men with type 2 diabetes had a greater postprandial lipemia compared a reduction in medium chain acylcarnitines, BCAA and ratios of acylcarni- to weight-matched healthy men (p<0.01). Despite this, postprandial VLDL-TG tine intermediates to substrates. The modulating effect of VO2max on age- storage was similar in the two groups in both upper body (p=0.46) and lower related fatty acid metabolite levels was reduced following weight loss but body (p=0.69) subcutaneous adipose tissue. The postabsorptive VLDL-TG revealed modulation of BCAA-related metabolites. The results suggest that secretion rate was similar in the two groups (p=0.78) and the two groups higher VO2max reduces the rate of metabolic aging and caloric restriction suppressed VLDL-TG secretion rate equally (≈50%) after the oral fat toler- also improves metabolism, consistent with a “younger” metabolic state. ance test (p=0.77). Postprandial VLDL-TG storage increased with increasing Supported By: P30-DK089503 regional depot fat mass in upper body subcutaneous adipose tissue, both in the entire depot (p<0.001) and per 109 fat cells (p=0.04). Fat cell size was 1836-P not a predictor of postprandial VLDL-TG storage. In conclusion, men with Sustained Effects of a Protein and Lipid Preload on Glucose Toler- type 2 diabetes have similar postprandial VLDL-TG storage and suppression ance in Type 2 Diabetes of VLDL-TG secretion compared to weight-matched controls. This argues DOMENICO TRICO’, EMANUELE FILICE, SIMONA BALDI, SILVIA FRASCERRA, against an impaired postprandial adipose tissue storage capacity as a cause ANDREA MARI, ANDREA NATALI, Pisa, Italy, Padova, Italy for ectopic fat storage. Small amounts of nutrients given as a “preload” reduce post-glucose gly- Supported By: Danish Diabetes Academy caemic peaks in type 2 diabetic patients by activating a number of mecha- nisms involved in glucose homeostasis. To ascertain whether the positive 1834-P effect of a nutrient preload on glucose tolerance is sustained throughout the Role of Brown-Fat-released Lipid Metabolites in Cold Adaptation whole post-prandial phase and to investigate the mechanisms involved, we LUIZ O. LEIRIA, MATTHEW D. LYNES, MICHAEL A. KIEBISH, TIM SCHULZ, YU- evaluated total, exogenous and endogenous plasma glucose, insulin secre- HUA TSENG, Boston, MA, Framingham, MA, Potsdam, Germany tion, insulin clearance, GLP-1, GIP and glucagon for 300 min after a 75 g Brown adipose tissue (BAT) is activated under cold condition to generate oral glucose load preceded by the ingestion of either water or a small non- heat. Both short- and long-term cold exposure result in improved glucose glucidic preload (50 g parmesan cheese, one boiled egg). After the nutrient and fatty acid metabolism, thereby producing an insulin-sensitizing effect. preload, we found a signifi cant reduction of peak glucose gradient (-49%, We hypothesize that once cold activated, BAT may release insulin-sensitiz- p<0.02), total plasma glucose (iAUC -28%, p<0.03), exogenous glucose ing agents in the circulation in order to improve peripheral glucose uptake. appearance (iAUC -30%, p<0.03), insulin clearance (-28%, p<0.04), and an Aiming to fi nd such agents, we performed serum lipidomics in mice kept at enhancement of insulin secretion (iAUC +22%, p<0.003) and β-cell glucose thermoneutrality (30ºC) or cold (5ºC) for 2 days and 11 days. Cold exposure sensitivity (+44%, p<0.01). These effects were associated to higher plasma yielded a marked increase in the serum levels of the omega (ω)-3 polyun- levels of GLP-1 (iAUC +463%, p<0.002), GIP (iAUC +152%, p<0.0003) and saturated fatty acids (PUFA) metabolites derivatives from eicosapentaenoic glucagon (iAUC +144%, p<0.0002). A protein and lipid preload signifi cantly acid (EPA), the so-called hydroxyeicosapentaenoic acid (HEPE). Several of improves glucose tolerance throughout the whole post-absorptive phase by the HEPE isomers were increased in cold conditions, especially the 12-HEPE, reducing the appearance of oral glucose and improving both beta-cell func- which were found to be increased approximately 40 times (p<0.001) in the tion and insulin bioavailability in type 2 diabetes. serum of cold-exposed animals. We found no signifi cant changes in the Figure. serum concentration of ω6-PUFA metabolites. As a result, the circulatory ω3/ω6 ratio was signifi cantly increased in cold adaptation. The increase in HEPE levels was absent in a mice model with severe brown fat paucity, indicating that brown fat is the potential source for this group of omega-3 metabolites in circulation. Expression of lipoxygenases (LOX), specially LOX- 12, and cycloxygenase-2 (COX-2), the key enzymes responsible for the con- version of EPA to HEPE, was elevated under cold exposure (p<0.05). In con- clusion, our data reveal a novel cold-induced pathway that coordinates the ω3-PUFA metabolism in BAT, resulting in the release of ω3 but not ω6 lipids in the circulation. Given the known effect of ω3 PUFA on improving insulin sensitivity, the ω3 metabolites likely mediate insulin-sensitizing effects of cold exposure. Supported By: American Diabetes Association (1-16-PDF-063 to L.O.L.); National Institutes of Health

1835-P Effects of Intrinsic Oxidative Capacity (VO2max) and Weight Loss on Aging-related Changes in Plasma Metabolite Levels 1837-P CHANISA THONUSIN, HEIDI B. IGLAY REGER, SHERVIN ASSARI, CHARLES R. TRPV1-dependent Synaptic Regulation of Liver-Related Neurons in Obesity EVANS, AMY E. ROTHBERG, CHARLES F. BURANT, Ann Arbor, MI the Hypothalamus Is Diminished in db/db Mouse POSTERS Individuals with higher VO2max have a decreased risk of metabolic dis- HONG GAO, KAYOKO MIYADA, ANDREA ZSOMBOK, New Orleans, LA eases and a reduced age-related mortality. Rats selected for increased Whole body glucose homeostasis is largely controlled by central mech- Integrated Physiology/ VO2max also show improved metabolic status and longevity and maintain a anisms. Preautonomic neurons in the paraventricular nucleus (PVN) of the higher mitochondrial capacity for fatty acid and branched-chain amino acid hypothalamus play pivotal role in determining the sympathetic and parasym- (BCAA) catabolism as they age. To understand whether these fi ndings trans- pathetic outfl ows, and thus infl uence homeostatic functions. Hormones and late to humans, we assessed VO2max (ml/min/kg fat free mass) and fasting nutrients alter the activity of neurons through direct effect or via modulation plasma metabolite levels by liquid chromatography-mass spectrometry in a of synaptic inputs. Transient receptor potential vanilloid type 1 (TRPV1) has group of lean (n = 28) and obese individuals (n = 124), the latter before and been shown to have benefi cial effects on whole body metabolism. In this after diet-induced weight loss (-17.8 +/- 6.3%, p < 0.001). Subjects consumed study, we identifi ed liver-related PVN neurons with the retrograde trans- an isocaloric, fi xed macronutrient diet for 3 days before testing. VO2max fell neuronal viral tracer, pseudorabies virus 152 and used whole-cell patch- with age, but was higher at any age in lean compared to obese. Acylcarniti- clamp recordings to determine the electrophysiological properties and the nes derived from fatty acid and BCAA catabolism and ratios of acylcarnitine TRPV1-dependent regulation of liver-related PVN neurons in control and

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A473 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE

db/db mice. We tested the hypothesis that TRPV1-dependent excitation of liver-related PVN neurons is diminished in db/db mice. Capsaicin, a potent TRPV1 agonist, increased the frequency of miniature excitatory post-syn- aptic currents (mEPSCs) in lean control mice. In contrast, capsaicin did not alter mEPSC frequency of liver-related PVN neurons in db/db mice. The aver- age basal frequency of mEPSCs and inhibitory PSCs (mIPSCs) was not differ- ent between control and db/db mice; however, we observed a decreasing trend in excitatory neurotransmission to liver-related neurons in db/db mice. These fi ndings are consistent with our previous data demonstrating altered hypothalamic autonomic circuitry in hyperglycemic conditions. Our data suggest that hyperglycemia and/or dysregulated insulin signaling affect TRPV1-dependent neurotransmission, which can contribute to autonomic dysfunction and dysregulation of neural control of glucose metabolism in obese-diabetic conditions. Supported By: National Institutes of Health (R01DK099598)

1838-P Consumption of either High-Fructose Corn Syrup or Sucrose Increases Uric Acid, Postprandial Triglycerides, LDL-cholesterol, and ApoB, in Men and Women CANDICE ALLISTER PRICE, KIMBER STANHOPE, VALENTINA MEDICI, ANDREW A. BREMER, VIVIEN LEE, MARINELLE V. NUNEZ, GUOXIA X. CHEN, NANCY L. KEIM, PETER J. HAVEL, Davis, CA, Sacramento, CA Fructose consumption in the U.S. has increased to >50 g/day per person since the 1970s. Previous studies from our laboratory have demonstrated that the consumption of fructose-sweetened beverages at 25% of daily energy requirements (Ereq) reduces insulin sensitivity, increases hepatic de novo lipogenesis (DNL) and elevates LDL-cholesterol and lipoproteins. These fi ndings were not observed in subjects consuming glucose-sweetened bev- erages, despite identical weight gain in both groups. These results indicate it is not simply the caloric content of dietary sugar, but rather the type of sugar that increases metabolic risk factors. The two most common fructose- 1840-P containing sweeteners include: 1.) high fructose corn syrup (HFCS), com- Evaluation of a Novel Paradigm to Quantify Metabolic Flexibility posed of monosaccharides fructose (55%) and glucose (45%) and 2.) sucrose, with Whole Room Calorimetry composed of equal proportions of fructose and glucose as a disaccharide. KAREN D. CORBIN, CHRISTOPHER BOCK, RICHARD E. PRATLEY, STEVEN R. Controversy remains whether there are differential effects of these two SMITH, Orlando, FL sources of added sugars in our diet. We studied 75 participants matched for Metabolic fl exibility is the ability to adjust substrate oxidation to match age and BMI in 3 sweetened beverage groups: 1.) aspartame, 2.) sucrose or substrate availability and energy needs. Persons with diabetes display a dual 3.) HFCS at 25% of Ereq. After 2 weeks consuming sugar-sweetened bever- defect in fat (overnight fasted) and carbohydrate (CHO; post-meal) oxidation. ages, only subjects consuming HFCS-sweetened beverages exhibited sig- Simultaneous assessment of this dual defect is key to fully understand its nifi cant weight gain compared with aspartame (+0.8 kg, p=0.03). Compared interrelatedness and mechanistic infl uence on outcomes. There are no stan- with aspartame, both sucrose and HFCS increased plasma concentrations dardized protocols for the measurement of metabolic fl exibility. To address of uric acid (+0.31 ± 0.09 mg/dL, p=0.02 and +0.46 ± 0.09 mg/dL, p=0.0005), this gap, we developed a controlled environment paradigm (whole room postprandial TG (54 ± 8 mg/dL, p<0.0001 and 37 ± 7 mg/dL, p=0.003) apo- calorimeter with scripted activities of daily living) coupled with dietary con- lipoprotein B (+6.2 ± 1.8 mg/dL, 0.04 and +10.1 ± 1.7 mg/dL, p=0.0003), and ditions to maximize fuel switching. Based on our earlier studies, we hypoth- plasma LDL (+12.2 ± 2.6 mg/dL, p=0.002 and +13.4 ±2.4 mg/dL, p=0.0004), esized that fat and CHO oxidation defects would co-exist. We used overnight respectively. These results demonstrate that HFCS- or sucrose-sweetened fasting respiratory quotient (RQ) as a measure of fat oxidation and substrate beverages consumed at 25% of Ereq are equally capable of increasing uric switching in response to a high CHO breakfast (increase in RQ; ΔRQ) as an acid and established lipid/lipoprotein risk factors risk for cardiovascular dis- integrated measure of CHO oxidation. Cohort: 36 healthy individuals (73% ease in only 2 weeks in young men and women. female), age 14-73 years, body mass index 17-39. The mean (SD) overnight Supported By: National Institutes of Health/National Heart, Lung, and Blood fasted RQ was 0.915 ± 0.06. Seventeen percent of participants only had a Institute (1R01-HL09133, 1R01-HL107256, R01-HL-075675, R01-HL-121324, 1R01- CHO oxidation defect (defi ned as lowest ΔRQ quartile), 17% only had a fat oxi- HL-121324-02S1); University of California, Davis; National Institutes of Health (K12 dation defect (highest fasting RQ quartile), and 8% displayed a dual defect. A HD051958 to K.S.) subset of 19 individuals were re-tested within 2.3 ± 1.1 days. Despite lack of a lead-in diet, resting post-absorptive RQ was highly reproducible between 1839-P days (CV: 8.5% [1st measurement] and 1.5% [post 24 h in the calorimeter)] as was post-breakfast “peak” RQ (CV 4.0%). However, delta RQ was highly vari- WITHDRAWN able and may need additional pre-calorimeter dietary control to reduce the interday variability. Our results establish an important paradigm to measure dual-substrate metabolic fl exibility and identify unique fuel metabolism pat-

Obesity terns between individuals. This could be of great relevance for developing

POSTERS targeted metabolic therapies for diabetes where there is a paucity of data related to metabolic adaptation to glycemic excursions. Integrated Physiology/ 1841-P Morning Hyperinsulinemia Stimulates Hepatic and Whole-Body Glucose Disposal Later in the Day MARY COURTNEY MOORE, MARTA S. SMITH, BEN FARMER, GUILLAUME KRAFT, PHILLIP WILLIAMS, ALAN D. CHERRINGTON, Nashville, TN Whether breakfast consumption vs. breakfast skipping enhances glucose disposal later in the day is an important question in management of diabe- tes. We previously showed that a 4 h morning (AM) duodenal glucose infusion signifi cantly enhanced net hepatic glucose uptake (NHGU) in conscious dogs during a hyperinsulinemic hyperglycemic clamp later in the day (PM). The cur-

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A474 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE

2 rent study was designed to differentiate the effect of AM hyperinsulinemia vs. and [ H5]glycerol. Liver biopsies were subsequently obtained during bariatric hyperglycemia on PM glucose disposal. We used hepatic balance and tracer surgery. Basal serum FGF-21 correlated positively with body fat (r = 0.39, p = (3H-glucose) techniques to examine 2 groups of chronically catheterized dogs. 0.04), fasting glucose (r = 0.39, p = 0.03), and basal endogenous glucose pro- After an overnight fast, the dogs underwent a pancreatic clamp (0-240 min) duction (EGP; r = 0.44, p = 0.02), and negatively with peripheral insulin sensi- with somatostatin and glucose infused via peripheral vein (IV) and insulin and tivity (r = -0.39, p = 0.05). After fructose ingestion, FGF-21 increased to 300 glucagon delivered into the hepatic portal circulation. During the AM, the condi- ± 28% of basal at 120 min (p < 0.01) and returned to basal levels at 300 min. tions existing were euglycemia with hyperinsulinemia (initially 10xbasal, taper- Fold FGF-21 increase following fructose ingestion correlated negatively with ing to 4xbasal; AM ins [n=6)] or hyperglycemia (1.4xbasal) with basal insulin body fat (r = -0.48, p =0.01), fasting insulin (r = -0.36, p = 0.05) and basal EGP (AM glc [n=5)]. The AM clamp ceased at 240 min, and it was followed by a (r = -0.38, p = 0.05), and positively with peripheral (r = 0.53, p < 0.01) and 3H-glucose equilibration period (240-330 min) and collection of mid-day basal adipose tissue insulin sensitivity (r = 0.55, p < 0.01). Basal and post-fructose samples (330-360 min). Subsequently all dogs underwent a PM clamp (360- serum FGF-21 did not correlate with mRNA expression of de novo lipogenesis 600 min) with infusion of IV somatostatin; intraportal insulin (4xbasal), glucagon genes (CHREBP, SREBP, FAS, ACC) in preliminary liver biopsy analyses (n = 20). (basal), and glucose (4 mg.kg-1.min-1); and IV glucose as needed to maintain arte- High basal serum FGF-21 is associated with increased basal EGP and insulin rial plasma glucose at ≈200 mg/dL. In the PM clamp, mean rates (mg.kg-1.min-1) resistance, whereas a more robust FGF-21 response to fructose ingestion is of IV glucose infusion (13.7±2.5 vs. 8.4±1.8), NHGU (6.9±0.9 vs. 3.5±0.5 mg.kg-1. associated with improved metabolic health in obese subjects. This is consis- min-1), tracer-determined hepatic glucose uptake (5.8±0.9 vs. 3.3±0.3), and tent with the possibility that FGF-21 may mediate an adaptive response to hepatic glycogen synthesis (6.0±0.8 vs. 2.9±0.5) were greater in AM ins vs. AM fructose ingestion. glc (P<0.05 for each). Phosphorylated/total liver glycogen synthase was signifi - cantly lower (P<0.01) in AM ins vs. AM glc. These fi ndings indicate that hyperin- 1844-P sulinemia but not hyperglycemia resulting from breakfast intake can markedly Mice Fed a High-Starch Diet Become Obese but Do Not Develop stimulate hepatic and whole-body glucose disposal later in the same day. Muscle Insulin Resistance Supported By: National Institutes of Health AMANDA E. BRANDON, TUONG-VI NGUYEN, EURWIN SURYANA, LEWIN SMALL, GREGORY COONEY, Sydney, Australia 1842-P Mice fed a high fat diet accumulate fat in adipose tissue (WAT) and non- Fasting and Postprandial Substrate Oxidation Differs in Insulin- adipose tissues and develop glucose intolerance and insulin resistance. A diet Treated Type 2 Diabetes Using Sodium-Glucose Cotransporter 2 high in starch also leads to signifi cant fat deposition but the impact on glucose Inhibitor metabolism is less defi ned. Three groups of mice were housed at thermoneu- KEN KANAZAWA, HIROSHI UCHINO, FUMIKA SHIGIYAMA, FUKUMI FUKUMI, trality (29oC) and fed a chow (20% protein, 11% fat, 69% carbohydrate), high SHUKI USUI, MASAHIKO MIYAGI, HIROSHI YOSHINO, NAOKI KUMASHIRO, starch (Hi-ST; 20% protein, 20% fat, 60% corn starch) or high fat diet (Hi-F; YASUYO ANDO, TAKAHISA HIROSE, Tokyo, Japan 20% protein, 60% lard, 20% corn starch). Body weight, food intake, body com- Hyperglycemia is known to impair energy substrate oxidation both fast- position (DEXA), and glucose tolerance were monitored and after 25-30 weeks ing and postprandial, as a result of glucose toxicity, due to exceeding cellu- mice had surgery to insert carotid and jugular catheters. One week later, mice lar energy source. Insulin treatment along with hyperinsulinemia have dem- underwent a hyperinsulinaemic-euglycaemic clamp (6 mU/kg/min insulin) with onstrated that, excessive extracellular glucose transmit into the cells which radioactive tracer infusion to determine whole body and tissue specifi c glu- already has abundant dense intracellular energy source. In this context, we cose metabolism. Hi-ST and Hi-F diet mice acquired signifi cantly more WAT examined whether reduction of unavailable plasma glucose to the urine with a than chow mice (9.4±0.5g; 11.5±0.7g; 3.4±0.2g respectively). Despite similar sodium-glucose cotransporter 2 (S) inhibitor may have any impact on 1.) fasting fatness, Hi-ST mice had better oral glucose tolerance than Hi-F mice (iAUC and postprandial energy expenditure (EE) and substrate oxidation, 2.) the period 364±36 vs. 657±69; P<0.001) and were similar to lean, chow-fed mice (iAUC and basal-bolus dosage of insulin that achieve euglycemia. Twenty T2D (M/F = 359±20). Clamp glucose infusion rate (GIR) was signifi cantly reduced in Hi-ST 17/3, age 49 ± 11.8, duration 2.4±3.0 years, BMI 27.0 ± 4.8, A1c 12.9 ± 1.3%) and Hi-F compared to chow indicating whole body insulin resistance. However were admitted to this 7 days, randomized, open-label, prospective study to GIR was higher in Hi-ST compared to Hi-F (Chow 30.1±2.1; Hi-St 19.6±1.9; Hi-F initiate basal-bolus insulin titration algorism (INS) or adding Dapaglifl ozin 5mg/ 11.1±1.4 mg/kg/min p<0.05). Analysis of the components of whole body insulin day to the INS (INS/S). We measured indirect calorimetry to provide EE and action showed that Hi-ST and Hi-F mice had liver insulin resistance (reduced respiratory quotient (RQ) during fasting and postprandial before and last day % suppression of HGO) but Hi-ST mice did not exhibit insulin resistance at the of the study. INS/S, compared with INS, the rate of achieved euglycemia was level of muscle glucose uptake (chow 22.1±1.8; Hi-ST 21.0±2.3; Hi-F 14.0±2.5 higher (50% vs. 27.8%, p<0.05), whereas total daily dose of insulin was 19% µmol/100g/min). These results indicate that despite development of signifi - lower and was accompanied by decreased basal/bolus ratio (p=0.02). Before cant obesity, mice fed a Hi-ST diet display a different pattern of glucose toler- and after the treatment, fasting EE and postprandial EE elevation was similar ance and insulin sensitivity compared to obese Hi-F fed mice. The Hi-ST model on both groups. Surprisingly, in INS/S, fasting RQ was signifi cantly increased offers new opportunities to help understand the mechanistic links between after the treatment (0.72±0.05 vs. 0.79±0.08 respectively, p=0.04) with post- obesity and reduced insulin action. prandial RQ elevation was abolished, while INS shows vice versa (p<0.02). Supported By: Diabetes Australia INS/S increased glucosuria and serum ketone body formation. Together, our data indicate that adding SGLT2 inhibitor to the insulin treated diabetes, 1845-P enhance fasting carbohydrate oxidation resulting lower basal insulin require- Alterations in Fatty Acid Composition after Worksite Wellness ment, and sustained energy expenditure regardless of the renal elimination of Intervention glucose as an energy substrate may lead to body weight loss. BERNARD V. MILLER, MARTIN GARRAFFO, PETER J. WALTER, Bethesda, MD Predictive factors of weight loss are needed to target those most likely to 1843-P benefi t from intervention. It is hypothesized that body fat loss is regulated by The FGF-21 Response to Fructose Predicts Metabolic Health in lipogenic activity that can be altered by changes in energy balance and nutri- Obese Humans ent composition. 70 women with obesity were stratifi ed into tertiles of weight

KASPER W. TER HORST, PIM W. GILIJAMSE, MARIETTE T. ACKERMANS, loss after a 6 mo worksite wellness intervention performed at NIH. Body com- Obesity

JOHANNES A. ROMIJN, MAX NIEUWDORP, ELEFTHERIA MARATOS-FLIER, position with visceral fat (VF), diet composition and free (FFA) and esterifi ed POSTERS MARK A. HERMAN, MIREILLE J. SERLIE, Amsterdam, Netherlands, Boston, MA (EFA) fatty acid concentrations and composition were measured. Indexes of

High fructose intake has been implicated in the development of obesity, de novo lipogenesis (DNL), Stearoyl-CoA desaturase-1 (SCD 16 and 18), Elon- Integrated Physiology/ insulin resistance and metabolic syndrome. Recent evidence shows that gase-6 (Elovl-6), Elongase-5 (Elovl-5), Δ5-desaturase (D5D) and Δ6-desaturase Fibroblast Growth Factor-21 (FGF-21) has multiple effects on glucose, lipid (D6D) activity were calculated. In the tertile of most weight loss (T1, n=20) and energy metabolism and may also mediate an adaptive response to fruc- BMI (-3.2 ± 0.4 kg/m2), FM (-7.2 ± 1.1) and VF (-33 ± 5%) decreased (p <.001). tose ingestion. Fructose acutely stimulates circulating FGF-21 consistent with In the tertile of least weight loss (T3, n=21) BMI (0.6 ± 0.2 kg/m2), FM (1.3 ± a hormonal response. We aimed to evaluate whether basal FGF-21 and the 0.4 kg) and VF (10 ± 4%) increased (p<.05). Total FFA concentration increased, FGF-21 response to fructose may be predictive of metabolic parameters. We while total EFA decreased in all tertiles. Total TG concentration decreased recruited 35 obese adults (22 ş, 13 š; age 44 ± 2 y; BMI 44 ± 1 kg/m2) and in T1. FFA-in T1 and T3 16:0, 18:0 and 18:1n-7 decreased, while 12- and 14-C assessed the FGF-21 response to a 75-g oral fructose dose (fructose toler- SFA, MUFA and 16:1n-9 increased. 18:1n-9 and SCD 18 were increased in T1. ance test), and basal and insulin-mediated glucose and lipid fl uxes during a In T3, 18:3n-6, 20:4, DNL, Elovl-6 and D6D decreased, while 18:3n-3, 22:6 and 2 two-step euglycemic hyperinsulinemic clamp with infusion of [ H2]glucose SCD (16 and 18) increased. Baseline SCD 16 predicted weight change (r=.45,

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A475 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

p<.05). EFA - In T1 and T3 12- and 14-C SFA and MUFA, 18:1n-7 and 18:3n-6 a HFD, as shown by glucose and insulin tolerance tests. In vitro studies in decreased. In T1 16:1n-7, 16:1n-9, SCD (16 and 18) and D6D decreased while C2C12 myotubes suggest that mitochondrial stress and ER stress can both D5D increased. In T3 17:0, DNL and D6D decreased, while 18:2 and 18:3n-3 induce FGF-21 expression and secretion, which is blunted when cells are increased. In T1 dietary 18:1n-9 and 18:3n-3 decreased while 17:0 increased. treated with PBA, an ER stress inhibitor. Taken together, these data sug- In T3 14:0, 16:0, 17:0, 18:0, 16:1n-7, 18:1n-9 and 18:2 decreased, while 14:1 gest that OPA1-elicited mitochondrial dysfunction in muscle activates stress increased. We conclude FFA and EFA composition are reciprocally altered by response pathways, including ER stress, inducing FGF-21 expression and changes in body and dietary fat composition. Initial FFA composition predicts secretion, which prevents diet-induced obesity and insulin resistance. bodyweight change and may be important in selecting patients that will ben- efi t most from interventions designed to reduce obesity-related risk. & 1848-P Supported By: National Institutes of Health Impaired Microvascular Blood Flow Accompanies Metabolic Infl ex- i bility in Healthy People with a Family History of Type 2 Diabetes RYAN D. RUSSELL, TIM GREENAWAY, STEPHEN RATTIGAN, STEPHEN M. RICH- INTEGRATED PHYSIOLOGY—MUSCLE ARDS, MICHELLE A. KESKE, Hobart, Australia Metabolic fl exibility (MF) is the capacity to switch fuel oxidation based on availability, and is related to cellular glucose uptake. MF is impaired in type 2 Moderated Poster Discussion: Integrated Physiology—Muscle diabetes (T2D) and in healthy people with a family history of T2D (FH+) vs. (Posters: 1846-P to 1853-P), see page 20. those with no family history of T2D (FH-). Muscle microvascular blood fl ow (MBF) is enhanced in response to insulin, and plays a key role in muscle glu- & 1846-P cose uptake and is blunted during insulin resistance and T2D. Whether MBF Lifelong NFkB Suppression Promotes Muscle Loss is impaired in healthy FH+ is unknown. JOSEPH M. VALENTINE, NING ZHANG, YOU ZHOU, HANYU LIANG, STEVEN E. We fed 14 (7FH-, 8FH+) age and BMI matched (Table 1) overnight-fasted SHOELSON, NICOLAS MUSI, San Antonio, TX, Boston, MA volunteers a liquid mixed meal (MM, 295 kcal). Plasma glucose and insulin A major risk factor for the development of diabetes is an increase in periph- levels were monitored every 30 min over 2 hrs following the MM. Brachial eral insulin resistance that develops with advancing age. Aging has been char- artery blood fl ow (Doppler ultrasound) and forearm muscle microvascular acterized as a “sterile” low-grade infl ammatory state, which is thought to play recruitment (contrast-enhanced ultrasound) was assessed at baseline and a role in skeletal muscle atrophy and insulin resistance. Therefore, we hypoth- 60 min following the MM. MF (indirect calorimetry) was calculated as the esized that suppression of canonical NFkB signaling (a master transcriptional sum of the change in RER from fasting through 60 min of the MM. regulator of the infl ammatory response) would protect against aging-related Both groups had similar plasma glucose and insulin levels before and dur- sarcopenia and insulin resistance. To test this hypothesis, we studied mice ing the MM (Table 1). Howe ver, FH+ had impaired MF and MBF responses to with muscle-specifi c overexpression of a superrepressor IkBα mutant (MISR). the MM, and a smaller brachial artery diameter than FH- (Table 1). MISR mice were protected against aging-related insulin resistance; however, This is the fi rst study showing reduced MBF accompanies impaired MF in contrary to our hypothesis, MISR mice displayed muscle loss compared with healthy FH+, and that both can be detected using a MM. Reduced MBF and wild type (WT) litter-mates. To better understand how life-long suppression MF in FH+ may explained heightened risk for T2D in this population. of NFkB leads to reduced muscle mass, we performed whole genome RNA Table 1. sequencing in muscle from WT and MISR mice. Gene expression of several proteins involved in the regulation of muscle differentiation and growth were upregulated in MISR mice, including myostatin (1.5-fold), GDF11 (4-fold), and p38α (1.2-fold), as well as, proteins within the non-canonical NFkB pathway [Tbkbp1 (3.5-fold) and NIK (2.5-fold)]. Accordingly, gene expression levels of TRAF2 and TRAF3 were suppressed (both by 1.5-fold), which further supports non-canonical NFkB pathway activation. Notably, transduction with an adeno- virus containing an IkbαDN plasmid to C2C12 myotubes for 12 h caused similar changes in protein content of p38α, NIK, TRAF2 and TRAF3 when harvested 48-72 h later. In summary, here we show that suppression of the canonical NFkB pathway, both in vivo and in vitro, results in the altered expression of genes and proteins involved in control of muscle differentiation and growth. Our fi ndings also indicate that the canonical NFkB pathway is important for normal muscle maintenance and its sustained suppression may have harmful effects.

& 1847-P Muscle-derived FGF-21 Prevents Diet-induced Obesity and Insulin Resistance in Mice Defi cient for OPA1 in Skeletal Muscle RENATA O. PEREIRA, SATYA M. TADINADA, FREDERICK ZASADNY, ANGELA C. OLVERA, JENNIFER M. JEFFERS, E. DALE ABEL, Iowa City, IA Optic Atrophy 1 (OPA1) is an inner mitochondrial membrane protein that plays a fundamental role in mitochondrial fusion and respiratory function, and reported to be reduced in some models of insulin resistance and obe- sity. We previously observed that inducible OPA1 defi ciency in adult skeletal muscle (mOPA1 KO) results in progressive non-lethal mitochondrial dysfunc- tion, increased fi broblast growth factor-21 (FGF-21) circulating levels and

Obesity resistance to diet-induced obesity and insulin resistance. We hypothesized POSTERS that the energy-stressed muscle produces and secrets FGF-21 into the circu- lation, conferring this favorable metabolic phenotype. To test our hypothesis

Integrated Physiology/ we generated muscle-specifi c OPA1/FGF-21 double knockout mice (DKO) and fed these mice a high-fat diet (HFD) for 10 weeks. Circulating FGF-21 as well as FGF-21 protein levels were completely normalized in DKO mice. Muscle explants removed from mOPA1 KO mice, DKO mice and their respective WT controls revealed that muscle can secrete FGF-21 in vitro in an autonomous manner. mOPA1 KO mice had a 2-fold increase in secreted FGF-21 relative to WT mice, which was entirely blunted in DKO mice. Upon high-fat feeding, weight gain was similarly increased between WT and DKO mice. Body fat was also equivalently increased in both WT and DKO mice fed a HFD relative Supported By: American College of Sports Medicine; Virtual Tasmanian Aca- to control-fed mice, and no differences were observed in total lean mass. demic Health Sciences Precinct; Royal Hobart Hospital Research Foundation DKO mice became as glucose intolerant and insulin resistant as WT mice on

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A476 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

& 1849-P & 1851-P Glucagon-like Peptide-1 and Insulin at Physiological Concentra- Cardiac Specifi c Inhibition of Mitochondrial Akt1 Signaling Caused tions Increase Cardiac Microvascular Perfusion Independently but Rapid Development of Cardiomyopathy and Insulin Resistance in Not Synergistically in Lean Healthy Adults Mice ALVIN WAI KIT TAN, SHARMILA C. SUBARAN, MATTHEW A. SAUDER, LINDA A. YUMAY CHEN, YU-HAN CHEN, WENJUN FAN, BRIAN A. PEDERSEN, MATTHEW JAHN, ANANDA BASU, ZHENQI LIU, Charlottesville, VA, Rochester, MN FUNSTEN, HSIAO-CHEN LEE, CHARITY JUANG, ROBERT EDWARDS, GRANT R. Both glucagon-like peptide-1 (GLP-1) and insulin (INS) increase micro- MACGREGOR, PING H. WANG, Irvine, CA vascular perfusion in cardiac muscle via different nitric oxide-dependent Mitochondrial dysfunction is associated with diabetic cardiomyopathy. mechanisms; thereby augmenting oxygen, nutrients and INS delivery and Akt1 translocation to mitochondria is reduced in diabetic myocardium. To enhancing peripheral INS actions. To examine whether GLP-1 and INS act investigate for evidence of a causal relationship between mitochondrial synergistically on cardiac microvasculature, 22 lean healthy adults under- signaling dysfunction and development of diabetic cardiomyopathy, we went 2 study protocols in random order after an overnight fast. Subjects generated mice that express mitochondria-targeted dominant negative Akt received either an intravenous infusion of GLP-1 (1.2 pmol/kg/min, protocol (mdnAkt) in an inducible manner into the Rosa26 locus (CAMDAKT mice). 1) or normal saline (protocol 2) for 150 min with a 2-hour euglycemic INS Cardiac specifi c expression was achieved using the Cre-ER-Lox strategy. clamp (1 mU/kg/min) superimposed from 30 min onwards. Cardiac muscle Both male and female mice were analyzed. Upon Tamoxifen injection, microvascular blood volume (MBV), fl ow velocity (MFV) and blood fl ow (MBF) mdnAkt was expressed in the myocardium, but was undetectable in other were determined using contrast-enhanced ultrasound at 0, 30 and 150 min. tissues. The mdnAkt protein product was detected in myocardial mitochon- Brachial artery (BA) diameter, fl ow velocity and blood fl ow were measured dria only, undetected in the cytosol or nucleus. Expression of mdnAkt caused at 0 and 150 min. At 30 min, GLP-1 alone increased cardiac MBV and MBF profound inhibition of mitochondrial Akt signaling. Wildtype mice, Cre mice by 53.6 and 46.2% respectively, and reduced MFV by 3.1% (all p<0.001). Co- and vehicle-injected CAMDAKT mice were used as controls. After Tamoxifen infusing insulin from 30 min onwards did not result in further changes in induction, CAMDAKT mice developed heart failure within 7 days. Histopa- MBV, MFV and MBF at 150 min. In the 2nd protocol, insulin alone increased thology revealed initial cardiomyocyte cell death and altered mitochondrial cardiac MBV and MBF by 70.0 and 68.9%, and reduced MFV by 5.3% (all structure, followed by reduction of muscle mass, infl ammation, cardiac fi bro- p<0.001). The percent change in cardiac MBV, MFV and MBF at 150 min sis, and biventricular heart failure. Liver, lung, and kidney congestion were from baseline were comparable in both protocols (p=0.26, 0.13 and 0.23 accompanied by generalized edema. Survival analysis showed increased respectively). INS-GLP-1 co-infusion signifi cantly increased BA diameter, mortality in the CAMDAKT mice only after Tamoxifen injection. These data fl ow velocity and blood fl ow by 6.2, 18.8 and 35.4% respectively. The cor- indicate that impaired mitochondrial Akt1 signaling can cause heart failure responding increases for INS alone were 7.0, 14.6 and 32.3%, with no sig- in vivo. To analyze the impact on glucose homeostasis, we performed OGTT. nifi cant differences between protocols. We conclude that GLP-1 and INS at Oral administration of glucose induced rapid translocation of phosoho-Akt1 physiological concentrations acutely increase cardiac microvascular perfu- from cytosol to mitochondria in wildtype mice. Plasma glucose levels during sion and dilate conduit artery in lean healthy adults but their effects are not OGTT were identical in the induced CAMAKT and control mice. However, synergistic. This suggests that both regulate oxygen and nutrients delivery higher postprandial insulin in the induced CAMAKT mice suggested insulin to cardiac muscle under physiologic conditions. resistance. This study shed new light on the role of mitochondrial Akt1 sig- naling in diabetic cardiomyopathy. & 1850-P Supported By: National Institutes of Health Lysophosphatidylcholines Activate PPARδ and Protect Skeletal Muscle Cells from Lipotoxicity & 1852-P CHRISTIAN KLINGLER, RAINER LEHMANN, TILL ADHIKARY, HANS-ULRICH Mesenchymal Stem Cell Transplant Improves Motor Function and HÄRING, ERWIN SCHLEICHER, CORA WEIGERT, Tübingen, Germany, Marburg, Modulates Markers of Muscle Atrophy in Diabetic Rats Germany MARIA AUGUSTA SABADINE, THIAGO L. RUSSO, GENOVEVA F. LUNA, ANGELA Lysophosphatidylcholines (LPC) are about to become an important bio- M.O. LEAL, São Carlos, Brazil marker in the prognosis of type 2 diabetes. Metabolomics studies demon- Diabetic neuropathy is the most common diabetic complication for what strate a negative correlation of plasma LPCs with insulin resistance, obe- there is no satisfactory therapy. Stem cell therapy is an attractive strat- sity, and type 2 diabetes. The physiological function of LPCs on metabolic egy, due to their capacity to differentiate into other tissues and their abil- pathways is largely unknown. Here we investigated the effects of LPCs on ity to modulate neurotrophic factors. The aim of this study was to evaluate human skeletal muscle cells differentiated to myotubes. First we validated the effects of mesenchymal stem cells (MSCs) transplant on neuromuscu- the concentrations of LPC (16:0) and LPC (18:1) which do not induce cell stress lar function, skeletal muscle morphology and the expression of genes and parameters. Concentrations higher than 50 µM were needed to induce the proteins associated with muscle atrophy. Diabetes mellitus was induced in release of intracellular creatine kinase and lactate dehydrogenase. Below adult male Wistar rats by a single intraperitoneal (i.p.) injection of streptozo- this concentration neither expression of ER stress marker ATF3 nor of tocin 50mg/kg body weight. Control animals (C) received the same volume of infl ammatory markers IL6 or CXCL3 were affected. Transcriptome analysis vehicle. Eight weeks after the induction of diabetes, diabetic animals were of human myotubes treated with 10 µM LPC for 24 h revealed enrichment divided into 2 groups. Diabetic treated animals (DM-MSCs) received four of up-regulated PPAR target transcripts, including ANGPTL4, PDK4, PLIN2 weekly i.p. injections of bone marrow-derived MSCs (BM-MSCs, 1x106cells/ and CPT1α. The increase in both PDK4 and ANGPTL4 RNA expression was injection) and diabetic non-treated animals received vehicle injections (DM- abolished in the presence of either PPARδ inhibitor GSK0660 and GSK3787. PBS). Anterior tibial muscle was collected 4 weeks after the fi rst injection The induction of PDK4 by LPCs was blocked with siRNA against PPARD. for morphological analysis and for the determination of mRNA (atrogin-1, Luciferase reporter gene assays demonstrated activation of the ligand bind- muRF-1 and myostatin) and protein (atrogin-1, murf-1) expression. Mechani- ing domain of PPARδ by LPC (16:0) and LPC (18:1). To address the direct bind- cal and thermal nociceptive thresholds, walking-track analysis and motor ing of LPCs to PPARδ, gel shift assays were performed using recombinant coordination activity were signifi cantly improved in DM-MSCs compared PPARδ and RXRα. Both LPC (16:0) and LPC (18:1) were able to enhance PPARδ with DM-PBS (P < 0.0001). Muscle mass decreased in diabetic animals and DNA binding activity. Further results show that the LPC-mediated activa- did not change after transplantation. Muscle mRNA and protein expression Obesity POSTERS tion of PPARδ can reduce fatty acid-induced infl ammation in skeletal muscle of atrogin-1 and muRF-1 was signifi cantly higher in diabetic animals than cells. Treatment of human myotubes with 10 µM LPC (16:0) and LPC (18:1) in control animals. Transplantation of BM-MSCs decreased signifi cantly ameliorated palmitate-driven ER-stress analyzed as ATF3 RNA expression the mRNA expression of atrogin-1 and muRF-1 and the protein expression Integrated Physiology/ and XBP1 splicing as well as palmitate-mediated cytokine induction. The of atrogin-1 (P < 0.0001). Myostatin mRNA expression did not differ in the protective effect of LPC was prevented in the presence of GSK0660. Tak- groups. The results demonstrate the improvement of motor function and the ing together, the results show that LPCs are PPARδ agonists and hence can modulation of markers of atrophy of skeletal muscle by BM-MSCs in diabetic contribute to PPARδ-dependent anti-infl ammatory effects. rats. Supported By: German Center for Diabetes Research Supported By: Fundação de Amparo à Pesquisa do Estado de São Paulo

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A477 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

1855-P & 1853-P Multifactorial Regulation of Skeletal Muscle β-F1-ATPase in Human Enhanced GLUT4-dependent Glucose Uptake Relieves Nutrient Obesity Stress through Changes in Amino Acid and Lipid Metabolism LEE TRAN, PAUL LANGLAIS, ELENA A. DE FILIPPIS, TONYA R. BENJAMIN, LORI R. ANN L. OLSON, JAMI GURLEY, Oklahoma City, OK ROUST, CHAD C. CARROLL, LAWRENCE J. MANDARINO, CHRISTOS KATSANOS, Impaired GLUT4-dependent glucose uptake is a contributing factor in the Scottsdale, AZ, Glendale, AZ development of whole body insulin resistance in obese patients and obese Obesity perturbs muscle ATP turnover, but the precise mechanism (s) are animal models. Mice engineered to express human Glut4 gene under the con- unclear. An important rate-limiting step in ATP synthesis is catalyzed by trol of the human Glut4 Promoter (hG4TG), are resistant to obesity-induced the β-subunit of ATP synthase (β-F1-ATPase). We hypothesize that obesity insulin resistance. This is likely due to increased glucose uptake in skeletal decreases the synthesis rate of β-F1-ATPase. Responses were evaluated muscle. To investigate the fate of Glut4-dependent glucose fl ux, we carried before and after parenteral administration of amino acids in age-matched out an analysis of intermediary metabolism in lean and obese hG4TG. We lean (BMI<25) and obese (BMI>30.0) volunteers. A primed, constant infusion found that hG4TG muscle showed down-regulation of multiple key mitochon- of D10-leucine was used to determine β-F1-ATPase synthesis rate in mus- drial enzymes with the exception of pyruvate carboxylase (PC), a potential cle, and an oral glucose tolerance test (OGTT) was used to evaluate insulin anaplerotic enzyme in skeletal muscle. Despite the reduction in key mito- sensitivity via Matsuda index. β-F1-ATPase protein, mRNA expression, and chondrial enzymes, mitochondrial number, size and function was unaffected. ATPase synthase specifi c activity were also quantifi ed. Although obesity Next, we tested the prediction that increased Glut4-dependent glucose fl ux was associated with increased mRNA expression, obese individuals trended would signifi cantly alter intermediary metabolites in muscle. We isolated towards decreased protein expression compared to lean controls indicative hind-limb skeletal muscle from obese non-transgenic (NT) and obese hG4TG of impaired translation. Analysis of β-F1-ATPase synthesis rates revealed mice that had been fasted overnight or allowed ad libitum access to food signifi cant decreases in only obese females, whereas decreased ATP syn- (Fed) (n=5 per group). A metabolomic profi le was obtained using quantitative thase activity was observed in all obese subjects and linked to insulin resis- mass spectrometry. The pool of Kreb’s cycle intermediates was decreased, tance (r=0.59, P<0.01). There was an increase in all parameters following however, citrate was not. The profi le demonstrated that Ala, Gly, Ser and hyperaminoacidemia except ATP synthase activity, which unexpectedly Pro levels were higher in hG4TG mice, while Glu and Gln were reduced. This decreased in lean subjects. Our data outlines important factors regulating is consistent with the notion that glucose fl ux promoted synthesis of amino β-F1-ATPase, which may contribute to poor muscle energy metabolism in acids arising from glycolysis, while synthesis of Glu and Gln arising from obese individuals. Kreb’s cycle intermediates were reduced. All long chain acyl carnitine lev- els were also signifi cantly lower in obese hG4TG mice indicating an effi - Table. cient beta-oxidation. Glut4-dependent glucose fl ux relieved nutrient stress through changes in lipid and amino acid metabolism. These data emphasize the utility of targeting Glut4-dependent glucose uptake for improving insu- lin-dependent glucose homeostasis and insulin sensitivity.

1854-P ApoA-I Provides Glycemic Control In Vivo by Direct Action on Both Insulin Secretion and Peripheral Tissue Glucose Uptake Supported By: National Institutes of Health; National Institute of Diabetes and JOAN DOMINGO-ESPIN, OKTAWIA WOLANIN, MARIA LINDAHL, SAMUEL W. Digestive and Kidney Diseases (DK094062) CUSHMAN, KARIN G. STENKULA, JENS O. LAGERSTEDT, Lund, Sweden Apolipoprotein A-I (apoA-I) of high-density lipoprotein is mainly known for its role in the reverse cholesterol transport pathway. ApoA-I also pro- 1856-P vides glucose control with demonstrated in vitro effects on beta-cell insulin The Role of FGF-21 in Glucose Uptake and Obesity Resistance in secretion and muscle glucose uptake. In addition, apoA-I injections of insulin Mice with Impaired Skeletal Muscle Fatty Acid Oxidation resistant diet-induced obesity (DIO) mice lead to increased glucose-stimu- JAYCOB D. WARFEL, RANDALL L. MYNATT, Baton Rouge, LA lated insulin secretion (GSIS) and peripheral tissue glucose uptake. However, Prevailing hypotheses suggest that fatty acid oxidation is thought to the relative contribution in vivo of apoA-I as an enhancer of GSIS and as a cause ectopic accumulation of lipotoxic species within muscle cells, and direct stimulator of insulin-independent glucose uptake is not known. that these species have the ability to interfere with insulin signaling. We DIO and chow diet mice were instantly and transiently blocked for insulin recently showed, however, that within a mouse skeletal muscle specifi c car- m-/- secretion and utilized in glucose tolerance tests and in positron emission nitine palmitoyl transferase-1b knockout model (Cpt1b ), a lipotoxic envi- tomography (PET) analyzes. Data demonstrates that apoA-I to equal extent ronment is created via increased intramyocellular lipid (IMCL) content, and enhances GSIS and acts as peripheral tissue activator (insulin-independent), yet the animals remain insulin sensitive and have low body fat percentage and verifi es skeletal muscle as an apoA-I target tissue. Interestingly, the as compared to control mice. The inhibition of fatty acid oxidation within heart is identifi ed as an important target tissue for the apoA-I-stimulated this model causes a reduction in mTORC1 signaling, which contributes to glucose uptake, with potential implications in diabetic cardiomyopathy. insulin sensitivity. Coordinately, mTORC2 activity is increased, leading to an m-/- Explorations of apoA-I as a novel antidiabetic drug should extend to treat- increase in FGF-21 expression. FGF-21 secretion is also increased in Cpt1b ments of diabetic cardiomyopathy and other cardiovascular diseases in dia- mice, and these responses are specifi c to skeletal muscle. A double knockout betes. mouse model consisting of a whole body knock out of FGF-21 and a knockout of Cpt-1b specifi cally in skeletal muscle was used to determine the effect Figure. of FGF-21 on the metabolic phenotype within Cpt1bm-/- mice. FGF-21 seems to contribute to the enhancement of glucose uptake and insulin sensitiv- ity of Cpt1bm-/- mice skeletal muscle by negatively affecting the basal phos- phorylation state of AKT2. Additionally, the actions of FGF-21 contribute to

Obesity the increased expression of “browning” markers of uncoupling, lipolysis and POSTERS fat oxidation in white adipose tissue of Cpt1bm-/- mice. However, neither the decreased fat pad weight nor decreased adipocyte size observed in Cpt1bm-/-

Integrated Physiology/ mice is corrected in the double knockout mice. Therefore increased circulat- ing FGF-21 and adipocyte browning are not responsible for the resistance to obesity seen in Cpt1bm-/- mice. Supported By: National Institutes of Health

Supported By: Swedish Research Council (K2014-54X-22426-01-3); Swedish Diabetes Foundation; Albert Påhlsson Foundation; Tage Blücher Foundation; Royal Physiographic Society in Lund

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A478 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

1857-P 1859-P FGF-21, Its Receptors and Target Genes in Skeletal Muscle and Adi- Physical Inactivity and High-Fat Diet Additively Induce Diacylgly- pose Tissue in Obesity and Type 2 Diabetes: Effects of Insulin and cerol Accumulation and Insulin Resistance in Muscle Exercise Training SAORI KAKEHI, YOSHIFUMI TAMURA, NAOKO KAGA, RYUZO KAWAMORI, HIRO- RIKKE KRUSE, SARA G. VIENBERG, BIRGITTE F. VIND, BIRGITTE ANDERSEN, KURT TAKA WATADA, Tokyo, Japan HØJLUND, Odense, Denmark, Måløv, Denmark Physical inactivity (PI) has been shown to impair muscle insulin sensitivity Pharmacologic doses of fi broblast growth factor-21 (FGF-21) improve glu- (M-IS); however, the mechanism is unknown. In addition, although PI is tend cose tolerance, lipid metabolism and energy expenditure in rodent models. to coincident with high fat diet (HFD), the combined effect of them has not Induced expression and secretion of FGF-21 from muscle increase brown- been examined. To elucidate them, we used 24 h hind-limb cast immobiliza- ing of white adipose tissue in a myokine-like manner. Recent studies have tion (HCI) for PI model. We divided C57BL6J mice into four groups; control, reported that insulin and training induce FGF-21 levels in muscle and plasma. 24 h HCI, 2 wk HFD, and 24 h HCI after 2 wk HFD. The M-IS was evaluated by Obesity and T2D are potentially FGF-21 resistant states, but to what extent ex vivo insulin-stimulated 2-deoxy glucose uptake. We found that 24 h HCI the FGF-21 responses to insulin and training are preserved, and whether increased intramyocellular diacylglycerol (IMDG) (Figure B), known to acti- FGF-21 receptors and downstream target genes are altered remains to be vate PKC and impair insulin signaling, while IM triacylglycerol (TG) was not established. Here, we investigated the effect of insulin and training on changed. Interestingly, whereas 2 wk HFD did not change IMDG and IMTG serum FGF-21, the effect of insulin on expression of FGF-21 and its recep- levels, 24 h HCI after 2 wk HFD dramatically increased IMDG (Figure B). In tors in skeletal muscle biopsies, and the infl uence of overweight/obesity and parallel with IMDG accumulation, PKCε activity and oxidative stress (OS) T2D on these responses as well as on expression of FGF-21 receptors and were increased, and M-IS (Figure A) and insulin signaling were reciprocally downstream target genes in fat biopsies. Insulin increased serum FGF-21 impaired. The IMDG accumulations were also accompanied by increased and FGF-21 mRNA levels in muscle with no infl uence of overweight/obesity muscle mRNA of adipose triglyceride lipase (ATGL) and Lipin1, enzymes for or T2D, and no effect of training. The increase in muscle FGF-21 expression DG synthesis. In addition, we identifi ed OS increased ATGL and Lipin1 mRNA correlated strongly with the increased serum FGF-21 in response to insulin. in vitro. Finally, we applied 24 h leg CI in human and found increased IMDG Serum and muscle FGF-21 tended to be higher in T2D compared with lean and mRNA of ATGL and Lipin1 in muscle. These results suggested that 24 and overweight/obese controls. In adipose tissue, expression of β-Klotho h HCI decrease M-IS, which is furthered by HFD. Increased IMDG and OS was reduced and FGFR1c expression was increased in overweight/obesity might be involved in the mechanisms. and T2D, whereas the expression of most target genes (ACACB, EGR1, LEP, Figure. LIPE, PCK1, ATGL, PPARGC1A, SCD or GLUT1) was unaltered. UCP1 was hardly expressed in adipose tissue, whereas CIDEA expression was decreased in T2D. Insulin stimulation of muscle FGF-21 expression correlates strongly with the increase in serum FGF-21, and this response appears intact in over- weight/obesity and T2D. FGF-21 resistance may involve reduced β-Klotho expression in adipose tissue. This seems to be compensated by increased expression of FGFR1c as the expression of FGF-21 target genes is unaltered in adipose tissue.

1858-P TGFβ Signalling Suppresses Key Mitochondrial Regulators in Human Skeletal Muscle Cells CHRISTOPH HOFFMANN, ANJA BOEHM, CLAUDIA AL-MARDINI, ANDREAS FRITSCHE, HANS ULRICH HÄRING, CORA WEIGERT, Tübingen, Germany Individuals differ signifi cantly in their response to comparable exercise loads, even after adjustment for basic anthropometric parameters. This includes a high variability in the improvement of insulin sensitivity. Whole genome expression analyses of muscle biopsies of high responders in insu- lin sensitivity compared to low responders point to a reduced activation 1860-P and abundance of PGC1α and AMPK in the muscle of low responders after The Defi ciency of Pyruvate Dehydrogenase in the Heart Alters Car- training and increased TGFβ signaling. Using primary human satellite cell diac Glucose Oxidation and Sensitizes Heart to Ischemic Insults culture, we studied the functional consequences of enhanced TGFβ signal- WANQING SUN, NANHU QUAN, LIN WANG, COURTNEY A. CATES, JI LI, Jackson, ing in human muscle and the potential infl uence of TGFβ on mitochondrial MS regulators. First we validated functional TGFβ signaling in differentiated Pyruvate dehydrogenase (PDH) plays a key role in aerobic energy metabo- human muscle cells and established the TGFβ-induced protein TGFBI as a lism and occupies a central crossroad between glycolysis and the tricarbox- time- and concentration-dependent marker of active TGFβ1. We showed ylic acid cycle. We generated inducible cardiac-specifi c PDH knockout mice that the primary myotubes were capable to either secrete TGFβ1 or activate that demonstrated a high mortality rate. It was hypothesized that pyruvate latent TGFβ1 in their surroundings. Moreover, we demonstrate that TGFβ dehydrogenase modulating cardiac glucose metabolism is crucial for heart signaling and PGC1α/AMPK expression are interconnected. TGFβ1 stimula- functions under normal physiological and/or stress conditions. The myocar- tion reduced abundance of PGC1α, TFAM and AMPK transcripts, while an dial infarction was conducted by a ligation of the left anterior descending antagonist had the opposing effect. TGFβ1 stimulation also suppressed coronary arteries. The results demonstrated that cardiac PDH defi ciency expression of key mitochondrial enzymes, while it upregulated cytokine caused a larger myocardial infarcts size and macrophage infi ltration in the expression. The impact of TGFβ1 as transcriptional suppressor was indepen- hearts (p<0.01 vs. WT mice). Wheat germ agglutinin and Masson Trichrome dent from the level of muscle cell differentiation, but we observed strong staining revealed a signifi cantly increased hypertrophy and fi brosis in PDH- effects on myotube formation. Notably, TGFβ1 signaling as such was neither defi cient hearts (p<0.05 vs. WT). The measurements of heart substrate Obesity impaired in cells obtained from high, nor from low exercise responders. Our metabolism in an ex vivo working heart perfusion system demonstrated a POSTERS results demonstrate that TGFβ1 blocks expression of mitochondrial enzymes signifi cant impairment of glucose oxidation in PDH-defi cient hearts during important for fuel oxidation and thus activation of TGFβ signaling in skeletal ischemia/reperfusion (p<0.05 vs. WT). However, Dichloroacetate, a PDH Integrated Physiology/ muscle can be involved in the different metabolic response to exercise. We activator, signifi cantly increases glucose oxidation in WT hearts during isch- suggest that an intramuscular cross-talk of different cell types resident in emia/reperfusion and reduces myocardial infarct size in WT, but not in PDH- skeletal muscle contribute to the release and activation of TGFβ. defi cient hearts. Furthermore, the Immunoblotting data showed that cardiac Supported By: German Center for Diabetes Research PDH defi ciency leads to an impaired ischemic AMPK activation through Ses- trin2-LKB1 interaction which is responsible for an increased susceptibility of PDH defi cient heart to ischemic insults. Thus, cardiac PDH defi ciency impairs ischemic AMPK signaling and sensitizes hearts to ischemic damage. Supported By: American Diabetes Association (1-14-BS-131 to J.L.); National Institutes of Health

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A479 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

1861-P 1863-P Insulin Resistance Causes Mitochondrial Lipid Remodelling Along Subjects with Type 1 Diabetes Demonstrate Metabolic and Micro- with Increased Respiration in C2C12 Myotubes vascular Insulin Resistance Compared with Healthy Controls LISA KAPPLER, JIA LI, CHUNXIU HU, STEFANIE HAUCK, ULI OHMAYER, HEIKE LINDA JAHN, AMANDA KLEINER, LEE HARTLINE, FARHAD HASAN, ZHENQI LIU, RUNGE, GUOWANG XU, HANS-ULRICH HÄRING, CORA WEIGERT, RAINER LEH- EUGENE J. BARRETT, Charlottesville, VA MANN, MIRIAM HOENE, Tübingen, Germany, Dalian, China, Munich, Germany, Arterial vascular disease causes morbidity and is the major cause of mor- Dalian, Germany tality for type 1 diabetic patients (DM1). Metabolic insulin resistance (IR) in The causalities of metabolic overfl ow, insulin resistance and mitochon- the general population raises CVD risk 1.5 to 3-fold. In obesity and type 2 dia- drial dysfunction are not resolved. Here, we compared the consequences betes metabolic IR is accompanied by vascular IR, characterized by impaired of insulin resistance (IR) in the absence or presence of high glucose concen- vasodilatory actions of insulin on resistance and microvascular vessels. trations for the mitochondrial lipidome, proteome, and respiration in C2C12 Vasodilatory responses of conduit and resistance vessels are reported to cells. IR was introduced by chronic high insulin concentrations (1 µM) com- be impaired in DM1 subjects. Microvascular IR has not been systematically bined with different levels of glucose (25 mM vs. control 5.5 mM). Cells were studied in DM1. also supplemented with 50 µM each of palmitate and oleate during the 48 We assessed the conduit artery stiffness (pulse wave velocity-PWV and h-treatment with 13C-labelled palmitate for the last 18 h to track fatty acid augmentation index-AI) and skeletal muscle microvascular blood volume incorporation into lipids. Functional respiratory analyses, stable isotope- (MBV) using contrast enhanced ultrasound before and after a 2 h euglycemic assisted lipidomics and proteomics analyses of isolated mitochondria were insulin clamp (1 mU/kg/min) in 8 DM1 (age 25 ± 0.7, BMI 23.8 ± 0.8) and 8 applied. healthy controls (age 25 ± 2, BMI 22.4 ± 0.6). VO2 max (Bruce protocol) was IR increased oxidative phosphorylation and maximal respiration through obtained on a separate day. complex II, which was signifi cant under high glucose conditions, potentially Despite a higher steady-state clamp plasma [glucose] in DM1 subjects indicating a compensatory mechanism during substrate overfl ow. Lipidom- (108 ± 5 vs. 78 ± 2, p<0.01) their glucose infusion rate (GIR) was lower (4.7 ± ics resulted in the detection of 492 lipid species including 118 incorporating 0.9 vs. 7.0 ± 0.5 p=0.05) indicating metabolic IR. MBV decreased in response one and 22 incorporating two 13C-palmitate residues. Phosphatidylethano- to insulin in DM1 (-10 ± 23% change from baseline to post-clamp) but rose lamines (PE) were increased by IR, and this effect was also signifi cant under (55 ± 12%, p=0.03) in controls. This lack of vasodilator response to insulin in high glucose conditions only. Importantly, mitochondrial PE content has pre- DM1 indicates microvascular IR. The change of MBV as well as the VO2max viously been shown to correlate with electron transport chain activity and correlated with GIR (r = 0.65, p<0.01; r =0.69, p< 0.01 respectively). Vascular respiration. In addition, we observed a higher incorporation of 13C-palmitate stiffness did not differ signifi cantly between groups and AI correlated with into cardiolipins (CL) in IR myotubes (signifi cant for low glucose). IR-induced PWV at baseline (r = 0.53, p< 0.05) and at 120 min (r = 0.54, p< 0.05). VO2 max changes were also visible by distinct changes in the proteomics pattern. Our also correlated with AI at baseline (r =0.60, p< 0.05) but not post clamp. results indicate that lipid remodeling contributes to the functional changes We conclude that compared to healthy controls DM1 subjects demon- caused by IR in mitochondria from C2C12 myotubes. strate both metabolic and microvascular IR. The microvascular IR is present Supported By: German Center for Diabetes Research even in the absence of microalbuminuria, indicating it occurs early in DM1. Microvascular IR contributes to metabolic IR and fi tness may mitigate the 1862-P impact of microvascular IR on metabolic IR. Characteristics of Insulin Sensitivity/Secretion, Body Composition, Supported By: National Institutes of Health and Physical Fitness Underlying IFG and IGT in Women with Recent Gestational Diabetes 1864-P ANDREAS LECHNER, NORA N. SOMMER, CHRISTINA HAWLITSCHEK, SABRINA Myokine Regulation of Insulin Secretion: Impact of Infl ammation REIF, ANNE POTZEL, DANIEL POPP, VANESSA SACCO, CARMEN WICHMANN, and Type 2 Diabetes FRIEDERIKE BANNING, INES FREIBOTHE, HARALD GRALLERT, HOLGER HETTER- ALEXANDER J. RYAN, ROBERT R. HENRY, THEODORE P. CIARALDI, San Diego, ICH, JOCHEN SEISSLER, UTA FERRARI, Munich, Germany CA, La Jolla, CA Type 2 diabetes often develops from one of two distinct prediabetic Skeletal muscle (SkM) secretes factors (myokines) that can exert multi- states, IFG and IGT. Understanding the pathophysiology of these conditions ple actions, including on β-cell mass and function. The secretion of certain can therefore help to clarify the origins of T2D, which are still not fully under- myokines has been shown to differ between nondiabetic (ND) and type 2 stood. diabetic (T2D) subjects. To study the control of myokine regulation of β-cell We cross-sectionally examined 197 women with recent gestational diabe- function, SkM biopsies were taken from ND and T2D subjects and satellite tes (GDM), a high-risk group for T2D, and asked the question, which abnor- cells cultured to myotubes (MT). Conditioned media (CM) was collected from malities of insulin sensitivity and secretion (oGTT/ivGTT) are associated MT after 24 hr and used to treat INS-1 cells for 24 hr. Cell viability, total insu- with isolated IFG (n=31), isolated IGT (n=23) and combined hyperglycemia lin content, glucose-stimulated insulin secretion (GSIS) and maximal (IBMX- (IFG+IGT and newly diagnosed T2D; n=18). We then tried to explain our fi nd- stimulated) IS were monitored. MT were treated with lipopolysaccharide ings by parameters of antropometrics, body composition (MRI/MRS; n=86) (infectious infl ammation - II) or a combination of glucose (11 mM), insulin and physical fi tness (ergospirometry; n=140). (120 pM), and palmitate (0.3 mM) (metabolic infl ammation - MI) to model the Fasting insulin resistance (IR) was present in all 3 hyperglycemic groups infl ammatory and metabolic conditions seen in vivo: media not exposed to (fasting serum insulin p<0.002 vs. NGT for all) but postabsorptive IR was only MT served as controls. Under baseline conditions, CM from ND and T2D MT found in IGT and combined hyperglycemia (2 h s-insulin in the oGTT p<0.0001 had no effect on INS-1 cell viability, insulin content, or GSIS. After exposure for both). 30 min insulin release in the oGTT (p=0.0004 for both) and fi rst- to II, CM from ND-MT augmented GSIS in INS-1 cells by 100 ± 25% over con- phase secretion in the ivGTT (p=0.03 and p=0.0004; n=86) were reduced in trol, T2D-CM had no effect. After exposure to MI T2D-CM suppressed GSIS IFG and combined hyperglycemia. In multivariate linear regression models, by 35 ± 5%, ND-CM was without effect. Under these conditions cell viability, high BMI was predictive of fasting s-insulin (R2=0.5) whereas high BMI, high total Ins content and maximal IS were unaffected. Effects of CM on GSIS intramyocellular lipid content in soleus muscle and low VO2max were pre- were lost after CM was boiled, implicating protein (s) as the responsible

Obesity dictive of 2 h s-insulin, albeit to a lower extent (R2=0.2). No relevant model factor (s). Both augmentation of GSIS by ND-CM from II-treated MT, and

POSTERS could be found for fi rst-phase insulin release. suppression by T2D-CM from MI-treated MT were inhibited by wortmanin In conclusion, fasting IR, strongly related to obesity, contributes to IFG and and SB203580, implicating PI3-K and p38 MAPK in mediating these effects.

Integrated Physiology/ IGT in young women with recent GDM. Reduced fi rst-phase insulin secre- In summary: 1.) ND-MT are able to augment GSIS when stressed, 2.) T2D-MT tion is found only in IFG whereas postabsorptive IR is the hallmark of IGT. responding to a diabetic-like environment secrete myokines that suppress Postabsorptive IR is linked to obesity, low physical fi tness and high muscle GSIS, 3.) Unknown protein myokines exert effects specifi cally on GSIS, pos- lipid content, but only weakly, and low fi rst-phase insulin secretion is not sibly through p38 MAPK. In T2D, both insulin resistance and a suppression explained at all by the parameters measured. Other causes for these two of adaptive increased insulin secretion are intrinsic properties of SkM that metabolic disturbances have to be sought in future studies. can contribute to the full T2D phenotype. Supported By: German Center for Diabetes Research Supported By: U.S. Department of Veterans Affairs (I01CX00635)

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A480 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

1865-P 1867-P Eight Weeks of Overfeeding Decrease Mitochondrial Content, Func- Decreased Irisin Levels and the Effect of Exenatide in Obese tion, SIRT6 and Mitochondrial SIRT3 Protein Content in Humans Patients with Newly Diagnosed Type 2 Diabetes JEFFREY COVINGTON, DARCY L. JOHANNSEN, CAITLIN HEBERT, ZHENGYU GUANG WANG, YING WANG, JIA LIU, YANJIN HU, YUAN XU, Beijing, China ZHANG, KEVIN CONLEY, CHARMAINE TAM, STEVEN R. SMITH, ERIC RAVUSSIN, Irisin is a novel myokine secreted by skeletal muscles during exercise. SUDIP BAJPEYI, Baton Rouge, LA, Seattle, WA, Sydney, Australia, Winter Park, FL, Abnormal circulating irisin levels have been associated with obesity and El Paso, TX type 2 diabetes. This study aimed to investigate serum irisin levels in obese Excess dietary fat intake can contribute to insulin resistance, which has patients with newly diagnosed type 2 diabetes before and after glucagon- often been shown to be associated with mitochondrial dysfunction in skele- like peptide-1 (GLP-1) receptor agonist-exenatide treatment. A total of 129 tal muscle. One of the purpose of this study was to investigate the effects of participants, including 54 obese patients with newly diagnosed with type 2 8-weeks of high-fat overfeeding on skeletal muscle mitochondrial content, diabetes and 75 age- and sex-matched healthy subjects with normal glucose function, and molecular markers known to regulate mitochondrial function: tolerance, were enrolled. All of the type 2 diabetic patients were assigned p65/RelA and SIRT6. to 12 weeks of exenatide treatment. Obese patients with newly diagnosed 35 volunteers (N=29m/6f; BMI: 25.5 ± 2.2 kg/m2; age: 26.7 ± 5.3 yrs) were type 2 diabetes had lower irisin levels than the control group (P < 0.01). overfed for 8 weeks (40% excess calories, 44% from fat). Mitochondrial Exenatide administration markedly decreased the elevated BMI, FBG, HbA1c content and functions was assessed by skeletal muscle OXPHOS protein and HOMA-IR and increased HOMA-β levels in obese patients with type 2 (western immunoblotting), and ex vivo mitochondrial respiration (Clarke- diabetes (all P < 0.01). A signifi cant increase in irisin levels was observed type electrode) respectively. Mitochondrial protein fraction was extracted after exenatide treatment (P < 0.01). The change in irisin levels was sig- using the Chapell-Perry method, p65/RelA level was measured using real nifi cantly correlated with the decreases in FBG and HbA1c after exenatide time qPCR, SIRT6 and SIRT3 protein content were measured using western treatment (FBG: r = -0.35; HbA1c: r = -0.37; all P < 0.05). Exenatide treatment immunoblotting. signifi cantly elevated the decreased irisin levels of type 2 diabetic patients. Eight weeks of overfeeding resulted in an increase in body weight (6.9 ± After exenatide treatment, the changes in irisin levels were correlated 2.0 kg. p<0.001). Overfeeding resulted in a decrease in all fi ve mitochon- with the decreases in FBG and HbA1c. The upregulation of irisin might be drial OXPHOS protein content (Complex I-V; all p<0.05) and maximal ex vivo a novel mechanism for the benefi cial effects of exenatide in type 2 diabetic mitochondrial respiration (32.29 ± 3.48 to 25.34 ± 2.13 nmol O2/min; P=0.04). patients. Overfeeding also resulted in signifi cant increase in p65/RelA mRNA (p=0.04) Supported By: National Natural Science Foundation of China (81270369, and a decrease in SIRT6 protein content (p=0.03). Although the whole tissue 81070244, 30770873); Beijing Natural Science Foundation (7142060); Beijing SIRT3 protein did not change, overfeeding led to a signifi cant decrease in Municipal Administration of Hospitals (QML20150308) mitochondrial fraction of SIRT3 protein content (p=0.05). In summary, signifi cant overfeeding in humans decreased mitochondrial 1868-P content and function. This dysregulation might be mediated through lower Sex Differences in Insulin Sensitivity in Youth without Diabetes Exist, levels of skeletal muscle SIRT6 and mitochondrial SIRT3 protein with over- but Have No Relationship with Muscle Mitochondrial Function feeding. MELANIE CREE-GREEN, LAURA PYLE, MARK S. BROWN, BRADLEY R. NEW- Supported By: National Institutes of Health COMER, BRANDY RINGHAM, KRISTEN J. NADEAU, DANA DABELEA, Aurora, CO, Harrisonburg, VA 1866-P Type 2 diabetes is increasingly prevalent in youth, more common in ado- Quantitative Proteomics Analysis Reveals Abnormalities of Global lescent females and forecasts early morbidity and mortality. Decreased Kinome Interactome in Obese Insulin Resistant Human Partici- insulin sensitivity (IS) is associated with muscle mitochondrial dysfunction pants in individuals with type 2 diabetes or prediabetes. However, it is not known YUE QI, ABDULLAH MALLISHO, DIVYASRI DAMACHARLA, NISHIT SHAH, MAJED if this relationship exists in youth without diabetes or if there is a difference ALHARBI, ALICE HU, MICHAEL A. CARUSO, DANJUN MA, XIANGMIN ZHANG, between sexes in adolescence. We studied 207 healthy youth from various ZAHER MSALLATY, BERHANE SEYOUM, ZHENGPING YI, Detroit, MI, Oklahoma Colorado cohorts: 101 males (age 17.1 years, BMI-Z 0.38, 68% Tanner 5) and City, OK 106 females (age 17.3 years, BMI-Z 0.5, 76% Tanner 5). IS was assessed with Protein kinases are key regulators which interact with other proteins for TG:HDL ratio and oral glucose tolerance test Matsuda index. Post-exercise signal transduction and other biological functions. Deregulation of protein muscle mitochondrial function was assessed by 31-P MR spectroscopy dur- kinases and their interaction partners are among main causes for insulin ing near maximal isometric calf exercise and summarized as the ADP time resistance and type 2 diabetes (T2D). Nonetheless, little has been reported constant and rate of oxidative phosphorylation (Oxphos). Data are expressed for global profi le of kinase interaction partners in human subjects with as median (25%ile, 75%ile). Compared to males, females had slower Oxphos or without insulin resistance. Here, we present a comparison of kinome [0.14 (0.08, 0.20) vs. 0.12 (0.06, 0.17) seconds, p=0.04], higher TG:HDL ratio interactome between 8 lean healthy control and 8 obese insulin resistant [1.77 (1.29, 2.7) vs. 1.53 (0.96, 2.0), p=0.007] and lower Matsuda score [3.7 participants using quantitative proteomics. Sixteen muscle biopsies were (2.5, 5.1) vs. 5.42 (3.4, 7.6), p<0.001]. Yet when adjusted for BMI-Z score and/ homogenized and labeled with ATP probes, while no probe labeling for or Tanner stage, only the Matsuda score remained signifi cantly different nonspecifi c controls. Protein kinases and their interaction proteins were between the sexes. Further, there was no relationship between any mea- pulled down and digested, followed by HPLC-ESI-MS/MS using an Orbitrap sure of IS and mitochondrial function in the entire cohort or within either sex, Elite. We identifi ed 95 protein kinases and 1507 kinase interaction partners with or without adjustment for BMI-Z or pubertal stage. In summary, healthy (with at least 2 unique peptides and are greater than 10 fold enrichment ratio female adolescents have decreased IS regardless of BMI-Z score compared compared to nonspecifi c controls), which provided the largest kinome inter- to males, but do not have decreased post-exercise mitochondrial function. actome in human skeletal muscle. We quantifi ed 58 kinases and 673 interac- Further, clinical measures of IS do not relate to muscle mitochondrial func- tion partners which were identifi ed with Stable Isotope Labeling by Amino tion across a large cohort of healthy youth. Mitochondrial function per se Acids in Cell Culture (SILAC) normalized peak area in more than half of the does not appear to be related to early abnormalities in IS and instead IS may

kinome pull down (i.e., >8 biopsies used). Interestingly, 147 of kinase inter- be related to factors such as activity, diet and adiposity which may lead to Obesity

action partners changed signifi cantly (p<0.05) between lean controls and vascular and mitochondrial dysfunctions. POSTERS obesities, and 125 of those in obesities showed elevated association (>1.5 Supported By: American Diabetes Association (7-11-CD-08 to K.J.N.);

fold) with kinases compared with lean controls while only 22 proteins exhib- R01DK068001 (to D.D.); National Center for Research Resources (K23 RR020038- Integrated Physiology/ ited decreased levels (<0.67 fold) in obesities. Pathways analysis for the 147 05 to K.J.N.); JDRF (11-2010-343 to K.J.N.); 1R56DK088971 (to K.J.N.); T32 interaction proteins revealed that 10 participated in diabetes pathways, 8 in DK063687, K12HD057022, K23 DK107871 (to M.C-G.) glucose metabolic process, 7 in protein degradation and 7 in infl ammation. These abnormalities of kinome interactome in subjects with insulin resis- tance provide novel insights to understand the molecular mechanisms for the pathogenesis of T2D.

ADA-Supported Research & Moderated Poster Discussion For author disclosure information, see page A696.

A481 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

INTEGRATED PHYSIOLOGY—OTHER HORMONES & 1871-P Excess Androgen Receptor Activation Produces Hyperinsuline- mia, Insulin Resistance, and Secondary Beta-Cell Failure in Female Moderated Poster Discussion: Integrated Physiology—Other Hor- Mice mones (Posters: 1869-P to 1874-P), see page 20. JAMIE MORFORD, CAMILLE ALLARD, FRANCK MAUVAIS-JARVIS, New Orleans, LA & 1869-P Women with polycystic ovarian syndrome (PCOS) have androgen excess A Novel Glucagon-like Peptide-1 and -2 Co-agonist Improves Glyce- and are predisposed to insulin resistance and type 2 diabetes. Studies exam- mic Control and Increases Intestinal Mass in Diabetic Mice ining this issue have focused on the role of testosterone excess as a risk factor SANNE MØLLER KNUDSEN, PERNILLE WISMANN, GITTE HANSEN, SØREN for obesity and insulin resistance in skeletal muscle. However, women with LJUNGBERG PEDERSEN, NIELS VRANG, JACOB JELSING, Hørsholm, Denmark hyperandrogenemia also show pancreatic beta-cell dysfunction. In these Bariatric surgery is currently the most effective treatment option for obe- women, hyperinsulinemia and beta-cell dysfunction are closely associated sity showing concomitant effects on glucose regulation. This has incited with testosterone concentrations, independent of insulin resistance. This a great interest in developing a pharmaceutical mimetic. Gastric bypass raises the possibility that in women with PCOS, excess testosterone induces leads to a marked hypertrophy and cell hyperplasia of the gut which may beta-cell hyperfunction and predisposes to secondary beta-cell failure. To be responsible for the increased levels of circulating gut hormones and test this, we generated an inducible β-cell AR knockout (βARKO) mouse by hence the effect on glucose homeostasis. We therefore hypothesize that crossing ARlox/lox mice with the β-cell specifi c Ins1-Cre/ERT transgenic the combination of the insulinotropic hormone, GLP-1, with the intestino- mouse. AR deletion was induced in adulthood using tamoxifen. We report tropic hormone, GLP-2, could have benefi cial effects on glucose homeosta- that control female mice exposed to chronic androgen excess exhibit fasting sis. A lead compound (GUB09-123) was generated using solid-phase peptide hyperinsulinemia associated to hyperglycemia demonstrating islet failure to synthesis. The compound exhibits potent agonistic effects on both GLP-1 compensate for insulin resistance as observed in hyperandrogenic women. and GLP-2 receptors. Subsequently, GUB09-123 (250 nmol/kg) was adminis- This effect of androgen excess was not observed in female βARKO mice that tered bi-daily s.c. to db/db mice for eight weeks, followed by assessment of remained normoglycemic and insulin sensitive. Testosterone exacerbated glucose homeostasis and intestinal morphometry as primary experimental glucose-stimulated insulin secretion in cultured female mouse and human endpoints. We fi nd, that chronic administration of GUB09-123 to db/db mice islets, an effect that was abolished in βARKO islets and human islets treated improve glucose tolerance by 62% after 4 weeks and by 71% after 8 weeks with the AR antagonist fl utamide, confi rming results obtained in vivo. This of treatment (assessed as total AUC during OGTT, p<0.001 vs. vehicle). Fur- study indicates that excess androgen action in beta-cells produces insulin thermore, GUB09-123 improve glycemic control through a decrease in freely hyper-production leading to secondary insulin resistance and beta-cell fail- fed blood glucose levels (15.9 mmol/l vs. 5.9 mmol/l, p<0.001) and a 1.8% ure with strong implications for the etiology of PCOS. reduction of HbA1c levels (p<0.001 vs. vehicle). Subsequent stereological Supported By: American Diabetes Association (7-13-BS-101 to F.M-J.); National analysis of the entire intestine reveal a marked increase in intestinal mass Institutes of Health (R01DK074970) (63%, p<0.01 vs. vehicle) after eight weeks of treatment. The present data suggests that a GLP-1/GLP-2 co-agonist has remarkably benefi cial effects & 1872-P on glucose homeostasis indicating its potential in treatment of type 2 dia- The Antidiabetes Effect of Leptin in Streptozotocin-treated Mice Is betes. Blocked by Dopamine Antagonism SHUQIN LUO, MICHAEL EZROKHI, SCOTT WATTERS, CHRISTINE CARDI, & 1870-P ANTHONY H. CINCOTTA, Tiverton, RI Increase in Serum Fibroblast Growth Factor-21 (FGF-21) Predicts Type 1 diabetes (T1D) induced by streptozotocin (STZ) in rodents is revers- Development of Diabetes in IGT Subjects and Pioglitazone Atten- ible by pharmacological administration of leptin (L), though the mechanism is uates the Increase in FGF-21 and Reduces Conversion of IGT to unknown. Studies suggest that central dopamine (D) may mediate in part the T2DM antidiabetes effects of leptin in type 2 diabetes animal models. The present RYAN ALONZO, ALBERTO CHAVEZ-VELAZQUEZ, ROBERT MARTINEZ, JORGE study investigated whether blockade of dopamine activity may attenuate VELEZ-GARZA, MARCO CARREON, NICOLAS MUSI, RALPH A. DEFRONZO, anti-hyperglycemic effects of L in STZ induced T1D mice. Male C57BL/6J DEVJIT TRIPATHY, San Antonio, TX mice were given two i.p. injections 4 days apart of STZ (120 mg/kg, N=30) FGF-21 reduces serum glucose and lipid levels in animal models of diabe- or vehicle (non-STZ, N=6). Four days after second STZ treatment, fasting tes. In obese and type 2 diabetic humans, IGF-21 levels are elevated, sug- plasma glucose (FPG) levels were elevated in STZ vs. non-STZ mice (433 ± gesting FGF-21 resistance. We examined the effect of pioglitazone on serum 10 mg/dl vs. 159 ± 6 mg/dl, p<0.0001). Thereafter, STZ mice were divided FGF-21 and the relationship between change in serum FGF-21 and glucose into three treatment groups (STZ/vehicle [STZ/V], STZ/L/V [recombinant tolerance in subjects with IGT. The ACT NOW study examined whether pio- mouse L continuously infused via subcutaneous osmotic pump (20 µg/day)], glitazone (PIO) could prevent/delay development of T2DM. 602 IGT subjects STZ/L/D antagonists [Leptin infusion plus dopamine D1 and D2 antagonists were randomized to PIO (45 mg/day) or placebo (PLAC) and followed for 2.4 (SCH 23390, 0.5 mg/kg; Haloperidol 1 mg/kg, administered i.p. daily one hour years. From the original cohort, we randomly chose 80 IGT subjects; PIO before the onset of waking)] (N=10/group). Six days later, relative to non-STZ (n=41, 29F/12M, age 53 ± 1.7 yrs, BMI 34 ± 0.8 kg/m2) and PLAC ( n=39, mice, STZ/V mice exhibited increased FPG by 3.6 fold (552 ± 16 mg/dl vs. 152 ± 26F/13M age 49 ± 1.9 yrs, BMI = 33±0.8 kg/m2). Indices of insulin secretion 6 mg/dl; p<0.0001), and reduced plasma insulin by 85% (0.13 ± 0.03 ng/ml and insulin sensitivity were derived from the plasma glucose and insulin con- vs. 0.84 ± 0.15 ng/ml; p<0.0001) and plasma L by 90% (0.11 ± 0.06 ng/ml vs. centrations during OGTT. FGF-21 was measured by ELISA. After 2.4 years, 1.15 ± 0.21 ng/ml; p<0.0001). Relative to STZ/V treatment, STZ/L/V treat- 13 subjects converted to T2DM; 11 in the PLAC group and 2 in the PIO group ment reduced hyperglycemia by 53% (262 ± 26 vs. 552 ± 16 mg/dl; p<0.0000), (p<0.005). There was no difference in FGF-21 between PLAC and PIO groups without altering plasma insulin level (0.11 ± 0.03 ng/ml vs. 0.13 ± 0.03 ng/ml), at baseline (324±31 vs. 357±30 pg/ml, p=ns). After 2.4 years, FGF-21 levels though dramatically increasing plasma L (9.0 ± 0.3 ng/ml). Furthermore, com- in PLAC group increased (324±31 to 460±52 pg/ml, p<0.05), while no change pared to STZ/L/V, STZ/L/D antagonist treatment blocked 77% of the effect

Obesity was observed in PIO group (357±30 to 377±30 pg/ml, p=ns). FGF-21 increased of L on hyperglycemia (486 ± 24 mg/dl vs. 262 ± 26 mg/dl; p<0.00001) without POSTERS from 328±59 to 609±89 pg/ml (p<0.05) in IGT subjects who converted to altering plasma insulin (0.12 ± 0.02 ng/ml) or L (7.7± 0.8 ng/ml). These fi ndings T2DM. In contrast, FGF-21 did not change in subjects who remained IGT indicate that the antidiabetes effect of leptin in insulin defi cient mice can be

Integrated Physiology/ (343±52 vs. 316± 64 pg/ml, p=ns) or reverted to NGT (366±39 vs. 376±49 pg/ largely attenuated by dopamine antagonism. ml, p=ns). At study end FGF-21 progressively increased from NGT to IGT and T2DM (p <0.05 for trend). FGF-21 inversely correlated with Matsuda index & 1873-P of insulin sensitivity at baseline (r= -0.232, p=0.02) and at study end (r=- Estrogens Stimulate Hepatic Fibroblast Growth Factor-21 Produc- 0.402, p=0.005). Increase in FGF-21 predicts development of T2DM and the tion in Mice increase is attenuated by pioglitazone. These results suggest that the insu- CAMILLE ALLARD, BEIBEI XU, KENNETH S. KORACH, ELLIS R. LEVIN, FRANCK lin sensitizing effect of pioglitazone is, in part, mediated by the drug’s effect MAUVAIS-JARVIS, New Orleans, LA, Durham, NC, Long Beach, CA to reduce FGF-21 resistance. Estrogens are involved in maintaining energy homeostasis in women. After menopause, women loose this protection which increases the incidence of metabolic syndrome. Fibroblast Growth Factor-21 (FGF-21) has emerged as

For author disclosure information, see page A696. & Moderated Poster Discussion ADA-Supported Research

A482 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES an appealing therapeutic drug to treat metabolic syndrome. FGF-21 improves quantify UGE for 3 days [D]. Empaglifl ozin [EMPA 25mg] was given in AM metabolic dysfunction in rodents with properties reminiscent to those of D2 and D3. Fasting plasma glucose [FPG], mean 24 h glucose [MDG] and estrogens in ovariectomized (OVX) females. We investigated the effects UGE were determined. Fasting plasma insulin [FPI], C-peptide [C-pep] and of estrogens on FGF-21 production in mice. After fasting, the physiological glucagon [Gcg] prior to D1 and on D3 were measured. In T2DM, UGE on D1 increase in circulating FGF-21 was blunted in OVX female mice compared (10±2g/d) rose to 96±12 [D2] and 97±8 [D3]. In NonD, UGE on D1 (0.2±0.2) to sham-operated ones. 17beta-estradiol (E2) treatment increased hepatic increased to 48±5 [D2] and 52±7 [D3]. In T2DM, on D1 FPG was 188±7mg/ mRNA and circulating levels of FGF-21 in OVX female mice, and enhanced dl, MDG 184±11mg/dl. On D2, FPG (185±11) did not change, but MDG (168±8) molecular markers of FGF-21 action in peripheral tissues. E2-induced FGF-21 decreased (p<0.05). On D3, FPG fell to 167±8, MDG 166±8 (p<0.05). In NonD secretion was dependent upon estrogen receptor (ER) alpha activation as on D1 FPG was 99±3mg/dl, MDG 109±3mg/dl and did not change (FPG 97±3 this effect was reproduced by treatment with the ER alpha agonist PPT, and and 99±3; MDG 104±3 and 105±4), respectively D2 and D3. There were lost in ER alpha knock-out mice. Conversely, agonists for the ER beta and no changes in FPI (T2DM, 12±2 to 11±2µU/ml; NonD 8±2 to 8±2) or C-pep G-protein coupled ER (GPER) failed to induce FGF-21 production. ER alpha (T2DM, 3.1±0.1 to 3.3±0.1ng/ml; NonD 3.0±0.1 vs. 2.8±0.1). In contrast, Gcg action is mediated via extranuclear and nuclear pools of receptors. In female increased in T2DM from 88±6 to 109±9pg/ml and in NonD from 71±2 to mice expressing only the membrane pool of ER alpha, E2 failed to increase 79±3pg/ml) after EMPA (p<0.05). There was a strong positive correlation the circulating FGF-21 concentrations. In contrast, in female mice expressing (r=0.62, p<0.0001) between ΔUGE and ΔGcg in all subjects. only the nuclear pool of ER alpha, E2 was able to induce FGF-21 production Conclusion: Acute exposure to EMPA: (i) induced marked glucosuria in all suggesting that nuclear recruitment of ER alpha is needed to induce FGF-21 subjects, yet fasting and day-long glycemia improved modestly in T2DM and production. Interestingly, in isolated female hepatocytes, E2 does not induce did not change in NonD; (ii) Insulin and C-pep were unchanged, while gluca- the expression of the FGF-21 gene, suggesting that E2 stimulation of FGF-21 gon was elevated in both groups. These results suggest the presence of a production by hepatocytes in vivo could be mediated via extra hepatic action renal-hepatic axis in which glucosuria triggers a compensatory rise in EGP, of ER alpha. In conclusion, we demonstrate that E2 is able to stimulate FGF- which matches the UGE in NonD and opposes plasma glucose reduction in 21 secretion in an ER alpha-dependent manner in female mice. This effect T2DM. This “paradoxical” stimulation of EGP is mediated, in part, by gluca- seems to be mainly mediated by nuclear events. Studies using conditional ER gon and possibly by the sympathoadrenergic system. alpha knock-out mice are ongoing to determine the target tissue of ER alpha Supported By: Boehringer Ingelheim action that induces hepatic production of FGF-21. Supported By: American Diabetes Association (1-16-PDF-004 to C.A.); National 1876-P Institutes of Health (R01DK074970); National Institute of Environmental Health Postprandial Transorgan Bile Acid Kinetics: Implications for TGR5 Sciences (1ZIAES070065) Agonism HANNAH M. EGGINK, SAMUEL VAN NIEROP, MARIEKE G. SCHOONEMAN, & 1874-P ANITA BOELEN, ANDRIES KALSBEEK, MARTIJN KOEHORST, GABRIELLA A.M. Evaluation of the Effect of Intranasal Glucagon on Endogenous Glu- TEN HAVE, MAX NIEUWDORP, ALBERT K. GROEN, JOHANNES A. ROMIJN, cose Production NICOLAAS E.P. DEUTZ, MAARTEN R. SOETERS, Amsterdam, Netherlands, Gron- SATYA DASH, CHANGTING XIAO, KHAJAG KOULAJIAN, GARY F. LEWIS, Toronto, ingen, Netherlands, College Station, TX ON, Canada The bile acid (BA) receptor Takeda G protein-coupled receptor (TGR5) is Background: Glucagon is known to acutely stimulate endogenous glucose seen as an important regulator of energy metabolism affecting obesity and production (EGP). Recent studies in rodents showed a paradoxical, suppres- type 2 diabetes mellitus (DM2). To investigate the potential of BA to activate sive effect of hypothalamic glucagon on EGP. Here we investigated whether TGR5 in different organs we studied fasting and postprandial glucose, insu- preferential elevation of CNS glucagon has a similar suppressive effect on lin and BA levels in a porcine transorgan fl ux model. Pigs were fed a mixed EGP in humans by assessing the effect of intranasal glucagon (ING) on EGP. meal and blood was sampled repetitively from the aorta and portal, caval, Previous studies have demonstrated that the intranasal route preferentially renal and hepatic veins during 4 hours. BA were measured by LCMS. A TGR5 delivers small peptide hormones directly to the CNS. activation index was calculated with published TGR5 EC50 concentrations. Methods: In this single blind randomized crossover study, we assessed Portal and venous blood samples from 11 obese and 5 DM2 patients permit- EGP for 6 hours following 1mg of intranasal glucagon (ING) or intranasal pla- ted translation of the pig data to humans. cebo (INP) in 10 healthy, lean, normoglycemic men. To prevent fl uctuations in Porcine postprandial areas under the curve (AUC) for glucose and insulin pancreatic hormone and growth hormone secretion, a pancreatic clamp was were highest in the portal vein. Fasting portal total BA concentrations were administered along with deuterated glucose. In pilot studies ING transiently ~6 fold higher compared to other sampling sites. After the meal, glyco-chen- increased plasma glucagon concentration within 1 hour of administration. odeoxycholic acid (gCDCA) and glyco-hyodeoxycholic acid (gHDCA) were To mimic this systemic spillover, we administered intravenous glucagon (IVG) the most prominent portal BA (AUC 2501 vs. 2403 µmol·min/L respectively). for 30 minutes along with INP. Peripheral gHDCA levels were higher compared with gCDCA (portal and Results: Plasma concentrations of glucagon were matched in 6 out of peripheral gHDCA: gCDCA-ratio: 0.95 vs. 3.04; p < 0.001). Fasting BA fl ux in 10 participants (mean plasma glucagon during 1st hour: INP 83±13 vs. ING the porta was high and doubled after the meal. All forms of lithocholic acid 88±30 ng/ml, p=0.8). In these subjects EGP showed no suppression with ING (LCA), a strong TGR5 agonist, were well detected in the portal vein, but to a vs. INP over 6 hours (mean EGP INP 11.5±0.6 vs. ING 12.1±0.6 µmol/min/kg, minimal degree elsewhere. TGR5 activation did not increase after the meal, p=0.05). In 4 out of 10 subjects plasma glucagon was signifi cantly higher in but was higher in the portal compared to the peripheral vein (TGR5 index: the fi rst hour with ING. This was associated with a transient increase in EGP 6.57 vs. 0.64 respectively; p < 0.001). In human portal samples gCA, gDCA without an increase in glucose (mean EGP in 1st hour: INP 13.3±0.8 vs. ING and gCDCA were ~30 fold higher compared to venous samples. LCA forms 15.8±1 µmol/min/kg, p=0.04) but no difference in EGP thereafter (INP 9.7±0.5 were only detected in the portal vein. TGR5 activation was much higher in vs. ING 9.7±0.4 µmol/min/kg, p=1). Analysis of EGP in all 10 subjects showed the portal vein compared to the peripheral vein. no difference in EGP between 60 and 360 minutes (mean EGP: INP 9.9±0.4 In conclusion, there are large differences between portal and peripheral vs. ING 10.1±0.4 µmol/min/kg, p=0.5). BA concentrations pointing at a modest role for BA in TGR5 activation at Conclusion: Intranasal glucagon, administered at this dose, does not sup- peripheral sites. Whether TGR5 is a therapeutic target in obesity or DM2 Obesity

press EGP in humans. remains to be studied with specifi c TGR5 agonists in humans. POSTERS Supported By: Canadian Institutes of Health Research

1877-P Integrated Physiology/ 1875-P Mechanism by which Leptin Reverses Diabetic Ketoacidosis in Acute Glucosuric Response and Impact on Glycemic Indices of Rats Empaglifl ozin in Nondiabetic and Diabetic Subjects RACHEL J. PERRY, LIANG PENG, LYNN KENNEDY, YUICHI NOZAKI, GERALD I. HUSSEIN AL JOBORI, GIUSEPPE DANIELE, ROBERT MARTINEZ, JOHN ADAMS, SHULMAN, New Haven, CT CURTIS TRIPLITT, RALPH A. DEFRONZO, EUGENIO CERSOSIMO, San Antonio, TX Recent studies have demonstrated that leptin replacement therapy We examined the effect of SGLT2 inhibition on urine glucose excretion reverses diabetic ketoacidosis (DKA) in an insulin-independent fashion in (UGE), daily glucose profi le and hormonal responses in 15 MET-treated T2DM rodent models of type 1 diabetes; however the mechanism by which this (age~55 y, 3F/12M, BMI 31±1kg/m2, A1c 7.8±0.2% duration~8 y) and 16 age occurs is unknown. In order to address this question we examined the acute sex-matched nondiabetic subjects [NonD]. Continuous glucose monitoring effects of leptin replacement therapy on fasting rates of hepatic glucose was used to record daily glycemic indices and 24 h urine was collected to production and ketogenesis in awake streptozotocin-treated rats in DKA.

ADA-Supported Research & Moderated Poster Discussion For author disclosure information, see page A696.

A483 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

Following a 6 hr arterial infusion of leptin designed to match plasma lev- tion (- 21.7%). When a SSTR2-antagonist was infused, baseline glucagon els in normal rats, plasma glucose concentrations and beta-hydroxybutrate secretion was dramatically increased (+162.9%) and the inhibitory effect (BOHB) concentrations decreased by 250 mg/dL (P<0.0001) and 60% of GLP-1 on glucagon secretion was lost despite a preserved somatosta- (P<0.001) respectively. These reductions were matched by 50% reductions in tin response to GLP-1 (+114.2%). The GLP-1 infusion triggered a moderate rates of hepatic glucose production and BOHB turnover (P<0.05 and P<0.001, increase in insulin secretion regardless of the presence of SSTR2-antagonist respectively) but were totally abrogated by co-infusion of corticosterone. (+101.7% and +62.3% without or with SSTR2-antagonist, respectively). At Leptin replacement therapy had no effect in fed diabetic animals infused very low glucose levels (1.5 mmol/L) GLP-1 also decreased glucagon secre- with a low dose of insulin in order to maintain plasma insulin concentra- tion (-11.5%), but this coincided with an increase in somatostatin secretion tions ~10 uU/mL. In order to examine the molecular mechanism by which (+92.7%). SSTR2-antagonist addition dramatically increased baseline secre- leptin replacement therapy reversed ketogenesis we assessed rates of tion (+118.8%), and the suppressive effect of GLP-1 was eliminated. There 13 2 adipocyte lipolysis refl ected by [U- C]palmitate and [ H5]glycerol turnover. was no insulin secretion. All reported changes were statistically signifi cant. Leptin treatment was associated with more than 50% reductions in rates Our study shows that GLP-1 increases the secretion of somatostatin and of lipolysis and hepatic acetyl CoA (P<0.001). Surprisingly, despite marked concomitantly decreases the secretion of glucagon at all glucose levels. reductions in ketogenesis, hepatic malonyl CoA remained unchanged (~10 However, the basal somatostatin secretion and the somatostatin response nmol/g). Furthermore, a 6 hr infusion of acetate to match hepatic acetyl CoA to GLP-1 are strongly dependent on ambient glucose concentrations. Fur- concentrations in the DKA rats fully abrogated the acute effects of leptin thermore, the decreased glucagon secretion in response to GLP-1 infusion is to decrease rates of hepatic gluconeogenesis and ketogenesis. These data completely eliminated in the presence of the SSTR2-antagonist. Our results demonstrate that leptin’s insulin-independent effect to acutely reverse DKA clearly illustrate the powerful control of glucagon secretion exerted by the is dependent on its ability to inhibit lipolysis by reductions in plasma cor- glucose-dependent secretion of somatostatin from the delta cells of the ticosterone concentrations. Furthermore these effects occur by reductions pancreatic islets. in rates of fatty acid delivery to the liver resulting in reductions in hepatic Supported By: Novo Nordisk Foundation acetyl CoA content independent of changes in malonyl CoA. Supported By: National Institutes of Health (R01 DK-40936, R24 DK-85638, R01 1880-P AG-23686, P30 DK-45735, U24 DK-59635, T32 DK-101019, R01 DK-92606) Glucagon Promotes Colon Cancer Cell Proliferation by Modulation of AMPK and MAPK/ERK 1878-P TAKASHI YAGI, NAOTSUKA OKAYAMA, MAYO HACHIYA, SATOSHI YASUDA, Early Use of Plasma Adiponectin Helps Identify Responders to Pio- ASAMI HOTTA, SHUNSUKE ITO, HIDEOMI OGUCHI, KENRO IMAEDA, EIJI glitazone (PIO) Therapy in Nonalcoholic Steatohepatitis (NASH) KUBOTA, TAKASHI JOH, Nagoya, Japan FERNANDO BRIL, PAOLA PORTILLO SANCHEZ, CHRIS WISHERD, BRENDA Many recent papers have shown that diabetes patients are highly predis- MCDERMOTT, SEAN CONLEY, SRILAXMI KALAVALAPALLI, KENNETH CUSI, posed to cancer. In addition, increased cancer mortality has been reported Gainesville, FL, Salem, NH in patients with diabetes. We hypothesized that hyperglucagonemia, one Treatment with PIO is safe/effective in patients with NASH, but about of the characteristic features of type 2 diabetes mellitus, might promote 1/3 do not respond as expected. Identifying potential responders may help colorectal cancer development in diabetes patients. The role of glucagon in avoid unnecessary drug exposure. We aimed to determine the predictive colon cancer progression was investigated. value of total and high molecular weight (HMW) adiponectin in response Glucagon receptor (GCGR) expression in mouse colon cancer cell lines to long term PIO in patients with NASH. Patients with prediabetes/T2DM (CMT93 and CT26) was fi rst examined by Western blotting. To evaluate the (n=67) were treated with PIO 45mg/d for 18 months and had paired liver effect of glucagon on colon cancer cell proliferation, cell viability of CMT93 biopsies to assess histological response (40 from a RCT and 27 from the and CT26 cells after exposure to 1 µM glucagon was measured using Cell open-label phase). Total and HMW plasma adiponectin were measured at Counting Kit-8. Furthermore, the small hairpin RNA (shRNA)-mediated GCGR 0, 1, 2, 3, 6, 12, and 18 months by Chemiluminescent ELISA (ALPCO, Salem, knockdown CMT93 cell was generated to analyze the effects of glucagon. NH). After 18 months of PIO, total and HMW plasma adiponectin had a 2- To investigate the signal transduction pathways related to the effect of glu- and 3.3-fold increase, respectively (both p<0.001), with early peaks at 1 cagon on cell proliferation, the activation of AMP-activated protein kinase month. Of note, HMW adiponectin continued to rise gradually, reaching a (AMPK) and its downstream effectors was examined by Western blotting. plateau in the HMW/total adiponectin ratio at month 6. When patients were Additionally, cells were exposed to glucagon in combination with 5-amin- divided based on histological response (i.e., resolution of NASH), respond- oimidazole-4-carboxamide 1-β-D-ribofuranoside (AICAR), an AMPK activa- ers had a larger increase in total plasma adiponectin at 1 month (2587±1268 tor, or PD98059, a MAPK/ERK inhibitor. ng/ml, p=0.04), which was consistent with response at month 18 (1845±959 Glucagon signifi cantly enhanced the proliferation of CMT93 and CT26 ng/ml, p=0.03). In contrast, HMW adiponectin had larger increases later on, cells. However, glucagon had no effect on the proliferation of GCGR knock- at months 12 (p=0.08) and 18 (p=0.03). Differences were more pronounced down CMT93 cells. Glucagon suppressed the phosphorylation of AMPK when response was defi ned as a rather large reduction (ɗ3 points) in the and the subsequent MAPK/ERK activation in CMT93 cells, but not in GCGR NAFLD activity score. Of note, early changes of total adiponectin (at months knockdown CMT93 cells. AICAR inhibited the cell growth enhanced by glu- 1-3) strongly predicted long term histological response (AUROCs of ~0.80- cagon, and PD98059 negated the effect of glucagon on cell proliferation. 0.83). Based on changes in total adiponectin in the fi rst 3 months we pre- In conclusion, glucagon promotes cell proliferation of colon cancer cells dicted long term response to PIO with a PPV=90% and NPV=80%, which through inactivation of AMPK and the subsequent MAPK/ERK activation. improved to PPV=100% and NPV=86% using HMW adiponectin. The present data suggest that targeting the glucagon signaling pathway in Conclusion: Early increases in total and HMW adiponectin levels after PIO the treatment of colorectal cancer might be benefi cial in diabetic patients. help identify NASH patients likely to respond to therapy. The nature of this increase (extent, timing and value of total vs. HMW) that best predicts clini- 1881-P cal response requires further evaluation. (NCT00994682). Soy Protein Rescues Ovariectomized-induced Insulin Resistance Supported By: American Diabetes Association (1-08-CR-08 to K.C.); Burroughs via ERα Wellcome Fund; U.S. Department of Veterans Affairs (1 I01 CX000167-01) Obesity TERESE M. ZIDON, REBECCA J. WELLY, OLIVIA E. STRICKLIN, MICHELLE L.

POSTERS GASTECKI, MAKENZIE L. WOODFORD, YOUNG-MIN PARK, DENNIS B. LUBAHN, 1879-P JAUME PADILLA, VICTORIA J. VIEIRA-POTTER, Columbia, MO

Integrated Physiology/ Why Doesn’t GLP-1 Inhibit Glucagon Secretion during Hypo gly- Loss of ovarian hormone production (i.e., menopause) increases adipos- cemia? ity and insulin resistance (IR), signifi cantly increasing type 2 diabetes risk. ANNE ØRGAARD, JENS JUUL HOLST, Copenhagen, Denmark Soy protein extract high in phytoestrogens (SOY) improves postmenopausal In this project, we wanted to explore the hypothesis that GLP-1 does metabolic parameters, but the mechanisms are unknown. We previously not inhibit glucagon secretion during hypoglycemia because the inhibition demonstrated that SOY reduced adiposity, infl ammation, and IR in female depends on somatostatin secretion, which in turn is dependent on glucose low-fi t rats with and without ovarian function which prompted our interest in levels. the molecular mechanism. We aimed to test the hypothesis that the insulin The project was carried out by infusing GLP-1 in the absence or the sensitizing effect of SOY is through estrogen receptor alpha (ERα), the major presence of a SSTR2-antagonist in the isolated perfused mouse pancreas ER in adipose tissue, which is known to exert positive metabolic effects model. At normal glucose levels (6 mmol/L), GLP-1 clearly increased soma- of estrogen. We ovariectomized (OVX) adult female mice, null (i.e., KO) for tostatin secretion (+111.8%) and concomitantly decreased glucagon secre- either ERα or ERβ and their C57BL/6 wild-type (WT) controls with intact ERs

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A484 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

(N=7-12/group; age ~1 yr). One week following OVX, mice from each group increase in both SD rats and SHR (p<0.05). GLP-1 increased diuresis 3-fold were randomized to receive either SOY (600 ppm phytoestrogen) or control SD rats and 2-fold in SHR and increased Na+ excretion 4-fold in SD and 2-fold diet without SOY (<15ppm phytoestrogen, C) creating 6 groups: ERαKO/C, in SHR. BP was only increased by GLP-1 in SHR. Increasing concentrations ERαKO/SOY, ERβKO/C, ERβKO/SOY, WT/C, and WT/SOY which were fol- of GLP-1 (1 pM-1 µM) induced signifi cant vasodilation in isolated renal ves- lowed for 12 wks. Before OVX, ERαKO mice were ~9% fatter than WT (via sels from SD rats whereas no vasodilation was seen in vessels from SHR. EchoMRI™, p<0.001), but OVX did not further increase adiposity; also, no SOY- Immunohistochemical staining showed GLP-1 receptor positive staining in mediated improvement in glucose tolerance was observed (p=0.9). However, the renal vasculature in SD rats but not in SHR. OVX increased adiposity in all mice with ERα (i.e., WT and ERβKO) (p<0.001). It is concluded that SHR lack expression of renal GLP-1 receptors. The In ERβKO only (presumably with greatest ERα expression), SOY tended to renal effects found in SHR were abolished by the GLP-1 receptor antagonist mitigate this increase (+12.2 ± 1.8% (C) vs. +7.3 ± 1.7% (SOY); p=0.06). Impor- exendin 9-39. Thus, activation of extrarenal receptors mediate the changes tantly, SOY signifi cantly improved IR (p=0.02) in WT with both intact ERs, but found in BP, RBF, diuresis and Na+ excretion. the effect in ERβKO did not reach signifi cance (p=0.09). Thus, SOY-mediated IR reduction requires both ERα and ERβ, and that this effect is indepen- 1884-P dent of SOY-mediated adiposity reduction, which is mediated through ERα. Diabetic Enteropathy Recognizes a IGFBP3-mediated Colonic Stem- In conclusion, SOY-mediated IR reduction is mainly ERα-mediated, yet not Cell Dysfunction dependent on SOY-mediated improvements in body composition. Analyses FRANCESCA D’ADDIO, STEFANO LA ROSA, ANNA MAESTRONI, PETER JUNG, of tissue-specifi c mechanisms are currently underway. ELENA ORSENIGO, MOUFIDA BEN NASR, SARA TEZZA, ROBERTO BASSI, GIO- Supported By: National Institutes of Health VANNA FINZI, ALESSANDRO MARANDO, ANDREA VERGANI, ROBERTO FREGO, LUCA ALBARELLO, ANNAPAOLA ANDOLFO, ROBERTA MANUGUERRA, EDI 1882-P VIALE, CARLO STAUDACHER, DOMENICO CORRADI, EDUARD BATLLE, DAVID Brain Endothelial Cells Take Up Glucagon-like Peptide 1 (GLP-1) via BREAULT, ANTONIO SECCHI, FRANCO FOLLI, PAOLO FIORINA, Milan, Italy, Varese, GLP-1 Receptors and Protein Kinase A (PKA) Signaling Italy, Barcelona, Spain, Boston, MA, New York, NY, Monza, Italy, Parma, Italy, San ZHUO FU, SARAH M. GRAY, KEVIN AYLOR, EUGENE J. BARRETT, ZHENQI LIU, Antonio, TX Charlottesville, VA The role of circulating factors in regulating colonic stem cells (CoSCs) and GLP-1 in addition to regulating glucose-dependent insulin secretion exerts colonic epithelial homeostasis is unclear. Individuals with long-standing anorexic and neuroprotective effects. It is unclear whether these central type 1 diabetes (T1D) frequently have intestinal symptoms, termed dia- nervous system actions were secondary to peripheral or neuron-derived betic enteropathy (DE), though its etiology is unknown. Here, we report T1D GLP-1. The observations that brain expresses abundant GLP-1 receptors and patients with DE exhibit abnormalities in their intestinal mucosa and CoSCs, dipeptidyl-peptidase-4) inhibitors, which do not cross the blood brain barrier, which fail to generate in vitro mini-guts. Proteomic profi ling of T1D+DE could improve brain function suggest that peripherally derived GLP-1 may patient serum revealed altered levels of insulin-like growth factor 1 (IGF- play an important role in regulating brain function. 1) and its binding protein-3 (IGFBP3). IGFBP3 prevented in vitro growth of To examine whether brain endothelial cells could take up GLP-1 and if so patient-derived organoids via binding its receptor TMEM219, in an IGF-1- the underlying pathways, we incubated rat brain endothelial cells (RBEC) independent manner, and disrupted in vivo CoSC function in a preclinical with FAM-labeled GLP-1 at 10 pM, 100 pM, or 1 nM for 3 min in the pres- DE model. Restoration of normoglycemia in patients with long-standing T1D ence or absence of GLP-1 receptor antagonist exendin 9-39 or PKA inhibitor via kidney-pancreas transplantation or in diabetic mice by treatment with H89. RBEC fl uorescent density was determined using confocal microscopy to an ecto-TMEM219 recombinant protein normalized circulating IGF-1/IGFBP3 quantify the amount of FAM-GLP-1 that was taken up by RBEC. The presence levels and reestablished CoSC homeostasis. These fi ndings demonstrate of GLP-1 receptors in different regions of the brain and cultured RBECs was that peripheral IGF-1/IGFBP3 control CoSCs and their dysfunction in DE. examined using Western blotting and immunohistochemistry staining. Supported By: Italian Ministry of Health GLP-1 receptors were expressed abundantly in brain cortex, hippocampus, brain stem and cerebellum as well as in the cultured RBECs. RBEC promptly 1885-P took up FAM-GLP-1 with the cytosolic fl uorescne being the highest at FAM- Measuring Thermogenic Fat Metabolism in Humans and Mice Using GLP-1 concentration of 1 nM. Treatment of the cells for 30 min with exendin Mass Spectrometry-based Lipidomic Profi ling 9-39 or H89 reduced RBEC uptake of FAM-GLP-1 by ~ 80% (p<0.01) or ~ 90% MATTHEW D. LYNES, FARNAZ SHAMSI, MICHAEL KIEBISH, RUIDAN XUE, TIAN (p<0.01) respectively when the media FAM-GLP-1 concentration was at 10 or LIAN HUANG, TIM J. SCHULZ, AARON M. CYPESS, YU-HUA TSENG, Boston, MA, 100 pM. This inhibitory effect was less potent at higher FAM-GLP-1 concentra- Framingham, MA, Nuthetal, Germany tions (1 nM, ~50% inhibition for both exendin 9-39 and H89, p<0.01 for both). Brown adipose tissue (BAT) increases energy expenditure in response We conclude that brain endothelial cells effectively take up GLP-1 via to cold exposure to maintain body temperature. In humans, BAT activity is the GLP-1 receptors and the PKA signaling pathway. Thus, peripheral GLP-1 inversely correlated to body mass index, prompting speculation that ther- could enter the brain parenchyma via transendothelial transport to exert its apies designed to enhance BAT activity could be used to combat obesity. anorexic and neuroprotective effects. Cold exposure is an effective way to activate BAT, and, once activated, can Supported By: American Diabetes Association (1-16-PDF-037 to Z.F.); National robustly regulate fatty acid and glucose homeostasis. Little is known about Institutes of Health the specifi c lipid metabolic signature in response to cold challenge. We report here a comprehensive analysis using a highly sensitive mass spectrometry 1883-P approach to measure changes in the circulating lipid composition of human GLP-1 Increases Renal Blood Flow and Sodium Excretion via Acti- subjects before and after a 1-hour cold challenge. These data, supported by vation of Extrarenal GLP-1 Receptors in Spontaneously Hyper- mouse models of cold exposure, demonstrated that cold exposure was corre- tensive Rats lated with changes in phosphatidylglycerol metabolism. To determine if dif- ELISA P. JENSEN, JONAS RONN, JENS JUUL HOLST, CHARLOTTE M. SORENSEN, ferences in serum lipidomics were associated with changes in thermogeni- Copenhagen, Denmark cally active adipose tissue, we profi led both BAT and white adipose tissue

Glucagon-like peptide-1 (GLP-1) has a range of extrapancreatic effects (WAT) from mice housed in either cold or thermoneutral conditions. We also Obesity which includes renal effects. The effects of GLP-1 vary according to the ani- analyze the lipidomics of WAT in a mouse model of enhanced thermogenesis POSTERS mal model employed, and normo- and hypertensive animals may react dif- in WAT. Our data show that the Cardiolipin synthesis and remodeling path-

ferently to GLP-1. We investigated changes in renal blood fl ow (RBF), blood ways are activated in adipose tissue with enhanced thermogenic capacity, Integrated Physiology/ pressure (BP), diuresis and Na+ excretion in normotensive Sprague Dawley which may contribute to changes in circulating phosphatidylglycerol, the (SD) rats and spontaneously hypertensive rats (SHR) during administration of major substrate for Cardiolipin synthesis. Cardiolipin Synthase (CRLS1) was GLP-1 (1 nM) or GLP-1 combined with the GLP-1 receptor antagonist, exendin expressed at higher levels in mouse and human BAT compared to WAT, and 9-39 (100 nM) via a catheter directly into the renal artery. The effect of GLP-1 in mice, CRLS1 was further increased by cold exposure. To determine the on pre-contracted renal vessels from SD rats and SHR was investigated in a role of CRLS1 in human adipose tissue, we have generated several human wire-myograph. Renal tissue from SD rats and SHR was immunohistochemi- preadipocyte cell lines with decreased CRLS1 expression and characterized cal stained with a thoroughly validated GLP-1 receptor specifi c antibody. them in vitro. These data provide a lipid bio-signature of thermogenesis and GLP-1 infusion (20 min) in SD and SHR rats increased RBF from suggest new pathways regulated by cold. 9.1±0.5 ml/min to 10.1±0.5 ml/min in SD rats (p<0.01) and 4.6±0.2 ml/min Supported By: American Diabetes Association (7-12-BS-191 to M.D.L.); National to 5.3 ml/min±0.3 ml/min in SHR (p<0.01) and exendin 9-39 reduced the Institutes of Health

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A485 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1886-P Figure. GLP-1 Increases Amylase and Lipase as a Consequence of Prolif- GLP-1 Action Attenuates Breast Cancer Growth and Progression eration. SHIHO KAKO-KOMATSU, TAKASHI NOMIYAMA, CHIKAYO IWAYA-OTA, YASU- NARI YOSHINAGA, SHINICHI YAMASHITA, TAKAKO KAWANAMI, YURIKO HAMA- GUCHI, TOMOKO TANAKA, AKINORI IWASAKI, TOSHIHIKO YANASE, Fuku oka, Japan Incretin therapy has emerged as one of the most popular treatment for type 2 diabetes. GLP-1R agonist, Exendin-4 (Ex-4), has received much atten- tion, because of its’ tissue protective effects beyond glycemic control. We have previously reported vascular protective effect (Diabetes 2010, BBRC 2011) and anti-prostate cancer effect (Diabetes 2014, PLOS ONE 2015) of Ex-4. On the other hand, breast cancer is one of the most popular cancers in female with patients with type 2 diabetes and obesity. Then, we next examined whether GLP-1 action could attenuate breast cancer in the pres- ent study. First, we observed abundant GLP-1R expression in early stage breast cancer tissue extracted from patients without diabetes, but not advanced breast cancer. In human breast cancer cell lines, MCF-7, MDA-MB-231 and KPL-1 cell, GLP-1R was expressed, and 0.1~10nM Ex-4 signifi cantly decreased breast cell number of breast cancer cells, in dose-dependent Supported By: Novo Nordisk Foundation; Augustinus Foundation; Aase and manner. Although Ex-4 did not induce apoptosis in breast cancer cells, Ejnar Danielsen Foundation; European Foundation for the Study of Diabetes; Euro- BrdU assay revealed that Ex-4 attenuates cell proliferation of breast can- pean Molecular Biology Organization; Faculty of Health and Medical Sciences cer cells dose-dependently. If we transplanted MCF-7 cells into nondiabetic nude mice subcutaneously and treated them with 300pM/kg/day Ex-4 for 6 1888-P weeks, we observed decreased tumor size of MCF-7 in Ex-4-treated mice, in The Effect of Diabetes-associated Genetic Variation in TCF7L2 on both male and female mice. Immunohistochemistry with Ki67, a marker of Hepatic Vein Glucagon Pulses during Fasting and in Response to cell proliferation, revealed that breast cancer cell proliferation was signifi - Intravenous Glucose Infusion cantly reduced in tumor extracted mice treated with Ex-4. Further, 60% high RON T. VARGHESE, JAMES C. ANDREWS, ANU SHARMA, CHIARA DALLAMAN, fat diet increased breast cancer growth in nude mice, and Ex-4 treatments ROBERT A. RIZZA, CLAUDIO COBELLI, ALEKSEY MATVEYENKO, ADRIAN VELLA, decreased tumor size signifi cantly to control diet level. Rochester, MN, Padova, Italy These data suggest that GLP-1 could attenuate breast cancer via inhibi- The diabetes-associated allele in TCF7L2 increases the rate of conversion tion of breast cancer cell proliferation. to diabetes, in part by impairing insulin secretion. We have recently dem- Supported By: Suzuken Memorial Foundation onstrated that postprandial glucagon suppression is impaired in subjects homozygous for the diabetes-associated allele (TT) at rs7903146 in this locus. 1887-P As part of a study examining whether TCF7L2 directly alters the frequency Glucagon-like Peptide-1 Increases Pancreatic Enzymes as a Conse- or mass of insulin pulses, we also measured the frequency and amplitude quence of Proliferation of Exocrine Acinar Cells of glucagon pulses in the hepatic vein. We did so in part because of recent NICOLAI J. WEWER ALBRECHTSEN, REIDAR ALBRECHTEN, LASSE BREMHOLM, data suggesting a reciprocal relationship between insulin and glucagon BERIT SVENDSEN, STEEN S. POULSEN, ELISA POULSEN, RUNE E. KUHRE, CHAR- pulses in subjects with normal, as opposed to impaired glucose tolerance LOTTE JANUS, LINDA HILSTED, CAROLYN DEACON, BOLETTE HARTMANN, (NGT vs. IGT respectively). We studied 33 nondiabetic subjects of whom 16 JENS JUUL HOLST, Copenhagen, Denmark were homozygous for the diabetes-associated allele (TT) at rs7903146. The Therapy with GLP-1 receptor agonists (GLP-1RAs) has been associated remainder were homozygous for the protective allele (CC). Genotype groups with an increased risk of acute pancreatitis, and the plasma levels of the were otherwise matched for age, gender and fasting glucose. After an inpa- pancreatic enzymes amylase and lipase have been shown to increase dur- tient, overnight fast, a hepatic vein catheter was placed to allow sampling ing therapy. We investigated the acute and long-term effects of GLP-1RA on of hepatic vein blood at 2 minute intervals. During the latter phase of the amylase and lipase levels in humans and in pancreatic cell lines. Intact GLP-1, experiment glucose was maintained at 150mg/dL during a hyperglycemic its primary metabolite (9-36NH2) or exendin-4 (EX4) were injected into eight clamp. Splanchnic blood fl ow was measured using indocyanine green. Inter- healthy subjects, but none acutely increased amylase (100±354 min × U/L, pulse secretion i.e., basal secretion of glucagon did not differ during fasting P=0.65) or lipase activity (67±242 min × U/L, P=0.74) compared to placebo (18.1 ± 1.5 vs. 20.2 ± 1.7 ng/L/min, p = 0.37, CC vs. TT respectively). Similarly, injection. GLP-1RAs stimulated proliferation of the exocrine acinar cell line pulse interval (5.4 ± 0.3 vs. 5.7 ± 0.3 min, p = 0.53) and amplitude (11.2 ± 1.1 (95% CI: 1.7.2.5, P=0.001) through interaction with GLP-1R. GLP-1RA-induced vs. 11.7 ± 1.5 ng/L/min, p = 0.79) did not differ during fasting. Hyperglycemia proliferation was dependent on c-Src (P =0.001) and epidermal growth factor suppressed glucagon concentrations equally (e.g.: basal 11.3 ± 1.0 vs. 12.6 receptor (EGFR)-mediated signaling (P=0.02). Neither amylase (P=0.72) nor ± 1.2 ng/L/min, p = 0.44) in both groups without changing pulse frequency. lipase (P=0.54) levels were directly stimulated by GLP-1RAs; however, amy- This data suggests that the effects of the TT genotype (rs7903146 at the lase and lipase levels increased concomitantly with proliferation (P=.007). In TCF7L2 locus), on glucagon secretion are not mediated by hyperglycemia per summary GLP-1RAs do not directly increase amylase or lipase in humans or se and may be mediated by enteric or other signals which will require further in pancreatic acinar cell lines; however, GLP-1RAs stimulate proliferation via study to elucidate. a c-Src/EGFR-mediated mechanism. Therefore, elevated pancreas-related Supported By: National Institutes of Health infl ammatory markers in patients may refl ect proliferation of acinar cells (see Figure). 1889-P

Obesity The Association between Changes in Thyroid Function and Incident

POSTERS Type 2 Diabetes: 7-Year Longitudinal Study JI EUN JUN, SEUNG-EUN LEE, SO YOUNG PARK, SANG-MAN JIN, KYU YEON

Integrated Physiology/ HUR, MOON-KYU LEE, JAE HYEON KIM, Seoul, Republic of Korea Thyroid hormones are important regulators of glucose homeostasis. How- ever, the association between normal levels of thyroid hormones and type 2 diabetes remains unclear. The aim of study was to clarify the incidence of type 2 diabetes according to the baseline level of thyroid stimulating hor- mone (TSH), free thyroxine (FT4) and triiodothyronine (T3) as well as changes of each hormones in euthyroid subjects. Among the participants who con- secutively underwent thyroid function tests between 2006 and 2012 through a yearly health check-up program, 6,235 euthyroid subjects (3,619 men and 2,616 women) without diabetes were enrolled in the study. The change in each hormone was calculated by subtracting the level at the end of follow-

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A486 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES up or 1 year before diagnosis of diabetes from the baseline level. During p<0.05] mRNA expression were decreased. In addition, Ghrelin was infl u- 25,692 person-years of follow-up, there were 229 new cases of type 2 dia- enced by Oxytocin treatment, i.e., Ghrelin mRNA expression was increased betes. After full adjustment for potential confounders including HbA1c in Cox [maximum 2.3-fold, 2 hours, 100 nM, p<0.05]. By contrast, oxytocin does not proportional hazards models, individuals in the highest tertile of TSH change alter mRNA expression of adiponectin, ucp-1, and leptin. Taken together, our (2.5 to 4.2 µIU/mL) had a greater risk of incident type 2 diabetes (hazard ratio study represents evidence for hitherto unrecognized direct effects of the [HR] =1.72, 95% confi dence interval [CI]: 1.22-2.43, p for trend=0.002) in com- hypothalamic nonapeptide oxytocin on adipocyte biology. parison with individuals in the lowest tertile (-4.1 to -0.5 µIU/mL). Simultane- Supported By: Federal Armed Forces of Germany (07K3-S-200506) ously, the highest tertile of T3 change (16.3 to 104.7 ng/dL) and FT4 change (0.2 to 1.6 ng/dL) conferred protective effects against diabetes (HR=0.59, 1892-P 95% CI: 0.40-0.87, p for trend=0.005 and HR=0.24, 95% CI: 0.16-0.36, p for Dopamine Agonists Reverse Hyperglycemia in Streptozotocin- trend <0.001, respectively) compared to those in the lowest tertile (-76.5 to induced Diabetic Mice -1.8 ng/dL and -0.6 to 0.0 ng/dL, respectively). These associations remained SHUQIN LUO, MICHAEL EZROKHI, YANG LI, SCOTT WATTERS, ANTHONY H. CIN- signifi cant when each hormones were analyzed as continuous variables. COTTA, Tiverton, RI However, baseline levels or tertiles of TSH and thyroid hormones were not Bromocriptine-QR is a dopamine D2 agonist approved for treatment of associated with the risk of type 2 diabetes. These results indicated that type 2 diabetes (T2D). In animal models of T2D, circadian-timed bromo- even slight changes in TSH and thyroid hormones could have a signifi cant criptine (BC) treatment reduces both insulin and leptin resistance and ele- effect on the development of type 2 diabetes in euthyroid subjects. vated sympathetic tone to improve hyperglycemia. The present study inves- tigated whether dopamine agonist (DA) treatment may reduce hyperglycemia 1890-P associated with type 1 diabetes resulting from insulin and leptin defi ciency The Potentiating Effect of GLP-1 on Glucose-induced Glucagon Sup- subsequent to streptozotocin (STZ) administration in mice. Male C57BL/6J pression in Healthy Individuals and in Patients with Type 1 and 2 mice (10 wks old) were fed regular chow and housed on a daily 14 hr photo- Diabetes period and given a one-time STZ (120 mg/kg, i.p) (N=22) or vehicle (non-STZ) JONATAN I. BAGGER, ASGER LUND, MAGNUS GRØNDAHL, JENS J. HOLST, TINA (N=10) administration. Five days later fasting plasma glucose (FPG) levels VILSBØLL, FILIP K. KNOP, Hellerup, Denmark, Copenhagen, Denmark were elevated in STZ vs. non-STZ treated mice (471 ± 39 vs. 163 ± 5 mg/dl, Glucagon-like peptide-1 (GLP-1) has been shown to suppress glucagon in p<0.0001). Thereafter, STZ mice were divided into two groups (N=11/group) a glucose-dependent manner. The glucagon-suppressive potency of GLP-1 and administered either D1 plus D2 DA (SKF 38393, 20 mg/kg; BC, 10 mg/kg) has not been investigated in patients with diabetes. We examined the dose- (STZ-DA) or vehicle (STZ-V) while non-STZ mice were vehicle injected i.p. one response relationship of GLP-1 on glucose-induced glucagon suppression hour before the onset of dark daily for 10 days. Six days after the initiation of in individuals with normal glucose tolerance (NGT), patients with type 1 such intervention, the STZ-DA group had a 43% reduced FPG level vs. STZ-V diabetes (T1D) and patients with type 2 diabetes (T2D). Ten subjects with (259 ± 27 vs. 456 ± 49 mg/dl; p<0.0001) (non-STZ mice FPG: 161 ± 4 mg/dl). At NGT (age: 56±4 (mean±SEM) years; BMI: 26±3 kg/m2; fasting plasma glu- day 10 of treatment, relative to non-STZ treated mice, STZ-V mice exhibited cose (FPG): 5.4±0.2 mmol/L), 8 C-peptide negative patients with T1D (age: increased FPG by 2.9 fold (471 ± 39 vs. 163 ± 5 mg/dl; p<0.0001) and reduced 57±4 years; BMI: 29±1 kg/m2; FPG: 7.3±0.2 mmol/L) and 10 patients with plasma insulin by 48% (0.62 ± 0.18 vs. 1.2 ± 0.2 ng/ml; p<0.05) and plasma T2D (age 60±3 years; BMI: 31±1 kg/m2; FPG: 8.3±0.2 mmol/L) underwent 4 leptin by 64% (0.53 ± 0.19 vs. 1.48 ± 0.25 ng/ml; p<0.01). At this time point, stepwise glucose clamps (30-min steps at fasting plasma glucose and 6, 7, relative to STZ-V treatment, STZ-DA treatment reduced hyperglycemia by 8, 10, 12 and 15 mmo/L plasma glucose) with a blinded infusion of saline 52% (471 ± 39 vs. 228 ± 42 mg/dl; p<0.005) without altering levels of plasma or GLP-1 at 0.2, 0.4, or 0.8 pmol/kg/min (randomized). Plasma glucose and insulin (0.62 ± 0.18 vs. 0.74 ± 0.16 ng/ml) or leptin (0.53 ± 0.19 vs. 0.55 ± glucagon were plotted and the slope of the linear regression line (α) was 0.1 ng/ml). DA treatment improves hyperglycemia in leptin defi cient STZ- used to evaluate glucose-induced glucagon suppression and the infl uence of treated mice having reduced plasma insulin levels. Though undelineated, GLP-1. In NGT subjects, glucagon was signifi cantly (P<0.01) and progressively this DA effect may involve improved sensitivity to insulin and/or leptin as suppressed during the glucose clamp with saline (α: -0.35), and GLP-1 did not well as reductions in elevated sympathetic tone. infl uence glucagon suppression further regardless of dose (α: -0.31, -0.36, -0.23). In patients with T1D, only marginal, but signifi cant (P<0.01), changes 1893-P in glucagon were observed during the glucose clamp with saline (α: -0.12), Vitamin D Is Not Related to Insulin Resistance in African-American but not during the GLP-1 infusions (α: -0.06, 0.15, -0.01). In patients with T2D, and African Immigrant Women glucagon was progressively but equally suppressed (P<0.01) during saline (α: BRIANNA A. BINGHAM, PAOLA C. ALDANA, MICHELLE T. DUONG, LILIAN -0.40) and GLP-1 infusions (α: -0.40, -0.51, -0.57). We conclude that in NGT MABUNDO, MADIA RICKS, JAMES T. REYNOLDS, ANNE E. SUMNER, STEPHA- subjects and patients with T2D, intravenous glucose exerts robust inhibi- NIE T. CHUNG, Bethesda, MD tion of glucagon secretion, and that this inhibition is not further augmented The role of vitamin D (25(OH)D) in mediating insulin resistance is contro- by GLP-1. In contrast, patients with T1D were insensitive to the glucagon- versial in African-Americans (AA) and unknown in African Immigrant (AI) suppressive effects of both glucose and GLP-1. women. Since vitamin D is a fat soluble vitamin, low vitamin D concentra- tions may be related to sequestration in adipose tissue but not to decreased 1891-P insulin mediated glucose uptake. Our goal was to examine the relationship of Oxytocin Exhibits Proinfl ammatory Properties and Induces Ghrelin 25(OH)D with 3 measures of insulin resistance in 83 African descent women Expression via a Mitogen-activated Protein Kinase Signalling Path- living in the DC metro area who self-identifi ed as healthy (35% AA and 65% way in Brown Adipocytes AI; age 42±10 (mean±SD), range: 22-61 y; BMI 29±5, range: 21-41kg/m2). MALTE GRÜNDER, SEBASTIAN KÜCHLER, DIANA GRÜNDER, SÖREN WEST- Insulin resistance was measured in 3 ways: (1) insulin sensitivity index (SI) PHAL, Ulm, Germany via minimal modeling as our reference standard, (2) Matsuda Index during an There is growing evidence that oxytocin is involved in glucose metabo- OGTT and (3) HOMA-IR from fasting measures. Total 25(OH)D was measured lism and possesses antilipolytic properties. Moreover, obesity and insulin by immunoassay and percent (%) fat by DXA. Total 25(OH)D was 22±8ng/mL

resistance are considered a chronic subinfl ammatory state. Thus, circulat- and did not differ by ethnicity (AA: 21±9 vs. AI: 23±7 ng/mL, P=0.3). Percent Obesity ing concentrations of monocyte chemoattractant protein-1 (MCP-1) and fat was similar in AA and AIs (38±86 vs. 37±7%, P=0.5). African Immigrant POSTERS interleukin- 6 (IL-6) are elevated in individuals with increased body weight, women had lived in the U.S. for 14±9 y. In the entire cohort, 25(OH)D nega-

whereas adiponectin and ghrelin levels are decreased. We investigated the tively correlated with both BMI (-0.3, P=0.01) and percent fat (r=-0.2, P=0.04) Integrated Physiology/ effect of the nonapetide oxytocin on adipocyte signaling, adipokine gene but had no relationship with SI (r=0.2, P=0.2). Total 25(OH)D correlated expression, and thermogenesis in a murine model of SV40T-immortalized with Matsuda and HOMA-IR (both r=0.2, P=0.03) but this relationship did adipocytes. In fully differentiated adipocytes, oxytocin induced a dose- and not remain signifi cant after adjusting for % fat (P=0.1). This is the fi rst com- time-dependent acute phosphorylation of p44/ p42 MAP- Kinase [maximum parison of vitamin D in AA and AI women showing no difference in 25(OH)D 2-fold, 5 min, 100nM, p<0.01]. No effect could be observed on STAT3 phos- concentrations. Overall, 25(OH)D was not related to SI, a sensitive marker of phorylation. Chronic oxytocin, 100 nM, incubation showed no impairment whole body insulin resistance. The relationships of 25(OH)D with Matsuda of cell lipid accumulation, as proved by fat-specifi c staining. Oxytocin acti- and HOMA-IR were mediated by adiposity and may not refl ect decreased vated mRNA expression of IL-6 in a pronounced and acute manner [maximum insulin mediated glucose uptake. 2.2-fold, 2 hours, 100 nM, p<0.01] whereas MCP-1 [minimum 0.85- fold, 8 Supported By: National Institute of Diabetes and Digestive and Kidney Diseases hours, 100 nM, p<0.01] and TNFalpha [minimum 0.7-fold, 8 hours, 100 nM,

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A487 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1894-P Figure 1. CTX-1 Expressed as Baseline-subtracted Area Under Curve Famine from Feast: Hyperglycemia Induces Low Red Blood Cell (bsAUC) (Mean±SEM) During 75g-OGTT and IIGI in Patients with T2D and Vitamin C and Red Blood Cell Fragility CTRLs (A), 75g-OGTT and IIGI in Totally Pancreatectomized (PX) Patients and YU WANG, HONGBIN TU, MAHTAB NIYYATI, YAOHUI WANG, MARK LEVINE, CTRLS (B) and 50g-OGTT and IIGIs + GIP, GLP-1, GLP-2 and a Combination of Bethesda, MD the Three Hormones in Patients with T2D (C). New approaches are needed to treat microvascular angiopathy. Because red blood cells (RBCs) are less deformable in diabetes, we explored a novel concept linking decreased RBC deformability to RBC vitamin C (ascorbate) and hyperglycemia. When whole blood from ascorbate defi cient Gulo -/- mice (unable to synthesize vitamin C) was centrifuged to obtain plasma, RBCs hemolized. In ascorbate depleted Gulo -/- mice, RBCs displayed an inverse relationship between ascorbate concentrations and osmotic fragility, appeared as spherocytes, and had decreased β-spectrin without changes in other RBC membrane-associated proteins. Ascorbate oral resupplementa- tion for 3 days reversed changes in RBC osmotic fragility, RBC morphology, and β-spectrin. Only the oxidized form of ascorbate, dehydroascorbic acid, entered mouse and human RBCs via glucose transporters (GLUTs), and was 1896-P immediately reduced internally to ascorbate. In mouse RBCs, GLUT 3 and 4 Xenin-25 Is Produced by a Subset of Enterochromaffi n Cells were responsible transporters, while in human RBCs GLUT 1 was responsi- SONGYAN WANG, ISMAIL GREGORY, DAN CRIMMINS, BURTON M. WICE, St. ble. Dehydroascorbic acid entry into RBCs was blocked by the GLUT inhibitor Louis, MO cytochalasin B, and was progressively inhibited by glucose concentrations Enteroendocrine (EE) cells are a rare and complex population of hormone found in diabetes. Plasma glucose concentrations were inversely related producing intestinal epithelial cells that regulate gastrointestinal and whole to RBC ascorbate concentrations in fed and fasted AZIP diabetic mice and animal physiology. Numerous subtypes of EE cells have been defi ned based corresponding wild type controls. In 14 healthy subjects, 13 subjects with on the specifi c hormone (s) produced by individual cells. Our lab has shown mild diabetes (HgbA1C 5.7-7.7), and 13 subjects with poorly controlled dia- in human studies that IV infusion of a 25-amino acid neurotensin-related EE betes (HgbA1C ≥ 7.8), RBC ascorbate concentrations were inversely corre- cell product called xenin-25 (Xen) delays gastric emptying, decreases post- lated with plasma glucose concentrations and RBC osmotic fragility. RBC prandial glycemia, regulates GLP-1 release, increases cholinergic input to ascorbate concentrations and β-spectrin protein levels decreased as dia- islets, and amplifi es effects of GIP on insulin and glucagon secretion. How- betes worsened. Taken together, hyperglycemia in diabetes produced lower ever, an ELISA developed in our lab detects exogenously administered but RBC ascorbate levels with increased RBC rigidity, a novel candidate to drive not endogenously released Xen in plasma. Our objectives are to determine microvascular angiopathy. If RBC ascorbate concentrations are increased in if Xen is a physiologic peptide and to identify the subtype of EE cells that humans with diabetes, there is new possibility that microvascular disease produces this hormone. Xen antibodies were generated, affi nity-purifi ed and could be delayed. used for immunohistochemical studies with paraffi n-embedded sections of Supported By: National Institutes of Health mouse and human small intestine. Immunoreactive Xen (IR-Xen) did not stain GIP-producing EE cells as reported by others. Instead, IR-Xen was detected 1895-P only in serotonin-producing cells confi ned to the proximal small intestine. Gut Hormones, Rather than Glucose or Insulin, Are the Main Drivers Immunoreactivity was blocked by pre-incubating antibodies with intact Xen, of Diminished Bone Resorption in the Postabsorptive State but not Xen(6-25). IR-Xen was detected in a serotonin-producing EE cell line. ASGER LUND, JONATAN I. BAGGER, MIKKEL CHRISTENSEN, MORTEN FROST, Within individual cells, IR-Xen and serotonin were detected in the same NIKLAS R. JOERGENSEN, JAN H. STORKHOLM, CARSTEN P. HANSEN, JENS J. and also distinct granules. In conclusion, Xen is produced by a subset of HOLST, TINA VILSBØLL, FILIP K. KNOP, Hellerup, Denmark, Odense, Denmark, Copen- enterochromaffi n cells. Because serotonin can be released into the intesti- hagen, Denmark nal lumen as well as the blood, endogenous Xen may also be secreted into The mechanisms behind diminished bone resorption after ingestion of the gut lumen. Alternatively, a modifi cation near its N-terminus may alter nutrients are uncertain, but may involve increments in plasma glucose, insu- its activity and prevent detection of endogenous Xen by our ELISA. These lin and/or the gut hormones glucose-dependent insulinotropic polypeptide results suggest there may be additional and novel functions for endoge- (GIP), glucagon-like peptide-1 (GLP-1) and glucagon-like peptide-2 (GLP-2). nously produced Xen. Furthermore, cell lines that produce Xen can be used We measured responses of the bone resorption marker C-terminal telo- to elucidate pathways that regulate production, processing, and release of peptide of type I collagen (CTX-I) during 1.) 75g-oral glucose tolerance test this important intestinal peptide. (OGTT) and isoglycemic intravenous (iv) glucose infusions (IIGI) in 10 patients with type 2 diabetes (T2D) and 10 nondiabetic control subjects (CTRLs); 2.) 1897-P 75g-OGTT and IIGI in 10 totally pancreatectomized (PX) patients (devoid of Palmitate Disrupts Circadian Synthesis and Secretion of Glucagon- endogenous insulin production) and 10 CTRLs and; 3.) 50g-OGTT and IIGI and like Peptide-1 from the Intestinal L Cell four additional IIGIs with iv infusion of physiological doses of GIP, GLP-1, ALEXANDRE MARTCHENKO, HYUN SEO ROBIN OH, PATRICIA L. BRUBAKER, GLP-2 and a combination of the three in 10 patients with T2D. We show that Toronto, ON, Canada OGTT induces robust reduction in CTX-I in both patients with T2D and CTRLs Glucagon-like peptide-1 (GLP-1) is an incretin hormone released by the as opposed to a marginal reduction during IIGI (Figure 1A). OGTT-induced intestinal L cell in response to nutrient intake. Recent studies in our labora- reduction in CTX-I was also present in PX patients during OGTT (Figure 1B). tory have identifi ed a circadian pattern of GLP-1 secretion in response to Lastly, combined infusion of GIP, GLP-1 and GLP-2 during IIGI, to some extent known secretagogues. However, chronic ingestion of a high-fat high-sucrose mimics the suppression of CTX-I observed during OGTT (Figure 1C). These diet, a large component of which is the saturated fatty acid, palmitate, was results indicate that gut hormones, rather than glucose or insulin, are main found to disrupt the circadian release of GLP-1 in rats. Furthermore, in

Obesity drivers of diminished bone resorption in the postabsorptive state. mGLUTag cells, an established model of the L cell, palmitate pre-treatment

POSTERS for 12 hr reduces GLP-1 secretion and cell content at 4 hr after synchroniza- tion, but does not alter mRNA levels for the GLP-1 precursor, proglucagon -

Integrated Physiology/ these changes occur in the absence of alterations in cell viability. We thus hypothesized that palmitate induces endoplasmic reticulum (ER) stress in the L cell, resulting in decreased levels of translation. Pre-incubation of mGLUTag cells with palmitate for 12 hr followed by synchronization and additional treatment for 4 and 16 hr did not increase the Bax/Bcl2 ratio, indicating no increase in apoptosis; preliminary data also showed no increase in reac- tive oxygen species. However, after both 4 and 16 hr, there was an increase in CHOP mRNA levels (p<0.05-0.01 resp.) indicating ER stress, and cellular GLP-1 content was reduced at both time points (p<0.05-0.01). GLP-1 content in the media was also reduced at 12-24 hr by palmitate treatment (p<0.05). Furthermore, total protein content in the mGLUTag cells was reduced at

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A488 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

4-24 hr by palmitate treatment (p<0.05). In combination with these data, 1900-P palmitate treatment for 12-24 hr also resulted in a decreased ability of The Effect of Mitochondrial Insuffi ciency in Endothelial Cell on mGLUTag cells to respond to a known GLP-1 secretagogue, glucose-depen- Systemic Glucose Homeostasis and Compensation for Metabolic dent insulinotropic peptide (p<0.05). In conclusion, our data suggest that Stress chronic exposure to the saturated fatty acid, palmitate, reduces overall pro- HYUN JIN KIM, SANG HYEON JU, JI MIN KIM, YEA EUM KANG, KYONG HYE tein content within the mGLUTag cells by inhibiting translation through ER JOUNG, BO RAM YOU, BON JEONG KU, Deajeon, Republic of Korea stress, leading to a specifi c decrease in GLP-1 production. Mitochondria of endothelial cell (EC) are believed to not act as an energy Supported By: Canadian Diabetes Association factory but sense the local environment and regulate the homeostasis. Endothelial dysfunction in diabetes and cardiovascular disease has been 1898-P studied mainly as possible mechanism for chronic complication. The purpose Regulation of Glucagon-like Peptide-1 Exocytosis by the SNARE of this study was to investigate the effect of mitochondrial insuffi ciency Protein Syntaxin1a in EC on systemic glucose homeostasis and compensation for metabolic SARAH E. WHEELER, HOLLY M. STACEY, HERBERT Y. GAISANO, PATRICIA L. stress.We examined the in vivo phenotypes of endothelial dysfunction in BRUBAKER, Toronto, ON, Canada EC-specifi c Crif1-insuffi cient mice (Crif1 EC+/- mice) under conditions of meta- Glucagon-like peptide-1 (GLP-1) is an intestinal L cell hormone that potenti- bolic stress induced by a HFD (High fat diet). CRIF1 (CR6-interacting factor-1) ates insulin secretion in a glucose-dependent manner. As such, GLP-1-based is a mitochondrial protein essential for the synthesis and formation of the mimetics are now widely used to treat hyperglycemia in patients with type 2 OxPhos complex in the inner mitochondrial membrane. IPGTT and ITT were diabetes. In attempts to develop novel therapeutic approaches for the treat- performed and serum concentrations of FGF (Fibroblast growth factor) 21, ment of type 2 diabetes, the capacity to enhance endogenous GLP-1 secretion GDF (Growth differentiation factor) 15, sICAM (soluble cell adhesion mol- from the L cell in a regulated fashion is currently being explored. Although ecule)-1 and lipid profi le were measured after a HFD for 22 weeks from 7 multiple secretagogues able to stimulate GLP-1 secretion have been identi- weeks of age on WT and Crif1 EC+/- mice.IPGTT and ITT showed Crif1 EC+/- mice fi ed, the mechanism of exocytosis remains unknown. We hypothesized that were more resistant to a HFD compared to WT mice, but not signifi cant the core SNARE protein, Syntaxin1a, is required for GLP-1 exocytosis from the statistically. There was no difference in serum lipid profi le, sICAM and primary adult murine intestinal L cell. We thus generated an intestinal epithe- HOMA-IR between WT and Crif1 EC+/- mice. Serum concentration of FGF-21 lial-specifi c inducible Syntaxin1a knockout mouse model (Syn1aKO), by cross- and GDF15 increased after a HFD compared to NCD, however, were lower ing Syntaxin1afl ox/fl ox mice with villin-creERT2 mice. Syn1aKO mice, in addition in Crif1 EC+/-mice than WT mice. Although there was no difference of body to age- and sex-matched Syntaxin1afl ox/fl ox, villin-creERT2, and Syntaxin1afl o x / - ; weight between WT and Crif1 EC+/- mice after a HFD, weights (g/100g) of liver villin-creERT2 controls were all treated with and without tamoxifen. Syn1aKO and subcutaneous WAT were lower in Crif1 EC+/- mice. Our results suggest and control mice were then challenged in vivo with an oral glucose tolerance that mitochondrial insuffi ciency in endothelial cell could trigger the protec- test, followed by determination of circulating levels of total plasma GLP-1. tive effect to metabolic stress through limiting ectopic lipid accumulation. Syn1aKO mice demonstrated elevated blood glucose levels 60 mins post glu- Further research is necessary to explore the mechanism and cause and cose administration in comparison to controls, 24.0 ± 1.9 vs. 16.0 ± 1.0 mM effect relationship. (P < 0.001), respectively. Accordingly, the area under the 2-hour blood glucose Supported By: National Research Foundation of Korea curve for Syn1aKO mice, 2162 ± 126 mM-min, was signifi cantly elevated com- pared to controls, 1783 ± 80 mM-min (P < 0.01). The glucose-intolerant state 1901-P of Syn1aKO mice was associated with a 17.2% reduction in basal (P < 0.05) Circadian Dopamine Activity at the Biological Clock Regulates GLP-1 levels. At t = 10 mins post-glucose, the increment in GLP-1 was even Insulin Sensitivity via the Hypothalamus further reduced (by 23.3%, P < 0.05), indicating impaired GLP-1 secretion in SHUQIN LUO, MICHAEL EZROKHI, YANG LI, ANTHONY H. CINCOTTA, Tiverton, RI response to glucose in Syn1aKO mice. In conclusion, therefore, Syntaxin1a The circadian peak in dopaminergic activity (DA) at the biological clock, plays a vital role in mediating the exocytosis of GLP-1. suprachiasmatic nuclei (SCN) is attenuated coincident with increases in nor- Supported By: Natural Sciences and Engineering Research Council of Canada epinephrine (NE) activity at the ventromedial hypothalamus (VMH) and para- ventricular nuclei (PVN) in insulin resistant (IR) states. Also, administration of 1899-P dopamine (D) to the SCN at the time of day it peaks there in healthy animals Glicentin and Oxyntomodulin Are Associated with Abnormal Glu- reverses IR while chronic administration of NE to the VMH or PVN induces cose Metabolism Secondary to Acute Pancreatitis IR. Whether D administration to the SCN area to restore insulin sensitivity SAYALI PENDHARKAR, VARSHA ASRANI, RINKI MURPHY, JOHN WINDSOR, (IS) is circadian-time dependent (CTD) and whether this effect can potentiate MAX PETROV, Auckland, New Zealand a reduction in VMH and/or PVN NE activity is unknown. This study investi- Pancreas, a dual gland, plays a key role in the development of diabetes gated 1.) the CTD impact of D administered locally at the SCN area (either and pancreatitis. Newly developed abnormal glucose metabolism (AGM) at a time to restore the natural circadian peak DA or at another time of day) after acute pancreatitis (AP), is present in nearly 40% of patients. Pancre- upon IR in spontaneously hypertensive rats (SHR) fed a high fat diet and 2.) atic diabetes is characterized by impaired insulin secretion and/or insulin whether such treatments had any impact to reduce elevated NE activity at resistance. To date, only GLP-1, GLP-2 and GIP have been studied and only the VMH or PVN. Male SHRs (12 wks old) were fed a 60% saturated fat diet in patients with diabetes secondary to chronic pancreatitis or pancreatic for 4 weeks, divided into 4 groups (N=10/group) and then treated daily at the cancer. Whether this is true for diabetes after AP is unknown. The aim of SCN either with D (2 nm in 0.2 µl artifi cial CSF) or vehicle (V) at 13 hours after the study was to investigate, for the fi rst time, the associations between a the onset of light (HALO) (natural circadian DA peak at SCN) or with the same comprehensive panel of gut hormones (glicentin, oxyntomodulin, PYY, GLP- V and D infusions at 19 HALO. Glucose tolerance test (GTT) was performed 1, GLP-2, and GIP) and glucose metabolism after AP. Eighty-three patients after 2 wks of treatment at 48 hrs prior to sacrifi ce for HPLC analysis of VMH with normoglycemia before AP were followed-up cross-sectionally. Fast- and PVN NE and 4-hydroxy-3-methoxyphenylglycol (HMPG) (NE metabolite). ing venous blood was collected for analysis of gut hormones, glucose, and Relative to V, D treatment at 13 HALO, but not 19 HALO, reduced GTT glu- HbA1c. Modifi ed Poisson regression and Spearman’s correlation were used cose AUC (24%, p<0.0001) and insulin AUC (36%, p<0.05) and increased IS

for statistical analyses. Age, sex, ethnicity, BMI, smoking, physical activity, (2.2-fold; Matsuda Method; p<0.05). D treatment vs. V at 13 HALO, but not Obesity recurrence, duration from fi rst attack, severity, and aetiology were adjusted at 19 HALO, reduced HMPG and HMPG x NE product in both VMH (by 46% POSTERS for. Decreased glicentin with a prevalence ratio (PR) of 0.26 (p-trend < 0.001), and 47% respectively, p<0.05) and PVN (33% and 43% respectively, p<0.05).

and oxyntomodulin with a PR of 0.60 (p-trend < 0.001) were signifi cantly CTD SCN D administration timed to restore the natural circadian peak in DA Integrated Physiology/ associated with AGM after AP in the unadjusted and/or adjusted analyses. at SCN, but not outside this time, ameliorates IR and reduces brain PVN and GLP-1, GLP-2, GIP, and PYY were not found to be statistically signifi cant. VMH NE activities that potentiate IR. Strong positive correlations were observed between pancreatic polypep- tide (PP) and PYY ('p = 0.64; p < 0.001), glucagon and PYY ('p = 0.62; p < 0.001), and PP and GIP (p ' = 0.59; p < 0.001). Oxyntomodulin and PP, and glucagon and oxyntomodulin were found to have signifi cant negative correlations (p = 0.004, and p = 0.026, respectively). Decreased glicentin and oxyntomodu- lin levels are characteristic of AGM secondary to AP, indicating desensitized β-cell receptors and consequent hyperglycaemia. Lack of grossly elevated PYY levels suggest insulin secretion may not be impaired after AP.

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A489 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1902-P 1904-P The Cardiac Hormone Atrial Natriuretic Peptide (ANP) Reduces Treatment with the Glucagon-like Peptide-1 Receptor Agonist Lira- Leptin Secretion in Healthy Men glutide Improves Glycemic Control in Women with Prior Gesta tional JENS JORDAN, PAMELA FISCHER-POSOVSZKY, JULIA REINKE, MARTIN M. Diabetes Mellitus: A Randomized, Placebo-Controlled Trial DANIELS, CEDRIC MORO, MARTIN WABITSCH, STEFAN ENGELI, ANDREAS L. SIGNE FOGHSGAARD, LOUISE VEDTOFTE, EMILIE BAHNE, CAMILLA ANDREA- BIRKENFELD, Hannover, Germany, Ulm, Germany, Berlin, Germany, Toulouse, France, SEN, ELISABETH R. MATHIESEN, JENS A. SVARE, LISELOTTE K. CHRISTIANSEN, Dresden, Germany JENS J. HOLST, TINE D. CLAUSEN, PETER DAMM, FILIP K. KNOP, TINA VILSBØLL, Natriuretic peptides (NP), which are released during physical exercise, Copenhagen, Denmark, Herlev, Denmark, Hillerød, Denmark promote lipid mobilization from adipose tissue and lipid oxidative capacity Women with prior gestational diabetes mellitus (GDM) are at high risk of thereby protecting from insulin resistance. In addition to these direct actions developing type 2 diabetes. For that reason, the group constitutes an oppor- on energy balance and substrate utilization, observational studies suggest tunity for studying the early stages of the disease as well as therapeutic that high NP levels are associated with reduced circulating leptin concentra- modulation of a high-risk condition. Overweight, nondiabetic women with tions. Whether or not this is a causal relationship is currently unknown. We prior GDM were randomized to liraglutide 1.8 mg subcutaneously once-daily tested the hypothesis that atrial natriuretic peptide (ANP) acutely decreases for one year (N=49; age: 38±5 (mean±SD) years; BMI: 32±6 kg/m2; fasting leptin levels in 12 healthy men. We infused ANP intravenously over 135 plasma glucose (FPG): 5.5±0.5 mmol/L; HbA1c: 33±4 mmol/mol (5.2±0.4%)) minutes while collecting venous blood and adipose tissue microdialysates or placebo (N=55; age: 38±5 years; BMI: 31±3 kg/m2; FPG: 5.4±0.4 mmol/L; at baseline and at the end of ANP-infusion. We obtained blood samples at HbA1c: 32±4 mmol/mol (5.1±0.4%)). The women underwent a 4-hour 75g oral identical time-points without ANP infusion in 7 age and BMI matched men. glucose tolerance test (OGTT) at baseline and after one year (N=83). Change Moreover, we assessed ANP effects on leptin in human SGBS adipocytes in in glucose tolerance (total area under the curve (tAUC) for plasma glucose vitro. With infusion, venous ANP concentrations increased ~ 10 fold. Sys- excursions during the OGTT) was defi ned as the primary endpoint. At base- temic and adipose tissue glycerol concentrations increased 70% and 80%, line, age, BMI, HbA1c, FPG and tAUC were similar in the two groups. FPG at respectively (P<0.01). ANP infusion decreased systemic leptin levels from one year was signifi cantly reduced in the liraglutide group (-0.4±0.4 mmol/L 3.6±1 ng/ml to 3.3±1 ng/ml (P=0.01, 12% decrease). In contrast, leptin did not (mean±SD), P=0.0003), but not in the placebo group (-0.03±0.4 mmol/L, change in the control group (P<0.05 vs. infusion). ANP markedly reduced lep- P=0.73). FPG was signifi cantly lower in the liraglutide group at one year vs. tin release into the supernatant of SGBS adipocytes, with no change in lep- placebo (5.1±0.4 vs. 5.3±0.5 mmol/L, P=0.01). Glucose tolerance (tAUC) at tin mRNA expression. We conclude that the cardiac hormone ANP acutely one year was signifi cantly better in the liraglutide group compared to the decrease systemic leptin concentrations in healthy lean subjects likely by placebo group (1,477±217 vs. 1,691±253 mmol/L × min, P<0.0001). Accord- inhibiting leptin secretion. We speculate that this mechanism may serve to ingly, signifi cant reductions in HbA1c (-2.2±4.0 mmol/mol (-0.2±0.4%), P=0.02) replenish energy stores following exercise and may limit the utility of treat- and change in glucose tolerance (tAUC) (-218±190 mmol/L × min, P<0.0001) ments targeting the NP system in type 2 diabetes and obesity in terms of were observed in the liraglutide group, but not in the placebo group ((-0.6±6.6 weight loss. mmol/mol (0.1±0.3%), P=0.65; -47±192 mmol/L × min, P=0.97). One-year treatment with the glucagon-like peptide-1 receptor agonist liraglutide in 1903-P women with prior GDM improves FPG, HbA1c and glucose tolerance signifi - Glucagon-like Peptide 1 Accelerates Heart Rate by Nonvagal, Non- cantly compared to placebo. sympathetic Mechanisms Supported By: Danish Diabetes Academy; University of Odense SIMON VEEDFALD, SIGNE TORÄNG, SARA A. LARSEN, ELISA P. JENSEN, LONNIE G. PETERSEN, ANNE NISSEN, BOLETTE HARTMANN, LARS B. SVENDSEN, JENS 1905-P J. HOLST, Copenhagen, Denmark Deletion of 14-3-3ζ Improves Oral Glucose Tolerance through a Treatment with glucagon-like peptide 1 (GLP-1) receptor agonists is GLP-1-dependent Mechanism accompanied by an increase in heart rate (HR). In chloralose anaesthetized GARETH E. LIM, MICAH PISKE, ANGEL F. LOPEZ, HAYLEY S. RAMSHAW, JAMES pigs (~30kg), we investigated the role of abdominal vagal afferents (I) and D. JOHNSON, Vancouver, BC, Canada, Adelaide, Australia efferent vagal pathways (II). In propofol anaesthetized pigs we investigated Metabolic hormones require complex signaling networks to control glu- the role of sympathetic pathways (III). cose homeostasis, but how such networks are coordinated within a cell is (I) GLP-1 was infused IV (3 or 30pmol/kg/min, 1 h) where after the vagal poorly understood. Molecular scaffolds, such as 14-3-3 proteins, are likely trunks were severed (at the level of the cardia) and the GLP-1 infusion candidates due to their ability to regulate the function of insulin signaling repeated. (II) GLP-1 was infused IV (12.5pmol/kg/min, 1 h) followed by sec- effectors. We previously identifi ed critical metabolic functions of 14-3- tion of the cervical vagi (n=3) and the GLP-1 infusion repeated. (III) GLP-1 3ζ, such as promoting in vitro β-cell survival and controlling adipogenesis. was infused IV (15pmol/kg/min, 1 h) followed by an infusion of saline (n=3), However, the overall contribution of 14-3-3ζ to glucose homeostasis has propranolol (n=5), phentolamine (n=6), or both blockers (n=5) and the GLP-1 not been studied in depth. Using cultured β-cells, we fi rst determined that infusion repeated. Cardiac output (CO) was determined by the thermo-dilu- siRNA-mediated depletion of 14-3-3ζ had no effects glucose-stimulated or tion method. KCl-stimulated insulin secretion, despite increasing insulin biosynthesis. (I) High dose GLP-1 infusions increased HR: baseline vs. peak (126 (14) vs. When compared with wildtype controls, 14-3-3ζ knockout mice exhibited 193 (25) beats/min (mean (SD), P<0.001 (t-test)); low dose: NS. Abdominal increased β-cell area, which was associated with elevated fasting insulin truncal vagotomy did not attenuate the increase in HR (iAUC/60 min (Δbpm), levels. Administration of intraperitoneal insulin tolerance bolus (1.5 U/kg) Wilcoxon tests); high dose experiments: 55 (36-68) vs. 40 (22-57) Δbpm revealed signifi cantly reduced insulin sensitivity in 14-3-3ζ knockout ani- (median (range), P=0.31). (II) Cervical vagotomy increased baseline heart rate mals. Although no differences were observed following an intraperitoneal but GLP-1 subsequently tended to cause larger increases in HR 18 (14-27) vs. glucose bolus (2 g/kg), glucose tolerance was signifi cantly improved in 14-3- 22 (15-24) Δbpm (median (range), P=0.75). (III) Adrenergic blockers did not 3ζ knockout mice after an oral glucose gavage (2 g/kg). This improvement attenuate the GLP-1 induced increase in HR: propranolol 30 (18-39) vs. 18 in glucose tolerance was associated with a potentiated incretin response (12-31) Δbpm (median (range), P=0.13); phentolamine 32 (12-55) vs. 30 (10- and elevated circulating glucagon-like peptide-1 (GLP-1), and no effects on

Obesity 35) Δbpm, (median (range), P=0.22), combined block 30 (11-40) vs. 28 (11-33) gastric emptying were observed. The impact of 14-3-3ζ on L cell function

POSTERS Δbpm (median (range), P=0.19). CO increased during GLP-1 infusions; 2.9 (0.7) was confi rmed in vitro, as knockdown of 14-3-3ζ in GLUTag L cells enhanced to 4.1 (0.4) L/min (mean (SD), P<0.001 (t-test)). In all (III) pigs, atropine ele- GLP-1 synthesis and secretion. Systemic inhibition of the GLP-1 receptor

Integrated Physiology/ vated HR indicating maintained vagal tone. We conclude, that the increase with Ex4(9-39) attenuated the improvement in oral glucose tolerance in in HR occurring during pharmacological GLP-1 receptor agonism, is neither 14-3-3ζ knockout mice. When taken together, these fi ndings demonstrate due to inhibition of vagal tone nor to stimulation of sympathetic drive. novel roles of 14-3-3ζ in the regulation of glucose homeostasis. Modulating Supported By: Novo Nordisk Foundation; University of Copenhagen 14-3-3ζ levels in intestinal L cells, and other tissues, may represent a novel therapeutic approach for the treatment of diabetes and obesity. Supported By: JDRF; Canadian Diabetes Association

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A490 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1906-P stood. Therefore, in this study we examined the role of L-PGDS in glucose Cellular Mechanism of Sexual Dimorphism for High Fat Diet-induced regulation using L-PGDS knock-out (KO), L-PGDS knock-in (KI) and C57BL/6 Whole-Body Insulin Resistance (wild type) mice that underwent either sham (n=5/group) or VSG (n=5/group). JOAO-PAULO G. CAMPOREZ, MICHAEL J. JURCZAK, KITT F. PETERSEN, GERALD Mice were kept on diabetogenic diet and all parameters were measured ini- I. SHULMAN, New Haven, CT, Pittsburgh, PA tially and 10 weeks post either sham or VSG. Protocol was approved by ani- In order to address the mechanisms associated with sexual dimorphism mal care ethical committee. Data were analyzed by unpaired t-test or one- and whole-body glucose and fat metabolism, we studied male and female way ANOVA wherever appropriate with the signifi cance (p<0.05). Results mice fed a high fat diet for 3 weeks. Female mice displayed increased of our intraperitoneal glucose tolerance test showed signifi cant glycemic whole-body insulin sensitivity (70%) assessed by hyperinsulinemic-eugly- improvement ten weeks post VSG in both C57BL/6 and L-PGDS KI groups cemic clamps, which could be attributed to both increased hepatic (140%) compared to the sham group. However, the L-PGDS KO group developed and peripheral (34%) insulin sensitivity whereas there were no differences glucose intolerance similar to the sham group ten weeks post VSG. Clearly in energy expenditure. Female mice also displayed increased insulin-stimu- L-PGDS is a limiting factor for the benefi cial effects of VSG. Similarly, HOMA- lated muscle and adipose tissue 2-deoxy-glucose uptake (70% and 100%, IR (Homeostatic Model Assessment-Insulin Resistance) value for C57BL/6 respectively), along with 40-50% reduction in hepatic and muscle triglycer- mice after sham surgery was 88626±6848, which was signifi cantly reduced ide and diacylglycerol (DAG) content. This improved liver and muscle insulin to 36055±22520 in response to VSG. The HOMA-IR value of the L-PGDS KI responsiveness was associated with ~2 fold increase in insulin-stimulated group was 59356±10748 in the sham group and was signifi cantly reduced AKT2 phosphorylation and reduced hepatic PKCε and muscle PKCθ activa- to 20627±16297 after VSG. In contrast, the HOMA-IR value increased from tion in liver and muscle respectively. In contrast there were no changes in 31384±2021 to 49387±7108 when comparing sham to VSG, respectively. We liver or muscle ceramide content between groups. Female mice also display also measured weight but did not fi nd any signifi cant difference in any of increased insulin-stimulated suppression of lipolysis, measured by in vivo the groups which implies that all the benefi cial effects were independent 13 2 [U- C]palmitate and [ H5]glycerol turnover, along with increased lipid uptake of weight change. In summary, VSG only showed its benefi cial metabolic in white adipose tissue (55%) and liver (23%) and without any changes in effects in the presence of L-PGDS. Further studies need to be performed muscle lipid uptake. In order to evaluate the specifi c effect of estradiol (E2), at molecular level to determine the actual mechanism of action of L-PGDS we studied high fat-fed male mice treated with E2 or vehicle. E2-treated in VSG. mice displayed increased insulin sensitivity, accounted by both increased hepatic and peripheral insulin sensitivity associated with increased insulin- 1909-P stimulated lipolysis suppression and reduced hepatic and muscle TG and Glucagon-like Peptide-1 Increases Cardiac Output Acutely in DAGs, and decreased PKCε and PKCθ activation. Taken together, these data Healthy Males Mainly due to Vasodilation in The Skeletal Muscles support the hypothesis that female mice are protected from lipid-induced and Adipose Tissue liver and muscle insulin resistance due to sex hormone-mediated reductions ALI ASMAR, LENE SIMONSEN, MEENA ASMAR, STEN MADSBAD, JENS J. in ectopic lipid content and reduction in lipolysis, leading to reduced PKCε HOLST, CHARLOTTE M. SORENSEN, BOLETTE HARTMANN, JENS BÜLOW, and PKCθ activation in liver and muscle, respectively. Copenhagen, Denmark During acute GLP-1 administration in healthy males, we recently demon- 1907-P strated a signifi cant increase in cardiac output (CO) associated with vaso- GLP-1 Action Attenuates Prostate Cancer Growth and Progression dilation. Simultaneously, renal plasma fl ow remained unchanged, thus, TORU SHIGEOKA, TAKASHI NOMIYAMA, SHINICHIRO IRIE, TAKAKO KAWAN- we hypothesize that GLP-1 induces vasodilation in the splanchnic circula- AMI, YURIKO HAMAGUCHI, TOMOKO TANAKA, MASATOSHI TANAKA, TOSHI- tion, skeletal muscles, and/or adipose tissue. On fi xed sodium intake, eight HIKO YANASE, Fukuoka, Japan healthy men were examined on two different occasions in random order. Incretin therapy has emerged as one of the most popular treatment for During a 2-hour infusion of either GLP-1 (1.5 pmol kg-1 min-1) or saline, CO type 2 diabetes. GLP-1R agonist, Exendin-4 (Ex-4), has received much atten- was continuously estimated non-invasively concomitantly with intra-arte- tion, because of its tissue protective effects beyond glycemic control. We rial blood pressure and heart rate recordings. After catheterizations of a have previously reported anti-prostate cancer effect (Diabetes 2014, PLOS femoral artery and vein and a hepatic vein, respectively, leg and splanchnic ONE 2015) of Ex-4. In our previous report, Ex-4 did not attenuate cell prolif- blood fl ow were measured by Fick’s Principle (in conjunction with a constant eration in a prostate cancer cell ALVA-41, which does not express GLP-1R. In infusion of indocyanine green). Regional subcutaneous abdominal adipose the present study, we examined anti-prostate cancer effect of Ex-4 in ALVA- tissue blood fl ow (ATBF) was measured by the 133Xe clearance technique. 41 forced expressed GLP-1R using lentivirus vector (ALVA-GLP-1R) and asso- The subjects remained supine during the experiments. During GLP-1 infusion, ciation between GLP-1R expression level and prostate cancer progression mean arterial pressure increased by 6% (5 ± 1 mm Hg, p=0.138) and CO even in human prostate cancer tissues. Firstly, we confi rmed abundant GLP-1R more, by 18% (1.1 ± 0.1 L/min, p=0.019). Arterial pulse pressure increased by expression in ALVA-GLP-1R using immunohistochemistry. Ex-4 signifi cantly 9 ± 2 mm Hg (p=0.025) and heart rate by 10 ± 1 bpm (p=0.005). Splanchnic decreased the proliferation of ALVA-GLP-1R in a dose dependent manner, blood fl ow remained unchanged during GLP-1 compared to saline infusion, but not in ALVA-41 transfected with control vector (ALVA-control). Further, whereas leg blood fl ow increased (41% vs. 22%, p=0.028) and regional ATBF we transplanted ALVA-GLP-1R and ALVA-control into nondiabetic athymic increased with maximum (115% vs. 28%, p=0.001) around 50 ± 5 minutes mice and treated them with vehicle or Ex-4 (300 pM/kg/day) for 4 weeks. As (range: 25-70 minutes). In healthy subjects, infusion of GLP-1 increases CO a result, tumor size of ALVA-GLP-1R was signifi cantly decreased compared acutely due to a positive chronotropic effect accompanied by vasodilation in with ALVA-control, with or without Ex-4 treatment. Immunohistochemistry the leg skeletal muscles and subcutaneous abdominal adipose tissue. of Ki67 revealed that ALVA-GLP-1R had a reduction of cell proliferation com- Supported By: Bispebjerg University Hospital; Danish Heart Foundation pared with ALVA-control. In addition, we next performed immunohistochem- istry of GLP-1R and Ki67 of 40 nondiabetic prostate cancer patients. Interest- 1910-P ingly, GLP-1R expression level was inversely associated with percentage of Glycodeoxycholic Acid Administration Increases Postprandial Ki67 positive cells and Gleason grading system of prostate cancers, signifi - GLP-1 Secretion in Healthy Humans cantly. These data suggest that GLP-1R expression and GLP-1 action attenu- SAMUEL VAN NIEROP, FRANK G. SCHAAP, FREDERIC VAZ, JOHANNES A. Obesity ates prostate cancer growth and progression in model mice and patients. ROMIJN, STEVEN W. OLDE DAMINK, MAARTEN R. SOETERS, Amsterdam, POSTERS Supported By: Suzuken Memorial Foundation Nether lands, Maastricht, Netherlands

Bile acids (BA) have come under investigation as hormones involved in Integrated Physiology/ 1908-P glucose and energy metabolism. Postprandial activation of the G protein- Lipocalin-type Prostaglandin d2 Synthase (L-PGDS): An Essential coupled BA receptor (TGR5) has been linked to increases in glucagon-like Component to Maintain Glucose Homeostasis after Vertical Sleeve peptide 1 (GLP-1) secretion, insulin secretion and energy expenditure. Gastrectomy We fi rst investigated plasma BA, GLP-1, insulin and glucose levels in 9 SUNIL KUMAR, RAYMOND G. LAU, THOMAS PALAIA, CHRISTOPHER E. HALL, healthy males after a mixed meal test. Insulin and glycodeoxycholate (gDCA) COLLIN BRATHWAITE, LOUIS RAGOLIA, Mineola, NY, East Northport, NY were positively correlated at 60’ after the meal (r 0.96; p < 0.001), but nei- Vertical Sleeve gastrectomy (VSG) ameliorates metabolic complications ther correlated with GLP-1. To establish the possible physiological role of in obese and diabetic subjects. Recently, L-PGDS has been considered an gDCA as a TGR5 agonist, we then investigated the effect of a single dose emerging target for diabetes and metabolic disorders; however, the mecha- of 750 mg gDCA. Subjects consumed a standard meal with or without gDCA nism involved in the regulation of glucose metabolism is not fully under- on separate occasions after which blood was sampled during 4 hrs and

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A491 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

assayed for GLP-1, glucose and insulin. Energy expenditure was measured 1913-P twice postprandially. Irisin Enhances Pancreatic Beta-Cell Survival and Insulin Secre- gDCA increased plasma GLP-1 at 60’ postprandially (24.1 vs. 13.6 µmol/L; tion and Can Mediate the Metabolic Cross Talk between the Skel- p < 0.01). There was a trend for higher postprandial insulin area-under-the- etal Muscle and Endocrine Pancreas curve in the fi rst hour (1055 vs. 741 min*pmol/L; p = 0.10), with no effect on NICOLA MARRANO, ANNALISA NATALICCHIO, GIUSEPPINA BIONDI, ROSSELLA plasma glucose levels (1182 vs. 1219 min*mmol/L; ns). gDCA plasma levels LABARBUTA, ANGELO CIGNARELLI, PIERO MARCHETTI, SEBASTIO PERRINI, were increased only after 240’, while other BA and fi broblast growth factor LUIGI LAVIOLA, FRANCESCO GIORGINO, Bari, Italy, Pisa, Italy 19 levels, as well as meal-induced energy expenditure were not affected by Skeletal muscle releases mediators that adjust insulin secretion to the gDCA administration. actual needs for appropriate peripheral glucose utilization. Saturated fatty The fi rst experiment showed a correlation between gDCA and insulin acids (SFA) affect pancreatic beta-cell function and modulate expression levels but not between either and GLP-1. However, in the second experi- of some cytokines in skeletal muscle. The aim of this study was to explore ment gDCA stimulated secretion of GLP-1. This may be mediated by intesti- the potential role of irisin, a secreted skeletal muscle protein derived from nal L-cell TGR5 activation after exposure to gDCA and fi ts the paradigm of cleavage of FNDC5, in the communication between skeletal muscle and BA as postprandial hormones in the gastrointestinal tract. TGR5-mediated beta-cells in response to excess SFA. Conditioned medium from rat L6 myo- increase in energy expenditure did not occur in this acute time frame. This tubes exposed to 0.5 mM palmitate for 24 h (PM24h) exerted detrimental acute experiment shows that BA administration may modulate GLP-1 secre- effects on INS-1E beta-cells by increasing apoptosis, decreasing cell prolif- tion in humans. However, timing of the BA administration as well as the eration, and reducing glucose-induced insulin secretion (GSIS). By contrast, form (i.e., conjugation) in which it is given may be crucial to leverage any when INS-1E cells were incubated with conditioned medium from myotubes benefi cial effects. exposed to palmitate for 4 h (PM4h), apoptosis markers, such as cytosolic Supported By: Dutch Diabetes Research Foundation release of oligosomes and cleaved caspase-3 levels, were reduced. Treat- ment of rat myotubes with palmitate for 4 h resulted in increased FNDC5 1911-P mRNA and protein content and 3-fold higher irisin release in the culture Dermatopontin Is a Novel Adipokine that Regulates Energy Homeo- medium. Palmitate also promoted release of irisin from human myotubes. stasis Mice fed a high fat diet showed a 2-fold increase in serum irisin levels within SHIXIONG TAN, YVONNE NG, HONGYU LI, MUHAMMAD MOHAMED, PIA ROE- 24 h. Importantly, the anti-apoptotic effect of the conditioned medium from DER, JILL THAM, WEI SUN, DONGHAI WU, SUE-ANNE TOH, AIMIN XU, WAN - PM4h was abrogated when INS-1E cells were cultured in the presence of JIN HONG, WEIPING HAN, Singapore, Singapore, Guangzhou, China, Hong Kong, an irisin neutralizing antibody. Finally, in human and rat pancreatic beta-cell China lines, as well as in human and murine primary pancreatic islets, recombi- Dermatopontin (DPT) was previously identifi ed as an extracellular matrix nant irisin prevented palmitate-induced apoptosis, by activating AKT/Bcl2 protein isolated from skin. Here we report that DPT is highly expressed and signaling, and directly stimulated insulin biosynthesis, GSIS and cell prolif- secreted from adipocytes via the classical ER-Golgi pathway, and can be eration. In conclusion, myotube-derived irisin emerges as a new player in readily detected in circulation. DPT levels in both subcutaneous and visceral the communication between skeletal muscle and beta-cells in response to adipose depots, and in blood positively correlate with body weights, and are short-term SFA challenge, since irisin can enhance GSIS and promote beta- higher in obese mice and humans. Administration of recombinant DPT by cell survival. intraperitoneal injection into mice on a high fat diet increases energy expen- diture, reduces weight gain, improves glucose clearance and insulin sensi- 1914-P tivity without affecting insulin secretion. In contrast, aged whole body DPT Bile Acids Stimulate Secretion of GLP-1 and PYY through Activation KO mice display reduced energy expenditure and exacerbated insulin resis- of Basolateral L-Cell GPBAR1 Receptors tance when compared with wild-type littermates. Transcriptomics analysis RUNE E. KUHRE, NICOLAI J. WEWER ALBRECHTSEN, OLAV LARSEN, SARA indicates that FGF-21 is highly upregulated in cells treated with recombinant L. JEPSEN, REIDAR ALBRECHTSEN, METTE M. ROSENKILDE, JENS J. HOLST, DPT. Our data show that DPT is a novel adipokine that regulates energy and Copenhagen, Denmark metabolic homeostasis. Glucagon-like peptide-1 (GLP-1) and peptide YY (PYY) are intestinal pep- Supported By: A*STAR Biomedical Research Council tides released from L-cells following nutrient intake and acting to inhibit appetite and optimize nutrient disposition. Here we show that bile acids 1912-P (BAs) stimulate their secretion from isolated perfused rat small intestine Methylglyoxal Induced Osteoclast Activation Mediated through (n=6 for all experiments). Intra-luminal (1mM) administration of taurodeoxy- MAPK/ERK Pathway cholate (TDCA) increased secretion (GLP-1: from 23±1 to 43±3 pM, PYY: 17±1 KEVIN K. YUE, DICKY K.M. LEE, CHING CHING TAM, GE ZHANG, Hong Kong, China to 55±3 pM, both P < 0.0001) and luminal instillation of unconjugated BAs Diabetes mellitus (DM) is characterized by chronic hyperglycemia and dia- (cholic acid (CA), deoxyCA (DCA) and chenoDCA (CDCA), all 1mM) resulted betic complications. Methylglyoxal (MG) is a reactive metabolite of glucose in comparable GLP-1 and PYY responses. TDCA depended on absorption via and one of the most reactive advanced glycation end-product precursors in the apical-bile acid transporter (ASBT) to stimulate GLP-1 and PYYs secre- which abnormal accumulation is found in the serum of diabetic patients. In tion, whereas specifi c inhibition of ASBT neither affected the absorption clinical studies, diabetic patients have higher risk to suffer from osteoporo- nor the GLP-1/PYY response to the mixture of (lipophilic) unconjugated BAs. sis. Osteoclast, which regulates the resorption of bone, is severely affected Depolarization generated by sodium-coupling of ASBT-mediated uptake of by MG. Hence the effect of MG leading to subsequent osteoporosis was conjugated BAs did not contribute to the secretory GLP-1 and PYY response investigated. Osteoclasts (cell line RAW 264.7) were treated with different to TDCA, as blockage of voltage-gated calcium channels had no effect. It concentrations and exposure time of MG. C57B mice were treated with MG therefore appears that BAs stimulate secretion of GLP-1 and PYY by mecha- by intraperitoneal injection for 4 weeks daily and hippocampus was col- nisms secondary to intestinal absorption, supposedly by direct activation of lected. The expressions of MAPK/ERK pathway molecules, together with the the Gαs-linked G-protein-bile-acid receptor 1 (GPBAR1), as shown here by mRNA expression of osteoclastic biomarkers were measured using Western its basolateral co-localization with GLP-1 producing cells in mouse and rat

Obesity blotting and RT-PCR respectively. For in vitro models, there was a signifi cant small intestine. In addition, BAs activated GPBAR1 transfected in COS7-cells POSTERS increase of phosphorylation of p44/42 MAPK (Erk1/2) after 24 hr of 200µM in following order of potency: lithoCA>DCA>CDCA for both human and rat and 400µM MG exposure, indicating activation of MAPK/ERK pathway. RT- GPBAR1. Supporting this fi nding, intra-arterial infusion in perfused intes-

Integrated Physiology/ PCR was performed to further confi rm the activation of osteoclast. The acti- tines of a poorly absorbable GPBAR1 agonist (1µM) robustly stimulated the vation of osteoclasts was demonstrated by increased mRNA expression of secretion of GLP-1/PYY, whereas intra-luminal infusion (10µM) was ineffec- CTSK (Cathepsin K), NFATC1 (Nuclear factor of activated T-cells, cytoplas- tive. Therefore, BAs are endogenous GLP-1/PYY secretagoges and GPBAR1 mic 1), OSCAR (Osteoclast-associated receptor), RANK (Receptor activator is an attractive pharmacologic target for enhancing the secretion of GLP-1 of nuclear factor kappa B) and TRAP (Tartrate-resistant acid phosphatase). and PYY with a view to treat type 2 diabetes and obesity. For in vivo models, the activation of osteoclast in tibia was demonstrated Supported By: Novo Nordisk Foundation Center for Basic Metabolic Research by increased mRNA expression of CTSK, OSCAR and TRAP. MG therefore activates the MAPK/ERK pathway, leading to increased expression of osteoclastic biomarkers and subsequent osteoclast activation. Activation of osteoclast may be an important event leading to imbalance bone remodeling resulting in osteoporosis.

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A492 OBESITY—ANIMALCATEGORY

1915-P & 1917-P Determinants of Fasting Hyperglucagonemia in Patients with Type 2 Harnessing GLP-1 and PYY Synergy for Bariatric Surgery-like Weight Diabetes and Nondiabetic Control Subjects Loss and Diabetes Remission MIA G. DEMANT, JONATAN I. BAGGER, MALTE P. SUPPLI, ASGER LUND, MORTEN STEPHANIE OLDHAM, SARAH WILL, LIHUAN LIANG, ASHA SETH, ISABELLE SER- HANSEN, METTE GYLDENLØVE, NICOLAI A. RHEE, KATRINE B. HANSEN, KRIS- MADIRAS, HAMID SALARI, JACQUELINE NAYLOR, ANDIE COLLINSON, BRAN- TINE J. HARE, KASPER AABOE, LISBETH BONDE, MIKKEL CHRISTENSEN, DAVID DON BOLAND, ELISE BERNARD, MARIA BEDNAREK, DAVID HORNIGOLD, DAVID P. SONNE, JENS JUUL HOLST, TINA VILSBØLL, FILIP K. KNOP, Hellerup, Denmark, BAKER, JAMES TREVASKIS, ANISH KONKAR, SIOBHAN O’BRIEN, ARTHUR T. Copenhagen, Denmark SUCKOW, Gaithersburg, MD Fasting hyperglucagonemia can be detrimental to glucose metabolism in Certain bariatric surgery procedures (e.g., Roux-en-Y) result in profound patients with type 2 diabetes (T2D) and may contribute to metabolic distur- weight loss, remission of diabetes and improved cardiovascular outcomes. bances in obese and/or prediabetic subjects. We evaluated the interrelation- Following meal consumption post-surgery, patients exhibit an exaggerated ship between fasting hyperglucagonemia and demographic and biochemical increase in the circulating levels of the enteroendocrine L-cell peptides GLP-1 parameters. Data from 106 patients with T2D (31% women, age: 57±9 years and PYY. Initial data in preclinical models utilizing GLP-1R and Y2R-selective (mean±SD); body mass index (BMI): 30.1 ± 4.4 kg/m2; fasting plasma glucose: PYY analogs revealed that combinations result in weight loss in diet-induced 9.61±2.39 mmol/L; hemoglobin A1c (HbA1c): 57.1±13.1 mmol/mol) and 163 obese mice (DIO) and glucose-lowering in diabetic db/db mice that mirror nondiabetic control subjects (29% women; age: 45±17 years; BMI: 25.8±4.1 the effects observed following bariatric surgery. This “bariatric surgery-like kg/m2; fasting plasma glucose: 5.2±0.4 mmol/L; HbA1c: 35.4±3.8 mmol/mol) effi cacy” could be achieved with combinations of doses that were signifi - were included. Multiple linear regression analysis was applied using a step- cantly lower than those required to elicit meaningful effi cacy with either of wise approach with fasting plasma glucagon as dependent parameter and the GLP-1R or Y2R analogs alone. To better understand the balance of phar- BMI, waist/hip ratio, blood pressure, HbA1c and fasting plasma glucose, macology required to achieve such impressive effi cacy, a series of 2-3 week insulin, glucose-dependent insulinotropic polypeptide and glucagon-like studies evaluating different ratios of Fc-GLP-1R and Fc-Y2R analogs were peptide-1 concentrations as independent parameters. Fasting plasma glu- conducted in db/db and DIO mice. In db/db mice, Fc-GLP-1R and Fc-Y2R mono- cagon concentrations where signifi cantly higher among patients with T2D therapy doses that had no effect on glucose homeostasis improved HbA1c by (13.5±6.3 vs. 8.5±3.8 mmol/L, P<0.001) together with HbA1c (P<0.001), fast- ~2% vs. vehicle when they were co-administered. This combination of doses, ing plasma glucose (P<0.001) and insulin (84.9±56.4 vs. 57.7±35.3 mmol/L, as well as several of the other ratios evaluated, resulted in preservation of P<0.001). When adjusted for T2D, HbA1c and insulin remained signifi cant circulating insulin levels in this model of β-cell failure. In DIO mice, combina- positive determinants for fasting plasma glucagon concentrations. Further- tions result in synergistic reductions in body weight that are driven primarily more, waist/hip ratio and BMI (only males) were signifi cant positive deter- by reductions in fat mass and associated with marked reductions in liver fat minants. We confi rm that fasting glucagon levels are abnormally high in T2D, and liver enzymes, as well as circulating cholesterol and triglycerides. These and show that fasting glucagon levels are infl uenced by waist/hip ratio, and studies have not only uncovered a unique interaction between GLP-1R and for males also BMI. This points to fat distribution as an important predictor Y2R agonists but, provide insight as to the balance of GLP-1R and Y2R activi- of fasting plasma glucagon concentrations, along with measures of glycemic ties that likely drive, and are most likely to mimic, the benefi cial effects of control and fasting plasma insulin. bariatric surgery.

& 1918-P OBESITY—ANIMAL Dual Amylin and Calcitonin Receptor Agonists (DACRA) Act Syner- gistically with GLP-1 on Food Intake and Body Weight SOFIE GYDESEN, SARA T. HJULER, KIM V. ANDREASSEN, MORTEN A. KARSDAL, Moderated Poster Discussion: Rodents Like Glucagon-Like Peptide-1 KIM HENRIKSEN, Herlev, Denmark (GLP-1) (Posters: 1916-P to 1923-P), see page 20. There is an urgent need for treatments which substantially reduce body weight. Even with the approval of high dose liraglutide as a treatment for & 1916-P obesity in the U.S., there is a limited availability of treatment options lead- GUB06-046, a Novel Secretin/GLP-1 Co-Agonist, Decreases Food ing to weight loss. KBP-089 is a dual amylin- and calcitonin receptor agonist Intake, Improves Glycemic Control and Preserves Beta-Cell Mass (DACRA) with superior activity over salmon calcitonin and amylin. We evalu- in db/db Mice ated tolerability, and the peptide as mono-therapy for obesity and in combi- SØREN B. VAN WITTELOOSTUIJN, LOUISE S. DALBØGE, GITTE HANSEN, KNUD nation with a GLP-1 molecule, liraglutide. 1. HFD and lean rats were four-step J. JENSEN, NIELS VRANG, JACOB JELSING, SØREN L. PEDERSEN, Hørsholm, dose-escalated (0.625, 2.5, 10 and 40 µg/kg) followed by 6 weeks of treat- Denmark, Frederiksberg, Denmark ment with either dose. 2. HFD rats received suboptimal doses of KBP-089 Bariatric surgery is currently the most effective treatment option for obe- s.c. (0.625 and 1.25 µg/kg), liraglutide (25 and 50 µg/kg) and the combinations sity, which has spurred an interest in developing a pharmaceutical mimetic (0.625 + 25 µg/kg and 1.25 + 50 µg/kg) and vehicle for 10 weeks. Dose esca- (a “medical bypass”). In a rat model of Roux-en-Y gastric bypass (RYGB) lation induced transient reduction in food intake at every escalation step. we identifi ed the gut hormone secretin (Sct) to be markedly upregulated in The following treatment with 2.5, 10 and 40 µg/kg resulted in a ~15% vehicle the alimentary limb, indicating a plausible role of secretin in the benefi cial corrected weight loss and a corresponding reduction adipose tissue (AT). effects of RYGB. Consequently, a library of novel Sct-based peptide co-ago- Moreover, all treatment groups improved oral glucose tolerance (p<0.01). A nists was developed and a lead compound (named GUB06-046) exhibiting single injection of KBP-089 (1.25 µg/kg) and liraglutide (50 µg/kg) reduced potent agonism of both the Sct receptor and the GLP-1 receptor was iden- 24 h food intake by 29% and 37% compared to vehicle, respectively. Notably, tifi ed. Acute single-dose administration of GUB06-046 (0.3 mg/kg, SC) to the combination reduced the 24 h food intake by 87%. 48 h post injection lean mice decreased food intake by 25% (p<0.001 vs. vehicle) and improved only the combination had signifi cant anorexic effect (30%). KBP-089 induced glucose tolerance by 38% assessed as total AUC during an oral glucose tol- a dose-dependent and sustained weight loss (13% vehicle-corrected in 2.5 erance test (p<0.001 vs. vehicle). Chronic administration of GUB06-046 (0.3 µg/kg group, p<0.001). KBP-089 (1.25 µg/kg) and liraglutide (50 µg/kg) low- ered body 8% and 2%, respectively, while the combination resulted in a 12% mg/kg, SC, BID) to db/db mice for eight weeks decreased cumulative food Obesity intake by 20% (p<0.001 vs. vehicle) and improved glycemic control as indi- body weight reduction. Moreover, the combination improved glucose toler- POSTERS cated by a decrease in fasting blood glucose levels (28.6 mmol/L vs. 15.6 ance (p<0.05) In conclusion, DACRAs acts synergistically with GLP-1 on food intake and body weight. KBP-089 was well tolerated, induced and sustained mmol/L, p<0.001 vs. vehicle) and a 1.6% reduction of HbA1c levels (7.2% Integrated Physiology/ vs. 5.6%, p<0.001 vs. vehicle). Subsequent stereological analysis revealed a a signifi cant weight loss and a reduction in AT in lean and HFD rats, under- marked increase in beta-cell mass (81%, p<0.01 vs. vehicle), with no impact scoring the potential of KBP-089 as an anti-obesity agent. on exocrine pancreas mass or pancreatic duct epithelial mass. The data sug- gest that Sct/GLP-1 co-agonism has benefi cial effects on appetite regula- tion, glucose homeostasis and beta-cell mass, without exhibiting prolifera- tive effects on the exocrine pancreas and the pancreatic duct epithelium.

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A493 OBESITY—ANIMALCATEGORY

& 1919-P of diastolic dysfunction and aortic stiffening, fi rst observed after 8 and 16 Teneligliptin Decreases the Uric Acids Levels by Reducing Xanthine weeks on WD, respectively. Herein, we determined whether and when the Dehydrogenase Expression in White Adipose Tissue of the Male DPP-4 inhibitor, linagliptin, prevents WD-induced development of diastolic Wister Rats dysfunction and aortic stiffness in female mice. We fed 4-week old C57BL/6 CHIHIRO TSUKAGOSHI MORIYA, HIROAKI SATOH, Fukushima, Japan mice a WD with or without a DPP-4 inhibitor, linagliptin (4mg·kg-1·day-1) for We have previously reported that teneligliptin improves not only glyce- 16 weeks and evaluated diastolic function and pulse wave velocity (PWV) mic control but lipid and uric acids metabolism in type 2 diabetic patients. after 4, 8 and 16 weeks on WD. Compared to lean control diet fed mice Recently it has reported adipose tissue produces and secretes uric acid (̆ in Figure), we observed early onset diastolic dysfunction after 8 weeks through xanthine oxidoreductase (XOR) and that the production is enhanced on WD (Áin Figure) and gradual aortic stiffening that was elevated after 16 in obesity. Therefore uric acid is also one of risk factor in the cardiovascu- weeks of WD without changes in blood pressure. Linagliptin prevented the lar diseases. In this study, we investigated the mechanism of the effects early onset of WD-induced diastolic dysfunction (◊ in Figure) and effi cacy of teneligliptin on the uric acids metabolism in the male Wister rats. Male persisted throughout the remainder of the feeding trial. Linagliptin also pre- Wister rats were fed normal chow diet (NCD), or 60% high fat diet (HFD) con- vented WD-induced aortic stiffening. Therapeutic targeting of DPP-4 with taining with either teneligliptin (~ 4.0 mg/kg/day) or not, for 4 weeks. Body linagliptin may be a promising strategy for prevention of CV complications weight was not signifi cantly different between the control and teneligliptin associated with obesity/diabetes in premenopausal women. groups, while body weight of the HFD-fed rats was signifi cantly greater than Figure. in the NCD-fed rats. The serum uric acid level did not signifi cantly differ between the control and teneligliptin groups under the NCD condition. How- ever, the serum uric acid level was signifi cantly decreased by 23% (from 0.34 ± 0.02 to 0.26 ± 0.02; P = 0.035) in the HFD-fed teneligliptin treated rats compared to the HFD-fed control rats. To investigate the molecular mechanisms of this effect of teneligliptin, we performed quantitative real- time PCR analysis. The expression level of xanthine dehydrogenase (Xdh), the major form of XOR in tissues, in either liver or WAT of NCD-fed rats was not altered by teneligliptin treatment. On the other hand, the Xdh expres- sion was reduced signifi cantly by 26% (P = 0.011) in the WAT of the HFD-fed teneligliptin treated rats compared that of HFD-fed control rats, whereas the Xdh expression in liver did not change signifi cantly in either group. In conclu- sion, these data suggest that teneligliptin had the effect of reducing the uric acid level by suppressing the Xdh expression in WAT of obesity.

& 1920-P Supported By: Boehringer Ingelheim Pharmaceuticals, Inc. Gemigliptin Ameliorates Western Diet-induced Metabolic Syn- drome in Mice & 1922-P SEUNG HEE CHOI, CHAE-MYEONG HA, BO-YOON PARK, HYEON-JI KANG, JI- Liraglutide Improves Insulin Sensitivity in Diet-induced Insulin MIN LEE, BYUNG-JUN CHOI, BYUNG-GYU KIM, HYE JIN HAM, KEUN-GYU PARK, Resistance Mice through Multiple Pathways IN-KYU LEE, Daegu, Republic of Korea JOSEPH ZHOU, PARATHAN CHANDRAMANI-SHIVALINGAPPA, ANIL POUDEL, Increasing attention has been focused on the extrapancreatic action of LIXIN LI, Montreal, QC, Canada, Mount Pleasant, MI dipeptidyl peptidase 4 (DPP-4) inhibitors, widely used oral anti-hyperglycemic Liraglutide, a full agonist of the GLP-1 receptor, has been used to treat type 2 agents in the treatment of metabolic and cardiovascular diseases. Here, we diabetes and was observed to improve glycaemic control, weight loss and liver investigated whether gemigliptin, a recently developed DPP-4 inhibitor, ame- enzymes. However, the underlying mechanisms of these effects are not clear. liorates features of metabolic syndrome. C57BL/6J mice were fed a Western To investigate these mechanisms, mice were fed with high fat high sucrose diet (WD) for 12 weeks and were subsequently divided into two groups: mice diet (HFHSD) for 3 months to induce insulin resistance, and diabetes. Subse- fed a WD diet alone or mice fed a WD diet supplemented with gemigliptin for quently, two groups of mice were injected with either liraglutide or vehicle an additional 4 weeks. Histological and biochemical examination of adipose daily for 14 days. Liraglutide signifi cantly inhibited food intake, improved gly- tissue and liver samples from mice was performed to determine the effect of cemic control and oral glucose tolerance test after the treatment period. Fur- gemigliptin on the development of metabolic syndrome. Gemigliptin treatment thermore, a signifi cant reduction of liver weight was observed in association attenuated WD-induced body weight gain, hypercholesterolemia, adipocyte with reduced protein levels of phosphorylated Acetyl-CoA carboxylase (ACC) hypertrophy, and macrophage infi ltration into adipose tissue, with the latter and upregulated cAMP-activated protein kinase (AMPK)-alpha protein expres- accompanied by an increased expression of uncoupling protein 1 in subcutane- sion. Brown adipose tissue plays a major role in controlling energy balance in ous fat. These events contributed to improved insulin sensitivity, as assessed rodents. Therefore, the potential effect of liraglutide on skeletal muscle brown by the homeostasis model assessment of insulin resistance and intraperitoneal fat differentiation was also examined. Our results indicated that liraglutide insulin tolerance test. Furthermore, gemigliptin reduced WD-induced hepatic treatment activated a full program of brown adipogenesis in skeletal muscle. triglyceride accumulation via inhibition of de novo lipogenesis and activation Specifi cally, expression of the brown fat-specifi c marker uncoupling protein-1 of fatty acid oxidation through AMP-dependent protein kinase activation. (UCP1), and PR domain-containing 16 (PRDM16), a transcription factor that reg- Gemigliptin ameliorated WD-induced hepatic infl ammation through suppres- ulates the thermogenic gene program in brown adipocytes, were signifi cantly sion of endoplasmic reticulum (ER) stress and oxidative stress with reduced upregulated. Furthermore, induction of mitochondrial biogenesis and function expression of ER stress markers and cytochrome P450 2E1, and decreased gene such as peroxisome proliferator-activated receptor γ (PPAR-γ), and cell DNA oxidation. This study demonstrates that DPP-4 inhibition with gemigliptin death activator-A (Cidea) were signifi cantly induced by liaglutide. Our data effectively ameliorates WD-induced metabolic dysfunctions. DPP-4 inhibitors indicates that liraglutide improves insulin sensitivity in diet-induced insulin may represent promising therapeutic agents for metabolic syndrome beyond Obesity resistant mice through many pathways. These pathways include suppression

POSTERS their current role as anti-hyperglycemic agents. of appetite, brown fat-winduced elevation of energy expenditure in skeletal Supported By: Korean Ministry of Health and Welfare muscle, and improvement of liver steatosis via suppression of hepatic lipogen-

Integrated Physiology/ esis in the AMPK-dependent pathway. & 1921-P Dipeptidyl Peptidase-4 Inhibition with Linagliptin Prevents Dia- & 1923-P stolic Dysfunction in Western Diet-Fed Female Mice Liraglutide and Exendin-4 Differentially Alter Body Weight Loss, VINCENT G. DEMARCO, JAMES R. SOWERS, MONA GARRO, ANNAYYA AROOR, Glucose Tolerance, Postprandial Dyslipidemia, and Gut Microbiota JAVAD HABIBI, GUANGHONG JIA, Columbia, MO in 60% High-Fat Diet-induced Obese Mice We previously reported improvement of diastolic dysfunction by DDP-4 FRANCOIS BRIAD, EMMANUEL BROUSSEAU, BENJAMIN LELOUVIER, FLORENCE inhibition in male Zucker Obese rats, a genetic model of diet-induced obe- SERVANT, MICHAEL COURTNEY, REMY BURCELIN, THIERRY SULPICE, Labege, sity. We also reported abrogation of cardiovascular (CV) protection in female France, Toulouse, France mice fed an obesogenic WD, compared to males, with earlier development Antidiabetic drugs need to demonstrate benefi ts, beyond the glucose con- trol, to target niches of type 2 diabetic patients. To differentiate the GLP-1

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A494 OBESITY—ANIMALCATEGORY receptor agonists liraglutide and exendin-4 (ex-4) and identify discriminant kinase which mediates activation of the noncanonical NF-κB pathway. We therapeutic features, we studied metabolic parameters in the 60% high fat here show that liver-specifi c deletion of NIK lowers hepatic glucose produc- diet-induced obese (DIO) mouse model. tion and hepatic steatosis in mice fed a high fat diet (HFD). NIKfl ox/fl ox mice DIO mice were treated i.p. with vehicle, ex-4 10µg/kg BID or liraglutide were crossed with mx1-cre mice, and liver NIK was inducibly deleted by poly 100µg/kg QD and drug effects were evaluated after 14 days of treatment. (I:C) injection. Liver-specifi c deletion of NIK did not alter insulin sensitivity, Compared with vehicle, ex-4 induced a signifi cant body weight loss (-3.6g) but decreased the hyperglycemic response to glucagon and hepatic glucose after treatment, while liraglutide showed a more pronounced effect (-6.2g). production in mice fed a HFD for 10 weeks. Mechanistically, deletion of Ex-4 showed better glycemic control over liraglutide after an oral glucose liver NIK increased the expression of hepatic PDE3B, thus decreasing the tolerance test, which was due to stronger impact on gastric emptying (as cAMP/PKA pathway in hepatocytes. Deletion of liver NIK also decreased the measured with acetaminophen oral administration). However, following a expression of hepatic lipogenic genes (e.g., LXRα, SREBP1c, Fasn, ACC, and test meal, both ex-4 and liraglutide signifi cantly reduced blood glucose by SCD1), thus protecting against HFD-induced hepatic steatosis in mice. These 40% and triglycerides levels by 25%. Ex-4 signifi cantly reduced intestinal data suggest that NIK in liver cells couple infl ammation to both hepatic glu- cholesterol absorption by 40% after a 14C-cholesterol labeled olive oil oral cose production and lipogenesis, contributing to aberrant glucose and lipid gavage, while liraglutide showed a more pronounced effect (50% reduction). metabolism in obesity. Fecal cholesterol mass excretion was signifi cantly increased by 62 and 75% Supported By: National Institutes of Health with ex-4 and liraglutide, respectively. Gut microbiota taxonomic profi ling further discriminated ex-4 and liraglutide with signifi cant changes in various & 1926-P bacteria taxa, such as bifi dobacteriaceae, clostridiaceae or lactobaciliceae. Chronic Treatment of Thyroid Hormone Reduces Obesity and Fatty These changes may be linked to the different impact of both drugs on gastric Liver without Affecting Infl ammation in Diet-induced Obese Mice emptying and immune system. TAEKYOON KIM, ALEXANDRA SHERIFF, HEE JOON KANG, RANDALL H. FRIED- In conclusion, ex-4 and liraglutide differentially alter microbiota, which LINE, HYE-LIM NOH, LEIGH ANDERSON, KUNIKAZU INASHIMA, ALYCIA QUICK, may in part contribute to the different mechanisms improving metabolic BIANCA ZARRELLA, CATHERINE SOLIMAN, CECILIA UONG, XIAODI HU, KAREN parameters. The DIO mouse model combined with our in vivo experiments KELLY, JASON K. KIM, Worcester, MA represents a useful tool to differentiate antidiabetics, and demonstrate their Thyroid hormone plays an important role in regulating energy metabolism, benefi ts beside glucose control. and active T3 (Triiodothyronine) hormone therapy may be used to treat obe- sity and metabolic disease. Here we examined the effects of chronic treat- ment of T3 on metabolic and infl ammation profi le in obese C57BL/6 mice fed Moderated Poster Discussion: Infl ammation and Immune Cells—Role a high-fat diet for 20 weeks. Mice received a daily intraperitoneal (ip) injec- in Obesity and Insulin Function (Posters: 1924-P to 1931-P), see page 14. tion of T3 (28 ng/g of bw) or DMSO vehicle (controls) for 20 days (n=6/group). During the T3 treatment, mice became less obese with a gradual decline in & 1924-P body weight and fat mass (Figure 1; *P<0.05). After 20 days, T3-treated mice Nrf2 Activation by Glucoraphanin Relieves Obesity and Insulin showed improved glucose tolerance, suggesting increased insulin sensitiv- Resistance through Increased Energy Expenditure and Alternative ity in these mice as compared to controls (Figure 2; ip glucose tolerance Activation of Macrophages test). Histological examination of liver samples showed the T3-treated mice NAOTO NAGATA, YUSUKE USHIDA, YUDAI AOKI, LIANG XU, FEN ZHUGE, HIRO- had reduced fatty liver as compared to controls (Figure 3). Despite dramatic YUKI SUGANUMA, SHUICHI KANEKO, TSUGUHITO OTA, Kanazawa, Japan, Nagoya, improvement in insulin sensitivity and fatty liver state, circulating levels Japan of infl ammatory cytokines, such as IL-6 and MCP-1, were not signifi cantly Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is a master regulator of altered in T3-treated mice (Figure 4). Local levels of cytokines, IL-6, IL-1beta, the antioxidant response and plays a role in maintaining glucose homeosta- IFNgamma, and TNF-alpha, in liver were also not affected by T3 treatment. sis. Although synthetic Nrf2 inducers alleviate obesity and insulin resistance These data indicate that chronic T3 treatment reduced obesity and fatty in animal models, they are not clinically available due to safety concerns. liver, resulting in improved insulin sensitivity in diet-induced obese mice We examined the effect of glucoraphanin (GR), a naturally occurring Nrf2 without affecting systemic and local infl ammation. inducer, on high-fat diet (HFD)-induced obesity and insulin resistance in Figure. mice. Eight-week-old male C57BL/6J mice were fed a HFD or a HFD contain- ing GR (300 mg/kg) for 14 weeks. GR suppressed HFD-induced weight gain by 15% (p < 0.01) without affecting food intake. Indirect calorimetry revealed that GR increased energy expenditure of mice fed HFD by 12% (p < 0.01). GR improved glucose tolerance, hyperinsulinemia (HFD, 5.6 ± 1.0 vs. HFD-GR, 2.9 ± 0.2 ng/ml, p < 0.05; fed state), and fatty liver and enhanced hepatic insulin signaling assessed by phospho-Akt. These changes were associated with attenuated excess lipid peroxidation (TBARS) and JNK and ERK activa- tion. A fl ow cytometry analysis revealed that GR decreased the number of liver macrophages identifi ed as CD45+CD11b+F4/80+ cells by 44% (p < 0.05) in mice fed a HFD. In addition, HFD-GR mice had 67% fewer CD11c+CD206− (M1) macrophages but 20% more CD11c−CD206+ (M2) macrophages com- pared with HFD mice, resulting in a predominance of the M2 compared to the M1 macrophage population in the liver. Although GR did not affect accu- mulation of adipose tissue macrophages (ATMs), it caused an M2 dominant phenotypic shift of ATMs. Importantly, the weight-reducing and insulin-sen- sitizing effects of GR were abolished in HFD-fed Nrf2 knockout mice. Thus, GR-mediated activation of Nrf2 relieved obesity, and alternatively activated Obesity

macrophages in the liver and adipose tissue, thereby attenuating obesity- POSTERS related infl ammation and insulin resistance in mice. Supported By: National Institutes of Health (R01-DK080756, U24-DK093000, Supported By: Japan Society for the Promotion of Science R24-DK090963) Integrated Physiology/

& 1925-P & 1927-P Liver NF-κB-inducing Kinase (NIK) Links Infl ammation to Glucose The Role of Free Fatty Acids in the Initiation of Hepatic Infl ammation Counterregulation and Hepatic Steatosis in Obesity and Oxidative Stress YAN LIU, LIANG SHENG, YI XIONG, HONG SHEN, LIANGYOU RUI, Ann Arbor, MI DANIEL HARMON, CHAO WU, IAN SIPULA, NICOLAS DEDOUSIS, GABRIELE Obesity is associated with chronic infl ammation. Infl ammation promotes SCHOISWOHL, ERIN KERSHAW, ERIC E. KELLEY, MAJA STEFANOVIC-RACIC, metabolic disease progression, including insulin resistance and hepatic ste- ROBERT M. O’DOHERTY, Pittsburgh, PA atosis; however, intracellular mediators, which couple infl ammation to glu- Obesity-induced hepatic steatosis associates with markers of liver dys- cose and lipid metabolism, are not fully understood. We previously reported function including infl ammatory cell infi ltration and increased free radical that obesity is associated with a marked increase in hepatic NIK, a Ser/Thr production. However, the mechanisms initiating hepatic infl ammation and

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A495 OBESITY—ANIMALCATEGORY

oxidative stress in vivo are unclear. In the present study, the impact of altered & 1929-P FFA delivery on liver infl ammation (macrophage (MAC) and dendritic cell (DC) Anti-infl ammatory Protection of Pancreatic Islets during Prolonged infi ltration) and oxidative stress (protein carbonyl, glutathione and/or DMPO High-Fat Diet (HFD) by NUPR1 through Enhanced Local Secretion radical adduct production) were assessed in fi ve in vivo mouse models (obe- of IL-1RA sity, lipid infusions, fasting, short-term high fat diets, and genetic impair- JOCHEN SEUFERT, AMIR E. MEHANA, JAN STRAETENER, JOHANNES BAU- ment of adipose tissue (AT) lipolysis). As expected, obesity was associated MANN, KATHARINA LAUBNER, NIKOLAOS PERAKAKIS, GÜNTER PÄTH, Freiburg, with pronounced DC/MAC infi ltration of the liver, and this was related to Germany substantial liver oxidative stress. Three-weeks, but not 1 week, of high- Metabolically unhealthy obesity confers a state of subclinical low grade fat feeding was suffi cient to induce hepatic infl ammation, and was also infl ammation that contributes to insulin resistance and beta-cell dysfunction observed in two models of acute lipid delivery (16 hr lipid infusion and over- in susceptive individuals. Previously, we demonstrated that mice with beta- night fasting). Furthermore, hepatic oxidative stress was observed after 16 cell-specifi c overexpression of the intracellular protein NUPR1 (NUPR1-Tg) hr fasting and within just one week of high-fat feeding. Finally, the deletion maintained normal glycemia during prolonged HFD (60% energy from fat) of ATGL (adipose triglyceride lipase) in AT, which prevented AT lipolysis, and while wild type (WT) controls became diabetic. Here we investigated insulin thus FFA delivery to the liver, protected the liver against obesity-induced secretion, apoptosis, and secretion of infl ammatory IL-1beta and its physio- and fasting-induced infl ammation. Together, these data demonstrate that logic receptor antagonist IL-1RA in isolated islets from normal and 20 weeks manipulation of FFA delivery to the liver leads to substantial increases in HFD-fed WT and NUPR1-Tg mice. liver immune cell content and oxidative stress. Furthermore, the occurrence Islets from normal diet-fed WT mice demonstrated signifi cantly increased of oxidative stress prior to increased hepatic infl ammation in response to apoptotic cleavage of the DNA repair enzyme PARP over time in culture and high-fat feeding point to FFA-induced oxidative stress as a mechanism for after 24 h exposure to STZ or IL-1beta. In contrast, PARP cleavage was not the initiation and maintenance of liver immune cell infi ltration during obesity much affected in NUPR1-Tg islets. Under same stress conditions, basal and development. stimulated insulin secretion was signifi cantly reduced in WT but maintained Supported By: National Institutes of Health in NUPR1-Tg islets. Secretion of IL-1beta was hardly detectable in islets from normal-fed mice but signifi cantly upregulated in islets from HFD-fed mice. & 1928-P In NUPR1-Tg islets, HFD-mediated induction of endogenous IL-1beta was PARP14 Plays a Key Role in Adipocyte Differentiation: A Potential reduced and less further inducible by STZ or IL-1beta. The reduced secretion Novel Target for Obesity and Insulin Resistance of IL-1beta from NUPR1-Tg islets was associated with signifi cant upregula- NORIYUKI INOUE, NAOAKI ARIMA, HIROSHI IWATA, MALLARY HOIDAL, tion of IL-1RA which was not observed in WT islets. ANDREW MLYNARCHIK, MARY WHELAN, MARK BOOTHBY, KEN MIZUNO, In summary, NUPR1 overexpression effi ciently preserves insulin secre- MASANORI AIKAWA, Boston, MA, Nashville, TN tory functions during HFD and cellular stress in vitro. During prolonged HFD, We demonstrated that poly ADP ribose polymerase 14 (PARP14), a mono- protective anti-infl ammatory mechanisms involve upregulation of IL-1RA. ribosylation enzyme, induces pro-infl ammatory activation of macrophages. NUPR1 may represent an endogenous molecular target for the preservation We also reported that PARP14 defi ciency suppresses adipocyte differentia- of islet function during low grade infl ammation in metabolically unhealthy tion and PARP14 overexpression enhances PPARγ transcriptional activity. obesity. The present study has further explored the mechanisms by which PARP14 regulates adipocyte biology. In the adipocyte-like cell line 3T3-L1 cells that & 1930-P stably express PARP14, enforced PARP14 expression enhanced adipocyte dif- Macrophage-specifi c Deletion of Major Histocompatibility Com- ferentiation (Figure) in contrast to the previous results of PARP14 defi ciency. plex II Does Not Improve Systemic Insulin Sensitivity because of These results suggest that PARP14 promotes adipocyte differentiation. We Adipocyte Compensation further examined the role of the catalytic domain and 3 macrodomains of ALECIA M. BLASZCZAK, TUO DENG, JOEY LIU, STEPHEN BERGIN, VALERIE P. PARP14. Mutant PARP14, which lacks catalytic activity of mono-ribosylation, WRIGHT, QING WANG, WILLA A. HSUEH, Columbus, OH, Houston, TX enhanced PPARγ activity as did wild-type PARP14. However, PARP14 lacking Both adipocytes and adipose tissue macrophages (ATMs) present antigen 3 macrodomains did not affect PPARγ transcriptional activity. These fi nd- to regulate differentiation and activation of adipose resident T cells (ARTs), ings indicate that PARP14 regulates PPARγ transcriptional activity through which is implicated as a key determinant of systemic insulin sensitivity. the macrodomains. Infl ammatory signaling in adipocytes contributes to obe- Within 2-3 weeks of high fat diet (HFD) in mice, major histocompatibility II sity. TNF-α and IL-1β induced PARP14 transcription and protein expression (MHCII) complex, which mediates antigen presentation, is increased in adipo- in 3T3-L1 adipocytes. In conclusion, PARP14 may link infl ammation and adi- cytes, leading to increased ARTs pro-infl ammatory Th1 cells and interferon-γ pocyte differentiation via PPARγ activity in adipocytes, serving as a novel (IFNγ) expression. Adipocyte-specifi c knockout of MHCII results in a marked therapeutic target for obesity and insulin resistance. increase in ARTs immunosuppressive T cells (Tregs) demonstrated both Figure. by fl ow and gene expression of Foxp3, a marker of Tregs, and decreased adipose infl ammation and systemic insulin resistance despite HFD. In con- trast, ATM MHCII expression is not altered during 8 weeks HFD in mice, and macrophage-specifi c knockout of MHCII (mMHCII-/-) did not improve insulin sensitivity during 12-14 weeks HFD, although bone marrow derived macrophages lacking MHCII were less responsive to LPS-induced infl amma- tion in vitro. To explain this phenotype, we found that mMHCII-/- vs. WT mice reveal a 2-fold (p< 0.05) increase in adipocyte MHCII expression, a 1.5-fold (p<0.05) increase in ARTs IFNγ expression, a 75% (p<0.01) decrease in ARTs Tregs, and a 60% (p<0.05) drop in Th2 cells. Moreover, Tregs expressing the IL-33 receptor and ILC-2 cells, which both regulate Tregs were decreased in adipose tissue of mMHCII-/- mice. Both the adipocytes and ATMs revealed

Obesity increased IL-1β and Nlrp3 expression. There was no difference in ATM Cd11c

POSTERS expression, a marker of macrophage infl ammation. Taken together, these data suggest that the adipocyte is a major adipose immune cell, regulat-

Integrated Physiology/ ing the ARTs subtype during HFD and compensating for macrophage loss of MHCII.

& 1931-P Colonic Proinfl ammatory Macrophage-Ccl2 Axis Regulates Meta- bolic Homeostasis through IL18 from Infl ammasome YOSHINAGA KAWANO, NOBUYUKI WATANABE, TETSUHIRO KIKUCHI, MASA- FUMI ONODERA, JUN NAKAE, HIROSHI ITOH, Tokyo, Japan High fat-diet (HFD) induces low-grade chronic infl ammation and insu- lin resistance via infl ammasome in insulin-responsive tissues. However,

For author disclosure information, see page A696. & Moderated Poster Discussion ADA-Supported Research

A496 OBESITY—ANIMALCATEGORY still little is known about the mechanism how HFD causes infl ammasome- insulin resistance. Ldlr-/- mice fed a high fat high sucrose diet with added dependent infl ammation in these tissues. Expression of Emr1 and infl am- cholesterol (HFHSC) develop obesity, dyslipidemia, and insulin resistance, masome gene (Il18, Il1b, Nlrp3) and population of F4/80+CD11b+CD11c- cells components of human metabolic syndrome. Ldlr-/- mice fed the HFHSC diet are increased in colon from 4-week HFD mice. Furthermore, ASC+cells, supplemented with 1% 10, 12 CLA are resistant to weight gain and have which were a hallmark of infl ammasome, in lamina propria and expression reduced subcutaneous fat, in stark contrast to weight matched calorically of Caspase-1 p10, which is a part of active enzyme of infl ammasome, of restricted mice that maintain subcutaneous fat. Despite weight loss, 10, 12 colonic CD11b+ cells were increased at 4-week HFD. These data indicate CLA-supplemented mice do not show improved insulin sensitivity as seen HFD induces an infl ammasome-dependent chronic infl ammation in colon at in calorically-restricted mice. Thiazolidinediones such as rosiglitazone are an early period. To investigate whether HFD-induce colonic infl ammation used clinically to improve insulin sensitivity. We therefore hypothesized that affects insulin sensitivity in vivo, we generated intestinal epithelial cell- rosiglitazone would improve 10, 12 CLA-mediated insulin resistance with- specifi c tamoxifen inducible Ccl2, which is one of important chemokines out compromising weight loss. Mice consuming 10, 12 CLA and rosiglitazone for intestinal macrophage, knockout mice (Vil-Ccl2KO). Vil-Ccl2KO exhibit (10 mg/kg) lost a similar amount of weight as mice fed 10, 12 CLA alone. improved glucose tolerance and insulin sensitivity despite of obesity. Gene Rosiglitazone also attenuated subcutaneous fat loss induced by 10, 12 CLA. expression of Emr1, Ccr2 and Il18 in colon from Vil-Ccl2KO were decreased Further, rosiglitazone improved 10, 12 CLA-induced insulin resistance param- compared with control. The concentration of IL-18, which is an infl am- eters such as fasting glucose and insulin, and glucose and insulin tolerance masome-dependent pro-infl ammatory cytokine, in portal vein of Vil-Ccl2KO tests. Rosiglitazone treatment also increased plasma and subcutaneous adi- was decreased. Interestingly, although weight and size of adipocytes in Vil- pose tissue adiponectin levels, without adversely affecting plasma lipids. Ccl2KO were increased as well as control mice, the numbers of crown-like We conclude that co-treatment of mice exhibiting characteristics of human structure and expression of Ccl2 in epididymal fat was decreased. Further- metabolic syndrome with 10, 12 CLA and rosiglitazone not only promotes more, administration of IL18 increased Ccl2 expression in epididymal fat. In fat loss, but also improves insulin sensitivity, unlike 10, 12 CLA treatment contrast, administration of IL18BP, which is an inhibitor of IL18, decreased alone. The notion that a compound as readily available as 10, 12 CLA could Ccl2 expression. These data indicate that colonic Ccl2 regulates glucose induce weight loss has great therapeutic potential, and when coupled with metabolism through pro-infl ammatory infi ltration of colonic macrophages an insulin-sensitizing agent such as rosiglitazone, the adverse side effect of via IL18 derived from an infl ammasome. insulin resistance is improved. Supported By: National Institutes of Health

Moderated Poster Discussion: Mechanisms of Insulin Resistance— & 1934-P Mouse Models (Posters: 1932-P to 1939-P), see page 15. Effect of High-Fat Diet on Weight Gain and Glucose Tolerance in Wild Type and Insulin-degrading Enzyme Knockout Mice & 1932-P FREDERICK G. HAMEL, GERRI L. SIFORD, RONDA L. SIMPSON, ROBERT G. BEN- A Novel Class of Rexinoid, UAB126, Ameliorates High-Fat Diet- NETT, Omaha, NE induced Obesity and Insulin Resistance through FGF-21-Dependent Human genetic studies have associated insulin-degrading enzyme (IDE) and -Independent Mechanisms in Mice with diabetes. However, studies of the IDE knockout (KO) mouse have dem- GUANG REN, TEAYOUN KIM, MARTIN E. YOUNG, VENKATRAM R. ATIGADDA, onstrated a variable phenotype with respect to glucose tolerance. We exam- DONALD MUCCIO, KIRK M. HABEGGER, JEONG-A KIM, Birmingham, AL ined the effect of diet-induced obesity, a standard method for inducing insu- Rexinoids, synthetic reagents that bind to retinoid X nuclear receptors lin resistance in rodents, on weight gain and glucose handling in C57BL/6 x (RXRs), have been suggested as potential pharmacological treatments for 129/Sv IDE+/+ (WT) and IDE-/- (KO) mice. Both male and female mice were metabolic syndrome. Previously known rexinoids, including bexarotene, studied. Age matched mice (approximately 90 days old), were provided ad have benefi cial effects on glucose homeostasis, but most of these rexi- lib, either a 25% fat content (by calories) diet (LabDiet 5015) or a 45% fat diet noids have side effects including elevation of serum triglyceride levels and (Research Diets D12451) for 56 days. Both had 20% protein (by calories). Glu- hepatomegaly. We developed a new class of rexinoids, UAB126, and evalu- cose handling was assessed with fasting blood glucose, oral glucose toler- ated whether UAB126 ameliorates high fat diet (HFD)-induced obesity and ance tests (OGTT) and HOMA-IR. The 25% diet marginally increased weight, insulin resistance. We observed that UAB126 prevented HFD-induced obe- with no signifi cant differences between WT and KO for either male or female sity, and improved glucose tolerance and insulin sensitivity in mice. Body mice. The 45% diet increased weight signifi cantly more in WT compared composition analysis revealed that fat mass was markedly reduced by to KO mice, in both males (16.5g ± 1.8 vs. 9.0g ± 1.3 p<0.05) and females UAB126 treatment while lean mass was not changed. UAB126 treatment (10.9g ± 1.3g vs. 3.3 ± 0.9 p<0.01). After 3 weeks, KO males on the 25% diet increased energy expenditure and fat oxidation without altering food intake. showed a signifi cantly increased fasting blood glucose (p<0.05) compared HFD mice treated with UAB126 had decreased serum triglycerides, free to the other groups. Both WT and KO males on the 45% diet had elevated fatty acids, and cholesterol levels compared to HFD mice. We also observed glucose levels. At the end of the study, the 25% diet showed no signifi cant that UAB126 reversed HFD-induced obesity and insulin resistance. Interest- differences among the groups, although fasting blood glucose of KO males ingly, mRNA expression of fi broblast growth factor (FGF) 21 was signifi cantly tended to be higher on both diets, but did not reach statistical signifi cance. increased in the livers of UAB126 treated mice. Since FGF-21 contributes to However, WT and KO male mice on the 45% diet showed insulin resistance glucose and lipid metabolism, we used FGF-21-null mice to examine the ben- by OGTT, whereas female mice were normal (2-way ANOVA, p=0.002). Simi- efi cial effects of UAB126 are mediated by FGF-21. The effects of UAB126 on lar results were seen with HOMA-IR (p<0.05). Our studies demonstrate a improving glucose tolerance and hepatic steatosis were reduced in FGF-21- sexual dimorphism in this mouse strain, with females being resistant to null mice compared to WT mice. However, the effect of UAB126 on weight developing insulin resistance. Surprisingly, both male and female KO mice loss was not reduced in FGF-21-null mice. These results suggest that UAB126 were resistant to weight gain on the higher fat diet. This indicates a hereto- has benefi cial effects on glucose homeostasis and lipid lowering through a fore unknown aspect of the function of IDE, and suggests a possible thera- FGF-21-dependent mechanism, while it ameliorates obesity through a FGF- peutic target for the treatment of obesity. 21-independent mechanism. From these results we concluded that UAB126 Supported By: U.S. Department of Veterans Affairs; Bly Memorial Research may be a novel therapeutic for prevention and treatment for obesity and Fund Obesity

insulin resistance. POSTERS Supported By: American Diabetes Association (1-12-BS-99 to J-A.K.); American & 1935-P Heart Association Spinophilin-defi cient Mice Are Protected from Diet-induced Obe- Integrated Physiology/ sity and Insulin Resistance & 1933-P HONGJUN WANG, YONG ZHANG, LILI SONG, JINGJING WANG, Charleston, SC Rosiglitazone Improves 10, 12 CLA-mediated Insulin Resistance Browning of white adipose tissue (WAT) has been shown to reduce obe- without Ameliorating Weight Loss in a Mouse Model of Metabolic sity and obesity-related complications suggesting that factors that promote Syndrome WAT browning may have applications in the development of therapeutic SHARI WANG, LEELA GOODSPEED, BARBARA HOUSTON, LAURA DEN HARTIGH, strategies for treating obesity. Here, we show that ablation of spinophilin Seattle, WA (SPL), a ubiquitously expressed, multidomain scaffolding protein, increases Trans-10, cis-12 conjugated linoleic acid (10, 12 CLA) is a dietary trans metabolism and improves energy balance. fatty acid reported to promote weight loss by unknown mechanisms. 10, 12 Male and female SPL knockout (KO) and wild type (WT) littermate controls CLA is associated with reduced adiposity, while simultaneously promoting were fed a Chow diet or a high fat diet (HFD). Body weight, hepatic steatosis,

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A497 OBESITY—ANIMALCATEGORY

glucose and insulin tolerance, physical activity and expression of browning slight reduction of food intake and there was an associated decrease of genes in adipose tissues were measured and compared. body weight. HbA1c and liver weight were also signifi cantly reduced in mice Male SPL knockout (KO) mice fed a Chow diet were signifi cantly leaner, treated with ASA19. These fi ndings indicate that T1R2/T1R3 contributes to had lower body weights, and exhibited better glucose tolerance and insulin regulation of insulin sensitivity and glucose metabolism in the peripheral tis- sensitivity than wild type (WT) littermate controls. When fed a high fat diet sues. Blocking this receptor might result in improvement of glucose metabo- (HFD), spl KO mice were protected from increased body fat, weight gain, lism, at least partly through increased hepatic insulin sensitivity and activa- hepatic steatosis, hyperinsulinemia, and insulin resistance. Physical activity tion of AMPK in skeletal muscle. Accordingly, T1R2/T1R3 may be a novel of spl KO mice was markedly increased compared to WT controls. Further- target for the treatment of metabolic syndrome. more, expression of the brown adipocyte marker, UCP-1, and the mitochon- drial activity markers, cd137 and c-idea, were signifi cantly increased in vis- & 1938-P ceral WAT (vWAT) of spl KO mice, suggesting that SPL knockout protected Dietary Curcumin Promotes Enhanced Insulin Sensitivity in Mice the mice from HFD-induced obesity and its metabolic complications, at least Given a High-Fat/High-Sugar Diet in part, by promoting the browning of white adipocytes in vWAT. MICHAEL ROUSE, JENNIFER FIORI O’CONNELL, ADRIA SUMMERS, JOSEPHINE Our data identify a critical role of SPL in regulating glucose homeostasis, M. EGAN, Baltimore, MD obesity, and adipocyte browning. These results suggest that SPL may serve Type 2 diabetes (T2DM) is a metabolic disease that affects an estimated as a drug target for obesity and diabetes. 348 million people worldwide. Although T2DM is primarily characterized by sustained levels of high glucose, other factors, such as obesity, chronic & 1936-P infl ammation, fatty liver, and islet dysfunction also signifi cantly contribute to High Fat Western Diet-induced Brain Insulin Resistance and Cogni- the development of this disease. Due to the putative therapeutic properties tive Impairment of curcumin (CUR), a natural polyphenol and primary component of turmeric, VISHAL KOTHARI, TALIA TORNABENE, YUWEN LUO, MICHAEL GREENE, GEETHA we investigated if daily consumption of curcumin (400 mg/kg) was protec- THANGIAH, RAMESH JEGANATHAN, Auburn, AL, Montgomery, AL tive of the metabolic consequences of a high fat/high sugar (HFS) diet given High fat and sucrose containing diet-induced obesity is associated with for 4.5 months in mice. Mice given HFS + CUR had reduced body weight insulin resistance and many other chronic, diet related illnesses. There is compared to mice on HFS alone, despite the fact that these mice actually growing evidence that insulin resistance and other features of the metabolic ate more food per week. This data was refl ective of the fact that HFS mice syndrome can infl uence cognitive health. This study was designed to deter- had elevated leptin levels in circulation, which were decreased in HFS + CUR mine whether diet-induced changes in the peripheral insulin sensitivity could mice. HFS + CUR mice also had with lower fasting blood glucose, insulin, contribute to alterations in brain insulin signaling and cognitive functions. and HOMA-IR compared to HFS mice. During IPGTT and ITT, we found that Four weeks old, male C57BL/6 mice were randomly assigned high-fat (40% HFS + CUR mice had lower blood glucose levels compared to HFS mice, prov- energy from fat) diet and liquid Sugar (42 g/L) or normal chow (12% kcal fat) ing that CUR improved insulin sensitivity. Additionally, HFS + CUR mice had diet for 14 weeks. This model was characterized for peripheral insulin resis- signifi cantly smaller islets compared to HFS in keeping with reduced insulin tance by glucose and insulin tolerance test. We also examined the changes requirement, and more closely resembled islets from mice on control diet. in protein expressions related to brain insulin signaling and cognitive func- HFS mice had increased neutral triglycerides and lipids in their livers com- tions. High fat western diet-fed mice exhibited a signifi cant increase in body pared to mice on a control diet, which were signifi cantly diminished by CUR weight, lower glucose tolerance and insulin tolerance compared to normal administration. CUR enhanced insulin signaling via increased hepatocyte chow diet animals. Brain tissues of western diet mice were insulin-resis- pAkt levels compared to HFS mice. In addition, we found that the expres- tance as evidenced by lower expression of insulin receptor phosphorylation sion of TXNIP mRNA, a pro-oxidant and pro-apoptotic protein is signifi cantly and hyper activation of Akt. Importantly, we found that expression of NGF elevated 2.3-fold in mouse and human islets cultured under high fat/high and its receptor TrkA, phospho-TrkA, p75, and MMP-7 which is necessary for glucose conditions, which was mitigated by CUR treatment. In conclusion, nerve growth factor signaling, were decreased in brain tissues of western the fi ndings of our study suggest that CUR is a potent, natural therapeutic diet mice. In addition, expression of PSD-95, a scaffolding protein enriched agent that can enhance insulin sensitivity in a multifaceted manner to pro- in post-synaptic densities, as well as insulin degrading enzyme, a principal tect against insulin resistance. regulator of amyloid β levels were also decreased. Stability of memory was also impaired in western diet mice brain as assessed by decreased expres- & 1939-P sion of cAMP response element-binding (CREB) protein. These results sug- A Novel Mechanism Mediating Diabetes and Obesity Protection gest that changes in the insulin sensitivity may contribute to cognitive with a Mechanism of Disruption of RGS14 and Increased Brown impairment associated with Western diet. Adipose Tissue Supported By: Alabama Agricultural Experimental Station DOROTHY E. VATNER, JIE ZHANG, HAIHONG ZONG, JEFFREY E. PESSIN, FREDRIC E. WONDISFORD, SALLY RADOVICK, STEPHEN F. VATNER, Newark, NJ, Bronx, NY, & 1937-P New Brunswick, NJ T1R2/T1R3 Is a Novel Target for Improving Insulin Sensitivity and Regulator of G protein signaling 14 (RGS14) knock out (KO) mice live 17% Glucose Metabolism longer than their wild type (WT) littermates. Since healthful aging is more YOSHIRO KITAHARA, SHIMPEI OGAWA, HARUKA KAWANABE, SEIJI KITAJIMA, important than simply aging, we examined protection against diabetes and KOJI OHSUMI, Kawasaki, Japan obesity, key components of healthful aging, in the RGS14 KO mice. Despite T1R2/T1R3 is a sweet taste receptor that is not only expressed in the taste similar food intake the RGS14 KO, compared to WT, had 15% lower body buds but also in various peripheral tissues. We investigated the role of T1R2/ weight, 28% lower adiposity index (based on weight of gonadal, retroperi- T1R3 in regulation of glucose metabolism by using T1R2 knockout (KO) mice. toneal, and inguinal fat pads), 47% increased brown adipose tissue and 32% We also synthesized T1R2/T1R3 antagonists and investigated the effect of reduction in the area under the glucose tolerance curve, all p<0.05. To con- these compounds on glucose metabolism in obese mice. There were no dif- fi rm protection against diabetes and obesity, the mice were fed a high fat ferences of body weight and fasting blood glucose between T1R2 KO mice diet for 3 months. With similar food intake on the high fat diet (HFD), the

Obesity and wild-type mice. However, glucose tolerance after an oral glucose load RGS14 KO, compared with WT, had 19% lower body weight and 28% lower

POSTERS was better in T1R2 KO mice than wild-type mice, although insulin secretion adiposity index, 24% lower area under the glucose tolerance curve, and 61% after an oral glucose load was comparable in both strains of mice. In primary increase in the insulin tolerance test, all p<0.05. RGS14 KO and WT on HFD

Integrated Physiology/ hepatocytes of wild-type mice, phosphorylation of Akt induced by insulin were also subjected to euglycemic-hyperinsulinemic clamps to determine (100 nM) was blocked by adding a T1R2/T1R3 agonist (1 mM sucralose), but the glucose infusion rate necessary to maintain euglycemia, which was this effect of sucralose was not seen in hepatocytes of T1R2 KO mice. In 150% greater in RGS14 KO along with 68% increase in whole-body glucose addition, activation of AMPK was detected in the gastrocnemius muscle of uptake. Of the organs tested, glucose uptake was increased the most in skel- T1R2 KO mice. Therefore, improvement of glucose tolerance in T1R2 KO mice etal muscle and heart of RGS14 KO. Thus, the RGS14 KO is a unique model of might be mediated via increased hepatic insulin sensitivity and activation of healthful aging, with protection against diabetes and obesity, via a potential AMPK in skeletal muscle. Then we synthesized T1R2/T1R3 antagonists and brown adipose tissue mechanism. investigated the effect of these compounds on glucose metabolism in KKAy mice and ob/ob mice. Blood glucose excursion after an oral glucose load was improved by a single dose of an antagonist (ASA19) in both KKAy mice and ob/ob mice. Administration of ASA19 to KKAy mice for 2 weeks caused

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A498 OBESITY—ANIMALCATEGORY

1940-P exposure were performed. Expression of factors involved in hypothalamic Top Models and Resources for Type 2 Diabetes Research from The feeding and brown/beige adipocyte thermogenic activity was performed by Jackson Laboratory real-time PCR and Western blot; MeDIP-sequencing was used to identify JIM YEADON, KATHY SNOW, ANDREW SCHILE, MELISSA OSBORNE, LAURE differential DNA methylation profi le in the hypothalamus of CB-O and STZ- CASE, CRYSTAL DAVIS, CATHLEEN LUTZ, MARJORIE STROBEL, Bar Harbor, ME O. The body weight of adult STZ-O was signifi cantly higher than CB-O. Daily Choosing the most appropriate model for diabetes research can be daunt- food intake and accumulated food intake during 12-hr refeeding after fasting ing. Many factors contribute to the complex disease presentation in humans was signifi cantly higher in STZ-O. After cold exposure, mRNA levels of ther- and currently no single mouse model recreates all aspects of the human mogenesis-related genes (UCP1, PGC1α, Cpt1b and Prdm16) and UCP1 pro- disease. Several mouse models can be utilized individually or jointly, how- tein levels were down-regulated in brown and subcutaneous white adipose ever, to provide a platform from which aspects of the human type 2 diabetes tissues from STZ-O. Furthermore, MeDIP-sequencing assay demonstrated phenotype, including insulin resistance, obesity, islet atrophy, neuropathy, paternal hyperglycermia altered overall methylome patterns in the hypo- nephropathy, infl ammation, dyslipidemia, impaired wound healing, and liver thalamus of adult offspring, with a large portion of differentially methylated steatosis can be studied. Comparisons between the most popular models genes focus on adipocytokine and endoplasmic reticulum protein process- for type 2 diabetes research, including the Lep ob/ob, Lepr db/db, and the ing pathways by GO and KEGG pathways analysis. Our study revealed that C57BL/6J diet induced obesity (DIO) mice, show that these models, depend- paternal hyperglycermia predisposes the offspring to developing obesity, ing on different genetic backgrounds and environmental conditions, are able which is possibly associated with altered hypothalamic methylome. to model various stages of the human disease from prediabetes through Supported By: National Program on Key Basic Research Project of China; phases I, II and III. Data comparing various metabolic and physical param- National Natural Science Foundation of China eters such as blood clinical chemistries, glucose tolerance, insulin resis- tance, body weight, body composition, food consumption, thermoregulation, 1943-P energy balance and behaviors, and the complications of diabetes (end-organ Reduced Liver-specifi c PGC-1α Expression Results in Increased system assays) are presented in the context of short and long term model 3-day High-Fat Diet-induced Weight Gain evaluation. E. MATTHEW MORRIS, COLIN MCCOIN, JOHN THYFAULT, Kansas City, KS Human and animal data suggest that a complex interplay of metabolic 1941-P factors infl uence susceptibility to acute high-fat diet (HFD)-induced weight A Neutralizing AgRP Antibody Reveals Leptin Dominates AgRP gain. Our previous work showed that a poly-genetic rat model of low aero- Effects on Feeding Behavior in Mice bic capacity displayed increased short-term HFD-induced weight gain due ARTHUR T. SUCKOW, JAMES TREVASKIS, MARION CORNU, FEENAGH KEYES, to both greater energy intake and feed effi ciency. This model also displayed PETER CARIUK, HAMID SALARI, SUSAN FOWLER, LEE BROWN, ARTHUR LEWIS, decreased hepatic fatty acid oxidation (FAO) and mitochondrial respiratory CARL WEBSTER, JOE GRIMSBY, ANISH KONKAR, Gaithersburg, MD capacity driven by decreased liver PGC-1α mRNA expression. Our fi ndings Agouti-related protein (AgRP) secreted from discrete neurons in the arcu- matched previous work showing that hepatic energy metabolism may regu- ate nucleus promotes food intake via MC4R receptors residing within the late energy intake and systemic metabolism. Herein, we examined if liver- paraventricular nucleus by acting either as a competitive antagonist of specifi c reductions in hepatic PGC-1α expression and expected reductions in the anorexigenic α-melanocyte stimulating hormone (α-MSH) or through hepatic FAO would increase short-term HFD-induced weight gain. Liver-spe- inverse agonism. Recent reports suggesting that increased AgRP tone drives cifi c, PGC-1α heterozygous (LPGC1a) and wild-type (WT) mice were placed in the hyperphagia and hyperglycemia evident in leptin-defi cient mice led us to indirect calorimeters, and fed open source low-fat diet (LFD, 10% fat, 3.5% hypothesize that the administration of an AgRP neutralizing antibody would sucrose, Research Diets) prior to the initiation of a 3 day HFD (45% fat, 17% result in pronounced weight loss and improved glucose control in ob/ob sucrose, D12451). As expected LPGC1a mice displayed reduced liver com- plete FAO and mitochondrial respiration of palmitoyl-carnitine (35 and 30%, mice. An antibody (AgRPAb) that binds to mouse AgRP [Kd=1.2 nM] and that prevents AgRP from blocking the effect of α-MSH on cells overexpressing respectively) compared to WT. 3-day HFD resulted in greater weight gain in MC4R was identifi ed. Intracerebroventricular (i.c.v.) delivery of AgRPAb prior both genotypes; however, HFD fed LPGC1a mice gained 50% more weight to i.c.v. dosing of AgRP completely inhibited its ability to promote food intake compared to WT. The greater HFD-induced weight gain in the LPGC-1a mice in DIO mice. A single dose of AgRPAb delivered either i.c.v. or via subcutane- tracked with a 30% greater feed effi ciency compared to WT. Only HFD fed ous injection did not signifi cantly inhibit food intake in ob/ob mice in an acute LPGC1a mice had greater food intake and demonstrated altered feeding pat- fast-refeed paradigm. Immunohistochemical analysis of AgRPAb distribution terns compared to LFD. While no signifi cant differences were observed in within the hypothalamus following an i.c.v. dose revealed it was present 4 energy expenditure and energy balance between genotypes, the WT dis- hours post-dose but, fully cleared by 24 hours. To understand the effects of played an appropriate HFD-induced reduction in respiratory quotient, while AgRP neutralization on body weight and glucose control and to overcome the this did not occur in the LPGC1a mice. Together, these data suggest that challenge of maintaining consistently high levels of antibody brain exposure liver-specifi c PGC-1α expression impacts HFD-induced weight gain through following peripheral or i.c.v. dosing, AgRPAb was delivered to ob/ob mice regulation of energy intake, feed effi ciency and substrate utilization. i.c.v. via an osmotic minipump for a period of 2 weeks. AgRPAb treatment did Supported By: National Institutes of Health; U.S. Department of Veterans not result in changes in food intake, body weight or glucose control relative Affairs; American Heart Association to a control antibody, whereas leptin delivered i.c.v. via osmotic minipump had pronounced effects on weight loss and glucose homeostasis. These 1944-P data indicate that AgRP neutralization by anti-AgRP antibody therapy can- Effects of Zinc Status on Obesity-induced Metabolic Disorder and not overcome the effect of leptin defi ciency on energy homeostasis, feeding Cardiac Hypertrophy in Young Mice behavior or obesity in mice. SHUDONG WANG, MANYU LUO, LU CAI, ZHIGUO ZHANG, Louisville, KY, Changc- hun, China 1942-P We investigated the effect of dietary zinc defi ciency and supplementation Paternal Hyperglycemia Reprograms Metabolic System and Exac- in mice consuming a high-fat diet (HFD) on metabolic disorder and cardiac erbates the Development of Obesity in Offspring hypertrophy using a murine model of childhood obesity. Childhood obesity Obesity XIAOQIU XIAO, XIAOQIN SHI, XINYU LI, YI HOU, XUEMEI CAO, HENG WANG, was induced in 4-week old mice fed with either HFD (60% kcal fat) or normal POSTERS HONGYIN WANG, CHUAN PENG, JIBIN LI, Chongqing, China diet (ND, 10% kcal fat) containing one of 3 different zinc quantities (low,

Previous studies showed that maternal obesity or diabetes exacerbated normal, and high zinc (ZD, ZN, and ZS are 10, 30, 90 mg zinc per 4057 kcal, Integrated Physiology/ the onset of obesity and other metabolic diseases in offspring. However, respectively) for 3 or 6 months. The mechanistic study was done in HFD- literatures as to transgenerational inheritance through male lineage are fed mice treated with and without p38MAPK inhibitor (SB203580). Results scarce. Our previous studies showed that paternal hyperglycermia exacer- showed that HFD induced signifi cant insulin resistance, hyperinsuline- bated the development of fatty liver in offspring. Current study investigated mia, hypertriglyceridemia, and cardiac hypertrophy. HFD also signifi cantly metabolic consequences of offspring born from hyperglycermic fathers. increased β-cell CLL/lymphoma 10 (BCL10)/caspase recruitment domain Male SD rats were ip injected with streptozotocin (STZ) or citrate buffer (CARD9) expression, p38 MAPK, and Nuclear factor kappa-B activation. (CB). STZ-injected rats with glucose levels higher than 16 mM were selected These obese effects were worsened in HFD/ZD mice and attenuated in HFD/ to breed with normal female rats. Offspring from STZ or CB treated fathers ZS mice, respectively. Immunoprecipitation results suggested the increase (STZ-O and CB-O) were maintained in the identical condition. We monitored in BCL10/CARD9 interaction in HFD group, which was further increased in body weight and food intake, and tests of fasting-refeeding and cold (4oC) the HFD/ZD group and attenuated in the HFD/ZS group. The cardiac patho-

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A499 OBESITY—ANIMALCATEGORY

logical changes were signifi cantly evidenced in a time-dependent manner 1947-P from 3 months to 6 months. Furthermore, inhibition of p38 MAPK, completely Fecal Proteomics Analysis Provides a Novel Window into the abolished HFD-induced, and even zinc defi ciency-worsened, cardiac effects Effects of Diet and Gut Microbiome on Host Gut Metabolism except for BCL10/CARD9 expression. In vitro time-course study showed that EMRAH ALTINDIS, SHIHO FUJISAKA, C. RONALD KAHN, Boston, MA, Toyama, palmitate treatment for 6-48 h increased the expression of BCL10 predomi- Japan nantly at 6-12 h, p38MAPK at 24-48 h, ANP at 48 h in primary cardiomyocyte. In both humans and rodents ingestion of a high fat diet (HFD) and the asso- These results suggest that HFD and zinc defi ciency synergistically induce ciated obesity alter systemic metabolism, in part through effects on the gut obesity-related systemic lipid profi le and cardiac hypertrophy probably via microbiome. We have recently shown that when obesity and diabetes-prone BCL10/CARD9 interaction with and activation of p38 MAPK-mediated mech- C57BL/6J mice on HFD are treated with the antibiotics vancomycin (Vanco) anism. Zinc supplementation prevented HFD-induced cardiac hypertrophy, or metronidazole (Metro), the gut microbiome is substantially altered result- suggesting the potential application of zinc to prevent cardiac hypertrophy ing in reduced infl ammation and improved insulin signaling and glucose in young obesity. metabolism. In the present study, we have explored the role of HFD and anti- Supported By: American Diabetes Association (1-15-BS-018 to L.C.) biotic treatment on the host gastrointestinal system by performing a quan- titative LC-MS/MS proteomics analysis of host proteins in fecal samples 1945-P of mice. In total, 236 host proteins could be identifi ed. These proteins fell Parkin Knockout Exacerbates Cardiac Dysfunction in High-Fat Diet- into 18 functional subgroups. Interestingly, only one host protein (prolactin induced Obesity through Interrupting Mitochondrial Dynamics receptor) was decreased in the HFD-fed animals, whereas 33 host proteins NAN HU, YINGMEI ZHANG, JUN REN, Laramie, WY, Shanghai, China were signifi cantly upregulated by feeding HFD. These included 6 protease Background: Obesity compromises cardiac function although the under- inhibitors, 5 proteases, 5 cell adhesion proteins and 4 proteins related to lying mechanism remains elusive. While mitochondria play a critical role lipid metabolism. The most up-regulated proteins were alpha-1-antitrypsin, in maintaining cardiac homeostasis, little is known with regards to mito- annexin A4, and serpin B6. Of the 33 upregulated proteins, 27 proteins were chondrial anomalies. Given the key role of mitophagy in the maintenance of further increased in Metro- or Vanco-treated HFD mice indicating effects of mitochondrial homeostasis, this study was designed to evaluate the impact both diet and microbiome. In order to identify the potential sites of release of of the mitophagy signaling molecule Parkin on high fat diet (HFD)-induced these proteins, we investigated their mRNA expression in liver, small intes- cardiomyopathy. tine and colon of the chow or HFD groups. qPCR analysis revealed signifi cant Method: Wild type (WT) and Parkin knockout (Parkin-/-) mice were fed increases for 11 of these 30 proteins in at least one host tissue, with jeju- low fat diet (LFD) or HFD for 20 weeks. Metabolic rate, glucose tolerance, num being the main site of production, exhibiting increases in mRNA for 8 echo, cardiomyocyte function, aconitase activity, ROS generation and mito- proteins. Taken together, these data indicate that the effects of HFD and chondrial respiration were assessed in WT and Parkin-/- mice fed LFD or the gut microbiome on the host intestinal proteome can be monitored in HFD. Western blot was performed to examine the possible cell signaling fecal samples. Understanding these changes in the fecal proteome will pro- pathway. vide a new window into defi ning the mechanisms underlying gut-microbiome Result: Parkin-/- failed to alter HFD-induced obesity and insulin resis- crosstalk in control of metabolism. tance, as evidenced by body weight gain and glucose intolerance. In addi- Supported By: National Institutes of Health tion, there was little difference in metabolic parameters (VO2, VCO2, RER, energy expenditure and physical activity) between WT and Parkin-/- mice 1948-P fed HFD. Nonetheless, Parkin-/- accentuated HFD-elicited cardiac con- Gastric Bypass Surgery in Mice Restores Glucose Tolerance but Is tractile dysfunction, as evidenced by echocardiographic, cardiomyocyte Accompanied by Endoplasmic and Cytosolic Stress in Islets contractile and intracellular Ca2+ properties. HFD feeding promoted more ROMAN VANGOITSENHOVEN, JOAO PAULO MONTEIRO CARVALHO MORI pronounced mitochondrial injury and ROS generation in Parkin-/- mice com- CUNHA, KATRIEN CORBEELS, MIRANDA VAN DER ENDE, MATTHIAS LANNOO, pared with WT mice. Mitochondrial respiration from HFD-Parkin-/- mice was ANN MEULEMANS, LUT OVERBERGH, CHANTAL MATHIEU, BART VAN DER reduced during ADP-dependent respiration compared with WT mice. HFD SCHUEREN, Leuven, Belgium down-regulated Fis1, Drp1, Parkin and Pink1 expression, denoting inhibited Diabetes remission after gastric bypass is well established, but the spe- mitochondrial fi ssion and mitophagy, the effect of which with exception of cifi c effects on islets of Langerhans remain controversial. Eight week old Pink1 was exacerbated by Parkin-/-. male B6 mice were fed a high fat diet for 14 weeks (OBESE) and subjected to Conclusion: Our present work revealed that Parkin-/- exacerbates HFD- Roux-en-Y gastric bypass (RYGB) or sham surgery. Sham operated mice were induced cardiac dysfunction and mitochondrial injury, possibly associated weight matched (WMS) or pair fed (PFS) to RYGB mice. Eight weeks post- with inhibition of appropriate mitochondrial fi ssion and cardiac autophagy. surgery, an intraperitoneal glucose tolerance test (IPGTT, n=6-8) was per- formed and ex vivo islet cell function was assessed by glucose stimulated 1946-P insulin secretion (GSIS, n=5-6), RNA sequencing (n=3) and pancreatic immu- nohistochemistry for insulin and Ki67, a proliferation marker (n=4). RYGB and WMS mice showed weight loss, and improved glucose tolerance compared WITHDRAWN to OBESE and PFS mice, but there were no signifi cant differences in ex vivo GSIS (Table 1). Pathway analysis of the islet transcriptome showed down- regulation of genes involved in beta-cell differentiation and proliferation in RYGB mice (Table 1). In all groups, Ki67 staining showed very low and simi- lar proliferation rates of islet cells and beta-cells specifi cally. Unexpectedly, endoplasmic reticulum and cytosolic stress genes were upregulated in RYGB mice (Table 1). In summary, although glucose homeostasis improves after RYGB surgery, islet function is unaltered ex vivo and the islet transcriptome suggests increased stress and decreased proliferation in RYGB mice.

Obesity Table 1. Metabolic and Islet Characteristics. POSTERS Integrated Physiology/

Mean±SD, *P < 0.05 vs. OBESE, $P < 0.05 vs. RYGB. Supported By: FWO G.0857.13

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A500 OBESITY—ANIMALCATEGORY

1949-P 1951-P Resistance to High Fat Diet-induced Obesity after Roux en-Y Gas- Roux-en-Y Gastric Bypass, but Not Vertical Sleeve Gastrectomy, tric Bypass Is Mediated by a Dynamic Program of Changes in Both Induces Intestinal Proliferation in Rodents Energy Expenditure and Calorie Intake PERNILLE WISMANN, PERNILLE BARKHOLT, DARLEEN A. SANDOVAL, NIELS SRIRAM MACHINENI, LEE HERZOG, JOSEPH BRANCALE, LEE M. KAPLAN, Bos- VRANG, JACOB JELSING, Hørsholm, Denmark, Ann Arbor, MI ton, MA, New York, NY, Charlestown, MA Today bariatric surgery is the most effective treatment option for obesity. High fat diets (HFD) cause alterations of multiple metabolic pathways Traditionally, Roux-en-Y gastric bypass (RYGB) has been the operation of that result in increased calorie intake (CI), decreased energy expenditure choice but most recently the less invasive vertical sleeve gastrectomy (VSG) (EE) and an increased fat mass. Despite substantial fat gain among diet- has become the preferred procedure. We have previously demonstrated that sensitive mice on a HFD, these mice become completely resistant to obesity RYGB leads to massive intestinal hypertrophy coupled with corresponding after Roux en-Y gastric bypass (RYGB) surgery. We sought to understand changes in endocrine cell numbers indicating that increased gut volume the physiological changes that drive this phenotype. Diet-induced obese could be responsible for the marked effects of surgery. However, the time C57BL/6 male mice underwent RYGB or a sham operation (SO) followed course for this effect remains unknown. Here we set out to investigate the by standard chow diet (SCD) to make the weights of the groups similar acute and chronic effects of both RYGB and VSG on intestinal volume in rats (32.8±1.1 and 34.2±1.7 g, respectively). Both groups of animals were then or mice. Rats were subjected to either RYGB or sham operation and termi- switched to HFD with close monitoring of energy balance. The RYGB group nated 10 and 60 days post surgery. Mice were subjected to either VSG or gained less weight over 2 weeks than the SO controls (1.1±1.7 vs. 8.1±1.2 g; sham operation and terminated 10 and 90 days post surgery. All intestines p<0.01). After accounting for the increased stool loss in RYGB (2.7±0.6 vs. were sampled using stereological principles and volume was estimated by 1.5±0.1 kcal/d in SO mice; p<0.01), RYGB mice increased net CI immediately point counting. Both RYGB and VSG surgery reduced body weight compared after switch to the HFD (7.9±1.5 to 11.8±2.8Kcal/d; p<0.01). When hourly EE to sham operation with weight loss apparent by day 10 post surgery. RYGB patterns were analyzed by deconvolution of random and circadian varia- surgery lead to a signifi cant increase in small intestinal volume both 10 and tions, the RYGB group exhibited a differential EE spike not associated with 60 days following surgery (1.3-fold increase at day 10; 2-fold increase at day increased physical activity, limited to the fi rst few days after switch to HFD. 60, p<0.001). In contrast, VSG operation in mice did not affect small intes- After 1 week, the continued resistance of RYGB mice to HFD-induced weight tinal volume, in neither the acute or chronic timeframe (831±81 vs. 862±30 gain was mediated primarily by a reduction in CI compared with SO animals mm3 at day 10; 652±37 vs. 712±45 mm3 at day 90). In conclusion, the data (8.7±0.8 vs. 12.7±0.9 Kcal/d; p<0.01). These observations demonstrate that demonstrates that the effects of VSG surgery on body weight is not directly the changes in CI and EE in response to dietary content evolve over time, and related to alterations in gut volume. that it is these dynamic changes that are most strongly modulated by RYGB. They suggest that multiple regulatory pathways are sequentially engaged 1952-P to mediate the changes in energy balance and adiposity induced by dietary miR-17 Upregulates 3T3-L1 Cell Differentiation via Repressing the content and bariatric surgery. Understanding these time-dependent effects Wnt Pathway Effector Tcf7l2 should facilitate the identifi cation of mechanisms most relevant to mimick- LILI TIAN, KEJING ZENG, ZHUOLUN SONG, WEIJUAN SHAO, JIANPING WENG, ing the effects of RYGB without surgery. BURTON B. YANG, TIANRU JIN, Toronto, ON, Canada, Guangzhou, China Supported By: Ethicon Endo-Surgery Obesity is a growing worldwide health problem, associated with increased risk of metabolic diseases, including diabetes. Understanding the molecular 1950-P mechanism of adipogenesis may lead to the discovery of novel therapeutic Tissue Specifi c Role of Orphan Nuclear Hormone Receptor Small targets for the treatment and prevention of obesity and diabetes. Wnt sig- Heterodimer Partner in the Development of Obesity and Diabetes naling is known to repress adipogenesis, while recent studies have shown YOONKWANG LEE, JUNG EUN PARK, MIKANG LEE, Rootstown, OH that a number of miRNAs are implicated in pre-adipocyte proliferation and Our early study reported that defi ciency of small heterodimer partner (SHP, differentiation. As a previous study indicated the stimulatory effect of miR- NR0B2) protected the mice from diet-induced obesity partly by increased 17/92 cluster on adipogenesis, we determined here that the fi rst member fatty acid oxidation. In addition, the SHP-/- mice displayed glucose intoler- of this cluster, miR-17, up-regulates mouse pre-adipocyte 3T3-L1 cell dif- ance due to impaired glucose-stimulated insulin secretion upon a 6-month ferentiation, as miR-17 precursor over-expression increased while miR-17 western diet regimen. Current study is aimed to defi ne specifi c roles of SHP inhibitor repressed 3T3-L1 differentiation. We then confi rmed the repressive in the development of obesity and diabetes furthermore. Using radiolabeled effect of the Wnt signaling pathway effector Tcf7l2 on adipogenesis and triacylglycerol, fecal analysis, and oil red-O staining, we found that SHP-/- revealed that Tcf7l2 is a miR-17 target, utilizing both the gain-of-function and mice had a signifi cant impairment in intestinal fat absorption. Gene expres- the loss-of-function approaches. The natural plant compound curcumin pos- sion (for example, pancreas specifi c transcription factor 1a (ptf1a), trypsin sesses the body weight lowing effect, and is a known suppressor of 3T3-L1 4 (try4) and pancreatic carboxyl ester lipase (cel) etc.) and fecal digestive differentiation. We found that curcumin attenuated miR-17 expression but enzyme activity indicated that pancreatic exocrine defi ciency is evident and stimulated Tcf7l2 expression in 3T3-L1 cells. Finally, high fat diet consump- may contribute to the observed lean and diabetic phenotypes in SHP-/- mice. tion in C57BL/6 mice increased miR-17 expression, associated with reduced We also assessed islet dysfunction using morphometric measurement of Tcf7l2 expression in mouse adipose tissue. Together, our observations sug- pancreatic sections and illumina beadchip array, which proved no signifi - gest that miR-17 is among the central switches of adipogenesis. It activates cant changes in size and population of islets, alpha cells, or beta-cells and in adipogenesis via repressing the Wnt signaling pathway, and its own expres- the expression of major genes involved in beta cell function. Nonetheless, sion can be nutritionally regulated in health and disease. along with impaired insulin secretion, glucose-stimulated Ca2+ infl ux was Supported By: Canadian Institutes of Health Research greatly diminished in SHP-/- islets. Interestingly, while the observed diabetic phenotype and pancreatic exocrine insuffi ciency in whole body SHP-/-mice 1953-P was recapitulated in the pancreas specifi c SHPKO (PSHPKO) mice, where The Effects of Metformin on Improving Obesity and Hepatic Steato- SHP deletion is confi rmed in both endocrine and exocrine tissues, the lean sis Are SIRT1-independent phenotype was reproduced only in liver-specifi c SHPKO mice but not in PSH- FEN XU, XIAOBIN ZHENG, HUA LIANG, JIANPING WENG, Guangzhou, China Obesity PKO mice. Even though detailed underlying molecular mechanisms for the Metformin is one of the most effective fi rst-line drugs to treat type 2 POSTERS observed phenotypes by SHP deletion remain to be elucidated, current data diabetes. Despite the wide use of Metformin, the mechanism of action of suggest that SHP plays distinctive and tissue-specifi c role in the develop- it has remained elusive. An earlier breakthrough study indicated that Met- Integrated Physiology/ ment of obesity and diabetes. formin led to the activation of the energy sensor AMPK to stimulate fatty Supported By: National Institutes of Health acid oxidation, impair fat synthesis, reduce gluconeogenesis of hepatocytes and thereby improve insulin sensitivity. An AMPK-interacted factor, SIRT1, also plays a vital role in the regulation of lipid metabolism and energy dis- sipation. We investigated the role of SIRT1 in the benefi cial effect of Met- formin on improving obesity and hepatic steatosis. In this study, eight-week old male SIRT1 heterozygous mice (SIRT1+/-) and their wild type littermates (WT) were subjected to a chow diet or a high fat diet (HFD) for 12 weeks. After the diet intervention, mice were randomly divided into groups as fol- lows: WT+chow diet, WT+HFD (WH), WT+HFD+Metformin (WHM), SIRT1+/-

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+chow diet, SIRT1+/-+HFD (SH), and SIRT1+/-+HFD+Metformin (SHM) group There was no signifi cant change in energy expenditure between SKIP-/- and for 6 weeks. The results showed that the body weight, fasting blood glucose WT. During 2 g/kg oral glucose tolerance test (OGTT), blood glucose levels (FBG), fat mass content and liver weight were statistically lower in WHM were lower in SKIP-/- at 15, 30 and 60 minutes; glucose-AUC was about 20% group comparing to WH group. decreased compared with WT. GIP secretion was about 1.5 fold increased Signifi cant reduction of hepatic lipid deposition was observed in WHM at 15 minute, and GIP-AUC was also about 1.5 fold increased in SKIP-/-. Six group according to the histology results. After heterozygous-knockout of g/kg OGTT resulted in a 1.4 fold increase in GLP-1 secretion of SKIP-/- com- SIRT1, we observed the same amelioration in SHM group with Metformin pared with WT. There was no difference in insulin sensitivity between treatment when compared with SH group. However, the body weight, FBG, SKIP-/- and WT. We also made double knock-in mice using SKIP-mCherry and fat mass content, liver weight and fat content were all higher in SIRT1+/- mice GIP-GFP mice (SKIP-/-GIP-/-). SKIP-/-GIP-/- gained less body weight than SKIP-/-, than those in WT mice under HFD feeding, regardless of Metformin use. whereas there was no difference in between SKIP-/-GIP-/-and WT. During 2 Collectively, the data indicate that the effects of Metformin on improving g/kg OGTT, SKIP-/- exhibited the highest GIP levels. SKIP-/- showed increased obesity and hepatic steatosis are independent of SIRT1. Quantitative real- adiponectin levels, while both SKIP-/- and SKIP-/-GIP-/- showed lower triglyc- time PCR revealed the expression of genes involved in fatty acid metabolism eride and total cholesterol levels. These results demonstrate that SKIP is and some lipoprotein-related genes were notably regulated after Metformin involved in incretin secretion, body fat composition and lipid metabolism. treatment in WT mice. Further study is necessary to uncover the relevant mechanisms in it. 1956-P 1954-P Sodium Glucose Transporter Type 2 Inhibitor, Empaglifl ozin, Im- WITHDRAWN proves Vascular Stiffness in Female Diabetic Mice Independent of Blood Pressure Reduction VINCENT G. DEMARCO, ANNAYYA AROOR, GUANGHONG JIA, RAVI NISTALA, MONA GARRO, GERALD MEININGER, LUIS MARTINEZ-LEMUS, ADAM WHALEY- CONNELL, ERIC MAYOUX, Columbia, MO, Ingelheim, Germany Vascular stiffness is considered to be an independent risk factor for the development and progression of cardiovascular disease. SGLT2 inhibitors (SGLT2i), which increase urinary glucose/sodium excretion to lower HbA1c and blood pressure (BP), are emerging as unique diabetes therapies and have been shown to reduce vascular stiffness. We examined whether the SGLT2i, empaglifl ozin (EMPA), ameliorates vascular stiffness and associated dysgly- cemia, microalbuminuria and hypertension in obese/diabetic female db/db mice. Eleven week old mice were fed a diet with or without EMPA (10mg/kg/ day) for 5 weeks. In vivo blood pressure (telemetry), aortic stiffness (pulse wave velocity, PWV) and renal artery resistivity were evaluated by ultra- sonography. Db/db had elevated BP that was not affected by SGLT2i. HbA1c, microalbuminuria and the renal resistivity index (RI) were elevated (P<0.001) in control (Db) mice vs. treated mice (Db-EMPA) and lean control mice. Ele- vated PWV and ex vivo aortic endothelial cell stiffness in Db were abrogated 1957-P with SGLT2i. Impaired acetycholine-induced vasodilation was improved in Dynamic Response in Plasma Metabolites Refl ect Effects of Diet, Db-EMPA. These results show that Empaglifl ozin ameliorates vascular stiff- Genetics, and the Gut Microbiome ness and dysfunction in obese diabetic female mice and these effects occur SHIHO FUJISAKA, JONATHAN M. DREYFUSS, CLARY CLISH, JULIAN A. independent of an effect on BP. PACHECO, LYNN BRY, ALEKSANDAR KOSTIC, C. RONALD KAHN, Toyama, Japan, Table. Established Vascular and Metabolic Abnormalities Are Improved with Boston, MA, Cambridge, MA SGLT2i in Diabetic Db/Db Mice. Diet, genetics and the gut microbiome are crucial factors in determining Blood Glycemia Vascular Stiffness Micro- individual metabolic status. This occurs, in part, through the production of Pressure albuminuria various metabolites by gut microbiota. We have recently shown that modi- SBP HbA1c Aortic PWV EC Stiffness Renal µAlb/Cre fying the gut microbiome with antibiotics (vancomycin or metronidazole) mmHg % m/s kPa Resistivity ratio improved diet-induced infl ammation and insulin signaling in C57BL/6J mice, but had no effect in 129 mice from either Jackson Labs (129J) or Taconic Con 136± 4 3.78± 0.15 3.20± 0.23 6.4± 0.7 0.58± 0.05 12± 2 Farms (129T). To explore the relationships between the metabolic pheno- Db 151± 3* 7.75±0.19* 4.50±0.30* 15.6± 2.0* 0.76±0.04* 231± 17* type, the gut microbiome and the other host factors, we performed LC-MS Db-EMPA 148± 4* 5.68±0.28† 3.65± 0.27† 3.9± 0.3† 0.62±0.05† 152± 14† based untargeted metabolomic analysis of the plasma of B6J, 129J and 129T *P<0.05 vs. lean Con, †P<0.05 vs. Db. mice on chow, high fat diet (HFD), and HFD with either vancomycin or metron- idazole and correlated these data with 16S rRNA sequence analysis of gut Supported By: Boehringer Ingelheim Pharmaceuticals, Inc. bacteria. Over 20,000 reproducible peaks representing low molecular weight (<1 kDa) metabolites could be detected in the serum of mice. Of these ~400 1955-P correspond to known metabolites and over 19,000 unknown metabolites. Sphingosine Kinase Type-1-interacting Protein (SKIP) Regulates Each strain of mice showed a unique pattern, with many plasma metabolites Incretin Secretion and Fat Composition showing 4- to 20-fold differences between strains of mice. This revealed YANYAN LIU, SHIN-ICHI HARASHIMA, YU WANG, NOBUYA INAGAKI, Kyoto, many unexpected changes in individual classes of metabolites associated

Obesity Japan with insulin resistance. For example, bile acid profi les were very different

POSTERS Gastric inhibitory polypeptide (GIP) and glucagon-like peptide-1 (GLP-1) among strains on both chow and further diverged on HFD. Likewise, some are the two primary incretins secreted from the intestine upon ingestion saturated and mono-unsaturated fatty acids went up on HFD in 129T mice,

Integrated Physiology/ of nutrients that stimulate insulin secretion from pancreatic b cells. Some but did not change or went down in B6J or 129J mice. On the other hand, common properties were reported in the secretion of the two incretin hor- short chain ceramides and C38 diacylglycerols went up on HFD in all strains, mones. We identifi ed SKIP, an A-kinase anchoring protein, as a molecule while long chain ceramides decreased. Both antibiotics further dramatically highly expressed in intestinal K and L cells by using GIP-GFP and GCG-GFP changed the bile acid and other metabolites. These changes strongly corre- knock-in mice. Immunohistochemical analysis of C57BL/6 mice showed that lated with specifi c bacterial taxa in the gut and their metagenomic content. SKIP is located in K and L cells. We have made SKIP-mCherry knock-in mice These data demonstrate the dramatic impact that diet, host genetics and (SKIP deletion, SKIP-/-) by inserting the mCherry gene into the SKIP gene. We the gut microbiome can have on plasma metabolites, and how these can confi rmed that mCherry was expressed in intestinal epithelial cells by immu- contribute to regulation of metabolic status. nohistochemistry. Starting from 5 weeks of age, SKIP-/- showed more body weight gain than WT (SKIP+/+). CT scan disclosed a 2.5 fold increase in visceral fat and a 1.6 fold increase in subcutaneous fat, but no fatty liver in SKIP-/-.

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A502 OBESITY—ANIMALCATEGORY

1958-P also communicates with the behavioral and activity centers of the brain. This Adiponectin Ameliorates Aging Phenotypes and Extends Life Span study investigated the possibility that inhibition of SuMN neurons (SuMNi), in Klotho-defi cient Mice including DA projections to SCN may change SCN interpretation of light and SHUICHI OTABE, XIAOHONG YUAN, TOMOKA FUKUTANI, KAYO TANAKA, HIT- induce MS. Rats maintained on regular chow and daily 14:10 LD cycles were OMI NAKAYAMA, YUJI TAJIRI, KENTARO YAMADA, Kurume, Japan infused at SuMN during the end of the daily light period (circadian time [CT] We have reported that the transgenic overproduction of adiponectin 10:00 to 14:00) with either vehicle or GABAa agonist/AMPA antagonist/ extends the lifespan of metabolic syndrome mice through the attenuation NMDA antagonist (SuMNi cocktail) (muscimol/CNQX/d-AP5: 3.6/1.8/120 of chronic infl ammation. However, the mechanism of the longevity effect nmol per day) to inhibit SuMN activity prior to being released into constant remains to be fully elucidated. Here we assessed the effect of hypera- darkness for 24 hours. Rats were then exposed to a light pulse (30 min, ~200 diponectinemia on a premature-aging syndrome in Klotho-defi cient mice. lux) at CT 15:00 and the brains collected at CT 16:00 and analyzed for SCN Klotho-defi cient mice stop growing at ~3 weeks of age and hardly gain body c-Fos immunohistochemistry. The SuMNi cocktail markedly reduced light- weight until they die around 8 week of age showing osteopenia, skin atrophy, induced c-Fos expression at the SCN core, but not the shell, region relative and impaired cognitive function. We created a Klotho-defi cient mouse line to controls (73%, p= 0.02). The SuMNi cocktail effect on peripheral metabo- expressing human adiponectin in the liver by repeated backcrossing more lism was investigated in high fat diet resistant (HFDR) rats. HFDR rats were than 20 generations. We have confi rmed that human adiponectin is biologi- infused at SuMN through an osmotic pump for 2 weeks with either vehicle or cally active in mice. By using a human adiponectin transgene, endogenous SuMNi cocktail. Such SuMNi increased HOMA-IR (169%, p=0.04), decreased and transgenic adiponectin could be separately measured. The plasma levels Matsuda insulin sensitivity index (49%, p = 0.023), increased body weight of endogenous adiponectin at 7 weeks of age were 71.1±13.9 and 82.2±2.3 gain (8%, P=0.001), retroperitoneal fat (69%, p=0.005), food consumption µg/mL in male and female Klotho-defi cient mice, respectively. The concen- (32%, p < 0.001), plasma leptin (163%, p =0.0015), and plasma triglyceride tration of human adiponectin in 7-week old male and female adiponectin- (75%, p=0.021). These fi ndings are the fi rst to identify a neuronal mechanism transgenic Klotho-defi cient mice was 648.6±426.0 and 839.8±592.9 µg/mL, that can alter the SCN clock interpretation of light and induce MS. respectively. Furthermore, endogenous adiponectin was also increased in adiponectin-transgenic mice; 189.6±36.8 and 210.6±48.6 µg/mL in males 1961-P and females, respectively. Male and female adiponectin-transgenic mice TCTP Modulates Lipid and Glucose Homeostasis in Mice by Sup- lived for 78.9±26.5 days (n=36) and 97.8±30.4 days (n=36), respectively. The pressing Hepatic Adiponectin Signaling and FGF-21 Expression life spans were signifi cantly longer than those of male and female non-trans- ZHE DAI, DELING ZHANG, KUN DONG, FENG DONG, XIAOBAN XIN, MUHAM- genic Klotho-defi cient mice; 53.9±10.0 (n=33) and 54.4±11.8 days (n=36), MAD ABDUL-GHANI, WEIPING JIA, FENG LIU, LILY Q. DONG, San Antonio, TX, respectively (both p<0.0001). Although no difference was observed in oxida- Shanghai, China tive stress markers or TUNEL staining in the heart, liver, kidney and brain, APPL1 and APPL2 act as integrated Yin-Yang machinery to modulate adi- aging phenotypes such as osteopenia and skin atrophy were ameliorated in ponectin pathway. To elucidate the mechanisms underlying the action of adiponectin-transgenic mice. Thus, hyperadiponectinemia may be protective APPL1/2, we screened a yeast two-hybrid cDNA library using APPL2 as bait. against premature death independent of the effect on metabolic syndrome. This screening led to identifi cation of TCTP (Translationally Controlled Tumor Protein) as a binding partner of APPL2, which is confi rmed in hepatocytes 1959-P by pull-down and co-immunoprecipitation experiments. The interaction with Neuronal DNA Methyltransferase 1 Defi ciency Attenuates Diet- TCTP enhances the binding of APPL2 to adiponectin receptors and thus inhib- induced Obesity its adiponectin signaling. To determine whether TCTP negatively regulates EMILY C. BRUGGEMAN, HANG SHI, BINGZHONG XUE, Atlanta, GA adiponectin signaling and function in vivo, we generated liver-specifi c TCTP Aberrant neuronal DNA methylation patterns have been implicated in knockout mice (TCTPLKO mice). Deletion of TCTP in liver enhanced hepatic adi- obesity development, but the role of neuronal DNA methyltransferases ponectin and insulin signaling, increased fatty acid oxidation, and reduced (Dnmts; enzymes that catalyze DNA methylation) in energy balance remains gluconeogenesis in mouse liver. TCTPLKO mice display increased resistance poorly understood. We investigated the role of neuronal Dnmt1 in normal to high fat diet-induced obesity, liver steatosis, hypoadiponectinemia, hyper- energy regulation and obesity development using a novel Dnmt1 knockout glycemia, and hyperlipidemia compared to fl oxed control mice. In addition, mouse model, Dnmt1fl / fl Synapsin1Cre (ND1KO), which specifi cally deletes liver-specifi c knockout of the tctp gene greatly increased hepatic insulin Dnmt1 in neurons. ND1KO and fl /fl control littermates were fed either a sensitivity as determined by hyperglycemic-euglycemic clamp studies. Inter- normal chow diet or a high fat diet (HFD). We found that neuronal Dnmt1 estingly, the TCTPLKO mice show signifi cantly increased serum and hepatic defi ciency attenuated HFD-induced obesity by reducing food intake and FGF-21 levels compared to the Loxp littermates. Suppressing hepatic FGF-21 increasing energy expenditure in male mice. In addition, HFD-fed ND1 KO expression reduced insulin sensitivity, increased triglyceride and free fatty mice had improved insulin sensitivity as measured by an insulin tolerance acid levels, and decreased hepatic fatty acid oxidation in the TCTPLKO mice. test. HFD-fed ND1KO mice had smaller fat pads and an upregulation of ther- Taken together, our study uncovers TCTP as a critical regulator of hepatic mogenesis-associated genes in brown adipose tissue. Further, ND1KO mice insulin sensitivity and glucose/lipid homeostasis. In addition, TCTP nega- had evidence of increased browning of white adipose tissue during a 7-day tively regulates hepatic glucose and lipid homeostasis by inhibiting adi- cold challenge. These data suggest that neuronal Dnmt1 deletion increased ponectin signaling and hepatic FGF-21 expression. diet-induced thermogenesis and cold-induced adipocyte browning, which Supported By: DK102965 may explain the lean phenotype in ND1KO mice. Interestingly, we found that ND1KO mice had elevated estrogen receptor-α (ESR1) gene expression in the 1962-P dorsomedial region of the ventromedial hypothalamus (VMH). ESR1 expres- Myeloid Cell-specifi c HIF-1α Deletion Decreases the Number of sion in the VMH promotes a lean phenotype in both male and female mice, Liver Tumors in High Fat-fed Mice and ESR1 DNA methylation has been linked to several diseases. Thus, ESR1 AKIKO TAKIKAWA, ISAO USUI, QUN ZHANG, KEISUKE OKABE, TOMONOBU in VMH may be a promising target mediating the obesity-protective effects KADO, ALLAH NAWAZ, SHIHO FUJISAKA, TAKASHI NAKAGAWA, KOICHI of neuronal Dnmt1 defi ciency. We conclude that neuronal Dnmt1 regulates TSUNEYAMA, KAZUYUKI TOBE, Toyama, Japan, Tokushima, Japan

energy balance, which may be mediated in part by ESR1 expression in the Obesity and diabetes are known to increase the incidence or growth of Obesity dorsomedial VMH. cancer. However the mechanisms are not fully clarifi ed. Macrophages resid- POSTERS ing inside and/or outside of cancers affect the occurrence and growth of the

1960-P cancer. In this study, we investigated the role of HIF-1α in macrophages in the Integrated Physiology/ Inhibition of the Supramammillary Nucleus Activity Alters Clock occurrence and growth of liver cancer in high fat-fed mice. To induce the occur- Photosensitivity and Induces Metabolic Syndrome rence of liver cancer, diethylnitrosamin (DEN), a carcinogenic substance, was YAHONG ZHANG, MICHAEL EZROKHI, SCOTT WATTERS, CHRISTINE CARDI, intraperitoneally injected to macrophage-specifi c HIF-1α-deleted mice and ANTHONY CINCOTTA, Tiverton, RI wild type controls. These mice were fed on high fat diet (HFD) or normal chow Seasonal induction of metabolic syndrome (MS) is driven in large part by diet from 6 weeks of age for 6 months. HIF-1α deletion did not affect body reduction in circadian peak dopaminergic (DA) input to the peri-suprachias- weight and liver weight. HFD treatment for 6 months increased the number, matic nuclei (SCN) (the clock pacemaker) without any change in photoperiod incidence and size of DEN-induced liver tumor. Macrophage-specifi c HIF-1α (PP). An endogenous mechanism to change the SCN interpretation of the PP deletion decreased the number of liver tumor by about 50%, while it did not and alter metabolism has not been elucidated. The DA input to the SCN origi- affect the incidence and size of the tumors in HFD-fed mice. These results nates primarily from the Supramammillary Nucleus (SuMN), a center that suggest that HIF-1α in macrophages is involved not in the growth but in the

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A503 OBESITY—ANIMALCATEGORY

occurrence of liver tumors. To know the mechanisms of how HIF-1α deletion 1965-P decreased the occurrence of liver tumors, we fi rst examined macrophage polarity. The expression of both M1 macrophage markers, such as CD11c and WITHDRAWN IL-1β, and M2 markers, such as CD206, was inhibited by HIF-1α deletion. Then, samples were separately collected from tumor area and non-tumor area of the liver. mRNA expressions of several genes related to cancer growth, such as IGF2 and EGFR, were inhibited by HIF-1α deletion. Although many mac- rophages were observed both inside and outside of the tumors, just a few cells were stained with antibody against pimonodazole, a hypoxia probe, in the liver of HFD-fed mice. These results suggest that HIF-1α in macrophages plays a stimulatory role in the initiation of liver tumor, where unknown factors other than tissue hypoxia may activate macrophage HIF-1α.

1963-P C2 Domain-Containing Protein CDP138 Plays an Important Role in Energy Balance and Fat Browning through Regulating Adrenaline Release QIONG ZHOU, YE SONG, JUN-YUAN HUANG, ANDRIA G. SHARMA, KAVIN ZHU, ZHANGPING LIAO, ZHEN Y. JIANG, Boston, MA CDP138 is a calcium- and lipid-binding protein that is known to be involved in membrane traffi cking. Here we report that mice without CDP138 develop obesity under both a normal chow diet (NCD) and a high fat diet (HFD) condi- tions. CDP138-/- mice have lower energy expenditure, oxygen consumption and body temperature in comparison with Wild-Type (WT) mice. Interest- 1966-P ingly, CDP138 is highly expressed in the medulla, but not cortex, of the adre- Liraglutide Protects Diet-induced Obesity through Induction of nal grand and is required for cold exposure-induced adrenaline secretion into Brown Adipogenesis in Mice the circulation. In addition, tissue cyclic adenosine monophosphate (cAMP) JOSEPH ZHOU, PRASHANTH CHANDRAMANI-SHIVALINGAPPA, LIXIN LI, Mon- levels, hormone sensitive lipase (HSL) phosphorylation, and adipose triglyc- treal, QC, Canada, Mount Pleasant, MI eride lipase (ATGL) protein expression were decreased in CDP138-/- mice. Research has shown that glucagon like peptide-1(GLP-1) agonist induces Furthermore, thermogenesis and inguinal fat browning in cold condition weight loss in type 2 diabetes subjects, the underlying mechanisms remains were also decreased in CDP138-/- mice. Our data indicate that CDP138 is a unclear. Brown adipocyte tissue (BAT) plays a major role in controlling novel regulator of acute stress response and plays an important role in lipid energy balance in rodents. The role of GLP-1 on the differentiation of BAT metabolism through regulating adrenaline secretion from adrenal grand. has not been studied. As such, we aimed to determine the protective effect Supported By: American Diabetes Association (7-11-BS-72 to Z.Y.J.); National of liraglutide, a full agonist of the GLP-1 receptor, on diet induced obesity. Institute of Diabetes and Digestive and Kidney Diseases Mice were fed with either chow diet or high fat high sucrose diet (HFHSD); liraglutide or saline were injected daily for fi ve weeks. Liraglutide signifi - 1964-P cantly attenuated weight gain along with epididymal fat mass in both diets PAS-domain-containing Protein Kinase (PASK) Is Required for the groups. Furthermore, improved oral glucose tolerant test was also observed Regulation of Food Intake and Glucose Homeostasis in liragltuide treated mice. Liraglutide had a transient suppression on food ANGELES MONDRAGON, DAVID J. HODSON, FRANCESCA SEMPLICI, TAKAHISA intake, which indicates the reduced caloric intake plays minor role on the MIKAMI, RYOKO ISHII, GUY A. RUTTER, GABRIELA DA SILVA XAVIER, London, weight loss. Importantly, brown fat-specifi c gene, uncoupling protein-1 (UCP- United Kingdom, Tokyo, Japan 1), was induced in white fat tissue (WAT) by liraglutide treatment. Further- Aims/Hypothesis: To explore the role of PAS-domain containing protein more, liraglutide, signifi cantly induced expression of several genes involved kinase (PASK) in energy homeostasis. in brown fat differentiation; including the PR domain-containing protein-16 Methods: We examined global (Pask-KO) and islet β-cell specifi c Pask null (PRDM16), a key drivers of brown fat differentiation and peroxisome pro- mice (βPask-KO). Intraperitoneal glucose tolerance was followed over circa- liferator-activated receptor-α (PPAR-α). Peroxisome proliferator-activated dian time in Pask-KO and βPask-KO mice. Open circuit indirect calorimetry receptor-coactivator-1-α (PGC-1 α) which regulates mitochondrial biogen- (CLAMS), whole body composition (EchoMRI), and gene expression analysis esis was also upregulated. Our results indicated that lirglutide activate the (real-time quantitative PCR) were performed. brown fat gene program and induce beige fat in WAT. In addition, HFHSD Results: Pask-KO mice displayed elevated (p<0.05) fasting glucose, and associated with an increase in caspase-3 expression, a key effector cas- increased feeding frequency and overall food intake (1.6±0.07-fold, p<0.05), pase that performs the apoptotic program in WAT. Liraglutide attenuates vs. wild-type littermate controls, whilst βPask-KO mice displayed only mild the HFHSD induced upregulation of caspase-3. Our study demonstrated that glucose dyshomeostasis. Glucose homeostasis, and circadian gene (Clock the protective effect of liraglutide on diet-induced obesity may act through and Bmal1) expression in pancreatic islets, were shifted by 12 h vs. control induction of beige fat which lead to elevation of energy expenditure. Lira- in Pask-KO mice but not βPask-KO mice. Circadian expression of the Ins2 glutide is a potential therapy for metabolic syndrome. gene was disrupted in global Pask-KO, and to a lesser extent in βPask-KO, Supported By: Central Michigan University islets. Disrupted Clock gene expression was also observed in the hypothala- mus of global Pask-KO mice. The glucagon-like peptide-1 (GLP-1) analogue, 1967-P liraglutide, failed to suppress food intake or lower blood glucose in these Differential Regulation of Metabolic and Protein Synthesis Path- Pask-KO mice. ways in Mouse Embryonic Fibroblasts and Induced Pluripotent

Obesity Conclusions/Interpretations: Our observations suggest an important role Stem Cells from Leptin Receptor Defi cient Mice POSTERS for PASK in the the modulation of food intake in response to incretins, and MANOJ K. GUPTA, HEIDRUN VETHE, VILAS WAGH, JUN SHIRAKAWA, SAMIR energy homeostasis. SOFTIC, HARALD BARSNES, MARC VAUDEL, TOMOZUMI TAKATANI, RACHEL

Integrated Physiology/ Signifi cance Statement: Our work demonstrates that the protein PASK is MARTINEZ, JIANG HU, HELGE RAEDER, ROHIT N. KULKARNI, Boston, MA, Ber- involved in the regulation of 24 h glucose homeostasis and food intake, i.e., gen, Norway that PASK is a regulator of circadian energy homeostasis. We also show that Leptin is a central regulator of metabolism, however, the role of leptin sig- PASK mediates the anorectic effect of the widely used antidiabetic drug, the naling in linking pluripotency with growth and development and metabolic GLP-1 mimetic, liraglutide, i.e., PASK is a potential drug target for the treat- disease progression is poorly understood. In the present study, we explored ment of diabetes and obesity. the relevance of the leptin receptor (LepR) in pluripotency and metabolism Supported By: Diabetes UK; European Foundation for the Study of Diabetes; UK using embryonic day (ED) 14.5 fi broblasts (MEFs) and their reprogrammed Medical Research Council induced pluripotent stem cells (iPSCs) from mice carrying the db/db mutation (LepRdb/db). A global quantitative proteomics approach, revealed key path- ways regulating pluripotency, metabolic homeostasis and protein synthesis during fetal growth and development. LepRdb/db MEFs exhibited metabolic

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A504 OBESITY—HUMANCATEGORY abnormalities and mitochondrial dysfunction as compared to control (Ctrl) glucose and glucose area under the curve response correlated negatively MEFs (n=3, p<0.05), while LepRdb/db iPSCs demonstrated altered Oct4 and with α-amylase (r=-0.72, P<0.01; r=-0.66, P<0.01; r=-0.79, P<0.01, respec- Stat3 pathways, which are involved in normal embryonic development, and tively) and positively with lipase (r=0.54, P<0.01; r=0.45, P=0.03; r=0.43, upregulation in the expression of the elongation initiation factor 4e (eIF4e) P=0.04, respectively). These data suggest that RYGB normalizes the pancre- implicating enhanced protein synthesis (n=3, p<0.05). Interestingly, ChIP atic exocrine function at the gene level, and this adaptation may contribute analysis revealed higher Stat3 binding on the eIF4e promoter in LepRdb/db to improved glycemic control after RYGB. iPSCs and polysome/monosome profi ling suggested higher protein synthesis Supported By: Cleveland Clinic (1009); National Institutes of Health (U24 compared to Ctrl iPSCs. Finally, correction of the point mutation in LepRdb/db DK076174) iPSCs using CRISPR/Cas9 gene editing reversed the alterations in pluripo- tency, metabolic changes and protein synthesis. These data provide evi- dence for a role of the LepR in regulating metabolic properties and key devel- OBESITY—HUMAN opmental pathways in embryonic fi broblasts and protein synthesis pathway and pluripotency of induced pluripotent stem cells. Moderated Poster Discussion: Bariatric Surgery, Bypass Liners, and 1968-P Blood Glucose in Humans (Posters: 1970-P to 1977-P), see page 21. Metabolic Burden in Mice of Erythropoietin Stimulation HEATHER M. ROGERS, LI WANG, CONSTANCE TOM NOGUCHI, Bethesda, MD, & 1970-P Macau, China Increased Postprandial Glucagon-like Peptide-1 Responses in Erythropoietin (EPO), the cytokine required for erythrocyte production, Obese Patients with Type 2 Diabetes Implanted for 26 Weeks with promotes erythroid differentiation and increases glucose uptake in eryth- the EndoBarrier Gastrointestinal Liner roid progenitor cells in culture. In adult male C57BL/6 mice, EPO stimulated ULRICH ROHDE, CECILIE A. FEDERSPIEL, EBBE LANGHOLZ, PETER VILMANN, erythropoiesis, reduced blood glucose and decreased body weight, suggest- STEFFEN U. FRIIS, JENS J. HOLST, TINA VILSBØLL, FILIP K. KNOP, Hellerup, Den- ing a metabolic burden associated with increased erythropoiesis. Decrease mark, Herlev, Denmark, Copenhagen, Denmark in body weight/fat mass has also been linked to EPO non-hematopoietic The endoscopically implanted and reversible duodenal-jejunal bypass response. Mice with EpoR restricted to erythroid tissue and Δ EpoRWAT sleeve, the EndoBarrier, results in weight loss and may improve glycemic mice with targeted deletion of EpoR in white adipose tissue (WAT) have control in type 2 diabetes (T2D). We investigated the effect of the EndoBar- increased blood glucose level and are glucose intolerance with an age rier on postprandial glucose tolerance and responses of the incretin hor- dependent increase in obesity and insulin resistance. The metabolic burden mones glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic associated with erythropoiesis is evident in young mice (12 weeks and 16 polypeptide (GIP) and the pancreatic hormones insulin and glucagon. Before weeks) treated with EPO (3000U/kg for 3 weeks), which increases hema- (Pre), one week (1 w) and 26 weeks (26 w) after implantation, 10 normal glu- tocrit, decreases blood glucose and improves glucose tolerance without cose tolerant (NGT) persons (age: 49 (16) years; body weight (BW): 101.1 (21.7) 2 change in body weight in wild type and Δ EpoRWAT mice. In contrast, in older kg; body mass index (BMI): 34, 3 (3.7) kg/m ; glycated hemoglobin (HbA1c) 31 mice at 8 months with increasing fat mass, EPO treatment increases hemat- (5) mmol/mol) and 9 matched T2D patients (age: 50 (23); BW: 103.7 (33.5) kg; 2 ocrit and glucose tolerance, and decreases body weight by 14% in wild type BMI: 38.6 (6.7) kg/m ; HbA1c: 49 (12) mmol/mol)) underwent a 4-hour mixed mice, in contrast to Δ EpoRWAT mice with increased hematocrit and mini- liquid meal test. Data are median values with interquartile range in brack- mal change in body weight, indicating that the non-hematopoietic metabolic ets. NGT and T2D subjects lost 6.8 (11.9) and 6.2 (5.8) kg, respectively. In response is mediated in part via EpoR expression in WAT. In older mice, this both groups, HbA1c remained unaffected. Fasting glucose, GLP-1, GIP, insulin, non-hematopoietic EPO response is dominant compared with the metabolic C-peptide and glucagon did not differ between Pre, 1 w and 26 w within both burden associated with EPO stimulated erythropoiesis. These data suggest groups. Postprandial GLP-1 responses (incremental area under curve (iAUC)) that the metabolic burden associated with EPO stimulated erythropoiesis among T2D patients, but not NGT subjects, were signifi cantly higher at 1 w results in decreased blood glucose and improved glucose tolerance and that compared to Pre (iAUC: 1330 (1078) vs. 2230 (3618) pM×min, P=0.0208) and in older mice with increased fat mass, EPO treatment also decreases body remained elevated at 26 w compared to Pre (2685 (1343) pM×min, P=0.0004). weight due in part to EpoR expressing WAT which can dominate the meta- Within each group, postprandial glucose, GIP, insulin, C-peptide and gluca- bolic response. gon excursions were similar on the three study days. Despite increased post- Supported By: National Institute of Diabetes and Digestive and Kidney Diseases prandial GLP-1 responses and weight loss in T2D patients after EndoBarrier implantation, postprandial glucose metabolism was not improved. 1969-P Normalization of Pancreatic Exocrine Function Is Associated & 1971-P with Better Glycemic Control following Roux-en-Y Gastric Bypass Postextractional Durability of Endocrine and Metabolic Effects of (RYGB) in Diabetic Rats Endoscopic Duodenal-Jejunal By-Pass Liner in Obese Type 2 Dia- ANNY MULYA, AMANDA R. SCELSI, J. DAVID MOSINSKI, ALI AMINIAN, betic Patients 3 Months after Its Removal ESAM BATAYYAH, HAZEL HUANG, OLIVIA DAN, PHILIP R. SCHAUER, STACY A. PETRA KAVALKOVA, MILOS MRAZ, PAVEL TRACHTA, DENISA HALUZIKOVA, BRETHAUER, JOHN P. KIRWAN, Cleveland, OH ZDENKA LACINOVA, MAREK BENES, ZUZANA VLASAKOVA, TOMAS PETR, LIBOR RYGB surgery re-establishes glycemic control in obese patients with type 2 VITEK, TEREZIE PELIKANOVA, MARTIN HALUZIK, Prague, Czech Republic diabetes; however, the mechanism remains unclear. Abnormal pancreatic Duodenal-jejunal by-pass liner (DJBL) is an endoscopically implantable function is a key determinant of type 2 diabetes. We hypothesized that RYGB device designed to mimic the effects of gastrointestinal by-pass opera- would cause molecular adaptations in pancreatic digestive enzymes that tions. It has been suggested that some effects of DJBL may persist after would correlate with improved glucose metabolism. Twenty, adult Zucker its removal. The aim of our study was to assess the effects of DJBL on body Diabetic Fatty (ZDF) rats were randomized into Sham (N=6) or RYGB (N=7) composition, glucose control and metabolic and hormonal profi le of subjects surgery, or pair fed to the RYGB (N=7) group. Zucker nondiabetic rats (N=3) with obesity and type 2 diabetes mellitus (T2DM) throughout implantation were used as Controls. At 30 days post-surgery, body weight and OGTT and after its removal. Thirty obese patients with T2DM (22 males, age 51.8 ± Obesity were assessed and the pancreas was harvested for subsequent qRT-PCR 1.8 years) underwent the implantation of the EndoBarrier DJBL (GI Dynam- POSTERS on genes that code for pancreatic digestive enzymes (α-amylase, lipase ics, U.S.) and a comprehensive metabolic and hormonal evaluation 1, 6 and and chymotrypsinogen). Post-intervention, the Sham group had signifi cantly 10 months after implantation and 3 months after its removal, respectively. Integrated Physiology/ higher body weights than RYGB (P<0.01) and Pair-Fed (P=0.03) groups. The Postprandial effects of DJBL were evaluated during a 2-hour liquid meal RYGB group had better glucose control than the Sham and Pair-Fed groups, test (LMT). Over 10 months the implantation of EndoBarrier decreased body primarily driven by a decrease in fasting glucose (P<0.05). There was no sig- weight (129.7 ± 4.4 vs. 117.3 ± 4.3 kg, p<0.05) and improved glucose control nifi cant difference in chymotrypsinogen mRNA between Sham, RYGB and (fasting blood glucose 12.3 ± 0.7 vs. 9.1 ± 0.7 mmol/l, p<0.05; HbA1c 75.0 ± Pair-Fed groups. Pancreatic α-amylase (P<0.01) was 5 and 1.5 fold higher 3.4 vs. 55.4 ± 3.8 mmol/mol, p<0.05). These effects were accompanied by in the RYGB (0.6+/-0.05) compared to Sham (0.1+/-0.06) or Pair-Fed (0.4+/- increased concentrations of a potent regulator of bile acid synthesis-fi bro- 0.1) groups. Pancreatic lipase (P<0.01) was reduced by 52 and 57% in RYGB blast growth factor 19 (FGF-19) and circulating bile acid levels as well as (0.8+/-0.08) compared to Sham (1.7+/-0.1) or Pair-Fed (1.8+/-0.4) groups, enhanced serum GLP-1, while glucagon levels were reduced. Changes in respectively. RYGB normalized both pancreatic α-amylase and lipase gene FGF-19, bile acids, glucagon and GLP-1 concentrations were completely lost expression when compared to Controls. Fasting blood glucose, the 2-hour 3 months after DJBL removal. HbA1c worsened from 55.4 ± 2.5 to 61.1 ± 3.3

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A505 OBESITY—HUMANCATEGORY

mmol/mol and body weight increased from 117.3 ± 4.3 to 120.3 ± 4.5 kg, higher in those with T2D duration < 8 yrs (61.1 vs. 38.9% in > 8 yrs). In surgi- but both parameters still remained markedly lower than the baseline val- cal groups combined (n= 97), baseline BMI was lower in the uncontrolled vs. ues. We conclude that the implantation of DJBL decreased body weight and controlled glycemia groups (35.8 ± 3.8 vs. 37.1 ±3 kg/m2, P < 0.05). All three improved glucose control most likely through increased FGF-19, bile acids glycemic status groups had similar baseline and 1 yr post intervention body and GLP-1 and decreased glucagon concentrations. While the improvements weights. However, markedly higher post-surgical body weight was noted in in body weight and glucose control partially persisted, all hormonal changes the relapse vs. controlled glycemia groups at 2 yrs (83.6±16 vs. 75±11 kg, P = were lost 3 months after DJBL removal. 0.03) and was not different between the controlled vs. uncontrolled groups. Supported By: RVO-VFN64165; AZV 15-26854A, AZV 15-27863A; In summary, refractory hyperglycemia post bariatric surgery is dependent SVV260019/2014 on procedure type (SG), duration of disease (>8 yrs) and lower baseline BMI. However, inadequate surgical weight loss at 2 yrs is associated with relapse & 1972-P in glycemic control. Thus, targeting weight loss is key to maintaining T2D High Glucose Variability in Type 1 DM Patients after Surgical Treat- control. ment of Obesity by Roux-en-Y Gastric Bypass Supported By: American Diabetes Association (1-11-CT-26 to S.R.K.) SHIRIKANT TAMHANE, VIKASH DADLANI, MANPREET MUNDI, PRABIN THAPA, TODD KELLOGG, MARIA COLLAZO-CLAVELL, YOGISH KUDVA, Rochester, MN & 1974-P Obesity is an increasing problem in individuals with type 1 diabetes (T1D). Altered Hypoglycemia Counterregulation following Gastric Bypass Roux-en-Y Gastric Bypass Surgery (RYGB), to treat morbid obesity in patients Surgery for Morbid Obesity with T1D has been shown to reduce BMI and comorbidities. Increased glu- NICLAS ABRAHAMSSON, JOEY LAU BORJESSON, MAGNUS SUNDBOM, cose variability (GV) is associated with worse quality of life. There have been ANDERS F. KARLSSON, JAN W. ERIKSSON, Uppsala, Sweden no prior studies of GV in T1D after RYGB. The aim of our study was to char- The gastric bypass (GBP) surgery is one of the most common bariatric acterize GV in patients with T1D after RYGB. procedures performed. It was recently observed that GBP patients have We studied 7 subjects with T1D on continuos subcutaneous insulin infu- frequent, often asymptomatic, episodes of hypoglycemia. To characterize sion therapy (CSII) who underwent RYGB from 2008. We retrieved CSII and counter-regulatory hormonal response during hypoglycemia, we subjected blood glucose meter (BGM) data from electronic records. GV measurements patients to hyperinsulinemic hypoglycemic clamps (80 mU/m2/min; glucose included mean ± SD, % time spent in various glucose ranges, indices of hypo nadir 2.7 mmol/L) before and 5 months after GBP. Subjective symptoms were (Low blood glucose index [LBGI)] and hyperglycemia (High blood glucose evaluated with the Edinburgh Hypoglycemia Score, and counter-regulatory index [HBGI)] and average daily risk range (ADRR). hormones and the incretins glucagon-like peptide-1 (GLP-1) and glucose- Age of the subjects was 49.5 (± 8.3) yrs, BMI 42.9 (± 6.9), and duration of dependent insulinotropic peptide (GIP) were measured. 12 obese, nondia- DM 22 (± 11.7) yrs. After mean duration of 1.9 (±1.7) yrs after RYGB, HbA1c betic patients, mean age 43 years, BMI 41 kg/m2, completed investigation was 8.14 (±1.10), mean blood glucose (BG) over 4 consecutive weeks for all before and after GBP. After surgery, the Edinburgh hypoglycemia scores dur- 7 subjects was 222.8 (±89.03). GV is described in Table. Total daily insulin ing hypoglycemia had been signifi cantly attenuated from 10.7 (6.4) to 5.2 requirement decreased on an average by 41%. (4.9). Blood pressure and heart rate responses were however similar before Table. and after surgery. Importantly, the patients exhibited markedly and signifi - cantly lower glucagon, cortisol, and catecholamine responses to hypoglyce- Obs BG BG LBGI HBGI ADRR % values % values % values HbA1c Hct mia after surgery. GLP-1 and GIP rose during hypoglycemia but less post- vs. mean STD inrange <70 (hypo) >180 (hyper) pre-surgery. After surgery, lipolysis (refl ected by plasma free fatty acids and 1 156.05 71.06 1.89 7.39 22.60 66 6 28 8.4 43.6 glycerol) was attenuated during the hyperinsulinemic hypoglycemic clamps, 2 307.28 118.59 0.16 36.53 72.69 12 0 88 10 40.8 and glucose infusion rates required were greatly enhanced. This indicates 3 261.86 115.07 0.37 27.21 60.26 24 1 75 8.5 38.9 a marked increase in insulin sensitivity. In conclusion, the present fi ndings demonstrate that GBP surgery is followed by a pronounced attenuation of 4 201.92 63.31 0.16 13.72 23.39 38 1 61 8.5 40.9 symptoms and hormonal responses to hypoglycemia. As several neurohor- 5 219.84 69.15 0.00 16.81 23.11 26 0 74 7.9 33.9 monal axes are involved, this suggests an altered CNS sensing of hypogly- 6 219.50 113.36 1.61 19.52 47.08 36 7 57 6.6 36.7 cemia and these fi ndings may explain the frequent asymptomatic hypoglyce- 7 193.29 72.70 0.19 12.85 24.05 47 0 53 7.1 45.8 mic episodes occurring after GBP. Avg. 222.82 89.03 0.62 19.15 39.03 35.57 2.14 62.29 8.14 40.09 & 1975-P STD 49.07 25.13 0.78 9.83 20.98 17.53 3.02 19.37 1.10 4.03 Neuronal Correlates of Altered Food Reward Processes in Bariatric Our study demonstrated that after bariatric surgery T1D patients experi- Surgery Patients with Improved Glycemic Control ence moderate to severe risk ADRR suggesting signifi cant GV. Further explo- SABINE FRANK, JAANA M. HEINZE, STEPHANIE KULLMANN, ANDREAS ration of GV with continuous glucose monitoring is warranted to develop FRITSCHE, KATARZYNA LINDER, MAXIMILIAN VON FEILITZSCH, ALFRED KÖNIG- RYGB as a systematic option for T1D. SRAINER, HANS-ULRICH HÄRING, RALF VEIT, HUBERT PREISSL, Tübingen, Ger- many & 1973-P Bariatric surgery (BS) results in substantial body weight loss and improve- Refractory Hyperglycemia Post-bariatric Surgery from a Random- ment of glycemic control in diabetic patients. In nondiabetic patients, BS ized, Controlled Trial of Surgery vs. Medical Therapy for T2DM was shown to “normalize” food and non-food related brain functions when (STAMPEDE) compared to lean and non-surgical obese control subjects. Thus, we aimed SANGEETA R. KASHYAP, DEEPAK BHATT, KATHY WOLSKI, CLAIRE POTHIER, to investigate the neuronal and behavioral correlates of improved glycemic PHILIP R. SCHAUER, Cleveland, OH, Boston, MA control after BS within a functional magnetic resonance imaging (fMRI) Previously we reported biochemical remission of T2D (HbA1c ɖ6%) with study. We applied fMRI measurement in 24 subjects with T2DM including 12 bariatric surgery (38% with RYGB, 24% with sleeve gastrectomy (SG)) vs. obese non-surgical subjects (control group) and 12 subjects who had under- Obesity 5% with intensive medical therapy (IMT) at 3 years. Refractory hypergly- gone Roux-en Y gastric bypass surgery (RYGB) at least 6 months ago. Groups POSTERS cemia post bariatric surgery is recognized but poorly characterized in ran- were matched for pre surgical HbA1c (7.0±0.4% vs. 7.1±0.5%, p=0.97 non- domized controlled trials. We defi ned glycemic status at 3 yr follow up of surgical vs. RYGB pre surgical) and post-surgical BMI (37.8±1.4 vs. 35.7±0.8

Integrated Physiology/ 137 T2D subjects (age 48±8 yrs, HbA1c 9±1%, BMI 36±2 kg/m2) randomized kg/m², p=0.21 non-surgical vs. RYGB) and differed therefore only in post- to RYGB, SG and IMT in terms of a) controlled (HbA1c ɖ6% at 1 and 3 yr), b) surgical HbA1c (7.0±0.4% vs. 5.7±0.2%, p=0.005 non-surgical vs. RYGB). uncontrolled (never reached HbA1c ɖ6% at 1 or 3 yr) and c) relapse in glyce- Brain activation was recorded during a food picture Wanting and Liking rat- mia (HbA1c ɖ6% at 1 and HbA1c >6% at 3 yrs) and made associations with ing task. RYGB patients showed lower Wanting and Liking ratings as well baseline characteristics and weight loss post randomization. In total, 32 as lower eating behavior-related pathologies compared to non-surgical sub- patients had controlled glycemia, 87 had uncontrolled glycemia and 18 with jects. Additionally, they showed substantial improvements in glycemic con- relapse in glycemic control. Uncontrolled T2DM was greatest in IMT (39.1%) trol from HbA1c of 7.1±0.5% to 5.7±0.2% (p<0.001). RYGB patients showed and similar in the SG (32.2%) and RYGB (28.7%) groups. Relapse of T2D was increased activation in visual, motor, cognitive control, memory, and gusta- more frequent in SG (50%), and lower in both IMT 22.2% and RYGB 27.8%. tory regions. In contrast, non-surgical patients showed increased activation Uncontrolled glycemia was more frequent in those with T2D duration ≥8 yrs in areas associated with inhibition, reward and in the precuneus, a major (60.9% vs. 39.1% in duration <8 yrs), but relapse in glycemic control was connectivity hub in the brain. Therefore, food reward related neuronal acti-

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A506 OBESITY—HUMANCATEGORY vation patterns show distinct differences between diabetic obese patients Moderated Poster Discussion: Benefi ts Galore of Bariatric Surgery and with improved glycemic control after BS and comparable obese (less well Bypass Liners—Human Trials (Posters: 1978-P to 1985-P), see page 15. controlled) diabetic non-surgical subjects. Overall, improved glycemic con- trol after RYGB infl uences food reward processing which might be a crucial & 1978-P factor for long-term outcome of BS. Duodenal-Jejunal Bypass Liner Reduces Nonalcoholic Fatty Liver Disease (NAFLD) in Obese Patients with Type 2 Diabetes (T2DM) & 1976-P KATHARINA LAUBNER, LEONARD NICOLAUS, DOMINIK BETTINGER, CHRIS- Changes in Adiposity, Insulin Secretion and Sensitivity, Glucose TOPH NEUMANN-HAEFELIN, NIKOLAOS PERAKAKIS, ROBERT THIMME, HEN- Homeostasis, and Diabetes Control after Roux-en-Y Gastric Bypass NING SCHWACHA, JOCHEN SEUFERT, Freiburg, Germany in Chinese Diabetes and Nonmorbid Obese Adult Patients: A 5-Year NAFLD increases mortality in obese T2DM patients. Bariatric surgery may Prospective Cohort Study improve NAFLD. The duodenal-jejunal bypass liner (DJBL) is a removable HONGKAI GAO, MOSHEN MAZIDI, Beijing, China endoscopic device that mimicks the metabolic effects of a surgical duodeno- To report on the changes over time in adiposity, insulin secretion and sen- jejunal bypass, and therefore holds the potential to improve NAFLD in obese sitivity, glucose homeostasis and improvement in diabetes control following T2DM patients. We tested this hypothesis prospectively in 59 obese T2DM Roux-en-Y gastric bypass (RYGB) in diabetes patients of Chinese ethnicity patients. with BMI of 28 to 35 kg/m2. This prospective cohort study involved 284 59 subjects with a mean age: 46.0 ± 9.8 y, mean diabetes duration: 8.0 ± patients who met the indications of bariatric surgery for obesity. Markers of 7.9 y, mean HbA1c: 7.8 ± 1.5%, and a mean BMI: 45.0 ± 7.3 kg/m² were endo- adiposity, insulin secretion and sensitivity, glucose homeostasis and diabe- scopically implanted with a DJBL. Anthropometric, metabolic, cardiovascu- tes control were monitored from baseline to 12 months in all participants and lar risk parameters and parameters that are linked to NAFLD were moni- up to fi ve years in a subset of participants. Mixed-linear and logistic regres- tored for 12 months. After this time period, the DJBL was removed as per sion models were used for follow up. Mean body mass index decreased by license. We calculated the change from baseline of the NAFLD fi brosis score 15.5% from 29.98 kg/m2 at baseline to 25.32 kg/m2 at 12 months; and fur- (Angulo P, et al., Hepatology 2007), a validated score to identify liver fi brosis ther decreased slowly between 12 months and fi ve years. Indices of insulin in patients with NAFLD. secretion and c-peptide (both fasting and at various time-points during oral Average HbA1c reduction was 1.2% (p=0.015), despite simultaneous glucose tolerance test) improved during follow-up, translating into improved reduction of antidiabetic medication. Mean BMI was reduced from 45.5 insulin sensitivity and glucose homeostasis. Improved diabetes control kg/m² to 39.8 kg/m² (p<0.005), corresponding to an excess weight loss of (HbA1c<6.5%) among those with diabetes was 64% (181/284) at six months 39.1% (p=0.011). The mean NAFLD fi brosis score was reduced from -0.04 ± and 66.9% (190/284) at 12 months and 51.2% (145/284) at 3 years and 48.6% 1.3 to -0.87 ± 1.9. In the patients with a NAFLD score above 0.676, indicat- (138/284) at 5 years. Findings were mostly similar in men and women. The ing advanced fi brosis at baseline, signifi cant improvements were observed. present study clearly shows sustained benefi t of bariatric surgery for up Reductions of HbA1c, BMI and improvement of the NAFLD score seems to be to fi ve years post-procedure on adiposity, insulin secretion and sensitivity, preserved 6 months after explantation of the DJBL. glucose homeostasis and diabetes control, in nonmorbidly obese Chinese The DJBL is an endoscopic device that improves glycemic control, body people. weight and parameters of NAFLD in the majority of obese T2DM patients. The DJBL represents an effective therapeutic option for morbidly obese & 1977-P patients with T2DM and NAFLD complementary to bariatric surgery and life- The Impact of Roux-en-Y Gastric Bypass on Continuous Glucose style modifi cations. Monitoring in Mildly Obese Asian Patients with Diabetes ESTER YEOH, CHUN HAI TAN, NITISH MISHRA, MA THAZIN, BENJAMIN LAM, & 1979-P MICHAEL WONG, BERNICE TAN, BOON KHIM LIM, SU CHI LIM, SUBRAMANIAM Postprandial Glycemia and Lipid Profi le Are More Profoundly Ame- TAVINTHARAN, CHEE FANG SUM, ANTON CHENG, Singapore, Singapore liorated after Gastric Bypass vs. Sleeve Gastrectomy Roux-en-Y gastric bypass (RYGB) is associated with improvements in met- ALEXANDER KOKKINOS, CHRISTOS LIASKOS, NICHOLAS TENTOLOURIS, JOHN abolic parameters and diabetes remission in those with obesity, body mass DOUPIS, KLEOPATRA ALEXIADOU, GEORGIA ARGYRAKOPOULOU, DESPOINA index (BMI) ≥35 kg/m2. Asians with lower BMI ranges are at higher metabolic PERREA, THEODOROS SERGENTANIS, THEODORA PSALTOPOULOU, NICHOLAS risk compared to Caucasians. We hypothesize that RYGB leads to improved KATSILAMBROS, THEODOROS DIAMANTIS, Athens, Greece, Nikaia, Greece glycemic patterns on continuous glucose monitoring (CGM) and aimed to Bariatric surgery leads to substantial improvements in morbid obesity compare RYGB against best medical therapy (including GLP-1 agonist use) comorbidities. The present study compares the improvement in postprandial in Asian diabetes patients with suboptimal glycemic control (HbA1c>8%) glycemia and lipid profi le after Roux-en-Y gastric bypass (RYGB) and sleeve within BMI 27-32 kg/m2. Eleven patients were randomized to RYGB vs. best gastrectomy (SG). Twenty eight patients undergoing RYGB (age: 37.5±9.3 medical therapy, age (mean±SD) 41.5 ±12.3 years and diabetes duration years, BMI: 46.9±5.3 kg/m2) and 43 undergoing SG (age: 37.7±9.3 years, 6.1±3 years. Anthropometric measurements and HbA1c were performed at BMI: 50.2±7.2 kg/m2, both p=NS vs. RYGB) were consecutively recruited, baseline and 12 weeks, while retrospective CGM was performed at baseline and examined preoperatively, 3, and 6 months postoperatively. Each sub- and 6 weeks. Patients (N=6) allocated to RYGB had marked reductions in ject consumed a 450 kcal test meal, and blood samples were taken at base- weight (80.7±9.8 to 67.9±13.6 kg, p=0.001), BMI (29.5±2.1 to 24.5±2.6 kg/m2, line and every 30 minutes up to 3 hours afterwards for the assessment of p=0.020), waist circumference (104±7.3 to 90.6±7.6 cm, p=0.001) and HbA1c glucose, total cholesterol (TC), triglycerides (TG), HDL (HDL-C), and LDL cho- (10.0±1.6 to 6.2±0.5%, p=0.010). CGM glycemic profi les improved with lesterol (LDL-C). Postprandial TG and glucose responses were calculated as reduced glycemic variability [SD (2.9±0.8 to 1.7±0.4, p=0.013)], improved area under the curve (AUC) using the trapezoid rule. Values are presented as average blood glucose (BG) (11.4±2.3 to 6.9±1.3 mM, p=0.002) and percent- mean±SD. There were no preoperative differences between the two groups. age duration within target BG range of 4-10 mM (41.8±23.6 to 88.7±10.8%, Both operations led to signifi cant and comparable weight loss after 3 (RYGB p=0.001). HbA1c improved in the medical group (N=5) but was not statisti- BMI: 38.7±4.6 vs. SG BMI: 41.9±6.8 kg/m2, p=NS) and 6 months (RYGB BMI: cally signifi cant (8.7±0.9 to 7.1±1.0%, p=0.056) despite reductions in weight 34.7±4.2 vs. SG BMI: 37.4±5.7 kg/m2, p=NS). After 6 months, there were sig-

(73±3.2 to 70.3±3.7 kg, p=0.017) and BMI (30.2±2.3 to 29.1±1.9, p=0.020). nifi cant decreases for the RYGB group in TC (168.9±26.6 vs. SG: 199.5±35.6 Obesity

Although CGM profi les in the medical group showed improved trends [SD mg/dl, p=0.004) and LDL-C (108.3±24.9 vs. SG: 135.5±34 mg/dl, p=0.007). POSTERS (2.3±0.2 to 1.7±0.6, p=0.093), average BG (9.2±2.5 to 7.9±1.9 mM, p=0.473) There were no differences in HDL-C. TG postprandial AUC was lower in the

and percentage duration within target BG range (64.4±30.6 to 81.8±22.6%, RYGB group at 3 (18881±4785.3 vs. SG: 26730±6314.6 min*mg/dl, p=0.0002) Integrated Physiology/ p=0.430)], these were not statistically signifi cant. In conclusion, in mildly and 6 months (15535.7±4230.7 vs. SG: 24198.8±7480.1 min*mg/dl, p=0.001). obese Asian patients with diabetes, RYGB is found to be more effective in The same was true for glucose AUC at 3 (RYGB: 16794±2028.1 vs. SG: improving metabolic parameters and glycemic profi les, compared to best 19212.9±3347.7 min*mg/dl, p=0.008) and 6 months (RYGB: 15785.4±1884 vs. medical therapy. SG: 17535±1833.3 min*mg/dl, p=0.016). RYGB is more effective than SG in improving postoperative glycemia and lipid profi le, despite similar weight loss. This difference is most probably attributable to the malabsorptive nature and the favorable postoperative gut peptide responses that charac- terize gastric bypass.

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& 1980-P & 1982-P Treatment of Obese T2DM and Prediabetes with Dual-Path Enteral Changes in Gut Microbiome after Medical vs. Surgical Weight Loss Bypass Created by an Incisionless Anastomosis System (IAS) in a Randomized Trial DONALD C. SIMONSON, EVŽEN MACHYTKA, MAREK BUŽGA, DAVID B. LAUTZ, CLARE LEE, LILIANA FLOREA, JAMES POTTER, CYNTHIA SEARS, NOWELLA MARVIN RYOU, CHRISTOPHER C. THOMPSON, Boston, MA, Ostrava, Czech Repub- DURKIN, MELLISA SCUDDER, NISA M. MARUTHUR, MICHAEL SCHWEITZER, lic, Concord, MA, West Bridgewater, MA THOMAS MAGNUSON, KIMBERLEY STEELE, JEANNE M. CLARK, Baltimore, MD Surgical procedures creating intestinal bypass are effective in treating Bariatric surgery, especially Roux-en-Y gastric bypass (RYGB), is highly obesity, and a less invasive approach would be desirable. This is the fi rst effective in improving obesity and diabetes compared to medical weight clinical series evaluating endoscopic creation of a dual-path enteral bypass loss (MWL). We hypothesized this may be partly related to the increase using an IAS that relies on magnetic compression to create an anastomosis in the diversity and presence of benefi cial gut microbes after surgery. We enabling a portion of ingested nutrients to bypass much of the small bowel. performed 16S rRNA sequencing to identify the microbiome composition at The IAS was preloaded into the biopsy channels of two endoscopes, and baseline and at 10% weight loss in 12 subjects with type 2 diabetes who simultaneous colonoscopy and enteroscopy was performed. The IAS was were randomized (1:1:1) to one of three weight loss interventions: MWL, endoscopically deployed forming magnetic octagons in the jejunum and adjustable gastric banding (AGB), or RYGB. Metastats Qiime was used to ileum, with magnetic coupling to create a compression anastomosis. Mated calculate taxonomic profi les from 16S rRNA reads. Metastats was then magnets were passed in the stool after the compression anastomosis was applied to analyze change in relative abundance of taxa across arms, and formed, typically in 1-2 weeks. Upper GI series performed 2 weeks post-pro- to generate a principal component analysis. All participants were female, cedure confi rmed anastomotic patency. The major side effect was diarrhea, 75% black with mean age of 51 and weight loss from 6.1 to 10.0%. Mean which resolved with dietary changes or standard medical treatment. Ten hemoglobin (Hb) A1c similarly decreased in all three arms. At follow-up, gut obese patients were studied (4 T2DM, 3 pre-DM, 3 non-DM; 6M/4F; age = microbial diversity increased in 2 of 4 participants after RYGB, 1 of 4 after 48 ± 11 yrs; weight = 121 ± 18 kg; BMI = 41.3 ± 4.4 kg/m²). In all patients, AGB and 1 of 4 after MWL. The relative abundance of a potentially benefi - weight loss was 9.1 ± 1.7 kg at 2 mos and 12.9 ± 2.4 kg at 6 mos (both p < cial microbe, Faecalibacterium prausnitzii, increased in 3 participants after 0.001). During 2-hr mixed meal tolerance tests, insulin AUC (p < 0.05) and RYGB, 1 after AGB and 1 after MWL. The principal component analysis did GIP were lower, and GLP-1 moderately higher, at follow-up. In the 4 T2DM, not show any pattern of clustering in bacterial members after weight loss baseline HbA1c (7.8 ± 1.2%) decreased by 1.3 ± 0.8% and 1.8 ± 1.1% at 2 regardless of the method of weight loss. Even at a similar weight loss amount and 6 mos (both p < 0.05), despite reduction or cessation of antidiabetic and HbA1c improvement, our results suggest that there is a difference in medications, and baseline FPG of 177 ± 54 mg/dl declined to 111 ± 3 mg/dl the gut microbial composition change after surgical vs. medical weight loss. at 6 mos (p < 0.01). All 3 pre-DM (baseline HbA1c = 6.1 ± 0.2%; FPG = 119 ± 2 After RYGB, there was a tendency towards increased gut microbial diver- mg/dl) reverted to non-DM status at 6 mos (HbA1c = 5.2 ± 0.1%; FPG = 105 ± 3 sity and increase in relative abundance of the potentially benefi cial bacte- mg/dl), both p < 0.05. rium, F. prausnitzii, suggesting a possible role of gut microbiome in metabolic Conclusions: This non-surgical approach to creating a dual-path enteral improvements after RYGB. Larger and longer follow-up studies are needed bypass resulted in 1.) signifi cant weight loss in obese T2DM, pre-DM, and to further explore the mechanisms of longer-term differential improvement non-DM patients, 2.) favorable changes in insulin and incretin responses to in weight and glucose. a mixed-meal, 3.) signifi cant improvement in HbA1c and FPG in T2DM, and 4.) Supported By: National Institutes of Health reversion of pre-DM to non-DM status. & 1983-P & 1981-P The Effect of Bariatric Surgery-induced Weight Loss on Different Decrease of Visceral vs. Subcutaneous Fat with Bariatric Surgery: Lymphocyte Populations in Peripheral Blood and Adipose Tissue of A Systematic Review and Meta-analysis Subjects with Severe Obesity CLAUDIA MERLOTTI, VALERIO CERIANI, ANTONIO E. PONTIROLI, Milan, Italy ANNA CINKAJZLOVA, ZDENKA LACINOVA, JANA KLOUCKOVA, PETRA KAVAL- With bariatric surgery there is a substantial decrease of visceral (VF) and KOVA, PAVEL TRACHTA, JARMILA KRIZOVA, MILOS MRAZ, MARTIN HALUZIK, of subcutaneous (SF) adipose tissue, that can be measured through com- Prague, Czech Republic puterized tomography scan and ultrasound (CT and US, thickness and area), Lymphocytes have been suggested to be one of the primary drivers of and magnetic resonance (MRI, weight). Aim of this systematic review and obesity-associated adipose tissue infl ammation; however, their exact role meta-analysis was to compare the effect of bariatric surgery on VF and SF. in this process in humans remains largely unexplored. The aim of our study We searched in MEDLINE, EMBASE, Cochrane Library, and in reference lists was to assess the changes in different lymphocyte populations in periph- of articles and systematic reviews; we considered clinical studies, published eral blood (PB) and subcutaneous adipose tissue (SCAT) of subjects with 3rd in any language as full articles, describing decrease of both VF and SF; of 21 grade obesity after surgically induced weight loss. studies (776 subjects), 13 studies (570 subjects) were included in the analysis Anthropometric, biochemical and hormonal analyses were performed in (random-effects model). Difference of effect was espressed as SMD (stan- 12 severely obese subjects undergoing bariatric surgery (BS) at baseline and dardized mean difference) with 95% confi dence intervals (C.I.). Appropriate 1 and 6 months after intervention. Flow cytometry was used to quantify methodology (PRISMA statement) was used. CD4+ and CD8+ T cells, CD19+ B cells and CD16+56+ NK cells in PB and SCAT At baseline SF area was greater than VF area. After 6 months/1 year, sampled from abdominal region. decrease of SF area was greater than VF area decrease, while decrease of BS induced a marked weight loss (BMI 46.6 ± 3.2 vs. 38.0 ± 2.7 kg/m2, VF thickness was greater than SF thickness. % VF decrease (weight, area, p<0.001 for baseline vs. 6 months after BS) along with improved glucose thickness) was greater than % SF decrease (Table 1). Heterogeneity was sig- control (HbA1c 48.5 ± 5.5 vs. 39.2 ± 2.4 mmol/mol, p=0.012) and reduced CRP nifi cant. These data indicate that with bariatric surgery percent decrease of levels (11.5 ± 2.9 vs. 6.5 ± 2.0 mg/l, p=0.016). The amount of CD4+ and CD8+ visceral fat is superior to percent decrease of subcutaneous fat. T cells in PB was not changed after BS, while in SCAT a decrease of CD4+ + Table 1. (35.6 ± 4.2 vs. 27.2 ± 4.1%, p=0.019) as well as CD8 cells (21.2 ± 2.1 vs. 15.8 ± 2.2%, p=0.044) was observed. Similarly, CD16+56+ NK cells remained Obesity studies subjects SMD p heterogeneity greater unchanged in PB, but decreased in SCAT (7.2 ± 1.2 vs. .4.6 ± 0.7%, p=0.026), POSTERS (n) (95% C.I.) (I-squared, p) decrease while CD19+ B cells decreased only in PB (7.6 ± 1.4 vs. 5.7 ± 1.1%, p=0.009). Δ area (cm2)1 3 112 -1.98 (-2.31;-1.66) 0.0001 0.0%, NS SF In SCAT, changes in CD4+ and CD8+ cells were positively associated with the Integrated Physiology/ + Δ area (%) 3 112 0.35 (0.09;0.62) 0.01 84.8%, 0.001 VF reduction of body weight and HbA1c and, in case of CD4 cells, also with the + + Δ thickness (mm)2 6 339 2.21 (1.98;2.43) 0.0001 98.7%, 0.001 VF decrease in serum insulin and leptin, while changes in CD16 56 cells corre- lated positively with the reduction of BMI, triglyceride and leptin levels. Δ thickness (%) 6 339 2.31 (2.09;2.54) 0.0001 98.5%, 0.001 VF In conclusion, bariatric surgery-induced weight loss is associated with the Δ weight (kg)3 6 119 -0.02 (-0.29;0.25) NS 64.7%, 0.05 similar reduction in the local adipose tissue content of CD4+ and CD8+ T lympho- Δ weight (%) 6 119 5.36 (4.61;6.13) 0.0001 97.3%, 0.001 VF cytes and NK cells, which might contribute to the amelioration of low-grade infl ammation and to metabolic improvements after BS. 1 = CT/US/MRI; 2 = CT/US; 3 = MRI; SF = subcutaneous; VF = visceral; one Supported By: RVO-VFN64165; AZV 15-26854A, AZV 15-27863A; study evaluated area and thickness; NS = not signifi cant. SVV260019/2014

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& 1984-P age 20-50 years meeting study criteria were placed on the HP or HC diet for 6 Regulation of the PPARγ-FABP4 Axis after Bariatric Surgery months (mo) with weekly food pickup and weight monitoring. OGTT, RMR, and CYRUS JAHANSOUZ, HONGLIANG XU, ANN V. HERTZEL, DAVID A. BERNLOHR, DXA were performed at baseline and after 6 mo on the HP or HC diet. Glucose SAYEED IKRAMUDDIN, Minneapolis, MN and insulin were assayed for determination of insulin sensitivity and irisin, FGF- Even with advances in modern medicine, bariatric surgery remains the 21) and UCP-1 were determined. 100% of the patients on the HP diet (n=12) con- most effective treatment for weight loss and type 2 diabetes mellitus (T2DM). verted to NGT whereas only 33.3% converted that were on the HC diet (n=12). Despite widespread clinical acceptance, the mechanism (s) responsible for Weight and BP signifi cantly decreased in both groups. HP group had an increase achieving such rapid improvement in insulin resistance, prior to substantial in % lean body mass and decrease in fat mass while both % lean and fat body weight loss, has remained elusive. One target of great interest has been Per- mass decreased in the HC group. Glucose tolerance and insulin sensitivity were oxisome Proliferator-Activated Receptor γ (PPARγ) and its activity as a means signifi cantly improved in the HP diet compared to the HC at 6 mo. Our results of modulating whole body insulin sensitivity. Herein we show that as early suggest that lean body mass preservation may be more important than total as one week following the Vertical Sleeve Gastrectomy (VSG; n=20), adipose weight loss in the conversion of IGT to NGT, possibly due to the high insulin sen- tissue PPARγ mRNA and protein levels decrease by nearly 50% (p=0.028 and sitivity of muscle cells. Additionally, the HP had increased levels of Irisin, and p=0.09, respectively). Several PPARγ downstream targets, notably Fatty Acid UCP-1 possible major regulators of metabolic rate and weight loss. Binding Protein 4 (FABP4/aP2), CD36, perilipin, Pde3B and Atgl decrease in Table. abundance as well. To evaluate the regulation of PPARγ in response to the VSG with other bariatric procedures, the observations were extended to a ran- HP (n=12) HC (n=12) domized controlled trial comparing patients with T2DM undergoing Roux-en-Y Parameters Baseline 6 months p- value Baseline 6 months P-value p* Gastric Bypass (RYGB; n=13) or hypocaloric restriction (HC diet; n=14). While HP vs. HC patients undergoing RYGB showed similar trends in decreased PPARγ expres- at 6 months sion as those undergoing VSG, patients randomized to HC diet exhibited mark- Age (yr) 43.1 ± 1.3 41.1 ± 1.7 edly increased PPARγ expression as indicated by a near fi ve-fold increase in Weight (lbs) 248.8 ± 12.2 229.4 ± 11.3 0.005 236.6 ± 6.8 211 ± 8.3 0.008 0.07 protein level (p=0.03). Furthermore, higher preoperative hemoglobin A1c cor- Blood Pressure (BP) 130/80 ± 3/2 116/72 ± 3/3 .01/.01 126/80±4/2 113/73±4/2 .01/.01 0.6 related with a diminished capacity to reduce PPARγ gene expression (r=0.60, p=0.015) in adipose tissue independently from the type of surgery performed. RMR 1571 ± 19 1862 ± 18 0.005 1569 ± 17 1598±15 0.6 0.001 Altogether, our data suggest a common pathway after bariatric surgery leading % lean body mass + 2.55 ± .3 .001 -3.0 ± .7 .01 .001 to the rapid down regulation of PPARγ activity and resolution of insulin resis- % fat body mass -2.49± .4 .001 -3.55± .8 .005 .02 tance prior to substantial weight loss and distinct from caloric restriction. Irisin ( ng/ml) 9.41 ± 0.7 11.92 ± 0.4 0.03 8.86 ± 0.8 8.99 ± 0.8 0.5 0.02 Supported By: National Institutes of Health (DK053189, ST-01, P30DK050456) FGF-21 (pg/ml) 166.4 ± 14 127.9 ± 11 0.02 168.6 ± 15 160.9 ±13 0.06 0.02 & 1985-P UCP-1 (ng/ml) 0.30 ± 0.06 0.63 ± 0.07 0.05 0.82 ± 0.35 0.62 ± 0.26 0.03 0.04 Aging Increases Monocyte Chemoattractant Protein-1 Level in p is signifi cance of change from Baseline to 6 mo; p* is signifi cance between Association with Bariatric Surgery Operating Time and Metabolic HP vs. HC at 6 mo. Severity Risk Supported By: American Diabetes Association (7-12-CT-41 to A.K.) STEVEN K. MALIN, JENNIFER L. KAPLAN, LINGJIN MENG, JAMES C. GARMEY, JENNIFER L. KIRBY, ANGELA TAYLOR, PETER HALLOWELL, COLEEN A. MCNA- 1987-P MARA, Charlottesville, VA Relationship between Gestational Weight Gain and Pregnancy Monocyte chemoattractant protein-1 (MCP-1) is an infl ammatory marker Complications or Delivery Outcome related to obesity and aging. Aging infl uences bariatric surgery operative WENJIA YANG, FEIFEI HAN, XIAOLING CAI, XUEYAO HAN, LINONG JI, Beijing, diffi culty, but the mechanism is unknown. Therefore, we assessed the role of China MCP-1 on operating time in young vs. old bariatric surgery patients. 56 young The aim of this study is to analyze the relationship between gestational (F:46, Age:34.9±0.5 y, BMI:48.1±1.0kg/m2) and 48 old (F:34, Age:57.0±0.7 y, 2 weight gain and delivery outcomes or pregnancy complications.This retro- BMI:46.7±1.0kg/m ) adults undergoing bariatric surgery (gastric bypass, spective study included 1,102 pregnant women. Data were extracted from sleeve gastrectomy) were included in this cross-sectional study. Blood pres- electronic medical records. Baseline weight and weight change during the sure, glycemia (fasting glucose/insulin, HbA1c), blood lipids (LDL, HDL and whole pregnance were recorded. For the statistical analysis, these partici- TG), and infl ammation (MCP-1) were assessed prior to surgery. Metabolic pants were divided into four groups based on the weight gain quartiles in severity z-scores were calculated from BMI, glucose, TG, HDL and blood different trimester of pregnancy. Weight gain in the second trimester of pressure. Omental adipose biopsies were collected at the time of surgery pregnancy was positively correlated with macrosomia, and negatively cor- for MCP-1 protein analysis. Operating time was used to depict surgical dif- related with neonatal death. Logistic analysis showed that weight gain in fi culty and was defi ned as time of incision to close. Old vs. young adults the second trimester was an independent risk factor of postpartum hemor- had higher HbA1c (7.1±0.2 vs. 6.2±0.2%, P=0.02) and LDL (325.6±31.5 vs. rhage (OR=1.067, P=0.04) and macrosomia (OR=1.145, P=0.02). Weight gain 242.3±24.5nM, P=0.03). There was no difference in BMI, metabolic severity in the third trimester showed signifi cantly positive correlation with neonate or insulin between groups, but operating time was longer in old vs. young weight and macrosomia, signifi cantly negative correlation with neonatal (164.2 ±10.8 vs. 136.4±8.3 min, P=0.04). Circulating MCP-1 (302.2±17.8 vs. death, preterm birth, gestational diabetes, infant of low-birth weight. Logis- 235.2±24.8pg/ml, P=0.04) was also elevated in old vs. young, but this was tic analysis showed that weight gain in the third trimester was independent not explained by omental fat differences. Nevertheless, increased serum risk factor of preterm birth (OR=0.770, P<0.01), infant of low-birth weight and omental fat MCP-1 was associated with increased metabolic severity (OR=0.813, P=0.02) and gestational diabetes (OR=0.828, P<0.01). Total ges- (R=0.25, P=0.02 and R=0.27, P=0.01, respectively). Longer operating time tational weight gain was positively correlated with neonate weight and was also linked to higher MCP-1 total protein (R=0.31, P<0.01) and HbA1c the morbidity of macrosomia, and negative correlation with the neonatal (R=0.30, P=0.01). Thus, elevated MCP-1 is associated with higher operating death, stillbirth, gestational diabetes, preterm birth and infant of low-birth time and metabolic risk severity in older bariatric patients. Pre-operative weight, which is an independent risk factor for the development of neona- Obesity treatment of adipose tissue infl ammation in older bariatric adults may be tal death (OR=1.063, P<0.01), postpartum hemorrhage (OR=1.096, P=0.01), POSTERS required to reduce surgical diffi culty and enhance surgery effectiveness. macrosomia (OR=0.890, P=0.04), infant of low-birth weight and gestational

diabetes (OR=0.922, P=0.01). According to this analysis, the weight gain dur- Integrated Physiology/ 1986-P ing pregnancy was associated with the gestational complications, adverse Remission of Prediabetes and Improved Metabolic Parameters with pregnancy outcomes, status of neonate in varying degrees. a High-Protein Diet FRANKIE B. STENTZ, ABBAS KITABCHI, Memphis, TN 1988-P Remission from prediabetes to normal glucose tolerance (NGT), weight loss, The Association of Clusterin and Leptin in Type 2 Diabetes Is Depen- body composition, and metabolic parameters were studied in this randomized dent upon the Degree of Obesity controlled clinical trial feeding study comparing a High Protein (HP) (30% pro- RAELENE E. MASER, M. JAMES LENHARD, RYAN T. POHLIG, Newark, DE tein, 40% carbohydrate, 30% fat) diet vs. High Carbohydrate (HC) (15% protein, Leptin, a hormone produced by adipocytes, acts in the hypothalamus to 55% carbohydrate, 30% fat) diet in obese, prediabetic patients. Subjects (n=24) decrease appetite and increase energy expenditure. Clusterin, a glycoprotein expressed in the central nervous system and plasma, has also been identi-

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fi ed as an anorexigenic neuropeptide. In rodents, clusterin has been shown 1990-P to be a regulator of leptin signaling in hypothalamic neurons. Clusterin has Identifi cation of Differential Circulating LncRNAs as Novel Meta- also been identifi ed as a leptin-binding protein in plasma. The relationship of bolic Biomarkers in Obese Patients clusterin and leptin in the circulation has not been addressed. In this study, JIA SUN, YUTING RUAN, MING WANG, NA YU, RONGPING CHEN, LEI SUN, we examined whether serum clusterin is independently associated with TIEMIN LIU, HONG CHEN, Guangzhou, China, Singapore, Singapore, Dallas, TX leptin levels in persons with type 2 diabetes (T2D). Fifty individuals with Circulating Long non-coding RNAs (lncRNAs) have been found to be valu- T2D (age=63±10 years, BMI=32.7±5.3 kg/m2) were examined. Serum clus- able biomarkers in a number of human diseases. However, lncRNAs biomark- terin levels were higher in females compared to males (225±38 vs. 200±30 ers have yet to be identifi ed in obesity. This study was designed to charac- µg/mL) but were not statistically signifi cant (p=0.094) after controlling for terize circulating lncRNA expression in obese and nonobese human subjects. leptin. There was no difference in BMI between males and females in this First, we assessed the genome-wide circulating lncRNA expression profi les study. Signifi cant Spearman rank correlations of clusterin included leptin in blood from 3 obese and 3 nonobese human subjects. We found that circu- (r=0.29, p<0.05) and gender (r=0.37, p<0.01). Linear regression, with leptin as lating levels of three lncRNAs (lncRNA-p5549, lncRNA-p21015 and lncRNA- the dependent variable, revealed that gender (p<0.001), BMI (p<0.001), and p19461) were signifi cantly decreased in obese human subjects only. Next, clusterin (p=0.023) were signifi cant correlates (model R2=0.64, p<0.001). The using RT-PCR we measured the expression levels of these three lncRNAs inclusion of a signifi cant interaction of BMI x clusterin (p=0.018) improved in 13 obese and 13 nonobese human subjects and found similar differences. the model (R2=0.68). This interaction indicated a strong positive association Moreover, the circulating levels of these three lncRNAs were negatively cor- of serum clusterin and leptin for those with the highest BMI, whereas a slight related with body mass index (BMI), waist circumference, waist to hip ratio negative relationship between clusterin and leptin for those with the low- and fasting insulin. The expression of lncRNA-p19461 was also negatively est BMI. Thus in T2D, serum clusterin and leptin are associated particularly correlated signifi cantly with homeostasis model assessment-estimated for those that are very obese. Clusterin does play a key role in hypothalamic insulin resistance. Finally, we tested the circulating levels of these three feeding regulatory pathways in rodents. Future investigations are needed to lncRNAs in 9 obese human subjects who lost weight after 12 weeks of a determine if the clusterin signaling pathway is linked to leptin signaling in diet-induced weight loss program. We found that only the circulating level of obese humans. Clusterin may offer a new target for better understanding of lncRNA-p19461 signifi cantly increased. In summary, lncRNA-p19461 may be obesity-related metabolic disorders. a potential candidate as a noninvasive metabolic biomarker of obesity.

1989-P 1991-P Characteristics of Individuals Developing Type 2 Diabetes in the Use of a Custom Genotyping Array to Identify Variants Associated SCALE Obesity and Prediabetes Randomized, Double-Blind, Lira- with BMI in Pima Indian Adults and Children glutide vs. Placebo Trial LESLIE J. BAIER, YUNHUA MULLER, SAYUKO KOBES, PENG CHEN, PAOLO PIAGGI, DAVID C.W. LAU, MATTHIAS BLUEHER, LUC VAN GAAL, DOMENICA M. RUBINO, ANUP NAIR, MICHAEL TRAURIG, WILLIAM C. KNOWLER, CLIFTON BOGARDUS, GERMAN GUERRERO, LINDA SHAPIRO MANNING, JOHN P.H. WILDING, Calgary, ROBERT L. HANSON, Phoenix, AZ AB, Canada, Leipzig, Germany, Antwerp, Belgium, Arlington, VA, Plainsboro, NJ, A custom Affymetrix Axiom array was designed to capture variation Søborg, Denmark, Liverpool, United Kingdom detected in whole genome sequence data from 266 full heritage Pima Indians. This 3-year trial examined the effect of liraglutide 3.0 mg, as an adjunct to This array was used to genotype participants of a longitudinal study of type 2 diet and exercise, in delaying the onset of T2D (primary endpoint) in adults diabetes among the Gila River Indian Community, where full heritage Pima Indi- with prediabetes and BMI ≥30 kg/m2, or ≥27 kg/m2 with comorbidities. ans (N= 3,625) and non-full heritage Pima Indians (N=4,113) were genotyped in Individuals were randomized 2:1 to once-daily s.c. liraglutide 3.0 mg or 2 stages. Genotypes from 548,206 variants passed quality control metrics and placebo, with a 500 kcal/day defi cit diet and 150 min/week exercise. Effi cacy tagged (r2 ≥0.85 within 300 kb windows) 92% of the 4.9M common variants data are observed means, with LOCF. (minor allele frequency >0.05) detected in the Pima genomes. Among subjects Individuals developing T2D were on average older, had more comorbidi- with genotypic data, 5,875 had at least one measure of BMI at age ≥15 years ties, with a higher baseline A1c, FPG and BMI than the entire randomized from an exam when the subject was nondiabetic, and 5,350 had at least one population (Table). Time to onset of T2D over 3 years was 2.7 times longer measure of body size between the ages of 5-20 years for calculation of an with liraglutide 3.0 mg compared to placebo (95% CI 1.9;3.9), with 79.3% age and sex adjusted childhood z-score. Using a mixed model to account for reduced risk of developing T2D with liraglutide 3.0 mg (p<0.0001). Mean family membership with adjustment for age, birth year, sex, European/Amer- weight loss at 3 years for the entire study population was 6.1% with lira- ican Indian admixture, fraction Pima, and using genomic control to account glutide 3.0 mg vs. 1.9% with placebo (estimated difference -4.3% [95% CI for residual stratifi cation, no SNP achieved genome-wide signifi cance (P <5 × -4.9;-3.7], p<0.0001). Most individuals who developed T2D (>90% in both 10-8) in its association with adult maximum BMI. However, SNPs (rs17806420, groups) lost less body weight than the treatment group mean. In those with rs11621059, rs615171, rs76623538, rs12766426, and chr22:31664134) near T2D, one hypoglycemic event was reported with liraglutide 3.0 mg vs. fi ve CYB5A, C14orf177, TNKS/MSRA, PALLD, ADAM12, and LIMK2 associated with with placebo, none severe. Liraglutide 3.0 mg was generally well tolerated. adult BMI with P-values <10-5, where the association was derived from both Liraglutide 3.0 mg, as an adjunct to diet and exercise, delayed the time full and non-full heritage Pima Indians (P <0.05 in each group). Among these to onset and reduced the risk of developing T2D vs. placebo over 3 years. SNPs, rs615171 near TNKS/MSRA had the strongest association with child- (NCT01272219). hood BMI z-score (P = 8 × 10-5). Individuals with genotypes T/T vs. C/C at Table. Baseline Characteristics. rs615171 had a 1.7 fold higher z-score during childhood and a 6.1% higher BMI in adulthood. This fi nding is consistent with a recent report that variants in this T2D by T2D by Entire Entire region strongly associate with extreme obesity in French and German adoles- week 160 week 160 population population cents. Additional replication and functional analyses are ongoing for all top Liraglutide Placebo Liraglutide Placebo n = 26 n = 46 n = 1505 N = 749 SNPs to provide further evidence that these loci affect BMI. Supported By: National Institute of Diabetes and Digestive and Kidney Dis- Female, number (%) 13 (50%) 33 (72%) 1141 (76%) 573 (77%) eases/National Institutes of Health Age, years (mean±SD) 48.4±8.3 49.3±13.2 47.5±11.7 47.3±11.8 Obesity 2 POSTERS BMI, kg/m (mean±SD) 40.2±8.6 40.4±7.0 38.8±6.4 39.0±6.3 1992-P A1c, % (mean±SD) 6.1±0.4 5.9±0.4 5.8±0.3 5.7±0.3 Weight Management during Short-Term Intensive Insulin Therapy

Integrated Physiology/ Fasting plasma glucose, mmol/L (mean±SD) 6.0±0.6 5.9±0.6 5.5±0.6 5.5±0.5 Was Associated with Better Clinical Outcomes in Patients with Newly Diagnosed Type 2 Diabetes Oral glucose tolerance test 2 hr plasma glucose, 7.8±2.0 7.9±1.8 7.4±1.8 7.4±1.7 XUESI WAN, ZHIMIN HUANG, LIEHUA LIU, WEIJIAN KE, YANBING LI, Guangzhou, mmol/L (mean±SD) China Comorbidities – – – – Obesity is common disadvantage for the insulin therapy with type 2 diabe- Dyslipidemia, number (%) 14 (54%) 21 (46%) 499 (33%) 249 (33%) tes mellitus (T2DM). The results of our previous studies suggest that short- Hypertension, number (%) 19 (73%) 18 (39%) 635 (42%) 312 (42%) term continuous subcutaneous insulin infusion (CSII) induced drug-free Supported By: Novo Nordisk A/S disease remission in patients with newly diagnosed T2DM. Weight manage- ment is so important during the CSII therapy but the determinants of weight changes were still unknown. We retrospectively investigated 160 patients with newly diagnosed T2DM (n=160, 103 male, aged 49.8±10.4 ys, HbA1c

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A510 OBESITY—HUMANCATEGORY

11±2.1%) underwent 2 weeks of CSII. After short-term CSII treatment, 51.9% diastolic dimension and LA volume index (all Ps<0.05). Hepatic steatosis of the patients had a reduction in body mass index (ΔBMI), while 26.9% amount showed a strong positive relationship with a parameter of LV fi lling were unchanged. The BMI decreased from 25.27±3.27 to 25.01±3.07. While pressure which refl ects diastolic dysfunction (P=0.033). Subjects without the Waist to hip ratio didn’t reduced signifi cantly, 0.93±0.06 both before NAFLD were more likely to have higher myocardial glucose uptake compared and after therapy. There was strong positive correlation between pro- to those with NAFLD. Furthermore, hepatic fi brosis had a signifi cant cor- intervention value and its corresponding change in BMI (r=0.265, p=0.001). relation with diastolic dysfunction and impaired myocardial insulin sensitiv- Daily insulin dosage and diet (kcal/kg body weight) had negative relationship ity. Multivariable logistic regression showed that diastolic dysfunction was with ΔBMI (r=-0.299, -0.345, p<0.01). But the exercise time and frequency independently associated with hepatic fi brosis (OR=1.47 to 1.64, all Ps<0.01) of hypoglycemia did not correlate with the ΔBMI. Moreover, the reduction and steatosis (OR=1.91 to 1.94, all Ps<0.05). of BMI had positive relationship with the improvements of HOMA-IR and In conclusion, hepatic steatosis and fi brosis are signifi cantly associated acute insulin response (AIR) during the intravenous glucose tolerance test with diastolic heart dysfunction. This association is linked with myocardial (r=0.228, 0.223, p<0.05). In multiple linear regression ΔBMI was associ- insulin sensitivity evaluated by PET. ated with 1-year glycemic remission (standardized coeffi cients were 0.28, F=6.132, p=0.001) after adjusted for age, sex and baseline GHbA1c.We con- 1995-P cluded that stricter weight management during CSII therapy was associated Effects of Exenatide on Proliferation and Differentiation of Human with better clinical outcomes in patients with newly diagnosed T2DM. And Primary Preadipocytes lower insulin dosage and hypo-caloric diet might be profi table. WEI JIANG, XIAOFENG LI, HONGYUN LU, SHUO LIN, LONGYI ZENG, Guangzhou, China 1993-P Exenatide is a glucagon-like peptide receptor agonist which could induce Comprehensive Assessment of the Safety and Toxicology of MOD- weight loss and regulate glucose homeostasis. However, its role on human 6031: A Novel, Long-Acting Dual GLP-1/Glucagon Agonist preadipocytes is still controversial. Here we evaluated the effect of exenatide LITAL ISRAELI YAGEV, AHUVA BAR ILAN, VERED LEV, OREN HERSHKOVITZ, GILI on proliferation and differentiation of human preadipocytes. Human primary HART, Nes Ziona, Israel preadipocytes were isolated and cultured from omental adipose tissues OPKO Biologics is developing long acting Oxyntomodulin (MOD-6031) for undergoing surgery. Then the preadipocytes were treated with graded the indication of weight management and potentially type 2 diabetes, utiliz- concentrations of exenatide (0, 10-10, 10-9, 10-8, 10-7mol/L) during differen- ing reversible PEGylaton technology. The core technology is a spacer that tiating to adipocyte. The proliferation of preadipocytes determined by MTT links between the PEG and the peptide. Once injected, a fi rst order hydro- detection was enhanced signifi cantly in 10-8 and 10-7mol/L groups after 72 h lysis slowly releases the intact OXM, enabling a prolonged exposure of the incubation (P<0.05). The mRNA and protein levels of preadipocytes specifi c peptide while maintaining its biological activity and ability to pass throw the marker pref-1 were elevated after exenatide intervention during preadipo- blood brain barrier. cytes differentiation (P<0.05). The morphological properties and the oil red A battery of comprehensive toxicological studies have been perform staining showed that exenatide inhibited preadipocytes differentiation in a to evaluate the potential toxicity of MOD-6031 following subcutaneous dose-dependent pattern (P<0.05). Glycerol released in the culture medium administration to rat and monkeys and to assess the reversibility of adverse was increased signifi cantly after exenatide treatment during preadipocytes effects following a recovery period. GLP-compliant safety pharmacology differentiation (P<0.05). Our study showed that exenatide could promote studies on the core organ systems have been conducted for extensive safety human preadipocytes proliferation, inhibit differentiation and accelerate evaluation. Additionally, in vitro Genotoxicity studies have been performed lipolysis which may contribute to lower body weight and increased insulin to evaluate MOD-6031 potential to induce genetic damages. sensitivity. Administrations of MOD-6031 up to 300 and 250mg/kg to rats and mon- Supported By: National Natural Science Foundation of China (81070661); Novo keys, respectively, demonstrated an excellent safety profi le without any Nordisk China (2014) unexpected adverse events at signifi cant margins above the clinical doses. The changes noted were related to the pharmacological activity of the drug. 1996-P No antibodies to MOD-6031 were detected in plasma samples. No adverse Brain Insulin Action Suppresses Endogenous Glucose Production effects on respiratory or central nervous system functional observational and Stimulates Glucose Uptake during Hyperinsulinemia in Lean parameters were observed after single SC doses in rats. In a single SC injec- but Not in Obese Men tion to telemetrized monkeys few non-signifi cant toxicologically fi ndings and MARTIN HENI, ROBERT WAGNER, STEPHANIE KULLMANN, HALIZA MAT HUSIN, minor increase of QTc interval were observed at the highest administered SOFIYA GANCHEVA, MICHAEL RODEN, NORBERT STEFAN, HUBERT PREISSL, dose (250 mg/kg), corresponding to exposure that is ~1250-fold higher than HANS-ULRICH HÄRING, ANDREAS FRITSCHE, Tübingen, Germany, Düsseldorf, the proposed starting dose in the Phase1. The Ames and Chromosomal Aber- Germany ration studies resulted with no genotoxicity effects of MOD-6031. The human brain is an insulin-sensitive organ and intranasal insulin spray The overall safety profi le of MOD-6031 supports the ongoing phase 1 application enables insulin delivery into the CNS. By this approach, specifi c study in over-weight and obese patients. insulin-sensitive brain areas have been identifi ed. Furthermore, brain insulin resistance was observed in obese subjects. The role of brain insulin action for 1994-P peripheral metabolism is still unclear. Twenty one young men (11 lean, 10 over- Association of Hepatic Steatosis and Fibrosis with Myocardial Dys- weight) participated in this randomized controlled crossover trial. Each subject 2 function in Relation to Myocardial Insulin Sensitivity underwent two hyperinsulinemic-euglycemic clamps with [6,6- H2] glucose JI-YEON LEE, YONG-HO LEE, KWANG JOON KIM, GYURI KIM, SO RA KIM, infusion. 90 min after clamp initiation, participants received either intranasal EUGENE HAN, MIJIN YUN, CHUL HOON KIM, BYUNG-WAN LEE, CHUL WOO insulin or placebo. Insulin spill-over into circulation after intranasal insulin appli- AHN, BONG-SOO CHA, EUN SEOK KANG, Seoul, Republic of Korea cation was mimicked by an adequate iv insulin bolus over 15 min during placebo Nonalcoholic fatty liver disease (NAFLD) is associated with increased car- day. Plasma insulin concentrations were comparable between both clamps, diovascular risk, including cardiac dysfunction. Among categories of NAFLD, both directly after spray application as well as thereafter. Higher glucose infu-

hepatic fi brosis is the most progressive condition which can affect mortal- sion rates were necessary to maintain euglycemia after administration of nasal Obesity ity. Myocardium requires energy from fatty acids and glucose, which can be insulin compared to placebo. This effect was only present in lean subjects in POSTERS altered under insulin resistant condition. We investigated the association whom the glucose infusion rate increased stronger after administration of

of hepatic steatosis and fi brosis with myocardial dysfunction in relation to nasal insulin than placebo (+52 ± 26% vs. +28 ± 12%, p<0.05). The suppression Integrated Physiology/ myocardial insulin sensitivity using [18F]-fl uorodeoxyglucose-positron emis- of endogenous glucose production by insulin was larger after administration sion tomography (PET). of nasal insulin than after placebo (p<0.005). Furthermore, glucose uptake into A total of 315 subjects (NAFLD, n=120) in a health promotion center of tissue tended to be higher after nasal insulin (p<0.1). In the overweight group, a tertiary hospital were enrolled. Myocardial glucose uptake was evalu- no such effects were detected. We provide novel evidence that brain insulin ated using PET. Hepatic steatosis and fi brosis were assessed by transient action improves glucose metabolism at systemic hyperinsulinemia by enhanc- liver elastography (Fibroscan®) which non-invasively quantifi es hepatic fat ing insulin-mediated suppression of endogenous glucose production and stimu- amount based on the properties of ultrasonic attenuation by fat. Cardiac lating of glucose uptake. In overweight subjects, these mechanisms appear to structure and function were examined by echocardiogram. be absent. Therefore, brain insulin resistance, as found in obesity, may have Compared to those without NAFLD, patients with NAFLD had alteration unfavorable effects for whole body glucose homeostasis. in cardiac remodeling, manifested by increased LV mass index, LV end- Supported By: German Federal Ministry of Education and Research

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1997-P 1999-P Associations between Visceral Adipose Tissue and Glucose Metab- Comparisons of Weight Changes between SGLT2 Inhibitors Treat- olism: The Role of Ethnicity ment and GLP-1 Analogs Treatment in Type 2 Diabetes Patients: A PERNILLE F. RØNN, GREGERS S. ANDERSEN, TORSTEN LAURITZEN, DIRK L. Meta-analysis CHRISTENSEN, METTE AADAHL, TORBEN HANSEN, NIELS GRARUP, BENDIX XIAOLING CAI, YIFEI CHEN, WENJIA YANG, LINGLI ZHOU, XUEYAO HAN, LINONG CARSTENSEN, MARIT E. JØRGENSEN, Aarhus, Denmark, Gentofte, Denmark, JI, Beijing, China Copenhagen, Denmark, Glostrup, Denmark Both the SGLT2 inhibitors treatment and GLP-1 analogs treatment may Visceral adipose tissue (VAT) has been identifi ed as a risk marker of meta- result in body weight reduction different from other antidiabetes treatments. bolic disturbances and evidence suggest that the risk differs between ethnic The aim of this study is to evaluate the effi cacy of weight changes from groups. The aim was therefore to examine the associations between VAT baseline of these two antidiabetes treatments compared with placebo in assessed by ultrasonography and indices of glucose metabolism in Inuit, type 2 diabetes patients and the associated factors. Studies were searched Africans and Europeans. We did a cross-sectional analysis of 5,053 partici- from when recording began, June 2004, until June 2015, and placebo-con- pants aged 17-95 years from 3 studies conducted in Greenland, Kenya and trolled randomized trials in type 2 diabetes patients with a study length ≥12 Denmark from 2005 to 2010. We used fractional polynomials to describe the weeks were included. A total of 53 RCTs were included. Compared with effect of VAT on indices of glucose metabolism (fasting and 2-h glucose, and placebo, treatment with SGLT2 inhibitors was associated with a signifi cant insulin resistance by homeostasis model assessment (HOMA-IR)) adjusting greater decrease in weight change from baseline, (weighted mean differ- for age, physical activity energy expenditure (PAEE), BMI, waist circumfer- ences (WMD), -1.86kg (95% CL, -2.04 to -1.68kg), P <0. 001). Compared with ence (WC) and TBC1D4 allele status. VAT was positively associated with placebo, changes with GLP-1 treatment was also associated with a compa- HOMA-IR across all ethnic groups and both genders (Figure 1) and with 2-h rable decrease in weight change from baseline (WMD, -1.36kg (95% CL, 1.74 glucose in Europeans only. A weak positive or no association was found for to -0.97kg), P <0.001). Meta-regression analysis indicated that the baseline fasting glucose, but Africans had signifi cantly lower absolute values of fast- age, gender, baseline HbA1c, diabetes duration, or baseline BMI, was not ing glucose for a given level of VAT. Ethnicity modifi ed the infl uence of VAT associated with the weight change from baseline in SGLT2 inhibitors or in on 2-h glucose (p=0.006) and HOMA-IR (p=0.014) with a steeper increase GLP-1 treatment corrected by placebo. Comparisons of weight changes from among Europeans but in men only, and not on fasting glucose for either gen- baseline corrected by placebo between SGLT2 inhibitors and GLP-1 treat- der. The results indicate that VAT has an effect on insulin resistance beyond ment indicated that the difference was not signifi cant (p>0.05). According BMI and WC in all ethnic groups with more pronounced associations for to this meta-analysis, treatment with SGLT2 inhibitors and treatment with Europeans than for Inuit and Africans. GLP-1 analogs led to comparable weight changes from baseline, which are Figure 1. both with signifi cance when compared with placebo treatment. 2000-P Long-Chain Saturated Fatty Acid Content Is Reduced in Deep Sub- cutaneous Adipose Tissue but Does Not Relate to Insulin Sensitivity in Humans with Type 2 Diabetes Mellitus KALMAN BODIS, JESPER LUNDBOM, TOMAS JELENIK, DANIEL MARKGRAF, VOLKER BURKART, KARSTEN MUESSIG, MICHAEL RODEN, JULIA SZENDROEDI, Düsseldorf, Germany Fatty acid (FA) composition of deep subcutaneous adipose tissue (DSAT) has gained attention as a potential regulator of insulin sensitivity (IS). The relevance of the FA unsaturation index (UI) of DSAT in type 2 diabetes (T2D) is currently unknown. We hypothesized that (i) UI is reduced and (ii) chain length of FA’s in DSAT is higher in glucose tolerant humans (CON) compared to T2D patients of similar body mass (BMI), and (iii) cannot be explained by alteration of FA oxidation capacity. Employing in vivo proton magnetic resonance spectroscopy, we measured UI and CH2/CH3 ratios, as markers of FA’s double bond content and saturated chain length of DSAT, in 17 T2D Supported By: Aarhus University and 13 CON of similar age and BMI. IS was assessed from euglycemic-hyper- insulinemic clamps (m-value) in 10 T2D and 11 CON. Capacity of octanoyl- 1998-P carnitine oxidation was measured ex vivo from high-resolution respirometry in DSAT biopsies. M-value was 45% lower in T2D than in CON (p<0.001). WITHDRAWN T2D had slightly lower CH2/CH3 ratios even upon adjustment for age, sex and BMI (5.4±0.1 vs. 5.6±0.1, p<0.05). UI and CH2/CH3 did not correlate with m-value (r=0.10, p=0.67 and r=0.11, p=0.63). Of note, CH2/CH3 and UI cor- related with HOMA-IR upon adjustment for age, sex and diabetes diagno- sis (r=0.51, p<0.05; r=-0.58, p<0.05), but not upon adjustment for BMI. Also, oxidation capacity of the octanoyl-carnitine in DSAT did not differ between T2D and CON (1.31±0.38 vs. 1.40±0.72 pmol.g-1.s-1). In conclusion, lower CH2/ CH3 ratios suggest that long chain saturated FA are reduced in DSAT. How- ever only under conditions of fasting but not postprandial hyperinsulinemia, whole-body glucose uptake may relate to FA composition of DSAT in T2D. Supported By: German Center for Diabetes Research Obesity

POSTERS 2001-P Altered Exercise Effects on Microvascular Function in Pediatric

Integrated Physiology/ Obesity BRIAN D. TRAN, ABRAHAM CHIU, GOUTHAM GANESAN, HOANG PHAM, PIETRO GALASSETTI, Irvine, CA While prior studies suggest altered vascular function in pediatric obesity, it is unclear if these alterations extend to physiological exercise-induced vasodilation, an important modulator of long-term cardiovascular risk. We hypothesized that exercise-induced microvascular dilation is reduced in obese (Ob) and/or overweight (OW) children relative to healthy-weight (NW) children. In 9 Ob (13 ± 2 yrs, 97 ± 1 BMI%, 4F), 7 OW (14 ± 2 yrs, 89 ± 3 BMI%, 3F), and 7 NW children (14 ± 2 yrs, 59 ± 15 BMI%, 3F) microvascu- lar function was assessed at the base of the middle fi nger via an optical

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A512 OBESITY—HUMANCATEGORY probe connected to a Perifl ux 5000 Laser Doppler system (Perimed, Sweden). the intervention, weight, waist circumference, visceral adiposity (Bioelectri- Laser doppler fl owmetry (LDF) measurements of skin perfusion were then cal impedance analysis), peripheral, vascular and central aortic BPs (Appla- obtained before, during, and after a 1-min blood fl ow occlusion induced at nation tonometry), as well a metabolic profi le were measured. BMI and low- the wrist at >200 mmHg. Subjects then performed an incremental cycling density lipoprotein cholesterol (LDL-C) were calculated. Data were analyzed exercise test to exhaustion on a metabolic cart/cycle ergometer system with Wilcoxon and Mann Whitney U tests. An Ethic Committee approved the (Carefusion, U.S.), with the incremental phase lasting ~10 min after 2 min protocol and a written informed consent was obtained from all volunteers. of unloaded pedaling. LDF measurements were then repeated at 8 min post In the dapaglifl ozin group, there were signifi cant decreasing of weight exercise. Pre-occlusive steady-state perfusion (Plat1), post-occlusive hype- (69.0 ± 4.7 vs. 66.1 ± 4.7 kg, p <0.001), BMI (28.3 ± 2.8 vs. 27.1 ± 2.8 kg/m2, p remic peak (pMax), post-occlusive steady-state perfusion (Plat2), and time <0.001), waist circumference (92.3 ± 8.1 vs. 88.5 ± 8.2 cm, p <0.04), visceral to reach pMAx (tMax) and Plat2 were obtained from perfusion tracings. The adiposity (83.3 ± 31.3 vs. 59.0 ± 23.7 cm2, p <0.01), uric acid level (291 ± 83 exercise-induced increase in pMax was signifi cantly greater in NW vs. Ob vs. 231 ± 59 µmol/L, p <0.01) and peripheral systolic BP (115 ± 14 vs. 106 ± (+56 ± 22% in NW, +3.7 ± 7% in Ob, p < 0.05). Similarly, the ratio of Plat2/ 11 mmHg, p <0.05; with a signifi cant increment of creatinine concentration Plat1 (%) signifi cantly increased following exercise in NW (from -0.1±7% pre- (35.3 ± 8.8 vs. 70.7 ± 17.6 µmol/L, p <0.001). exe to +35±11% post-exe, p <0.05) and OW (-13 ± 8% pre-exe vs. 50 ± 25% In conclusion, dapaglifl ozin for 3 months in patients with overweight or post-exe, p<0.05), but remained unchanged in Ob (8±10% pre-exe vs. 15±8% obesity without DM, decreased weight, BMI, waist circumference, visceral post-exe, NS). tMax as well as the time of onset of Plat 2 was similar follow- adiposity and peripheral systolic BP, with increment in the creatinine con- ing exercise across all groups. Overall, our cohort of obese children, despite centration. displaying signifi cant adaptations to exercise, appear to have lost the ability to: 1.) further increase maximum dilation, and 2.) sustain the dilation for a 2004-P prolonged period after exercise. Increased Body Weight Provokes Cortisol and Brain Responses to Supported By: National Institutes of Health (UL1 TR001414, P01 HD048721, K24 High Calorie Food Images DK085223) ANIA M. JASTREBOFF, DONGJU SEO, CHERYL LACADIE, ROBERT S. SHERWIN, ROBERT T. CONSTABLE, RAJITA SINHA, New Haven, CT 2002-P Glucocorticoid (GC) exposure increases food intake. However, it is not estab- Plasma Adiponectin Is a Determinant of Treg Abundance in Human lished whether exposure to high-calorie food (HCF) cues (e.g., advertisements) Adipose Tissue and increased adiposity induce changes in endogenous cortisol release which, DAVID BRADLEY, JOEY LIU, STEPHEN BERGIN, ALECIA M. BLASZCZAK, QING in turn, may infl uence brain reward-motivation response and motivation to eat. WANG, VALERIE WRIGHT, BRADLEY NEEDLEMAN, SABRENA NORIA, DEAN In this study, we used functional MRI (fMRI) to assess neural responses to MIKAMI, WILLA A. HSUEH, Columbus, OH sustained exposure to HCF vs. non-food (NF) images in 13 lean (LN) (BMI 22 kg/ Obesity is associated with numerous metabolic complications related to m2) vs. 15 OW/OB (BMI 29 kg/m2) individuals while simultaneously sampling AT infl ammation. A comprehensive system of pro- and anti-infl ammatory cortisol levels and behavioral responses 2 hrs after a standardized meal. HCF mediators and immune cells is required to maintain normal adipose stor- vs. NF produced sustained increases in hunger, wanting and liking (p’s<.0001) age, endocrine function, and systemic insulin action, all critical to whole and increases in brain activation in the hypothalamus, amygdala, striatum, body metabolism. Although adipose tissue macrophages (ATMs) are argu- anterior cingulate cortex, thalamus, insula, and visual regions in whole brain ably the best understood immunomodulators of AT health, adipose resident corrected (WBC), BMI covaried, contrast maps in all subjects (p<.05, WBC). T lymphocytes (ARTs), including pro-infl ammatory Th1 and Th17 cells and Increases in cortisol with HCF relative to NF cue exposure were seen only in anti-infl ammatory Th2 and regulatory T cells (Tregs), have recently been OW/OB and correlated with increases in brain activation during HCF vs. NF cue identifi ed as important metabolic mediators in mice. Tregs protect against exposure in the amygdala, striatum, and insula (Figure). These results raise the obesity-associated infl ammation and related comorbidities. However, little intriguing possibility that in OW/OB individuals GC dysregulation and cortisol is known about ARTs in humans. VAT and SAT biopsies were obtained from increases during HCF cue exposure play a role in greater activation in brain obese (n=119, age 43.7 ± 10.9 yo, BMI 48.0 ± 8.9 kg/m2) and lean (n=13, age reward-motivation pathways and, in turn, increase desire to consume HCF, 42.9 ± 13.8 yo, BMI 23.2 ± 2.0 kg/m2) patients during elective surgery. Adipo- thereby contributing to obesity. cytes and stromal vascular fraction (SVF) were isolated, and SVF was subject Figure. Whole Brain Correlation of Relative Increases in Cortisol with Neu- to fl ow analyses. We found a dramatic increase in Tregs as % of CD4+ cells ral Response to High-Calorie Food vs. Non-Food Images. in lean compared to obese SAT (26.8 ± 8.8% vs. 9.27 ±1.1%; p <0.001) and VAT (29.5 ±12.3% vs. 6.9 ±0.9%; p<0.001), and a decrease in VAT Th2s (1.8 ± 1.1% vs. 9.9 ± 1.2%; p<0.03). BMI and adiponectin in SAT (r = -0.38, p<0.005 and r=0.37, p=0.02) and VAT (r= -0.40, p <0.001 and r=0.40, p= <0.003) strongly correlated with % Tregs. However, we found no signifi cant relation- ship between these phenotypes and % Th1 or Th2 cells. Multivariate linear regression revealed that adiponectin levels were strongly predictive of both SAT and VAT Tregs (p=0.03 and 0.05, respectively). Our data thus demon- strates for the fi rst time that lean vs. obese individuals have signifi cantly higher SAT and VAT Treg abundance and that adiponectin is independently Supported By: National Institutes of Health (R01-DK099039, UL1-DE019586, PL1- predictive of VAT and SAT Treg composition. Further studies are required to DA024859, K23-DK101694, R01-DK20495) determine if adipose tissue T cell subtype changes are responsible for meta- bolic regulation of obese and lean human phenotypes. 2005-P Association between Decreased Insulin Clearance and Initial 2003-P Changes Predisposing toward Obesity in Apparently Healthy Men Effect of Dapaglifl ozin on Visceral Adiposity and Blood Pressure in KAGEUMI TAKENO, YOSHIFUMI TAMURA, SAORI KAKEHI, TAKASHI FUNAYAMA, Patients with Overweight or Obesity without Diabetes Mellitus YASUHIKO FURUKAWA, HIDEYOSI KAGA, RURIKO SUZUKI, YUKI SOMEYA, Obesity MANUEL GONZÁLEZ-ORTIZ, CRISTAL DÍAZ-CRUZ, ANAYELI J. PATIÑO-LAGUNA, RYUZO KAWAMORI, HIROTAKA WATADA, Tokyo, Japan POSTERS LUIS D. LÓPEZ-MURILLO, ESPERANZA MARTÍNEZ-ABUNDIS, Guadalajara, Mexico Hyperinsulinemia is observed in obese subject with insulin resistance and Dapaglifl ozin has shown improve glycemic control, weight and blood pres- caused by not only increased insulin secretion, but also decreased metabolic Integrated Physiology/ sure (BP) in type 2 diabetic patients, however, these effects have not been clearance of insulin (MCRI). Previous studies have suggested that decreased studied in patients with obesity without diabetes mellitus (DM). MCRI is caused by obesity. However, genetic deletion of carcinoembryonic The aim of this study was to evaluate the effect of dapaglifl ozin on vis- antigen-related cell adhesion molecule (CEACAM)-1, impairs MCRI, thus, ceral adiposity and peripheral, vascular and central aortic BPs in patients induces hyperinsulinemia and subsequently developed central obesity and with overweight or obesity without DM. insulin resistance. Therefore, in healthy subjects, reduction of MCRI could A randomized, double-blind, placebo-controlled, clinical trial was carried be the primary change that induces hyperinsulinemia, obesity and insu- out in 32 adults with overweight or obesity [Body mass index (BMI) 25.0-34.9 lin resistance. However, to date, the presence and feature of apparently kg/m2] without DM. Sixteen patients recived 10 mg of dapaglifl ozin (Forxiga, healthy humans with decreased MCRI has not been elucidated yet. Thus, Bristol-Myers Squibb, Humacao, Puerto Rico, U.S.) daily before breakfast we studied 51 nonobese, nondiabetic (BMI<25kg/m2) Japanese men with- during 3 months and the others 16 placebo. At the benning and at the end of out any metabolic risk factors. Using a hyperinsulinemic euglycemic clamp

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with glucose tracer, we evaluated insulin sensitivity (IS) and MCRI at steady that increased visceral adiposity with normal weight is a strong predictor for state of clamp study. Based on the median value of MCRI, we divided the the prevalence of NAFLD in Japanese patients with type 2 diabetes. subjects into low MCRI group and high MCRI group. Compared with high Supported By: Grants-in-Aid for Scientifi c Research from the Ministry of Educa- MCRI group, low MCRI group showed lower muscle IS (6.2±1.5 vs. 7.6±1.8 tion, Culture, Sports, Science and Technology of Japan mg/kg/min, P<0.01) and higher fasting IRI (6.1 (4.8-7.0) vs. 3.7 (2.4-4.6) µIU/ ml, P<0.01) and higher subcutaneous fat area (SFA). Correlation analysis in all 2008-P subjects revealed that MCRI was most signifi cantly correlated to fasting IRI Liver Protein Profi les in Obese Humans With and Without Fatty (r=-0.55, P<0.01) and also correlated to muscle IS (r=0.37, P<0.05), visceral fat Liver Reveal PLIN2 and Novel Protein Markers Associated with area (r=-0.30, P<0.05) and SFA (r=-0.52, P<0.01) respectively. In conclusion, Insulin Resistance we found decreased MCRI in metabolically normal nonobese Japanese men SOFIYA GANCHEVA, MITCHELL PLESSER, SONG NIE, MARIA A. GRITSENKO, and it was closely associated with slightly impaired IS and slightly elevated CHRYSI KOLIAKI, JULIA SZENDROEDI, KIRTI KAUL, TOMAS JELENIK, RICHARD insulin and adiposity levels. Taken together with previous reports, decreased D. SMITH, MATTHIAS SCHLENSAK, WEI-JUN QIAN, MICHAEL RODEN, CHRIS- MCRI may be involved in the initial changes predisposing toward obesity and TOPH BUETTNER, Düsseldorf, Germany, New York, NY, Richland, WA insulin resistance. Nonalcoholic fatty liver disease (NAFLD) is the most frequent liver pathol- ogy in western countries and is tightly linked to obesity and insulin resis- 2006-P tance. To gain better insight into the pathophysiological mechanisms leading Obesity Indices vs. 2-hour Plasma Glucose for Predicting Incident to metabolic liver disease, we performed quantitative proteomic profi ling Diabetes in a 16-Year Prospective Study of Hong Kong Chinese by 2D liquid chromatography-tandem mass spectrometry coupled with KAREN S.L. LAM, YU-CHO WOO, PAUL C.H. LEE, MICHELE M.Y. YUEN, CAROL TMT (tandem mass tags) labeling of liver biopsies to identify differentially H.Y. FONG, ANNETTE W.K. TSO, BERNARD M.Y. CHEUNG, Hong Kong, China expressed proteins. Obese patients with (NAFL+; n=5, age 40±10 yrs, BMI Obesity is an established risk factor of type 2 diabetes. Here we inves- 48.6±3.3 kg/m2, liver fat content, HCL 38±4%) and without hepatic steato- tigated in a relatively nonobese, community-based Hong Kong Chinese sis (NAFL-; n=4, age 36±11 yrs, BMI 48.2±9.7kg/m2, HCL 2±2%) and healthy Cohort, whether obesity indices could replace glycemic measures in pre- lean humans (CON; n=5, age 38±8 yrs, BMI 25.8±3.3 kg/m2, HCL1±2%) were 2 dicting incident diabetes, in a 16-year prospective study. We studied 1505 studied with hyperinsulinemic euglycemic clamps using [6,6- H2] glucose subjects from the Hong Kong Cardiovascular Risk Factor Prevalence Study and underwent a liver biopsy during bariatric or elective abdominal surgery. (CRISPS) who were nondiabetic at baseline (CRISP1: 1995-6), but had dia- Both obese patient groups exhibited peripheral insulin resistance (M-value; betes at follow-up assessments: CRISP2 (2000-4), and/or CRISP3 (2005-8), NAFL+: 2.9±0.4 mg/kg/min, NAFL-: 2.8±0.8 mg/kg/min vs. CON: 7.6±1.9 mg/ and/or CRISP4 (2010-2); or were confi rmed nondiabetic by CRISP4. Glycemic kg/min; p=0.001), but hepatic insulin sensitivity was similar between groups status was tested by 75g OGTT at baseline and follow-up visits. Incident dia- (p=0.64). A total of 178 proteins were differentially expressed in livers from betes was defi ned as FG ≥ 7 mmol/L or 2hG ≥ 11.1 mmol/L or use of antidia- NAFL+ vs. CON, but only 22 of which were also different between NAFL+ and betic agents. By CRISP4, 281 participants had developed diabetes while 1224 NAFL-. Among these 22, PLIN2 expression was 38 and 95% higher in NAFL- remained nondiabetic. On multiple logistic regression including either body and NAFL+ vs. CON, respectively (p=0.0002), and correlated with peripheral mass index (BMI), waist circumference (WC) or waist-hip-ratio (WHR) in the insulin sensitivity (r=-0.61, p=0.028) and HCL (r=0.77, p=0.002) in all partici- model, the most signifi cant predictors of incident diabetes (all p<0.001) were, pants. Furthermore, expression of AVIL (advillin, Ca2+-regulated actin bind- in order of standardized odds-ratios, baseline 2hG, BMI or WC, FG and trig- ing protein), AADAT (transaminase with broad substrate specifi city) and ECI1 lyceride levels. Other signifi cant predictors included age (p=0.001), smoker (mitochondrial enoyl-CoA isomerase involved in unsaturated fatty acid oxi- (p=0.009), hypertension (p=0.033), sex (p=0.037) and WHR (p=0.046). Their dation) was reduced in NAFLD+ vs. CON and related positively with M-value performance in diabetes prediction was assessed by the area under the and negatively with HCL. These results identify proteins and molecular path- curve (AUC) of ROC (receiver-operator curve) analysis. The AUC-ROC of BMI ways that are important in human insulin resistance and NAFLD. and WC were 0.732 and 0.733 respectively, being less favorable than 2hG (0.799; p<0.002; DeLong) but comparable to FG (0.714; p>0.3). However, the 2009-P AUC-ROC of BMI + age (0.773) or WC in women (0.749) did not differ signifi - Suppression of AMPK Is Linked to Activation of TGF-β1/Smad3 Sig- cantly from 2hG. The optimal cut-off by Youden J index for BMI, WC (women) naling in White Adipose Tissue of Obese Humans: Possible Involve- 2 and age were >24.5 kg/m , >79 cm and >45 years respectively. In conclusion, ment in Obesity-induced Insulin Resistance BMI and age or, in women, WC alone, can potentially replace the use of the TING LUO, ALLISON NOCON, JESSICA FRY, ALEX SHERBAN, XIANLIANG RUI, OGTT to identify subjects at increased risk of diabetes in our population for BINGBING JIANG, X. JIAN XU, JINGYAN HAN, YUN YAN, QIN YANG, MENGWEI intensifi ed preventive measures by life-style modifi cation. ZANG, Boston, MA, Kansas City, KS, Irvine, CA Fibrosis is emerging as a hallmark of metabolically dysregulated white adi- 2007-P pose tissue (WAT) in obesity. However, little is known about the pathological Increased Visceral Adiposity with Normal Weight Is Associated mechanism of adipose tissue fi brosis. Here we characterize a role of AMPK in with Prevalence of Nonalcoholic Fatty Liver Disease in Japanese the regulation of extracellular matrix (ECM) in WAT. Histological and patholog- Patients with Type 2 Diabetes ical changes in omental fat depots were compared between obese subjects (n TAKATO TAKEUCHI, RYOTARO BOUCHI, NOHIRHIKO OHARA, YUJIRO NAKANO, = 8) and age-matched normal weight subjects (n = 10). Masson’s Trichrome and MASANORI MURAKAMI, MASAHIRO ASAKAWA, YURIKO SASAHARA, MASU- Sirius Red staining showed elevated collagen fi bers around adipocytes of the YUKI NUMASAWA, ISAO MINAMI, HAJIME IZUMIYAMA, KOSHI HASHIMOTO, pathologically expanded adipose tissue of obese subjects. Immunofl uorescent TAKANOBU YOSHIMOTO, YOSHIHIRO OGAWA, Tokyo, Japan analysis also revealed increased collagen I staining surrounding adipocytes To investigate the impact of increased visceral adiposity with normal in visceral adipose tissue, suggesting excessive fat collagen deposition as a weight (OB(-)VA(+)) on the prevalence of nonalcoholic fatty liver disease critical pathological process in human obesity. Phosphorylation and activity (NAFLD) in patients with type 2 diabetes. This was a cross-sectional study of of AMPK was diminished in adipocytes of obese subjects without affecting 140 Japanese patients with type 2 diabetes (mean age 65 ± 11 years; 44.6% AMPKα. Conversely, positive cells for TGF-β1 and phosphorylated Smad3 2 Obesity female). Visceral fat area (VFA, cm ) and liver attenuation index (LAI) were were highly increased and some inappropriately activated α-smooth muscle

POSTERS assessed by abdominal computed tomography. The patients were divided actin (α-SMA)+ myofi broblast-like cells were observed. Consequently, fasting into 4 groups by VFA and body mass index (BMI, kg/m2) as follows; BMI < 25 glucose levels were elevated in obese subjects as compared to controls. To 2 2 2 2 Integrated Physiology/ kg/m and VFA < 100 cm (OB(-)VA(-)), BMI ≥ 25 kg/m and VFA < 100 cm determine a causal relationship between WAT fi brosis and insulin resistance (OB(+)VA(-)), BMI < 25 kg/m2 and VFA ≥ 100 cm2 (OB(-)VA(+)), and BMI ≥ 25 related to obesity, our in vitro studies showed that TGF-β1 caused an impair- kg/m2 and VFA ≥ 100 cm2 (OB(+)VA(+)). Multivariate linear regression and ment of insulin-induced phosphorylation of Akt and suppression of lipolysis in logistic regression analysis were done to determine the impact of OB(-)VA(+) 3T3L1 adipocytes. TGF-β1-induced insulin resistance was attenuated by AMPK on LAI. In this study, 25.0% were OB(-)VA(+) patients, where the LAI lev- agonists including AICAR and Metformin. The effect of Metformin on insulin els were lower (1.09 ± 0.22) than those in OB(-)VA(-) patients (1.23 ± 0.15), sensitivity was recapitulated by the constitutively active AMPK. Collectively, and were equivalent to those in OB(+)VA(+) patients (1.03 ± 0.26). In multi- inhibition of AMPK likely integrates with overproduction of TGF-β1 to promote variate linear regression analysis, OB(-)VA(+) was independently associated the development of aberrant collagen deposition, myofi broblast-like cell acti- with LAI (standardized β -0.212, p = 0.014). In multivariate logistic regression vation, and insulin resistance in WAT of obese individuals. analysis, OB(-)VA(+) was a signifi cant predictor of LAI < 0.9 (odds ratio 5.88, Supported By: American Diabetes Association (1-15-BS-216 to M.Z.); National 95% confi dence interval 1.03-33.52, p = 0.046). This study provides evidence Institutes of Health

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2010-P 2012-P Increasing Adiposity Is Associated with Increasing Pro- and Body Composition and Type 2 Diabetes Prevalence in the MESA and Decreasing Anti-infl ammatory Adipokines: A Role for Leptin MASALA Studies MARY HELEN BLACK, YU-HSIANG SHU, JUN WU, CORINNA KOEBNICK, ADRI- ELENA FLOWERS, MATTHEW ALLISON, JEFFREY CARR, JINGZHONG DING, ENNE MACKAY, RICHARD M. WATANABE, THOMAS A. BUCHANAN, ANNY H. DAVID HERRINGTON, NAMRATHA KANDULA, KIANG LIU, KEVIN MILES, RAVI XIANG, Pasadena, CA, Los Angeles, CA SHAH, ALKA M. KANAYA, San Francisco, CA, San Diego, CA, Nashville, TN, Win- Higher pro- and lower anti-infl ammatory adipokine levels have been ston-Salem, NC, Chicago, IL, Boston, MA shown to be associated with obesity in cross-sectional studies, but longi- A previous study using data from the Multi-Ethnic Study of Atheroscle- tudinal data on changes in adipokines and adiposity are limited. We exam- rosis (MESA) and Mediators of Atherosclerosis in South Asians Living ined these relationships in 352 BetaGene participants with baseline and in America (MASALA) cohorts found that South Asians have signifi cantly follow-up measures (73% female; mean±SD age 34.9±8.2 yrs, BMI 29.3±5.9 higher prevalence of type 2 diabetes (T2D) (26%) compared to whites (6%), kg/m2, follow-up 4.6±1.5 yrs). Total body and trunk fat were assessed by African- (18%), Hispanic- (17%), and Chinese-Americans (13%). A high pro- DXA. Adiponectin, IL-1β, IL-6, IL-1Ra, leptin, lipocalin, MCP-1, resistin and portion of ectopic adipose tissue is hypothesized to be an underlying cause TNF-α were assayed using Millipore multiplex kits, and apelin, CRP, DPP- of the T2D burden in South Asians. Using data collected from participants in 4, visfatin, SFRP4 and SFRP5 with ELISA. Generalized estimating equations MESA and MASALA, we evaluated whether differences in body composition (GEE) were used to test associations between rates of change in each of 3 between race/ethnic groups explain the differences in T2D prevalence. Iden- adiposity measures and rates of change in 15 adipokines, adjusted for age, tical abdominal and cardiac computerized tomography protocols were used sex and baseline adiposity. Canonical correlation analysis (CCA) was used to for body composition measures. We calculated multivariate adjusted preva- assess the joint multivariate association between changes in adiposity and lence of T2D by race/ethnicity and z-scores to determine if the estimates adipokines. By GEE, increasing BMI, total body and trunk fat were associ- differed from South Asians. Model 1 was adjusted for age, sex, study site, ated with decreasing adiponectin (all p<0.001) and SFRP5 (all p<0.0001), and education, income, smoking, alcohol, exercise, BMI, HDL, triglycerides, and increasing CRP (all p<0.0001), IL-6 (all p<0.02), leptin (all p<0.0001), IL-1Ra (all hypertension. Model 2 included the prior variables and liver fat attenuation, p<0.004) and SFRP4 (all pɖ0.05). CCA revealed that 81% of the shared varia- abdominal visceral fat area, abdominal intermuscular fat, and pericardial fat tion between rates of change in adiposity and adipokines was explained by volume. Inclusion of body composition variables did not meaningfully atten- a single underlying component of each (p<0.0001). The adiposity component uate the differences in T2D prevalence between race/ethnic groups. The was well captured by rates of change in BMI, total body and trunk fat (ρ=0.88 higher T2D prevalence among South Asians remains unexplained by body to 0.96), while the adipokine component was most represented by rates of composition and other measured risk factors. change in leptin (ρ=0.87), CRP (ρ=0.42), SFRP5 (ρ=-0.38), IL-1Ra (ρ=0.33) and Table. Prevalence (95% CI, p-value) of Type 2 Diabetes by Race/Ethnic Group. adiponectin (ρ=-0.31). Joint association between changes in adiposity and adipokines was most accounted for by changes in leptin, followed by CRP, South Asian White African American Hispanic Chinese American n=724 n=741 n=390 n=484 n=243 SFRP5, IL-1Ra and adiponectin. These data suggest that increasing adiposity contributes to a growing imbalance in pro- and anti-infl ammatory adipokines Model 1 over time, in which leptin may have an important role. Men 23 (17, 29) 6 (4, 9; p<0.001) 17 (10, 23; p=0.10) 17 (12, 23; p=0.09) 14 (7, 20; p=0.02) Supported By: National Institute of Diabetes and Digestive and Kidney Diseases Women 19 (11, 28) 5 (2, 7; p=0.001) 17 (11, 24; 0.74) 9 (6, 13; p=0.03) 7 (2, 11; 0.006) Model 2 2011-P Men 26 (18, 34) 6 (4, 9; p<0.001) 18 (11, 26; p=0.28) 17 (12, 23; p=0.16) 13 (6, 20; p=0.02) Risk of Incident Hypertension According to Metabolic Health and Obesity: Defi nition of Metabolic Health Does Not Matter Women 18 (10, 27) 5 (2, 7; p=0.001) 20 (13, 28; p=0.77) 9 (6, 12; p=0.03) 6 (2, 11; p=0.006) YU MI KANG, CHANG HEE JUNG, JUNG EUN JANG, JOONG-YEOL PARK, HONG- Supported By: 1R01 HL093009, K24HL112827, UL1 RR024131, N01-HC-95159, KYU KIM, WOO JE LEE, Seoul, Republic of Korea N01-HC-95160, N01-HC-95161, N01-HC-95162, N01-HC-95163, N01-HC-95164, Metabolically healthy obese (MHO) phenotype refers to obese individuals N01-HC-95165, N01-HC-95166, N01-HC-95167, N01- HC-95168, N01-HC-95169, with a favorable metabolic profi le. Its prognostic value remains controversial UL1-TR-000040, UL1-TR-001079, 5R01HL085323-04, RO1-HL088451 and may partly depend on differences in defi ning methods. The aim of the present study was to investigate whether the MHO phenotype is associated 2013-P with an increased risk of incident hypertension in a Korean population while Adipose Tissue Macrophage Burden: The Relationship to Insulin adapting various defi nitions of metabolic health. The study population com- Sensitivity, Physical Fitness, and Weight Loss prised 31033 Koreans without hypertension. Participants were stratifi ed by 2 MARIA MORGAN-BATHKE, ANA ESPINOSA DEYCAZA, ESBEN SONDERGAARD, body mass index (cut-off value, 25.0 kg/m ) and metabolic health state, using MICHAEL D. JENSEN, La Crosse, WI, Rochester, MN 4 different defi nitions: Adult Treatment Panel (ATP)-III criteria, Wildman cri- It has been suggested that adipose tissue infl ammation in obesity is related teria, Karelis criteria, and the homeostasis model assessment (HOMA) index. to insulin sensitivity, but whether this includes adipose tissue lipolysis insu- Over the median follow-up period of 35.0 months (range, 4.5-81.4 months), lin action, as expressed by the insulin concentration needed to suppress FFA 4589 of the 31033 individuals (14.8%) developed incident hypertension. fl ux by 50% (IC ), has not been examined. We measured IC physical fi tness Compared with the metabolically healthy nonobese group, the MHO group 50 50, (VO2 peak) and abdominal and femoral subcutaneous adipose tissue mac- showed increased risk of incident hypertension with multivariate-adjusted rophage burden (fl ow cytometry) in 22 adults pre-weight loss (12 males) and hazard ratios of 1.42 (95% confi dence interval [CI], 1.30-1.55), 1.44 (95% CI 5 post-weight loss (2 males). CD68 was used to identify total macrophage 1.32-1.58), 1.41 (95% CI 1.32-1.58), and 1.33 (95% CI 1.22-1.45), when defi ned numbers, CD14 for pro-infl ammatory (M1) and CD206 for anti-infl ammatory by ATP-III criteria, Wildman criteria, Karelis criteria, and HOMA index, (M2) macrophages. Surprisingly, we found no relationship between IC50 and respectively. Metabolically unhealthy obese individuals were at the highest abdominal (female p = 0.3, male p = 0.47) or femoral (female p = 0.98, male risk of incident hypertension. MHO subjects showed a substantially higher p = 0.62) macrophage burden. There was no relationship between physical risk of incident hypertension regardless of the defi nition of metabolic health fi tness (VO2 peak/kg FFM) and abdominal (female p = 0.38, male p = 0.3) used. Thus, it is important to consider both metabolic health and obesity or femoral (female p = 0.9, male p = 0.69) total macrophage burden. How- Obesity when evaluating the risk of hypertension. POSTERS ever, we did fi nd a negative relationship between physical fi tness (VO2 peak/ kg FFM) and abdominal pro-infl ammatory macrophage content for males

(p = 0.07). There was a decrease in the ratio of M1 to M2 macrophages Integrated Physiology/ following weight loss in both abdominal (p = 0.09) and femoral (p = 0.03) depots. These results suggest that adipose macrophage content is not a strong predictor of adipose tissue insulin resistance, but that decreases in pro-infl ammatory relative to anti-infl ammatory adipose macrophages results from weight loss. Supported By: National Institutes of Health

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A515 OBESITY—HUMANCATEGORY

2014-P angiogenesis is associated with over-expression of anti-antigenic factor Impact of Brown Adipose Tissue on Visceral Obesity and Percent VEGF-A165b within the adipose milieu and in blood. In the present study, Body Fat: A Prospective Matched Case-Control Study we postulated that up-regulation of Wnt5a is associated with the blunted JU HEE CHOI, HYE SOO CHUNG, HYE JIN YOO, JI A. SEO, SIN GON KIM, NAN HEE adipose tissue angiogenesis and examined the role of Wnt5a in regulating KIM, DONG SEOP CHOI, HYUN JUNG LEE, SEI HYUN BAIK, KYUNG MOOK CHOI, VEGF165b expression in human obesity. We biopsied subcutaneous and vis- Seoul, Republic of Korea ceral adipose tissue from 37 obese individuals (BMI 43±3 kg/m2, age 41±3 Objective: This prospective matched case-control study aimed to evaluate years) during planned bariatric surgery and characterized depot-specifi c if brown adipose tissue (BAT) is associated with visceral obesity, percent protein expression of VEGF-A165b and Wnt5a using Western blot analysis. body fat (PBF), and cardiometabolic risk factors in humans. In both subcutaneous and visceral fat, VEGF-A165b expression correlated Research Design and Methods: In total, 45 BAT-positive and 45 BAT-nega- positively with Wnt5a (r2=0.8, p<0.001). In subcutaneous fat where adipose tive subjects were enrolled. All subjects were matched with respect to age, tissue angiogenic capacity is greater than visceral fat, exogenously admin- sex, and season/year of 18F-fl uorodeoxyglucose-positron emission tomogra- istered recombinant wnt5a increased whole adipose tissue and vascular phy/computed tomography (FDG-PET/CT). All follow-up measurements were endothelial cell VEGF-A165b expression (p<0.05). This was associated with performed after a median time of 23.0 (16.0, 31.0) months. markedly blunted adipose tissue angiogenic sprout formation capacity in Results: Among the 6,877 consecutive FDG-PET/CT scans in 4,736 sub- fat pad tissue explants from the subcutaneous depot. Moreover, recombi- jects, 146 subjects (3.1%) had a positive BAT scan. The BAT-positive group nant Wnt5a increased soluble Flt-1 concentration, a negative regulator of and the matched negative group did not differ signifi cantly in any component angiogenesis, in the angiogenic sprout media (p<0.05). We demonstrated a of metabolic syndrome (all P >0.05), with the exception of triglyceride levels signifi cant link between Wnt5a and anti-angiogenic VEGF-A165b expression (P = 0.04). Furthermore, free T4 (P = 0.66), TSH (P = 0.91), homeostasis model in the fat of obese subjects which was associated with impaired angiogenic assessment of insulin resistance (HOMA-IR) (P = 0.16), and high-sensitivity capacity. Wnt5a over-expression in obesity may be a negative regulator of C-reactive protein (hsCRP) values (P = 0.90) were not different between the angiogenesis in obesity. two groups. However, multiple stepwise regression analysis revealed that Supported By: National Institutes of Health presence of BAT, age, waist circumference, HDL, and TSH were indepen- dently associated with the visceral fat area (VFA) as measured by abdominal 2017-P CT (R 2 = 0.60). Furthermore, the presence of BAT, waist circumference, and Weight Loss followed by Weight Maintenance Obtained With or HOMA-IR were independent factors affecting bioelectrical impedance anal- Without GLP-1 Receptor Agonist Treatment Decreases Branched- ysis (BIA)-measured PBF (R 2 = 0.70) after adjusting for confounding factors. Chain Amino Acids Conclusion: The presence of BAT, as detected by FDG-PET/CT, was inde- SIGNE S. TOREKOV, LINE ENGELBRECHTSEN, EVA P. IEPSEN, EHM ANDERSSON, pendently associated with VFA and PBF in adult Asian women during a rela- YUVARAJ MAHENDRAN, JULIE LUNDGREN, ANNA JONSSON, STEN MADS- tively short-term follow-up period. BAD, JENS JUUL HOLST, HENRIK VESTERGAARD, TORBEN HANSEN, Copenha- gen, Denmark, Hvidovre, Denmark 2015-P Increased levels of circulating branched chain amino acids (BCAAs) and Role of the Autonomic Nervous System Function in the Adaptative tyrosine have been suggested to be involved in the pathogenesis of insu- Capacity of Left Ventricle Contractility to Hypervolemia in Obese lin resistance. However, it is unknown how metabolites are affected during Patients with Glucose Intolerance long-term weight maintenance after weight loss, and how treatment with AMEL REZKI, BADREDDINE MERIOUD, MARINOS FYSEKIDIS, SABRINA CHI- glucagon-like peptide 1 receptor agonists (GLP-1 RAs) can affect metabolite HEB, ISABELA BANU, EMMANUEL COSSON, PAUL VALENSI, Bondy, France, Paris, profi les. Thus, we aimed to characterize amino acid and glycolysis related France metabolite changes after an acute weight loss intervention followed by Hypervolemia, an increase in cardiac output and alterations in cardio- long term weight maintenance in a group of GLP-1 RA-treated obese indi- vascular autonomic activity are clearly demonstrated in obese patients. This viduals and control participants. Fifty-eight obese individuals underwent a study aimed to compare hemodynamic parameters in obese patients with diet-induced 12% body weight loss during 8 weeks. Participants were ran- normal (NGT) or impaired glucose tolerance (IGT), and examine their relation- domized to weight maintenance with or without administration of the GLP-1 ship with autonomic nervous system activity. We included 66 nondiabetic RA liraglutide (1.2 mg/day) for 52 weeks. Metabolomic profi ling by a high- obese patients with normal blood pressure and without cardio-vascular his- throughput proton nuclear magnetic resonance spectroscopy platform was tory, 38 NGTs and 28 IGTs. Cardiac vagal and sympathetic activity (HF-HR, used for quantifi cation of metabolites. The weight loss was maintained in LF-HR) were assessed by spectral analysis of heart rate variations. Cardiac both groups and was associated with 9-32% decreases in plasma concen- index (CI), stroke volume (SV) and thoracic fl uid content (TFC) were measured trations of alanine, phenylalanine, histidine, tyrosine and the BCAAs leucine, in 39 of them by thoracic impedance (Task Force Monitor® fi n g e r p l e t h y s - isoleucine and valine (p<0.05). Plasma citrate levels increased during weight mography). In IGTs, cutaneous blood fl ow (CBF, mean and standard devia- loss (p=5.2 x 10-15) and showed inverse correlation with HOMA-IR levels of tion) was assessed by laser doppler method, and glucose variability (GV) was -0.318 (p=0.025), refl ecting decrease in insulin resistance. The control group evaluated by calculating CONGA from 24 h-continuous glucose monitoring. had lower levels of valine compared to the GLP-1RA group during weight IGTs were separated into 2 groups according to HF-HR peak: below (G1-IGTs) maintenance (p=0.005). We demonstrate that acute weight loss is associ- and over (G2-IGTs) the median value. IGTs had similar sex-ratio, age, BMI but ated with marked changes in plasma concentrations of eight amino acids higher CI, SV and TFC (p<0.04 to <0.0001) than NGTs. G2-IGTs were younger and glycolysis related metabolites and that the changes in plasma concen- than G1-IGTs patients (p<0.0001). In G2-IGTs, CI, SV and CONGA were higher trations of the suggested T2D risk markers (BCAAs and tyrosine) remain low than in G1-IGTs (p<0.01 to p<0.0001) with higher LF-HR peak (p<0.05), and during long-term weight maintenance, consistent with a decreased T2D risk. similarly after age adjustment. In IGTs LF-HR correlated with CI (p<0.02) and GLP-1 RA treatment may infl uence the concentration of valine, pointing to a negatively with standard deviation of CBF (p<0.01), and CONGA correlated link between GLP-1, valine and glucoregulation. with CI and SV (p<0.05 for both). To conclude, among obese patients, IGTs Supported By: The Danish Research Council for Health and Disease; Danish exhibit higher thoracic fl uid content than NGTs. IGTs with preserved cardiac Ministry of Science, Technology and Innovation; Danish Diabetes Academy; Cam- bridge Weight Plan

Obesity autonomic activity preserve an adaptative capacity to increase left ventri-

POSTERS cle contractility probably due to a greater sympathetic activity which also results in a higher glucose variability and lower CBF variability. 2018-P

Integrated Physiology/ Exenatide Led to a Signifi cant Decrease in Liver Fat Content and 2016-P Epi cardial Adipose Tissue in Obese Type 2 Diabetic Subjects: A Wnt5a in Adipose Tissue Regulates Angiogenesis via Activation of Prospective, Randomized Clinical Trial ANNE DUTOUR, INES ABDESSELAM, PATRICIA ANCEL, FRANK KOBER, GEORGES Antiangiogenic Isoform VEGFA165b in Obese Subjects SHAKUN KARKI, DOAN T.M. NGO, MELISSA G. FARB, BRIAN CARMINE, DONALD MRAD, PATRICE DARMON, OLIVIA RONSIN, VINCENT PRADEL, NATHALIE T. HESS, NOYAN GOKCE, Boston, MA LESAVRE, ALEXIS JACQUIER, MONIQUE BERNARD, BÉNÉDICTE GABORIT, Mar- Experimental studies suggest that Wnt5a is a pro-infl ammatory secreted seille, France protein that is associated with metabolic dysfunction in obesity. Impaired Objective: Ectopic adiposity has been associated with type 2 diabetes angiogenesis in adipose tissue has been implicated in development of (T2D) complications. Glucagon-like-peptide-1 analogues are currently used adipose tissue capillary rarefaction, hypoxia, infl ammation and metabolic in T2D, but no prospective randomized trial has investigated their effect on dysfunction. Our recent work demonstrated that impaired adipose tissue ectopic fat storage.

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A516 OBESITY—HUMANCATEGORY

Research Design and Methods: 44 obese type 2 diabetics uncontrolled on (GLP-1), peptide YY (PYY), cholecystokinin (CCK)], hunger hormone [ghrelin], oral-antidiabetic-drugs were randomly assigned to receive Exenatide or ref- pancreatic hormones [Insulin, glucagon and amylin (active and total)] and erence treatment according to French guidelines. Epicardial adipose tissue leptin were collected at baseline and after 30, 60, 90, 120, 180 and 240 (EAT), myocardial (MTGC), hepatic (HTGC), and pancreatic (PTGC) triglyceride minutes from starting the meal. Positive area under the curve (AUC0-240) for content were assessed 45 minutes after standardized meal with 3T mag- each hormone was calculated. Positive AUC0-240 for GLP-1 and PYY were netic resonance imaging and 1H-magnetic resonance spectroscopy before signifi cantly higher after GL and UGC than after OM (p<0.001 for both), but and after 26 weeks of treatment. was not different between the 3 meals for CCK and ghrelin. Although, posi- Results: Study population had a mean HbA1c= 7.5±0.2% and mean tive AUC0-240 for insulin was not different between the 3 meals, positive 2 BMI=36.1±1.1 kg/m . Ninety fi ve percent had hepatic steatosis at baseline AUC0-120 was signifi cantly higher after UGC than after OM (p=0.02). Posi- (HTGC≥5.6%). Exenatide and reference group led to similar improvement in tive AUC0-240 for glucagon was signifi cantly higher after GL and UGC than HbA1c (-0.7±0.3 vs. -0.7±0.4%, p=0.29) whereas only Exenatide group led after OM (p<0.001 for both), but was not different between the 3 meals for to signifi cant weight loss (-5.5±1.2 vs. -0.2±0.8 kgs, p=0.001 for the differ- amylin (active and total), and leptin. This study shows that intake of DSNFs ence between groups). Remarkably, Exenatide induced signifi cant reduction signifi cantly increases secretion of 2 gut satiety hormones, in addition to in EAT (-8.8±2.1%), and HTGC (-23.8±9.5%), compared to reference treated insulin and glucagon. This may partially explain their effect on blood glu- group, respectively (-1.2±1.6%, p=0.003 and +12.5±9.6%, p=0.007). No sig- cose and body weight. However, these results warrant further evaluation nifi cant difference was observed in other ectopic fat stores PTGC or MTGC. in longitudinal trials. In the group treated with Exenatide, reductions in liver fat and EAT were not associated with HOMA-IR, adiponectin, HbA1c, or fructosamin change, 2021-P but were signifi cantly related to weight loss (r=0.47, p=0.03, and r=0.50, Overall and Central Obesity in Relation to Insulin Sensitivity and p=0.018, respectively). Secretion: Genetic Evidence from a Han Chinese Population Conclusion: Our data indicate that Exenatide is an effective treatment to FENG JIANG, CHENG HU, WEIPING JIA, Shanghai, China reduce liver fat content and epicardial fat in obese patients with type 2 dia- We aimed to validate genetic variants as an instrument for measuring betes, and these effects are mainly weight loss dependent. overall and central obesity and to characterize the role of these variants in insulin sensitivity and secretion in a Chinese population. We genotyped a 2019-P total of 38 established loci associated with body mass index (BMI; a surro- Infl uence of Metabolically Healthy Obesity on the Progression of gate of overall obesity) and waist circumference (WC; a surrogate of central Coronary Artery Calcifi cation obesity). Then, using the genetic risk score (GRS), we examined the associa- JI WON YOON, CHAN-HYEON JUNG, MIN-KYUNG KIM, HYO EUN PARK, KYONG tions of these loci with a range of glycaemic traits in 2884 individuals from a SOO PARK, HAK CHUL JANG, MIN KYONG MOON, SU-YEON CHOI, BOKYUNG community-based population with Han Chinese ancestry. Our analyses vali- KOO, Seoul, Republic of Korea, Seongnam, Republic of Korea dated the effects of BMI-GRS based on 32 BMI loci and WC-GRS based on There have been confl icting data whether metabolically healthy obesity 6 WC loci in a community-based population comprised of middle-aged Han (MHO) increases the risk of cardiovascular disease. We investigated the Chinese adults (P=4.62×10-13 and P=0.007, respectively). More importantly, infl uence of MHO on the progression of coronary artery disease using the we found a stronger predisposition for increased BMI with more subcutane- coronary artery calcium score (CACS). A retrospective cohort of men with ous fat area, higher insulin levels and greater insulin resistance, as assessed the serial measurement of CACS at least 12 months apart (n=1,856) was with the homeostatic model assessment of insulin resistance (HOMA-IR) assessed; CACS progression was defi ned considering the annual CACS without adjustment for central obesity (P range from 0.007 to 0.03). We change and baseline CACS. MHO was defi ned as obesity not accompanied also demonstrated associations between increased BMI-GRS and increased by metabolic syndrome. Infl uence of number of metabolic abnormalities Stumvoll 1st phase and Stumvoll 2nd phase insulin secretion (P=3×10-4 and (triglyceride ≥ 150 mg/dL, HDL cholesterol <40 mg/dL, fasting glucose ≥ 100 7×10-4, respectively), which were independent of central obesity. In con- mg/dL, and blood pressure ≥ 130/85 mmHg) accompanying obesity as well trast, each additional WC-increasing allele in the GRS was associated with as MHO on CACS progression was investigated by cox regression analy- higher glucose (P<0.018) and lower insulin sensitivity (P=0.030), as assessed sis. When compared to nonobese/metabolically healthy subjects (reference with the Gutt index. Our results indicate that the combined effects of group), the MHO group had an increased risk of CACS progression (age- genetic variants conferring obesity susceptibility on type 2 diabetes mellitus adjusted hazard ratio [HR] 1.351, 95% confi dence interval [CI] 1.111-1.643, (T2DM)-related traits varied between overall obesity and central obesity in P = 0.003). Of the subjects with MHO at baseline, 31% developed meta- Han Chinese populations. This fi nding suggests that predisposition to overall bolic syndrome during the follow-up period, which was signifi cantly higher obesity might have an effect on insulin secretion, whereas predisposition to compared to that of the reference group (HR 1.767, 95% CI 1.353-2.308, P = central obesity may affect insulin sensitivity. 0.001). By contrast, obesity without any metabolic abnormality did not Supported By: National Natural Science Foundation of China (81322010, increase the risk of metabolic syndrome (HR 1.102, 95% CI 0.555 - 2.186, P = 81570713) 0.782) or CACS progression (HR 1.366, 95% CI 0.911 - 2.047, P = 0.131) during the follow-up period. In conclusion, MHO was unstable state; about 30% of 2022-P subjects with MHO experienced metabolic deterioration during the follow- Alterations in Visceral Adipose Tissue Innate Type 2 Lymphoid Cells up period. It also increased the risk of coronary artery disease progression (ILC2s) as Determinants of Metabolic Health in Obese Patients compared to nonobese/metabolically healthy subjects. More strict defi nition STEPHEN BERGIN, JOEY LIU, DAVID BRADLEY, ALECIA BLASZCZAK, QIAN of MHO might differentiate the benign phenotype of obesity. WANG, VALERIE WRIGHT, RYAN JUDD, MICHAEL CALIGIURI, BRADLEY NEEDLE- MAN, SABRENA NORIA, DEAN MIKAMI, WILLA A. HSUEH, Columbus, OH 2020-P A balance of pro- and anti-infl ammatory mediators and immune cells in adi- The Effect of Diabetes-specifi c Nutritional Formulas on Gut Satiety pose tissue (AT) maintains normal adipose storage, endocrine function, and and Hunger Hormones, Pancreatic Hormones, and Leptin systemic insulin action, all critical to whole body metabolism. Multiple cells ADHAM MOTTALIB, BARAKATUN-NISAK MOHD-YUSOF, MAHMOUD SAKR, in lean AT in mice including alternatively-activated macrophages, eosino-

WAEL MOHAMED, DAVID M. POBER, JOHN TROUP, JOANNA MITRI, OSAMA phils, type 2 innate lymphoid cells (ILC2s), invariant natural killer T (iNKT) Obesity

HAMDY, Boston, MA, Aliso Viejo, CA cells, and CD4+ Th2 and Treg cells work in concert to establish a type 2 anti- POSTERS Diabetes-specifi c nutritional formulas (DSNFs) are used by patients with infl ammatory microenvironment. ILCs are a family of innate immune cells

diabetes as part of medical nutrition therapy with the aim of improving that belong to the lymphoid lineage but do not respond in an antigen-specifi c Integrated Physiology/ glycemic control and reducing body weight. However, their mechanisms manner, as they lack a B or T cell receptor. Lack of ILC2s in mice results in of action are unclear. This study was designed to evaluate the effect of weight gain and insulin resistance, and ILC2 cells promote beiging of fat. AT DSNFs on gut satiety and hunger hormones, pancreatic hormones and lep- ICL2s are decreased in obese vs. lean individuals. In order to better under- tin. We compared postprandial (PP) effect of 2 DSNFs; Glucerna (GL, Abbott stand the role of ILC2s in human AT, VAT and SAT biopsies were obtained Nutrition) and Ultra Glucose Control (UGC, Metagenics) vs. oatmeal (OM, from obese (n=43, age 45.9 ± 10.7 yo, BMI 46.6 ± 11.1 kg/m2) patients dur- Quaker). After an overnight fast and withholding diabetes medications, 22 ing elective surgery. Adipocytes and stromal vascular fraction (SVF) were patients with type 2 diabetes and BMI≥25 kg/m² (mean age 62.3±6.8 years; isolated, and SVF was subjected to fl ow analyses. In obese patients, the A1c 6.8±0.7%, body weight 97.4±21.3 kg and BMI 33.2±5.9 kg/m²) were % ILC2s (CD161+CD25+CD294+) in VAT were negatively associated with given 200 kcal of each of the three meals in random sequence on 3 sepa- both plasma insulin (r= -0.429, p=0.026) and insulin resistance measured by rate days. Blood samples for gut satiety hormones [glucagon-like peptide-1 HOMA-IR (r= -0.464, p=0.015), but not age, BMI, or plasma adiponectin. The

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A517 OBESITY—HUMANCATEGORY

% ILC2s also trended to correlate with immunsuppressive Tregs (as % CD4+ Table. 4-Month Changes by % Weight Loss (N =317; Metformin Users cells) and to inversely correlate with % CD4+ pro infl ammatory Th1 cells. In Excluded from this Analysis). contrast, % ILC2s in SAT demonstrated no associations. These data suggest 2.5-4.9 p-value 5-6.9 p-value ≥7 p-value that in humans VAT ILC2s may be a determinant of hyperinsulinemia and (N=67/21%) (n=69/22%) (N=142/45%) insulin resistance, and provide a therapeutic target in diseases associated Total Cholesterol (mg/dl) -5.5 (20.8) 0.06 -11.1 (25.1) 0.0002 -17.9 (25.7) <.0001 with insulin resistance. HDL Cholesterol (mg/dl) -4.7 (9.0) <.0001 -4.5 (11.7) 0.001 -6.6 (9.1) <.0001 2023-P Triglycerides (mg/dl) -12.7 (42.6) 0.01 -23.5 (48.0) <.0001 -26.7 (48.7) <.0001 Targeting Infl ammation with Salicylate (Salsalate) Improves Glyce- LDL Cholesterol (mg/dl) +3.2 (22.1) 0.22 -1.6 (20.6) 0.30 -6.1 (24.5) 0.005 mia over 30 Months in a Randomized Trial of Overweight Patients Systolic BP (mmHg) -5.9 (16.0) 0.01 -8.3 (17.6) 0.0001 -11.5 (14.8) <.0001 without Diabetes and with Stable Cardiovascular Disease Diastolic BP (mmHg) -2.5 (8.3) 0.04 -4.0 (8.4) 0.0002 -7.1 (8.4) <.0001 NINAD SALASTEKAR, TANVI DESAI, ERNST SCHAEFER, THOMAS HAUSER, MEL- Glucose (mg/dl) +1.4 (8.5) 0.25 -0.21 (11.4) 0.34 -2.5 (7.6) 0.0003 VIN CLOUSE, STACEY JOSEPH, KRISTEN FOWLER, STEVEN E. SHOELSON, ALLI- SON B. GOLDFINE, Boston, MA Data cells contain Mean (SD). Type 2 diabetes (T2D) is more common in patients with cardiovascular Supported By: National Institute of Diabetes and Digestive and Kidney Diseases disease, and infl ammation may underlie pathophysiology of both conditions. We assessed long-term effi cacy and safety of targeting infl ammation using salsalate to improve glycemia in statin-treated overweight persons with 2025-P Ethnic Difference in Glucose Effectiveness and Disposition Index: A stable coronary heart disease, without T2D. Compensatory Mechanism for Glucose Homeostasis in Overweight/ Glycemia was assessed in 192 participants without T2D at baseline in a Obese African American and White Women with Prediabetes pre-specifi ed secondary analysis from Targeting INfl ammation Using SAL- TRUDY R. GAILLARD, KWAME OSEI, Cincinnati, OH, Columbus, OH salate in CardioVascular Disease (TINSAL-CVD), a multi-center, double- Objective: Prediabetes varies among ethnic populations. Therefore, we masked, randomized (1:1), placebo-controlled, parallel clinical trial. compared the pathophysiologic mechanisms of prediabetes in overweight/ Participants were mostly Caucasian males, age 60±7 yrs, BMI 31.4 ± 3.0 kg/ obese African American (AA) and white American (WA) women. m2, fasting blood glucose (FBG) 92.8 mg/dL ± 11.0 mg/dL, and HbA1c 5.8% ± Methods: We recruited 95 women (67 AA, 28 WA) with prediabetes. Stan- 0.3%. Reductions in mean FBG -5.70 mg/dL (95% CI: -7.44 to -3.97 mg/dL, dard OGTT and FSIVGTT were performed in each subject. Insulin sensitivity P<0.001), HbA -0.11% (95% CI: -0.210 to -0.002%, P=0.046) and glycated 1c (Si), glucose effectiveness (Sg), beta-cell function (acute insulin response serum protein (GSP) -81.8 µg/mL (95% CI: -93.7 to -69.9 µg/mL, P<0.001) to glucose (AIRg) and disposition index (DI: Si x AIRg) were calculated using were seen in salsalate compared to placebo over 30 months. FBG and GSP Bergman’s Minmod. decreased in salsalate compared to placebo in participants with and with- Results: Mean BMI was greater in AA vs. WA with prediabetes (38.3±8.2 out prediabetes (normal glucose tolerance (NGT)) with greater decreases vs. 34.6±8.5 kg/m2, p=0.05). Mean fasting serum glucose, and insulin levels in the prediabetes (FBG -7% vs. -5%, and GSP -37% vs. -29%, all P<0.001) tended to be lower in AA vs. WA. Similarly, mean peak serum glucose levels compared to the NGT sub-cohort. Salsalate lowered total white blood cell were lower while peak insulin levels were higher at 30 and 60 mins in AA vs. counts (mean difference -0.7x103/µL, 95% CI: -1.0 to -0.4x103/µL, P<0.001) WA. In contrast, mean fasting and peak serum c-peptide levels at 60 and 90 and increased adiponectin (mean difference 1.8 µg/mL, 95% CI: 0.9 to 2.6 mins were signifi cantly lower in AA vs. WA. Mean AIRg, was higher in AA vs. µg/mL, P<0.001) compared to placebo, consistent with anti-infl ammatory WA (633±502 vs. 414±246, p=0.01). Although, Si (2.9±3.2 vs. 2.5±1.8 (mu/I) effects, but increased albuminurea (16.7 µg/mg, 95% CI: 6.4 to 27.1µg/mg x10-4/min-1, p=0.483) was not different, DI was signifi cantly higher in AA vs. creatinine, P<0.001). WA (1381 ±1126 vs. 901.9±477x10- 2xmin-1, p=0.01). In addition, mean Sg was Salsalate improves glycemia with sustained decreases in FBG, HbA , GSP 1c signifi cantly higher in AA vs. WA (2.5±1.2 vs. 1.9±0.72 x (min-1, p= 0.02). levels in obese patients and hence may decrease risk of T2D. Salsalate may Conclusions: We found that in overweight/obese prediabetic AA and WA inform new therapeutic approaches for T2D prevention, but renal safety may women with similar Si, the mean AIRg, Sg and DI were signifi cantly higher in limit clinical utility. AA. We conclude that the pathophysiologic mechanisms of prediabetes dif- Supported By: National Institutes of Health; National Heart, Lung, and Blood fer in the overweight/obese AA when compared to WA women. Institute Supported By: American Diabetes Association (1-11-CT-39 to K.O.) 2024-P Cardiometabolic Risk Reduction with Lifestyle Intervention: Pitt 2026-P Association of Androgen with Gender Differences in Serum Adipo- Retiree Study Initial Results cyte Fatty Acid Binding Protein Levels Induced by the Fat Content ELIZABETH M. VENDITTI, MARSHA D. MARCUS, RACHEL G. MILLER, VINCENT C. and Distribution ARENA, SUSAN L. GREENSPAN, BONNIE P. GILLIS, LAURA S. KINZEL, ELIZABETH XIANG HU, XIAOJING MA, XIAOPING PAN, YUQI LUO, YITING XU, QIN XIONG, CWENAR, MARY O. RACEK, KRISTIN L. SCHROEDER, Pittsburgh, PA YUQIAN BAO, WEIPING JIA, Shanghai, China Elders with obesity are at higher risk for hyperglycemia and other comor- Adipocyte fatty acid binding protein (A-FABP) is a transport protein in bidities. Evidence suggests 3-5% mean weight loss may improve cardiomet- mature adipocytes. Clinical investigations have indicated women have abolic risk but ≥ 7% is the more commonly cited goal. We studied the impact higher levels of A-FABP than men. In consideration of sex hormones and the of a 12-session behavioral group diet/activity intervention in 322 elders with fat content and distribution, the present study aims to identify the factors obesity and ≥ 1 additional cardiometabolic risk factor (mean age 71; mean related to gender differences in serum A-FABP levels. BMI 33; 23% male; 13% non-white). Month 0-4 changes were analyzed by Serum A-FABP levels were measured by the sandwich enzyme-linked baseline glycemic status (fasting fi nger stick) and by % weight loss cate- immunosorbent assay. An automatic bioelectrical impedance analyzer was gory. The sample was 69% (N=221) normoglycemic and 31% (N=101) glu- used to measure the fat mass and percentage of the total body, trunk, arms cose impaired (IFG 100-125 mg/dl). Groups showed signifi cant mean weight and legs. Obesity loss independent of baseline glycemia (≥6.5%; p <.0001) and had similar A total of 507 subjects were enrolled, including 194 men, 132 premeno- POSTERS benefi ts in waist circumference, blood pressure (BP), triglyceride, and total pausal women, and 181 postmenopausal women. Serum A-FABP levels cholesterol. Only the IFG group had signifi cant glucose reduction (-5.7±8.7; increased in the order from men, premenopausal women to postmenopausal

Integrated Physiology/ p <.0001). No LDL changes were found for either glycemic group and HDL women in either body mass index category (< 25.0 or 25.0 kg/m2) (all P < worsened (58% men, 49% women used statins). In the full sample (Table) as ≥ 0.05). Multiple stepwise regression analyses showed that after the adjust- % weight loss increased, signifi cant linear benefi ts were observed for most ment of the factors related to serum A-FABP levels, the trunk fat mass was cardiometabolic indices except HDL and LDL. Only participants with ≥7% an independent and positive factor of serum A-FABP levels (Standardized loss had signifi cant LDL and glucose improvements suggesting that larger β = 0.354, 0.565, and 0.406 for men, pre- and postmenopausal women, respec- weight losses may be needed for cardiometabolic benefi ts in a mixed risk tively; all P < 0.001). For men, total testosterone (Standardized = -0.183, P = group of elders. Long term follow-up data are needed to document the sus- β 0.002) was associated independently and inversely with serum A-FABP lev- tainability of the health benefi ts observed. els. For pre- and postmenopausal women, bioavailable testosterone (Stan- dardized β =0.194, P = 0.007) and total testosterone (Standardized β =0.126,

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A518 OBESITY—HUMANCATEGORY

P = 0.049) were independent and positive factors associated with serum results are relevant not only for understanding the physiology of ASCs in A-FABP levels, respectively. terms of cell-based therapies but also for their role as key regulators of the In conclusion, serum A-FABP levels increased in men, premenopausal immune response. women and postmenopausal women progressively. Androgen was identi- Supported By: Spanish Ministry of Economy and Competitiveness/Euro- fi ed as an independent and negative factor of serum A-FABP levels in men pean Regional Development Fund (PI14/00228, SAF2012-36186, PI15/00143, and an independent and positive factor of serum A-FABP levels in women. CB07708/0012, RYC2013-13186, CP10/00438) With the effects on the fat content, especially the trunk fat, androgen might contribute to the gender differences in serum A-FABP levels. 2029-P Supported By: National Basic Research Program of China (2013CB530606); Effect of Ursolic Acid on Metabolic Syndrome, Insulin Sensitivity, Shanghai Health and Family Planning Commission (2013ZYJB1001); Shanghai and Infl ammation Jiao Tong University (15ZH2010); National Natural Science Foundation of China MANUEL GONZÁLEZ-ORTIZ, ALEJANDRA M. RAMÍREZ-RODRÍGUEZ, NATALHIE (81100563); Key Project of Science and Technology of Shanghai (13XD1403000) ACUÑA-ORTEGA, ESPERANZA MARTÍNEZ-ABUNDIS, Guadalajara, Mexico Ursolic acid is a promising compound to treat the metabolic syndrome 2027-P (MS), because its positive effects in vitro and in vivo on insulin, glucose and Changes in the Metabolic Gene NRIP1 Expression in Response to lipid metabolisms. Weight Loss and Exercise The aim of this study was to evaluate the effect of ursolic acid administra- YANG DE MARINIS, JIANGMING SUN, ANETTE UNTERMANN, BENGTA WULFF tion on MS, insulin sensitivity and infl ammation. KAMPMANN, PRADEEP BOMPADA, MARTIN RIDDERSTRALE, Malmö, Sweden, A randomized, double blind, placebo-controlled clinical trial was per- Copenhagen, Denmark formed. Twenty-four adults with diagnosis of MS in accordance with the Nuclear receptor interacting protein 1 (NRIP1) is an important regulator International Diabetes Federation criteria, without drug treatment were of energy expenditure. Here we investigated changes in NRIP1 profi le in included. They were randomly assigned in two groups of 12 patients each to adipose tissue, serum and skeletal muscles in response to weight loss and receive orally 150 mg. of ursolic acid or placebo once a day during 90 days. In exercise. Fourteen obese adults were subjected to a 3-month weight loss the beginning and at the end of the study, waist circumference, body weight (WL) followed by 6-month weight maintenance (WM) intervention. Subcu- and blood pressure were measured, as well glucose and insulin concentra- taneous adipose tissue biopsies were obtained at baseline, after WL, and tions in an oral glucose tolerance test; the values were used to calculate after WM. We observed signifi cantly increased NRIP1 mRNA levels after insulin sensitivity (Matsuda Index). Triglycerides, total cholesterol, high-den- WL, which remained to be elevated after WM. Gas chromatography-mass sity lipoprotein cholesterol, interleukin-6 and C reactive protein concentra- spectrometry metabolomics analysis revealed positive association between tions were also assessed. The tolerability was investigated throughout the NRIP1 expression and HDL cholesterol and glycine levels; and negative asso- study. Data were analyzed with Wilcoxon and Mann Whitney U tests. An ciation with triglycerides, BMI, fat mass, and waist circumference. In the Ethic Committee approved the protocol and a written informed consent was serum of nonobese subjects, males (n=42) have 3-fold higher NRIP1 than obtained from all volunteers. the females (n=71), which is negatively associated with BMI, waist-hip ratio, After ursolic acid administration, the remission of the MS occurred in 50% acute myocardial infarction, age, etc. To investigate the changes in NRIP1 of patients treated with ursolic acid, while in the control group no patients expression in response to exercise, we analyzed skeletal muscle transcrip- had remission (p = 0.014). There were signifi cant differences in insulin sen- tome from database GSE59088. NRIP1 expression increased 80% in the sitivity (3.1 ± 1.1 vs. 4.2 ± 1.2, p = 0.003), body weight (75.7 ± 11.5 vs. 70.0 ± sedentary rest group and ~25% in the strength training group compared to 17.8 kg, p = 0.001), body mass index (29.9 ± 3.7 vs. 27.3 ± 6.4 kg/m2, p = 0.001), before sessions; while no changes in the endurance training group. Further- waist circumference (95.2 ± 9.4 vs. 86.6 ± 10.0 cm, p = 0.001) and fasting more, NRIP1 expression became sensitive to insulin stimulation during a eug- glucose (6.0 ± 0.5 vs. 4.7 ± 0.4 mmol/L, p = 0.002). The rest of evaluations lycemic clamp after sedentary living, whereas re-training led to decreased were not modifi ed. NRIP1 levels. These observations imply that the expression of NRIP1, a sup- In conclusion, the ursolic acid administration during 90 days leads to pressor of mitochondrial function, is elevated in states characterized by low remission of the MS, increasing insulin sensitivity and reducing body weight, energy demand, either due to lack of surplus energy (after weight loss), or BMI, waist circumference and fasting glucose. under non-oxidative conditions (resistance training), while during endurance training where the major energy comes from oxidative metabolism of car- 2030-P bohydrate and lipid, NRIP1 expression is suppressed to ensure high energy Body Mass Index (BMI), Waist Circumference (WC) and Waist output. NRIP1 may therefore serve as an important marker and target for Height Ratio (WHtR) Cutoff Values, and the Impact of Their Combi- various interventions. nations on Future Diabetes among Japanese Individuals Supported By: Swedish Research Council RISA IGARASHI, KAZUYA FUJIHARA, YORIKO HEIANZA, MASAHIRO ISHIZAWA, MEGUMI TSURUTA, MARIKO HATTA, SAKIKO YOSHIZAWA, CHIKA HIRIKAWA, 2028-P SATOSHI MATSUNAGA, OSAMU HANYU, SATORU KODAMA, KOJI SATO, KIMI- Obesity and Type 2 Diabetes Alters the Immune Properties of Human NORI KATO, HIROHITO SONE, Niigata, Japan Adipose-derived Stem Cells BMI and WC are associated with the development of diabetes. A BMI CAROLINA SERENA, NOELIA KEIRAN, VICTORIA CEPERUELO-MALLAFRE, MIR- cutoff value of 25.0 kg/m2 or WC cutoff value of 90/80 cm in men/women IAM EJARQUE, ROSA FRADERA, KELLY ROCHE, CATALINA NUÑEZ-ROA, JOAN have been established to identify obesity among Asian individuals. How- VENDRELL, SONIA FERNANDEZ-VELEDO, Tarragona, Spain ever, there is no consensus the optimal cutoff values for BMI and WC for Adipose tissue-derived stem cells (ASCs) are proposed as an alternative the development of diabetes. Moreover, the impact of WHtR on incident dia- stem cell source to bone marrow-derived cells for immune cell therapy. How- betes is unclear. We investigated the optimal cutoff value for BMI, WC and ever, microenvironmental factors may contribute to the functionality of the WHtR and their combined effects on the development of diabetes by sex. stem cell population in human adipose tissue (AT). We hypothesized that the Diagnosis of diabetes was indicated by fasting glucose ≥ 126 mg/dl, HbA1c fat depot in addition to the donor phenotype controls the immunomodulatory ≥6.5% or a self-reported history of clinician-diagnosed diabetes. During a

capacity of ASCs. Focusing on obesity and type 2 diabetes (T2D) as metabolic 4-y follow-up, 1343 individuals developed diabetes among 39254 individu- Obesity disorders which might affect the immune response of ASCs, we compared als. According to the ROC curve in our study, the optimal cutoff value for BMI POSTERS the infl ammatory response of ASCs isolated from subcutaneous and visceral or WC to predict diabetes was ≥24.0 kg/m2 for both sexes, or ≥87/82cm in

AT from age-matched donors (lean n=4; obese n=4 and T2D n=4). Obese and men/women, respectively. Furthermore, the optimal cutoff value for WHtR Integrated Physiology/ T2D-derived ASCs showed increased expression of infl ammatory markers, was ≥ 0.47 for both sexes. In men, WC was signifi cantly associated with dia- activation of the NLRP3 infl ammasome and higher migration, invasion and betes only among individuals with BMI ≥24.0 kg/m2. Compared to persons phagocytosis capacities than those derived from lean donors. Remarkably, with BMI ≥24.0 kg/m2 and WC <87 cm, those with BMI ≥24.0 kg/m2 and WC ASCs derived from obese and T2D subjects exhibited a reduction in typical ≥87 cm had a 2.78 (2.41, 3.21) times increased risk of diabetes. Conversely, immunosuppressive activities attributed to stem cells. Accordingly, obese WC ≥82 cm was associated with the risk of diabetes even if the BMI was and T2D-ASCs were less effective in suppressing T cell and B cell prolif- <24.0 kg/m2 in women. WHtR ≥0.47 was also associated with the risk of eration, activating the M2 macrophage phenotype as well as in increasing diabetes even if the BMI was <24.0 kg/m2 in both sexes. Persons with BMI TGF-beta1 secretion than lean-derived ASCs. Overall, these data indicate ≥24.0 kg/m2 and WHtR ≥ 0.47 had a 3.94 (3.15, 4.92) times increased risk of that the metabolic phenotype of the donor compromises the immunomodula- diabetes in men and, a 5.71 (3.67, 8.89) greater risk in women compared to tory properties of ASCs, particularly in cells derived from visceral AT. These those with BMI ≥24.0 kg/m2 and WHtR ≥0.47. Our fi ndings suggested that

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a lower BMI than the current criterion for obesity was associated with the 2033-P development of diabetes. We also noted that the WHtR cutoff value to pre- Sex-specifi c U-Shaped Associations between Birthweight and dict future diabetes could apply to both men and women. As predictors of Adult Adiposity the incidence of diabetes, the combined effect of BMI and WHtR was stron- VIVIAN C. LUFT, BRUCE B. DUNCAN, GABRIELE ROCKENBACH, NOEL T. MUEL- ger than that of BMI and WC. LER, MARIA INÊS SCHMIDT, Porto Alegre, Brazil, Santa Catarina, Brazil, Baltimore, MD 2031-P Recent evidence from high-income countries suggests that abnormal Genetic Variations Enriched in Pima Indians Near GCLM Are Asso- fetal growth has sex-specifi c metabolic consequences. We investigated ciated with BMI sex-specifi c associations of birthweight with adult body mass index (BMI), MICHAEL TRAURIG, PANKAJ KUMAR, PAOLO PIAGGI, SAYUKO KOBES, ROBERT waist circumference (WC) and waist-to-hip ratio (WHR) in the Brazilian Lon- L. HANSON, CLIFTON BOGARDUS, LESLIE J. BAIER, Phoenix, AZ gitudinal Study of Adult Health (ELSA-Brasil), a multicenter cohort study of Pima Indians from the Gila River Indian Community (GRIC) in Arizona have a 15,105 adults 35 to 74 years of age who were born before the nutritional high rate of obesity. To identify loci with an enriched population-level effect transition in Brazil. Socio-demographics and birthweight were obtained by in this community, we used whole-genome sequence data from 267 full-her- interview. Anthropometry was directly ascertained. After adjusting through itage Pima Indians to identify variants with allele frequencies signifi cantly ANCOVA for socio-demographics, history of maternal diabetes and history different in Pimas as compared to the HapMap populations. One of the vari- of paternal diabetes, we found higher mean BMI, WC and WHR for par- ants, rs7520019 (A/G), maps upstream of GCLM and has an ancestral allele ticipants with high (≥ 4 kg) vs. normal birthweight (2.5 kg-4 kg). For those (G) frequency of 92.1% in Pimas and 47.3%, 61.3%, and 50.0% in the CEU, with low (< 2.5 kg) vs. normal birthweight, results were heterogeneous, with CHB, and MEX populations, respectively. GCLM codes for the glutamate- women with maternal history of diabetes having greater overall and central cysteine ligase, modifi er subunit important for the synthesis of glutathi- obesity (P interaction < 0.05). (Figure: red line, maternal diabetes; green line, one, an antioxidant involved in infl ammation, insulin resistance, and energy no maternal diabetes; bars=95% CIs). In conclusion, U-shaped associations metabolism. GCLM-/- mice are protected from HFD-induced obesity and have are present among women, particularly among those with a maternal history increased basal metabolic rate due to increased hepatic mitochondrial respi- of diabetes. Our fi ndings support the hypothesis that an adverse intrauterine ration. Genotyping of rs7520019 in 6624 subjects from the GRIC who partici- environment has sex-specifi c effects on adiposity that last into mid-to-late pated in a longitudinal study and who had data on maximum BMI measured adulthood. when the subjects were nondiabetic and ≥15 years of age shows that the G Figure. allele is associated with higher BMI (β = 2.4% per copy; P = 7.0 × 10-4 after genomic control, with adjustment for age, sex, birth-year, and admixture). The BMI association replicated in a sample of 3080 urban American Indians who are predominately from Phoenix and have self-reported data on BMI at maximum weight (β = 2.3%; P = 1.1 × 10-3); combining both datasets showed the strongest association with BMI (N = 9704; β = 2.3%; P = 2.5 × 10-6). A subset of Pima Indians also participated in inpatient metabolic studies when they were nondiabetic. Among these subjects the G allele was also associ- ated with a lower resting metabolic rate (N = 502; β = -0.12 kcal/day; P = 0.05 adjusted for age, sex, fat mass, fat free mass, and admixture). In vitro func- tional studies are being carried out to determine whether rs7520019 or other variants in high linkage disequilibrium are affecting GCLM expression. Supported By: National Institutes of Health; National Institute of Diabetes and Digestive and Kidney Diseases

2032-P With Increasing BMI, Liver Enzymes, and Frequency of Ultrasound Abnormalities Describe a U-Shaped Curve ALICE FANIN, ALESSIA GIORGINI, ALBERTO BENETTI, LAURA FOLINI, AHMED S. ZAKARIA, MASSIMO ZUIN, ANTONIO E. PONTIROLI, Milan, Italy Liver enzymes are commonly used as markers of liver steatosis. Eleva- tion of liver enzymes occurs in obesity, hyperlipidemia and diabetes mellitus, but also in lean subjects, especially in anorexia nervosa; however, in the latter, prevalence of elevated enzymes widely ranges among studies, and its causes remain under-evaluated. Aim of this study was to analyze ami- notransferases (ALT, AST), liver ultrasound (US), liver magnetic resonance (MRI), and metabolic data [insulin, blood glucose (and HOMA-IR index), total, HDL, LDL cholesterol, triglycerides] of a population of 813 Italian sub- jects, including anorexia patients (Class 1, BMI < 19 kg/m2, n = 107), normal- weight people (Class 2, <25 kg/m2, n = 243), overweight (Class 3, < 30 kg/ m2, n = 46), obese (Class 4, < 35 kg/m2, n = 67), and severely obese patients (Class 5, > 35 kg/m 2, n = 350) and to correlate these data with body mass index (BMI) classes. The relationship between BMI and ALT/AST, and between BMI and US/MRI data is visually represented by a U-shaped curve: obese (Class Supported By: Brazilian Ministry of Health

Obesity 4 and Class 5) and anorexia patients (Class 1) have higher percentages of

POSTERS abnormal fi ndings, compared with normal weight people (p 0.04 to 0.001 vs. 2034-P Class 2). Variables in study also showed signifi cant correlations with other Intranasal Insulin Enhances Brain Connectivity in Peripheral Insu-

Integrated Physiology/ metabolic and nutritional indices [ALT vs. total cholesterol (p=0.001), HDL lin-Sensitive Individuals cholesterol (p=0.002), and triglycerides (p=0.0001); AST vs. LDL cholesterol STEPHANIE KULLMANN, MARTIN HENI, RALF VEIT, KLAUS SCHEFFLER, HANS- (p=0.04), HDL cholesterol (p=0.01), and triglycerides (p=0.001); US vs. HDL ULRICH HAERING, ANDREAS FRITSCHE, HUBERT PREISSL, Tübingen, Germany cholesterol (p=0.0001), and triglycerides (p=0.0001); MRI vs. LDL cholesterol Brain insulin resistance (BIR) is a possible link between metabolic and cog- (p=0.005), HDL cholesterol (p=0.005), and triglycerides (p=0.006)]. In con- nitive disorders. Even in young adults, obesity-associated BIR was recently trast, insulin and HOMA-IR index are lower in lean subjects and progres- identifi ed in specifi c brain regions. However, BIR in one region may lead to sively higher in normal weight and overweight, highest in morbidly obese alterations in functional connectivity in brain networks. Studies in type 2 patients. In conclusion, both liver enzymes levels and US/MRI analyses are diabetes and dementia suggest that intranasal insulin can enhance func- abnormal in anorexia and in obese patients; these fi ndings can be attribut- tional connectivity (FC) within the default-mode-network, which includes able to insulin resistance in obese, not in underweight subjects. brain regions especially prone to dementia. Whether brain insulin action can infl uence FC in healthy lean and obese individuals is still illusive.

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We performed functional magnetic resonance imaging in 25 lean and 23 T2DM) obese subjects undergoing bariatric surgery (n=8 per group). ECM- obese participants (body mass index range 19.5-46.5 kg/m2; age range 19-37 receptor interaction, focal adhesion and cell adhesion molecules were the years), on two separate days, to measure resting-state FC of the brain before 3 most highly represented biological pathways in OM vs. SC comparisons and 30 min after intranasal insulin or placebo application. Additionally, we derived from both T2DM and non-T2DM subjects. Integrin receptors such as assessed peripheral insulin sensitivity during a 75g oral glucose tolerance ITGB8 and ITGB1 that are crucial for cell attachment to the ECM, regulation of test to evaluate possible interactions between brain and peripheral insulin kinase activity and cell signalling, were two genes that markedly overlapped sensitivity. Based on recent fi ndings, we evaluated FC in response to intra- in all three pathways. When comparing T2DM vs. non-T2DM in SC and OM nasal insulin. depots respectively, ECM components were positively enriched in T2DM, We identifi ed a signifi cant interaction between brain insulin action and but not in non-T2DM subjects. LRP2, KLK7, WT1, WT1-AS and LRRN4 were peripheral insulin sensitivity. With higher peripheral insulin sensitivity, we among top genes differentially upregulated whereas SIM1, HOXC10, PAX3, observed a signifi cant increase in FC between the hypothalamus and pre- IRX5 and MYEOV were genes signifi cantly downregulated in OM AT from both frontal regions of the default-mode-network after intranasal insulin (p<0.05, T2DM and non-T2DM subjects, indicating its importance in obesity. Our fi nd- corrected for multiple comparisons). No such relationship was observed ings show that excessive ECM components and AT fi brosis appeared to play after placebo administration. a vital role in the differences observed between depots and T2DM status, In conclusion, we were able to show that brain insulin action induced by potentially contributing to AT dysfunction and insulin resistance. intranasal insulin enhances functional connections in the CNS. This mecha- Supported By: National Medical Research Council of Singapore nism possibly contributes to the benefi cial effects of brain insulin action on metabolism and cognition. However, whole-body insulin resistant persons 2037-P fail to show this effect, suggesting elevated risk for not only type 2 diabetes Pancreatic Fat Accumulation Is Driven by Environmental Rather but also cognitive impairments. than Genetic Factors: A Classic Twin Study GYÖRGY JERMENDY, MÁRTON KOLOSSVÁRY, ALEXISZ PANAJOTU, SÁRA PAPP, 2035-P ÁDÁM L. JERMENDY, ÁDÁM D. TÁRNOKI, DÁVID L. TÁRNOKI, SZILÁRD VÖRÖS, High Risk Obesogenic Environments Accentuate Genetic Suscepti- BÉLA MERKELY, PÁL MAUROVICH-HORVAT, Budapest, Hungary, Atlanta, GA bility to Obesity Several studies showed that intrapancreatic fat accumulation is associated JESS TYRRELL, HANIEH YAGHOOTKAR, ROBIN BEAUMONT, SAMUEL E. JONES, with beta-cell dysfunction. Pancreatic fat can be quantitatively assessed by RYAN AMES, MARCUS A. TUKE, KATHERINE S. RUTH, ZOLTAN KUTALIK, RACHEL non-contrast-enhanced computed tomography (CT). The role of genetic and M. FREATHY, ANNA MURRAY, ANDREW R. WOOD, MICHAEL N. WEEDON, TIM- environmental factors in pancreatic lipid accumulation is unclear. Therefore, OTHY M. FRAYLING, Exeter, United Kingdom, Lausanne, Switzerland we sought to evaluate the contribution of genetics and the environment on Susceptibility to obesity and type 2 diabetes in today’s environment has pancreatic lipid content within a cohort of healthy adult twin pairs. a strong genetic component. However, little is known about how genetic We investigated 77 twin pairs (154 twin subjects; 98 women, 56 men) variation interacts with the modern environment to predispose some indi- with a 256-slice CT-scanner, of whom 47 were monozygotic (MZ) and 30 viduals to obesity and type 2 diabetes whilst others remain slim. Previous were dizygotic (DZ) same gender pairs. Using non-enhanced CT images we gene-obesogenic environment studies have been limited by the need to measured the average value of pancreatic attenuation (Hounsfi eld [HU] unit) perform meta-analyses of many heterogeneous studies. We aimed to use in three regions of interest (head, body and tail). Blood samples were col- 120,000 individuals from the UK Biobank to test the hypothesis that high risk lected before the CT scan. Intra-pair correlations were calculated and struc- obesogenic environments accentuate genetic susceptibility to obesity. We tural equation model was used for evaluating additive genetic (A), dominant selected 10 self-reported measures of the obesogenic environment including genetic (D) and unique environmental (E) components. measures of TV watching, westernised diet, physical activity and rural vs. Main clinical and laboratory fi ndings of the twins were: age 56.1±9.4 years, urban home. We generated a 69-variant genetic risk score (GRS) for obesity BMI 27.7±5.3 kg/m2, fasting glucose 97.5±25.7 mg/dL, HbA1c 5.5±1.0%, and tested its association with BMI in high and low exposure groups and C-peptide 2.3±1.4 ng/mL, LDL-cholesterol 134±39 mg/dL, HDL-cholesterol tested for interactions. Higher BMI was associated with 10 self-reported 62±14 mg/dL, triglycerides 132±73 mg/dL (mean±SD). Average pancreatic measures of the obesogenic environment (all P<0.001). We found evidence attenuation was 47.2±11.3 HU in MZ and 47.6±11.8 HU in DZ twins. The intra- -5 of gene-environment interactions with TV-watching (Pinteraction=7x10 ), phys- pair correlation between HU values were stronger in MZ as compared to DZ -6 ical (Pinteraction=5x10 ), and sedentary activity (Pinteraction=0.03). In each case, twins (rMZ=0.498, p<0.001; rDZ=0.080, p=0.674). Using the structural equa- a higher risk environment accentuated the effect of the GRS on BMI. For tion model, a predominant environmental infl uence (E: 59%) and a moderate example, within individuals watching ≥4 hours TV per day, carrying 10 addi- additive genetic dependence (A: 41%) was found. Dominant genetic infl u- tional BMI-raising alleles was associated with 4.0kg extra weight in some- ence was not identifi ed (D: 0%). one 1.73m tall. In contrast, within individuals watching <4 hours TV per day, We found a moderate genetic and a much stronger environmental carrying 10 additional BMI-raising alleles was associated with 3.1kg extra dependence of pancreatic lipid accumulation in our twin cohort indicating weight. Our fi ndings suggest that many aspects of the obesogenic environ- that environmental factors and lifestyle characteristics are predominantly ment accentuate the risk of obesity in genetically susceptible individuals. involved in the development of fat accumulation in the pancreas. Supported By: Diabetes Research and Wellness Foundation (to J.T.); Medical Supported By: European Foundation for the Study of Diabetes Research Council (to S.E.J.); Wellcome Trust UK (to R.M.F., M.A.T., M.N.W., A.M., R.B., R.A.); European Research Council (A.R.W., T.M.F., H.Y.) 2038-P Weight Trajectories during the Transition to Adulthood in the U.S., 2036-P 1994-2008 Transcriptomic Analysis Reveals a Role for Extracellular Matrix SOLVEIG ARGESEANU CUNNINGHAM, LINDA S. GEISS, GLORIA L. BECKLES, and Fibrosis in Maintaining Human Adipose Tissue Function GIUSEPPINA IMPERATORE, Atlanta, GA CHIA ROU YEO, MADHUR AGRAWAL, SHAWN HOON, VANNA CHHAY, JONA- Given the links between obesity and diabetes onset, we examine weight THAN CALEB YOU XING QUEK, ASIM SHABBIR, MUHAMMAD SHABEER, E. SHY- trajectories during the developmentally critical transition to adulthood. We

ONG TAI, CHIN MENG KHOO, JIMMY BOK YAN SO, DAVIDE LOMANTO, SUE- used data from the National Longitudinal Study of Adolescent Health, a U.S. Obesity

ANNE E.E. SHIOW TOH, Singapore, Singapore nationally representative prospective cohort of individuals recruited in high POSTERS Adipose tissue, which is closely bounded by extracellular matrix (ECM) school in 1994 with 3 follow-up visits through 2008 (analytic sample=14,135).

components, is now recognized as an important endocrine organ in support- Weight and height were measured at each visit. We defi ned overweight as Integrated Physiology/ ing metabolic health. While ECM is crucial in maintaining structural integrity 85th percentile ɖ BMI [Body Mass Index] z-score <95th at ages <19 y and of adipose tissue (AT), excessive ECM components can lead to reduction in AT 25ɖ BMI<30 at >18 y) and obesity (z-score ≥95th and BMI ≥30, respectively). expandability, development of AT fi brosis and fi nally metabolically dysfunc- Between ages 16 and 26 y, youths gained on average 0.46 BMI point annu- tional AT. We aimed to screen transcriptomic differences in AT derived from ally. Of those who were normal weight in adolescence (75%), less than half different depots and disease status via RNA sequencing, and subsequently remained normal weight in adulthood (mean age 26 y), 34% became over- identify the most relevant pathways and ontologies using impact analysis weight and 22% became obese (see Table). Of overweight adolescents (15%), (ipathway) and gene set enrichment analysis (GSEA). Genome-wide transcrip- 68% became obese and only 8% achieved normal weight. Among obese tomic analysis enabled us to have an unbiased, hypothesis-free approach to adolescents (10%), 89% stayed obese and only 2% were normal weight in our data set. Human adipose tissue were obtained from omental (OM) and adulthood. Very few youths who were overweight or obese in adolescence subcutaneous (SC) adipose depot from diabetic (T2DM) and nondiabetic (non- achieved normal weight in young adulthood, and the majority of those who

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A521 OBESITY—HUMANCATEGORY

were normal weight in adolescence no longer were so in adulthood. Efforts to 2040-P encourage youth to reach and maintain a healthy weight are important. Human Subcutaneous (SC) White Adipose Tissue (WAT) Beiging Table. Weight Status in Adulthood According to Weight Status in Adolescence. after Cold Exposure: Role of Mast Cells, Histamine, and IL-4 Weight Status in Adolescence PHILIP A. KERN, BRIAN FINLIN, BEIBEI ZHU, AMY L. CONFIDES, BRIANNA D. HARFMANN, ESTHER E. DUPONT-VERSTEEGDEN, Lexington, KY Normal weight 95% CI Overweight 95% CI Obese 95% CI Mice adapt to the cold through thermogenesis in both brown and beige (74.8%) (15.1%) (10.1%) adipose tissue mediated through the PKA-PGC1α-UCP1 pathway. The beig- Weight Status in Adulthood ing of human WAT is not well studied. In recent studies, a single 30 min Normal weight 43.68% 42.01-45.35 7.59% 5.88-9.30 2.30% 1.20-3.41 cold exposure increased PGC1α and UCP1 mRNA in SC WAT of control sub- Overweight 34.42% 33.09-35.74 23.01% 20.50-25.53 8.71% 6.61-10.81 jects. To examine UCP1 protein levels, protein abundance (IHC) for UCP1 was examined before and after 10 days of 30 min cold exposure/day (Figure). Obese 21.90% 20.64-23.17 69.39 66.51-72.28 88.99% 86.46-91.52 There was an increase in beiging evidenced by a signifi cant increase in Note: Estimates are survey-adjusted. UCP1. We have studied immune cells that control SC WAT beiging; changes in mast cell markers and IL-4 correlate with UCP1 expression. To examine 2039-P the interactions between mast cells and adipocytes, TIB64 mast cells were Regional Body Composition Measured by Dual-Energy X-Ray Ab- cocultured with 3T3-L1 adipocytes. When mast cells were exposed to cold sorptiometry (DXA) Relates to Insulin Resistance in a Sedentary they secreted higher levels of IL4 and Metrnl as well as histamine. The cold- Overweight and Obese Population stimulated mast cell conditioned medium increased UCP1 in adipocytes, and ANNE E. BANTLE, TYLER A. BOSCH, LISA S. CHOW, Minneapolis, MN this was blocked by a degranulation inhibitor. Some of this increase in UCP1 DXA is a common tool for assessing body composition, with new capability was histamine mediated, and occurred through PGC1α, and other increases to assess regional fat distribution. Using a cross-sectional design we evalu- in UCP1 was IL4 mediated and independent of PGC1α. ated the correlation between DXA regional body fat composition and insulin The beiging of human WAT may be important in energy regulation and resistance in overweight/obese sedentary adults. DXA measures included involves interactions with mast cells and PGC1α-independent mechanisms, fat free mass, visceral fat mass, visceral fat percent of total fat, android perhaps involving histamine, and IL4, resulting in increased UCP1 abundance. fat mass, and gynoid fat mass. Two indices of insulin resistance, HOMA-IR Figure. and Matsuda Index, were used. Linear correlation was tested using Pear- son’s correlation coeffi cient. Participants (n=68: 50 women/18 men) had mean age 28.8 ± 6.6 years and mean BMI 33.3 ± 7.1 kg/m2. Participants were categorized as insulin sensitive (mean ± SD: HOMA-IR 1.1 ± 0.4, Matsuda Index 8.3 ± 4.8) or insulin resistant (HOMA-IR 2.7 ± 2.0, Matsuda Index 3.6 ± 2.1). Of the DXA based body composition measures, visceral fat mass and visceral fat percent most consistently had a signifi cant negative correlation (-0.30 to -0.48, P<0.05) with insulin resistance by Matsuda Index; this asso- ciation was strongest in the insulin resistant group (R=-0.48, p<0.02). We conclude that DXA based measurement of visceral fat mass and percent of total fat could be used to predict insulin resistance by Matsuda Index in sedentary overweight/obese adults, reinforcing the expanding utility of DXA in clinical and research settings. Table. Correlation of Body Composition Measurements with HOMA-IR and Matsuda Index.

Supported By: National Institutes of Health (DK107646)

2041-P Changes in Skeletal Muscle Mitochondrial Function after Bariatric Surgery Depend on Surgery Type and Dissociate from Improvement of Insulin Sensitivity SOFIYA GANCHEVA, CHRYSI KOLIAKI, TOMAS JELENIK, JULIA SZENDROEDI, PETER NOWOTNY, KIRTI KAUL, MATTHIAS SCHLENSAK, MICHAEL RODEN, Düs- seldorf, Germany It has been suggested that skeletal muscle mitochondrial function improves after bariatric surgery. However, the time course of changes in mitochondrial respiratory capacity and insulin sensitivity as well as the impact of type of surgery remain unclear. Here, we quantifi ed skeletal muscle mitochondrial respiration using high resolution respirometry and peripheral insulin sensitivity (M-value) using hyperinsulinemic-euglycemic

Obesity clamps in obese patients before, two weeks, three, six and twelve months POSTERS after gastric sleeve (GS; n=8 age 41±12 yrs, BMI 52.6±10 kg/m2) or gastric bypass (GB; n=8, age 40±10 yrs, BMI 46.6±4.6 kg/m2) surgery. Normal weight

Integrated Physiology/ humans studied at baseline served as controls (CON; n=7, age 38±4 yrs, BMI 24.9±1.1 kg/m2). M-values were higher in CON than in GS and GB groups at baseline (GS: 2.8±0.7 vs. GB: 2.6±1.7 vs. CON: 8.7±3.2 mg*kg-1*min-1, ANOVA p=0.001) and improved faster in GB patients by doubling already at 6 months and further increasing to 158% at 12 months, whereas GS group improved by 121% at 12 months only. Fasting free fatty acids (FFA) transiently increased 2 weeks after GB surgery and consequently returned to baseline at 3 months (ANOVA p=0.003). Maximal uncoupled (state u) respiration linked to tricar- boxylic acid cycle decreased by 37% at 3 months in GB patients and only Supported By: National Institutes of Health (1R01DK098203-01) returned to baseline at 12 months (ANOVA p=0.001), independent of changes in M-value and FFA. In GS group state u respiration decreased by 44% at 6

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A522 ISLET BIOLOGY—APOPTOSISCATEGORY months and remained reduced at 12 months (ANOVA p=0.0007). We con- and human islets. This suggests modulating Cx36, or Ca2+ through other clude that changes in muscle mitochondrial function following GB and GS means, may reduce the decline in β-cell mass and function in type 1 diabe- surgery depend on surgery type and dissociate from changes in insulin sen- tes, thereby delaying or preventing the onset of disease. sitivity and lipid availability. Supported By: National Institutes of Health (R00 DK085145, R01 DK102950, R01 DK106412, F32 DK102276-01A1); JDRF (5-CDA-2014-198-A-N)

ISLET BIOLOGY—APOPTOSIS & 2044-P Single Insulin Gene Mutation in Neonatal Diabetes Provokes Endo- plasmic Reticulum Stress and Demise of Insulin-Secreting β-Cells Moderated Poster Discussion: Beta-Cell Life and Death GUO DONG LI, SHU MENG, CHUI SUN YAP, CHAUR LEE TEO, YONG MONG BEE, (Posters: 2042-P to 2047-P), see page 18. Singapore, Singapore Neonatal diabetes mellitus is characterized by the onset in a neonate or & 2042-P infant below 6 months of age and caused usually by mutations in one of the Beta-Cell Dedifferentiation Plays a Central Role in Beta-Cell Fail- genes essential for pancreatic islet β-cell development and function, such as ure in a Model of Wolfram Syndrome insulin and KATP channel. We identifi ed a novel heterozygous missense muta- KIKUKO SHIINOKI, KATSUYA TANABE, MASAYUKI HATANAKA, MANABU KONDO, tion of insulin gene, C109Y, in a family with permanent neonatal diabetes. YUKIO TANIZAWA, Ube, Japan This study aimed to understand the molecular mechanism underlying the β-cell failure is central in the pathogenesis of both type 1 and type 2 dia- pathogenesis due to this mutation. An in vitro transformed rat insulinoma betes. It results from reduced β-cell mass and function. Wolfram syndrome cell line (INS-1) capable of normally synthesizing and secreting insulin alike is classifi ed as one of the monogenic forms of diabetes. Its causative gene, adult islet β-cells was transfected by vectors encoding wild-type (WT) or WFS1, is involved in the regulation of β-cell number and function through mutant C109Y type (MT) human proinsulin, followed by examinations of insu- maintaining endoplasmic reticulum homeostasis. WFS1 is also one of the lin expression, biosynthesis, subcellular localization and cell survival. Immu- type 2 diabetes susceptibility genes. We investigated the contribution of nofl uorescence of human proinsulin C-peptide showed a lower intensity in these two processes to β-cell failure using mice lacking Wfs1. Wfs1 ablation INS-1 cells expressing MT than those expressing WT after 24-h transfection. causes hyperglycemia with reduced β-cell mass with aging. Wfs1-defi cient The number of cells expressing MT proinsulin was signifi cantly less (-30%), β-cells suffered from oxidative and endoplasmic stresses, and reverted to indicating the loss of living cells. RT-PCR analysis showed a decrease of endocrine progenitor-like cells expressing neurogenin3, when the mice were human proinsulin transcript, and immunoblotting also exhibited dramatic still normoglycemic. Also, they exhibited reduced MafA expression and reduction of human C-peptide in MT-expressing cells. The subcellular local- insulin degranulation. However, this phenomenon could not be seen in the ization of biosynthesized WT and MT human proinsulin was examined by mice under nursing, indicating Wfs1 defi cient β-cells become de-differen- double immunostaining. Cells expressing WT proinsulin displayed a punctate tiated after weaning. Lineage-tracing experiments demonstrated that loss pattern of human C-peptide at cell periphery, suggesting its predominant of β-cells was mainly due to β-cell de-differentiation and that a subset of destination in the secretary granules. By contrast, cells expressing mutant β-cells adopted α cell fate. β-cell de-differentiation was more apparent proinsulin displayed scarce secretary granules and that C-peptide immunos- in Wfs1-defi cient mice carrying Ay/a (Wfs1-/-;Ay/a) in which β-cell loss was taining was largely co-localized with ER. Phosphorylated elF2α, an ER stress accelerated. We next asked whether β-cell de-differentiation was revers- marker, was enhanced in MT-expressing cells. Our data demonstrated that ible by preventing hyperglycemia, using SGLT2 inhibitor. Wfs1-/-;Ay/a mice the C109Y mutation caused misfolding and retention of the misfolded pro- given SGLT2i exhibited preservation of insulin-positive β-cell mass without insulin in ER. This would induce ER stress and islet β-cell demise, leading to increase in α cell number. However, even under this condition, unusual neu- development of neonatal diabetes. rogenin3 expression indistinguishably remained in β-cells, and at the same time, reduced MafA expression was not reversed. Moreover, insulin secre- & 2045-P tion in response to glucose challenging was not improved in the mice treated Noncoding RNA Expression Related to Human β Cell ER Stress and with SGLT2i. Taken together, β-cell de-differentiation is central of the patho- Death in Humanized Obese and Diabetic Mice genesis of diabetes in Wolfram syndrome, providing evidence that it may be REGAN ROAT, SHALINI JAIN, JENICA CHRISTOPHERSON, COLETTE FREE, MUNIR involved in human diabetes. HOSSAIN, ZHIGUANG GUO, Sioux Falls, SD We established a humanized obese and diabetic mouse model by trans- & 2043-P planting human islets into immunodefi cient, diabetic and obese db/db mice, Connexin36 Gap Junction Coupling Protects against Cytokine- and investigated human islet non-coding RNA expression in the context of induced Beta-Cell Death and Dysfunction with Ca2+ Dependence increased metabolic demand, ER stress, and β-cell death. Immunodefi cient NIKKI L. FARNSWORTH, AUDREY HEINTZ, RICHARD K. BENNINGER, Aurora, CO obese db/db-Rag-1 mice had increased body weight (58 vs. 31 g; p<0.05), Pro-infl ammatory cytokines mediate the decline in β-cell mass and islet fasting plasma cholesterol (144 vs. 77 mg/dL; p<0.05), and non-fasting function in type 1 diabetes. Connexin36 (Cx36) gap junctions regulate β-cell plasma insulin (91.3 vs. 0.7 µg/L; p<0.05) compared to the lean control mice. Ca2+ and insulin secretion. Cx36 gap junctions can also protect against β-cell Human islets were transplanted under the left kidney capsule of obese and apoptosis, and are decreased by pro-infl ammatory cytokines, disrupting Ca2+ STZ-induced diabetic lean controls. Blood glucose before transplantation signaling. We aim to understand how Cx36 gap junctions protect against and at tissue collection in obese mice was 354±20mg/dL and 209±45mg/ cytokine-induced apoptosis and disruption to insulin secretion, and the role dL, and in lean mice was 506±24mg/dL and 83±9mg/dL. OGTT showed that intracellular Ca2+ plays in this protection. blood glucose level of the obese mice was signifi cantly higher than that of Cell death and insulin secretion were measured in isolated islets from the lean mice at all time points (p<0.05) and human C-peptide at 15 minutes control (Cx36+/+); Cx36 knockout (Cx36-/-); Cx36 over-expressing (RIP-Cx36) was 1.39±0.47µg/L in obese mice and 1.15±0.18µg/L in lean mice. qRT-PCR mice, and human islets. Islets were treated for 24 h with a cocktail of cytok- of the human islet graft showed an increase in ER stress markers BIP, CHOP, ines (10ng/mL TNF-α, 5ng/mL IL-1β, 100ng/mL IFN-γ) alone or in combination and XBP-1 in the obese mice compared to the lean mice (p<0.05). Caspase with either 250µM Diazoxide, 200µM TEA, 6µM Ochratoxin A, 1µM Thapsi- 3 expression and TUNEL+ β-cells were increased in the human islet grafts gargin, or 50µM Modafi nil. of obese mice compared to lean mice (p<0.05). miRNAs (miR-146a-5p, miR- Cytokine-induced cell death was signifi cantly increased in Cx36-/- islets 148a-3p, miR-15b-5p, miR-199a-3p, miR-200a-3p, miR-200b-3p, miR-24-3p, and signifi cantly reduced in RIP-Cx36 islets compared to Cx36+/+ controls. miR-34a-5p, miR-375, and lncRNA (POLG2-1:1) were signifi cantly decreased Similarly, cytokine-induced disruption to insulin secretion was increased in and lncRNAs (CTRB1-1:1, SRPK1-1:1, GTF3C5-1:1, CTRB1-1:5, BCAR1-2:1, and Cx36-/- islets and reduced in RIP-Cx36 islets compared to Cx36+/+ controls. CTRB1-1:4) were signifi cantly increased in the grafts of obese mice compared In cytokine-treated human islets, increasing Cx36 function with Modafi nil to lean mice (p<0.05). Our results indicate that human β-cell ER stress and signifi cantly reduced cell death. Furthermore, natural variation in Cx36 cou- apoptosis are increased in the setting of obesity and diabetes and expres- pling correlated (ρ=0.78) with insulin secretion following cytokine treatment. sion of some non-coding RNAs is associated with the changes. Our model 2+ Decreasing intracellular Ca with the KATP activator Diazoxide or SERCA can be used to study the molecular mechanisms of ER stress and apoptosis POSTERS activator Ochratoxin A reduced cytokine-induced cell death in Cx36-/- islets in human β-cells in the setting of obesity, diabetes, and insulin resistance Islet Biology/

to similar levels as in Cx36+/+ islets. and evaluating the effi cacy of drug therapies. Insulin Secretion In conclusion, Cx36 gap junctions protect against cytokine-induced β-cell death and islet dysfunction through Ca2+-dependent mechanisms in mouse

ADA-Supported Research & Moderated Poster Discussion For author disclosure information, see page A696.

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