INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

1719-P INTEGRATED PHYSIOLOGY—INSULIN SECRETION Glucose-Induced Extracellular Matrix Production is Negatively IN VIVO Regulated by Extracellular Signal Regulated Kinase 5 (ERK5) YUEXIU WU, BIAO FENG, SHALI CHEN, SUBRATA CHAKRABARTI, London, ON, Canada Increased extracellular matrix protein production is a characteristic fea- Guided Audio Tour: Genetic and Human Metabolic Predictors of Diabetes ture of diabetic complications. Fibronectin (FN), a key extracellular matrix (Posters 1721-P to 1728-P), see page 17. protein is known to be upregulated in diabetes. ERK5 plays a critical role in cardiovascular development and in maintaining endothelial cell integrity. The aim of the study is to investigate the regulation of ERK5 signaling on & 1721-P glucose-induced FN overproduction.Human microvascular endothelial cells Mexican American Subjects With Impaired Fasting Glucose (IFG) were used. FN mRNA and protein levels were measured using ELISA and are Characterized by Hepatic and Adipocyte Insulin Resistance and real-time PCR respectively. Transforming growth factor beta 1(TGF`1) ex- Beta Cell Dysfunction pression and Smad2 proteins were investigated. Constitutively active MEK5 RUTH ARYA, ZANDRA PEREZ-CADENA, ANDREA HANSIS-DIARTE, DEIDRE WIN- (CAMEK5) adenovirus was transducted to upregulate ERK5 signaling. ERK5 NIER, LAUREN CORTEZ, RALPH DEFRONZO, DEVJIT TRIPATHY, CHRISTOPHER siRNA and dominant negative MEK5 (DNMEK5) transfections were used JENKINSON, San Antonio, TX to downregulate ERK5.Glucose caused a dose dependent increase in FN The aim of the current study was to characterize the metabolic defects production. Such increase was associated with upregulation of TGF`1 and associated with prediabetic states IFG and impaired glucose tolerance (IGT). Smad2 phosphorylation. ERK5 expression peaked at 24 hrs. Signifi cant de- We compared adipocyte, hepatic and whole body insulin resistance and beta crease of basal and glucose induced increased FN mRNA and protein levels cell function in subjects with normal glucose tolerance NGT n=36 age 46±2 were observed after CAMEK5 transduction. In contrast, ERK5 siRNA and BMI 30±1, IFG n=30 age 47±2 BMI 33±1, IGT n=20 age 49±3 BMI 34±1, IFG/ DNMEK5 treatment led to an increase of FN synthesis. Moreover, West- IGT n=36 age 49±3 BMI 34±1, T2D n=81 age 58±1 BMI 35±1). Indices of in- ern blot analysis showed that phosphorylated Smad2 was suppressed by sulin secretion and insulin sensitivity were derived from plasma glucose, CAMEK5 transduction with or without glucose treatment. On the other hand insulin and FFA concentrations during the OGTT and from hyperinsulinemic siERK5 transfection enhanced TGF`1mRNA expression. Furthermore, we in- euglycemic clamp (80/mU/m2/min) with indirect calorimetry and tritiated vestigated neurotrophins, eg. nerve growth factor (NGF) and brain-derived glucose infusion (non-diabetics, n=54) to measure hepatic glucose produc- neurotrophic factor (BNDF). Both are known upstream regulators of ERK5. tion. Adipocyte IR (AR) was the product of fasting FFA and fasting insulin. IFG The alterations of neurotrophins paralleled that of ERK5. Exogenous NGF subjects had similar whole body glucose uptake but higher hepatic insulin re- supplementation resulted in elevated phosphorylated and total ERK5 with or sistance (BGP x Fasting insulin) vs NGT (21.2±4 vs 7.6±1 mg/kg/min, p<0.05). without glucose treatment.Taken together, our experiments demonstrated a Whole body glucose disposal (skeletal muscle) was lower in subjects with novel mechanism of FN regulation under high glucose conditions. Decreased IFG/IGT primarily due to reduced non-oxidative glucose metabolism. Plasma ERK5 signaling contributes to glucose-induced FN overproduction. fasting FFA and AR progressively increased from NGT to IFG to IFG/IGT and Supported by: Canadian Diabetes Association DM. Adipocyte IR correlated with whole body glucose disposal (r=-0.621, p<0.005), hepatic insulin resistance (r=0.766, p<0.005), insulin secretion/ insulin resistance (disposition) index 6I0-120/6G0-120 X MI (r=-0.248, p=0.01) 1720-P and the non-oxidative glucose disposal during the insulin clamp (r=-0.361, Exenatide Ameliorates Insulin Resistance by Upregulating SIRT1 p=0.03). Compared to NGT subjects the disposition index (6I0-120/6G0-120 x MI) Expression of Adipose Tissue in db/db Mice was reduced by 18%, 68%, 80% in IFG, IGT, and combined IFG/IGT respec- ZONGLAN CHEN, HUIMIN GU, FEN XU, JINHUA YAN, HUA LIANG, JIANPING tively. In conclusion subjects with IFG are characterized by hepatic insulin WENG, Guangzhou, China resistance while those with IGT primarily have skeletal muscle insulin resis- Although Exenatide can improve insulin sensitivity of obesity and type 2 tance and are characterized by marked beta cell dysfunction. Therapeutic diabetes, the underlying mechanisms are still poorly understood. Growing interventions to preserve beta cell function should be started early. evidence suggests that SIRT1 has a positive role in the metabolic pathway through its direct or indirect involvement in insulin signaling. In this study, we investigated the regulation of Exenatide on SIRT1 expression in white adipose tissue of db/db mice. The 7-8 weeks male db/db mice and their & 1722-P heterozygous were divided into 3 groups of 10 animals each as follows: NC Insulin Sensitivity, Insulin Secretion and Disposition Index in Per- group (db/m mice, normal control), DM group (db/db mice), EX group (db/ sons With Normal and “Pre-Diabetic” HbA1c Levels: The Pathobiol- db mice treated with Exenatide 24nmol/kg/d for 8 weeks). Intraperitoneal ogy of Prediabetes in A Biracial Cohort (POP-ABC) Study insulin tolerance test (IPITT) indicated Exenatide could improve insulin sen- CHIMAROKE EDEOGA, SOTONTE EBENIBO, SAMUEL DAGOGO-JACK, Memphis, sitivity and lower fasting glucose levels signifi cantly in db/db mice. Western TN blotting analysis demonstrated that DM group had decreased expressions of An A1c range of 5.7-6.4% has been suggested for the diagnosis of predia- SIRT1, IRS1 and increased expression of p-JNK as compared with NC group. betes. In this report, we compared metabolic data in persons with normal All of these changes were attenuated or reversed when the diabetic mice (N=189) or “prediabetic”(N=98) status based on A1c. We studied 287 nondia- were treated with Exenatide. Our data suggested that Exenatide treatment betic adult offspring (147 black, 140 white) of type 2 diabetes (T2DM) par- ameliorated insulin resistance in db/db mice potentially through regulating ents enrolled in POP-ABC, a study of incident dysglycemia among offspring SIRT1-p-JNK-IRS1 axis. of T2DM parents. Their mean (+ SD) age was 44.2 + 10.6 y and BMI was 30.2 + 7.25 kg/m2. Baseline measurements included anthropometrics, 75-g OGTT and A1c level, followed by IVGTT and DEXA within 3 months of enrollment. Insulin action (M) was measured using euglycemic clamp, and beta-cell func-

tion was assessed using acute insulin response (AIR) during IVGTT. The dis- Obesity

position index (DI) was calculated as the product of AIR and M. Basal homeo- POSTERS stasis was assessed by calculating HOMA-IR and HOMA-B. Compared to

the normal controls (A1c < 5.7%), subjects with “prediabetic”A1c levels (5.7- Integrated Physiology/ 6.4%) were older (47.9 + 9.2 (SD), P=0.0004), and had higher fasting glucose (92.2 + 6.45 vs. 90.2 + 7.25 mg/dl, P=0.011), BMI (31.5 +7.97 vs. 29.2 + 13.2 kg/m2, P=0.03), total body fat (P=0.02) and trunk fat mass (P=0.016). How- ever, there were no signifi cant differences between the two groups with regard to HOMA-IR, HOMA-B, AIR, M value or DI in African Americans or Supported by: Program for Changjiang Scholars and Innovative Research Team Caucasiand (Fig 1). We conclude that an A1c range of 5.7-6.4% is associated with increased adiposity, but does not predict established glucoregulatory impairments that enable distinction from persons with lower A1c values.

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A443 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

false positive fi ndings. Fortyfi ve SNPs, selected to cover over 90% of com- mon genetic variability, were genotyped in 8 MODY (HNF4A, GCK, HNF1A, PDX1, HNF1B, NEUROD1, KLF11, CEL) and 2 neonatal diabetes (NDM) (KCNJ11, ABCC8) genes. Allelic variants of 4 SNPs (rs1303722 and rs882019 of GCK, rs7310409 of HNF1A and rs5219 of KCNJ11, the latter being a known type 2 diabetes risk variant) exerted independent signifi cant effects on DC of beta- cell function (p=0.007-0.03). Allelic variants of 5 other SNPs (rs2869084 and rs6031544 of HNF4A, rs10774580 of HNF1A, rs1801262 of NEUROD1, and rs7129639 of ABCC8) were found to infl uence signifi cantly PC of beta-cell function (p=0.001-0.04). Only 1 (rs6721191 of KLF11) out of 45 SNPs was asso- ciated to insulin sensitivity (p=0.047). In multivariate models, combining GCK, HNF1A and KCNJ11 SNPs accounted for ~2,5% of DC of beta-cell function, whereas combining HNF4A, HNF1A, NEUROD1 and ABCC8 SNPs accounted Supported by: NIH-NIDDK for ~3.6% of PC of beta-cell function. We conclude that common variability of MODY and NDM genes signifi cantly affects beta cell function in patients with type 2 diabetes, thereby potentially playing a role in the pathophysiol- & 1723-P ogy and/or the “metabolic prognosis” of the disease. Short Term Mild Hyperglycemia Improves Oral Glucose Disposal Supported by: European Foundation for the Study of Diabetes/Novartis Grant and Ƶ-Cell Function in Subjects With Impaired Glucose Tolerance (IGT) LORENA A. WRIGHT, KRISTINA M. UTZSCHNEIDER, JENNY TONG, SEDA SUVAG, & 1725-P STEVEN E. KAHN, Seattle, WA, Cincinatti, OH Mutations in ABCA1 and Enhanced Ƶ-cell Secretory Capacity in Hu- Mild 24 h hyperglycemia improves insulin sensitivity, `-cell function and mans: Preliminary Results glucose disposal in subjects with normal glucose tolerance. To determine MICHAEL R. RICKELS, EUGENE S. GOESER, CARISSA FULLER, CHRISTINE LORD, whether such an intervention has similar benefi ts in subjects with IGT, 6 ANNE M. BOWLER, MARINA CUCHEL, Philadelphia, PA such subjects (4F/2M, 52.4±5.7 y [x±sem], BMI 35.1±6.5 kg/m2 ) were studied Loss-of-function mutations affecting the cholesterol transporter ABCA1 twice, 4 weeks apart. They received infusions of glucose (10% dextrose; impair cellular cholesterol effl ux and are associated with reduced HDL-C D10) or normal saline (NS) in random order. After 24 h of infusion, a fre- levels that are nearly absent in the homozygous state (Tangier disease). Pub- quently sampled IV glucose tolerance test (FSIGT) was performed, followed lished data suggest that ABCA1 may be important in regulating `-cell cho- by an OGTT. Insulin sensitivity index (SI), acute insulin response (AIRg), dis- lesterol homeostasis and insulin secretion. To determine whether loss-of- position index (DI= SI x AIRg) and IV glucose tolerance (Kg) were computed function mutations in ABCA1 affect `-cell secretory capacity we performed from the FSIGT. Incremental areas under the curve (iAUC) for glucose and glucose-potentiated arginine tests in 6 subjects with heterozygous loss-of- insulin and iAUC insulin/iAUC glucose as a measure of `-cell function were function mutations in ABCA1 and 6 healthy control subjects (HDL-C 28 ± 3 vs. computed from the OGTT.Plasma glucose increased after 24 h of the D10 70 ± 7 mg/dl, p < 0.001; LDL-C 110 ± 4 vs. 100 ± 9 mg/dl, p = NS) pair-matched infusion (5.85±0.35 to 6.87±0.58 mM; p<0.004), but did not change with NS for age, gender, race, BMI and fasting glucose, as well as in one homozygous (5.59±0.40 to 5.59±0.38 mM). SI, AIRg nor DI or Kg changed signifi cantly subject (HDL-C 2 mg/dl, LDL-C 62 mg/dl). The acute insulin and C-peptide re- with D10 (Table). However, with D10 the iAUC glucose decreased and iAUC sponses to 5 g iv arginine were measured fasting (AIRarg & ACRarg) and during insulin increased, so that iAUC insulin/iAUC glucose increased.In summary, 230 mg/dl (AIRpot & ACRpot) and 340 mg/dl (AIRmax & ACRmax) glucose clamps. mild short term hyperglycemia improved `-cell function during the OGTT by AIRarg was not different between ABCA1 heterozygotes and controls (33 ± 240%, while the response to IV glucose (AIRg) increased by only 48%. We 11 vs. 26 ± 7 μU/ml), while under glucose-potentiated conditions both AIRpot conclude that in IGT, the `-cell retains its ability to respond to a physiological (209 ± 43 vs. 116 ± 27 μU/ml, p < 0.05) and AIRmax (226 ± 38 vs. 164 ± 41 μU/ increase in glucose. The fact that this benefi cial effect of short term hyper- ml, p < 0.05) were greater in ABCA1 heterozygotes than controls. There was glycemia was more apparent during the OGTT suggests that under these a trend toward greater acute C-peptide responses in ABCA1 heterozygotes conditions in IGT, the incretin response is preserved. than controls under all conditions (ACRarg: 1.8 ± 0.4 vs. 1.3 ± 0.2; ACRpot: 8.4 ± 2.2 vs. 6.4 ± 1.3; ACRmax: 8.3 ± 1.5 vs. 6.0 ± 0.7 ng/ml; p ) 0.1 for all). There Variable Saline (NS) Glucose (D10) P value was no difference in insulin sensitivity (M/I). The homozygous subject had ~ Infusion Infusion 4-fold increased acute insulin (AIRarg: 121; AIRpot: 794; AIRmax: 707 μU/ml) and S (x 10-5 min-1/pM) 0.31±0.04 0.41±0.13 0.38 I C-peptide (ACRarg: 4.8; ACRpot: 26; ACRmax: 26 ng/ml) responses as compared AIRg (pM.10 min) 2792±1593 4025±2425 0.24 with controls. These data indicate that loss-of-function of ABCA1 may be DI (x102 min-1) 753±331 1062±409 0.23 associated with enhanced `-cell secretory capacity that was dramatic in the homozygous subject, and support the importance of cellular cholesterol Kg (%/min) 1.38±0.28 1.92±0.22 0.23 homeostasis in regulating `-cell insulin secretion in humans. iAUC glucose (mM.120 min) 344±67 226±58 0.03 Supported by: PHS Grants UL1RR024134 and P30DK19525 iAUC insulin (pM.120 min) 28986±7088 45871±8927 0.009 iAUC insulin/iAUC glucose (pM/mM) 85±19 240±41 0.009 & 1726-P Data are x±SEM; paired t-test Decreased Activities of Proinsulin Convertase Enzyme (PC)3 Rather than PC2 in Pancreatic Beta-Cells Could Predict an Increased Risk of Type 2 Diabetes & 1724-P HIDENORI KATSUTA, KIYOSHI SUZUKI, SACHIHIKO OZAWA, TOSHIHIKO SADA- Common Variants of Monogenic Diabetes Genes Infl uence Beta Cell KARI, AKIKO SHIDA, TOSHITAKA MURASHIMA, HIROHISA ONUMA, KAORI Obesity Function in Patients With Newly Diagnosed Type 2 Diabetes MIYOKAWA-GORIN, YOSHIKAZU SUMITANI, TOSHIAKI TANAKA, SUSUMU POSTERS MADDALENA TROMBETTA, SARA BONETTI, LINDA BOSELLI, MARCO DAURIZ, NISHIDA, HITOSHI ISHIDA, Tokyo, Japan GIOVANNI MALERBA, ELISABETTA TRABETTI, PIER FRANCO PIGNATTI, ENZO Relative hyperproinsulinemia in the fasting state has long been character- Integrated Physiology/ BONORA, RICCARDO C. BONADONNA, Verona, Italy ized as a manifestation of impaired beta-cell function often seen in type 2 We tested the hypothesis that common genetic variability of “beta cell” diabetes (T2D). We investigated whether the altered activities of proinsulin genes responsible for monogenic diabetes may affect beta cell function convertase enzyme (PC)3 and PC2 would participate in the prevalence of in type 2 diabetes (DM2). We studied 590 drug-naive GAD-negative pa- T2DM in subjects with various stages of glucose intolerance. The circulating tients with newly diagnosed DM2 (age: median=60.0 yrs; I.Q. range: 52-66; levels of intact proinsulin (i-PI) were determined using a CLIA and those of BMI:29.3 kg/m2; 26.5-32.9). Beta cell function was assessed by state-of-art intermediate des-31,32-proinsulin (des-31,32-PI) were evaluated from a RIA mathematical modeling of glucose/C-peptide curves during a 240’ frequently for total proinsulin (t-PI) that had also a 95% cross-reactivity to des-31-32- sampled OGTT, to provide the beta cell responses to: i. the rate of increase PI, using identical blood samples taken from subjects of 207 normal glyce- of glucose (derivative control: DC) and ii. glucose concentration itself (pro- mic tolerance (NGT), 287 IFG/IGT and 201 T2D classifi ed according to the portional control, PC). Insulin sensitivity, measured in all patients by the WHO criteria. Based on the above measurements, the PC3 and PC2 activi- insulin clamp technique, was used as an internal control for the number of ties were estimated by the following formulas:des31,32-PI=(t-PI-i-PI)/0.95,

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A444 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

PC3 activity=des31,32-PI/i-PI, PC2 activity=IRI/des31,32-PI. The levels of clinical deterioration are unclear. In children, an association was previously i-PI/IRI were signifi cantly higher in the T2D group than those in the NGT reported between current and future clinical deterioration and HG (increased group (p<0.01). The levels of des-31,32-PI and PC2 activity were not differ- glucose excursions and especially glucose at 60min during an oral glucose ent among the all groups but PC3 activity was signifi cantly decreased in tolerance test (OGTT)) or HI (reduced insulin excursion during an OGTT). We both of IFG/IGT and T2D groups (p<0.01). Moreover, we examined whether hypothesized that HG or HI during an OGTT could also be associated with PC3 was an independent predictor of the prevalence or development of clinical deterioration in adults. First, we studied, at baseline, 225 adults with T2D as HOMA-` or Oral Disposition Index (DIO) was suggested. A lower CF (without known CFRD) who are part of an ongoing observational cohort. PC3()1.7), HOMA-`()30) and DIO(<1.1) were associated with an increased Increased glucose levels at the 60thmin-OGTT were associated with de- risk of T2D (unadjusted odds ratio [OR] 1.94 [95%CI 1.36-2.78], p<0.001, 1.95 creased lung function (spirometry: FEV%; r = -0.241, p <0.01), and decreased [95%CI 1.39-2.71], p<0.001, 11.2 [6.78-18.4], p<0.001 ). In multivariate stud- insulin levels at that time point were associated with decreased BMI (kg/ ies, PC3()1.7) was still associated with an increased risk of T2D (2.14 [1.25- m2; r = -0.298, p <0.01). Second, the results of the OGTT were used to clas- 3.64], p<0.05) after adjustment for age, sex, BMI, Cre, HDL-C, TG, HOMA-` sify the CF patients as either normal glucose tolerant (NGT), glucose intoler- and DIO. These results strongly suggest that lower PC3 activity rather than ant (IGT), diabetic de novo (CFRD) or as indeterminate HG (INDET; glucose PC2 could be a useful marker of an increased risk of T2D through an impaired <7.8mM at 0min and 120min, *11.1mM at 60min). Similarly to IGTs, INDETs proinsulin conversion. had increased glucose area under the curve during the OGTT and a trend towards decreased lung function compared to NGTs. Our results suggest th & 1727-P that 1) HG and HI at the 60 min-OGTT could be good indicators of decreased lung function and weight loss, respectively, and 2) the study of IGT & INDET Baseline Insulin Resistance May Not Contribute Per Se to Progres- patients could allow the identifi cation of early markers of clinical deteriora- sive Loss of Ƶ-Cell Function: The Insulin Resistance Atherosclero- tion. Prospective analyses are now required to establish predictors of future sis Study (IRAS) clinical deterioration in adults with CF. STEVEN M. HAFFNER, ANTHONY J. HANLEY, LYNNE E. WAGENKNECHT, MAR- Supported by: Cystic Fibrosis, Canada IAN J. REWERS, ANDREW J. KARTER, CARLOS LORENZO, San Antonio, TX, To- ronto, ON, Canada, Winston-Salem, NC, Aurora, CO, Oakland, CA Hyperglycemia may result from an intrinsic `-cell defect which results in Guided Audio Tour: Insulin Secretion and Sensitivity—Humans—Type 1 inadequate compensation for insulin resistance. There are few data on the and Type 2 Diabetes (Posters 1729-P to 1735-P), see page 17. impact of insulin sensitivity, adiposity, and plasma glucose levels on future changes in `-cell function. Therefore, we examined the association of insulin & sensitivity index (SI), glucose levels, and body mass index (BMI) with the 1729-P 5-year change in acute insulin response (AIR) in 741 non-diabetic subjects. A Comparative Assessment of the Effect of Caloric Load on Meta- bolic Parameters Evaluated in a Mixed Meal Tolerance Test SI and AIR were measured at the baseline and follow-up visits by the fre- quently sampled intravenous glucose tolerance test. In a generalized linear SUDHA S. SHANKAR, LORI MIXSON, DAVID E. KELLEY, Rahway, NJ mixed model analysis, baseline glucose levels were inversely associated The Mixed Meal Tolerance Test (MMTT) offers a physiologically relevant with the 5-year change in AIR (defi ned as follow-up AIR minus baseline AIR), situation to assess effects of disease and of treatment on insulin secretion. Beta cell sensitivity to glucose ( ) is one of the key parameters of insulin whereas baseline SI and BMI were not (Model 1). Five-year changes in BMI, \ secretion that is particularly refl ective of beta-cell impairment and response SI, and 2-h glucose were independently associated with the 5-year change in AIR (Model 2 and 3). In conclusion, weight gain and longitudinal declines to therapy. However, there is scant information on the contribution of meal in insulin sensitivity are associated with increases in `-cell function. This size and composition upon \ and other parameters of insulin secretion in supports the concept that `-cell function adapts to insulin action. Insulin patients with Type 2 diabetes mellitus (T2DM). We tested the hypothesis resistance may not contribute per se to `-cell dysfunction. Elevated plasma that while the metabolic responses to a lower caloric load would differ from glucose levels may have a deleterious effect on the `-cell and/or may signal that to a meal with higher caloric load, beta-cell sensitivity to glucose would an intrinsic `-cell defect. be an “intrinsic” property that does not vary in relation to meal size. A reto- Table: Multiple mixed regression models with the 5-year change in log AIR spective cross-sectional comparative analysis of data was performed, using as the dependent variable MMTT data from trials in T2DM patients with similar baseline character- istics, who had received either a low calorie meal of 460Kcal (LCM) or a Model 1 Model 2 Model 3 high calorie meal of 680 Kcal (HCM). Both meals had similar macronutrient ` ± SE ` ± SE ` ± SE composition of ~ 66% carbohydrate, ~18% fat, and ~16% protein. MMTT BMI - 0.005 ± 0.024 - 0.035 ± 0.024 - 0.035 ± 0.023 (180 minutes, with 9-point sampling) data from four trials, two with a LCM (N=114) and two with a HCM (N=125) were compared. Data are presented as Log S 0.036 ± 0.042 - 0.099 ± 0.034 † - 0.175 ± 0.034 † I LCM vs HCM, mean ± SEM. Signifi cant differences were observed in incre- Fasting glucose - 0.073 ± 0.026 † - 0.089 ± 0.024 ‡ - 0.082 ± 0.024 ‡ mental AUC(0-3hr) glucose (72.9 ± 3.3 vs 89.7 ± 3.8 mg.h/dL, p<0.05), AUC(0-2hr) 2-h glucose - 0.064 ± 0.024 † - 0.080 ± 0.025 † - 0.117 ± 0.026 ‡ insulin (71.4 ± 9.2 vs 145.3 ± 12.1 μIU.hr/mL, p<0.01) AUC(0-2hr) C-peptide (10.1 5-year change in BMI — 0.042 ± 0.021 * 0.045 ± 0.020 * ± 0.4 vs 15.2 ± 0.7 ng.hr/mL, p<0.05) and AUC insulin secretion rate (513.3 ± 25.8 vs 609.8 ± 21.0 pmol.h/min, p<0.01). However, the measure of beta cell 5-year change in log S — - 0.147 ± 0.025 ‡ - 0.203 ± 0.026 ‡ I sensitivity to glucose (\) was comparable between the two caloric loads 5-year change in fasting glucose — — - 0.043 ± 0.026 (9.6 ± 0.9 vs 8.9 ± 0.8, p=NS). Our fi ndings suggest that acute metabolic 5-year change in 2-h glucose — — - 0.112 ± 0.027 ‡ responses such as glycemic excursion, insulin and C-peptide differ between MMTTs having different caloric loads, whereas intrinsic parameters of beta ` ± SE expressed for 1 SD unit increase; * p <0.05; † p <0.01; ‡ p <0.001 Age, cell function/beta cell responsivity to a mixed meal challenge do not. sex, race/ethnicity, clinic, family history of diabetes, and baseline AIR were also included in all three models. Supported by: NHLBI (HL-47887, HL-47889) & 1730-P

The Graded Glucose Infusion is a Simple yet Sensitive Tool to Detect Obesity

Differences in Insulin Secretion due to Metabolic Dysfunction and POSTERS Treatment of Disease

& 1728-P SUDHA S. SHANKAR, RAVI SHANKAR, LORI MIXSON, DEBORAH L. MILLER, BAR- Integrated Physiology/ Association of Hyperglycemia or Hypoinsulinemia During an Oral NALI PRAMANIK, CHAN R. BEALS, HELMUT O. STEINBERG, DAVID E. KELLEY, Glucose Tolerance Test With Worse Clinical Profi le Before the On- Rahway, NJ set of Cystic Fibrosis-Related Diabetes in Adults We have previously shown in healthy volunteers, that the Graded Glu- ADÈLE CORIATI, MARIE-SOLEIL GAUTHIER, SOPHIE ZIAI, LINDA BELSON, YVES cose Infusion (GGI) is comparable to the hyperglycemic clamp to measure BERTHIAUME, RÉMI RABASA-LHORET, Montreal, QC, Canada changes in glucose-dependent insulin secretion (GDIS), while being opera- Approximately 40% of adults with cystic fi brosis (CF) develop CF-related tionally simpler. We tested the hypothesis that the GGI can distinguish dif- diabetes (CFRD), a complication that increases mortality. Two to four years ferences between metabolic states, and in response to treatment in Type 2 before its diagnosis, CFRD is preceded by an accelerated clinical deteriora- DM (T2DM), by comparing GDIS between lean (L, n=8), obese non-diabetic tion (decreases in weight and lung function). CFRD is characterized by hy- (OB, n=12) and T2DM (T2DM, n=12); in T2DM, GDIS was also measured after perglycemia (HG) and hypoinsulinemia (HI), but their respective roles in this a single subcutaneous dose of liraglutide 0.6mg (LIRA). All subjects under-

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A445 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

went a 160-minute GGI with fi xed stepwise increments in glucose infusion Diabetes Prevention Program at baseline and after placebo (PLC), intensive rates (GIR) from 2 to 12 mg/kg/min. Data are presented as mean ± SEM lifestyle (ILS), metformin (MET), or troglitazone (TROG) treatment. Follow-up (Table 1). At similar GIR, signifi cant differences (p<0.01) were observed be- in all arms was 2 years, except for TROG, which was stopped after 1 year. tween all groups in maximal glycemic excursion (G), insulin secretion (ISR), At baseline, log (NT-proBNP) did not differ across groups (Table, p=0.07). and beta cell sensitivity to glucose (ISR/G). Following LIRA, there were sig- At follow-up, log (NT-proBNP) increased in TROG and ILS groups only (non- nifi cant (p<0.001 over PBO) changes in G (222.3 (12.4) mg/dL), ISR (3.65 (0.44) transformed change in NT-proBNP is shown below). Changes in BMI and in- ng/min), and ISR/G (0.023 (0.004)) in T2DM, achieving treatment effects sulin sensitivity (as log-transformed 1/HOMA-IR) are shown below. Change similar to PBO-treated OB (p=NS for comparison). These data indicate that in NT-proBNP was correlated with the change in insulin sensitivity (partial compared to L, OB show compensatory increases in insulin secretion, while correlation coeffi cient = 0.12, p<0.0001, after adjustment for change in BMI, T2DM have impaired beta-cell function; the effect of liraglutide reveals a age, sex, race, and baseline values of NT-proBNP, BMI, and log [1/HOMA-IR]) latent functional reserve in T2DM, responsive to incretin stimulation.These in all groups, and only with the change in BMI in the placebo group (-0.10, fi ndings demonstrate for the fi rst time in a systematic comparison, that the p=0.007). Within the DPP, percentage of TROG participants with heart fail- GGI is sensitive enough to detect differences in GDIS due to metabolic dys- ure (0%) or edema (4.6%) did not differ from PLC.TROG, ILS, and MET data function, as well as treatment. suggest that changes in insulin sensitivity have a greater infl uence on NT- Table 1. GGI data in lean, obese and diabetic (mean ± SEM) proBNP than changes in BMI. Parameter Lean Obese T2DM PLC ILS MET TROG (PBO) (PBO) (PBO) Baseline NT-proBNP (pg/mL) Maximal glycemic excursion during the GGI, 153.3 (13.0) 185.2 (12.8) 277.6 (9.2) Median [IQR] 33.42 [17.12-62.28] 30.90 [13.99-59.89] 29.68 [14.04-54.17] 29.11 [15.66-52.93] as a change from baseline (G), mg/dL N 762 916 959 583 Time-weighted average (TWA) of the insulin secretion 2.85 (0.40) 3.93 (0.47) 1.59 (0.37) ৫ Nt-proBNP (pg/mL) rates (ISR) at the highest glucose infusion rate, as a Median [IQR] +1.32 [-12.4-19.81] +3.20 [-8.81-19.99] +1.45 [-10.5-15.66] +6.28 [-5.39-6.84] change from baseline, in ng/min N 692 832 887 285 Slope of the insulin secretion rate vs ambient glucose 0.015 (0.006) 0.021 (0.004) 0.006 (0.001) p-value 0.19 0.008 0.82 0.0003 over the duration of the GGI (ISR/G) ৫ BMI (kg/m2) Mean±SEM +0.03±0.08 -1.96±0.09 -0.75± 0.07 +0.37± 0.13 N 689 837 887 279 p-value 0.69 0.0001 0.0001 0.005 & 1731-P ৫ Log [1/HOMA-IR] Effects of Acylated and Unacylated Ghrelin on Insulin Secretion and Mean±SEM -0.02± 0.02 +0.22±0.02 +0.11±0.02 +0.31±0.03 Glucose Tolerance in Healthy Humans N 673 818 860 281 JENNY TONG, RON L. PRIGEON, MARTIN BIDLINGMAIER, ELIZABETH STAM- p-value 0.35 0.0001 0.0001 0.0001 BROOK, MATTHIAS TSCHOEP, DAVID D’ALESSIO, Cincinnati, OH, Baltimore, MD, 6-change Munich, Germany Supported by: NIDDK (U01-DK048489) Unacylated ghrelin (UAG) is the predominant ghrelin isoform in the circula- tion, and despite an identical peptide sequence with acyl-ghrelin (AG) does not activate the classical ghrelin receptor (growth hormone secretagogue & 1733-P receptor, GHSR 1a). AG inhibits insulin secretion in humans, but preclinical 50-Year Medalist Study: Pattern of Insulin Positive Cells and its Cor- studies suggest that UAG may promote `-cell function. We hypothesized relation With C-peptide Levels and Age at Onset that UAG would have the opposite effect of AG on insulin secretion and SUSAN BONNER-WEIR, HILLARY A. KEENAN, BROOKE MORRIS, STEPHANIE glucose tolerance. AG (1 μg/kg/h), UAG (4 μg/kg/h), combined AG and UAG, HASTINGS, SARA TUREK, JENNIFER K. SUN, GEORGE L. KING, Boston, MA or saline were infused to 9 healthy subjects (5M/4F; age 27 ± 2 y; BMI 25 ± The Medalist Study provides a unique opportunity to compare physiologi- 1 kg/m2, fasting plasma glucose 92 ± 2 mg/dl, mean ± SEM) on 4 separate cal factors and post-mortem pancreatic morphology of T1DM patients with occasions in randomized order. AG and UAG were infused for 30 min to >50 years duration. We studied 21 Medalist pancreases and all were found achieve steady-state levels and continued through a 3-h frequently sam- to have insulin positive beta cells. The distribution and quantity of insulin pled IV glucose tolerance test (FSIGT). Acute insulin response to IV glucose positive cells were categorized as: C1) few scattered singlet or clusters (n=6), C2) similarly scattered with additional occasional insulin positive cells (AIRg), insulin sensitivity index (SI), disposition index (DI = SI x AIRg) and IV glucose tolerance (Kg) were computed from the FSIGT. AG and the combined within islets (n=11) and C3) at least 30% of the islets having signifi cant num- AG+UAG infusion increased serum GH 20-fold from baseline, while UAG and ber of insulin positive cells (n=4). Most islets in Medalists were atrophic saline had no effect on GH levels. Only AG infusion increased fasting glucose comprised mainly of glucagon positive cells; some scattered glucagon cells levels (p <0.05); none of the treatments had any effect on fasting plasma were in or near ducts. Clinical data was analyzed across categories, only age insulin. Both AG and AG+UAG decreased AIRg (saline 451 ± 93, AG 328 ± of onset and c-peptide levels were signifi cantly different (p ) 0.01). Those 80, AG+UAG 299 ± 80 pM/l, p = 0.005). UAG also showed a trend towards in C1 were the youngest at onset (3.8±2.0 y) and had the lowest c-peptide (0.07±0.07 ng/dl) while those in C2 had a mean age at onset of 9.3±7.1 y and a lower AIRg (364 ± 79 pM/l, p =0.08) as compared to saline. SI was not af- fected by any of the ghrelin treatments. The AG+UAG treatment decreased a c-peptide of 0.21±0.01 ng/dl, and C3 had a later onset at 25.3±6.3 y and a DI and Kg (p < 0.05 for both). In summary, AG suppresses insulin secretion higher c-peptide level, 2.2±2.2 ng/dl. All but 2 (in C2) had high risk DR3 and/ and impairs glucose clearance as previously reported. UAG has effects that or DR4 alleles. Antibody positivity (IA2 or GAD65) spanned categories with approach those of AG and the combination of AG+UAG may have additive 2 in C1, 4 in C2 and 1 in C3. C3 had the most diverse morphology: 2 having at effects on `-cell function. In contrast to the preclinical studies, these data least half the pancreas with few scattered insulin positive cells and no islets suggest similarities of the two isoforms of ghrelin in the regulation of glu- with insulin positive cells and the rest of the pancreas having islets com- cose metabolism. posed of at least 30% insulin positive cells and many with amyloid deposits. One of these was IA2+.Analysis of the Medalist cohort (n=659) by catego- Obesity Supported by: NIH (k23DK80081, R03DK89090)

POSTERS ries defi ned by mean c-peptide level of C1, C2 and C3 showed a similarly signifi cant increase in age at onset (p=0.04). Insulin positive cells persist in & 1732-P very long-term diabetes even in the absence of measurable c-peptide and a Integrated Physiology/ Preventive Therapies for Type 2 Diabetes that Improve Insulin Sen- greater number of insulin positive cells correlates with higher c-peptide and sitivity Raise NT-proBNP later onset of disease. Beta cells persisted as scattered singlets more com- GEOFFREY A. WALFORD, YONG MA, RONALD B. GOLDBERG, PETR JAROLIM, ED- monly than islets in T1DM patients of extreme duration. WARD S. HORTON, KIEREN J. MATHER, ELIZABETH BARRETT-CONNOR, JACLYN DAVIS, JOSE C. FLOREZ, THOMAS WANG, DPP RESEARCH GROUP, Boston, MA, Rockville, MD, Miami, FL, Indianapolis, IN, San Diego, CA Individuals with obesity and insulin resistance have reduced circulating levels of natriuretic peptides (NPs), which may contribute to increased sus- ceptibility to hypertension and cardiac disease. The effect of interventions that reduce metabolic risk on NP concentrations is unclear.We analyzed N-terminal B-type natriuretic peptide (NT-proBNP) concentrations in the

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A446 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

Guided Audio Tour: Insulin Secretion and Action in Animal Models (Posters & 1734-P 1738-P to 1742-P), see page 13. BMI is the Major Driver of Beta Cell Loss in RecentOnset Type 1 Diabetes: a Multicentre, Longitudinal Study 1736-P ADAM BARKER, ANGELO LAURIA, NANETTE SCHLOOT, NORA HOSSZUFALUSI, Triglyceride-Rich Lipoprotein Metabolism in the Central Nervous PAL PANCZEL, JOHNNY LUDVIGSSON, CHANTAL MATHIEU, DIDAC MAURICIO, System Regulates Brain Lipid Sensing and Plays an Essential Role in Body Weight RegulationWITHDRAWN ESTER RUBINAT, MARIA NORDWALL, BART VAN DER SCHUEREN, THOMAS MA- HONG WANG, CHARLES W. REHRER III, MATTHEW D. TAUSSIG, AMBER A. DRUP-POULSEN, WERNER SCHERBAUM, NICK WAREHAM, ILSE WEETS, FRANS PALUMBO, KALYANI G. BHARADWAJ, IRA J. GOLDBERG, ROBERT H. ECKEL, Au- GORUS, DAVID LESLIE, PAOLO POZZILLI, Cambridge, United Kingdom, Rome, Italy, rora, CO, New Yorks, NY Düsseldorf, Germany, Budapest, Hungary, Linköping, Sweden, Leuven, Belgium, Lipoprotein lipase (LPL) is rate-limiting in the hydrolysis of triglycerides (TG) and uptake of fatty acids from circulating dietary lipids. We have recent- Lleida, Spain, Copenhagen, Denmark, Brussel, Belgium, London, United Kingdom ly created mice with neuron-specifi c LPL defi ciency (NEXLPL-/-) that have C-peptide is a valuable indicator of residual `-cell function in type1 diabe- reduced uptake of TG-rich lipoprotein-derived fatty acids and decreased tes (T1D) patients. Although there is great heterogeneity in C-peptide levels levels of n-3 long chain polyunsaturated fatty acids (n-3 PUFAs) in the hypo- thalamus. Of great interest these mice become obese on chow by 16 wk (Cell at clinical onset, factors infl uencing the subsequent decline of C-peptide Metabolism, 2011). We have now conducted experiments in 10 wk old mice to are not well understood.The aim of this study was to investigate the natu- test if dietary supplementation of n-3 PUFAs prevents obesity. After 40 wk ral course of residual beta cell function in 4411 T1D patients diagnosed in on a synthetic high carbohydrate diet (HC, 70% CHO, 10% fat, n-6:n-3 ratio European centers with longitudinal data available up to 10 years following of 7:1) w/o n-3 PUFA supplement (n-6:n-3 ratio 1:3), NEXLPL-/- mice have an average of 35% fat mass vs. 27% in WT mice on both diets. After 24 wk on diagnosis. The patients were grouped according to age at diagnosis (0-4, synthetic high fat diets (HF, 45% fat) w/o n-3 PUFA supplement, both NEX- 5-9, 10-18 and >18years). Linear regression models with change in fasting LPL-/- and WT mice gain less body weight (38.0 g vs. 39.1 g on HF/n-3 PUFA; C-peptide as an outcome were used to identify variables measured at diag- 50.1 g vs. 47.6 g on HF diet; respectively). This reduction in body weight for both groups on the HF/n-3 PUFA diet is likely an effect of larger amounts of nosis including body mass index (BMI) [expressed as z-scores vs. a matched n-3 PUFA on palatability. Noticeably, HF feeding enhanced the body weight control population], HbA1c and insulin dose that were signifi cant predictors and fat mass gain in NEXLPL-/- mice much less than in WT mice, rendering of fasting C-peptide decline over the following years.There were no signifi - similar body compositions at the end of the HF feeding (42.6% vs. 42.0% cant predictors of fasting C-peptide decline in those with an age of onset fat mass, respectively). Taken together, these data suggest that when LPL is defi cient in neurons, n-3 PUFA supplementation in the diet cannot pre- between 0-4 years or 5-9 years. In individuals diagnosed between 10-18 vent obesity; moreover, NEXLPL-/- mice become insensitive to the amount years of age, the rate of C-peptide decline increased by 0.047 nM per year of fat in the diet and fatty acid composition. Overall, results from the feeding for each standard deviation increase in baseline BMI. In individuals diag- studies further support our hypothesis that LPL regulates TG-rich lipoprotein metabolism, providing important signaling molecule(s) such as n-3 PUFAs nosed at >18 years of age the most consistent predictor of fasting C-peptide that are involved in lipid sensing in the CNS, and plays an essential role in change was gender with all models showing that in women the decline over regulation of energy balance and body weight. 1-year was greater by approximately 0.13 nM.A higher baseline BMI isas- Supported by: NIH K-12 BIRCWH award, NIH-NIDDK R01 sociated with a greater rate of fasting C-peptide decline in adolescent T1D. This fi nding implies that in patients diagnosed between 10 and 18 years of age,decreased insulin sensitivity can play a key role in disease progression 1737-P and suggests thattherapies that target reduction in BMI might be helpful in preserving beta cell function. WITHDRAWN & 1735-P Healthy Pre-Monopausal Women With Type 1 Diabetes Have In- creased Peripheral Insulin Resistance Compared to Non Diabetic Women TALIA L. BROWN, RACHEL M. SIPPL, JANET K. SNELL-BERGEON, Aurora, CO Increased insulin resistance (IR) exists in adults and adolescents with type 1 diabetes (T1D) and appears to be independent of metabolic syndrome Guided Audio Tour: Insulin Secretion and Action in Animal Models (Posters factors, but has not been investigated in healthy premenopausal women 1738-P to 1742-P), see page 13. with T1D. The objective was to determine relative IR in premenopausal women with and without diabetes, using the gold standard hyperinsulinemic euglycemic clamp technique.Women with T1D (n=7, age 33±8 yr, diabetes & 1738-P duration 17±7 yr) and women without diabetes (Non-DM) (n=11, age 29±5 yr) Genetic Evidence that Hyperinsulinemia Causes Diet-Induced Obe- from the Women, Sex Hormones and Insulin study were recruited into the sity in Mammals by Preventing “Browning” of White Adipose Tis- IR sub-study. Inclusion criteria were 18-45 years of age with regular men- sue strual cycles, euthyroid, and A1c <9.5% for participants with T1D. Three- ARYA E. MEHRAN, NICOLE M. TEMPLEMAN, KWAN-YI CHU, XIAOKE HU, SU- stage (4, 8, and 40 mu/m2) euglycemic clamps were performed. Adipose and SANNE M. CLEE, JAMES D. JOHNSON, Vancouver, BC, Canada peripheral IR were measured as %free fatty acid suppression (FFA) at the We are taught that obesity, with its elevated lipids and low-grade infl am- end of the second stage and glucose infusion rate (GIR) during the fi nal 30 mation, fi rst leads to insulin resistance, then subsequently to compensatory minutes of the third stage, respectively.Women with T1D had both impaired insulin hypersecretion, followed by diabetes when the `-cells fail. However, FFA (41%±36 vs 86%±16 for Non-DM) and decreased GIR (4.9±3 (mg/kg/ there has long been clinical evidence that fasting hyperinsulinemia can pre- min) vs 12.4±5 (mg/kg/min) in Non-DM). Using multivariable linear regres- cede obesity, and clinical trials with some drugs that reduce insulin secretion sion models, diabetes status was an independent predictor of both GIR and show weight loss. To date, there has been no direct genetic demonstration FFA (Table). Models were adjusted for diabetes, stage-specifi c blood glucose that insulin itself modulates obesity in mammals. Using Ins1-/- and Ins2-/- levels and fi nal insulin serum concentrations, body mass index, triglycerides, mice as ideal negative controls, we found Ins1 is restricted to pancreatic HDL cholesterol, and systolic blood pressure.Healthy premenopausal wom- `-cells, while Ins2 mRNA and protein is expressed more widely (thymus, en with T1D are more likely to have adipose and peripheral insulin resistance brain). By selectively reducing the gene dosage of Ins1, we generated mice Obesity POSTERS than Non-DM women, independent of metabolic syndrome parameters. that did not exhibit fasting hyperinsulinemia in response to a prolonged high- fat diet. Reducing Ins1 gene dosage prevented islet hyperplasia in response Models for GIR and FFA. Beta coeffi cient and p-value represent diabetes sta- to high fat feeding, confi rming an essential role for autocrine/paracrine in- Integrated Physiology/ tus variable. sulin signaling in this process. Compared to Ins1+/+:Ins2-/- littermate con- Model Beta Coeffi cient P-Value trols, Ins1+/-:Ins2-/- mice were completely protected from obesity, insulin Glucose Infusion Rate resistance and hepatic steatosis caused by high fat feeding, in the absence Diabetes Status 7.702 0.0022 of sustained changes in glycemia. Genetic prevention of hyperinsulinemia increased energy expenditure and uncoupling protein expression in white Diabetes Status + Full Model 10.42 0.0007 adipose tissue (but not brown adipose tissue or muscle), while reducing adi- % Free Fatty Acid Suppression pose infl ammation and fatty acid spillover. Together, our data demonstrate Diabetes Status 41.99 0.0036 that the increase in fasting circulating insulin is required for diet-induced Diabetes Status + Full Model 53.33 0.0034 obesity in mammals.

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A447 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

standard diet (SD). After one week glucose levels in CORT-HFD rats were & 1739-P elevated (>11 mM) and animals were 15-fold more insulin resistant than con- Differential Development of Insulin Resistance and Beta Cell Adap- trols indicated by HOMA-IR levels (15.1±1.64 vs 1.0±0.12), whereas HOMA-` tation in Multiple Diet Induced Obesity Models indicated lower ` cell function compared to CORT-SD animals (3.72±0.64 BILAL OMAR, GIOVANNI PACINI, BO AHRÉN, Lund, Sweden, Padova, Italy vs 1.64±0.22). Moreover, ` and _ cell mass were 1.5- and 0.6-fold higher, The C57BL/6 mouse becomes insulin resistant and glucose intolerant af- respectively, in CORT-HFD treated animals compared to controls. Surpris- ter feeding with a high fat diet (>40% kcal) and is a widely used model for ingly, CORT-HFD isolated islets were found to be highly glucose responsive studying type 2 diabetes (T2D). Many different high fat diets exist and dif- as they released 10-fold more insulin compared to controls. Voluntary wheel ferences in the composition of macronutrients in the diets could induce dif- running showed that CORT-HFD animals had less visceral adiposity (16.2±3.5 ferent metabolic phenotypes. In order to determine which model produces vs 22.4±1.7mg/g) than controls. Blood glucose levels in sedentary CORT-HFD a phenotype most like that of human T2D, the diet models need to be evalu- rats were elevated (12.8±0.15mM) compared to running CORT-HFD animals ated in head-to-head comparisons. We evaluated the two most widely used (9.1±0.55mM). Exercised animals had improved glucose tolerance compared high fat diets in mice, the standard 60% high fat diet (HFD) and the high to sedentary CORT-HFD animals (600.1±121.5 vs 988.4±137.9 following oral fat-high sucrose Surwit diet (HFHS), using intravenous glucose tolerance glucose load, p<0.05). Moreover, insulin sensitivity was improved in running tests (IVGTT) and the euglycemic-hyperinsulinemic clamp, to determine ef- CORT-HFD animals compared to sedentary CORT-HFD animals (53.1±12.0 vs fects on insulin resistance and beta cell adaptation. Mice fed a HFD gained 92.3±26.4mmol/L). We conclude that ROD animals exhibit severe insulin re- more weight than HFHS fed mice (9.5 g v 6.7g, p<0.05) despite near identical sistance that overwhelms ` cell function. However, exercise intervention energy intake. Both high fat diet models show impaired glucose tolerance introduced in this model greatly improved glucose and insulin tolerance re- after 8 weeks but show marked differences in insulin secretion. Mice fed sulting in better glucose control and body fat composition. the HFD had elevated basal plasma insulin(+111nM, p<0.01), which was not seen in the HFHS fed group. The acute insulin response (AIR) was unchanged & 1742-P in the HFD group, but slightly increased in the HFHS diet group. The HFHS Partial Deletion of the Novel Antioxidant SEPS1 in Ƶ-Cells Leads to diet group had a 3 fold greater insulin secretion in response to the glucose Impairment in Insulin Secretion load during the IVGTT compared to the HS control diet group (iAUC 15nM CHIA-HENG WENG, TANYA SAMARASEKERA, NICOLE STUPKA, KEN WALDER, v 3nM, p<0.001), while no differences were seen in the HFD group. Insulin JOSEPH PROIETTO, SOF ANDRIKOPOULOS, NICOLE WONG, Heidelberg, Australia, sensitivity, as determined by euglycemic-hyperinsulinemic clamp, was de- Geelong, Australia creased 4-fold in the HFD group (M/I 0.24 μmol/g/nM ND v 0.06 μmol/g/nM Selenoprotein S (SEPS1) is an antioxidant enzyme that has been shown HFD, p<0.01) but not in the HFHS diet group after 8 weeks. After 16 weeks, to protect against oxidative stress (H O ) induced cell death in a pancreatic glucose tolerance was not signifi cantly impaired in either the HFD or HFHS 2 2 `-cell line. We have preliminary data that showed that `-cell specifi c over- diet groups. Basal insulin was signifi cantly elevated in both HFD and HFHS expression of SEPS1 protected against alloxan- and streptozotocin-induced groups but to a greater extent in the HFD group. We conclude that the HFD diabetes. Based on these fi ndings, we hypothesised that the deletion of and HFHS diet models show differential effects on the development of insu- SEPS1 will predispose mice to oxidative stress induced diabetes. The aim of lin resistance and beta cell adaptation. this study was to generate and characterise global and `-cell specifi c SEPS1 deleted mice. Global and `-cell specifi c SEPS1 deleted mice were generated & 1740-P by PGK-Cre and RIP-Cre respectively and fed a standard chow diet. At eight The SIRT1 Activator Resveratrol Prevents Fat-Induced Ƶ-Cell Dys- weeks of age, plasma glucose and insulin responses were examined follow- function In Vivo ing an intravenous glucose tolerance test (IVGTT) and an oral glucose toler- TEJAS DESAI, ALEX IVOVIC, BERNARD HAU, KHAJAG KOULAJIAN, DANNA ance test (OGTT) in deleted mice and negative littermates. Global deletion BREEN, ADRIA GIACCA, Toronto, ON, Canada of SEPS1 did not alter plasma glucose and insulin levels during the IVGTT Over the last decade, sirtuins and SIRT1 in particular, have been identifi ed when compared with the negative littermates. Homozygous `-cell-specifi c as key regulators of nutrients and metabolism. Within the `-cell, there is evi- deletion of SEPS1 led to embryonic lethality. Mice with a partial deletion dence that SIRT1 plays a benefi cial role on insulin secretion. It has also been of SEPS1 in the `-cell (+/-) had impaired glucose tolerance (1796.8±159.2 established that excess circulating fat as seen in obesity can be detrimental vs 1349.4±78.3 mmol/Lx120min, p < 0.05, n=5-10) associated with reduced to `-cell function (“`-cell lipotoxicity”), an effect that may involve decreased insulin levels (87.3±33.7 vs 168.2±21.6 ng/mLx120min, p<0.05, n=6-8). The SIRT1 activity. Consequently, it would seem logical that SIRT1 activation IVGTT confi rmed the reduced glucose mediated insulin secretory response may have a benefi cial role on `-cell function in conditions of nutrient excess. in `-cell specifi c SEPS1 (+/-) mice (36.2± 5.2 vs 80.0±7.2 ng/mLx30min, p< Here we attempted to mitigate lipotoxicity induced `-cell dysfunction using 0.005, n= 12-19). In conclusion, mice with a partial deletion of SEPS1 in the a pharmacological model of SIRT1 activation. Female Wistar rats (n= 4-8 `-cell displayed impaired insulin secretion and glucose intolerance, further per group) were infused intravenously for 48h with 1) Saline (SAL, control), emphasising the important role of SEPS1 in glucose homeostasis. 2) Oleate (OLE) at 1.4μmol/min to elevate plasma FFA by 50%, 3) Oleate + the SIRT1 activator Resveratrol (RSV, 0.025mg/kg.min) and 4) RSV alone 1743-P (0.025mg/kg.min). The infusion period was followed by assessment of -cell ` Micro-RNA 122 Regulates Insulin Secretion in Diet-Induced Obese function by hyperglycemic clamp. OLE infusion resulted in decreased plasma Mice insulin and c-peptide levels in response to the rise in glucose that were RANDALL H. FRIEDLINE, GREGORY WU, HWI JIN KO, YONGJIN LEE, XIAODI HU, signifi cantly decreased (>50%) compared to SAL control infusion (p<0.05). HSUN-FAN WANG, YOSHIHIRO AZUMA, SARAH R. NATHAN, YUN HEE NOH, In the group coinfused with OLE+RSV, complete prevention of the oleate- KLAUS PECHHOLD, JUN XIE, GUANGPING GAO, JASON K. KIM, Worcester, MA, induced decrease in insulin and c-peptide was seen compared to the SAL Seoul, Republic of Korea control infusion. No signifi cant difference was seen between SAL and RSV Micro-RNAs are integrally involved in many facets of cell differentiation, infusion alone. These results indicate that RSV prevents the -cell dysfunc- ` growth, and function, and are highly expressed in metabolic organs including tion caused by 48h OLE infusion, an effect that may be SIRT1 mediated. adipose tissue, liver, and pancreas. Here we examined the role of micro-

Obesity RNA-122 (miR-122) on glucose homeostasis by generating mice with adeno-

POSTERS & 1741-P associated viral knockdown of miR-122. Male C57BL/6 mice were treated Rapid-Onset Diabetes Induced by High Corticosterone Exposure with AAV-miR-122 or scrambled vector (Scr), and started on a high-fat diet

Integrated Physiology/ and High-Fat Feeding Impairs In Vivo Islet Function in Rats (HFD) 4 wks later. miR-122 treatment did not affect body weight or insulin JACQUELINE L. BEAUDRY, ANNA D’SOUZA, YANIV SHPILBERG, TREVOR TEICH, sensitivity in chow-fed mice. During 16 wks of HFD, miR-122 and Scr mice ROBERT TSUSHIMA, MICHAEL RIDDELL, Toronto, ON, Canada became obese with similar whole body fat mass. Basal plasma insulin lev- We have recently established a new rapid-onset-diabetic (ROD) rodent els tended to be lower in miR-122 mice compared to Scr mice after HFD. A model, induced by elevations in the stress hormone, corticosterone (CORT) 2-hr hyperglycemic clamp was performed in awake mice to assess glucose- and a high-fat diet (HFD). Together these treatments induce severe insulin induced insulin secretion in vivo (n=5/group). Plasma glucose levels were resistance however, rarely in concert have they been investigated on ` cell quickly raised and maintained at ~350 mg/dl in both groups of mice. Plasma function. Regular exercise alleviates insulin resistance and preserve ` cell insulin levels were similarly elevated during clamps in chow-fed mice (Fig. mass. We examined the short-term effect of these stressors and voluntary 1). In contrast, HFD-fed miR-122 mice showed lower plasma insulin levels exercise on ` cell dynamics in male Sprague-Dawley rats (~6 weeks) given during clamps compared to HFD-fed Scr mice (Fig. 2). This resulted in ~20% CORT pellets (400 mg/rat) or wax pellets (control) and placed on a HFD or reduction in area-under-curve of insulin levels in miR-122 mice, suggesting

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A448 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO blunted insulin secretion in these mice (Fig. 3). This was further confi rmed mg/kg/min, NHP showed robust glycemic and C-peptide responses through- by measuring total serum C-peptide levels which were 2-3 fold lower in out the 160-minute GGI (Fig.1). Data are represented as mean ± S.E. At the HFD-fed miR-122 mice following glucose injection (Fig. 4; *P<0.05). Overall, highest GIR, signifi cant (P<0.001) glycemic (124.2 mg/dL) and C-peptide (9.70 our fi ndings indicate that miR-122 knockdown impairs insulin secretion in ng/mL) responses expressed as a change from baseline, were observed. diet-induced obese mice and further identify a novel role of miR-122 in the In a similar experiment, HV had robust glycemic and C-peptide responses regulation of pancreatic function in obesity. (Fig.1). Comparison of data was done between NHP and HV (Fig.1). While at the highest GIR, NHP and HV had similar [geometric mean ratio (90%CI)] maximal glycemic [0.85 (0.63, 1.16), p=0.18], and C-peptide [1.62 (1.04, 2.53), p=0.04] responses, overall quantitative responses to a GC in NHP appear more robust than in humans. Together, these fi ndings demonstrate for the fi rst time in a systematic evaluation, that in adult healthy NHP, the GGI is able to detect metabolic responses to a glycemic challenge, and the re- sponse patterns are highly similar to those in healthy human volunteers.

Supported by: NIH (R01-DK080756)

1744-P GRP78 Over-Expression in Pancreatic Ƶ-Cells Protects from High Fat Diet-Induced Diabetes in Mice TRACY TEODORO, IRMGARD SCHUIKI, LILING ZHANG, ALLEN VOLCHUK, Toronto, ON, Canada Endoplasmic reticulum (ER) stress is a mechanism involved in type 2 dia- betes pathology and has been shown to contribute to both insulin resistance in liver and adipose tissue and pancreatic `-cell dysfunction. To examine whether ER stress in pancreatic `-cells contributes to the development of type 2 diabetes we generated a mouse model that over-expresses the resident ER chaperone and Unfolded Protein Response regulator GRP78 in pancreatic `-cells. Under the control of a rat insulin promoter GRP78 was overexpressed by approximately 8-fold in pancreatic `-cells and the mice had normal weight gain and metabolic physiology on a regular chow diet. We hypothesized that increased chaperone capacity would protect `-cells from dysfunction resulting from obesity and improve glucose homeostasis in mice on a chronic high fat diet (HFD). As expected, control transgene negative mice on a 20-week HFD developed obesity, glucose intolerance and insulin resistance. Remarkably, GRP78 transgenic mice tended to be leaner than their transgene negative littermates and were protected from glucose intol- erance, hyperinsulinemia and insulin resistance. Islets from transgenic mice were also protected from the hypertrophy characteristic of obesity-induced insulin resistance. Compared to transgene negative animals on a HFD, islet 1746-P area (mass) was reduced and was comparable to islet mass in regular chow Quantifi cation of Glucose-Stimulated Insulin Secretion Rate in fed mice. HFD feeding caused mild ER stress in islets and islets from GRP78 Normal, Pre-Diabetic, and Diabetic Cynomolgus Monkeys (Macaca transgenic mice were protected from ER stress. In summary, increased Fascicularis) Using a Graded Glucose Infusion chaperone capacity in pancreatic `-cells protects from the pathogenesis YI-XIN (JIM) WANG, PAUL B. HIGGINS, CHEN WANG, JIAN WANG, YUPENG of obesity-induced type 2 diabetes by improving pancreatic `-cell function, FANG, FENGLAI DU, BINGDI WANG, XIAOLI WANG, FRANCINE M. GREGOIRE, which ultimately improves whole body glucose homeostasis. BARBARA C. HANSEN, Taicang, China, Kannapolis, NC, Tampa, FL Supported by: CIHR (MOP-114992) Graded glucose infusion (GGI) procedures have been used to evaluate ` cell function independently of insulin sensitivity. Quantifi cation of insulin se- cretion rate (ISR) using the GGI provides insight into the trajectories of ` cell 1745-P Obesity

Metabolic Response Patterns to a Parenteral Hyperglycemic Chal- dysfunction in diabetes development and can be used to directly evaluate POSTERS lenge in Lean Healthy Non-Human Primates Resemble those in Lean the effects of therapeutic agents on ` cell function. ISR was determined by deconvolution of serum C-peptide concentrations measured in a 5 stage GGI Healthy Human Volunteers Integrated Physiology/ EFFIE TOZZO, LORI MIXSON, STACEY L. CONARELLO, BARBARA C. HANSEN, (glucose infusion rates of 2, 4, 8, 16, and 32mg/kg/min for 40mins each) in 6 CHAN R. BEALS, DAVID E. KELLEY, SUDHA S. SHANKAR, Rahway, NJ, Tampa, FL normal (6.0±0.2yrs, 7.6±0.7kg, fasting glucose: 60±4mg/dL), 14 prediabetic We have previously shown that the Graded Glucose Infusion (GGI) method (15.0±0.8yrs, 9.0±0.9kg, fasting glucose: 80±4mg/dL), and 4 untreated newly can successfully detect glucose-dependent insulin secretion (GDIS) re- diabetic (16.0±1.2yrs, 9.0±1.8kg, fasting glucose: 169±22mg/dL) cynomolgus sponses to a parenteral stepwise glycemic challenge (GC) in healthy human macaques. After correction for circulating glucose concentrations, group volunteers (HV). We tested the hypothesis that the GGI can detect GDIS in mean ISRs were compared by one-way ANOVA with Tukey’s post hoc com- healthy adult NHP, with response patterns resembling those in HV, by com- parisons. A signifi cant mean difference in ISR was found between the groups paring GDIS between NHP (N=6, weight: 10.7±1.0 kg, FPG: 62.4±5.6 mg/dL, (F=6.18, p=0.001). ISR was signifi cantly less in the diabetic group compared mean ± SD) and HV (N=8, BMI: 23.3±2.1 kg/m2, FPG: 86.9±7.3 mg/dL). After to the normal (p<0.01) and prediabetic groups (p<0.05). No signifi cant differ- an overnight fast, in response to glucose infusion rates (GIR) from 2 to 16 ences in ISR were noted between the normal and prediabetic groups (Figure

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A449 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

1). Observations are consistent with those in rhesus macaques and humans. 1748-P We conclude that the GGI with ISR calculation produced predictable mea- Signifi cant Early Changes During Time Course Assessment of Lipid sures of ` cell responsiveness in normal, prediabetic, and diabetic cyno- and Metabolic Profi le Following Bariatric Surgery molgus macaques and could be of utility in assessing the effi cacy of agents ROBERT V. CONSIDINE, LORI A. MIXSON, AIZZA HASSAN, ROSEMARIE JONES, targeting improvements in ` cell function. SAMER G. MATTAR, MANU CHAKRAVARTHY, CHAN R. BEALS, DAVID E. KELLEY, SUDHA S. SHANKAR, Indianapolis, IN, Rahway, NJ Studies of the acute metabolic improvements in Type 2 diabetes after Roux-en-Y gastric bypass surgery (RYGB) have largely focused on single time-points, with changes attributed primarily to altered gut hormone re- lease. Scant data exist on the time course of metabolic changes, especially lipid parameters, in the intermediate period between recovery from RYGB, but before signifi cant weight loss. We tested the hypothesis that the pattern of change in fasting and prandial glucose homeostasis and lipid parameters between 1-4 weeks post RYGB (POST) refl ect the interaction of multiple prevailing metabolic processes. Fasting parameters were measured at 1, 2 and 4 wks POST, and enteroinsular response to a meal (~350 kcal; 25.9g carbohydrate, 8.4g fat, 44g protein) at 2 and 4 wks POST, in 18 women with Type 2 diabetes (BMI 52.7±14.2 kg/m2, 44.0±10.1 yr; mean±SD). Compared to presurgery (PRE), at 1 wk there were signifi cant (p<0.01) decreases in fasting glucose (20%), insulin (47%), and C-peptide (32%), which were maintained at 2 and 4 wks. During meal test at 2 and 4 wks, a return to pre-meal values of glucose, insulin and C-peptide was observed at 120-minutes, in contrast to PRE excursions which remained well above pre-meal values at 120-minutes. Robust increases in circulating prandial PYY AUC (p<0.005) occurred at 2 (253%) and 4 wks (136%). Decreases (p<0.01) in triglycerides (11%), and HDL- C (23%) at 1 wk were maintained at 4 wks, with LDL-C signifi cantly reduced (14%) at 4 wks. The pattern of changes in fasting and prandial glucose ho- meostasis at 2 and 4 wks POST are consistent with improved enteroinsular axis function. Changes in PYY may be associated with earlier satiety, while 1747-P lipid profi le changes at 1 and 4 wks POST are consistent with caloric restric- Differences between Japanese and Caucasians in Insulin Sensitiv- tion, both of which may also lead to improved glucose homeostasis. Taken ity and Beta-Cell Function: Contribution of BMI together, our fi ndings suggest that multiple interactive processes may con- MITSURU OHSUGI, HARUHIKO TANAKA, MARIA PEDERSEN, JONAS B. MØLLER, tribute to the weight loss independent metabolic improvement after RYGB. RUNE V. OVERGAARD, JAN LYNGE, KATRINE ALMIND, NINA-MARIA VASCON- CELOS, PERNILLE POULSEN, CHARLOTTE KELLER, KOHJIRO UEKI, STEEN H. IN- 1749-P GWERSEN, BENTE K. PEDERSEN, TAKASHI KADOWAKI, Tokyo, Japan, Copenha- Pancreatic Fat Does Not Impair Beta-Cell Function in a Non-Dia- gen, Denmark, Bagsvaerd, Denmark betic Cohort The pathophysiology of type 2 diabetes (T2D) is perceived to be differ- BETTINA NOWOTNY, ROSHAN LIVINGSTONE, SABINE LINK, BIRGIT KLÜPPEL- ent in Japanese compared to Caucasians. Japanese have been reported HOLZ, PETER J. NOWOTNY, JONG-HEE HWANG, GIOVANNI PACINI, GUIDO GI- to have higher insulin sensitivity but less ability to compensate for insulin ANI, MICHAEL RODEN, Düsseldorf, Germany, Padova, Italy resistance by increased beta-cell function, although the supportive scien- Beta cell dysfunction contributes to the development of type 2 diabetes tifi c data are limited.We undertook a cross-sectional study in 120 Japanese on the basis of insulin resistance. Ectopic lipid storage has been shown to living in Japan and 150 Caucasians living in Denmark, with the objective be associated with insulin resistance of liver and muscle. Whether elevated to compare insulin sensitivity and beta-cell function, and to investigate the pancreatic fat contents are related to impaired beta cell function in humans role of demographic, genetic, biochemical and life-style related factors. remains unclear. Thus, this study analyses the association of pancreatic fat Participants were grouped according to their glucose tolerance: normal or content and beta cell function in a non-diabetic cohort. 75non-diabetic sub- impaired glucose tolerance (NGT, IGT) or T2D, with comparable distributions jects (43f/32m; mean age 58±11 years; body mass index (BMI) 26.0±3.8kg/m2) of high/low BMI according to regional obesity defi nitions. Insulin sensitiv- underwent a 75g oral glucose tolerance test (OGTT). Blood glucose, insulin ity and beta-cell function were estimated from oral glucose tolerance test and C-peptide levels were determined at various time points, and pancreatic data.Mean glucose profi les were similar in Japanese and Caucasian NGT, fat (median 6.5%, range 0.3-41.9%) content was measured using 1H MRS IGT and T2D participants respectively, whereas the insulin and C-peptide using a 3TPhilips scanner with stimulated echo acquisition mode (STEAM) responses were lower in Japanese. The Japanese participants were less sequence. Indices of beta cell function [Insulinogenic index (IGI, pmolINS/ insulin resistant as determined by HOMA-IR and Matsuda index, and had mmolGLUC), adaptation (AI) and disposition (DI) index (common units)] and in- lower beta-cell function as determined by HOMA-B, insulinogenic index and sulin sensitivity (OGIS, ml/min/m2) were calculated. Pancreatic fat contents insulin secretion ratio than Caucasians. Disposition indices, expressing be- ranged from 0.25 to 41.89 % (mean 9.31±9.26%), and means±SEM of beta ta-cell function relative to insulin resistance, were similar in Japanese and cell function were 215±36 for IGI, 0.45±0.01 for AI and 2.60±0.12 for DI, while Caucasians.BMI and other measures of body size such as weight and waist, insulin sensitivity (OGIS) was 444±8. Pancreatic fat correlated signifi cantly were major determinants for both insulin sensitivity and beta-cell function. with weight (r=0.35, p=0.002), BMI (r=0.37, p=0.0003) and waist circumfer- Accordingly, following BMI-adjustment, the differences between Japanese ence (r=0.30, p=0.005), which was stronger in females than in males. No and Caucasians were substantially reduced in all groups and were no longer correlations were found with fasting or 2h whole blood glucose and insulin statistical signifi cant in NGT and IGT participants.In conclusion, our results

Obesity or C-peptide levels throughout the OGTT. No index of beta cell function was

POSTERS confi rm that Japanese generally have higher insulin sensitivity and lower associated with pancreatic fat content, nor we observed a clear correlation beta-cell function than do Caucasians. The major part of these differences with OGIS (r=-0.18; p=0.08). Linear regression analyses with adjustment for can be explained by differences in body size (BMI).

Integrated Physiology/ sex, age and BMI did not improve the associations. In conclusion, pancreatic Supported by: Japan Sci. & Techn. and Danish Sci. Techn. Innov. Agencies; Novo fat content is associated with increased body weight and body fat mass, but Nordisk A/S cannot serve as a surrogate of impaired beta cell function in a non-diabetic population.

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A450 INTEGRATED PHYSIOLOGY—INSULINCATEGORY SECRETION IN VIVO

1750-P 1752-P Vietnamese Americans Demonstrate Markedly Impaired Insulin Se- Beta Cell-Specifi c Overexpression of the Intracellular Protein p8 cretion Across Glycemic Status Preserved Glucose Tolerance during High Fat Diet- or Insulitis-In- LAN CHI T. LUU, TIMOTHY ALLERTON, GABRIEL UWAIFO, ROBERT DUBIN, WIL- duced Beta Cell Damage In Vivo LIAM T. CEFALU, New Orleans, LA AMIR E. MEHANA, INGO H. PILZ, BIANCA DUFNER, CHRISTINA JÄGER, SAN- Vietnamese Americans (VNA) represent one of the fastest growing minor- DRA SOJKA, JOHANNES BAUMANN, MARCUS ALT, JOCHEN SEUFERT, GÜNTER ity populations in the United States with a 38% population growth since PÄTH, Freiburg, Germany 2000. Previous studies have shown that Asian Americans, as a group, are OBJECTIVE: p8 was initially described to protect exocrine acinar tissue at greater risk to develop Type 2 Diabetes (T2DM) than Caucasians. How- from infl ammatory damage. To evalute the protective potential in the en- ever, there is currently a dearth of research investigating the unique meta- docrine pancreas, we generated transgenic (tg) mice with beta cell-specifi c bolic risk factors that contribute to the development of T2DM in VNA. We p8 overexpression. Previously, we introduced the tg phenotype and demon- sought to evaluate insulin secretion (IS), insulin sensitivity (Si) and metabolic strated that upon 10 weeks high fat diet (HFD) tg and wild type (wt) mice fl exibility [(MF), the switch in substrate oxidation, i.e., utilization of fat to gain weight, develop insulin resistance and became glucose intolerant. carbohydrate] across glucose classifi cations of VNA [normal glucose toler- Importantly, tg mice preserved a signifi cantly enhanced 1st phase insulin ance (NGT), impaired glucose tolerance (IGT), and T2DM}. In vivo physiologic secretion. Here, we analysed beta cell mass in HFD-mice and investigated measures were conducted in 30 VNA subjects of mixed gender (15 NGT, 8 the effects insulitits and STZ-induced beta cell damage combined with HFD. IGT, and 7 T2DM), ages 18 - 69, and BMI of 18-40 kg/m2. Subjects completed RESULTS: After 10 weeks of HFD (60%) mice demonstrate signifi cant loss of a mixed meal tolerance at which time indirect calorimetry was performed beta cell mass. This loss was signifi cantly reduced in HFD-tg mice. We next pre and post meal to assess MF by measuring resting quotient (RQ). During analysed effects of 5 x 40 mg/kg STZ-induced insulitis. Unfasted blood glu- a separate visit, subjects underwent a frequently sampled intravenous glu- cose levels do not signifi cantly change. In contrast, glucose tolerance mea- cose tolerance test to calculate IS and Si. First phase IS was 43.0+18.0mu/L sured by fasted ipGTT dramatically decreases in wt mice (1st and 2nd phase) in NGT, 17.1+ 9.9mu/L in IGT, and 11.8+9.2 in T2DM (P < 0.0001 across while tg mice demonstrate levels similar to untreated tg controls. Next, we groups). Beta cell function was reduced(P<.03) across NGT 120+45.6 mU/ fed mice with normal diet or HFD and inject a single dose of 150 mg/kg STZ mM, IGT 73.2+26.6 mU/mM, and T2DM 73.6+46.7mU/mM. The disposition after 4 weeks resulting in acute diabetes (unfasted blood sugar above 400 index (DI) was positively correlated with 2-hour plasma glucose (r2=0.74, mg/dl). Within the next 4 weeks normal and HFD fed wt do not change their P<0.0001). At glucose values indicative of IGT (2hr value of 140 mg/dl), DI diabetic state while tg recover. Interestingly, recovering in normal-fed tg values were markedly impaired by 80%. Insulin resistance increased from mice was less pronounced (n.s.) than in HFD-fed tg mice, which signifi cantly 0.75+0.3 in NGT, 0.88+0.32 in IGT, to 2.0+2.1 in T2DM. After RQ was adjusted ameliorate their diabetic blood sugar levels down to values below 300 mg/ for Si the 6RQ values for NGT, IGT, and T2D were (0.075, 0.010, and 0.00) dl in weeks 3 and 4.CONCLUSION: p8 protects beta cell mass from damage respectively.In conclusions, Vietnamese Americans have markedly impaired by HFD-induced lipotoxicity and insulitis. p8 appears to be also regenerative. insulin secretion when already diagnosed as IGT with further progression To further evaluate its anti-infl ammatory and regenerative potential we are to T2DM. Vietnamese Americans have severe impairment in metabolic fl ex- currently investigating insulitis-induced lymphocyte infi ltration into islets ibility as they progress from NGT to T2DM. and beta cell proliferation after STZ-damage. Supported by: LSUHSC School of Medicine Supported by: Deutsche Forschungsgemeinschaft (DFG) Grant PA1663-2-1 (G.P.)

1751-P 1753-P Decreased Insulin Responses to IV Glucose and Tolbutamide in Sub- GIP Elicits Additive But Not Synergistic Effects of Glucose Lower- jects With Impaired Fasting Glucose: Clues to Cellular Mechanisms ing Over Sub-Optimal Doses of GLP-1 for Ƶ-Cell Failure? ANNE M. CHRISTIANSEN, SARAH WILL, OVER CABRERA, DAVID TESS, XIANG- SEDA SUVAG, KRISTINA UTZSCHNEIDER, LORENA A. WRIGHT, BRIAN FISH, STE- PING LI, SNEHA NARAYANAN, SHENA PATEL, BHAVNA PARATALA, I-CHEN YU, VEN KAHN, Seattle, WA JESSIE DOW, MYLENE PERREAULT, MARGARET JACKSON, JULI JONES, Cam- In subjects with impaired fasting glucose (IFG), the 1st phase insulin re- bridge, MA, Groton, CT sponse to IV glucose is impaired. Whether the 2nd phase insulin response to Glucose-dependent insulinotropic peptide (GIP) and glucagon-like pep- glucose and the response to the K-ATP channel blocker tolbutamide are simi- tide-1 (GLP-1) are both insulin secretagogues and metabolic modulators but larly impaired, is not known. To address this, 233 subjects with normal fast- have differing biological effects. The purpose of the current studies were ing glucose (NFG) (n=156, 53M/103F, age 52±0.8 y, BMI 26±0.3 kg/m2, fast- to examine if GIP can elicit an additive or synergistic effect over a GLP-1 ing glucose 92.8±0.4 mg/dl) or IFG (n=77, 45M/32F, age 53±1.1 y, BMI 28±0.5 agonist (Exendin-4 (EX4)) for glucose lowering in vivo in mice and ex vivo in kg/m2, fasting glucose 106.3±0.6 mg/dl, x±sem) underwent an IV glucose human islet cultures. In all of these studies we choose doses based on PK/ tolerance test with tolbutamide given at 20 min. First phase insulin response PD modeling in order to address combination agonism. To look at the effects was calculated as the incremental area under the curve (iAUC) insulin from of GIP and GLP-1 in vivo, lean, fasted C57/B6 mice were injected with varying 0-10 min; 2nd phase response as iAUC insulin/iAUC glucose from 10-19 min doses of GIP (sc) or EX4 (ip) prior to an ip glucose tolerance test. Based on the and tolbutamide response as iAUC insulin/iAUC glucose from 19-30 min. modeled dose effect relationship, the ED50 and ED75 of EX4 and the ED90 Insulin sensitivity (SI), determined by the minimal model, was lower in IFG of GIP were chosen for combination studies. We found that when EX4 was subjects (median [IQR]: 2.5 [2.1] vs 3.8 [3.0] x 10-4 min-1/(μU/mL), p<0.001). As administered at the ED50, the addition of ED90 GIP could demonstrate an ad- SI determines the magnitude of insulin responses, analyses were adjusted ditive effect on glucose lowering. However, a combination of ED75 EX4 and st for SI. Subjects with IFG had lower 1 phase (33.7 [31.9] vs 52.6 [39.5] μU/ ED90 of GIP could not provide greater than maximal GLP-1 glucose lowering. mL; p<0.001), 2nd phase (0.17 [0.17] vs 0.22 [0.20] μU/ml per mg/dl, p<0.001) To further understand the human biology of GIP and GLP-1, we performed ex and tolbutamide insulin responses (0.69 [0.65] vs 0.75 [0.69] μU/ml per mg/ vivo experiments in human islet cultures. Islets were isolated from pancreas dl, p=0.004). The 1st phase and tolbutamide responses were strongly corre- recovered from normal human subjects and cultured in CMARL base media lated in all subjects (r2=0.57; p<0.001) and in NFG (r2=0.69; p<0.001) and IFG for 3-7 days. Islets were treated acutely with a maximal concentration of 2 nd

(r =0.56; p<0.001). In contrast, the 2 phase insulin response did not corre- EX4 with and without a maximal concentration of GIP at varying concen- Obesity st late with either 1 phase or tolbutamide responses in any group. Thus, the `- trations of glucose and insulin secretion was measured. Similar to the in POSTERS cell defect in IFG affects both glucose- and tolbutamide-induced responses. vivo studies, a combination of GIP and EX4 did not demonstrate synergistic st

The strong relationship between the 1 phase and tolbutamide responses effects. Taken together, these data help elucidate the complex combinatory Integrated Physiology/ suggests that these share a common mechanism through the K-ATP channel, effects of GIP and GLP-1 on metabolism and suggest that these effects may while the lack of a relationship between these and the 2nd phase suggests be similar in both in vivo mouse and ex vivo human model systems. that an alternative or additional mechanism underlies this latter response.

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A451 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1754-P INTEGRATED PHYSIOLOGY—LIVER Prolyl Endopeptidase (Prep) Regulates Insulin Secretion and Glu- cose Metabolism in Mice JUNG DAE KIM, GIUSEPPE D’AGOSTINO, JIN KWON JEONG, RICHARD KIBBEY, Guided Audio Tour: Physiological Studies of Hepatic and Lipid Metabolism OWEN CHAN, SABRINA DIANO, New Haven, CT (Posters 1756-P to 1763-P), see page 15. Prolyl endopeptidase (Prep)is a highly conserved serine protease impli- cated in regulation of neuronal functions, including hypothalamic processes. & 1756-P However, Prep is still the subject of studies attempting to elucidate its Liver Glycogen Loading Amplifi es Counterregulatory Hormone Re- physiological role. Here we report that Prep expression in the hypothala- lease and the Hepatic Response to Hypoglycemia mus is predominantly presented in the ventromedial nucleus (VMN) which JASON J. WINNICK, GUILLAUME KRAFT, BEN FARMER, MARGARET LAUTZ, is involved in the regulation of energy and glucose metabolism. Thus, we ERIC ALLEN, ALAN D. CHERRINGTON, Nashville, TN gt/gt analyzed glucose metabolism in Prep null (Prep ) mice compared to wild The purpose of this study was to determine whether the hepatic glycogen gt/gt type controls (WT). Although Prep mice fed on a standard chow diet level infl uences the response to hypoglycemia. During the fi rst 4h of each showed no signifi cant difference in body weights, body composition and study animals received somatostatin and basal intraportal infusions of both food intake compared to wild types, glucose tolerance test (GTT) showed insulin and glucagon. Plasma glucose was doubled by glucose infusion into gt/gt that Prep mice had a marked increase of blood glucose level after 1g/kg a peripheral vein and liver glycogen loading was either enhanced (L+; n=5; intraperitoneal glucose challenge and circulation insulin measurement dur- estimated liver glycogen content of ~82±7 mg/g) due to glucokinase translo- gt/gt ing GTT showed signifi cant lower insulin levels in Prep mice compared to cation brought about by intraportal fructose infusion, or not (saline infusion; gt/gt WT, indication that Prep mice have a decreased insulin secretion by the L-; n=5; ~54±4 mg/g). The 4h glycogen loading period was followed by a 2h pancreas. Consistent with decreased glucose-induced insulin secretion in control period during which basal replacement of hormones was continued gt/gt gt/gt Prep mice, Prep mice have shown less phosphorylation level of insulin but fructose infusion was stopped. In the fi nal 2h, the somatostatin and receptor by glucose challenge in VMN compared to WT. In agreement with glucagon infusions were stopped and plasma insulin levels were increased these fi ndings, the hyperinsulinemic-euglycemic clamp study has also shown 16-fold in both groups. During the ensuing 2h hypoglycemic period the ar- gt/gt that the Prep mice have an impaired insulin secretory response to a hy- terial plasma glucose was clamped at 47±1 and 47±2 mg/dL in L+ and L-, gt/gt perglycemic challenge. Moreover, the Prep mice require less glucose to respectively. The hepatic sinusoidal insulin levels were 427±18 and 402±34 maintain the hyperglycemic plateau which is consistent with the lower insu- μU/mL in L+ and L-, respectively. However, the amount of glucose required lin response. However, when in vitro pancreatic insulin release was studied, to maintain the hypoglycemic clamp was signifi cantly less in L+ compared gt/gt gt/gt no differences were found between WT and Prep mice. Finally, Prep to L- (1.8±0.2 and 4.7±0.2 mg/kg/min, respectively; p<0.001), as a result of mice showed increased sensitivity to insulin compared to WT controls by greater net hepatic glucose output in L+ than L- (5.1±0.6 and 1.5±0.2 mg/kg/ ITT, indicating that insulin-mediated glucose uptake is not impaired rather is min, respectively; p<0.001). Plasma levels of both glucagon and epinephrine enhanced in the absence of PREP. Altogether these data reveal a previously were basal and not different between L+ and L- prior to the hypoglycemic unsuspected role of PREP in hypothalamic control of glucose metabolism. challenge. However, the average hepatic sinusoidal plasma glucagon level during the hypoglycemic period was higher in L+ compared to L- (113±30 vs 1755-P 63±81 pg/mL, respectively; p=0.07) as was the average arterial plasma epi- Poor Compensatory Insulin Secretion against Insulin Resistance nephrine level (1736±43 and 942±274 pg/mL, respectively; p=0.08). These during Oral Glucose Tolerance Tests in Korean Subjects With Im- data suggest that the increase in hepatic glucose output was secondary paired Glucose Tolerance or Type 2 Diabetes Mellitus to augmented epinephrine and glucagon secretion and that the brain can TAE JUNG OH, KYONG SOO PARK, SEONG YEON KIM, HONG KYU LEE, EUN KY detect the hepatic glycogen content. KIM, MIN KYEONG KIM, YOUNG MIN CHO, Seoul, Republic of Korea It has been known that the amount of insulin secretion during oral glu- & 1757-P cose tolerance tests (OGTTs) is much lower in Asian subjects with impaired PEPCK-C is Required for Normal Mitochondrial Substrate Shuttling glucose tolerance (IGT) or type 2 diabetes than in Caucasian counterparts. JOAO DUARTE, JOHN G. JONES, SHAWN C. BURGESS, Dallas, TX, Coimbra, Por- Oral disposition index (DI), which is the product of 1/fasting plasma insulin tugal and insulinogenic index during the standard 75 g OGTTs, refl ects the pan- Cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C) catalyzes the creatic beta cell function corrected for insulin sensitivity. In this study, we conversion of oxaloacetate (OAA) to phosphenolpyruvate and has long been addressed the beta cell function in Korean subjects with normal glucose associated with the transcriptional regulation of gluconeogenesis. We’ve tolerance (NGT), IGT, or type 2 diabetes in terms of oral DI. A total of 269 demonstrated that this pathway is also critical for the regulation of he- subjects (93 men and 176 women) aged 30-80 years were recruited by local patic energy metabolism and the tricarboxylic (TCA) cycle. The importance advertisement. Standard 75g oral glucose tolerance tests were performed. of PEPCK in non-gluconeogenic pathways is underscored in PEPCK-C liver Plasma glucose and insulin levels during OGTTs were measured. Oral DI, in- knockout (LKO) mice which have a severely impaired hepatic TCA cycle, sulinogenic index, and Matsuda index were calculated. We analyzed three fasting hepatic steatosis and markedly higher concentrations of some TCA groups according to degrees of glucose tolerance (183 NGT, 57 IGT, and 29 cycle intermediates, including a 10-fold elevation in liver malate. To further type 2 diabetes subjects). The oral DI was remarkably reduced in type 2 investigate the role of PEPCK-C in the regulation of lipid metabolism we diabetic patients (0.6±3.5 in type 2 diabetes vs. 1.5±1.3 in IGT vs. 2.9±3.1 in examined lipogenic fl ux in mice with varying levels of PEPCK-C expression NGT, P < 0.001). Interestingly, the regression curves for IGT and type 2 dia- and found that PEPCK-C has a robust and positive effect on lipid synthesis. betes (Fig 1) were fl at rather than hyperbolic, indicating poor compensatory We hypothesized that this occurs through a metabolic mechanism involving insulin secretion to overcome insulin resistance. There was virtually absent PEPCK-C as a regulator of cataplerotic fl ux through mitochondrial substrate compensatory insulin secretion to overcome prevailing insulin resistance in shuttles. To investigate this possibility, livers were excised from control and Korean subjects with IGT or type 2 diabetes, which may be the unique and PEPCK-C LKO animals and mitochondrial and cytosolic compartments were predominant pathophysiology of type 2 diabetes in this population. analyzed by 1H NMR metabolomic analysis. PEPCK-C LKO mice had had a

Obesity substantial accumulation of aspartate, malate, glutamate and glutamine in POSTERS the cytosol suggesting a blunted malate-aspartate shuttle. Gene expression analysis revealed reduced expression of the tricarboxylic acid transporter

Integrated Physiology/ but otherwise normal expression of enzymes of the shuttle system. We sug- gest that PEPCK-C loss of function suppresses lipid synthesis by impairing the mitochondrial substrate shuttle system, including anaplerotic infl ux via the malate-aspartate shuttle and cataplerotic export of citrate, a required pathway for lipogenesis. Supported by: NIH (RO1DK078184)

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A452 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

for age (49±2 vs 46±2 yrs), gender (85 vs 68% women) and PNPLA3 genotype & 1758-P (CC in 71 vs. 63%). Needle biopsies of subcutaneous AT were taken for qPCR Transaldolase Exchange is the Primary Mechanism for Asymmetric gene expression studies.AT expression of MCP1, TNF_, and the macrophage Enrichment of Glucose from 13C-triose Phosphate Precursors marker CD68, were comparable between the GG and CC groups. The anti- AMAN RAJPAL, JOHN JONES, FILIPA CARVALHO, ANA R. SIMOES, SIMMI DUBE, infl ammatory cytokine adiponectin (1.11 vs. 1.00, p<0.05 for GG vs CC) and CRISTINA BAROSA, ANANDA BASU, ROBERT A. RIZZA, RITA BASU, Rochester, transcription factor Twist1 (1.21 vs. 1.00, p=0.1) tended to be upregulated MN, Coimbra, Portugal in GG group (genetic NAFLD). In obese vs. non-obese NAFLD, MCP1 (1.2 vs. Transaldolase exchange (TA) contributes to plasma glucose enrichment 1.0. p<0.05), TNF_ (1.42 vs. 1.00, p<0.05), CD68 (1.3 vs 1.0, p<0.1) were up- from gluconeogenic tracers resulting in overestimation of gluconeogenesis. regulated, while expression of adiponectin (0.85 vs. 1.00, p<0.05) and Twist1 TA was recently revealed by reduced enrichment of carbon 3 relative to car- (0.78 vs. 1.00, p<0.02) was signifi cantly downregulated. Conclusions: Sub- bon 4 (C3/C4 < 1.0) of glucose from [1-13C] acetate infusion. A key untested jects who have genetic NAFLD due the PNPLA3 I148M variant lack AT infl am- assumption is that triose phosphate isomerase (TPI) exchange is complete. mation in contrast to those with obese NAFLD. Incomplete TPI exchange would contribute to C3/C4 < 1.0 because [1-13C] 13 glyceraldehyde-3-phosphate is initially labelled from [1- C] acetate infusion & 1760-P (Figure 1 below). If dihydroxyacetone phosphate was initially labelled from Impact of African-American Ethnicity and Insulin Resistance in the [U-13C] glycerol, incomplete TPI exchange would preferentially enrich glu- Development of NAFLD cose C3, resulting in C3/C4 > 1.0. However, if TPI exchange was complete ROMINA LOMONACO, CAROLINA ORTIZ-LOPEZ, BEVERLY ORSAK, JOAN FINCH, and TA was active, then C3/C4 would be < 1.0, as with [1-13C] acetate.[U-13C] JEAN HARDIES, NORMA DIAZ, FERMIN TIO, KENNETH CUSI, Gainesville, FL, San glycerol was infused in 5 subjects with mean age of 60 ± 5 yrs, BMI of 31 ± Antonio, TX 3 kg/m2 and FFM of 57 ± 5 kg. Plasma glucose 13C-enrichment was measured The role of African-American (AA) ethnicity and metabolic factors associ- by 13C NMR following an overnight fast and during a 0.35 mU/KgFFM/min ated with the development of nonalcoholic fatty liver disease (NAFLD) has insulin infusion. Somatostatin and glucagon were also infused during the never been well characterized in this population. To this end, we studied clamp to ensure comparable and equal portal concentrations in both groups. 300 patients (M/F=59/41%; age=50±1, AA=8%, Hispanic [Hisp]=60%, Cau- At baseline, carbon 3 and 4 excess 13C-enrichments were 0.34 ± 0.04 % and casians [Cauc]=32%, BMI=32.6±0.2kg/m2) in whom we measured: 1) liver 0.36 ± 0.04 %, respectively, resulting in C3/C4 of 0.93 ± 0.03. During clamp fat (%LFAT) by MRS; 2) total body fat (TBF) by DXA; 3) liver and muscle (Rd) these values were 0.27 ± 0.07 % and 0.31 ± 0.09 %, respectively, resulting in insulin sensitivity (insulin clamp with 3-3H glucose); 4) hepatic insulin resis- C3/C4 of 0.90 ± 0.04. These data suggest that the asymmetrical enrichment tance (IR; HIRi=EGP x fasting insulin [FPI]); 5) adipose tissue IR (AdipoIRi= of glucose from labeled triose-phosphate precursors is primarily due to TA. fasting FFA x FPI); 6) liver histology (biopsy; n=185). Clinical characteristics were similar among all ethnic groups. Patients with NAFLD (n=243) vs. no- NAFLD (n=57) were characterized by a higher prevalence of MetS/T2DM and for being more IR. Hisp and Cauc pts with NAFLD had a similar prevalence of obesity/MetS, adiposity (BMI/TBF), %LFAT, hepatic/muscle IR and se- verity of NASH on histology, so were grouped together (Hisp/Cauc). In AA, mean plasma AST/ALT were not signifi cantly increased in contrast to being elevated in Hisp/Cauc with NAFLD. AA compared to Hisp/Cauc with NAFLD had lower LFAT (16.0±2.0% vs. 23.6±0.8%, p=0.01) and more benign liver histology, with a trend towards less steatohepatitis (NAFLD activity scores= 3.7±0.4 vs. 4.2±0.1) and fi brosis (0.4±0.2 vs. 0.9±0.1, p=0.07), respectively. When examining the role of insulin resistance, AdipoIRi was less severe in AA vs. Hisp (6±1 vs. 9±1 mmol/L · μU/mL, p=0.03) with a similar trend vs. Cauc (7±1 mmol/L · μU/mL). HIRi also showed a trend to be less severe in AA vs. Hisp/Cauc (16±2 vs. 21±1 mg/kg−1 · min−1 · μU/mL, p=0.29), but muscle (Rd) was not different. LFAT correlated strongly with AdipoIRi in AA (r=0.59, p<0.01) but not in Hisp/Cauc. Conclusion: Patients of AA ancestry with NA- FLD have less severe hepatic steatosis and liver histological disease than Hisp/Cauc, which may be explained, at least in part, to less severe adipose tissue insulin resistance. Supported by: Burroughs Wellcome Fund

Supported by: DK29953 & 1761-P Impact of Glucose Tolerance Status on Insulin Sensitivity and Dis- & ease Severity in Patients With NAFLD 1759-P CAROLINA ORTIZ-LOPEZ, ROMINA LOMONACO, BEVERLY ORSAK, JOAN FINCH, Adipose Tissue is Infl amed in NAFLD Due to Obesity but not in JEAN HARDIES, FERMIN TIO, KENNETH CUSI, San Antonio, TX, Gainesville, FL NAFLD Due to Genetic Variation in PNPLA3 The metabolic factors contributing to nonalcoholic fatty liver disease (NA- SUSANNA LALLUKKA, KSENIA SEVASTIANOVA, ELINA M. PETÄJÄ, JULIA PERT- FLD), and its more severe form with steatohepatitis (NASH), in prediabetes TILÄ, PERTTU ARKKILA, MARJU ORHO-MELANDER, NINA LUNDBOM, ANTTI (IFG, IGT) or T2DM are unclear. To examine this we recruited 282 middle-aged HAKKARAINEN, VESA M. OLKKONEN, HANNELE YKI-JÄRVINEN, Helsinki, Fin- pts diagnosed with NAFLD by MRI and spectroscopy (MRS; ethn: Hisp=57%/ land, Malmö, Sweden Cauc=31%/AA=8%/other=4%) divided by an OGTT into normal controls (n=18 The rs738409 C>G SNP in the PNPLA3 (adiponutrin) gene leads to a mis- NGT), IFG (n=27), IGT (n=78), and T2DM (n=125). They were well-matched for sense mutation (I148M). Over 20 studies have shown this variant to markedly

BMI: 34.3±1.0 vs. 33.8±0.6 vs. 33.8±0.5 vs. 33.5±1.2 kg/m2; total body fat Obesity increase liver fat [“genetic non-alcoholic fatty liver disease (NAFLD)”] and (TBF by DXA): 33±2 vs. 34±1 vs. 33±1 vs. 34±2%, respectively (all NS). We POSTERS risk of NASH but not insulin resistance. In contrast, NAFLD due to obesity is measured: 1) liver/muscle (Rd) insulin sensitivity (insulin clamp with 3-3H associated with insulin resistance and adipose tissue (AT) infl ammation. We

glucose) and hepatic insulin resistance (IR) (Hep-IR index [HIRi]= EGP x fast- Integrated Physiology/ compared groups with similar increases in liver fat due to either obesity or ing plasma insulin [FPI]); 2) adipose tissue IR (Adipo-IR index [AdipoIRi]= the I148M variant to establish whether lack of systemic insulin resistance in fasting FFA x FPI); 3) liver histology (biopsy; n=183). Pts with NAFLD had sig- genetic NAFLD is accompanied by lack AT infl ammation.We studied 82 sub- nifi cant liver/adipose tissue/muscle IR. Despite similar BMI/TBF, T2DM had jects with either the GG (n=27) or the CC (n=55) genotype at rs738409. Liver worse IR at all sites (liver/adipose tissue/muscle; all p<0.01) compared to fat (1H-MRS) was 2.1-fold increased in GG vs CC (11±2 vs 5±1%, p<0.001) NGT and IFG. IFG had similar insulin sensitivity compared to NGT, while IGT but fasting insulins similar (9±1 vs 9±1 mU/L, NS). Of the 82 subjects, 41 had intermediate results between T2DM and NGT/IFG. Liver AST/ALT were were obese (BMI >30 kg/m²) and 41 non-obese. Liver fat (10±1 vs. 5±1%, not signifi cantly different between groups, but liver fat by MRS was higher p=0.007) was 1.9-fold and fasting insulin (11±1 vs. 7±1 mU/L, p<0.001) 1.6- in IGT/T2DM vs. NGT (24±1% vs. 17±3%, p<0.05). Having IGT or T2DM was fold increased in obese vs. non-obese. The GG and CC groups were matched associated with more severe liver histology compared to NGT or IFG (NAFLD for age (49±3 vs 47±2 yrs), gender (74 vs 78% women) and BMI (31±1 vs 31±1 activity score: 4.3±0.2 vs. 3.5±0.3, fi brosis: 1.0±0.1 vs. 0.3±0.1, respectively; kg/m²). The obese (36±1) and non-obese (26±1 kg/m²) groups were matched

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A453 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

both p<0.05). Hyperglycemia (2h OGTT glucose) correlated the strongest Gene Gene DNA DNA DNA DNA with parameters of adipose tissue IR (AdipoIRi, insulin or 2h-OGTT suppres- expression, expression, methylation, methylation, methylation, methylation, sion of FFA; r=0.35-0.41, p<0.0001) and more weakly with necroinfl ammation liver liver liver (%) liver (%) sperm (%) sperm (%) or fi brosis (r=0.22-0.24, p<0.01), but not with steatosis. Plasma infl ammatory Control IUGR-F2 Control IUGR-F2 Control IUGR_F1 markers were not different between groups. Conclusion: Hyperglycemia in Fasn 1 +/- 0.20 0.4 +/- 0.12* 3.2 +/- 0.30 3.0 +/- 0.13 n.d. n.d. NAFLD may play a signifi cant role in the more severe IR and liver histology of obese T2DM vs. NGT or IFG pts, independent of adiposity. Acc1 1 +/- 0.19 0.6 +/- 0.10* n.d. n.d. n.d. n.d. Supported by: Burroughs Wellcome Fund Acc2 1 +/- 0.20 0.6 +/- 0.11 n.d. n.d. n.d. n.d. Scd1 1 +/- 0.11 0.4 +/- 0.08* n.d. n.d. n.d. n.d. & 1762-P Srebf1 1 +/- 0.25 0.3 +/- 0.04* 5.6 +/- 0.30 5.4 +/- 0.35 n.d. n.d. Elevated Lipogenesis in NAFLD is Associated With Lower Hepatic Lxra 1 +/- 0.07 0.7 +/- 0.05* 9.1 +/- 0.47 7.2 +/- 0.29* 6.1 +/- 0.54 4.8 +/- 0.27* Export of Plasma Free Fatty Acids Secreted in VLDL-TG JENNIFER E. LAMBERT, JOSEPH J. LEE, YELENA HOVHANNISYAN, MARESSA J. Supported by: BFU2008-3759/BFI (MICINN, Spain), Fundacion2000, Spain VALDEZ, DORA L. BRADFORD, MARIA A. RAMOS-ROMAN, JEFFREY D. BROWN- ING, ELIZABETH J. PARKS, Dallas, TX De novo lipogenesis (DNL) contributes signifi cantly to excess intrahepatic triglyceride (IHTG) in Caucasians, but the infl uence of DNL in other ethnic Guided Audio Tour: Liver Metabolism—Animal Studies (Posters 1764-P to groups is unknown. Here, we quantifi ed the fatty acid (FA) sources used for 1771-P), see page 15. hepatic-TG in Hispanics (Hisp) and African Americans (AA) with elevated IHTG (liver fat = 18 ± 4%; HighLF; 9 Hisp, 3 AA) and normal liver fat (liver & 1764-P fat = 3 ± 2%; LowLF; 3 Hisp, 7 AA; P<0.001) matched for age and body fat. Impact of Patatin-Like Phospholipase Domain-Containing 3 Expres- Multiple stable isotopes were fed and infused to measure contribution of sion on Non Alchoholic Fatty Liver Disease Associated Hepatic In- DNL, dietary, and plasma FFA fl ux to liver lipid synthesis. Subjects consumed sulin Resistance a standardized, high-fat (40g) evening meal, and then fasted for the next 18h. NAOKI KUMASHIRO, ROMY KURSAWE, DANIEL F. VATNER, SACHIN K. MAJUM- IHTG was measured by MRS and VLDL-TG FA sources by GC/MS. After the DAR, FITSUM GUEBRE-EGZIABHER, JENNIFER L. CANTLEY, IOANA FAT, DEREK 18h fast, in both the HighLF and LowLF groups, the labeling strategy resulted M. ERION, DONGYAN ZHANG, TORU YOSHIMURA, PRASAD MANCHEM, SAN- in identifi cation of the same quantity of total FA sources (65 ± 13%; P=0.97) JAY BHANOT, GLENN S. GERHARD, KITT F. PETERSEN, GARY W. CLINE, VARMAN and a similar proportion of VLDL-TG derived from the previous evening meal T. SAMUEL, GERALD I. SHULMAN, New Haven, CT, Carlsbad, CA, Danville, PA, (5.7 ± 2.4% HighLF vs. 4.9 ± 1.9% LowLF; P=0.41). By contrast, in subjects West Haven, CT with HighLF, the proportion of VLDL-TG arising from DNL was signifi cantly Genome wide array studies have associated nonalcoholic fatty liver higher (22 ± 10% vs. 10 ± 8%; P=0.008) while that from the plasma FFA pool disease (NAFLD) with polymorphisms in the Patatin-like Phospholipase Do- was lower (38 ± 15% vs. 51 ± 13%; P=0.04) compared to LowLF. There was main-containing 3 (PNPLA3) gene. However, the mechanisms whereby PN- no difference in plasma FFA turnover rates (P=0.33) between groups, but PLA3 regulates hepatic lipid metabolism are unclear. We measured hepatic the plasma FFA concentration was higher in HighLF (0.73 ± 0.1 vs. 0.53 ± 0.1 PNPLA3 mRNA expression from human liver biopsy samples and related mmol/L; P=0.002), also suggesting a defi cit in FFA utilization. No signifi cant this with hepatic lipid content and HOMA-IR (n=28). Hepatic PNPLA3 mRNA effects of race were observed (P>0.05) for any FA source. Even though the expression was strongly correlated with hepatic triglyceride, diacylglyc- incidence of NAFLD has been found to be high in Hisp and low in AA, these erol (DAG) content and HOMA-IR [R=0.62; P=0.0006, R=0.63; P=0.0004, and data suggest that when it occurs in either population, it will be associated R=0.47; P=0.01, respectively] suggesting that increased PNPLA3 expression with high DNL. In summary, the data support a dominant role for DNL in etiol- favors hepatic lipid accumulation. To test this hypothesis, we treated nor- ogy of IHTG, irrespective of race. Further, elevated DNL is associated with mal SD rats fed with high-fat diet with specifi c antisense oligonucleotides a reduction in the hepatic export of plasma FFA via VLDL secretion, which (ASO’s) to decrease PNPLA3 expression in liver and adipose tissue. PNPLA3 would lead to an exacerbation of NAFLD. expression was decreased by ~50% in the fasted and ~90% in the refed Supported by: NIH (5RL1DK081187), CTSA (UL1RR024982), TORS (UL1DE019584) livers and epididymal adipose tissue by PNPLA3 ASO. Body weight and epididymal adipose tissue weight were not changed. To assess the effects & 1763-P of PNPLA3 knockdown on liver and muscle insulin sensitivity we performed Prenatal Undernutrition in Male Mice Programs Expression of Lipo- hyperinsulinemic [4 mU/(kg-min)]-euglycemic clamps. Knockdown of PNPLA3 genic Genes in the Second-Generation Offspring: Potential Role of expression protected rats from lipid-induced hepatic insulin resistance as Epigenetic Mechanisms refl ected by an approximately twofold greater suppression of endogenous DEBORA MARTINEZ, SILVIA RIBO, CHRISTIAN DAVIAUD, VINCENT W. BLOKS, glucose production [48±4 vs. 81±8% (P=0.002)] during the clamp without any THAIS PENTINAT, JUDITH CEBRIA, RUBEN DIAZ, TORSTEN PLOSCH, JORG TOST, changes in insulin-stimulated peripheral glucose uptake. PNPLA3 ASO treat- JOSEP C. JIMENEZ-CHILLARON, Esplugues, Barcelona, Spain, Evry, France, Gron- ment did not change hepatic triglyceride content [11.2±1.1 vs. 8.9±1.0 mg/ ingen, The Netherlands (g-liver), P=0.17] but did reduce hepatic DAG content by ~40% [936±134 vs. In utero undernutrition increases the diabetes risk later in life. In addi- 535±39 nmol/g-liver, P=0.02]. Taken together these data suggest that PN- tion, the diabetes risk may be transmitted to subsequent generations, in PLA3 may function primarily in a lipogenic capacity and inhibition of PNPLA3 part through epigenetic mechanisms. We have developed a mouse model of activity may be a novel therapeutic approach for the treatment of NAFLD intrauterine growth restriction (IUGR) by 50% maternal caloric deprivation and hepatic insulin resistance. during gestation. Male offspring (IUGR-F1) developed glucose intolerance and altered hepatic lipid metabolism with ageing.Here we aimed to explore & 1765-P whether prenatal undernutrition infl uences expression of lipogenic genes Pancreas-Specifi c PANDER Over Expression Induces Hepatic Insu- through epigenetic modifi cations, primarily DNA methylation, in the second- lin Resistance

Obesity generation offspring.We determined that, in livers from IUGR-F2 mice, ex- GRACE C. DOUGAN, SHARI L. MOAK, MARK G. ATHANASON, WHITNEY A. POSTERS pression of lipogenic genes (Fasn, Acc1, Acc2 and Scd1) was reduced while DANSE, MELANIE N. KUEHL, CLAUDIA E. ROBERT-COOPERMAN, BRANT R. BUR- expression of genes that regulate lipid oxidation (Cpt1a, Cpt1b and Cpt2) KHARDT, Tampa, FL

Integrated Physiology/ was unaltered (Table 1; *P< 0.05). Srebf1, which regulates expression of all PANcreatic-DERived Factor (PANDER) is a recently characterized secreted previously described lipogenic genes, was also reduced in IUGR-F2 mice. protein with hormone-like functions linked to glucose homeostasis that is Likewise, Lxra, which in turn regulates Srebf1, was signifi cantly down-regu- strongly expressed from the endocrine pancreas. The PANDER knockout lated as well. DNA methylation of the regulatory regions of Fasn and Srebf1 demonstrated increased hepatic insulin sensitivity with subsequent de- was normal in IUGR-F2 mice. In contrast, promoter DNA methylation of Lxra creased hepatic glucose production (HGP) during hyperinsulinemic-euglyce- was signifi cantly altered. Strikingly, this epigenetic signature was already mic clamp (HEC) conditions. To further characterize the function of PANDER present in sperm samples from their progenitors, IUGR-F1.In conclusion, this in-vivo, a transgenic mouse model was created that overexpressed PANDER study supports the hypothesis that prenatal nutrition may induce epigenetic (PANTG) specifi cally from the endocrine pancreas as has been described changes, presumably in the male germ cells, that remain stable during game- earlier. To examine the impact of PANDER expression on the liver, HEC stud- togenesis and are inherited into the following generation offspring. ies were performed on 3-4 month old PANTG and wild-type (WT) mice fed either a normal (16.7% kcal fat) or high-fat (45% kcal fat) diet for 10 weeks.

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A454 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

To measure hepatic insulin sensitivity and hepatic glucose production, we ganelle for lipid degradation these analyses identifi ed that in alb-SREBP-1a utilized HEC studies. HEC experiments revealed similar glucose infusion rate mice 10-fold more peroxisomal proteins than mitochondrial proteins are reg- or rate of disposal in PANTG versus WT mice on normal or high fat diet (n=8 ulated. Accordingly, the lipid profi le in serum and liver biopsies revealed that per PANTG and WT). This is consistent with our previous insulin tolerance alb-SREBP-1a6P mice were protected against dyslipidemia. Correspondingly tests and indicates peripheral insulin resistance is not present in the PANTG only alb-SREBP-1a developed an insulin resistance. These data highlight the model. However, there is a signifi cant increase in HGP in PANTG as com- role of phosphorylation of SREBP-1a as a regulation principle and further pared to WT mice under clamp conditions at low dose insulin infusion (30.2 stress the role of peroxisomes in hepatic lipid accumulation. ± 3.0 mg/kg/min vs. 17.9 ± 2.6 mg/kg/min, respectively; P<0.05). Increased HGP was also observed in PANTG as compared to WT mice on high fat diet & 1768-P (30.5 ± 3.6 mg/kg/min versus 21.9 ± 1.4 mg/kg/min, respectively; P<0.05). Hepatic DEPTOR Expression Improves Systemic Insulin Resistance Furthermore, percentage of suppression of HGP under clamp conditions is YUMIKO CHIBA, KENJI UNO, HIDEKI KATAGIRI, Sendai, Miyagi, Japan signifi cantly higher in WT as compared to PANTG mice as found under nor- DEPTOR is an inhibitor of the mammalian target of rapamycin (mTOR) sig- mal chow (80.4% ± 3.2% vs. 66.9% ± 3.4%, respectively; P<0.05) or high fat naling pathway which contributes to cell growth, proliferation, survival and diet (75.2% ± 1.4% vs. 63.9% ± 4.2%, respectively; P<0.05). These data dem- metabolism. However, it remains to be clarifi ed whether DEPTOR in the liver onstrate that PANDER overexpression induces hepatic insulin resistance in plays any roles in glucose metabolism.First, levels of endogenous DEPTOR our transgenic model and further indicates a biological function in hepatic expressions in the liver and white adipose tissue (WAT) were decreased in insulin signaling counter-regulation. obese mice. Then, to examine whether hepatic DEPTOR has any metabolic effects, we expressed DEPTOR in the livers of lean mice (DEP-mice) by us- 1766-P ing the adenovirus-mediated gene delivery. Glucose tolerance tests (GTTs) revealed that DEP-mice exhibited better glucose tolerance and lower plasma insulin levels during GTTs. Next, to explore the effect of DEPTOR in obesity, WITHDRAWN we expressed DEPTOR in the livers of mice with diet-induced obesity (DEP- DIO-mice) or genetic obesity (KKAy) (DEP-K-mice). Hepatic DEPTOR expres- sion in obese mice markedly improved glucose intolerance and decreased plasma insulin levels during GTTs, suggesting improvement of systemic in- sulin resistance. In detail, both insulin-stimulated tyrosine phosphorylation of the insulin receptor (IR) and serine-473 phosphorylation of Akt in the liver were enhanced in DEP-DIO mice. Levels of hepatic gluconeogenic enzyme, phosphoenolpyruvate carboxykinase were decreased in DEP-DIO- and DEP- K-mice. These results indicate that hepatic DEPTOR expression suppressed gluconeogenesis in the liver, thereby lowering fasting blood glucose. In ad- dition, periodic acid-schiff (PAS) staining revealed that DEPTOR expression increased glycogen accumulation in the livers of obese mice. Furthermore, body weight and WAT weight on day 7 after adenovirus administration were reduced, and additionally, plasma leptin levels and food intakes for 7days were decreased in DEP-DIO-mice, suggesting improved leptin resistance. In conclusion, hepatic DEPTOR expression improves systemic insulin resis- tance in the states of obesity, constituting a potential therapeutic target for the metabolic syndrome.

& 1769-P SRT3025, a Novel Small Molecule SIRT1 Activator, Reduces Hepat- ic Steatosis in Diet Induced Obese (DIO) Mice Through Suppression of Lipogenesis and Increased Fatty Acid Oxidation QING NIE, YONG QI, MEGHAN L. DAVIS, MARC O. JOHNSON, ELDEN O. LAINEZ, ANGELA M. COTE, GEORGE P. VLASUK, JAMES L. ELLIS, VIPIN SURI, Cambridge, MA Non alcoholic fatty liver disease (NAFLD) and Non Alcoholic Steatohepa- titis (NASH) are frequently associated with metabolic dysfunction including Type 2 diabetes and obesity. The protein deacetylase SIRT1 plays a critical role in the regulation of metabolic and infl ammatory processes. Transgenic mice overexpressing SIRT1 are strikingly resistant to high fat diet induced hepatic lipid accumulation as well as infl ammation while mice lacking SIRT1 & 1767-P are highly susceptible to diet induced hepatic steatosis.The effect of a novel Molecular Mechanisms of Ectopic Lipid Accumulation in Liver SIRT1 activator, SRT3025, on hepatic steatosis in DIO mice was studied BIRGIT KNEBEL, STEFAN LEHR, JUTTA HAAS, SONJA HARTWIG, ULRIKE NITZ- following high fat diet feeding (60% KCal from fat) to 6 week old C57/BL6 GEN, SYLVIA JACOB, DIRK MÜLLER-WIELAND, JÖRG KOTZKA, Düsseldorf, Ger- mice for 12 weeks followed by treatment with 100 mg/kg/day SRT3025 for 7 many, Hamburg, Germany weeks. SRT3025 treatment reduced body weight, improved insulin sensitiv- The transcription factor SREBP (sterol regulatory binding protein)-1a plays ity, reduced serum triglycerides by 41% and hepatic triglycerides by 50% a dominant role in intracellular lipid metabolism. Previously we have shown compared to vehicle treated DIO mice. Oil red O staining confi rmed a signifi - that SREBP-1a is a substrate of ERK, JNK and p38 MAP kinases. To provide cant reduction in lipid accumulation in SRT3025 treated mice.To understand

direct evidence that phosphorylation is of biological and clinical relevance, the time course of these effects, livers from DIO mice were analyzed after 1, Obesity we generated transgenic mice expressing mature SREBP-1a variant lacking 4 and 7 weeks of treatment with 100 mg/kg/day SRT3025. SRT3025 reduced POSTERS all phosphorylation sites designed as alb-SREBP-1a6P and wild type SREBP- hepatic lipid accumulation after 1 week and substantially resolved steato-

1a liver specifi c under control of the albumin promoter and a liver specifi c sis after 4 and 7 weeks. Biochemical measurement of livers from SRT3025 Integrated Physiology/ enhancer. Macroscopic examination showed that preventing of SREBP-1a treated mice revealed a reduction in triglycerides of 17% at week 1, 31% at phosphorylation protects mice not only for fatty liver disease but also from week 4 and 63% at week 7. Analysis of hepatic gene expression and metabo- obesity. To understand molecular mechanisms related to hepatic lipid accu- lite profi les showed SRT3025 reduced expression of several lipogenic genes, mulation a holistic gene expression analyses in liver was performed. These including ATP citrate lyase and acyl CoA synthetases, elongases and desatu- investigation revealed mainly genes involved in lipid metabolism (p= 3.98E- rases. In addition, metabolite profi ling suggested that SRT3025 increased 04) which were differentially regulated in alb-SREBP-1a and alb-SREBP-1a6P fatty acid oxidation.These data strongly suggest therapeutic potential for mice. Inasmuch as organelle specifi c transcripts were regulated we focus pharmacological SIRT1 activation with SRT3025 in the treatment of hepatic on mitochondria or peroxisomes. Accordingly, proteome analyses using 2D- steatosis associated with metabolic dysfunction in humans. DIGE and mass spectroscopy of highly purifi ed liver mitochondria and peroxi- somes were performed. Although mitochondria are the major subcellular or-

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A455 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1772-P & 1770-P Foxo3 Modulates Cholesterol Biosynthesis via Suppression of Effects of Perilipin 2 Antisense Oligonucleotide Treatment on He- Srebf2 Gene Expression patic Lipid Metabolism and Gene Expression RONGYA TAO, XIWEN XIONG, RONALD A. DEPINHO, XIAOCHENG C. DONG, In- YUMI IMAI, SIOBHAN BOYLE, GLADYS M. VARELA, RAJESH PATEL, EMILIE CA- dianapolis, IN, Houston, TX RON, XIAOYAN YIN, RAINA DHIR, RAVINDRA DHIR, MARK GRAHAM, REXFORD Foxo transcription factors have pleiotropic biological functions including S. AHIMA, Norfolk, VA, Philadelphia, PA, Carlsbad, CA their expanding roles in metabolism. Although Foxo3 hasbeen implicated in Non-alcoholic fatty liver disease (NAFLD) is the most common liver dis- cholesterol metabolism, the underlying molecular mechanisms are not clear. ease worldwide, and may progress to liver cirrhosis and cancer. NAFLD is In this work, we attempted to address this problem by using both cell-based characterized by insulin resistance, and closely associated with obesity, dys- and whole-animal systems. Initially, we observed that Foxo1/3/4 liver-specif- lipidemia, hyperglycemia, and cardiovascular morbidity. We have previously ic triple knockout mice (LTKO) exhibited 26% and 30% increases in the liver shown that Perilipin 2 (Plin2), also known as Adipose Differentiation Related and serum cholesterol levels, respectively; and these abnormalities were Peptide (ADRP), a member of the family of lipid droplet proteins, is markedly largely reversed by hepatic overexpression of Foxo3. We then analyzed increased in NAFLD. Reducing plin2 in the liver of diet-induced obese (DIO) Foxo3 liver-specifi c knockout mice and also found an signifi cant increase mice by anti-sense oligonucleotide (Plin2 ASO) treatment decreased hepatic in cholesterol levels in both liver and serum. Promoter sequence analyses steatosis and enhanced insulin sensitivity. We studied the role of Plin2 in the and luciferase reporter assays revealed that Foxo3 could downregulate liver by performing a microarray analysis of gene expression and validating the masterregulator of cholesterol biosynthesis_SREBP-2 encoded by the the results using RT PCR. Male C57BL/6J mice were fed a high fat (45%) Srebf2 gene_through direct binding to an insulin-response element in the diet for 4 weeks to induce steatosis, and were treated simultaneously with proximal promoter of the Srebf2 gene. This was also confi rmed by chroma- Plin2 ASO or scrambled (control) ASO (25 mg/kg IP twice weekly). Plin2 ASO tin immunoprecipitation analysis.Furthermore, epigenetic analysis showed specifi cally decreased Plin2 in the liver without affecting other lipid droplet that suppression of Srebf2 geneexpression by Foxo3 could be attributed to proteins. Plin2 ASO decreased hepatic and serum triglyceride levels, and in- a decrease in histone acetylation in the chromatin surrounding the Srebp2 creased ketone levels. Plin2 ASO decreased the expression of key lipogenic gene promoter. To further test whether Foxo3 might improve insulin resis- enzymes (dgat2, fas, acc) as well as enzymes involved in fatty acid metabo- tance induced dyslipidemia, we overexpressed Foxo3 in the liver of mice lism (acsl1, lipe) and steroid metabolism (hmgcr, hsd3b5, hsd17b2), indicating fed high-fat diet via adenovirus-mediated genetransfer. As expected, Foxo3 that Plin2 affects hepatic lipid metabolism beyond triglycerides. Plin2 ASO overexpression signifi cantly reduced expression of hepatic Srebf2 and its increased the expression of genes involved in the regulation of hepatocyte target genes including Hmgcr and Hmgcs1; as a result, hepatic and serum proliferation (afp, H19), mitosis (ccna2, incenp, sgol1), and extracellular cholesterol levels were lower by 30% and 20% as compared to control mice, matrix (col1a1, col3a1, mmp8). These results indicate that Plin2 has major respectively. Taken together, our data suggest a critical role of Foxo3 in cho- structural effects on hepatocytes, stellate cells, and matrix proteins. Further lesterol homeostasis. studies will determine whether the Plin2-mediated changes in hepatic lipids Supported by: NIDDK (R00DK077505) and structure are causally related, and whether this has implications for the pathogenesis of liver cirrhosis and cancer. Supported by: NIH R01DK062348, P01DK049210, and P30DK19525 (R.A.), 1773-P R01DK090490 (Y.I.) Ƶ-catenin Antisense Oligonucleotides Protect Mice Against Diet- Induced Hepatic Steatosis and Hepatic and Peripheral Insulin Re- sistance & 1771-P VIOLETA B. POPOV, FRANCOIS R. JORNAYVAZ, EMIN O. AKGUL, SHOICHI KANDA, PNPLA3 is Regulated by Glucose via CHREBP and its I148M Mutant MICHAEL J. JURCZAK, SACHIN MAJUMDAR, BLAS GUIGNI, KITT FALK PETERS- Slows Down Triglyceride Hydrolysis in Human Hepatocytes EN, PRASAD MANCHEM, SANJAY BHANOT, GERALD I. SHULMAN, VARMAN T. JULIA PERTTILÄ, CAROLINA HUAMAN SAMANEZ, SANDRINE CARON, BART SAMUEL, New Haven, CT, Carlsbad, CA STAELS, HANNELE YKI-JÄRVINEN, VESA M. OLKKONEN, Helsinki, Finland, Lille, Genetic studies have linked mutations in the Wnt signaling pathway France with the metabolic syndrome and type 2 diabetes. However, the cellular Adiponutrin, encoded by the PNPLA3 gene, belongs to the family of pa- mechanism by which this pathway affects insulin sensitivity is unknown. tatin-like domain containing enzymes and is expressed in liver and adipose To examine this question, we used a specifi c antisense oligonucleotide tissue. At least 20 independent studies have shown that a missense muta- (ASO) to decrease hepatic and adipose expression of `-catenin, the fi nal tion, encoding an isoleucine to methionine substitution at amino acid 148 downstream mediator of the canonical Wnt signaling pathway. We quanti- (rs738409; I148M), is associated with an increased hepatic fat content. Minor fi ed the impact on: lipid metabolism, whole body energy metabolism, and allele homozygotes (GG, approximately 5% of the population) display a 70% liver and muscle insulin sensitivity by hyperinsulinemic-euglycemic clamp elevated liver fat content and also have an increased risk of non-alcoholic studies. After four weeks of treatment, `-catenin ASO decreased expres- steatohepatitis. Adiponutrin is reported to show triglyceride lipase and acyl- sion of `-catenin by 80% in the liver and 70% in WAT relative to control ASO transferase activities, but its physiologic function and regulation are poorly treated mice. Both groups had similar body weight and body composition as understood.We studied transcriptional regulation of PNPLA3 in human hepa- assessed by1H magnetic resonance spectroscopy. `-catenin knockdown did tocytes and the impact of PNPLA3 I148M mutant on hepatocyte triglyceride not alter whole-body energy metabolism, food intake or locomotor activity. (TG) metabolism. Silencing of the carbohydrate responsive element binding However, `-catenin ASO improved the plasma lipid profi le with a 17% de- protein (ChREBP) abolished induction of PNPLA3 mRNA by glucose. Glucose- crease in cholesterol (P=0.03), a 37% decrease in triglycerides (P=0.03) and dependent binding of ChREBP to a carbohydrate response element in the a 20% decrease in fatty acids (P=0.03). Furthermore `-catenin knockdown PNPLA3 promoter was demonstrated by chromatin immunoprecipitation. protected mice from lipid-induced liver and muscle insulin resistance as re- [3H]acetate or [3H]oleate incorporation with 1 h pulse labeling, or 18 h [3H] fl ected by greater suppression of hepatic glucose production (31% vs. 82%, oleate labelling in HuH7 cells showed no effect of PNPLA3 I148M on TG syn- p= 0.03) and a 30% increase in peripheral glucose uptake (P=0.01) during thesis in the absence of free fatty acid (FFA) loading. Increased [3H]oleate the hyperinsulinemic-euglycemic clamp. This improvement in hepatic insu- accumulation into triglycerides in I148M expressing cells was observed after

Obesity lin sensitivity was associated with reductions in hepatic triglyceride (44%, 18 h of labelling in the presence of 200 μM FFA-albumin complexes. This POSTERS P<0.05) and diacylglycerol (60%, P<0.01). In contrast, hepatic ceramide con- was accompanied by increased PNPLA3 protein levels. The rate of hydroly- tent was increased 30% (P<0.001). CONCLUSIONS: ASO mediated knock- sis of [3H]TG during lipid depletion was signifi cantly decreased by PNPLA3

Integrated Physiology/ down of `-catenin in liver and adipose tissue decreases hepatic steatosis I148M.In conclusion, PNPLA3 is regulated in human hepatocytes by glucose and protects mice from lipid-induced liver and peripheral insulin resistance. via ChREBP. PNPLA3 I148M enhances cellular accumulation of [3H]TG in the Targeted inhibition of `-catenin activity may be a novel therapeutic strategy presence of excess FFA, which is known to stabilize PNPLA3 protein. These for treatment of NAFLD and hepatic insulin resistance. data do not exclude an effect of PNPLA3 I148M on hepatocyte lipogenesis, Supported by: NIDDK (P30-34989) but show that the mutant increases the stability of triglycerides.

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A456 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1774-P 1776-P GPR40 Agonist Treatment Shows Disease Reversal at the Gene Ex- Metformin Inhibits Proliferation and Colony Formation Ability pression Level in Adipose and Liver of Goto Kakizaki Rats through AMP-Activated Protein Kinase in Hepatocellular Carci- ERIC MUISE, MARIA E. TRUJILLO, MELISSA E. KIRKLAND, DANIEL KOSINSKI, MI- noma Cells CHELE J. PACHANSKI, JOHN R. THOMPSON, MARGARET VAN HEEK, ANDREW LONG YI ZHENG, DA JIN ZOU, Shanghai, China D. HOWARD, DAVID E. KELLEY, PAUL E. CARRINGTON, Rahway, NJ The anti-diabetic drug metformin reduces human cancer incidence and G protein-coupled receptor 40 (GPR40) is a free fatty acid receptor that improves the survival of cancer patients, including those with hepatocellu- is highly expressed in pancreatic ` cells and mediates free fatty acid-in- lar carcinoma. However, its biological and molecular activity against HCC is duced and glucose-dependent, insulin secretion (GDIS). Chronic treatment still not well understood. Here, we report that metformin treatment signifi - of diabetic Goto Kakizaki (GK) rats with a GPR40 agonist decreased basal cantly inhibited proliferation of diverse HCC cancer cell lines (SMMC7721, and fasted glucose levels, without measurable changes in insulin raising HepG2, HCC-LM3, Huh7) in a time- and dose-dependent manner. It also the question of effects in target tissues aside from the pancreas. RNA induced apoptosis, reduced colony formation ability, and caused cell cycle from livers and epididymal white adipose tissue (EWAT) were profi led on arrest accompanied by decreased cyclin D1. The growth inhibition was as- Affymetrix microarrays following chronic treatment with GPR40 or PPARa sociated with decreased IL-6/STAT3 signaling pathway. At the molecular agonists. GK rats treated with GPR40 agonist or Rosiglitazone for 21 days, level, metformin treatment led to activation of AMP-activated protein ki- showed ~20% glucose and ~5% HbA1C reductions. However, Rosiglitazone nase. Transfection of siRNA against AMPK_ eradicated the inhibitory ef- treatment resulted in an almost two-fold greater increase in body weight fects of metformin on IL-6/STAT3 signaling pathway, and rescued cells from over vehicle or GRP40 agonist treated animals.Comparison of expression metformin-induced growth inhibition, implying the essential role of AMPK. data from liver or adipose of vehicle treated GK rats and Wistar controls, Importantly, oral treatment with metformin led to a signifi cant reduction revealed a distinct “disease-associated” signature. Both Rosiglitazone and of tumor growth in mice bearing xenografts of SMMC7721 and HCC-LM3, GPR40 agonist treatment provided robust gene signatures in EWAT and accompanied by inhibition of tumor cell proliferation (as assessed by Ki-67 liver. Comparison of Rosiglitazone and GPR40 treatment data showed some immunohistochemical staining). In conclusion, our study provides a strong overlap, but at least half of the GPR40 agonist-mediated gene changes in evidence for antineoplastic effects of metformin in HCC treatment. each tissue were unique. Between 10 and 45% of the “disease-associated” Supported by: National Natural Science Foundation of China signatures, were also regulated by GPR40 agonist or Rosiglitazone treat- ment. Comparing changes between disease and treatment, the vast majority 1777-P were oppositely regulated by both compounds in liver, suggestive of disease The Role of Glucose-Dependent Insulinotropic Polypeptide (GIP) in reversal. In EWAT, the Rosiglitazone treatment effected an almost equal High Glycemic Index (GI) Diet Induced Hepatic Fat Accumulation split between disease amelioration and exacerbation. Conversely, GPR40 and Insulin Resistance agonist treatment resulted in a nearly complete reversal of the subset of FARNAZ KEYHANI-NEJAD, FRANK ISKEN, ANDREAS L. BIRKENFELD, ANDREAS the “disease-associated” signature. Together with the lack of weight-gain F. PFEIFFER, Nuthetal, Germany, Berlin, Germany these data would suggest that GPR40 agonists may provide robust glucose High GI diets induce hepatic fat accumulation and insulin resistance; also, lowering, without some of the liabilities associated with chronic treatment high GI diets strongly stimulate GIP secretion. The role of GIP in the devel- with PPARa agonists. opment of non alcoholic fatty liver disease (NAFLD) and insulin resistance has not been studied in vivo. Here, we hypothesized that altered GIP secre- 1775-P tion in mice fed different GI diets contributes to the development of NA- Amelioration of Excessive Hepatic Oxidative Metabolism and In- FLD, without affecting body composition.Weight matched, male C57Bl/6J fl ammation by Reducing PEPCK-C Activity in Diet Induced Obese mice were fed a high or low GI diet identical in macro and micronutrient Mice composition 40% Kcal from fat, 40% Kcal from carbohydrate. Body weight SANTHOSH SATAPATI, NISHANTH SUNNY, TIANTENG HE, SHAWN C. BURGESS, was assessed weekly, body composition was measured by nuclear magnetic Dallas, TX resonance spectroscopy. IPGTT was performed with 2mg/g glucose. Liver Hepatic insulin resistance is accompanied by elevated gluconeogenesis Triglyceride (TG) was determined from livers in fasted mice. A feeding test (GNG), lipid accumulation, oxidative stress and infl ammation. We previously was performed with the high or low GI diets after an overnight fast. Studies demonstrated that (i) nonalcoholic fatty liver disease (NAFLD) and insu- were repeated in GIP receptor KO (Gipr-/-) mice.After 22 weeks of a high fat, lin resistance occurs in the presence of a markedly elevated hepatic TCA high or low GI diet, body weight, body composition and food intake did not cycle metabolism and (ii) that gluconeogenesis (cataplerotic fl ux) catalyzed differ between groups. Postprandial plasma GIP level was 2.3 - fold higher by PEPCK-C robustly induces anaplerotic and oxidative TCA cycle function. compared to the low GI group (AUC 49640 ± 3770 vs. 21730 ± 3380, p<0.001). Therefore, we hypothesized that suppression of PEPCK-C fl ux during hepatic Hepatic TG content was increased 2- fold in the high GI group (3.1 ± 0.4 vs. insulin resistance might protect the liver from elevated oxidative metabo- 1.6 ± 0.3 mg/mg, p<0.01). During an IPGTT, the high GI group showed an lism and potential collateral oxidative damage. To test this hypothesis, we increased glucose and insulin excursion (AUC 3787 ± 310 vs. 2873 ± 175 and examined the effect of whole body PEPCK-C knockdown (~70%) in mice in- 113360 ± 18530 vs. 66273 ± 7003, p<0.05; respectively) compared to the low duced to obesity and insulin resistance by a 60% high fat diet (HFD). Male GI diet group, indicative of aggravated insulin resistance in the high GI group. Pcklox/lox (WT) and Pcklox/neo (KD) mice were fed either a control or HFD for Deletion of GIP receptor in Gipr-/- mice completely abolished the metabolic 16 weeks and in vivo hepatic mitochondrial oxidative and anaplerotic fl uxes derangements compared to control mice on a high GI diet.Together, these were determined using 2H/13C tracers detected in plasma metabolites by data suggest that in the setting of a high fat diet, high GI feeding aggravates NMR and LC-MS/MS. As expected, WT mice fed a HFD had signifi cantly hepatic steatosis and insulin resistance at least in part through an exag- elevated glycemia, endogenous glucose production (EGP) and GNG. In ad- gerated release of GIP. Our data show sugars with a low GI as attractive dition, hepatic mitochondrial anaplerotic and oxidative TCA cycle fl ux and nutritional ingredients for energy rich, highly refi ned foods. infl ammatory responses were elevated by diet induced obesity. KD mice had normal glycemia and rates of GNG on a control diet, as we previously re- 1778-P ported, but were markedly protected from elevated EGP, GNG, mitochondrial Lipid Metabolism, Oxidative Stress and Cell Death are Regulated by Obesity anaplerotic and oxidative TCA cycle fl ux on a HFD. In addition to suppressed PKC Delta in Dietary Models of Steatohepatitis POSTERS oxidative metabolism, the infl ammatory response on HFD was normalized MICHAEL W. GREENE, CHRISTINE M. BURRINGTON, DARIN T. LYNCH, SAMAN- by 70% knock down of PEPCK-C. We conclude that, in diet induced obese THA K. DAVENPORT, ANDREW K. JOHNSON, MELISSA J. HORSMAN, SALEEM Integrated Physiology/ mice, suppression of PEPCK-C fl ux reduces TCA cycle oxidative metabolism CHOWDHRY, PAUL C. TIRRELL, Cooperstown, NY and that this is suffi cient to protect against hepatic insulin resistance and Nonalcoholic steatohepatitis, which is seen in association with obesity, infl ammation. insulin resistance, and diabetes, is characterized by fatty liver, infl amma- Supported by: RO1DK078184 tion, oxidative stress, apoptosis, and fi brosis. We previously have shown that hepatic PKCb protein levels and activation correlate with steatohepa- titis progression in methionine and choline-defi cient (MCD) diet fed mice, an animal model of non-alcoholic steatohepatitis. To determine the role of PKCb in the pathophysiology of steatohepatitis, we fed PKCb null mice and wildtype (WT) littermates a control or MCD diet. No difference in hepatic steatosis was observed between WT and PKCb null mice fed the MCD diet.

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A457 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

However, the expression of key regulators of `-oxidation and plasma triglyc- NGT had more frequently elevated plasma triglycerides, low HDL, or both eride metabolism was differentially modulated in MCD fed PKCb null mice. (43% vs. 24%, 72% vs. 38% and 36% vs. 12%, respectively, all p<0.05-01) Consistent with these results, the MCD diet-induced reduction in serum trig- and increased markers of subclinical infl ammation (plasma MCP-1, sVCAM, lyceride was blocked in PKCb null mice. The ability of fatty acids to directly IL-6, IL-8 and TNF-_). Conclusion: the prevalence of NAFLD and NASH in modulate lipid metabolism gene expression was confi rmed using primary T2DM is much higher than previously believed, frequently with normal AST/ hepatocytes isolated from WT and PKCb null mice. MCD diet-induced he- ALT. This is associated with worse insulin resistance and an unfavorable CV patic oxidative stress, as measured by elevated staining for 4-HNE and an risk profi le compared to NGT pts with NAFLD. increase in TBARS in the liver, was blocked in PKCb null mice. Similar results Supported by: Burroughs Welcome Fund in db/db PKCb null mice confi rmed our fi ndings in an obese model of steato- hepatitis. Basal gene expression of the NADPH oxidase complex was signifi - 1781-P cantly elevated in control diet fed PKCb null mice, and the MCD diet had no Relationships of Circulating Cytokeratin-18 Fragment With Liver effect on its expression, suggesting that basal oxidative stress may play a Fat Content and Insulin Sensitivity before and after a Lifestyle protective role in PKCb null mice. Hepatocyte apoptosis and expression of Intervention pro-apoptotic genes was reduced in PKCb null mice compared to WT mice KONSTANTINOS KANTARTZIS, ANDREAS PETER, JÜRGEN MACHANN, FRITZ fed a MCD diet. Further, fatty acid-induced stimulation of cell cycle and pro- SCHICK, ANDREAS FRITSCHE, HANS-ULRICH HÄRING, NORBERT STEFAN, Tübin- apoptotic genes was reduced in PKCb null hepatocytes. Our results suggest gen, Germany that PKCb plays a role in the progression of diet-induced steatohepatitis and Cytokeratin-18 fragment (CK-18), a major intermediate fi lament protein in may be a target for therapeutic treatment of steatohepatitis. hepatocytes that is being cleaved by caspases during apoptosis, may be a useful marker for the prediction of nonalcoholic steatohepatitis. To date 1779-P there is little information whether circulating CK-18 also predicts the more Hypothalamic Orexin Defi ciency Causes Age Related Insulin Resis- prevalent condition nonalcoholic fatty liver (NAFL) and whether it can be tance via Enhancement of ER Stress in the Liver of Mice used as a marker of impaired glucose metabolism in NAFL. Furthermore, it TOSHIYASU SASAOKA, EMI TOKAI, YUYA NAKAMURA, MIKIO FUJITA, TSU- is unknown whether circulating CK-18 changes when improvement in liv- TOMU WADA, HIROSHI TSUNEKI, Toyama, Japan er fat is being observed. We investigated the relationships of circulating Increased hepatic glucose production (HGP) is a crucial cause of fast- caspase-generated neoepitope CK-18 with liver fat content (1H-MRS), body ing hyperglycemia in patients with diabetes. However, the integration fat distribution, (MRT) and insulin sensitivity [75-g OGTT and euglycemic, mechanism between central and peripheral regulation of HGP is unknown. hyperinsulinemic clamp (n=168)] in 220 subjects before and after a 9-month Therefore, we examined the involvement of hypothalamic orexin system in lifestyle intervention.At baseline circulating CK-18 was higher in men than the mechanism. Intracerebroventricular injection of orexin A at low doses in women (p=0.006). After adjustment for sex circulating CK-18 correlated caused decrease in the fasting blood glucose levels and the mRNA levels positively with total body fat (r=0.16, p=0.015) and visceral fat (r=0.16, encoding hepatic gluconeogenic genes PEPCK and G6Pase, whereas these p=0.021) and most strongly with liver fat (r=0.30, p0.067). Unexpectedly, levels were increased by high doses of orexin A in C57BL/6J mice. The de- a negative correlation with adjusted fasting glycemia was found (r=-0.19, creasing and increasing effects of orexin A were selectively blocked by a p=0.005), which became even stronger after additional adjustment for liver muscarinic antagonist atropine and an alpha-blocker phenoxybenzamine, fat (r=-0.26, p<0.0001). Also, a negative relationship with HbA1c levels was respectively. On the other hand, orexin knockout (OXKO) mice at 6 months found (r=-19, p=0.004). During the intervention these parameters improved. old demonstrated elevated glucose levels in pyruvate tolerance test, and Circulating CK-18 was found to decrease strongest in subjects who had the showed abnormal expression of IRS1, IRS2, and PEPCK in the liver. Insulin- largest decrease liver fat.In conclusion, our data suggest that circulating induced phosphorylation of Akt was impaired in the liver but not skeletal CK-18 may be a useful parameter to estimate liver fat content, already in muscle of 6-month-old OXKO mice. Moreover, the mRNA levels of CHOP, an moderate stages of hepatic steatosis, and that it may help to understand endoplasmic reticulum (ER) stress-associated proapoptotic factor, were in- the complex relationship between liver fat content, hepatic apoptosis and creased, whereas those of XBP1s and ATF6 regulating the expression of ER hepatic glucose production. chaperone were decreased in the liver of OXKO mice. These results indicate that hypothalamic orexin contributes to the maintenance of ER homeostasis 1782-P in the liver and dually regulates HGP by controlling vagus-sympathetic bal- Ezetimibe Prevents the Hepatic Steatosis Induced by a High-Fat But ance. Since orexin expression is reduced under hyperglycemic condition, the Not a High-Fructose Diet lack of orexin may be involved in a vicious cycle to develop hepatic insulin MASATERU USHIO, YOSHIHIKO NISHIO, YOSHIO NAGAI, YASUHIRO MAENO, resistance in type 2 diabetes. SHINJI KUME, TAKESHI YOSHIZAKI, KATSUTARO MORINO, SATOSHI UGI, OS- AMU SEKINE, ATSUNORI KASHIWAGI, HIROSHI MAEGAWA, Otsu Shiga, Japan, 1780-P Kagoshima, Japan, Kawasaki Kanagawa, Japan, Higashiomi Shiga, Japan NAFLD is Common in Patients With T2DM and Associated With Non-alcoholic fatty liver disease is the most frequent liver disease. Severe Insulin Resistance (IR) and Risk Factors for Cardiovascular Ezetimibe, an inhibitor of intestinal cholesterol absorption, was reported Disease (CVD) to ameliorate hepatic steatosis in human and animal model. To explore the ROMINA LOMONACO, CAROLINA ORTIZ-LOPEZ, BEVERLY ORSAK, JOAN FINCH, mechanism how ezetimibe reduces hepatic steatosis, we investigated the ROSE KAMINSKI-GRAHAM, KENNETH CUSI, San Antonio, TX, Gainesville, FL effects of ezetimibe on the expression of lipogenic enzymes and their tran- The true prevalence and metabolic factors contributing to the development scription factors in the mice fed with a high-fat or high-fructose diet. CBA/JN in T2DM of nonalcoholic fatty liver disease (NAFLD), and of its severe form mice were fed on a high-fat diet or a high-fructose diet for 8 weeks with or with steatohepatitis (NASH) remain unknown. To this end, we screened for without ezetimibe (0.008% wt/wt). A high-fat diet induced hepatic steatosis NAFLD by the gold-standard magnetic resonance spectroscopy (MRS) 81 pa- accompanied with hyperinsulinemia. The treatment with ezetimibe reduced tients with T2DM (age: 56±1; M/F: 72/28%; ethnicity: Hisp=56%, Cauc=35%, hepatic steatosis as well as insulin, fasting plasma glucose, and glucose AA=10%; BMI: 33.6±0.5 kg/m2; A1c= 7.2±0.1%) and compared to 34 controls production from pyruvate, suggesting the reduction of insulin resistance in

Obesity with normal glucose tolerance (NGT), all being unaware of having any liver the liver. Intestinal mRNA expression showed that ezetimibe reduced the

POSTERS disease. We measured: 1) total body fat (TBF) by DXA; 2) liver and muscle expression of fatty acid transfer protein4 in the mice fed with a high-fat diet. (Rd) insulin sensitivity (insulin clamp with 3-[3H]glucose); 3) hepatic IR The treatment with ezetimibe, however, prevented neither hepatic steatosis

Integrated Physiology/ (HIRi=EGP x fasting insulin [FPI]); 4) adipose tissue IR (AdipoIRi= fasting FFA x nor hyperinsulinemia in the mice fed with a high-fructose diet. Ezetimibe de- FPI). While pts with T2DM and NGT were well matched for ethnicity/gender/ creased LXR_ binding to SREBP-1c promoter and hepatic SREBP-1c but not BMI/TBF, the prevalence of NAFLD (80% vs. 53%; p<0.01) and NASH (80% fatty acid synthase mRNA expression in the mice fed with a high-fructose vs. 44%; p<0.01) in T2DM was much higher than in NGT. Most NAFLD pts in diet, suggesting that ezetimibe did not reduce hepatic lipogenesis induced both groups had normal AST/ALT (79% and 72%, respectively, NS). T2DM by a high-fructose diet. Thus, the treatment with ezetimibe prevented the compared to NGT pts with NAFLD had worse hepatic steatosis (MRS= 22±1 hepatic steatosis in the mouse fed with high-fat diet but not high-fructose vs. 16±2; p=0.03) and more severe IR at all levels: hepatic (HIRi= 22.5±2.3 vs. diet. The elevation of hepatic nuclear expression of ChREBP only in the mice 10.6±3.6 mg · kg−1 · min−1 · μU/mL and insulin suppression of EGP= 40±3% fed with a high-fructose diet may partly explain the differences in the effect vs. 54±5%), adipose tissue (AdipoIRi= 7.2±0.6 vs. 2.4±1.0 mmol/L · μU/mL of ezetimibe. and insulin suppression of FFA= 47±3% vs. 63±5%) and muscle (Rd= 5.7±0.6 vs. 10.1±0.8 mg · kgLBM-1 · min-1, all p<0.001). Pts with NAFLD and T2DM vs.

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A458 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1783-P tionships are yet unclear. We examined hepatic and muscular metabolism in Anti-Obesity and Anti-Diabetic Effects of Lichochalcone A wild-type mice (CON) and two transgenic mouse strains with different forms HAI YAN QUAN, SOO JUNG KIM, GO WOON KIM, DO YEON KIM, SUNG HYUN of NAFL, either a primary form due to liver- (alb-SREBP-1c: ALB), or a second- CHUNG, Seoul, Republic of Korea ary form, due to adipose tissue- (aP2-SREBP-1c: AP2) specifi c overexpression Licochalcone A (LA) has been shown to exert multiple pharmacological of the sterol regulatory-element binding protein-1c (SREBP-1c). Mitochon- effects. In this study, we examine anti-diabetic and anti-obesity activities drial oxidative capacity was measured in permeabilized tissues using high- of LA and action mechanisms in the ICR mice. They were randomly divided resolution respirometry (Oroboros Instruments). Mitochondrial content was into four groups; regular diet fed group (RD), high fat diet fed group (HFD), assessed from the ratio of mtDNA to nuclear DNA, using quantitative PCR. HFD plus LA (5, 10 mg/kg) groups. Body weight, blood glucose, triglyceride, Insulin sensitivity was quantifi ed at fasting from homeostasis model assess- cholesterol and non-esterifi ed fatty acid levels in the group given 10 mg/kg ment-estimated insulin resistance (HOMA-IR) and with the hyperinsuline- LA were signifi cant reduced by 14.0, 25.7, 48.2, 58.9 and 73.5%, respectively, mic-euglycemic clamp. AP2 mice with secondary NAFL were markedly more compared to HFD control group. We found that LA decreased the blood glu- insulin resistant as indicated by greater HOMA-IR (26.4±20.6, ALB: 5.2±3.0, cose level through inhibition of gluconeogenesis via AMPK pathway in the CON: 2.2±1.3; p<0.05 AP2 vs. CON) and lower insulin-mediated whole-body mice liver. LA also inhibited lipogenesis via suppression of sterol regulatory insulin sensitivity (glucose infusion rates: 17.3±9.4, ALB: 45.4±5.3, CON: element binding protein-1c (SREBP-1c) and its target genes, which enhanced 49.0±18.0 mg/kg/min; p<0.05 AP2 vs. ALB and CON). Moreover, AP2 mice insulin sensitivity. In the epididymal fat, LA signifi cantly reduced lipid ac- had 37% greater (p<0.001) liver weight than ALB and CON, accompanied cumulation and down-regulated the expression of peroxisome proliferator- by almost doubled pyruvate-dependent mitochondrial respiration capacity in activated receptora (PPARa), CCAAT/enhancer binding protein _ (C/EBP_), liver (p<0.01) and fatty acid-dependent mitochondrial respiration in muscle SREBP-1c and their target genes associated with adipogenesis. Therefore, we (p<0.01). Mitochondrial content was not affected by any of the conditions. propose LA as a potent therapeutic agent for type 2 diabetes and obesity. Hepatic steatosis does not necessarily lead to impaired mitochondrial func- tion, but rather increases oxidative capacity to compensate for elevated en- ergy supply. On the other hand, insulin resistance may develop progressively with increasing severity of fatty liver. Supported by: German Diabetes Association Menarini, Dr. Skröder Foundation

1785-P Additive Effects of Nicotine and High Fat Diet on Hepatic Steatosis in Mice Involve Oxidative Stress and Hepatocellular Apoptosis and are Mediated by Abdominal Lipolysis THEODORE C. FRIEDMAN, INDRANI SINHA-HIKIM, MEHER PARVEEN, MICHAEL MANGUBAT, CHANG-SUNG SHIN, ALEXEI LYZLOV, RASHEED IVEY, SAMUEL W. FRENCH, AMIYA P. SINHA-HIKIM, Los Angeles, CA, Torrance, CA Smoking is a major risk factor for diabetes and cardiovascular disease and may contribute to non alcoholic fatty liver disease (NAFLD). NAFLD has also been identifi ed as an independent risk factor for insulin resistance, athero- sclerosis and cardiovascular disease. We hypothesized that nicotine plus a high-fat diet (HFD) would have additive effects on the severity of hepatic steatosis in obese mice. Adult C57BL6 male mice were fed with normal chow diet (NCD) or HFD with 60% of calories derived from fat and given twice daily injections of nicotine (0.75 mg/kg BW, ip) or saline for 10 weeks. Light and electron microscopy revealed markedly increased lipid accumulation in hepatocytes from mice on a HFD plus nicotine, compared to mice on a HFD alone. The additive effects of nicotine plus a HFD was associated with great- er oxidative stress, increased incidence of hepatocyte apoptosis, inactiva- tion (dephosphorylation) of (AMPK, and activation of its downstream target acetyl-COA-carboxylase (ACC), leading to increased lipogenesis. Treatment with acipimox (an abdominal lipolysis inhibitor) completely blocked nicotine plus HFD-induced fat accumulation in the liver. We conclude that: 1) greater oxidative stress coupled with inactivation of AMPK may be critical for the additive effects of nicotine and HFD on the severity of hepatic steatosis in obese mice and 2) lipolysis of abdominal fat is an important contributor to the accumulation of lipids in the liver. We surmise that nicotine plus HFD is likely to be a very toxic combination in patients. Obesity POSTERS

Supported by: Basic Science Research Program through the NRF Integrated Physiology/ 1784-P Effects of Lipotoxicity on Insulin Sensitivity and Mitochondrial Function During the Development of Hepatic Steatosis TOMAS JELENIK, GILLES SÉQUARIS, JULIA SZENDRÖDI, JÖRG KOTZKA, ESTHER PHIELIX, BRIGIT KNEBEL, PETER NOWOTNY, HANS-JOACHIM PARTKE, DIRK MÜLLER-WIELAND, MICHAEL RODEN, Düsseldorf, Germany Non-alcoholic fatty liver (NAFL) and ectopic lipid storage in skeletal muscle Supported by: R24DA017298 have been associated with insulin resistance and abnormal mitochondrial function in obesity and type 2 diabetes mellitus. The underlying causal rela-

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A459 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1786-P protein expressed by hepatocytes which upregulates MMPs, are induced in Portal Vein Glucose Entry Triggers a Coordinated Cellular Response fi brosis but their role in NASH progression in diabetes is unclear. How hepa- that Potentiates Hepatic Glucose Uptake and Storage in Dogs tocyte CD147 induces stellate cell (HSC) MMPs is also not known. Therefore KATIE C. COATE, GUILLAUME KRAFT, MARTA S. SMITH, JOSE M. IRIMIA, BEN we investigated in vivo and in vitro the effect of lipid and glucose on liver FARMER, DOSS W. NEAL, WANDA SNEAD, PETER J. ROACH, MASAKAZU SHIO- MMPs and CD147. The ability of hepatocyte CD147 to induce HSC MMPs TA, ALAN D. CHERRINGTON, Dallas, TX, Nashville, TN, Indianapolis, IN was also studied. C57BL/6 male mice were fed a high fat diet (HFD:45%kCal Our goal was to identify the molecular events linking the pleiotropic ac- fat) or chow (CHOW:12%kCal fat) at 15wks some animals were made dia- tions of portal vein glucose delivery to increased hepatic glucose uptake betic (CHOW+DM or HFD+DM: STZ, 2x65mg/kg) and at termination (37wks) (HGU) and glycogen synthesis (GSYN) in vivo. Hyperinsulinemic hyperglyce- liver or hepatocytes were obtained. The cellular localisation of MMP activity mic (HIHG) clamps with (+) or without (-) portal vein glucose (PoG) infusion was studied by in situ zymography and MMP and CD147 were co-localised were conducted on 18-h-fasted conscious dogs (n=5/group) two weeks after by immunohistochemistry. Their gene and protein levels were measured by portal/hepatic vein catheterization. Experiments had a 100-min equilibra- qRT-PCR and Western immuno-blot and MMP activity was measured fl uori- tion, a 20-min control, and two 90-min clamp periods (P1 and P2). In P1 and metrically. In vitro an anti-CD147 mAb was used to examine the ability of P2, insulin and glucagon were infused intraportally at four-fold basal and CD147 either cell membrane bound (CM) or secreted in microparticles (MP) basal rates, respectively, while the hepatic glucose load (HGL) was doubled to induce HSC MMPs. By in situ zymography MMP activity localised to he- by a peripheral glucose infusion. In P2, saline (-PoG) or glucose (+PoG) was patocytes and the fi brotic septa, in contrast CD147 expression was only in infused intraportally, the latter to activate a feeding signal. Liver biopsies hepatocytes. Compared with controls HFD+DM increased hepatic and he- were obtained prior to and at the end of the clamp. The addition of portal patocyte MMP activity, MMP 2, 3, 9 and 14 mRNA, as well as CD147 mRNA vein glucose infusion to HIHG, without a change in HGL, triggered a signifi - and protein (P<0.05). CD147 was also increased by HFD alone. In vitro lipid cantly greater increase in the expression and activity of hepatic glucokinase increased hepatocyte CD147 expression and secretion in MPs(P<0.05) but (GK) than saline infusion (GK mRNA and activity, 26-fold basal and 5.0±0.3 had no effect on MMPs. Hepatocyte CM and MP increased HSC MMPs 1, U/g liver in +PoG vs. 6-fold basal and 3.1±0.2 U/g liver in -PoG, respectively). 2 and 14 an effect prevented by addition of anti-CD147 mAb (all P<0.05). In Augmented GK activity was associated with greater HGU in the +PoG group conclusion MMPs and CD147 are increased by HFD+DM induced NASH and (3.0±0.3 vs. 1.6±0.1 mg/kg/min in +PoG and -PoG, respectively; P < 0.05). in a pattern similar to other chronic liver diseases. Our results also impli- In addition, the glycogen synthase to phosphorylase activity ratio was in- cate hepatocyte CD147 in induction of MMPs in HSC. Together these results creased to a signifi cantly greater extent with portal vein glucose infusion suggest that the hepatocyte CD147 plays a key role in orchestrating matrix than saline infusion (10-fold basal vs. 6-fold basal in +PoG and -PoG, respec- remodelling by MMPs in NASH. tively). Accordingly, the increment in liver glycogen was greater in +PoG than Supported by: NH and MRC Australia -PoG (25±4 vs. 12±4 mg/g; P < 0.05). Thus, glucose sensing in the portal vein triggers an increase in the activity of GK and GS, which results in an augmen- 1789-P tation of HGU and GSYN, thereby ensuring the effi ciency of hepatic glucose Lowering Free Fatty Acid Levels Restores Central Regulation of Glu- clearance so as to minimize post-prandial glycemic excursions. cose Production in Type 2 Diabetes Supported by: NIH SYLVIA KEHLENBRINK, SUDHA KOPPAKA, DEEKSHA MEHTA, OANA SANDU, 1787-P YIXING LI, HARSHA JAYATILLAKE, DANIEL STEIN, PREETI KISHORE, MEREDITH Nateglinide Prevents SREBP1c Overexpression in HepG2 Cells via HAWKINS, Bronx, NY Suppression of LXR-Mediated Pathways Diazoxide (DZX) decreases endogenous glucose production (EGP) by ~30% SHIMPEI OGAWA, HARUKA KAWANABE, KYOKO MIURA, AKIKO OONUKI, AKIRA in nondiabetic humans, with complementary rat studies suggesting this is MITSUI, AKIRA MITSUI, YOSHIRO KITAHARA, Kawasaki, Japan, Kanagawa, Japan due to activation of hypothalamic KATP channels (JCI 121: 4916, 2011). Since Nateglinide (NT) is known to prevent the hepatic accumulation of triglyc- EGP is inappropriately increased in type 2 diabetes mellitus (T2DM), we erides (TG) in patients with type 2 diabetes. We previously reported that a examined whether this central regulation by diazoxide is intact. Moreover, single oral dose of NT suppressed postprandial de novo fatty acid synthe- since increased plasma free fatty acid (FFA) levels affect the regulation of sis via down-regulation of hepatic SREBP1c expression in KKAy mice. This glucose fl uxes, we determined whether lowering plasma FFA with nico- effect was only observed with NT among various glinides, indicating that tinic acid (NA) would impact the response to DZX in T2DM.We performed it was not a consequence of improved glycemic control and improvement paired 4-hour euglycemic ‘pancreatic clamp’ studies under basal hormonal 2 of postprandial insulin pattern. In the present study, we investigated the conditions in n=15 T2DM subjects (11M/4F, 50±2 yrs, BMI=30.2±1.3 kg/m , mechanism by which NT down-regulates hepatic expression of SREBP-1c HbA1c=9.7±0.5%). Three treatment conditions were compared to paired pla- both in vivo and in vitro. After an overnight fast, KKAy mice were given either cebo (PL) studies: DZX (6 mg/kg, n=8), DZX following overnight NA infusion the vehicle or NT (50 mg/kg) by oral gavage, and then were fed for 1 hr. In (DZX+NA; 0.01 mg/kg/min; n=5), and overnight NA infusion alone (n=7).DZX vehicle-treated mice, hepatic expression of SREBP1c, fatty acid synthase, alone failed to suppress EGP in these poorly controlled T2DM subjects. NA and acetyl-CoA carboxylase was up-regulated to about 3 times the level lowered FFA to ~100 μU/ml and restored suppression of EGP in response to seen before feeding at 2 or 3 hrs after feeding. Similarly, CYP7A1 was also DZX, comparable to that seen in nondiabetic individuals (DZX=1.52±0.13 vs. elevated about 2-fold. Oral administration of NT just before feeding signifi - DZX+NA=0.98±0.06 mg/kg/min, P=0.006). Lowering FFA with NA alone did cantly suppressed the elevation of SREBP1c and its downstream enzymes. not decrease EGP under these euglycemic conditions, presumably due to au- Up-regulation of CYP7A1 after food intake was also prevented by NT. Since toregulation of glucose fl uxes (JCI 86: 489, 1990).These studies demonstrate both SREBP1c and CYP7A1 are known to be up-regulated by LXR_ in rodents, loss of regulation of EGP by diazoxide under fi xed hormonal conditions in we next investigated the effect of NT on induction of SREBP1c expression T2DM and its restoration upon lowering plasma FFA levels. This suggests a by the LXR agonist T0901317 in HepG2 cells. T0901317 induced a dose- reversible loss of central regulation of EGP secondary to the chronic meta- dependent increase of SREBP1c expression with sub-maximal elevation of bolic defects of T2DM. SREBP1c expression by 4-fold above the basal level at a concentration of 0.3 μM. In this condition, NT at 10 μM or higher concentrations suppressed

Obesity T0901317-induced up-regulation of SREBP1c and its downstream enzymes.

POSTERS ABCA1, which is also regulated by LXR_, was suppressed by NT as well. These fi ndings indicate that NT suppresses LXR-mediated up-regulation of

Integrated Physiology/ hepatic gene expression. Since expression of LXR_ per se was not affected by NT, this drug might suppress LXR-mediated transcriptional activity of SREBP1c and thus prevent accumulation of TG in the liver. Supported by: NIDDK 1788-P Hepatocyte CD147 Induces Matrix Metalloproteinases (MMPs) in a Model of NASH Progression in Diabetes AUVRO MRIDHA, ALISON MORGAN, JAMES BONNER, LISA LO, STEPHEN TWIGG, NICHOLAS SHACKEL, SUSAN MCLENNAN, Sydney, NSW, Australia We have reported that high fat diet combined with diabetes increases NASH progression and fi brosis in a mouse model. The MMPs and CD147, a

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A460 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1790-P 1792-P Metformin Prevents High-Glucose Induced Fetuin A Release from Betulinic Acid Inhibits Hepatic Glucose Production via Activation HepG2 Cells of AMP-Activated Protein Kinase Signaling Pathway RUDY J. VALENTINE, LAUREN E. DISE, JOACHIM H. IX, NEIL B. RUDERMAN, Bos- SOO JUNG KIM, HAI YAN QUAN, GO WOON KIM, DO YEON KIM, SUNG HYUN ton, MA, La Jolla, CA CHUNG, Seoul, Republic of Korea Fetuin A is a liver-secreted protein known to impair insulin signaling. Data In the present study, we aimed to determine the effect of betulinic acid from clinical studies and animal models suggest that plasma fetuin A levels (BA) on hepatic gluconeogenesis and its mechanism of action in vivo and in are elevated in patients with metabolic syndrome-associated diseases, in- vitro. The result shows that BA signifi cantly inhibited hepatic glucose pro- cluding diabetes, CVD and NAFLD. Despite this, studies of factors that limit duction and AMP-activated protein kinase (AMPK) phosphorylation in time- fetuin A production by the liver and the mechanism by which they are limited and dose-dependent manners. BA increased cAMP-regulated transcriptional may provide potential disease targets. It has been demonstrated that incu- co-activator2 (CRTC2) phosphorylation and decreased cAMP regulatory bation of human hepatoma (HepG2) cells in a high glucose medium dimin- element-binding protein (CREB) phosphorylation, triggering dissociation of ishes AMP-activated protein kinase (AMPK) and causes insulin resistance. CREB-CBP-CRTC2 complex and thus reduces gluconeogenesis enzyme gene In addition, the anti-diabetic drug metformin increases AMPK activity and expression. The effects were signifi cantly abolished by treatment with com- restores insulin sensitivity. We examined here whether hyperglycemia led pound C, the AMPK inhibitor. In animal study, we examined the effects of to high fetuin A release, and whether metformin protects against glucose-in- oral administration of BA on the plasma glucose level and hepatic expres- duced fetuin A production in HepG2 cells. Cells were incubated in either low sion of gluconeogenesis-related genes of ICR mice fed with high fat diet. BA glucose (5.5 mM) or high glucose (25 mM) +/- 2 mM metformin for 24 hours at 5 and 10 mg/kg signifi cantly lowered the blood glucose level by inhibition and fetuin A levels in the media were determined by ELISA. High glucose sig- of hepatic glucose production through activation of AMPK signaling path- nifi cantly increased (69%) fetuin A released into the media. The increase in way. In conclusion, this study suggested that BA ameliorates the hypoglyce- fetuin A release under high glucose was completed prevented by metformin mia via AMPK signaling pathway in the liver, and this highlights the potential (6.5 vs 2.8 μg/ml). Interestingly, metformin also signifi cantly reduced fetuin value of BA as a therapeutic agent for type 2 diabetes. A release under low glucose conditions (3.9 vs 2.4 μg/ml). Treatment with another AMPK activator, AICAR, revealed similar results, reducing fetuin A release by 30 and 58% in low and high glucose, respectively. Since AMPK is down-regulated by high glucose and activated by metformin we investi- gated whether cellular AMPK activity (p-AMPK (Thr 172), assessed by West- ern blot) was associated with fetuin A release. Metformin signifi cantly in- creased cellular p-AMPK, which was inversely correlated with media fetuin A (r = -0.66, p<0.05). In conclusion, these data suggest that hyperglycemia increases fetuin A production in HepG2 cells. They also demonstrate that metformin inhibits this effect, most likely by activating AMPK. Supported by: NIH

1791-P Metabolomic Profi le Associated With Hepatic Insulin Resistance in Nondiabetic Subjects: Results from the RISC Study AMALIA GASTALDELLI, ANDREA NATALI, WALTER GALL, MELANIA GAGGINI, BEVERLEY BALKAU, JOHN PETRIE, MARTINE LAVILLE, RAPHAEL GABRIEL SAN- CHEZ, CHRISTIAN ANDERWALD, ELE FERRANNINI, RISC INVESTIGATORS, Pisa, Italy, Durham, NC, Villejuif, France, Glasgow, United Kingdom, Lyon, France, Madrid, Spain, Vienna, Austria Hepatic insulin resistance (Hep-IR) is the major determinant of elevated fasting plasma glucose (FPG) and therefore characterizes subjects at risk for type 2 diabetes. Since the assessment of Hep-IR requires tracer infusion, we Supported by: Basic Science Research Program through the NRF used untargeted global metabolomic analysis to identify potential biomark- ers in a nondiabetic population. The RISC cohort comprised 1,222 nondia- betic, normotensive European subjects (age 44±8 years (mean±SD) [range 1793-P 30-60], BMI 25.5±4.1 kg/m2, [range 16.9-43.9]) phenotyped for glucose tol- Characteristics of Non-Obese Japanese Men With Muscle and/or erance by OGTT, peripheral insulin resistance (by euglycemic hyperinsuline- Liver Insulin Resistance mic clamp), and Hep-IR index (validated against tracer infusion in a subgroup KAGEUMI TAKENO, YOSHIFUMI TAMURA, MINAKO KAWAGUCHI, TAKAHIRO of 380 subjects). In the entire cohort we measured by mass spectrometry a WATANABE, SATOSHI FUNAYAMA, YUKO SAKURAI, FUMIHIKO SATO, RISAKO panel of metabolites and correlated them with Hep-IR index, liver enzymes YAMAMOTO, MAENGKYU KIM, SAORI KAKEHI, SHIN-ICHI IKEDA, ADRIA GI- (AST/ALT), and fatty liver accumulation assessed using the recently validated ACCA, YOSHIO FUJITANI, TAKAHISA HIROSE, RYUZO KAWAMORI, HIROTAKA fatty liver index (FLI).After adjusting for center, sex, and age, the metabolites WATADA, Tokyo, Japan, Toronto, ON, Canada that remained independently associated with Hep-IR index by multiple linear Despite low body mass index (BMI), Asian people frequently develop type regression analysis were: _-hydroxybutyrate and creatine in a positive man- 2 diabetes. In addition to reduced insulin secretion, etiological difference ner (partial r = 0.12 and 0.17 respectively), and `-hydroxybutyrate, oleoyl- of insulin resistance (IR) between Caucasian and Asian might be involved LPC, AST/ALT, linoleoyl-lysophosphocholine (LPC), and glycine in a negative in this phenomenon. Recent data suggested that intrahepatic lipid (IHL) of manner (partial r = -0.16, -0.15, -0.15, -0.13, -0.07, p=0.01-0.0001). Similar non-obese Japanese subjects is higher than Caucasian, that may contribute

correlations were found also when ln(FLI) was used as dependent variable: to hepatic IR. To elucidate these mechanisms, we recruited 80 non-obese Obesity

_-hydroxybutyrate, creatine, and palmitoyl-LPC correlated in a positive (BMI<25kg/m2), non-diabetic Japanese men. Using a 2-step hyperinsuline- POSTERS manner (partial r = 0.12, 0.15 and 0.07), `-hydroxybutyrate, linoleoyl-LPC, mic euglycemic clamp with glucose tracer, we evaluated IR in muscle and

AST/ALT, serine and glycine, in a negative manner (partial r = -0.23, -0.22, liver, respectively. We also measured intramyocellular lipid (IMCL), IHL and Integrated Physiology/ -0.20,-0.09, -0.05, p=0.01-0.0001).Conclusion. In nondiabetic subjects, me- visceral fat volume (VFV) by MRS and MRI. Based on IR in liver and muscle, tabolites associated with oxidative stress and metabolic overload are new we divided the subjects into 4 groups: no IR in muscle nor liver (CON, n=44); biomarkers for hepatic insulin resistance and fatty liver. IR only in liver (L-IR, n=7); IR only in muscle (M-IR, n=17); IR both in muscle Supported by: EU contract QLG1-CT-2001-01252, AstraZeneca and liver (LM-IR, n=12). Characteristics of L-IR were similar to CON, except for high alcohol consumption and increased blood pressure (BP). On the other hand, in M-IR, BMI, triacylglycerol and FFA were signifi cantly higher and HDL cholesterol and VO2max were signifi cantly lower than CON. Interestingly, IHL was 3 times higher than CON, while IMCL was similar. In addition to the characteristics of M-IR, LM-IR showed higher fasting blood glucose, higher BP and greater VFV than CON. These characteristics and the degree of IR

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A461 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

were similar to Japanese obese men with metabolic syndrome. Multivariate analysis revealed that only alcohol consumption was a signifi cant indepen- dent determinant of L-IR (r=0.41, P<0.001). In addition, simple linear regres- sion analysis indicated that IHL was best predictor of IR in muscle (r=-0.59, P<0.0001). These data provide the unique characteristics of non-obese Japa- nese men with liver and/or muscle IR and suggest the importance of alcohol consumption as a cause of hepatic IR. In addition, IHL is a useful marker for IR in muscle, rather than a cause of hepatic IR in non-obese Japanese men. Supported by: The Ministry of Education, Culture, Sports, Science, and Technol- ogy of Japan

1794-P Nonalcoholic Fatty Liver Disease Is Associated With Atherosclero- sis in Middle-Aged and Elderly Chinese YU XU, YUN HUANG, YUFANG BI, MIN XU, ZHIMIN MA, TIANGE WANG, MIAN LI, YU LIU, JIELI LU, YUHONG CHEN, FEI HUANG, BAIHUI XU, JIE ZHANG, WEIQING WANG, GUANG NING, Shanghai, China 1796-P To evaluate the associations between nonalcoholic fatty liver disease Differential Subcellular Phosphorylation of Hepatic Fatty Acid Syn- (NAFLD) and atherosclerosis, a total of 8,971 participants aged 40 years and thase Channels Lipids to Regulate PPARƴ in Response to Feeding older from Baoshan district, Shanghai were included in the present study. ANNE P. JENSEN-URSTAD, HAOWEI SONG, IRFAN J. LODHI, CLAY F. SEMENK- The presence of NAFLD was evaluated by ultrasonography. Carotid intima- OVICH, St. Louis, MO media thickness (CIMT) and brachial-ankle pulse wave velocity (ba-PWV) PPAR_, a ligand-activated nuclear receptor, is targeted by drugs used to were measured in each participant. The prevalence of NAFLD was 30.0% treat diabetic dyslipidemia; it is also required for ketogenesis during fast- in the total population, with that of 30.3% in men and 29.8% in women, ing. Fatty acid synthase (FAS) activates PPAR_ in the liver by promoting the respectively. Subjects with NAFLD had remarkably higher CIMT and ba-PWV synthesis of an endogenous lipid ligand. FAS is thought to be stimulated by as compared with those without NAFLD (0.594±0.105 mm vs. 0.578±0.109 nutrients and insulin, so its activation of PPAR_ during fasting when insu- mm, P<0.0001; 1665±425 cm/sec vs. 1558±430 cm/sec, P<0.0001). Subjects lin levels are low is paradoxical. We demonstrate that hepatic FAS exists with both NAFLD and metabolic syndrome had signifi cantly higher CIMT and in two distinct subcellular pools that are differentially regulated. One pool ba-PWV as compared with those with neither of or either of these two dis- is localized to the cytoplasm while a second consists of FAS as a periph- eases after adjustment for age and sex (P all <0.05). Logistic regressions eral membrane protein in the ER and Golgi. FAS protein does not appear also revealed that NAFLD conferred 29% and 20% increased odds ratios of to exchange between pools based on subcellular localization and pulse elevated CIMT and ba-PWV independent of conventional risk factors and the chase experiments. ER/Golgi-associated FAS is active with feeding in mice presence of metabolic syndrome. In conclusion, NAFLD was associated with and with addition of insulin in a cultured hepatocyte cell line. Conversely, elevated CIMT and ba-PWV independent of conventional CVD risk factors cytoplasmic FAS is active with fasting and inhibited by feeding or insulin and metabolic syndrome. The effects of NAFLD and metabolic syndrome on treatment. Mass spectrometry experiments showed that cytoplasmic FAS, atherosclerosis might not fully overlap. but not ER/Golgi FAS, is phosphorylated during feeding or insulin treatment at two sites, Thr-1029 and Thr-1033, that fl ank the Asp-1032 catalytic resi- due of the FAS dehydratase domain. Mutation of these sites to alanines in- 1795-P creased cytoplasmic FAS specifi c activity and promoted PPAR_ target gene Serotonin Transporter Availability Within the Diencephalon of expression. These fi ndings suggest that hepatic FAS is compartmentalized Healthy Lean Male Subjects Does Not Correlate With Insulin Sen- to allow for multiple physiological functions with distinct regulation. ER/ sitivity Golgi-associated FAS is active during feeding and likely produces fatty acids KARIN E. KOOPMAN, SUSANNE E. LA FLEUR, MARIËTTE T. ACKERMANS, ERIC channeled to triglyceride synthesis. Cytoplasmic FAS is active during fasting FLIERS, JAN BOOIJ, MIREILLE J. SERLIE, Amsterdam, The Netherlands and appears to channel lipids whose primary function is nuclear receptor An association between brain serotonergic systems and glucose metabo- signaling. Preferentially manipulating distinct FAS pools could represent a lism has become evident. Functional imaging studies in humans have shown novel approach to the treatment of fatty liver and hyperlipidemia in obesity that binding to the serotonin transporter (SERT) in the brain correlates with and diabetes syndromes. BMI, while modulation of the serotonergic system alters insulin sensitivity. However, the physiological role of SERT in glucose metabolism in healthy 1797-P lean subjects remains unknown.We therefore measured SERT and glucose Olmesartan Improves Hepatic Insulin Sensitivity via Reducing He- metabolism in 18 healthy male volunteers with a normal BMI and normal in- patic PGC-1alpha Expression in Zuker fa/fa Rats sulin sensitivity. We determined SERT availability in the diencephalic region HIROAKI SATOH, SATORU YAMAZAKI, YOSHIYUKI SUGAYA, NORITAKA MACHII, (including hypothalamus) by calculating specifi c-to-nonspecifi c (SNS) bind- TSUYOSHI WATANABE, Fukushima, Japan ing ratios using [123I]FP-CIT SPECT and performed a 2-step hyperinsulinemic Evidence has accumulated that some of the angiotensin II AT1 receptor euglycemic clamp with a labelled glucose tracer to measure peripheral and antagonists have insulin sensitizing property. However, the acute effect of hepatic insulin sensitivity.All subjects had a normal BMI (median 21.9 kg/ the angiotension II AT1 receptor antagonists on glucose and insulin metabo- m2 (range 19.6 - 25.7)). Basal endogenous glucose production (EGP) was 12.5 lism has not been fully elucidated. We thus examined olmesartan, which is (10.7-14) umol/kgFFM/min. Insulin-mediated suppression of EGP was 72.8 a kind of angiotensin II AT1 receptor antagonists, on glucose and insulin me- (47 - 89) %, while rate of glucose disappearance (Rd) was 67.4 (44-93) μmol/ tabolism were assessed in the insulin-resistant obese Zucker rats, whereby Obesity kg.min. Median SERT SNS in the diencephalon was 0.58 (0.36 - 1.19). There insulin sensitivity was measured directly using the hyperinsulinemic-eug- POSTERS was no signifi cant correlation between SERT SNS and basal EGP, EGP sup- lycemic glucose clamp studies (at 25 mU/kg/min insulin infusion rate) after pression or Rd.Our fi ndings suggest that in metabolically healthy lean male a 8 hours fast. Male obese Zucker rats were intravenously administrated Integrated Physiology/ subjects, neither basal glucose production nor hepatic and peripheral insulin either low dose olmesartan or vehicle for 6 hours. Blood pressure was no sensitivity are regulated by diencephalic SERT. Whether changes in SERT in difference between two groups during the clamp studies. In basal state, the presence of obesity are associated with changes in insulin sensitivity low dose olmesartan administration had a marked effect to lower fasting remains to be studied. glucose levels from 153.0 ± 4.1 to 142.6 ± 2.5 mg/dl. (-6.8%; P<0.05), and to lower basal hepatic glucose output (HGO) from 11.6 ± 0.5 to 7.2 ± 0.2 mg/kg/ min (-38%; P<0.01). During the clamp studies, the glucose infusion rate (Ginf) required to maintain euglycemia, and the insulin stimulated glucose disposal rate (ISGDR), were no difference between two group. However, the insulin suppression of HGO is increased in olmesartan administrated rats compared to control rats (76.4 ± 2.3 v.s. 62.3 ± 3.6 %; P<0.05). Consistent with the clamp data, the insulin-stimulated phosphorylation of Akt was signifi cantly

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A462 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER enhanced in liver (by 55%), but not in skeletal muscle. Furthermore, PGC-1_ 1800-P expression in liver, which is a negative regulator of hepatic insulin signaling, Loss of SIRT1 in Liver Suppresses FGF21 Expression and Inhibits was markedly reduced in olmesartan group compared with control group, Fatty Acid Oxidation and Ketogenesis, Leading to Decreased Energy as assayed by QRT-PCR. In conclusion, these results indicate that acute ef- Expenditure and Increased Susceptibility of Obesity in Mice fect of olmesartan has benefi cial effects on hepatic insulin sensitivity in an YU LI, KIMBERLY WONG, AMBER GILES, JONGWOO LEE, HUNG-CHUN CHANG, insulin resistant state. LEONARD GUARENTE, MENGWEI ZANG, Boston, MA, Cambridge, MA Fibroblast growth factor 21 (FGF21) is the hepatocyte-derived hormone 1798-P that regulates fatty acid metabolism and has potential to treat obesity and Role of Sdf2l1, a Novel ER Stress-Related Protein, in the Regulation diabetes. We recently indicate that hepatic overexpression of SIRT1 in dia- of Insulin Sensitivity betic mice attenuates hepatic steatosis and insulin resistance. However, the TAKAYOSHI SASAKO, KOHJIRO UEKI, MITSURU OHSUGI, NAOTO KUBOTA, KA- in vivo long-term consequence of hepatic SIRT1 ablation in liver physiology ZUYUKI TOBE, TAKASHI KADOWAKI, Tokyo, Japan, Toyama, Japan remains unknown. We showed that hepatocyte-specifi c SIRT1 knockout Dynamic changes in glucose metabolism occur in the liver during the (SIRT1 LKO) mice with the albumin Cre-loxP system exhibited a striking phe- transition between fasting and feeding, and dys-regulation of the processes notype with greater propensity for obesity on a chow diet, characterized by leads to metabolic disorders. To further elucidate these changes, we com- increased whole body mass and fat mass, reduced energy expenditure, and pared global gene expressions in livers of B6J mice in 24h-fasted and 6h- unaltered food intake and physical activity. The obese phenotypes of SIRT1 refed states by microarray analysis. Interestingly, several ER stress-related LKO mice were associated with reduced hepatic and circulating levels of genes were up-regulated by refeeding. Among them, we focused on Sdf2l1, fasting FGF21. Hepatic impairment of FGF21 repressed expression of key en- a component of chaperone complex, which showed the second largest in- zymes involving fatty acid oxidation such as CPT1_ and MCAD, and inhibited crease of all the genes. Indeed, expression of ER stress markers as well as expression of ketogenic enzymes including ACAT1, HMGCS2, HMGCL, and Sdf2l1 was elevated prominently and transiently after refeeding, while it BDH1, thereby reducing plasma `-hydroxybutyrate levels in SIRT1 LKO mice. was attenuated by feeding protein-free diet or treatment with STZ. In or- Moreover, transcriptional activity of a FGF21 promoter-driven luciferase der to explore the role of Sdf2l1, we investigated the promoter activity by reporter was stimulated by SIRT1 activators, resveratrol and SRT1720, in luciferase assay, indicating that it was regulated by XBP1 and ATF6 upon SIRT1+/+ MEFs, but not in SIRT1-/- MEFs. The ability of resveratrol and ER stress. Moreover, adenovirus-mediated knocking down of Sdf2l1 in the SRT1720 to stimulate FGF21 protein was abolished by SIRT1 H335A inactive liver resulted in greater elevation of ER stress markers after refeeding. It mutant or by nicotinamide and splitomicin in HepG2 cells. Induction of FGF21 also impaired glucose tolerance in OGTT, attenuated insulin signaling and in- by SIRT1 activators enhanced expression of key enzymes for fatty acid oxi- creased triglyceride content in the liver. These urged us to hypothesize that dation and ketogenesis. These in vivo and in vitro fi ndings characterize 1) dys-regulation of Sdf2l1 might contribute to insulin resistance in obesity. hepatic SIRT1 as a master regulator of FGF21; 2) SIRT1-dependent activation Actually in livers of db/db mice, although ER stress sensors were activated, of FGF21 in liver as a component for adaptive fasting response; and 3) defec- Sdf2l1 was down-regulated. By contrast, adenovirus-mediated restoration tive hepatic SIRT1 and FGF21 signaling as a key pathological determinant of of hepatic Sdf2l1 expression in db/db mice improved glucose tolerance, in- energy metabolic abnormality and obesity susceptibility. sulin signaling and fatty liver. These data suggest that feeding induces tran- Supported by: NIH/NIDDK RO1 sient ER stress physiologically in the liver, while Sdf2l1 regulated by XBP1 and ATF6 may switch off the ER stress response, maintaining hepatic insulin 1801-P sensitivity and fatty acid metabolism during feeding. On the other hand, sup- Inhibition of Hepatic Notch Signaling Reverses Obesity-Induced In- pressed expression of Sdf2l1 in obesity may contribute to the development sulin Resistance and Fatty Liver of systemic insulin resistance, providing the possibility of Sdf2l1 to be a UTPAL B. PAJVANI, DOMENICO ACCILI, New York, NY novel therapeutic target of obesity-induced diabetes. Notch receptors have traditionally been thought to regulate normal devel- Supported by: The Ministry of Education, Culture, Sports, Science, and Technology opment and then remain quiescent unless deregulated in cancer. We recent- of Japan ly demonstrated that Notch inhibition improves insulin sensitivity, partially through interaction with the transcription factor FoxO1, paving the way to 1799-P exploit Notch-based therapeutics from cancer biology for use in obesity and Evaluation of the Liver Ketogenesis During Hypoglycemia Induced diabetes. To test the hypothesis that hepatic Notch inhibition can indepen- by Insulin in Alloxan-Diabetic Rats dently promote insulin sensitivity, we generated mice with a liver-specifi c FABIANA M. SCHIAVON, HELENTON C. BARRENA, MARCIA R. BATISTA, VAN- deletion of the Notch transcriptional effector, Rbp-Jk (L-Rbpj). L-Rbpj mice ESSA R. VILELA, ROBERTO B. BAZOTTE, Maringá, Brazil are protected from obesity-induced insulin resistance and demonstrate de- Insulin and the counterregulatory hormones released during long term in- creased hepatic glucose production. L-Rbpj mice are additionally protected sulin-induced hypoglycemia (LT-IIH) inhibits and activates the liver ketogen- from obesity-induced fatty liver, despite similar body weight and adiposity esis, respectively. Thus a question can be raised: which effect predominate as control animals. The combination of improved glucose homeostasis and during LT-IIH? To clarify this question the liver ketogenesis, glycogenolysis, hepatic lipids in L-Rbpj mice suggested that therapeutic inhibition of hepatic gluconeogenesis, ureagenesis and the production of L-lactate were mea- Notch signaling would be benefi cial in diabetic patients. Consistent with our sured, and their correlation with blood levels of ketone bodies (KB), L-lactate, fi ndings in L-Rbpj mice, pharmacologic inhibition of Notch signaling in vivo glucose and urea were investigated in male Wistar alloxan-diabetic rats and with gamma-secretase inhibitors (GSIs) markedly improved glucose toler- compared with nom-diabetic rats. Diabetes (DBT group) was induced with ance in wildtype, obese mice. This insulin-sensitizing effect of GSIs was ab- an intravenous injection of alloxan (40 mg/kg). LT-IIH was obtained with an sent in L-Rbpj mice, confi rming that hepatic Notch signaling is the tissue and intraperitoneal injection of Detemir insulin (1 U/kg). Blood KB, glucose, L- molecular target of GSIs. Unexpectedly, we observed that GSI-treated mice lactate and urea were evaluated at 0, 2, 4, 6, 8 or 10 after insulin injection. develop fatty liver. GSI-treated L-Rbpj mice were similarly susceptible to fat- Because LT-IIH was well established 2 h after insulin injection this time was ty liver, suggesting that while the glucose-lowering effect of GSIs is Notch- used for liver perfusion experiments where the production of KB, L-lactate, dependent, the steatotic effect of GSIs is likely Notch-independent. Thus,

urea and pyruvate were evaluated. In addition, the ketogenic capacity was we are now pursuing three parallel paths: (i) understanding the mechanism Obesity measured during the infusion of octanoate (0.3 mmol/L). LT-IIH decreased of GSI-induced steatosis to unravel its pathogenesis; (ii) exploring different POSTERS (P<0.05) blood KB and glucose levels, but there was an increased lactate- classes of Notch inhibitors for their clinical application; and (iii) determining

temia and a rebound increase in ketonemia during LT-IIH. Moreover, the ca- whether hepatic Notch signaling is altered in human obesity and diabetes. Integrated Physiology/ pacity to produce KB from octanoate was increased in the livers of DBT rats. Supported by: NIH/NIDDK (1K08DK093604) and The Louis Gerstner, Jr., Foundation Te elevated lactatemia in DBT rats could be attributed to the higher (P<0.05) glycogenolysis when part of glucose from glycogenolysis enters in the glyco- 1802-P lysis, producing L-lactate. In contrast, except glycerol, liver gluconeogenesis Regulation of Adipose Triglyceride Lipase (ATGL) and GOS2 Lipid was negligible in DBT rats. LT-IIH increased the liver ketogenic capacity and Droplet Proteins by FoxO Transcription Factors and Insulin in the the blood lactate levels in DBT rats increasing the risk of ketoacidosis and Liver lactic acidosis, respectively. WENWEI ZHANG, SO YOUNG BU, INSUG O-SULLIVAN, C.R. KAHN, DOUGLAS G. Supported by: CNPq (Grants 563870/2010-9) and Araucaria Foundation MASHEK, TERRY G. UNTERMAN, Chicago, IL, St. Paul, MN, Boston, MA FoxO transcription factors are major targets of insulin action. In the liver, FoxOs proteins promote gluconeogenesis and contribute to regulation of

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A463 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

glycolytic and lipogenic gene expression (JBC 281:10105, 2006). Studies in ing these functions are unclear. We show here that in the human HepG2 cell lower organisms show FoxOs promote lipid mobilization in adipose tissue line, GLP-1(28-36)amide treatment increased the phosphorylation of cAMP through regulation of adipose triglyceride lipase (ATGL), and recent fi ndings response element-binding protein (CREB) at Ser133. In addition, GLP-1(28-36) indicate that ATGL is important in regulating lipid turnover in liver. We asked amide repressed the expression of two rate-limiting gluconeogenic enzymes, whether FoxO proteins also regulate of ATGL and its inhibitor, the G0/G1 phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase switch gene protein 2 (G0S2), in liver. Studies in wild type mice show liver (G6Pase), and repressed the expression of a luciferase reporter driven by ATGL mRNA and protein levels are abundant after an 18 hr fast and mark- PEPCK promoter. In primary mouse hepatocytes, GLP-1(28-36)amide re- edly suppressed 6 hr after refeeding. Studies in liver-specifi c knockout mice pressed glucose production, associated with reduced expression of PEPCK show FoxO proteins are required for increased expression of ATGL in fasting. and G6Pase. The repression of glucose production along with the activation Conversely, transgenic mice expressing constitutively active FoxO1 in liver of CREB by GLP-1(28-36)amide are contradictory to the well-characterized show that suppression of FoxO function is required for reducing ATGL ex- signaling cascade of glucagon involving PKA/CREB in stimulating glucose pression and triglyceride lipase activity after refeeding. Gene array and ex- production in response to starvation. Our observations, however, are analo- pression studies reveal constitutively active FoxO1 also suppresses hepatic gous to the actions of glucagon and GLP-1 in similarly activating cAMP/PKA expression of G0S2, an inhibitor of ATGL. In vitro studies confi rm that FoxO1 signaling, yet resulting in opposite effects in regulating glucose homeosta- stimulates ATGL and suppresses G0S2 expression in isolated hepatocytes. sis. The identifi cation of the receptor that mediates the effect of GLP-1(28- To determine if FoxO proteins mediate effects of insulin on ATGL and G0S2 36)amide in hepatocytes will enhance our understanding of cAMP signaling expression, we examined ATGL and G0S2 expression in liver-specifi c insulin and its complexity in the regulation of glucose production and homeostasis. receptor knockout mice (LIRKO), and LIRKO mice in which FoxO1 also was Supported by: Canadian Institutes of Health Research inactivated in the liver (LIRFKO). ATGL expression was increased 4-fold in LIRKO mice compared to control and this effect was lost in LIRFKO mice. 1805-P Conversely, G0S2 expression was decreased by 80% in LIRKO mice, and Loss of Hepatic Rho-Kinase Ameliorates Insulin Resistance and Pre- disrupting FoxO1 partially reversed this effect. These studies indicate that vents the Development of Hepatosteatosis in Diet-Induced Obese FoxO proteins play an important role in regulating ATGL and G0S2 expression Mice and thereby mediate the ability of insulin to suppress lipid catabolism and HU HUANG, SEUNG-HWAN LEE, INES S. LIMA, IL JI PAEK, YOUNG BUM KIM, promote lipid storage in the liver. Boston, MA Non-alcoholic fatty liver disease is common and associated with obesity 1803-P and insulin resistance, ultimately leading to cirrhosis and its complication. Correction of Postprandial Hyperglycemia (P-HG) by Inhibiting However, the pathogenesis of this disease is not fully understood. To test SGLT2 Normalizes Glucose-Induced Glucokinase (GK) Transloca- the possibility that Rho-kinase 1 (ROCK1) regulates hepatic glucose homeo- tion (GK-T) from the Nucleus (N) to the Cytoplasm (C) in Liver at an stasis and lipid metabolism, mice lacking ROCK1 in liver were studied. Under Early Stage of Diabetes (ESD) in Zucker Diabetic Fatty Rats (ZDF) a normal chow diet, deletion of hepatic ROCK1 in mice enhanced systemic GREGORY A. MCCOY, TRACY P. O’BRIEN, ERIN C. HEALEY, ANTONIO CASTANE- insulin sensitivity, as revealed by lower insulin levels in the fasted state DA, KIICHIRO UETA, RICHARD L. PRINTZ, MASAKAZU SHIOTA, Nashville, TN and an increase in the area above the curve during an insulin-tolerant test. Impaired action of glucose and/or insulin at an ESD in ZDF leads to defec- These effects were independent of changes in adiposity. Consistent with tive GK-T and failure to suppress endogenous glucose production (EGP). We this, mice with liver-specifi c knockout of ROCK1 remained insulin sensitive sought to determine if these defects are linked to chronically elevated P-HG. under a high-fat diet (12 weeks treatment). In addition, the ability of insu- Male ZDF and lean control rats (ZCL) were catheterized at 8 wks of age; then lin to stimulate insulin receptor and Akt phosphorylation was increased in at 9 wks, canaglifl ozin (CA, a SGLT2 inhibitor; 10 mg/kg body weight (BW), these mice. Interestingly, liver-specifi c deletion of ROCK1 prevented high fat- q.d.) or vehicle (V) was given for 1 wk. Mixed meal tolerance tests (MTT; 4.7 induced obesity (Male, control mice 50.9 ±3.7, Liver-ROCK1 defi cient mice g glucose, 2.2 g protein and 0.6 g lipid per kg BW) and glucose-pancreatic 38.9±3.2g, p<0.01), along with a signifi cant decrease in fat mass assessed clamp studies using tracers ([3- and 2-3H]-glucose) were performed 6 h after by a MRI. Histological analysis indicated that hepatic steatosis by high-fat the last dosing in 6-h fasted conscious rats.Compared to ZCL, ZDF-V exhib- feeding was signifi cantly reduced in the absence of hepatic ROCK1. Fur- ited near normal basal levels of plasma glucose (B-PG, 7.9 ± 0.5 mM in ZDF-V thermore, hepatic triglyceride and total cholesterol in liver-specifi c ROCK1 vs 6.3 ± 0.2 mM ZCL) but elevated plasma insulin (B-PI, 8.4 ± 1.0 vs 0.9 ± 0.2 defi cient mice were also greatly decreased by ~40 -50%, compared with ng/ml), MTT-PG (AUC, 545 ± 82 vs 155 ± 25 mM/2h) and MTT-PI (AUC, 752 ± 1 control mice. However, mitochondrial functions in hepatocytes, including 45 vs 330 ± 66 ng/ml*2 h). These parameters in ZDF-CA were lowered to oxygen consumption and oxidative phosphorylation, were unaltered by loss near normal levels (B-PG, 6.2 ± 0.5 mM; B-PI, 3.3 ± 0.6 ng/ml; MTT-PG, 240 ± of ROCK1. These data demonstrate that genetic deletion of ROCK1 in liver re- 48 mM/2h; MTT-PI, 355 ± 72 ng/2 h). Basally, N localization of GK and GKRP, sults in hypersensitivity to insulin and protects against diet-induced obesity EGP and glucose cycling (GC) did not differ between ZDF-V and ZDF-CA. But and hepatosteatosis in mice. Thus, our studies identify ROCK1 as important in response to a hyperglycemic (HG, 13.5 mM)-hyperinsulinemic (HI, 15 ng/ regulator of insulin sensitivity and fat metabolism in liver. ml) clamp, in ZDF-CA vs ZDF-V, EGP was suppressed (-26 ± 11 vs 23 ± 9 μmol/ kg/min) with increased GC (93 ± 17 vs 46 ± 7 μmol/kg/min) and GK-T (N/C, 1806-P 1.19 ± 0.06 vs 1.54 ± 0.08). In ZDF-CA, the suppression of EGP (-4 ± 7 μmol/ Mechanisms of Glucose Lowering With DPP-4 Inhibitor Sitagliptin kg/min) with increased GC (89 ± 12 μmol/kg/min) and GK-T (N/C, 1.13 ± 0.08) Used Alone or With Metformin in T2DM: A Double-Tracer Study were also seen in response to a HG (13.5 mM)-euinsulinemic (4 ng/ml) clamp CAROLINA SOLIS-HERRERA, CURTIS TRIPLITT, JESUS GARDUÑO-GARCIA, JOHN but not a euglycemic (6 mM)-HI (15 ng/ml) clamp (HGP, 45 ± 11 μmol/kg/min; ADAMS, RALPH DEFRONZO, EUGENIO CERSOSIMO, San Antonio, TX GC, 49 ± 11 μmol/kg/min; GK-N/C, 1.48 ± 0.09).Reduction of P-HG with CA To assess glucose lowering mechanisms of sitagliptin(S), metformin(M) treatment at an ESD in ZDF lowers mixed meal-induced PG excursions and or in combination (S+M), we randomized 8 T2DM (age~52yrs, 4M/4F, 1yr restores HG-induced GK-T improving hepatic glucose fl ux. 2 duration, BMI~32 kg/m , HbA1C=8.4±1.2%) to 4 therapy periods (6wks + 2wk washout) with either Placebo (P), (M), (S) or (S+M) at maximum dos-

Obesity 1804-P es. After each 6wk period, they received a meal tolerance test (MTT, 600

POSTERS GLP-1-Derived Nonapeptide GLP-1(28-36)amide Reduces Hepatic kcal, 20g protein, 25g fat and 75g glucose) labeled with [14C-glu] to calcu- Glucose Production and Represses the Expression of Gluconeo- late oral glucose appearance rates (RaO); [3H-glu] IV infusion for glucose

Integrated Physiology/ genic Genes appearance rates (RaT) & disappearance (RdT). Endogenous glucose pro- WILFRED IP, WEIJUAN SHAO, TIANRU JIN, Toronto, ON, Canada duction (EGP) equals RaT-RaO. Fasting plasma glucose (FPG), insulin (FPI), The incretin hormone glucagon-like peptide-1 [GLP-1, GLP-1(7-36)amide or glucagon (FPGn) and post-MTT levels were measured over 360min. FPG GLP-1(7-37)] is encoded by the proglucagon gene and produced by intestinal decreased from 165±10 (P) to 149±9 (S), to 145±6 (M) and to 125±5 mg/dl endocrine L cells. Its release into the circulation in response to food intake (S+M); mean post-MTT levels in (P) of 208±15 mg/dl decreased to 182±13 facilitates its primary role in stimulating insulin secretion from pancreatic (S), 181±11 (M) and 155±9 (S+M) [p<0.01]. Basal EGP fell from 2.1±0.1 (P) to beta-cells. It has been suggested that GLP-1(28-36)amide is a cleavage prod- 1.8±0.1 (S), 1.9±0.2 (M) and to 1.5±0.1 mg/kg.min (S+M) [p<0.05]. Mean RaT uct of GLP-1(7-36)amide and possesses properties in lowering weight gain post-MTT was lower in (S)=2.5±0.1, (M)=2.7±0.1 and (S+M)=2.3±0.1 than in and attenuating diabetes and hepatic steatosis in diet-induced obese mice. (P)=2.9±0.2 mg/kg.min [p<0.05]. RaO was similar after all therapies (~1.9-2.3 In mouse hepatocytes, GLP-1(28-36)amide attenuates cAMP/dexametha- mg/kg.min), so the decline in RaT was explained by a drop in EGP post-MTT sone-stimulated glucose production, while molecular mechanisms underly- from 0.8±0.1 (P) to 0.6±0.1 (S), 0.4±0.1 (M) and 0.4±0.1 (S+M) [p<0.05]. Mean

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A464 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

RdT post-MMT was equal (~2.1-2.3 mg/kg.min). FPI did not change (~12 μU/ revealed HP20 exists mainly in choroid plexus, heart, kidneys and testes, but ml), and mean post-MTT insulin increased after (S+M)=76±9 vs. (P)=51±9, not in the spleen. Furthermore, stimulation with purifi ed HP20c increased (S)=52±8 and (M)=56±3 μU/ml [p<0.05]. FPGn did not change (~70 pg/ml), but phosphorylation of Akt in BAEC. Conclusions: Bovine HP20c is likely a se- a greater suppression of plasma glucagon post-MTT was seen in (S)=26±3% creted protein complex produced in the liver and delivered to specifi c organs and (S+M)=32±3% than in (P)=14±2% and (M)=13±4% [p<0.05]. Sitagliptin via the bloodstream. The bovine HP may be a useful tool for investigating therapy reduces FPG and post-meal glucose, which are further enhanced in HP function. combination with metformin. These are explained by a fall in basal EGP and more effi cient EGP suppression. The benefi cial effects of sitagliptin on EGP 1809-P are the result of a decline in postprandial plasma glucagon [(S) & (S+M)] and Hepatic Insulin Resistance is a Predictor of Cardiometabolic rise in plasma insulin [(S+M]. Disease Supported by: Merck and Co. FRANCESCO ANDREOZZI, AMALIA GASTALDELLI, GAIA MANNINO, SUBHASH KAMATH, ELENA SUCCURRO, FRANCO ARTURI, GIUSEPPE DANIELE, ANGELA 1807-P SCIACQUA, FRANCESCO PERTICONE, FRANCO FOLLI, GIORGIO SESTI, San Anto- Regulation of Fatty Acid Oxidation by Phosphatidylcholine Transfer nio, TX, Catanzaro, Italy, Pisa, Italy Protein (PC-TP): Potential Role for PC-TP-Thioesterase Superfamily Hepatic insulin resistance (Hep_IR) is a strong risk factor for the develop- Member 2 (Them2) Interactions ment of fasting hyperglycemia and type 2 diabetes. Moreover, subjects with YUKI KAWANO, DAVID E. COHEN, Boston, MA T2D and Hep_IR are more prone to develop non alcoholic fatty liver disease PC-TP is a phosphatidylcholine binding protein, which interacts with (NAFLD) and cardiovascular diseases (CVD). However, the assessment of Them2, a mitochondria-associated fatty acyl-CoA thioesterase. In mouse Hep_IR is not always feasible since it requires tracer infusion. The aim of hepatocytes and HEK 293E cells, endogenous Them2 expression promotes this study was to validate Hep_IR as a marker of presence of cardiometabol- fatty acid oxidation. This study was designed to ascertain whether PC-TP ic disease. Hep_IR (by Vangipurapu index recently validated vs tracers) was regulates rates of fatty acid oxidation and whether regulation is infl uenced calculated in a large group of subjects (n=756, age 42±12y, BMI 30±7 kg/m2, by Them2 expression. Primary hepatocytes were isolated from Pctp+/+ and mean±SD), CATAMERIS study, characterized for glucose tolerance by OGTT, Pctp-/- mice. PC-TP was inactivated using compound A1, a small molecule peripheral insulin resistance (by euglycemic hyperinsulinemic clamp, M val- inhibitor that was previously developed in our laboratory. Knockdown of ue, and Matsuda index), hepatic steatosis by ultrasound and fatty liver index endogenous PC-TP expression in HEK 293E cells was achieved using siRNA (FLI), liver enzymes (AST, ALT, GGT), lipid profi le, blood pressure (SBP and technology. Fatty acid oxidation rates were determined by exposing cells to DBP) and intima media thickness (IMT). The population comprised 492 NGT 14C-palmitic acid and then measuring the 14C-contents of acid soluble metab- subjects, 75 IFG, 75 IGT, 76 IFG/IGT and 38 T2D. Hep_IR was signifi cantly as- olites (ASM) and CO2. Protein expression levels were evaluated by immunob- sociated, after adjustment for gender and age, with peripheral IR measured lot analysis. Changes were considered signifi cant for P < 0.05. In Pctp-/- hepa- as Matsuda index (r=-0.733) and M value (r=-0.255), waist circumference 14 tocytes, C-incorporation rates into ASM and CO2 were decreased by 29% (r=0.611), both fasting (r=0.153) and 2h (r=0.263) glucose concentrations, all and 43%, respectively. In the absence of PC-TP expression, protein levels p<0.0001. Higher values of Hep_IR were associated with presence of he- of Them2 were similarly decreased by 40%. Without altering expression of patic steatosis and FLI (r=0.708) and increased liver enzymes, AST (r=0.139), Them2, exposure of Pctp+/+ hepatocytes to compound A1 resulted in a dose- ALT (r=0.343), GGT (r=0.126), all p<0.0001. Moreover, Hep_IR was associated 14 dependent increases in the C-incorporation rates into ASM (EC50 = 8.7 nM) with markers of CVD such as SBP and DBP (both r=0.256), CRP (r=0.238), and CO2 (EC50 = 9.1 nM) by up to 14% and 32%, respectively. Compound A1 fi brinogen (r=207), all p<0.0001, LDL (r=0.083, p<0.03) and with IMT, even did not infl uence fatty acid oxidation rates in Pctp-/- hepatocytes. Knockdown after correcting for gender, age, waist, SBP and LDL (partial r=0.214). The of PC-TP in HEK 293E cells, which did not alter Them2 expression, increased multivariate analysis for IMT was not signifi cant when Matsuda index was 14 C-incorporation rates into ASM and CO2 by 13% and 41%, respectively. We substituted to Hep_IR. In conclusion Hep_IR index is strongly associated conclude that reduced fatty acid oxidation rates in Pctp-/- hepatocytes were with metabolic and CVD factors and could be used for identifying subjects at likely attributable to decreased expression of Them2. Based on increased risk for developing cardiometabolic disease. oxidation rates following selective inhibition or knockdown of PC-TP, we speculate that PC-TP limits fatty acid oxidation in hepatocytes via interac- 1810-P tions with Them2. Regulation of Hepatic de novo Lipogenesis Supported by: NIH (R01DK48873, R01DK56626) QING CHEN, JOSHUA D. RABINOWITZ, MORRIS J. BIRNBAUM, Philadelphia, PA, Princeton, PA 1808-P Hepatic de novo lipogenesis is upregulated during insulin resistance, con- Identifi cation of Bovine Hibernation-Specifi c Protein Complex and tributing to the development of metabolic disorders such as hepatic steato- Evidence of its Regulation in Fasting and Aging sis and Type II diabetes. In recent years, the signaling pathway that controls SATOSHI FUJITA, RYUJI OKAMOTO, MASAYA TANIGUCHI, KATSUYA KONISHI, lipogenesis in response to insulin and nutrients has been extensively char- ITARU GOTO, KAZUSHI SUGIMOTO, MASHIO NAKAMURA, KATSUYA SHIRAKI, acterized. However, dysregulation in the biochemical pathway that occurs CHRISTA BUECHLER, MASAAKI ITO, Tsu, Japan, Regensburg, Germany during insulin resistance and consequently causes an abnormal increase Background: Hibernators such as chipmunks can tolerate hypothermic, in lipogenesis has yet to be identifi ed. One enzymatic step of particular hypoglycemic and ischemic stresses. Hibernation-Specifi c Protein (HP) is a interest is pyruvate dehydrogenase (PDH), where pyruvate is irreversibly plasma protein that regulates hibernation in chipmunks and ground squirrel converted to acetyl-CoA , the building block of fatty acids. We measured hibernators. The HP complex (HP20c) consists of three homologous proteins, de novo lipogenesis in primary mouse hepatocytes using 14C tracers and HP20, HP25 and HP27, which are all produced by the liver and belong to the concluded that lactate, rather than glucose, is the major lipogenic substrate. C1q family. We assessed the hypothesis that non-hibernating species such When hepatocytes isolated from fasted, fed, and leptin-defi cient (ob/ob) as cattle and human possess similar proteins to chipmunk HP. Methods: We animals are cultured under the same condition, ob/ob hepatocytes exhibit cloned a bovine HP20 gene from liver and bovine aortic endothelial cells the highest lactate-to-lipid fl ux, suggesting that PDH fl ux is dysregulated

(BAEC) and developed specifi c antibodies against bovine HP20. Native HP20 in diabetic models. In addition, we labeled hepatocytes with U-13C lactate Obesity was purifi ed from the bovine serum. We identifi ed binding proteins with and examined the relative PDH fl ux using an isotopomer analysis of citrate. POSTERS HP20 using mass spectrometry (MS), performed the immunostaining of mul- Using the ratio of the C5 citrate to the total amount of citrate produced from

tiple organs and evaluated the effect of bovine HP20c on BAEC. We exam- C3 oxaloacetate [C5 citrate: (C5 citrate + C3 citrate)], our analysis revealed Integrated Physiology/ ined the effect of fasting on the expression of HP20 in bovine. Results: The that even during fasting when PDH is believed to be inactive, there is a small bovine HP20 has 12 Gly-XY repeats followed by the C terminus having high percentage of C2 acetyl-CoA, indicating that a low level of PDH fl ux is main- similarity with adiponectin, collagens X and VIII and C1q. The total homology tained. More surprising, during feeding, when fatty acid oxidation is thought between bovine and chipmunk is 63%. The gene expression was highest in to be inactive, we found that there is a pool of unlabeled acetyl-coA, indicat- the liver and mild in the pancreas and kidneys. The native HP20 was purifi ed ing that signifi cant fatty acid oxidation remains or there is an alternative from serum as a complex containing 25kDa and 33kDa proteins. MS revealed source of acetyl-CoA under this condition. These data suggest the need for they were the orthologues of chipmunk HP25 and HP27. WB showed bovine a reevaluation of hepatic carbon fl ux into the citric acid cycle during various HP20 exists in fetal bovine, calf and adult serum, with the highest concen- nutritional and pathological conditions. tration found in adult serum. Interestingly, the concentration of HP20 was gradually increased when cattle were fasted. The immunostaining study

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A465 INTEGRATEDCATEGORY PHYSIOLOGY—LIVER

1811-P 1813-P A Plasma Metabolomic Fingerprint Differentiates Insulin Sensitive Inter-Species Differences Contributing to Poor Translatability of from Insulin Resistant Subjects With Nonalcoholic Fatty Liver Sustained Glucose Lowering Effects of Glycogen Phosphorylase RAINER LEHMANN, HOLGER FRANKEN, KONSTANTINOS KANTARTZIS, AN- Inhibition from Rodent to Human DREAS PETER, JÜRGEN MACHANN, FRITZ SCHICK, ERWIN SCHLEICHER, AN- XIAO WANG, STEVE WATERS, SHONNA MOODIE, ANURAAG KANSAL, San Ma- DREAS FRITSCHE, HANS-ULRICH HÄRING, NORBERT STEFAN, Tübingen, Ger- teo, CA, Bethesda, MD many Inhibitors of glycogen phosphorylase (GP), a key enzyme that regulates Nonalcoholic fatty liver (NAFL) predicts insulin resistance, subclinical in- the mobilization of glucose from glycogen, have demonstrated sustained fl ammation and dyslipidemia more strongly than general- or visceral obesity. glucose lowering in rodent models of type 2 diabetes (T2D). The therapeutic However, some individuals with NAFL, unexpectedly, do not develop such potential of GP inhibition for the treatment of T2D in human was therefore derangements of metabolism. The question now is, can broad systematic evaluated using Entelos’ Metabolism PhysioLab® platform, a large-scale blood screening approaches help to fi nd mechanisms and/or markers of the mathematical model of whole-body metabolism. We determined the mag- dissociation between NAFL and insulin resistance? We investigated param- nitude of GP inhibition required to lower A1C to a level competitive with cur- eters separating a metabolically benign from a malignant NAFL applying a rent standards of care (metformin) in a diverse group of T2D virtual patients targeted metabolomics approach.A total of 265 plasma metabolites were (VPs) over 4 weeks. Simulations predict that GP inhibition would not be clini- analyzed in 40 subjects with NAFL who were separated into two groups cally competitive (average A1C reduction is 0.15%). We subsequently inves- based on their insulin sensitivity. Liver fat content was measured by 1H- tigated the mechanisms underlying the predictions by creating “rodent-like MRS, and insulin sensitivity during a 75g OGTT before and after a lifestyle VPs” with appropriate physiological and behavioral changes. These changes intervention.At baseline, the insulin sensitive and the insulin resistant include 1) implementing a 10 fold increase in the capacity of hepatocytes to groups had almost identical mean liver fat content. At baseline, as well as store glycogen by altering feedback regulation of glycogen synthesis, and at follow-up, the lyso-phosphatidylcholines (lyso-PCs) separated the insulin 2) reproducing rodent-like frequent feeding behavior with nutrient composi- sensitive from the insulin resistant NAFL groups. The strongest correlations tions refl ective of rat chow. These results explain the success of the therapy with insulin sensitivity, both at baseline and at follow-up, were also found in rodents (average A1C reduction is 0.57%) and the physiological underpin- for lyso-PCs. The best separation of the two groups was achieved by a me- nings of the lack of successful drug development programs in GP inhibition. tabolite pattern including lyso-PC C16:0, ornithine, leucine, isoleucine, C3:0-, The ability to directly examine inter-species differences using in silico mod- C16:0- and C18:0-carnitine (ROC-AUC: 0.77, p=0.00023 at baseline and ROC- eling can be used to streamline therapeutic target validation and improve AUC: 0.80, p=0.000019 at follow-up).In conclusion, the lyso-PCs concentra- the success rate translating from pre-clinical studies to the clinic. tions differed most strongly between insulin sensitive and insulin resistant subjects with NAFL. These relationships were still apparent after 9 month of lifestyle intervention. Thus, in the future a metabolomic fi ngerprint, most robust in the class of lyso-PCs, may be able to differentiate insulin sensitive from insulin resistant subjects with NAFL and these fi ndings may highlight novel and interesting pathways for understanding the pathogenesis of insu- lin resistance in NAFL.

1812-P Role of Pin1 in the Pathogenesis of Non-Alcoholic Steatohepatitis in a Rodent Model YUSUKE NAKATSU, HIDEYUKI SAKODA, MIDORI FUJISHIRO, AKIFUMI KUSHI- YAMA, TOSHIAKI FUKUSHIMA, FUSANORI NISHIMURA, TAKAFUMI UCHIDA, TOMOICHIRO ASANO, Hiroshima, Japan, Tokyo, Japan, Sendai, Japan Prolyl isomerase (Pin1) is a unique enzyme which recognizes the pSer/ Pro or pThr/Pro motif and changes the conformations of target proteins. We previously reported that Pin1 binds to Crtc2 and suppresses CRE transcrip- tional and gluconeogenic activity in the liver. In addition, Pin1 associates with IRS-1, a major insulin receptor substrate, and enhances insulin-induced metabolic actions. These events ultimately contribute to the incorporation of excess nutrients into tissues. In this study, we speculated that Pin1 also plays a role in the pathogenesis of metabolic syndrome, since Pin1 is report- 1814-P edly involved in activations of NF-gB and IRAK1, key regulators of cytokine The Production of Short-Chain Fatty Acids and Their Role in Glu- production and infl ammation. Interestingly, Pin1 expression was markedly cose and Lipid Metabolism in Mice increased in the livers of a rodent model with MCD (Methionine choline- GIJS DEN BESTEN, KAREN VAN EUNEN, THEO H. VAN DIJK, BERT K. GROEN, defi cient diet)-induced NASH. In addition, Pin1 KO mice were highly resistant BARBARA M. BAKKER, DIRK-JAN REIJNGOUD, Groningen, The Netherlands to MCD-induced NASH. To elucidate whether Pin1 in hematopoietic cells or Obesity is a major risk factor to develop diabetes type 2, and preventing that in hepatocytes plays a role in the development of NASH, wild-type and obesity is considered to be important in combating diabetes. Short-chain Pin1 KO mice were irradiated and reconstituted with either Pin1 KO or wild- fatty acids (SCFA), acetate, propionate and butyrate, produced by bacterial type bone marrow. The mice with deletion of non-hemopoietic Pin1 showed fermentation of fi bers in the colon are shown to prevent diet-induced obesity decreased fat accumulation in the liver. In contrast, those with deletion of and insulin resistance. The in vivo SCFA production rates and the fate of hemopoietic cells showed a similar amount of fat accumulation in the liver, SCFA in the host, however, are still largely unknown and are of major im- but suppressed cytokine production and fi brosis. Thus, it is likely that he- portance to understand the obesity-preventing role of SCFA.To this purpose,

Obesity patic and hemopoietic Pin1 contribute to hepatic fat accumulation and in- we developed an in vivo mouse model by continuously infusing labeled SCFA

POSTERS fl ammation, respectively, and that both are necessary for full manifestation for 6 h at 40 μmol/min/kg into the cecum after a dietary period of 6 weeks. of NASH. These fi ndings suggest that inhibition of Pin1 might provide a novel In mice the cecum is the major production site of SCFA. Production rates of

Integrated Physiology/ therapy for NASH. the different SCFA, their interconversion rates and incorporation into end Supported by: Young Scientists 20790648 (N.Y.) from the Ministry of Education, products of liver metabolism of the host (glucose, cholesterol, lipids) were Science calculated applying GC-MS and mass isotopologue distribution analysis.Pro- duction rates were 6.7, 0.7 and 0.2 μmol/min/kg for acetate, propionate and butyrate, respectively. Interconversion was observed between acetate and butyrate (0.6 μmol/min/kg), propionate and butyrate (0.17 μmol/min/kg) but not between acetate and propionate. In the host, fractional glucose synthe- sis by gluconeogenesis was 69% for propionate. Label from butyrate was incorporated into 34% of newly synthesized glucose. In contrast, no label of acetate was found in glucose. Acetate and butyrate, but not propionate, were used to the same extent for de novo synthesis and chain elongation of

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A466 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE fatty acids.Using our newly developed cecal infusion model we were able vs. 29.5±6.2 vs. 28.4±6.4 nmol, p=0.85) arm despite decreased glucose con- to determine in vivo SCFA production and interconversion rates by the gut centrations. We conclude that caloric restriction signifi cantly ameliorates microbiota and incorporation of SCFA into glucose and lipids by the host. glucose metabolism and insulin action in people with type 2 diabetes with This allows us to measure SCFA utilization by the host and elucidate the role or without AGB. of SCFA in prevention of obesity and type 2 diabetes. Supported by: NIH (RO1-DK82396) Supported by: The Netherlands Consortium for Systems Biology 1817-P 1815-P Glucagon and the Gut Hormones GLP-1 and Oxyntomodulin Increase Identifi cation of Niacin Receptor GPR109A as a Novel Downstream Resting Energy Expenditure in Man Target of Glucagon Receptor in Liver JONATAN I. BAGGER, JENS J. HOLST, TINA VILSBØLL, FILIP K. KNOP, Hellerup, I-CHEN YU, ANN MARIE RICHARD, SARAH WILL, YINGJIANG ZHOU, BHAVNA S. Denmark, Copenhagen, Denmark PARATALA, DONGMEI LI, TIFFANY GARESKI, DAVID KUBASIAK, RUTH GIMENO, The satiety hormone oxyntomodulin (OXM) is a proglucagon product with JULI JONES, MYLENE PERREAULT, Cambridge, MA, Andover, MA body weight lowering effect. It binds to the glucagon-like peptide-1 (GLP-1) Non-alcoholic fatty liver disease, with its associated dyslipidemia, has receptor and the glucagon receptor, but the mechanisms behind the body been linked to several disorders including obesity and type 2 diabetes. He- weight reducing effect of OXM remain elusive. We measured resting energy patic steatosis induced by high-fat feeding is associated with reduction of expenditure (REE) during infusions of: 1) saline, 2) GLP-1, 3) glucagon, 4) OXM, glucagon receptor density in hepatocytes, and hepatic glucagon action is and 5) GLP-1+glucagon in healthy subjects.Indirect calorimetry was used to demonstrated to be essential for exercise-induced reversal of fatty liver in measure oxygen consumption (VO2), carbon dioxide production (VCO2), respi- mice. In hyperlipidemic rats, acute and chronic glucagon administration sig- ratory quotient (RQ) and REE after a 3h-continuous infusion of either saline, nifi cantly decreases serum cholesterol, triglycerides, and very low-density GLP-1 (1 pmol×kg-1×min-1), glucagon (0.86 pmol×kg-1×min-1), OXM (3 pmol×kg-1× lipoprotein cholesterol, while glucagon is reported to acutely regulate he- min-1) or glucagon+GLP-1 (same doses) in 15 healthy male volunteers (age: patic lipoprotein particle metabolism in humans. Although evidence points 22±2 years (mean±SEM); BMI: 23±0.5 kg/m2; HbA1c: 5.8±0.1%) after an over- to a central role of glucagon in hepatic lipid metabolism, the underlying mo- night fast.Plasma glucose (5.1±0.1 mM) and triglyceride (1.0±0.1 mM) concen- lecular mechanism remains elusive. To understand the molecular basis of trations were similar during the calorimetric measurements. REE was signifi - glucagon’s action on hepatic lipid metabolism, we conducted genome-wide cantly elevated in response to all peptide infusions compared to saline, with analysis of glucagon-induced transcriptional profi les in mouse liver after OXM eliciting the most pronounced increase in REE (Fig. 1A). GLP-1 alone and acute intraperitoneal glucagon injection. Besides expected gluconeogenic in combination with glucagon elevated RQ compared to saline whereas OXM genes, we identifi ed several novel genes regulated by glucagon, such as LIPG, decreased RQ (Fig. 1B).These data suggest that GLP-1, glucagon and OXM in- GPR109A, ANGPTL4,and LRP1. Our transcriptional profi ling data showed that crease REE. Also, our RQ results indicate that oxyntomodulin increases REE GPR109A expression was rapidly induced by 20 fold one hour post-glucagon by other mechanisms (perhaps lipid metabolism) than GLP-1, which is known injection, and real-time qPCR corroborated that fi nding at one- and six-hours to shift the metabolic substrate utilization towards carbohydrate metabolism. post-injection. The induction of GPR109A by glucagon was abolished in glu- cagon receptor knockout (GCGRKO) mice. Notably, serum nonesterifi ed free fatty acid levels were signifi cantly reduced in wild-type mice one hour after glucagon injection, but not in GCGRKO mice. Moreover, glucagon directly induced GPR109A expression in primary cultured mouse hepatocytes. To- gether, these fi ndings provide new insights to the molecular basis of gluca- gon’s action on hepatic lipid metabolism, and identifi ed GPR109A as a new downstream effector of glucagon in liver.

INTEGRATED PHYSIOLOGY—MACRONUTRIENT METABOLISM AND FOOD INTAKE

1816-P The Effect of Caloric Restriction Alone or in Combination With Ad- justable Gastric Banding on Fasting and Postprandial Glucose Me- tabolism in Diabetic Subjects 1818-P MATHENI SATHANANTHAN, PAULA GIESLER, JEANETTE LAUGEN, CHIARA Differences in Systemic and Splanchnic Metabolism of Saturated, DALLA MAN, CLAUDIO COBELLI, ALAN R. ZINSMEISTER, ADRIAN VELLA, Roch- Monounsaturated and Polyunsaturated Fatty Acids During Meal ester, MN, Padua, Italy Absorption In the absence of equivalent caloric restriction, Adjustable Gastric Band- EKTA SINGH, RITA BASU, JAIME ALMANDOZ, C.M. JOHNSON, BARBARA NOR- ing (AGB) is superior to pharmacotherapy in achieving glycemic control in BY, ROBERT H. NELSON, ROBERT H. NELSON, JOHN MILES, Rochester, MN type 2 diabetes. However, the results compared to other bariatric proce- Diets enriched in polyunsaturated fatty acids (FA) are protective against dures are inferior, suggesting that the benefi ts of AGB are mediated to a cardiovascular disease, but differences in the metabolism of various classes large extent by caloric restriction. We therefore studied subjects with type of FAs have been little explored. We measured systemic and splanchnic 2 diabetes who underwent AGB (n=6) and matched diabetic controls (n=8); at kinetics of palmitate (P), oleate (O), and linoleate (L) in 8 postabsorptive baseline, 6 & 12 weeks after initiation of caloric restriction similar to that af- subjects (age 39±3 y, BMI 25±2 kg/m2). After placement of femoral artery ter AGB. All subjects consumed a pureed diet (~800Kcal/day) over the period and hepatic vein catheters, subjects were given aliquots of a mixed liquid of study. A labeled mixed meal which included 35g of Jell-O labeled with meal containing triglyceride labeled with [1-13C] P, [1-13C] O, and [3H] L q 15 13 3 2 [1- C]-glucose. [6- H] and [6,6- H2] glucose infused intravenously enabled min to achieve steady-state suppression of lipolysis. At 270 min, intravenous Obesity measurement of the rate of meal appearance (Meal Ra), endogenous glu- infusions of a 3H triolein-labeled lipid emulsion and [U-13C]-labeled P, O and POSTERS cose production (EGP) and glucose disappearance (Rd). Integrated glucose L were initiated and continued until the end of the study (390 min). Splanch-

concentrations decreased signifi cantly in the diet arm (4.2±0.7 vs. 2.3±0.1 nic blood fl ow was measured with indocyanine green. Blood samples were Integrated Physiology/ Mol vs. 2.4±0.2 Mol, baseline vs. 6 vs. 12 weeks respectively, p=0.01). This taken over the last hour of the study. Plasma P, O and L concentrations was not observed in the AGB+diet arm (2.7±0.4 vs. 2.8±0.6mol vs. 3.0±0.9 (13±1, 32±3 and 28±3 μmol/L, respectively) and rates of appearance (23±3, Mol, p=0.94). However, fasting EGP (diet - 18.6±0.2 vs. 13.9±0.1 vs. 15.2±0.2 57±6 and 47±6 μmol/min, respectively) were each different from the other μmol/kg/min, p=0.03), (AGB+diet - 19.3±0.3 vs. 17.1±0.2 vs. 18.2±0.2 μmol/ 2 FAs, p<0.01. Systemic fractional spillover of dietary FAs into the plasma kg/min, p=0.04) and nadir postprandial EGP (diet - 5.8±1.1 vs. 4.9±0.8 vs. free fatty acid pool was lower for L (12±3%) than for P (24±4%), P<0.05, 4.8±0.5 μmol/kg/min, p=0.04), (AGB+diet - 6.4±1.7 vs. 5.1±0.7 vs. 5.4±0.4 but not compared to O (16±3%). Splanchnic uptake of P, O and L were also μmol/kg/min, p=0.05) were decreased. No differences in Rd or Meal Ra each different from the other 2 FAs (5±1, 10±2 and 12±2 μmol/min respec- were apparent in either arm. Integrated, incremental insulin concentra- tively, all p<0.01). Splanchnic release of L (13±2 μmol/min) was greater than tions decreased over the course of the study in the AGB+diet (24.7±4.9 vs. O and P (7±2 and 5±1 μmol/min, respectively, p<0.002), which did not differ 15.5±3.1 vs. 13.7±3.8 nmol, p=0.01) but did not change in the diet (29.8±10.0 from each other. Splanchnic clearance of P, O and L were different from the

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A467 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE

other 2 FAs (410±65, 333±54 and 480±82 ml/min respectively, each p<0.01). 1821-P The contribution of the splanchnic bed to systemic uptake was greater for Roles of Short-Chain Fatty Acid Receptors, GPR41 and GPR43, in Di- L (27±3%) than for O (18±2%) and P (23±3%), both p<0.05). In conclusion, L is etary Fiber Induced Weight Loss in High-Fat Diet Fed Obese Mice taken up more avidly by the splanchnic bed than are O or P. This preferential CHRISTOPHER C. NYSTROM, DALLAS K. CROOM, BOB P. WIARD, ANDREW A. uptake may be due to differences in cellular transport, greater uptake into a YOUNG, MARK A. PAULIK, LIHONG CHEN, Durham, NC splanchnic phospholipid pool, or other mechanisms. Obesity has reached epidemic proportions globally. Although overweight and obese are recognized as a major risk factor for diseases such as car- 1819-P diovascular diseases and diabetes, currently only one anti-obesity medicine Long-Term Effects of Low-Carbohydrate/High-Fat Liquid Diets is approved on the market. Other alternative approaches including dietary (LCHFD) on Diurnal Glucose Variations and Insulin Dose in Type 2 fi bers have been explored in both preclinical and clinical studies. Dietary fi - Diabetic Patients on Tube Feeding Who Require Insulin Therapy bers are edible parts of plants that are resistant to digestion and absorption YUTAKA MORI, TERUO OHTA, JUNICHI YOKOYAMA, KAZUNORI UTSUNOMIYA, in the small intestine, but are fermented in the large intestine to produce Komae, Japan, Utsunomiya, Japan short-chain fatty acids (SCFA). Oligofructose saccharide (OFS) is a dietary Objective: In this study, we investigated the usefulness of a low-carbo- fi ber that is known to reduce body weight in obese rodent models. To ex- hydrate/high-fat liquid diet (LCHFD) in long-term glucose control in type 2 plore potential roles of SCFAs in dietary fi ber-induced weight loss effect, we diabetic patients on tube feeding who require insulin therapy.Methods: The tested the effects of OFS in GPR41 and GPR43 knockout mice. High-fat (HF) study included a total of 10 patients requiring insulin therapy in whom PEG chow signifi cantly increased body weight in WT, GPR41-KO and GPR43-KO had been performed due to diffi culty swallowing and who were being tube- mice in four weeks. There was no difference between WT and KO mice. fed with high-carbohydrate liquid diets (HCD). With stable glucose control From the fi fth week, subgroups of HF-fed mice were switched to HF diet maintained, the patients were monitored for glucose levels for 6 consecutive with 10% OFS supplement for four weeks. OFS signifi cantly decreased body days by using CGM. The patients were continued on HCD during the fi rst 3 weight compared to HF alone in all strains. However, compared to its ef- days and were switched to LCHFD during the latter 3 days. The patients fect in WT mice, OFS induced signifi cantly more body weight reduction (56% were then continued on LCHFD and monitored for glucose levels for 3 con- more) in GPR41-KO mice but was less effi cacious (26% less) in GPR43-KO secutive days by using CGM after 3 months of feeding with LCHFD. Either mice. The body weight effects of OFS were not associated with signifi cantly diet was given with the daily calorie intake set at 800-1200 kcal/day to al- decreased food intake in all strains. In summary, our results suggested 1) low comparisons. Insulin regimens included basal-bolus insulin in 8 of the SCFAs might be a mechanism for dietary fi ber induced weight loss in high-fat 10 patients and intermediate-acting insulin in 2 patients. Variables exam- diet induced obese mice; 2) GPR41 and GPR43 might have opposite effects ined included 24-hour mean glucose levels and their SDs (mg/dL), total area on SCFA-mediated body weight effects. for the range of glucose variations (mg · hr/dL), MAGE (mg/dL), proportions of time in hyperglycemia and hypoglycemia as calculated from CGM data, 1822-P HbA1c values, and required insulin doses before and after switching of diets. Infl uence of Carbohydrate Beverage on Stress Hormones and Results: The SDs of 288 glucose levels over 24 hours, total area for the range Interleukin-6 Responses Associated With the Mood State in Elite of glucose variations and MAGE were signifi cantly decreased immediately Runners and 3 months after switching to LCHFD. Despite the signifi cant decrease in MAYSA SOUSA, RONALDO ZUCATELLI, ROSA FUKUI, ARITANIA SANTOS, MARIA required insulin dose 3 months after switching, the HbA1c values were sig- ELIZABETH SILVA, São Paulo, Brazil nifi cantly decreased 3 months after switching.Conclusions: LCHFD reduced In this study we evaluated the effects of carbohydrate (CHO) supplementa- not only the range of glucose variations but also the required insulin doses tion on glutamine, interleukin-6 (IL-6) and stress hormones levels associated and HbA1c values, suggesting LCHFD may be useful in long-term glucose with the mood state in elite runners undergoing an overload training program control in these patients on tube feeding who require insulin therapy. (days1-8) followed by a session of high intensity running consisted of 10x800 m at 100% of Vm3km and 2x1000 m maximal bouts (day 9). Twenty-four elite 1820-P male athletes (28.0±1.2 yr) were randomized to receive CHO (maltodextrin The Intake of Fiber Suppresses the High Fat High Carbohydrate Meal solution) or zero energy placebo solution (control group). The profi le of mood Induced Endotoxemia, Oxidative Stress and Infl ammation state (POMS) was performed before and after the overload training. Blood HUSAM GHANIM, SANDEEP DHINDSA, MANAV BATRA, AJAY CHAUDHURI, SA- samples were collected before, during, and post-running. Glutamine levels NAA ABUAYSHEH, KELLY GREEN, PARESH DANDONA, Buffalo, NY increased in the CHO group after the overload training (from 35.2±2.2 mg/L Our previous work has shown that the intake of a high fat high carbohydrate to 48.3±1.7 mg/L, p<0.05) in contrast to the control group (48.3±1.7 mg/L (HFHC) meal results in an increase in endotoxin (LPS) concentrations and in to 45.6 mg/L, p>0.05). When responses of the intermittent running session the expression of TLR4, the receptor for LPS, and CD-14, which facilitates the (10x800m) performed on day 9 were evaluated, a signifi cant increase in the binding of LPS to its receptor. In addition, there is an increase in ROS genera- stress hormones was observed in both groups. However, its concentrations tion and NFgB binding. Our work has also demonstrated that an equicaloric were blunted in the CHO group post-running in relation to control group (Cor- meal rich in fi ber and fruit (American Heart Association (AHA) recommended tisol: 22.4±0.9 pmol/L vs. 27.6±1.4 pmol/L, Norepinephrine: 1647.6±85.2 pg/ meal) does not induce these changes. We hypothesized, therefore, that the mL vs. 36.7±144.6 pg/mL; p<0.05). IL-6 levels were lower in the CHO group addition of fi ber to a HFHC meal will prevent the changes induced by HFHC compared with the control group (13.6±1.1 pg/mL and 26.5±1.1 pg/mL, re- meal. Eight fasting normal subjects (BMI<25Kg/m2) were given 900 Calorie spectively; p<0.05). Regarding the POMS test, no signifi cant drink treatment meals of either HFHC or AHA or HFHC with fi ber (30g) (Fiber One Original) on effects were detected for the total components score (fatigue, tension, de- 3 visits one week apart in a randomized crossover design. The HFHC meal pression, anger, confusion and vigor) after training. However, the individual induced an increase in LPS concentrations (by 69±14%) with an increase in subcomponents of the POMS test revealed an increased overall time effect ROS generation (by 120±24%) and the expression of TLR4, CD14 and IL-1` for fatigue in both groups (p<0.05) and a decreased for vigor state in the (by 71±14%, 86±14% and 127±14%, respectively; p<0.05 for all) in MNC. The control group (p<0.01) after training. The fi ndings of this study showed that AHA meal did not induce any of these changes. The addition of fi ber to HFHC CHO beverage resulted in less catabolic stress and may have enhanced the

Obesity meal signifi cantly reduced the increases observed with HFHC meal alone mental function, preventing performance deterioration.

POSTERS in LPS concentrations (by 58±10%; p<0.05), ROS generation (by 47±14%; Supported by: FAPESP p<0.05) and the expression of TLR4, CD14 and IL-1` (by 37±11%, 46±12%

Integrated Physiology/ and 52±15%, respectively; p<0,05 for all). Additionally, the intake of fi ber 1823-P with the meal reduced postprandial glucose excursion and increased insulin Diet Interventions of Different Composition Result in Similar Weight concentrations compared to meal alone. We conclude that the addition of fi - Loss But Alter Fed-State Fuel Oxidation ber to a pro-infl ammatory HFHC meal has a benefi cial anti-infl ammatory and MARESSA J. VALDEZ, JENNIFER E. LAMBERT, JOSEPH J. LEE, YELENA HOVHAN- metabolic effect. Thus, the fi ber content of the AHA meal may account for NISYAN, DORA L. BRADFORD, MARIA A. RAMOS-ROMAN, ELIZABETH J. PARKS, its non-infl ammatory nature. Since ROS, TLR-4 and IL-1` potentially interfere Dallas, TX with insulin signal transduction and are known to be pro-infl ammatory in the Controversy exists whether low-carbohydrate (LCD) or low-fat (LFD) diets arterial wall, this action of dietary fi ber may contribute to its benefi ts in the induce greater weight loss and metabolic improvement. The purpose of this prevention of insulin resistance, type 2 diabetes and atherogenesis. study was to determine whether LCD or LFD: 1) produces greater weight loss, and 2) differ in effect on substrate oxidation. Non-diabetic overweight His- panic (Hisp) and African American (AA) adults were randomized to either a

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A468 INTEGRATED PHYSIOLOGY—MACRONUTRIENTCATEGORY METABOLISM AND FOOD INTAKE

5-month LCD (4 Hisp/4 AA; 40% fat, 34% CHO, 26% protein) or LFD (3 Hisp/4 invasively in C57/B6 male mice fed with a chow diet. The TG turnover mea- AA; 28% fat, 52% CHO, 23% protein). At baseline and follow up, fat (FatOx) surement was done using a 2H-labeled TG clearance approach. Following and glucose (GlucOx) oxidation were measured using indirect calorimetry at adipose TG pool labeling achieved via 10% D2O in drinking water, the time fasting and 3 hours after consumption of a standardized meal (42g fat, 100g course of labeled adipose TG clearance was followed longitudinally in vivo CHO). Insulin sensitivity (SI) was measured by IVGTT. At baseline, groups did in mice for 5 weeks with the 2H-NMR approach. Assuming the labeled TG not differ in SI (LCD 2.5 ±0.5 vs. LFD 2.0 ±0.2 10^-4/min per uU/mL; P=0.45) clearance process can be modeled as a fi rst order kinetic process, the rate or body weight (LCD 96 ±7 vs. LFD 102 ±11 kg; P=0.69). Groups achieved of TG turnover was estimated from the kinetic parameters determined from signifi cant (P<0.05 for both) and similar weight loss (LCD -11 ±0.6 vs. LFD -9 time course curves of labeled TG and water clearance. Whole body lean and ±0.7%; P=0.07), and fasting FatOx and GlucOx did not change in either group fat masses of these mice were assessed with echo NMR and correlated (see Figure). GlucOx increased from fasting to fed in both groups before and with their rates of adipose TG turnover. Average rate of adipose TG turnover after weight-loss (P<0.05 for all). Fed-state FatOx and GlucOx were not dif- in these chow-fed lean mice (C57/B6) was found to be: 0.0367±0.0019 day-1 ferent after the LCD. After the LFD, fed-state FatOx increased while GlucOx and average body weight was 44.1±1.6 g (N = 8 / group, male C57 mice). decreased compared to baseline. In summary, this study suggests that LCD When comparing body weights of all mice to their corresponding rates of TG and LFD cause signifi cant and similar weight loss; thus, macronutrient com- turnover, we found a signifi cant negative correlation between body weight position may be less important than reducing overall calories and improving and rate of TG turnover (R2 = 0.863). In conclusion, we evaluated the rate of diet quality. Further, habitual macronutrient consumption rather than weight adipose TG turnover of lean C57 mice fed with a chow diet non-invasively. loss per se may infl uence postprandial substrate utilization. The results showed that old chow fed mice exhibited a certain degree of variation in the rate of adipose TG turnover. In addition, the rate of TG turn- over is found to be tightly correlated to both body weight and fat mass, sug- gesting rate of adipose TG turnover may play a role in determination of body weight phenotype. Furthermore, this study demonstrated a practical in vivo method for quantifying adipose TG turnover.

1826-P Central Fructose Impairs the Anorexic Effect Mediated by the Cen- tral Glucagon-Like Peptide 1 Receptor (Glp1r) MELISSA A. BURMEISTER, JENNIFER E. AYALA, DANIEL J. DRUCKER, JULIO E. AYALA, Orlando, FL, Toronto, ON, Canada Supported by: NIH (5RL1DK081187), CTSA (UL1RR024982), TORS (UL1DE019584) Increased sucrose consumption is implicated as a culprit in the obesity ep- idemic. Daily food intake is greater in sucrose- vs. starch-fed mice (2.74±0.33 1824-P vs. 1.78±0.20 g). The aim of this study is to identify mechanisms by which Substrate Regulation of Gluconeogenesis (GNG) by Branched-Chain sucrose stimulates food intake. Activation of hypothalamic AMP activated Amino Acid (BCAA) Oxidation in Mice protein kinase (AMPK) increases food intake, and we show that higher food PENGXIANG SHE, ZHIYOU ZHANG, MASAKAZU SHIOTA, CHARLES H. LANG, intake in Glp1r knockout mice correlates with hypothalamic AMPK activa- ROBERT A. HARRIS, CHRISTOPHER J. LYNCH, Hershey, PA, Nashville, TN, India- tion. We also show that the Glp1r agonist Exendin-4 (Ex4) inhibits hypotha- napolis, IN lamic AMPK, whereas the sucrose metabolite fructose activates AMPK. We Diabetes is characterized by fasting hyperglycemia resulting from elevat- hypothesize that fructose blunts the satiety effects mediated by the central ed hepatic GNG, which is thought to be controlled by the so-called rate- Glp1r via modulation of hypothalamic AMPK. To test this, 5h-fasted C57Bl/6 limiting gluconeogenic enzymes, phosphoenolpyruvate carboxylase (PEPCK) mice received intracerebroventricular (ICV) delivery of artifi cial cerebrospi- and glucose-6-phosphatase. However, this assertion has been recently nal fl uid (ACSF, 2 μl), Ex4 (0.01 μg), fructose (400 μg), a subthreshold dose of challenged, leading us to hypothesize that gluconeogenic substrate avail- the AMPK activator AICAR (1 μg), fructose+Ex4 or AICAR+Ex4 just prior to ability can play a major role in controlling GNG. Alanine is a major substrate the onset of the dark cycle. Food intake was continuously monitored for 18h. for net glucose synthesis. Previous in vitro studies have shown that during Food intake in response to ICV ACSF (1.85±0.19 g) was signifi cantly reduced oxidation BCAA serve as a nitrogen donor for alanine synthesis in muscle. by ICV Ex4 (0.73±0.17 g). Compared to ICV Ex4, food intake was greater in However, the link between altered BCAA oxidation and GNG in vivo has not mice receiving ICV fructose+Ex4 (1.35±0.18 g) or ICV AICAR+Ex4 (1.45±0.28 been demonstrated. BCAA oxidation is tightly controlled by a rate-limiting g). ICV fructose or AICAR alone had no effect. We found that Glp1 levels fol- step catalyzed by the branched-chain keto acid dehydrogenase (BCKD) en- lowing a sucrose gavage were not different compared to glucose (5.94±0.67 zyme complex containing E1, E2 and E3 subunits. BCKD is inhibited by BCKD vs. 4.78±0.20 pM) and that hypothalamic Glp1r mRNA levels were actually kinase (BDK)-catalyzed E1 phosphorylation. We studied glucose metabolism 1.5-fold higher in sucrose-fed mice, indicating that fructose exposure does in E2R (liver rescue) and BDK knockout (KO) mice, which respectively exhibit not impair Glp1 action by decreasing Glp1 secretion or Glp1r availability. In blocked and enhanced BCAA oxidation. Fasting blood glucose concentra- sum, these data show that central fructose attenuates the anorexic effect tion and endogenous glucose production (EGP) were decreased ~35% in E2R mediated by the central Glp1r and suggest that sucrose promotes central Glp1 KO mice but increased ~46% in BDK KO mice, compared to their wild-type resistance and increased food intake by activating hypothalamic AMPK. This (WT) controls. The decrement in hepatic glycogen content during fasting did demonstrates that dietary carbohydrate composition signifi cantly impacts not differ between WT and BDK KO mice, suggesting that enhanced EGP in central mechanisms that control feeding behavior. BDK KO mice was not due to elevated glycogenolysis. Hepatic PEPCK protein abundance and activity were unaltered in both E2R and BDK KO mice, sug- 1827-P gesting unchanged hepatic gluconeogenic capacity in these animals. BCAA p-Synephrine Stimulates Glucose Consumption via AMPK in L6 R concentrations were increased 3-7 fold in liver, muscle and plasma of E2 Skeletal Muscle Cells KO mice, while alanine levels in plasma and tissue were decreased ~40%. DEOKBAE PARK, DAEHO LEE, YOUNGKI LEE, Jeju, Republic of Korea In contrast, plasma BCAA and alanine concentrations were respectively de- Interest in p-synephrine, the primary protoalkaloid in the extract of bit- Obesity creased and increased ~50% in BDK KO mice, compared to WT mice. These ter orange and other citrus species, has increased due to its various phar- POSTERS data suggest that BCAA oxidation may control GNG and fasting glycemia macological effects and related adverse effects. The lipolytic activity of through regulating alanine synthesis. p-synephrine has been repeatedly revealed by in vitro and in vivo studies and Integrated Physiology/ p-synephrine is currently marketed as a dietary supplement for weight loss. 1825-P The present study investigated the effect of p-synephrine on glucose con- Body Weight Found to be Negatively Correlated to the Global Rate sumption and its action mechanism in L6 skeletal muscle cells. Treatment of of Adipose TG Turnover L6 skeletal muscle cells with p-synephrine (0 - 100 μM) did not affect cell vi- HAIYING LIU, DAN ZHOU, Rahway, NJ ability and increased basal glucose consumption up to 50% over the control The objectives of the study are: 1) to evaluate the feasibility of measur- in a dose-dependent manner. Insulin-stimulated glucose consumption was ing the global rate of adipose TG turnover in vivo non-invasively; and 2) to signifi cantly increased by the addition of p-synephrine. p-Synephrine stimu- investigate whether body weight is related to the rate of triglyceride (TG) lated the phosphorylation of AMPK but not of Akt. p-Synephrine-induced turnover. Whole body the rate of TG turnover due to beta oxidation was glucose consumption was sensitive to the inhibition of AMPK but not to the evaluated using nuclear magnetic resonance spectroscopy (NMRS) non- inhibition of PI3 kinase. p-Synephrine also stimulated the translocation of

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A469 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

Glut4 from the cytosol to the plasma membrane; this stimulation was sup- p=0.06), a transcription factor that mediates the synthesis of several proteins pressed by the inhibition of AMPK, but not of PI3 kinase. Taken together, involved in lipid accumulation. Together our fi ndings indicate that alterations p-synephrine can stimulate glucose consumption (Glut4-dependent glucose in skeletal muscle metabolism may not contribute meaningfully to the marked uptake) by stimulating AMPK activity, regardless of insulin-stimulated PI3 insulin resistance observed after 2 wks of overeating. kinase-Akt activity in L6 skeletal muscle cells. Supported by: R01DK71955 Supported by: NRFK (2010-0022036) & 1830-P Skeletal Muscle Uncoupled Mitochondrial Respiration is De- INTEGRATED PHYSIOLOGY—MUSCLE creased by Short Term Very Low Calorie Diet DOUGLAS R. MOELLERING, QINGLAN LIU, DANA Y. RIGSBY, YONGBIN YANG, VEERADEJ PISPRASERT, JAYLEEN M. GRAMS, W. TIMOTHY GARVEY, Birming- Guided Audio Tour: Skeletal Muscle Metabolism—Humans (Posters 1828- ham, AL P to 1834-P), see page 13. Mitochondrial dysfunction is involved in obesity, insulin resistance and diabetes while the cause and effect relationship is still under investigation. & 1828-P Mitochondrial uncoupling due to proton leak can contribute up to 30% of The Novel Adipo-Myokine Chitinase 3-Like-1 (YKL-40) Is a Potential basal metabolic rate, and may protect against reactive oxygen species (ROS) Factor in the Infl ammatory Cross-Talk between Fat and Muscle production shielding cells from damaging oxidation of proteins, lipids, and MANUELA ELSEN, SVEN W. GÖRGENS, SILJA LAMBERND, ANDREA CRAMER, mtDNA. Our group previously reported weight loss induced by a short term KRISTIN ECKARDT, HENRIKE SELL, JÜRGEN ECKEL, Düsseldorf, Germany very low calorie diet (VLCD) improved insulin sensitivity while decreasing YKL-40, a member of the glycosyl hydrolase family 18, plays a role in tis- IMCL content. The short term effects on mitochondrial function after VLCD sue remodeling and infl ammatory processes. Elevated serum levels of YKL- have not been investigated and our goal was to evaluate mitochondrial bio- 40 are found in patients with cardiovascular disease and type 2 diabetes. energetics, insulin sensitivity, and energy expenditure after VLCD. Insulin Proteomic profi ling of secreted proteins from human adipocytes and human resistant women were equilibrated on an isocaloric diet for one week and skeletal muscle cells (hSkMC) identifi ed YKL-40 as an adipokine and myokine baseline studies were obtained, then they were given a VLCD (800 kcal/day) (adipo-myokine). However, the overall biological function of YKL-40 remains for 7 days and studies were repeated. Needle biopsies of the vastus lateralis elusive. The aim of the study was to characterize YKL-40 as an adipo-myokine were performed and in situ/ex vivo mitochondrial function was evaluated and to investigate the regulation of YKL-40 in human adipocytes and hSkMC. in permeabilized muscle bundles using high-resolution respirometry. Three During differentiation of hSkMC, the protein levels of YKL-40 decreased by different substrate/inhibitor protocols were utilized: Malate and pyruvate 70 % and secretion declined by ~40 %. Human adipocytes showed a signifi - (P/M) to evaluate fl ux through pyruvate dehydrogenase, the integrity of the cant decrease of YKL-40 protein level by 50 % and a decrease in secretion TCA cycle, and the electron transport chain (ETC); malate, pyruvate, and suc- by ~80 % during differentiation. In comparison to differentiated hSkMC, hu- cinate (P/M/S) to assess convergent electron fl ux from complexes I and II; man adipocytes secrete 30 times more YKL-40 (13.85 ± 3.14 ng/ml vs. 0.43 ± and malate with palymitoyl-carnitine (P-C/M) to evaluate fatty acid beta 0.12 ng/ml). To analyze the cross-talk between adipose tissue and skeletal oxidation. The data revealed signifi cant decreases in both State 2 and State muscle, hSkMC were incubated with adipocyte-conditioned medium (CM) 4o leak rates after 1 week VLCD using P/M. Interestingly non-mitochondrial resulting in a signifi cant increase of YKL-40 protein level (3.5 fold). Treatment respiration was signifi cantly decreased after 1 week VLCD using P-C/M. In of hSkMC with IFN-a, TNF-_ and IL-1` increased YKL-40 protein level (2-3 conclusion: 1) One week of a VLCD signifi cantly reduced skeletal muscle fold). Preliminary data showed an increased phosphorylation of the stress mitochondrial proton leak; 2) This effect would be benefi cial by producing kinase p38 MAPK (1.5 fold) after incubation of hSkMC with recombinant increased fl ux through the ETC and decreased ROS production but would human YKL-40 for 24 h.In conclusion, YKL-40 is a new adipo-myokine with also predict lower resting energy expenditure per lean mass. substantially higher secretion from adipocytes. CM, that contains the secre- Supported by: NIH (DK038746, DK083562, P60 DK 079626), Merit Review Dept. tory output of adipocytes, as well as isolated cytokines are able to increase Veterans Affairs YKL-40 protein level in hSkMC. In turn, YKL-40 stimulates proinfl ammatory pathways in hSkMC, presumably in an auto-/endocrine manner. Thus, we as- & 1831-P sume that YKL-40 could be a new important factor in the cross-talk between Exercise Improves Skeletal Muscle Mitochondrial Capacity in Mor- adipose tissue and skeletal muscle, promoting infl ammation and potentially bidly Obese Subjects After Bariatric Surgery contributing to the development of insulin resistance. ELIZAVETA V. MENCHIKOVA, PAUL M. COEN, MAJA STEFANOVIC-RACIC, FRED- ERICO TOLEDO, JOSEPH A. HOUMARD, VLADIMIR B. RITOV, BRET H. GOODPAS- & 1829-P TER, Pittsburgh, PA, Greenville, NC Changes in Skeletal Muscle Metabolism May Not Contribute to the Severalstudies have thoroughly examined the effect of dramatic weight Early Development of Insulin Resistance After 2 Weeks of Over- loss throughbariatric surgery on a variety of health parameters related to eating the risk for type2 diabetes (T2D). Very little is known, however, about the ef- ANDREA CORNFORD, ARIEL BARKAN, JEFFREY F. HOROWITZ, Ann Arbor, MI fects of large surgery-inducedweight loss on mitochondria, likely a key fac- Prolonged overeating and the resultant weight gain are clearly linked with tor in the energetics of obesityand T2D. The current study was undertaken the development of insulin resistance, but metabolic adaptations that occur to determine the effects ofsurgery-induced weight loss, either alone or in after relatively short periods of overeating are not completely understood. combination with regularexercise, on mitochondrial capacity. One to three The primary aim of this study was to characterize changes in muscle lipid months following gastricbypass surgery, patients were randomized to ei- metabolism that may accompany the development of insulin resistance after ther control (CON; n=15) ormoderate structured exercise (EX; n=15). Muscle 2 wks of overeating. Healthy, non-obese men (n=8) and women (n=2) were biopsies were taken before andafter 6 months of intervention (~9 month admitted to the hospital for 2 wks, during which time they ate ~4000 kcals/ after surgery) for measurements ofelectron transport chain (rotenone-sen- day (70 kcals/kg fat free mass/day). Insulin sensitivity was estimated during sitive NADH-oxidase) and TCA cycle (citrate synthase; CS) activities. Tis-

Obesity a meal tolerance test, and a muscle biopsy was obtained after an overnight suecardiolipin (CL) content was measured as independent non-enzymatic

POSTERS fast to assess muscle lipid accumulation, as well as protein markers associ- marker ofmitochondrial mass. Both surgery groups lost a signifi cant amount ated with impaired insulin signaling, infl ammation, and the regulation of lipid (21.7±1.8 kgP=<0.001 and 21.8±1.8 kg P=<0.001 for CON and EX, respectively)

Integrated Physiology/ metabolism. Whole body insulin sensitivity declined markedly after 2 wks of ofweight, but there was no difference in weight loss between groups. With overeating (Matsuda composite index: 8.3±1.3 vs. 4.6±0.7, p<0.05). In contrast, weightloss alone (CON) there were no statistically signifi cant changes in CL muscle markers linked with insulin resistance and infl ammation (e.g., phospho- content,CS or NADH-oxidase activities. The addition of exercise, however, rylation of IRS-1-ser312, Akt-ser473, c-Jun N-terminal kinase (JNK)) were not signifi cantlyincreased citrate synthase and specifi c NADH-oxidase activities altered by overeating. The concentration of intramyocellular lipids tended to followingbariatric surgery (from 20±2 to 26±3 U/ww (N=12, P=0.012) and increase after 2 wks of overeating (triacylglycerol: 7.6±1.6 vs. 10.0±1.8 nmol/ 2.5±0.2 to3.4±0.2 U/mg CL (N=15, P=0.001) for CS and NADH-oxidase, re- mg ww, p=0.11; diacylglycerol: 104±10 vs. 142±23 pmol/mg ww; p=0.26), but spectively). Inconclusion, exercise is important to induce robust increases in these changes did not reach statistical signifi cance. We also did not fi nd sig- mitochondrialcapacity during surgery-induced weight loss. Additional stud- nifi cant changes in the protein abundance of the key enzymes within the lipid ies are needed toexamine whether these mitochondrial changes are related synthesis pathway (GPAT or DGAT), and we actually found a trend for a reduc- to improved insulinresistance, altered energy metabolism or long-term main- tion in protein abundance of sterol response element bind protein 1 (SREBP-1a tenance of weight lossfollowing bariatric surgery.

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nance spectroscopy (MRS). 31-phosphorus MRS was measured during and & 1832-P after a 90 second 70% maximal volitional contraction isometric calf exercise Lipopolysaccharide Induces Insulin Resistance in Human Muscle to asses ADP production with exercise and depletion with rest. Perceived Cells through Toll-Like Receptor 4 maximal effort was corrected with standard force vs area calculation based HANYU LIANG, ALICIA SANCHEZ-AVILA, EUGENIO CERSOSIMO, RALPH DE- on maximal calf cross sectional area, and force output monitored through- FRONZO, NICOLAS MUSI, San Antonio, TX out the exercise bout. Subjects included 14 children with T1D (6 female, 8 Lipopolysaccharide (LPS), or endotoxin, is a major component of the outer male; Age 15.4±0.1 years; mean±SE) and 10 lean healthy controls (6 female, membrane of Gram-negative bacteria. Emerging evidence suggest that LPS 4 male; Age 15.2±0.2) who were matched for BMI, tanner stage and activity plays an important role in the pathogenesis of insulin resistance. For ex- level. Youth with T1D were signifi cantly more insulin resistant than controls ample, we and others have reported that obese nondiabetic and diabetic (glucose infusion rate 9.6±0.1 vs 18.6±0.2 mg/kg/min; p<0.001). The time subjects have elevated plasma LPS concentrations. The goal of the present for half of the ADP made during exercise to convert to ATP was signifi cantly study was to determine whether LPS directly induces insulin resistance in longer in T1D (18.4±0.3 vs. 14.6±0.2 seconds; P<0.05), despite the fact that cultured human myotubes. Treatment of human myotubes derived from lean the subjects with T1D had a lower increase in ADP concentration above pre- nondiabetic subjects with 100 ng/ml LPS for 12 hr caused an infl ammatory exercise concentrations than controls (18±0.4 vs. 29±0.5 mmol/l; p<0.05). In response, as evidenced by signifi cant increases in IL-6 (5.9-fold) and MCP1 summary, youth with T1D performing exercise at an equal workload, had less (9.6-fold) gene expression and JNK phosphorylation (2-fold). The infl amma- conversion of ATP to ADP, and slowed post-exercise ATP synthesis. Youth tory response caused by LPS signifi cantly reduced insulin-stimulated IRS-1 with T1D also have signifi cantly greater peripheral insulin resistance com- (46%), Akt (45%) and AS160 (57%) phosphorylation, as well as insulin-stim- pared to controls. Abnormal mitochondrial function in skeletal muscle in T1D ulated muscle glucose uptake (18%). Notably, TAK-242, a TLR4 antagonist, may be related to an alteration in the intracellular substrate environment, prevented LPS-induced increases in IL-6 and MCP1 mRNA expression, and and further work is needed to address this question. JNK phosphorylation. Moreover, TAK-242 protected human myotubes from Supported by: NIH/NIDDK 1R56DK088971-01, JDRF5-2008-291, K23 RR020038- the deleterious effect of LPS on insulin action (IRS-1, Akt and AS160 phos- 05, CCTSI Co-Pilot phorylation, and glucose uptake). In summary, LPS directly causes an infl am- matory response that leads to insulin resistance in human muscle cells. In 1835-P addition, blocking TLR4 prevents the LPS-induced infl ammatory response Adiponectin Activates Endothelial Nitric Oxide Synthase (eNOS) and improves LPS-triggered insulin resistance. Pharmacologic blockade of and Increases Muscle Microvascular Perfusion TLR4 could be a useful strategy for the treatment of insulin resistance. LINA ZHAO, WEIDONG CHAI, ZHENHUA DONG, EUGENE J. BARRETT, ZHENQI Supported by: NIH (AG030979, DK080157) LIU, Charlottesville, VA Adiponectin, circulating as either the full length (fAd) or globular (gAd) & 1833-P form, regulates endothelial function, insulin sensitivity and energy homeo- Fenofi brate Increases Lipid Metabolism in Human Skeletal Muscle stasis. Hypoadiponectinemia is an independent risk factor for coronary KRISTEN E. BOYLE, JOSEPH A. HOUMARD, JACOB E. FRIEDMAN, NEDA RASOU- artery disease in patients with diabetes. Recent evidence suggests that LI, Aurora, CO, Greenville, NC, Denver, CO adiponectin causes vasodilation via a nitric oxide-dependent mechanism. PPAR_ agonists such as fenofi brate (FENO) are commonly used to treat To examine whether adiponectin activates eNOS, bovine aortic endothelial dyslipidemia although the mechanisms of action are not fully understood. cells were treated with fAd (0, 0.1, 1, 3, 10 μg/ml) or gAd (0, 0.1, 1, 2, 3 μg/ml) We hypothesized that FENO might increase lipid oxidation in skeletal muscle for 15 min. Both fAd and gAd dose-dependently increased eNOS phospho- of obese humans at risk for diabetes. Obese subjects (age 47.7±2.5, BMI, rylation at ser1177 with a maximal effect observed at a concentration of 3 34.8±1.0, n=11) with impaired fasting or glucose tolerance were treated with μg/ml for fAd (58 ± 13%, p<0.05) and 2 μg/ml for gAd (73 ± 27%, p<0.05).To FENO (145 mg/day) for 10 weeks and muscle biopsies obtained pre- and post- assess the effect of adiponectin on muscle microvascular function in vivo, treatment. FENO reduced plasma triglyceride levels from 170±33 to 112±11 overnight fasted, adult male SD rats received an intraperitoneal injection of mg/dl, p=0.04; however, insulin sensitivity remained unchanged. To measure either fAd (50 μg), gAd (50 μg), or normal saline. Hindlimb muscle microvas- skeletal muscle lipid oxidation, the gene expression of candidate genes (n=9) cular blood volume (MBV) and microvascular blood fl ow velocity (MFV) were as well as corresponding functional assays (n=2) were performed. The mRNA measured using contrast-enhanced ultrasound before and every 30 min after levels of MCAD, PGC-1_, PPAR_, UCP3, CPT1, FoxO1and NADH dehydroge- the injection for 2 hrs. Microvascular blood fl ow (MBF) was calculated as nase [ubiquinone] iron-sulfur protein 3 (NDUFS3) remained unchanged with the product of MBV and MFV. Injection of fAd increased muscle MBV (~1.8- FENO treatment. However, FENO increased 14C labeled palmitate oxidation fold, p<0.05) and MBF (~2.2-fold, p<0.05) at 60 min with no change in MFV in fresh tissue homogenates by 50%. In addition, the rate of O2 consumption and the values returned back to baseline at 90 min. On the contrary, gAd (state 3) in isolated mitochondria (MT) measured by the Seahorse metabolic increased muscle MBV at 30 min and this effect lasted throughout the 120 fl ux analyzer using carbohydrate donors glutamate or succinate decreased min study with the maximum effect seen at 90 min (~2-fold, p<0.05). MFV by 55% & 70% respectively. To determine direct effects of FENO in human did not change, leading to increased MBF. Saline treatment did not alter any myocytes, 14C labeled palmitate metabolism and state 3 O2 consumption of of the microvascular parameters.We conclude that both fAd and gAd are palmitoyl carnitine were measured in cells cultured from human vastus lat- capable of activating eNOS and recruiting muscle microvasculature, and gAd eralis tissue (HSkMC) (n=6) under 24h lipid (100uM oleate:palmitate, 2:1) or appears to have more rapidly onset of action and prolonged effect than fAd. lipid + FENO (12.5uM) treatment. Similar to our clinical data, FENO improved As microvasculature regulates substrate and insulin delivery and exchange lipid oxidation by 70% in HSkMC exposed to lipid. Together, these data indi- in muscle, these effects may contribute to the insulin-sensitizing action of cate that FENO treatment alters MT metabolism by decreasing carbohydrate adiponectin. and increasing lipid fl ux. These effects are likely direct in nature. The lack of Supported by: NIH changes in the gene expression of enzymes regulating lipid metabolism de- spite a signifi cant improvement in lipid oxidation suggests that FENO might 1836-P activate these enzymes post translationally. Palmitate or High-Fat Feeding Suppresses Autophagic Flux in Car- Supported by: VA Merit Grant

diomyocytes by Impairing Autophagosome Turnover Obesity

BHARAT JAISHY, STEPHEN JENKINS, E. DALE ABEL, Salt Lake City, UT POSTERS & 1834-P The molecular mechanisms regulating myocardial autophagy in obesity

Delayed Skeletal Muscle Mitochondrial ADP Recovery in Children and diabetes are incompletely understood. This study examined the regula- Integrated Physiology/ With Type 1 Diabetes tion of autophagy by fatty acids (FA) in cultured H9C2 cardiomyocytes (CMs) MELANIE CREE-GREEN, MARK BROWN, BRADLEY R. NEWCOMER, DEBRA SIN- and in adult murine hearts. H9C2 CMs were treated with vehicle or 500 μM GEL, JANE E. REUSCH, KRISTEN J. NADEAU, Denver, CO, Birmingham, AL Palmitate for 4 h in DMEM with 5 mM glucose. Eight-week-old wild type or Peripheral insulin resistance (IR) has been associated with mitochondrial mCherry-LC3 transgenic male mice were placed on a high fat, high sucrose dysfunction in type 2 diabetes (T2D). Muscle IR is also seen in patients with diet (HFD, 45% fat) for 12 weeks and compared with chow-fed controls (CFC). type 1 diabetes (T1D), but mitochondrial ATP regeneration rates following ex- Autophagosome numbers were estimated by LC3 immunoblot or by counting ercise in this patient population is unknown. We hypothesized that muscle IR GFP-LC3 puncta in H9C2 CMs and mCherry-LC3 puncta in heart sections. in T1D would be associated with decreased muscle mitochondrial function. Lysosomes were counted as lyostracker-red stained puncta. In H9C2 CMs, Peripheral IR was assessed with a high dose (80 u/m2/min) hyperinsulinemic- palmitate signifi cantly increased autophagosome abundance as measured euglycemic clamp. Mitochondrial function was assessed by magnetic reso- by LC3-II/I ratio (2.2 fold vs. veh, p<0.02) and GFP-positive puncta (4 fold,

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A471 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

p<0.05), but reduced lysosome abundance by 35% (p<0.05). Chloroquine multivariate ANOVA). A threshold 6^ for a sharp increase in H2O2 produc- (CQ) inhibits autophagosome-lysosome fusion and the increase in autopha- tion was evident and lower in the diabetic than control mitochondria. When gosome number is an index of autophagosome formation rate. CQ increased sequential additions of ATP were used instead of ADP, the same results autophagosome abundance in control cells by 2.4 fold (LC3-II/I ratio) and were observed, further confi rming the effectiveness of recycling of ATP to 16-fold (LC3-GFP puncta), (p<0.05). Despite increased autophagosome abun- ADP. These data show that ROS production from insulin-defi cient diabetic dance at baseline, the addition of CQ in palmitate treated CMs did not in- mitochondria is increased relative to control mitochondria when considered crease autophagosome abundance above that seen in CQ-treated control in relation to respiration and 6^ and suggest that mitochondrial reactive CMs. Consistent with in vitro data, HFD signifi cantly increased autophago- oxygen production is best interpreted in relation to the major driving forces; some abundance as evidenced by a 1.4-fold increase in LC3-II/I ratio and a i.e. electron transport and mitochondrial potential. 2-fold increase in mCherry-positive puncta formation in the heart (p<0.05). Supported by: Veterans Affairs Medical Research Funds Whereas CQ treatment increased LC3II/I ratios in CFC by 2-fold, the change in CQ-treated HFD mice was only 28%. These data suggest that the increase 1839-P in autophagosome abundance induced by FA exposure or HFD is not second- TAK-242, a Small-Molecule Inhibitor of Toll-Like Receptor-4 Sig- ary to increased autophagosome formation but to impaired autophagosome naling, Protects Against Lipopolysaccharide- and Lipid-Induced turnover via a mechanism that involves reduced lysosome abundance. Infl ammation and Insulin Resistance in Muscle Cells Supported by: NIH SOPHIE E. HUSSEY, RALPH A. DEFRONZO, ALICIA SANCHEZ-AVILA, NICOLAS MUSI, San Antonio, TX 1837-P Emerging evidence suggests that toll-like receptor-4 (TLR4) and down- High-Fat Diet-Induced Insulin Resistance in Mice With Carnitine stream signaling pathways (MAPK, NFgB) play a role in the pathogenesis of Palmitoyltransferase 1b Defi ciency skeletal muscle insulin resistance. Lipopolysaccharide (LPS) and saturated TEAYOUN KIM, LAN HE, YAN LI, MARIA S. JOHNSON, YISHU DING, TIM R. NAGY, fatty acids (SFA) bind and activate TLR4, and plasma levels of these endog- PHILIP A. WOOD, QINGLIN YANG, Birmingham, AL, Orlando, FL enous ligands are elevated in obesity and type 2 diabetes (T2D). The present CarnitinePalmitoyltransferase 1 (CPT1) has been proposed as an attrac- study utilized TAK-242, a cyclohexene derivative which selectively binds and tive therapeutictarget against diabetes by restricting the entry of excessive inhibits TLR4, to determine whether TLR4 mediates LPS- and SFA-induced long-chain fattyacids into mitochondria. We hypothesized that the benefi - insulin resistance. L6 myotubes were pre-incubated with/without TAK-242 cial effects of CPT1 area short-term effect, and long-term CPT1 defi ciency (1μM) for 1h, prior to stimulation with 100 ng/ml LPS for 1h or 24h, or 400uM will lead to exacerbatedinsulin resistance (IR) due to lipotoxicity. We tested stearic acid for 6h. LPS and stearic acid caused an infl ammatory response, this hypothesis by usinga molecular genetics approach to avoid non-spe- as evidenced by increased mRNA expression of IL-6 (3.5- and 10-fold re- cifi c effects of CPT1 inhibitors onthe development of diet-induced IR; the spectively) and MCP-1 (detected in LPS- and stearic acid- treated cells only). heterozygous CPT1b knockout mice(CPT1b+/-) and wild type littermates in TAK-242 prevented both LPS and stearic acid-induced IL-6 and MCP-1 gene short- ()2 months) and long-term(*6months) high fat diet experiments (HFD, expression. LPS reduced insulin stimulated p-Akt by ~36%, and this was 60 % kcal% fat). As expected, [14C]-palmitateoxidation capacity of isolated completely prevented by TAK-242. Stearic acid reduced insulin stimulated mitochondria from skeletal muscle was decreasedby 40 % in CPT1b+/- p-Akt by ~20%, and TAK-242 partially prevented this reduction. LPS reduced micerelative to controls (n=4, p<0.05), confi rming the CPT1 defi ciency. Insu- insulin-stimulated 2-deoxyglucose (2DG) uptake by ~25%, and this was com- lin and glucosetolerance tests showedno differences at baseline, whereas pletely prevented by TAK-242. Stearic acid reduced insulin-stimulated 2DG CPT1b+/- miceshowed improved insulin sensitivity under the short-term uptake by ~20%, and this reduction was no longer signifi cant when cells HFD condition comparedto controls (n=8). In contrast, CPT1b+/- mice exhib- were pre-treated with TAK-242. SUMMARY: i) TAK-242 protects against LPS- ited an exacerbated IR underthe long-term HFD condition. Hyperinsuline- and SFA-induced infl ammation and insulin resistance in muscle cells; ii) LPS mic euglycemic clamp studies revealedthat the glucose infusion rate was causes infl ammation and insulin resistance by activating TLR4; iii) SFA also 4-fold decreased in CPT1b+/- mice (p<0.001,n=4) after 5 months on a HFD. work through TLR4 to induce infl ammation and insulin resistance, although The body weights of CPT1b+/- micewere 25% and 14% less in both short- other mechanisms likely are involved in this process. CONCLUSION: Pharma- andlong-term HFD compared to controls. Body composition analysis (QMR) cological inhibitors of TLR4 may represent a novel therapeutic approach to revealedthat CPT1b+/- mice had 50 % less fat mass and 14 % lesslean mass reduce infl ammation and improve insulin action in patients with T2D. than control after 5 months of HFD (n=4, p<0.05), but there were no signifi - cantdifferences in body composition after correcting for difference in body 1840-P mass(ANCOVA). These data suggest that CPT1b is benefi cial only inshort- term, but detrimental in long-term HFD-induced IR. Therefore, the current- study provides important insights into the therapeutic strategy on the use ofCPT1 inhibitors for the treatment of insulin resistance. WITHDRAWN Supported by: 1R01HL085499, 1R01HL084456, R21 AT003734 from NIH, T32 HL007457-28

1838-P Superoxide Production from Muscle Mitochondria of Insulin Defi - cient Rats: Relationship to Respiration and Membrane Potential BRIAN D. FINK, JUDITH A. HERLEIN, WILLIAM I. SIVITZ, Iowa City, IA Superoxide production from isolated mitochondria is diffi cult to under- stand quantitatively since it is strongly infl uenced by simultaneous dia- betes-induced changes in respiration and potential (6^). We studied this relationship in isolated gastrocnemius muscle mitochondria of control and

Obesity streptozotocin (STZ)-diabetic rats. We used the 2-deoxyglucose (2-DOG) en-

POSTERS ergy clamp method to clamp 6^ at different levels dependent on added ADP. We measured respiration simultaneously with 6^ using a respiratory cham-

Integrated Physiology/ ber fi tted with a potential sensitive electrode. We measured superoxide (in- directly as H2O2) in parallel using the same mitochondrial preparations. Mito- chondria were fueled by combined glutamate, malate, and succinate. In the presence of excess hexokinase (HK) and 2-DOG, 6^ decreased in stepwise fashion reaching a plateau with each sequential increase in ADP (consistent with rapid recycling of ATP back to ADP through 2-DOG phosphorylation). No such effects were noted in the absence of HK. Absolute H2O2 produc- tion (per mg mitochondria) was decreased in mitochondria of STZ diabetic muscle, but increased when expressed relative to respiration (proportional to electron transport). Moreover, H2O2 per unit respiration was increased in diabetic compared to control mitochondria at each level of 6^ (p < 0.01 by

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A472 INTEGRATED CATEGORYPHYSIOLOGY—MUSCLE

1841-P antilipolytic drug acipimox, which improves insulin sensitivity, preferentially Carnitine Palmitoyltransferase-1 Defi ciency Impairs Fatty Acid Oxi- reduces plasma concentrations of saturated FAs. Plasma concentrations of dation in Skeletal Muscle, but Preserves Insulin Sensitivity saturated (C14:0, C16:0, C18:0) and unsaturated (C16:1, C18:1 trans, C18:1 cis SHAWNA E. WICKS, KIMBERLY R. HAYNIE, BOLORMAA VANDANMAGSAR, and C18:2) FAs and insulin sensitivity were measured by mass spectrometry JINGYING ZHANG, RANDALL L. MYNATT, Baton Rouge, LA and a hyperinsulinemic (160 mU/m2.min) euglycemic clamp, respectively, in Accumulation of lipid in skeletal muscle has been implicated in the de- 9 obese [age=49±3 y, BMI=31.5±1.0 kg/m2, glucose=92 mg/dl, total glucose velopment of insulin resistance, possibly through reduced fatty acidoxida- disposal (TGD)=8.5±0.8 mg/kg.min], 12 T2D (age=48±3, BMI=34.2±0.9, glu- tion. To examine this hypothesis, we generated mice with muscle-specifi c cose=148, TGD=4.1±0.4) and 11 lean (age=39±2, BMI=24.7±0.6, glucose=93, deletion of carnitine palmitoyltransferase-I (CPT-1m-/-), an essential enzyme TGD=10.8±0.9) subjects. Obese and T2D subjects received acipimox (200 mg for mitochondrial fatty acid import for beta-oxidation. While impaired fatty four times/day) for one week. Acipimox treatment increased TGD in obese acid oxidation (FAO) is thought to cause accumulation of lipid intermediates and T2D subjects by 9% and 56%, respectively. At baseline, species of satu- that impair insulin signaling, CPT-1m-/- mice accumulate less body fat and are rated (8-25%, P=NS) and unsaturated (7-23%, P=NS) FAs tended to be elevat- more insulin sensitive, as measured by both insulin (ITT) and glucose (GTT) ed in obese versus lean subjects. In T2D subjects both saturated (73-102%) tolerance tests. Serum levels of glucose and insulin are lower, indicative of and unsaturated (89-124%) FAs also were elevated at baseline (P<0.05 for all better glycemic control. Additionally, CPT-1m-/- mice have higher RER values, species vs. lean). Acipimox reduced all FA species by 37-72% and 20-59% in refl ecting increased systemic carbohydrate oxidation. Consistent with the in obese and T2D subjects, respectively (P<0.05 for all species). In summary, (i) vivo data, pyruvate oxidation is elevated, while FAO in muscle homogenates insulin resistant subjects have elevated plasma concentrations of both satu- and isolated mitochondria is lower. Isolated mitochondria respirate nor- rated and unsaturated FAs; and (ii) the insulin-sensitizing effect of acipimox mally, indicating that mitochondrial function is not compromised, but citrate is associated with reductions in the concentration of all plasma FA species. synthase activity and mitochondrial DNA content are elevated, suggesting Our data challenge the notion that saturated, rather than unsaturated FAs, mitochondrial biogenesis. Gene expression studies show marked up regula- are preferentially linked to insulin resistance in human subjects. tion of pyruvate dehydrogenase, electron transport chain and FAO enzymes. Supported by: NIH (AG030979, DK080157) Consistent with increased mitochondrial biogenesis, the mRNA levels of PC- G1a are increased in CPT-1m-/- mice. Long-chain acylcarnitine (LCAC) species 1844-P are signifi cantly reduced in muscle of CPT-1m-/- mice. This leads to the intrigu- Effects of Short-Term Insulin Intensive Therapy on Peripheral Insu- ing possibility that impaired skeletal muscle FAO, possibly through reduced lin Resistance in Patients With Newly Diagnosed Type 2 Diabetes LCAC, elicits a signal that triggers induction of mitochondrial biogenesis and XUESI WAN, ZHIMIN HUANG, LING MA, LIEHUA LIU, MINHUA LIANG, JUAN LIU, preserves insulin sensitivity. YANBING LI, Guangzhou, China The intramyocellular lipid (IMCL) is now fi rmly established as a tissue 1842-P marker for peripheral insulin resistance. The aim of our study was to evalu- Diurnal Variation of Skeletal Muscle and Liver Glycogen Concentra- ate the effects and mechanisms of short-term intensive therapy by continu- tion in Well Controlled Type 2 Diabetes ous subcutaneous insulin infusion (CSII) on insulin resistance in patients with MAVIN MACAULEY, FIONA E. SMITH, SARAH STEVEN, PETE THELWALL, ROY newly diagnosed type 2 diabetes (T2DM).40 Patients with newly diagnosed TAYLOR, Newcastle upon Tyne, United Kingdom T2DM (25M/15F, age 50.8±9.7yrs, BMI 25.0±3.1 kg/m2, FPG 11.9±3.2 mmol/L, Skeletal muscle plays a major role in glucose homeostasis in normal and PPG 18.4±5.5 mmol/L) were enrolled and received CSII therapy, which health, with approximately 30% of meal carbohydrate being stored as mus- were terminated when normoglycaemia had been maintained for 14 days. 20 cle glycogen after the fi rst meal of the day. Liver stores approximately 20% NGT subjects (11M/9F, aged 50.8±7.3yrs, BMI 23.7±2.7 kg/m2, FPG 4.8±0.4 of meal carbohydrate after a single meal in healthy subjects. Together, the mmol/L, and PPG 5.6±1.0 mmol/L) were recruited, too. HOMA IR, intramyo- glycogen depots behave as a dynamic buffer allowing rapid storage of os- cellular lipid (IMCL) in soleus muscle (SOL) and tibialis anterior muscle (TA) motically active glucose and maximum concentration in both in both muscle were used as indices of insulin resistance.The NGT subjects and patients and liver are seen after the evening meal. However, the effectiveness of with T2DM were comparable with respect to age and sex ratio. At baseline, this diurnal mechanism has not been previously studied in type 2 diabetes. IMCL concentration was obviously higher in patients with T2DM vs. NGT Changes in skeletal muscle and liver glycogen concentration were measured subjects (12.1±3.4mmol/kg vs. 9.07±1.63mmol/kg in SOL, 3.5±1.9 mmol/kg during normal daily eating in type 2 diabetes.13C Magnetic Resonance spec- vs. 2.25±0.95 mmol/kg in TA, p<0.01). After therapy, HOMA IR of patients troscopy at 3.0- Tesla, was used to quantify calf muscle and liver glycogen reduced signifi cantly from 4.0±1.7 to 2.2±1.1. And the IMCL in SOL and TA in the fasting state (0830h) and at 2000h after three defi ned meals (60% were also restored substantially normalized (9.92±2.44 and 2.59±1.39mmol/ carbohydrate, 20% protein, 20% fat). On a separate day, insulin sensitivity kg, respectively, p>0.05 compared with NGT subjects).In conclusion, early was measured by hyperinsulinaemic euglycaemic clamp.In the group of 11 intensive insulin therapy recovered insulin sensitivity in patients with newly type 2 diabetic subjects (7 males, 4 females), the mean fasting blood glu- diagnosed T2DM. Follow-up observation will be needed to study whether cose was 7.9±0.3mmol/l, HbA1c 6.4±0.1% and BMI 31.4±1.1kg/m2. Fasting the improvement will be maintained. muscle glycogen was 62.8±8.4mmol/l, and there was a complete lack of increase over the day of eating. Four hours after the evening meal there 1845-P was no change in mean muscle glycogen (62.6±8.1mmol/l). Fasting liver gly- Pigment Epithelial-Derived Factor (PEDF) and Follistatin-Like 1 cogen was 248.3±24.6mmol/l, and there was a small increase (10.4%) over (Fstl1): Two Novel Contraction-Regulated Myokines the same time period (274±18.4 mmol/l, p=0.12). Within the low range of SILJA LAMBERND, KRISTIN ECKARDT, SVEN W. GÖRGENS, JORGEN JENSEN, diurnal increments in liver glycogen observed, there was no signifi cant cor- JÜRGEN ECKEL, Düsseldorf, Germany, Oslo, Norway relation between glycogen increment and insulin sensitivity (Rd 3.1mg/kg/ Physical activity exerts major benefi cial effects regarding prevention of min; R 0.6, p=0.056).In type 2 diabetes, there is no signifi cant contribution chronic diseases. Most likely, myokines mediate these effects by affect- of skeletal muscle to the storage of glucose during diurnal food intake, and ing muscle physiology and exerting systemic effects on other tissues and minimal contribution of liver glycogen. This failure of the buffering capacity organs. During recent years, increased efforts have focused on analyzing

of glycogen stores contributes to post prandial hyperglycemia and has not the secretory output of skeletal muscle cellsWe sought to characterize the Obesity previously been quantitated. secretome of primary human skeletal muscle cells (hSkMC) by proteomic POSTERS profi ling and identifi ed PEDF and Fstl1 to be secreted by muscle cells among

1843-P others. To analyze the regulation of these myokines by contraction, we used Integrated Physiology/ Effect of Insulin Resistance and Acipimox on Free Fatty Acid Com- our recently established contraction model of hSkMC by applying electric position in Human Subjects pulse stimulation (EPS; 1 Hz, 2 ms, 11.5 V). Similar to in vivo contracting skel- HANYU LIANG, PUNTIP TANTIWONG, RALPH A. DEFRONZO, APIRADEE SRIWI- etal muscle, EPS induced contractile activity in hSkMC combined with the JITKAMOL, JOHN DUBÉ, NICOLAS MUSI, San Antonio, TX, Pittsburgh, PA formation of sarcomeres, activation of AMPK and increased IL-6 secretion. Elevations in plasma free fatty acid (FA) concentration have been associ- PEDF protein abundance in cell lysates increased by 1.25fold after EPS (n=7) ated with insulin resistance. Cell culture and animal studies suggest that and PEDF secretion increased by 1.56fold (n=4). In contrast to this, 60 min saturated FAs have a more deleterious effect on insulin action than unsatu- cycling at 70% of VO2 max decreased PEDF serum levels of healthy young rated FAs. One goal of this study was to examine whether insulin resistant men by 20.1% at 30 min post-exercise (n=8). Fstl1 secretion was signifi cantly [obese and type 2 diabetic (T2D)] subjects have a greater elevation in satu- elevated after 24 h EPS (2.6fold) and inhibited by brefeldin A. Serum levels of rated versus unsaturated FAs. Another goal was to determine whether the Fstl1 in healthy young men were increased immediately after 60min cycling

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A473 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

at VO2 max of 70% (1.2fold, n=8). First results of the incubation of hSkMC control subjects (P=0.01 and P=0.11, respectively). The incretin effect [100% with recombinant Fstl1 indicate that Fstl1 positively regulates insulin sig- × (AUCC-peptide,OGTT - AUCC-peptide,IIGI) / AUCC-peptide,OGTT] amounted to 17±5% and naling by increased insulin-stimulated Akt and GSK3_ phosphorylation and 26±3% in patients with MODY3 and MODY2, respectively (P=NS) and 36±4% enhanced insulin-stimulated glucose uptake.We identifi ed Fstl1 and PEDF in the control subjects (P=0.018 and P=NS, respectively).Our data suggest as novel contraction-regulated myokines, although PEDF serum levels are that MODY2 patients are the fi rst group of diabetes patients ever seen with decreased after exercise. Most likely this refl ects altered PEDF secretion normal incretin effect. In contrast, MODY3 patients were characterized by from adipose tissue, a major source of PEDF. Fstl1 seems to have a positive an impaired incretin effect when compared to control subjects. The differ- autocrine effect on skeletal muscle cells and may be one of the myokines ences are most likely explained by differences in the etiology and glucose mediating the benefi cial effect of physical exercise. tolerance between MODY3 and MODY2 patients.

1846-P & 1848-P Cathepsin D Accumulation Is Responsible for Hyperglycemia-In- Glucagon-Like-Peptide-1 (GLP-1) is Important for the Improved duced Angiotensin II Synthesis in Cardiomyocytes Ƶ-Cell Function in Type 2 Diabetic Subjects After Roux-en-Y Gastric SATORU KOBAYASHI, XIANMIN XU, KAI CHEN, DEREK TIMM, QIANGRONG LI- Bypass (RYGB) ANG, Sioux Falls, SD NILS B. JORGENSEN, CARSTEN DIRKSEN, SIV H. JACOBSEN, KIRSTINE N. BOJS- Although cardiovascular disease is the major cause of death among dia- EN-MØLLER, DORTE WORM, DORTE L. HANSEN, JENS J. HOLST, STEN MADS- betic patients, there are no effective preventive or therapeutic strategies to BAD, Hvidovre, Denmark, Copenhagen, Denmark ease this problem. This is partly due to a lack of understanding of the under- ` cell function is known to improve in type 2 diabetic (T2D) subjects after lying mechanisms that mediate diabetic cardiac complications. We previ- RYGB, but whether this is explained by an increased GLP-1 secretion is not ously showed that the expression and distribution of Cathepsin D (CTSD), an known. We used the GLP-1 receptor specifi c blocker, exendin (9-39) (EX9), to aspartic protease normally located within the lysosome, were dramatically evaluate the role of GLP-1 on post RYGB ` cell function.A liquid meal test was altered in cultured cardiomyocytes exposed to high glucose and in type 1 performed on two separate days before and 1 wk after RYGB in 7 subjects diabetic mouse hearts. We also demonstrated that the CTSD inhibitor pep- with T2D. Fasting blood samples were drawn followed by a bolus-infusion statin A (PepA) protected cardiomyocytes against high glucose-induced (0.14 mL/kg, 0.18 mL/min/kg) of EX9 (1 mg/mL) or isotonic saline (NaCl). The toxicity, indicating that CTSD accumulation contributes to hyperglycemia- order of EX9 and NaCl infusion was randomized. After 30 min of infusion, the induced cardiomyocyte injury. However, it is largely unknown how CTSD meal was ingested over 30 min, and blood was sampled frequently from 30 could cause cardiac injury under hyperglycemic conditions.Recent studies min before to 4 h after meal start. `-cell function was evaluated using the C- have suggested an association between intracellular Angiotensin II (Ang- peptide response above baseline and the insulinogenic index (IGI).After RYGB II) production and cell death in high glucose-treated cardiomyoctes and in fasting P-glucose decreased numerically (pre: 8.5 ± 0.94 mM, post: 7.0 ± 0.57 diabetic rat hearts. In the present study, we investigated if increased Ang-II mM (p=0.078)) and S-insulin levels decreased signifi cantly (127 ± 17 pM, 66 ± synthesis correlates with CTSD toxicity in cardiomyoctes cultured under high 4 pM (p<0.05)). HOMA-IR was halved after RYGB (6.5 ± 0.6, 3.0 ± 0.4 (p<0.05)) glucose conditions. Our results showed that intracellular Ang II levels were and did not differ between the day of EX9 infusion and the day of NaCl infu- increased in high glucose-treated cardiomyoctes, and were associated with sion neither pre nor post RYGB.EX9 infusion increased the area under the elevated CTSD levels and cell death as measured by multiple parameters. glucose curve from 2.2 ± 0.2 M · min during NaCl infusion to 2.7 ± 0.3 M · min Importantly, both PepA treatment and siRNA-mediated knockdown of CTSD (p< 0.05) before and from 1.9 ± 0.1 M · min to 2.4 ± 0.2 vs. (p< 0.05) after RYGB. reversed the increased Ang II levels and attenuated high glucose-induced The C-peptide responses above baseline with NaCl infusion were 251 ± 52 cardiomyocyte death. These results suggest that high glucose-induced nM · min before and 364 ± 69 nM · min (p<0.05) after RYGB, and decreased accumulation of CTSD promotes intracellular Ang-II synthesis, and that in- during EX9 infusion to 241 ± 48 nM · min before and 245 ± 53 nM · min after creased Ang-II levels may be responsible for CTSD cardiotoxicity in diabetes. RYGB, corresponding to 4 ± 6% and 34 ± 4% (p<0.05) reduction. EX9 infusion Future studies will investigate the mechanism by which CTSD triggers Ang-II reduced IGI from 359 ± 46 to 290 ± 54 (p=0.16) before and from 595 ± 83 to 330 synthesis, and confi rm the role of CTSD-Ang-II pathway in diabetic cardiac ± 62 (p<0.05) after RYGB.Infusion of a GLP-1 receptor specifi c blocker blunts injury using whole animal models. the `-cell response during a meal more after than before RYGB resulting in impaired glucose tolerance, suggesting a greater role of GLP-1 in maintaining glucose tolerance and insulin secretion in T2D patients after RYGB. INTEGRATED PHYSIOLOGY—OTHER HORMONES Supported by: Desiree og Niels Ydes Fond

& 1849-P Guided Audio Tour: Incretin Physiology, Pathophysiology, and Humans Post-Prandial Brain Responses to High Calorie Visual Food Cues are (Posters 1847-P to 1854-P), see page 13. Associated With Meal Induced GLP-1 Secretion YEE S. CHEAH, SARAH LEE, FERNANDO O. ZELAYA, MICHAEL J. BRAMMER, & 1847-P STEPHANIE A. AMIEL, London, United Kingdom Incretin Effect in Patients With Maturity Onset Diabetes of the Young Understanding appetite regulation by the gut-brain axis may provide SIGNE H. ØSTOFT, JONATAN I. BAGGER, TORBEN HANSEN, OLUF PEDERSEN, insight into the etiology of obesity and related disorders, including Type 2 JENS J. HOLST, FILIP K. KNOP, TINA VILSBØLL, Hellerup, Denmark, Copenhagen, diabetes. We investigated the normal human responses to food and food Denmark cues using Blood Oxygenation Level Dependent (BOLD) functional magnetic Maturity onset diabetes of the young (MODY) is a clinically and geneti- resonance brain imaging. Nine healthy non-diabetic males (age 33.1±6.6 cally heterogeneous subgroup of non-autoimmune diabetes, constituting yr, BMI 24.2±2.2 kg/m2) were studied twice after an overnight fast. BOLD about 1-2% of all diabetes. The incretin effect (the enhancement of glucose- responses to viewing images of high (HC) and low calorie (LC) foods and induced insulin secretion following oral glucose tolerance test (OGTT) com- non-food objects (O), compared to blurred images presented in a block de- pared to isoglycemic intravenous glucose infusion (IIGI)) is unclear in MODY sign, were measured in a 1.5T MRI scanner 32 mins after a 630 kcal mixed

Obesity patients. In the present study we studied the incretin effect in patients meal (fed) or 50 ml oral water (fasted) using XBAM program for whole brain

POSTERS with MODY2 (glucokinase gene mutations) and MODY3 (hepatocyte nuclear activation analyses. Measuring on a visual analog scale, subjects felt more factor 1_ gene mutations) and in a group of matched control subjects.Ten full (p = 0.004) and less hungry (p = 0.04) immediately pre-BOLD data col-

Integrated Physiology/ MODY3 patients (age: 31±3 years (mean±SEM); body mass index (BMI): lection in the fed compared to fasted state. Effects of meal ingestion on 2 24±1 kg/m ; fasting plasma glucose (FPG): 8.4±0.8 mM; HbA1c: 7.0±0.3%), brain responses to LC and O images were similar, with greater precuneus 9 MODY2 patients (age: 43±5 years; BMI: 24±2 kg/m2; FPG: 7.3±0.3 mM: activation, involved in visuospatial imagery and self processing operations, 2 HbA1c: 6.7±0.2%) and 9 control subjects (age: 41±5 years; BMI: 24±1 kg/m ; and less anterior cingulate cortex and caudate activation, involved in atten- FPG: 5.1±0.2 mM; HbA1c: 5.3±0.1%) were examined on 2 separate occasions: tion, confl ict monitoring and motor control, in the fed compared to the fasted 4h 50 g-OGTT and IIGI.MODY3 patients exhibited more severe glucose in- state. On viewing HC images, meal ingestion led to greater activation of the tolerance (evaluated from area under curve during OGTT) than MODY2 right orbitofrontal cortex (OFC), involved in representing food reward value patients (3,041±302 mM×4h vs. 2,139±59 mM×4h, P=0.0126) compared to and sensory specifi c satiety, compared to the fasted state. In preliminary the control subjects (1,351±20 mM×4h; P<0.001 and P<0.001, respectively). analysis, OFC responses in the fed state were inversely associated with the Isoglycemia was obtained using 37±4 g and 30±3 g of glucose during IIGIs post-prandial incremental area under the curve for GLP-1 concentrations in patients with MODY3 and MODY2, respectively (P=NS), and 24±2 g in (n=5, r = -0.75, p = 0.05). Our data suggest that whilst prior ingestion of a

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A474 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES high calorie meal has similar effects on responses in attention and visual processing networks to viewing images of less rewarding, LC and O, such & 1852-P a meal may induce activation of an appetite control network on viewing Impaired Myocardial Response to Glucagon-Like Peptide 1 in Hu- further high calorie foods, that is associated with GLP-1 secretion. mans With Type 2 Diabetes Mellitus Supported by: Diabetes UK STEVEN P. MOBERLY, GARY D. HUTCHINS, JOHNATHAN D. TUNE, KEVIN PERRY, ROBIN L. CHISHOLM, ROBERT V. CONSIDINE, KIEREN J. MATHER, Indianapolis, IN & 1850-P Effects of GLP-1 to drive myocardial glucose uptake and cardiac work have been reported in animal models but not directly tested in humans. We The Effect of GLP-1 on Food Intake is Lost in Truncal Vagotomized measured effects of GLP-1(7-36) (1.5 pmol/kg/min IV) on myocardial glucose Subjects uptake (MGU), oxygen consumption (MVO2), and blood fl ow (MBF) using ASTRID PLAMBOECK, SIMON VEEDFALD, CAROLYN DEACON, ANDRÈ WETTER- dynamic positron emission tomography. Diabetic subjects underwent 4 wk GREN, LARS SVENDSEN, SØREN MEISNER, CLAUS HOVENDAHL, JENS HOLST, treatment intensifi cation using diet/exercise and insulin. Studies took place TINA VILSBØLL, FILIP KNOP, Hellerup, Denmark, Copenhagen, Denmark, Odense, under fasting conditions following a 10 hour study infusion.Ten lean and 7 Denmark T2D subjects were studied under GLP-1 stimulation, and 6 lean subjects The rapid dipeptidyl peptidase 4-mediated inactivation of glucagon-like served as saline controls. Lean and T2D subjects differed as expected (Table peptide-1 (GLP-1) has led to the hypothesis that the hormone may act locally 1) but by design were matched on fasting fuel concentrations. GLP-1 infusion by activating vagal afferents residing in the intestinal mucosa. We aimed increased plasma GLP-1(7-36) concentrations equally in both groups.GLP-1 to evaluate the effect of exogenous GLP-1 on postprandial plasma glucose had no effect on MBF. GLP-1 signifi cantly increased MVO2 in lean subjects (PG) excursions, gastric emptying (GE), insulin secretion and food intake in (Table 2). MGU was elevated 4-fold by GLP-1 in lean subjects. MGU in the truncally vagotomized subjects.Ten truncally vagotomized subjects (due to GLP-1 treated T2D group was not different than the lean saline control group, duodenal ulcer) with pyloroplasty (age: 70±2 years (mean±SEM); fasting and markedly lower than GLP-1 treated lean subjects despite matched glu- PG (FPG): 5.7±0.1 mM), and 10 matched healthy control subjects (age: 67±1 cose and NEFA levels and hyperinsulinemia. These data indicate that GLP-1 years; FPG: 5.5±0.1 mM) received 300-min infusions of GLP-1 (1.2 pmol×kg- effects on myocardial glucose uptake and cardiac work are impaired in the 1×min-1) or saline. At time 30 min the subject ingested a fi xed liquid meal with setting of T2D. acetaminophen (for evaluation of GE), and after 240 minutes of infusion, an ad libitum meal was served.In the vagotomized group, GLP-1 only lowered Table 1 PG concentrations at time 60 min (9.5±1.0 vs. 7.4±1.0 mM, P<0.05) whereas Lean T2D p value GLP-1 reduced postprandial glucose concentrations below baseline in the BMI (kg/m2) 24.6±3.5 36.8±2.8 0.03 control group (5.4±0.1 vs. 4.2±0.2 mM , P<0.0001). GE was accelerated in vagotomized subjects compared to controls (time to peak plasma acetamino- MAP (mmHg) 84±3 104±7 0.02 phen: 71±16 vs. 152±7 min, P<0.001) on the saline day. GLP-1 decreased GE in Insulin (pmol/L) 76.8±11.4 169.8±36.0 0.03 both groups (time to peak paracetomol: 87±17 vs. 222±5 min, p<0.0001) but NEFA (mmol/L) 0.15±0.07 0.20±0.06 0.18 to lesser extent in the vagotomized group than in the controls. There was Glucose (mmol/L) 5.0±0.3 5.0±0.2 0.91 no effect of GLP-1 on insulin secretion in the vagotomized subjects whereas insulin secretion was suppressed by GLP-1 in the control group (most likely GLP-1 pre-infusion (pmol/L) 6.2±1.6 5.0±0.9 0.59 due to the strong GLP-1-induced inhibition of GE). GLP-1 infusion reduced ad GLP-1 infused (pmol/L) 85±9.6 81.1±10.7 0.86 libitum food intake compared to saline (367±37 vs. 318±29 g, P<0.05) in the control group whereas it had no effect in the vagotomized group (249±31 vs. Table 2 246±44 g, P=NS).The effect of GLP-1 on postprandial PG, GE and food intake is diminished in vagotomized subjects. Thus, our data could indicate that Lean-Sal Lean-GLP1 T2D -GLP1 p p (1) (2) (3) (1 vs 2) (2 vs 3) intact vagal innervation is important for the effects of GLP-1. MVO2 (mL O2/min) 88±3 108±6 93±7 0.03 NS & 1851-P MGU (umol/100mL/min) 21±13 90±27 12±6 0.03 0.02 Effects of Glucagon-Like Peptide-1 on Vasodilation and Glucose Supported by: NIH (HL092799) Uptake in Coronary Arteries JACOB SIVERTSEN, TINA VILSBØLL, SØREN GALATIUS, JAN S. JENSEN, FILIP & 1853-P KNOP, JAYA ROSENMEIER, Hellerup, Denmark Glucose Transport Via SGLT1 is Critical for Post-Prandial GIP Se- Confl icting results exist regarding the benefi cial cardiovascular effects of cretion in Rats and Humans the incretin hormone glucagon-like peptide-1 (GLP-1). Preclinical studies sug- ROBERT L. DOBBINS, LIHONG CHEN, YAPING J. LIU, DANA P. DANGER, SUSAN gest that GLP-1 may affect the cardiovascular system by increased glucose M. ANDREWS, JEFF A. WALD, ANN WALKER, CHARI D. SMITH, Research Tri- uptake in the heart, but this has not been confi rmed in humans. Our aim was angle Park, NC to assess the effect of GLP-1 on human coronary metabolism.We included GSK1613235 and KGA2727 are potent, selective inhibitors of the gastro- 35 patients with and with without diabetes (age: 57 (35-70) years (mean intestinal SGLT1 sodium-dependent glucose transporter. GSK1614235 is in 2 (range)), body mass index: 28 (18-38) kg/m , fasting plasma glucose: 6.2 (4.5- development for improving glucose control in diabetes. Nonclinical stud- 12.1) mM) in a double-blinded randomized crossover design. Patients were ies (KGA2727) and clinical trials (GSK1614235) assessed the translation of referred for coronary angiography and during this procedure they received SGLT1 inhibitor effects in rats to normal human physiology. Following a 16 h two different doses of GLP-1 (physiological (0.1 pmol/kg/min) or supra-phys- fast, Sprague Dawley rats received KGA2727 (0.1 mg/kg) or vehicle 15 min iological (1.0 pmol/kg/min)) and saline for 10 min through a catheter inserted prior to oral gavage with 0.8 g/10 μCi/kg of 3H-3-O-methylglucose (3-OMG). in the left coronary artery. Coronary perfusion was measured using an ultra- Blood and urine samples were collected at 0.025, 0.5, 0.75, 1, 1.5, 2, 4, 6, 8 sonic wire in the same coronary artery. Arterial-venous differences (a-v) of and 24 hours to measure tracer absorption and distribution. SGLT1 inhibition plasma glucose between the left coronary artery and coronary sinus were reduced urine 3-OMG recovery from 731±34 to 292±33 umol and increased calculated thus representing myocardial glucose uptake.Neither physiologi- excretion in the feces. Separate rats were treated with vehicle or 0.1 mg/ Obesity cal nor supra-physiological GLP-1 infusions had any impact on myocardial kg KGA2727 prior to free access to standard chow. Plasma glucose, insulin POSTERS glucose uptake during physiological or supra-physiological GLP-1 infusion and GIP concentrations were reduced in the 4 hour period after the meal (a-v glucose before and after physiological GLP-1 infusion: 0.1±0.2 mM and but total GLP-1 concentrations were unchanged. Using a cross-over design, Integrated Physiology/ 0.2±0.3 mM; P=NS; and supra-physiological GLP-1 infusion: 0.1±0.1 mM and twelve healthy human subjects received GSK1614235 20 mg or placebo im- 0.2±0.1 mM; P=NS). No changes were seen in coronary artery fl ow (fl ow mediately before a mixed meal (550 kcal, 45% CHO). Five minutes into the before and after physiological GLP-1 infusion: 24±7 ml/min and 24±7 ml/min; meal, 3 g of 3-OMG in water was ingested. GSK1614235 delayed and re- P=NS; and supra-physiological GLP-1 infusion: 27±6 ml/min and 28±7 ml/min; duced tracer absorption, with an AUC(0-10h) of 231±31 ug.h/mL relative to P=NS).Intracoronary infusions of GLP-1 do not seem to stimulate endothelial 446±31 ug.h/mL, for placebo. The recovery of tracer in urine was 1.2±0.7 g or myocardial GLP-1 receptors in a way that causes metabolic effect or va- after SGLT1 inhibition relative to 2.2±0.1 g for placebo. Peak glucose con- sodilation in the human heart at fasting glycemic levels. Thus, the present centrations and post-prandial concentrations of insulin, C-peptide and the results do not support an effect of GLP-1 receptor activation on human coro- K-cell product, GIP, were reduced with GSK1614235 for 2 hours following nary perfusion as previously reported in preclinical studies. breakfast. Opposite effects were observed for L-cell products, as total GLP-1 Supported by: Merck Sharpe & Dohme Limited concentrations were signifi cantly increased and a trend for increased PYY

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was noted. SGLT1 inhibitors block intestinal glucose absorption and reduce GIP secretion in rats and humans, suggesting SGLT1 glucose transport is & 1856-P critical for GIP release. Conversely, GLP-1 and PYY secretion are enhanced Activation of GPR119 Receptor With n-Acylethanolamines— by SGLT1 inhibition in humans. Evidence for Linoleoyl Ethanolamide as a Physiological Ligand for Supported by: GlaxoSmithKline GPR119 Receptor SAMREEN K. SYED, HAI HOANG BUI, LISA S. BEAVERS, THOMAS B. FARB, & 1854-P JAMES V. FICORILLI, AMY K. CHESTERFIELD, MING-SHANG KUO, KRISTER B. BOKVIST, ALEXANDER EFANOV, Indianapolis, IN The Effect of GIP on Abdominal, Subcutaneous Adipose tissue Blood GPR119 receptor plays an important role in the secretion of incretin Flow in Obese Subjects hormones in response to nutrient consumption. The endocannabinoid-like MEENA ASMAR, LENE SIMONSEN, JENS JUUL HOLST, JENS BÜLOW, Copen- lipid, oleoylethanolamide (OEA), and similar compounds have been shown hagen, Denmark to activate GPR119 receptor. However, the identity of particular endogenous Recently, we showed that Glucose-dependent insulinotropic polypep- compound(s) that mediate physiological activation of GPR119 in the intestine tide (GIP) in combination with hyperinsulinemia and slight hyperglycemia is not known.We have studied activation of GPR119 with an array of naturally increased abdominal, adipose tissue blood fl ow (ATBF), glucose uptake, occurring endocannabinoid-like compounds using heterologously expressed and free fatty acid re-esterifi cation, thus resulting in increased triglyceride GPR119 in a cell- based cAMP assay. The potencies for the studied n-acyle- deposition in abdominal, subcutaneous adipose tissue in lean humans. The thanolamines in activating GPR119 ranked as follows: OEA linoleylethano- present experiments were performed in order to elucidate the effects of * lamide (LEA)> palmitoylethanolamide (PEA) dihomo-gamma-linolenoyleth- GIP on ATBF in obese subjects with either normal or impaired glucose toler- * anolamide > stearoylethanolamide (SEA) > arachidonoylethanolamide (AEA). ance.16 obese (BMI>30 kg/m2) subjects were studied. The subjects were Interestingly, P450 oxidative metabolites of AEA - epoxyeicosatrienoic acid divided into 2 groups, based upon their plasma glucose response to an oral ethanolamides (EET-EA) are signifi cantly more potent in activating GPR119 glucose challenge: (i) normal and (ii) impaired glucose tolerance. ATBF was receptor than parental AEA. For lipids containing oleoyl fatty acid chains assessed during GIP (1.5 pmol/kg/min) infused intravenously in combination and different polar heads, OEA was the most potent GPR119 agonist, while with a hyperinsulinemic-hyperglycemic clamp.In the subjects with impaired oleoylserotonin, oleoylglycerol and oleoylacetylglycerol displayed lower glucose tolerance, ATBF remained virtually constant throughout the study potencies.Endogenous levels of the most active n-acylethanolamines (OEA, (1.1±0.1 ml min-1 (100 g tissue)-1). In the subjects with normal glucose tol- LEA, PEA, SEA, AEA and EET-EA) have been measured in different sections erance ATBF increased transiently but signifi cantly during the fi rst 60 min of the intestine in fasted and re-fed mice. Only two lipids, OEA and LEA, (2.01±0.3 ml min-1 (100 g tissue)-1), P=0.03) and then began to decrease to showed signifi cant increases in their levels upon re-feeding. OEA was in- reach the basal level again after 150 min. However, compared to ATBF in the creased in the fed state in the duodenum and jejunum. In the fed state a lean subjects from our previous study the increase in ATBF was signifi cantly strong 4-fold increase in OEA levels was seen in the jejunum, with a doubling lower (P<0.0001). In conclusion, GIP in combination with hyperinsulinemia of OEA levels observed in duodenum. LEA levels were increased in fed ani- and slight hyperglycemia increases ATBF in healthy lean subjects. This ef- mals in all four of the intestinal sections examined. The strongest increase fect is blunted in obese subjects with normal glucose tolerance and virtually in LEA levels was observed in the jejunum (34-fold). In the colon, which has absent in obese subjects with impaired glucose tolerance. This blunted ATBF the highest level of GPR119 expression, LEA levels were increased 4-fold in response may have an impaired effect on the handling of free fatty acids. the fed state. Fed levels of LEA were the highest of all measured n-acyle- thanolamides in any portion of the intestine. Compared to OEA levels, fed LEA levels were 4-15 fold higher.Our data demonstrate that LEA is a potent Guided Audio Tour: Regulations of Insulin and Glucagon Secretion GPR119 agonist with high intestinal levels, which are regulated by feeding. (Posters 1855-P to 1862-P), see page 15. Therefore, LEA is a conceivable candidate for a physiologically relevant en- dogenous GPR119 ligand. & 1855-P Up-Regulated Insulin Secretion After GPR119 Activation in Insulin & Resistant Mice: Dual Mechanisms Involving both Enhanced GLP-1 1857-P GPR43 Agonists Stimulate Secretion of GLP-1 Secretion and Exaggerated GLP-1 Receptor Expression KELLY WILBUR, DAVID B. WAINSCOTT, CHARLIE CHANGZHI HU, DAVID BARRETT, LINDA AHLKVIST, KATHLEEN BROWN, BO AHRÉN, Lund, Sweden, Research Tri- DONALYN SCHEUNER, YANYUN CHEN, ANNE REIFEL MILLER, Indianapolis, IN angel Park, Durham, NC Glucagon-like peptide-1 (GLP-1) is a gut hormone that plays a vital role We previously showed that the `-cell sensitivity to glucagon-like pep- in glucose homeostasis through its effects on glucose-dependent insulin tide 1 (GLP-1) is increased in insulin resistant mice. This is mechanistically secretion and gastric emptying. Stable GLP-1 mimetics, as well as small mol- still unresolved but may be therapeutically leveraged by increasing GLP-1 ecule agents that inhibit degradation of GLP-1 by DPPIV have demonstrated levels via activation of G-protein coupled receptor 119 (GPR119). This study therapeutic utility in the treatment of patients with type II diabetes. Agents therefore examined, fi rst, whether GPR119 activation elicits a stronger in- that stimulate endogenous GLP-1 secretion offer an attractive alternative to crease in GLP-1 and insulin levels in insulin resistant vs. control mice, and, these approaches. In the search for such agents, we have investigated the second, whether the increased `-cell sensitivity to GLP-1 in insulin resis- ability of agonists to G-protein coupled receptor 43 (GPR43, FFA2) to stimu- tance is associated with increased GLP-1 receptor (GLP-1R) expression.Fe- late GLP-1 secretion. GPR43 is a type 1 GPCR found in human ascending male C57BL/6J mice were fed a control (CD, 10% fat) or high-fat (HFD, 60% colon, adipocytes, pancreas and immune cells and activated by short chain fat) diet for eight weeks. Then, anesthetized mice were orally given a GPR119 fatty acids. Real-time polymerase chain reaction was utilized to determine agonist (GSK706A; 10 mg/kg, GlaxoSmithKline) or vehicle (saline). After 10 GPR43 mRNA levels in a murine intestinal tumor-derived cell line (STC-1) min, the mice were orally given a mixture of glucose (60%), protein (20%) and in ileum, duodenum, jejunum and colon of normal mice. Using a wheat and fat (20%) resembling a mixed meal (0.285 kcal) with blood collected for germ agglutinin, [35S]GTP S binding assay with membranes from HEK293 determination of glucose, insulin and active GLP-1. Pancreases were isolated a cells expressing cloned human GPR43, we confi rmed the GPR43 ago-allos- for GLP-1R and actin expression analysis with Western blot.HFD mice had teric modulation reported by Amgen for phenylacetamide compound A (EC50 Obesity higher baseline insulin than CD mice (322±23 vs. 179±11 pM, P<0.001), as of 2.1 uM, 129% effi cacy compared to acetate), as well as the orthosteric POSTERS evident for insulin resistance. In HFD vs. CD mice, GPR119 agonism mark- agonism reported by Euroscreen for pyrrolidine carboxylic compound O (EC edly exaggerated the increase in both AUC GLP-1 (81±9.6 vs. 37±6.9 pM x 50 of 0.16 uM and effi cacy of 120%). We then investigated the ability of both

Integrated Physiology/ 20 min, P=0.002) and AUC insulin (253±29 vs. 112±19 nM x 60 min, P<0.001) compounds to stimulate GLP-1 secretion in STC-1 cells and in C57BL/6 mice. after mixed meal. Pancreatic GLP-1R expression was higher in HFD (6.9±1.1 Our data demonstrate that Compound A provides robust stimulation of GLP-1 vs. 3.6±1.2 arb units, P=0.06).Hence, GPR119 agonism up-regulates insulin secretion both in vitro and in vivo’ whereas, the secretory effects of Com- secretion in insulin resistance which is likely caused by both increased GLP- pound O are less apparent. These data indicate that activation of GPR43 1 levels and exaggerated responsiveness to GLP-1, partly mediated by en- stimulates GLP-1 secretion and that the nature of the agonist may play a hanced GLP-1R expression. These results provide both a mechanistic basis role in this function. Finally, these data suggest that GPR43 agonists offer for increased `-cell sensitivity to GLP-1 in insulin resistance and a rationale potential as therapeutics in the treatment of type II diabetes through their for targeting GPR119 in diabetes therapy. action as GLP-1 secretogogues.

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A476 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

& 1858-P & 1860-P GPR40 Agonists Lower Glucose via Improvements in Insulin Secre- Interleukin-6 Amplifi es Adrenergic-Dependent Glucagon Secretion tion, Hepatic Glucose Production, and Hepatic Gene Expression in TAMMY M. LUNDBLAD, ANASTASIA COLDREN, MARCELA BRISSOVA, VALEN- the Goto Kakizaki Rat TINE CHUKWUMA, TASNEEM ANSARI, OWEN P. MCGUINNESS, Nashville, TN MARIA E. TRUJILLO, ERIC S. MUISE, MELISSA E. KIRKLAND, DANIEL KOSIN- Circulating levels of interleukin-6 (IL-6) and glucagon (GCG) are elevated SKI, MICHELE J. PACHANSKI, JIN CAO, CORIN MILLER, ALEKSANDER PETROV, in infl ammatory disease. Our prior work demonstrated that IL-6 knock-out GEORGE J. EIERMANN, MARGARET VANHEEK, ANDREW J. NICHOLS, LIHU mice (IL-6 KO) have a blunted GCG response to endotoxin that is restored by YANG, ANDREW D. HOWARD, DAVID E. KELLEY, PAUL E. CARRINGTON, Rahway, the acute replacement of IL-6. We hypothesized that IL-6 is also required for NJ the response to hypoglycemia. Using chronically catheterized (carotid artery GPR40 agonists lower glucose via improvements in insulin secretion, he- and jugular vein) conscious mice, we performed hyperinsulinemic (10mU/kg/ patic glucose production, and hepatic gene expression in the GK ratGPR40 is min) hypoglycemic (~60 mg/dl) clamps. The GCG response was blunted in IL-6 a free fatty acid receptor that is highly expressed in the pancreas and regu- KO mice (213±74 vs. 867±294 pg/ml; IL-6 KO vs. WT), despite an amplifi ed lates glucose in part through augmentation of glucose dependent insulin epinephrine (1049±298 vs. 308±123, pg/ml) response. No signifi cant differ- secretion (GDIS). Recent publications showed treatment of diabetic humans ences in insulin concentration, corticosteroids, or norepinephrine response with a GPR40 agonist (GPR40ag) lowered fasted glucose, suggestive of an were observed. To assess if IL-6 could directly alter alpha cell function, islets additional non-GDIS mechanism. To test this hypothesis, we administered isolated from IL-6 KO mice were placed in a cell perifusion system. Islets GPR40ag to GK rats. Acute treatment with a GPR40ag at 10 mg/kg resulted were perifused in low glucose (1.7 mM) or in low glucose plus known secret- in improvement in glucose AUC in an OGTT by 35% (vs. vehicle, p< 0.001) agogues of GCG (1 μM epinephrine or 20 mM arginine), either in the absence without corresponding changes in plasma insulin. Chronic treatment of GK or presence of recombinant mouse IL-6 (200 ng/ml). While IL-6 had no ef- rats for 14 d with 10 or 30 mg/kg of a GPR40ag decreased glucose under fect on GCG secretion (pg/islet equivalents) in low glucose alone (85±32 vs. both basal (1h post lights on, ~20%, p< 0.01) and 5h fasted (~30%, p< 0.01) 67±11; without IL-6 vs. with IL-6) or with arginine (117±17 vs. 121±45), IL-6 conditions. These effects were not associated with changes in plasma in- augmented (p<0.05) the GCG response to epinephrine (765±76 vs. 1017±55). sulin. Since high hepatic extraction and low insulin secretion could obscure Insulin secretion was unaltered by IL-6. Thus, IL-6 may act as a modulatory GPR40-mediated effects, we tested the effects of a GPR40ag on insulin se- cytokine to sensitize the alpha cell to mediators of the sympathetic nervous cretion in situ using pancreatic perfusions. GPR40ag increased basal insulin system. Further work is needed to defi ne the receptor populations necessary secretion and stimulated GDIS in the otherwise non-glucose responsive GK for IL-6-mediated amplifi cation of alpha cell adrenergic signaling. rat pancreas. To test for extra-pancreatic effects, we measured GPR40ag ef- fects on the conversion of [2-13C] pyruvate to glucose in vivo. Both acute and chronic treatment with GPR40 agonist decreased 13C glucose production. To explore mechanisms mediating these effects, livers were harvested from GK rats after chronic treatment and compared to vehicle treated and Wistar (non-diabetic) rat controls. Of the ~3400 genes differentially expressed be- tween Wistar and GK rats, >400 genes were also regulated in the liver with GPR40 treatment, of which 85% were in the direction of disease correction. Together, these data in the GK rat provide further mechanistic insights un- derlying the effi cacy of GPR40 agonism.

& 1859-P Identifi cation of Gut Factors that Mimic the Metabolic Benefi ts Seen After Gastric Bypass Surgery LEI LING, DAN KAPLAN KAPLAN, DARRIN A. LINDHOUT, MAZI SABERI, JIAN LUO, QING XIE, SHANNON GAO, PENG ZHANG, DAVID BAI, JIM AKINS, HONG YANG, R.M. LEARNED, THOMAS F. PARSONS, HUI TIAN, JIN-LONG CHEN, ALEX M. DEPAOLI, San Francisco, CA Gastric bypass (GB) leads to acute remission of T2D in many patients be- fore signifi cant weight loss. This metabolic improvement is heralded by both an increase in insulin secretion, perhaps explained in part by GLP-1, and an improvement in insulin sensitivity, at this point unexplained.To identify genes from the gut that might explain this observation, we conducted a systematic genomic study of Zucker Fatty rats either before or after a weight neutral GB procedure, the duodenal-jejunal bypass. Potential mediators of the surgi- cal effect were identifi ed based on a comparative analysis of genes up- or down-regulated by at least 2-fold in the gut. Using a predictive algorithm for Supported by: DK043748, DK059637, DK020593, DK007563 secreted factors, 150 proteins were selected for metabolic phenotyping us- ing an in-vivo protein expression system in a diet induced obese (DIO) mouse & 1861-P model of insulin resistance and diabetes. Based on this approach, NP258 and FGF21 is Expressed in Pancreatic Alfa-Cells and Contributes to El- NP266 were identifi ed as regulated secreted molecules that signifi cantly evated Circulating Levels of FGF21 in the Glucagon Receptor Knock- impact glucose metabolism.NP258 signifi cantly reduced or normalized glu- out Mice cose in DIO, ob/ob, db/db and TALLYHO mice and signifi cantly reduced body BIRGITTE ANDERSEN, JACOB HALD, KAREN AREVAD, RICHARD GELLING, BO weight in DIO and ob/ob mice, but not in db/db and TALLYHO mice. In obese AHRÉN, ERICA NISHIMURA, Maaloev, Denmark, Singapore, Singapore, Malmoe, diabetic patients NP258 serum levels were found to be reduced compared Sweden Obesity POSTERS with lean controls. NP258 levels increased after GB, although not after gas- Fibroblast growth factor (FGF21) is a metabolic regulator of glucose tric banding, consistent with the superior acute glycemic benefi t seen with and lipid metabolism. Treatment of rodent models of Type 2 diabetes with the GB procedure.NP266 is a gut hormone that elevates fasting and fed glu- rFGF21, lowers blood glucose, improves dyslipidemia and increases energy Integrated Physiology/ cose levels and worsens glucose tolerance with apparently diminished insu- expenditure. FGF21 is highly expressed in the liver and in the pancreas. He- lin secretion. In diabetic patients NP266 is increased and interestingly down patic FGF21 is induced by fasting and is a downstream target of PPAR_. In regulated after GB. Neutralizing antibodies are being developed.NP258 and this study we investigated the cellular localization of FGF21 in pancreatic NP266 are secreted GI factors that are regulated by gastric bypass and may islets and studied the effect of _-cell hyperplasia on plasma levels of FGF21 mediate the metabolic improvements seen with the procedure. The substan- in glucagon receptor knockout mice (GRKO).Islets from adult mice (NMRI, tial glucose and weight loss benefi ts of NP258 provide support for initiation GRKO and control mice) were isolated, embedded in paraffi n and stained of clinical trials with an optimized form of NP258 in patients with T2D. for insulin, glucagon and FGF21 (Abcam-ab66564). To test the FGF21 anti- serum specifi city a blocking experiment with rFGF21 was performed and showed that the FGF21 anti-serum was specifi c for FGF21. Plasma samples from GRKO (n=6) and control mice (n=6) were measured for FGF21 using an

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ELISA.FGF21 was found to be expressed in the islet _-cells. Beta-cells also compared to ZT2, along with a 70% increase in the insulin response during stained positive for FGF21, however this was at a lower level of expression the fi rst 30 min. In contrast, glycaemia and 30 min glucose/insulin were not compared to that observed in the _-cells. GRKO mice have previously been different. No circadian pattern was found in the L cells treated with me- shown to display _-cell hyperplasia and staining of the pancreas from GRKO dia alone. However, similar circadian patterns were observed in the GLP-1 mice demonstrated a very high level of FGF21 expression in the _-cells. Fur- response to both bethanechol [period (Tau)=28.6±1.2 h, amplitude (A)=41±7 thermore, plasma FGF21 was signifi cantly (p=0.0072) increased in the GRKO pg/ml] and PMA (tau 23.6±1.1 h, A=32±9 pg/ml). Together, our data demon- (9730 ± 2660 pg/ml) compared to control mice (630 ± 603 pg/ml).The dem- strate a daily variation in in vivo GLP-1 responses, being greater in the period onstration of FGF21 expression in pancreatic _-cells indicates that in addi- of time (ZT14) when food intake is greatest (night for rodents). The fi nding of tion to glucagon expression, _-cells produce another hormonal regulator of a circadian rhythm in vitro in response to both bethanechol and PMA sug- glucose and lipid metabolism. Furthermore, _-cell hyperplasia in the GRKO gests regulation by downstream pathways mediating GLP-1 exocytosis in mice can lead to an increase in plasma levels of FGF21. It is of great interest the intestinal L cell. that streptozotocin treatment of the GRKO mice further increases the _-cells Supported by: Canadian Diabetes Association mass and that these mice do not develop diabetes. This suggests that FGF21 may play a major role in the anti-diabetic effect in these mice. & 1864-P Electrical and Mechanical Stimulation of the Duodenum Increases & 1862-P GLP-1 Levels DARLEEN A. SANDOVAL, ADAM DUNKI-JACOBS, JOYCE SORRELL, RANDY J. Fibroblast Growth Factor 21 and Farnesoid X Receptor Mediate SEELEY, DAVID D’ALESSIO, Cincinnati, OH Chronic Glucagon Action Increases in GLP-1 are proposed to serve as a negative feedback signal for KIRK M. HABEGGER, KERSTIN STEMMER, DAVID SMILEY, TIMO D. MUEL- postprandial changes in gastric emptying and/or motility. Previous ex vivo LER, NICKKI OTTAWAY, JAZZMINN HEMBREE, RADHA KRISHNA, AYMAN M. data suggested that direct electrical stimulation (E-stim) of intestinal L-cells ARAFAT, STEVEN WOODS, SUSANNA M. HOFMANN, DAVE D’ALESSIO, PAUL increases release of GLP-1. This suggests potential feed-forward increases PFLUGER, DIEGO PEREZ-TILVE, NOBUYUKI ITOH, JOACHIM SPRANGER, RICHARD in GLP-1 driven by intestinal neuronal and/or motor activity. To determine if DIMARCHI, MATTHIAS H. TSCHOP, RANDY J. SEELEY, Cincinnati, OH, Blooming- E-stim could increase GLP-1 levels in an in vivo setting, we studied male Long ton, IN, Munich, Germany, Berlin, Germany, Neuherberg, Germany, Kyoto, Japan Evans rats (300-350g) under general anesthesia. The peritoneal cavity was Glucagon, an essential regulator of glucose homeostasis, also modulates opened and two ~1cm long electrode cuffs were placed around the outside lipid metabolism and promotes weight loss, as refl ected by the wasting of the duodenum, 2cm distal to the pylorus, or around the ileum, 4 cm proxi- syndrome observed in patients with glucagon producing tumors. Recently, mal to the cecum. A catheter was placed after the electrodes for nutrient novel single molecule co-agonist drugs that include glucagon agonism have infusion (3ml) of a mixed liquid diet (Ensure). Each animal had two 30min emerged as promising therapeutic candidates for obesity and diabetes. We back to back experimental periods. The 1st 30min consisted of either direct developed a novel stable, soluble and highly specifi c, long-acting glucagon intestinal E-stim or nutrient infusion. During the 2nd 30min period all animals receptor agonist, which made detailed in vivo dissection of the chronic glu- had a combination of E-stim plus nutrient infusion. In both the ileum and duo- cagon action profi le more feasible. Chronically enhanced glucagon recep- denum, nutrient infusion alone, but not E-stim alone, signifi cantly increased tor agonism in mice resulted in hyperglycemia, lower body fat and reduced plasma GLP-1 levels (p<0.05). However, the combination of E-stim and nutri- plasma cholesterol, but notably also raised expression as well as circulating ent infusion, in either the ileum or duodenum, signifi cantly increased plasma protein levels of fi broblast growth factor 21 (FGF21), a secreted liver protein GLP-1 when compared to nutrient infusion alone (p<0.05). E-stim could be in- and obesity drug candidate. We confi rmed this link in healthy human volun- creasing GLP-1 via increased intestinal muscle activity. Therefore, in another teers, where infusion of natural glucagon increased plasma FGF21 within series of experiments we studied the impact of extra-luminal mechanical hours. Functional relevance became evident in mice with genetic deletion of manipulation (M-stim) on GLP-1 levels. In the duodenum, but not the ileum, FGF21, where glucagon receptor activation failed to induce the body weight M-stim plus nutrient infusion also signifi cantly increased GLP-1 over nutrient loss, hyperglycemia and lipid metabolism changes observed in wt mice. infusion or M-stim alone (p<0.05). Thus, E- and M-stim of the duodenum, and Since we also observed that glucagon receptor activation induced hepatic E-stim of the ileum augment nutrient-stimulated GLP-1 release. Future stud- expression of the farnesoid X receptor (FXR), we challenged FXR defi cient ies are aimed at determining whether E-stim-induced changes in neuronal mice with glucagon, where it failed to induce the body weight loss and lipid activation and/or intestinal motility are the mechanism(s) by which E-stim metabolism changes observed in wt mice. Taken together, these data show increases in GLP-1. for the fi rst time that glucagon controls glucose, energy and lipid metabolism Supported by: Eithicon Endo-Surgery via FXR and FGF21 dependent pathways.

& 1865-P Guided Audio Tour: Preclinical Models of Incretin Physiology and Thera- Role of GLP1 Receptor in DGAT-1 Induced Glucose Lowering in Dia- peutics (Posters 1863-P to 1869-P), see page 17. betic Rodents JUDITH N. GORSKI, STEPHANIE CRAW, CAROL ANN KEOHANE, XIAORUI YAO, & 1863-P GAIL FORREST, JOYCE HWA, SHIRLY PINTO, Rahway, NJ Circadian Variations in Glucagon-Like Peptide-1 Responses Current literature has focused on the anti-obesity therapeutic potential MANUEL GIL-LOZANO, ERLI MINGOMATAJ, PATRICIA L. BRUBAKER, Toronto, ON, of inhibiting diacylglycerol acyl transferase-1 (DGAT1), key enzyme in trig- Canada lyceride resynthesis during intestinal absorption. Here we focus on antidi- Daily rhythms in glucose tolerance and insulin secretion have been re- abetic potential following DGAT1 inhibition. DGAT1-defi cient (-/-) mice are ported. As a gut L cell hormone stimulating insulin release, glucagon-like resistant to diet-induced obesity (DIO) and have improved insulin sensitivity. peptide-1 (GLP-1) may play a role in this pattern. Accordingly, previous stud- However, their lean phenotype may contribute to increased insulin sensitiv-

Obesity ies have shown that the GLP-1 response to a meal varies during the day in ity. We show chronic DGAT1 inhibition signifi cantly lowers non-fasting glu- POSTERS humans. Although various pathways are known to regulate nutrient-induced cose in dbdb mice independent of weight loss (Fig.1). Since DGAT1 inhibition GLP-1 secretion, the vagus nerve is an important mediator of the rapid L cell signifi cantly increases postprandial GLP1 levels; we further evaluated the

Integrated Physiology/ response to a meal. The present study was thus conducted to determine potential mechanism of action for the antidiabetic effects in Glp1r -/-mice. the regulation of circadian GLP-1 responses, using in vivo and in vitro ap- DGAT1 inhibition led to signifi cant elevation of post prandial plasma insulin proaches. In vivo, the bioactive GLP-1, insulin and glucose responses follow- levels in WT mice, which was devoid in Glp1r -/- mice (Fig.2). There was a ing an OGTT (3 g/kg) were examined at ZT2 and ZT14 (lights on: ZT0-ZT12) in signifi cant reduction in body weight and food intake and a postprandial de- male Wistar rats fasted for 4 h. Each animal was studied at both time points lay in gastric emptying in both WT and HF-fed Glp1r -/-mice suggesting that (n=6). Blood was sampled at t = 0, 10, 20, 30 and 60 min. In vitro, GLP-1 GLP1 signaling alone is not responsible for these effects. Taken together, secretion by GLUTag L cell was determined in response to bethanechol, a our studies suggest the benefi cial and potential use of DGAT1 inhibitors as a muscarinic receptor agonist that activates PKC (1 mM), and PMA, a receptor- novel therapeutic approach for the treatment of diabetes. independent PKC activator (1 μM), vs media alone. Two h secretion assays were performed in synchronized cells every 4 h for 48 h (n=8). A 100% in- crease in the in vivo GLP-1 response to oral glucose was observed at ZT14 as

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A478 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

2 weeks. Morphometric analysis revealed no difference in the number of L cells in distal ileum of control and Ex-4 treated mice. Ex-4 did not induce L cell proliferation, since GLP-1 positive cells of Ex-4 mice, like those of con- trol, did not express the proliferation marker Ki67. The length of the transit amplifying (TA) region of the crypts was similar in control and Ex-4 mice.To assess whether GLP-1 may act synergistically with other hormones that are also degraded by DPP4, two month old CD-1 mice were fed ad libitum for two months with either a normal chow diet (rodent chow no. 5015 Purina, Re- search Diets) or diet containing 4g/kg of the DPP4 inhibitor MK0626 (Merck). This drug has been shown by Merck to signifi cantly reduce DDP4 activity in circulation in mice one week after the initiation of treatment. The treatment did not affect body weight or random blood glucose levels. Morphometric analysis indicated that inhibition of DPP4 activity induced an augmentation in the villus surface area, crypt depth and in the number of L cells per surface area in the distal ileum (control vs DPP4 inhibitor: 0.16+/- 0.013 and 0.249+/- 0.035 per surface area, p<0.05). DPP4 inhibition induced a 25% increase in the length of the TA region and the appearance of few L cell clusters ex- pressing Ki67. In contrast to L cells, the number of CCK cells was similar in both groups of mice in the distal ileum. We conclude that inhibition of DPP4, but not the administration of Exendin 4, results in an increase in L cell num- ber in distal ileum. These fi ndings provide a promising approach to regulate GLP-1 levels in diabetic patients.. Supported by: Merck

& 1868-P Combinatorial Anti-diabetic Effects of Exenatide and Dapaglifl ozin in Diabetic Mice KRYSTYNA TATARKIEWICZ, CLARA POLIZZI, EMMANUEL SABLAN, DAMON MC- CUMBER, DAVID PARKES, San Diego, CA Exenatide (Ex), a GLP-1 receptor agonist, has been shown to improve gly- cemic control in patients with T2DM. Dapaglifl ozin (Da), a sodium-glucose & 1866-P co-transporter 2 inhibitor, regulates glucose reabsorption by the kidney and Resistin Knockout Mice Exhibit Impaired Adipocyte Glucose-De- has been proposed as a novel treatment for diabetes. It is not known if the pendent Insulinotropic Polypeptide Receptor (GIPR) Expression combination of Ex + Da (E+D) has greater glucose-lowering effects than ei- SU-JIN KIM, CUILAN NIAN, MITCHELL A. LAZAR, CHRISTOPHER H.S. MCIN- ther compound alone. Hence, we investigated this combination in diabetic TOSH, Vancouver, BC, Canada, Philadelphia, PA ob/ob mice.Basal glucose lowering, oral glucose tolerance (OGTT) and 24 h Glucose-dependent Insulinotropic Polypeptide (GIP) and Glucagon-like urine glucose excretion were measured after a single dose of Ex (0.03 mg/ Peptide-1 (GLP-1) are the two major incretin hormones that constitute the kg, SC), Da (1 mg/kg, PO), E+D, or vehicle (V).Ex (7.2 nmol/kg/d, SC infusion), hormonal arm of the enteroinsular axis and there is strong evidence that GIP Da (1 mg/kg/d, PO), E+D, or V were dosed for 4 wk. A1C, body weight (BW), also plays an important regulatory role in fat metabolism. In earlier studies food intake (FI), and terminal glomerular fi ltration rate (GFR) were measured. on the stimulatory effect of GIP on 3T3-L1 adipocyte lipoprotein lipase (LPL), Data are presented as mean±SEM; P<0.05 by ANOVA.V-corrected (corr) bas- resistin was demonstrated to be a key mediator of a GIP-mediated pathway. al glucose reduction (mg/dL) with Ex alone (-322±14) or Da alone (-286±60) The current study was initiated in order to determine whether resistin plays was less than that observed with the E+D (-507±42) at 1 h post dose and additional roles in GIP-mediated actions on adipocytes. Using primary adi- stayed signifi cantly lower for all groups vs. V during 3 h measurements. pocytes isolated from resistin homozygous knockout (Retn-/-) mice, heterozy- During the 2 h OGTT, glucose excursion (mg/dL) for E+D ranged between gotes (Retn+/-) and wild-type (WT) littermates (Retn+/+), GIP stimulated the 119-275, and was signifi cantly lower than for Ex (498±29) or Da (681±124) PKB/LKB1/AMPK/LPL pathway in Retn+/+ adipocytes, but not in Retn+/- or alone, or V (761±51) 30 min post dose. All groups reached peak glucose at 15 Retn-/- adipocytes, suggesting that a critical level of resistin signaling is es- min post dose (Ex=528±32, Da=895±62, E+D=275±92, V=1112±71). Glucose sential for GIP-regulated LPL activity. In parallel studies, both GIP receptor excretion (g/24 h) was reduced by the E+D (1.1±0.1) vs. V (2.2±0.3) but not by (GIPR) protein and mRNA were shown to be down-regulated in Retn+/- and Ex (1.4±0.2) or Da (2.3±0.3) alone.After 4-wk treatment, Ex, Da, and E+D simi- Retn-/- adipocytes, compared to Retn+/+ adipocytes. During screening to larly decreased V-corr A1C by 2.0±0.2, 1.6±0.4, and 2.4±0.1%, respectively. identify modulators of adipocyte GIPR expression, genes encoding tumor V-corr BW reduction by Ex and E+D ranged between 5-7%. Da alone had no necrosis factor (TNF) and one of its cognate receptors (TNFRS1b), as well effect on BW or FI vs. V. GFR (mL/h) was similar for Ex (20±2), Da (27±3), and V as the downstream signaling molecule, SAPK/JNK were found to be down- (19±2) and increased in the E+D group (33±3).In summary, Ex combined with regulated in Retn-/- epididymal white adipose tissue (eWAT). It was subse- Da regulated basal and postprandial glucose better than either compound quently established that phosphorylated levels of c-jun and SAPK/JNK were alone at doses tested. Optimized combinatorial doses may result in additive decreased in Retn-/- eWAT, when compared to Retn+/+. Using ChIP assays, glucose-lowering benefi ts for patients with T2DM. a functional element responsible for c-jun-mediated transcriptional activa- tion of the GIPR has been identifi ed. Blunted GIP responsiveness observed & +/- -/- 1869-P in Retn and Retn adipocytes is therefore at least partially due to reduced Brain Action of Amylin Increases Regional Sympathetic Nerve Traffi c GIPR expression, resulting from decreased c-jun-mediated transcriptional CAROLINE FERNANDES-SANTOS, DONALD A. MORGAN, KAMAL RAHMOUNI, Obesity +/- -/- activation of Gipr in Retn and Retn adipocytes. Iowa City, IA POSTERS Supported by: CDA, CIHR, and NIDDK The pancreatic amylin acts in the brain to decrease food intake and pro-

mote energy expenditure. We hypothesized that amylin action in the brain Integrated Physiology/ & 1867-P promotes energy expenditure by increasing the activity of the sympathetic The Number of Enteroendocrine L Cells Increases Following Inhibi- nervous system. First, we examined whether amylin modulate neuronal ac- tion of DPP4 but not After Exendin 4 Administration tivity in the brain nuclei that are critical for sympathetic nerve regulation. In MARINE GRIGORYAN, MAMDOUH H. KEDEES, YELENA GUZ, GLADYS TEITEL- C57BL/6J mice, intraperitoneal administration of amylin (10 μg/kg) signifi - MAN, Brooklyn, NY cantly (P<0.01 vs. vehicle) increased c-Fos immunoreactivity in hypothalamic To date, the effect of activation of GLP-1 receptor signaling on intesti- (+65% increase in the dorsomedial hypothalamic nucleus) and brainstem nal epithelial cell differentiation is unknown. To elucidate this question, we (+250% increase in the nucleus of the solitary tract) regions involved in compared the effects of GLP-1 receptor agonist Exendin 4 (Ex-4) and Dipepti- sympathetic regulation. Consistent with the involvement of brain amylin in dyl peptidase-4 (DPP4) inhibition on L cell number. Two-month old CD-1 mice energy homeostasis, intracerebroventricular (ICV) administration of amylin were injected ip twice daily with Ex-4 (1 nmol/Kg) or vehicle (0.9% saline) for (0.1 and 0.2 nmol) caused a signifi cant (P<0.05) and dose-related decrease

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A479 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

in food intake and body weight, measured 4 and 24 hrs after treatment in sion. We conclude that BMP signaling regulates regional patterning of hu- C57BL/6J mice. Using direct multifi ber sympathetic nerve recording, we also man intestine during development and that intestine generated in vitro from investigated the ability of ICV amylin to increase sympathetic nerve activ- PSCs is capable of region-specifi c incretin expression. This work will have a ity (SNA) in C57BL/6J mice. ICV amylin elicited a signifi cant (P<0.01) and signifi cant impact on the design and application of new treatment options dose-dependent increase in SNA subserving thermogenic brown adipose for diabetes, including therapeutic transplantation of intestinal tissue and tissue (BAT), with a 250% increase after the 0.2 nmol dose. ICV injection of the testing of potential incretin secretagogues. amylin also evoked a signifi cant (P<0.01) and dose-related increase in lumbar and renal SNA with a 338% and 203% increase, respectively, after the 0.2 1872-P nmol dose. Importantly, ICV pre-treatment with the amylin receptor antago- The Infl uence of 25-Hydroxyvitamin D Levels on Response to Anti- nist (AC187, 20 nmol) abolished the BAT SNA response (-13±14%) induced Diabetes Medications by ICV amylin (0.2 nmol) indicating that the sympathetic effects of amylin LIANA K. BILLINGS, A. SOFIA WARNER, VARINDERPAL KAUR, GEOFFREY A. are receptor-mediated. Our data demonstrate that brain action of amylin in- WALFORD, JOSE C. FLOREZ, Boston, MA, Cambridge, MA creases thermogenic sympathetic nerve activity and reveal an unexpected Association studies have revealed that people with low 25-hydroxyvitamin stimulatory effect of amylin on overall sympathetic nerve traffi c including to D (25OHD) levels have elevated insulin resistance and T2D risk. Animal stud- tissues involved in cardiovascular regulation. ies show that 25OHD infl uences beta cell function and insulin sensitivity, but the precise mechanism remains unclear in humans. We aimed to clarify the 1870-P mechanism linking 25OHD and glucose metabolism through pharmacological Oxyntomodulin Gene Delivery by Recombinant AAV Vector Attenu- perturbation by two anti-diabetes medications. Baseline 25OHD levels were ates Body Weight Gain and Improves Insulin Sensitivity in Mice measured in 314 male or non-pregnant female adults at risk of developing YANYUN CHEN, KEYUN QING, Indianapolis, IN future T2D. Fasting subjects were given 5mg of glipizide and glucose and in- Oxyntomodulin(OXM) is a 37aa preproglucagon-derived peptide hormone sulin levels were drawn at regular intervals up to 240 minutes. After a 6-day that is released along with GLP-1 from the L-cells of the small intestine in washout period, subjects began a 2-day course of metformin 500 mg twice proportion to nutrient ingestion. Recent literature reports suggest that OXM daily, followed by fasting glucose and insulin measurements. No association is involved in the regulation of food intake and body weight. Unfortunately, was found between 25OHD and glipizide response (measured by insulin area native OXM is a very weak anorectic agent and has a short half-life of 8 under the curve at 60 and 240 minutes). Higher 25OHD levels were associ- minutes because of its rapid degradation in the circulation by dipeptidyl pep- ated with an improved metformin response (measured by post-metformin tidase 4 and other endoproteinase. These properties limit our ability to study HOMA-IR adjusted for baseline HOMA-IR; P=0.02), although in a model with chronic effects of OXM in animal models.To investigate the effects of chroni- multivariate adjustment for age, sex, body mass index, race/ethnicity, and cally elevated OXM levels in mice, we constructed and packaged a recombi- season this association was no longer seen. In secondary analyses, where nant adeno-associated virus (AAV) 2 vector containing the mouse OXM gene subjects were dichotomized into defi cient (25OHD<20 ng/mL) or suffi cient and a reporter gene encoding a green fl uorescence protein (AAV-OXM-GFP) (25OHD*20 ng/mL) groups, post-metformin HOMA-IR (adjusted for baseline driven by the CMV promoter. Approximately 1x1010 particles of AAV-OXM- HOMA-IR) was greater in 25OHD defi cient subjects (2.3 [SD 2.6]) versus suf- GFP or the control vector AAV-GFP were injected through the tail vein into fi cient subjects (1.7 [SD 1.9], t test P=0.04). Among 209 suffi cient subjects, 2-month old C57BL/6 mice. Throughout the 6-month study period, the AAV- mean post-metformin HOMA-IR was lower than pre-treatment HOMA-IR OXM-GFP-injected mice had signifi cantly decreased food intake (662.7±7.8 (P<0.0001), a difference that was not seen among 77 defi cient subjects vs 692.1±10.0 g) and body weight gain (11.3±1.0 vs 18.6±0.8 g) when com- (P=0.8). These data suggest that 25OHD defi ciency may impair response pared with that of the AAV-GFP-injected mice. Body composition analysis to metformin and may infer that 25OHD infl uences insulin sensitivity in the indicated that decreased weight gain was due to lack accretion of fat mass liver. Further investigation is needed to determine whether raising 25OHD while fat free mass was not affected. Signifi cantly decreased plasma leptin levels would improve metformin response in people with 25OHD defi ciency. concentrations (35.3±2.7 vs 49.0 ±1.2 ng/ml) confi rmed decreased body adi- Supported by: The Endocrine Society’s Lilly Endocrine Scholars Award posity in the AAV-OXM-GFP-injected mice. Six months post the initial AAV injection, the AAV-OXM-GFP group showed signifi cantly improved insulin 1873-P sensitivity in response to an oral glucose challenge. In conclusion, chronic ERK1/2 Signaling is Required for Ghrelin-Induced Cardiomyocyte overexpression of OXM can decrease food intake, body weight, and improve Differentiation from Human Embryonic Stem Cells insulin sensitivity. OXM has the potential to be an anti-diabetic agent with RUI WEI, MEIJUAN GAO, JIN YANG, GUOQIANG LIU, LIN ZHANG, HAINING weight reduction effi cacy. WANG, GUANG WANG, HONGWEI GAO, GUIAN CHEN, TIANPEI HONG, Beijing, China 1871-P Ghrelin, an endogenous ligand for growth hormone secretagogue recep- Regional Incretin Hormone Expression in Human Pluripotent Stem tor (GHS-R), shows cardioprotective activity. The aim of this study was Cell Derived Intestine to investigate the effect of ghrelin on cardiogenesis and its underlying JONATHAN C. HOWELL, RYAN LAUF, JASON R. SPENCE, KYLE MCCRACKEN, mechanism in human embryonic stem cells (hESCs) cultured in an embryoid CHRISTOPHER N. MAYHEW, NOAH SHROYER, MICHAEL HELMRATH, JAMES M. body (EB)-based differentiation protocol. Functional cardiomyocytes were WELLS, Cincinnati, OH, Ann Arbor, MI induced from hESCs and were identifi ed by phase contrast microscope, Incretins, including glucose-dependent insulinotropic peptide (GIP) and pharmacological responses, PCR and immunofl uorescence staining. In order glucagon-like peptide-1 (GLP-1), are secreted by enteroendocrine cells that to clarify the potential mechanism, the levels of GHS-R1_ and the intracel- are largely localized to distinct regions of intestine (K cells in the proximal lular signaling pathways were detected, and the possible roles were tested gut secrete GIP while L cells secrete GLP-1 distally) and augment pancreatic by adding their specifi c antagonist or inhibitors. Furthermore, the effects of insulin release after a carbohydrate meal. The molecular mechanisms that ghrelin on the differentiation of hESCs were verifi ed in rat hearts. Our results govern normal incretin secretion, diminished secretion in type 2 diabetes, showed that functional cardiomyocytes could be differentiated from hESCs and proximal-distal patterning of embryonic intestine and enteroendocrine and ghrelin promoted cardiac differentiation in a dose- and time-dependent -9 Obesity cells during development are not well understood. We used an in vitro mod- manner with the optimum concentration 10 mol/L. Although GHS-R1_

POSTERS el of human intestinal development, in which human intestinal organoids was expressed in differentiated EBs, the effects of exogenous ghrelin were (HIOs) are generated in vitro from pluripotent stem cells (PSCs), to test the unchanged by a specifi c GHS-R1_ antagonist. Moreover, des-acyl ghrelin,

Integrated Physiology/ hypothesis that the bone morphogenetic protein (BMP) signaling pathway, which does not bind to GHS-R1_, displayed similar effects with ghrelin. Im- components of which are expressed throughout embryonic intestine in vivo, portantly, activation of ERK1/2, but not Akt, was induced by ghrelin. The regulates the proximal-distal identity of early intestinal endoderm. We found ghrelin-induced effects of cardiomyocyte differentiation were abolished by that modulation of BMP signaling was suffi cient to direct proximal-distal fate adding specifi c ERK1/2 inhibitor, but not specifi c PI3K inhibitor. Moreover, of HIOs such that stimulation of BMP signaling promoted expression of the ghrelin promoted the differentiation of grafted hESCs into Sox9- and Flk1- distal-enriched genes CA1, HOXA13, and MUC2 compared to controls, while positive mesodermal/cardiac progenitor cells in rat hearts. In conclusion, BMP inhibition induced expression of the proximal-specifi c genes CCK and ghrelin induced cardiomyocyte differentiation from hESCs via the activation LCT. Furthermore, we demonstrated that formation of incretin-expressing of the ERK1/2 signaling pathway. Our study, therefore, provides insight into enteroendocrine cells is dependent on global patterning events, as GLP-1 the mechanism of how ghrelin regulates cardiogenesis and indicates that was only detected in distalized HIOs in vitro. When transplanted into mice using ghrelin may be an effective strategy to promote the differentiation of for 6 weeks, regionalized HIOs were capable of both GIP and GLP-1 expres- hESCs into cardiomyocytes.

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A480 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1874-P Xen delays gastric emptying, inhibits food intake, and increases gut motility. Postsynaptic GABA(B) Receptor Function are Enhanced Within We recently showed that during graded glucose infusions, administration of Nucleus Tractus Solitarii in Diabetic Rats Xen alone had no effect on insulin secretion rates (ISRs) but increased ISRs YINGYING TAN, QI ZHANG, FANRONG YAO, Xianyang, China, Fargo, ND in combination with pharmacologic levels of GIP in humans with NGT and Diabetic autonomic neuropathy, a frequent and serious complication of IGT, but not T2DM. This study examines the effects of Xen on postprandial diabetes, affects autonomic nerve functions. Although its signifi cant nega- plasma glucose levels, ISRs, and gastric emptying in humans.Each fasted tive impact has been seriously considered on survival and quality of life in subject ingested a liquid mixed meal plus acetaminophen (ACM) at time zero people with diabetes, the mechanisms underlying the diabetic autonomic (ACM absorption is a validated technique to assess gastric emptying). On 2 neuropathy are still poorly understood. In this study, we investigated the separate visits, a primed-continuous IV infusion of either vehicle or Xen (12 pmol/kg/min) was started at time zero and continued for 300 min. Plasma glu- function and expression of h-aminobutyric acid-B (GABAB) receptor in the nucleus tractus solitarii (NTS) neurons, a key central integration site for cose, insulin, C-peptide, GIP, and ACM were measured throughout the study. arterial baroreceptor afferents, from streptozotocin-induced diabetic rats ISRs were calculated by deconvolution of C-peptide levels. Postprandial GIP between 4 and 8 weeks after onset of diabetes. Second-order barorecep- levels were similar in all 3 groups. Plasma glucose levels [incremental area tor neurons in NTS were retrogradely labeled by by a transported fl uores- under the curves (iAUCs) from 0-120 min] declined by 35%, 33% and 23% in cent dye, Dil. Using whole cell patch-clamp recording in brain slices, we subjects with NGT, IGT, and T2DM, respectively (p <0.05 for all groups) which was not attributable to higher ISRs. ACM absorption (iAUCs from 30-180 min) found that application of the GABAB receptor agonist baclofen signifi cantly decreased the spontaneous fi ring activity of labeled NTS neurons in both was reduced by 41%, 36%, and 48%, in subjects with NGT, IGT, and T2DM, diabetic and control rats. However, the magnitude of reduction in the fi ring respectively (all p values <0.05), indicating that Xen delays gastric emptying. rate was signifi cantly greater in diabetic rats than in control (87.1±3.2% vs. Thus, Xen amplifi ed ISRs in response to exogenously administered, pharma- 52.6±1.4%, n=12, P<0.05). Furthermore, baclofen produced larger membrane cologic levels of GIP in our prior study but does not augment the effects of en- hyperpolarization and outward currents in labeled NTS neurons in diabetic dogenously released GIP. However, Xen reduces postprandial glucose levels in humans with NGT, IGT, and T2DM by delaying gastric emptying. rats than in control. The EC50 of the baclofen effect was greater in control neurons (9.8±2.7 μmol/L; n=7) than diabetic neurons (2.4±0.6 μmol/L; n=9). Supported by: NIH/NIDDK (RC1DK086163 and R01DK088126) and The Blum Kov- The baclofen-induced current was abolished by either including G protein ler Foundation inhibitor in the pipette solution or bath application of the GABAB receptor an- tagonist in both diabetic and control rats. Blocking N-methyl-D-aspartic acid 1877-P receptors had no signifi cant effect on baclofen elicited outward currents Overexpression of AQP7 Contributes to Improve Insulin Resistance in diabetic rats. These fi ndings suggest that postsynaptic GABAB receptor in Adipocytes function is upregulated in second-order baroreceptor neurons in diabetic XUEMEI GU, WEI PAN, XIONG CHEN, FEI-XIA SHEN, Wenzhou, Zhejiang Province, rats. Changes in postsynaptic GABAB receptors in the NTS may contribute to China the impairment of barorefl ex in diabetes state. Aquaglyceroporin 7 (AQP7) is required for effl ux of glycerol from adipo- Supported by: National Natural Science Foundation of China (No. 81100175) cytes. In this study, we aimed to analyze expression profi les of AQP7 in the different differentiation phase of adipocytes and the role of AQP7 in the in- 1875-P sulin resistance of adipocytes. 3T3-L1 preadipocyte cells were induced fully Regulation of Glucagon Secretion in Pancreatic Alpha-Cells: Com- differentiated. Insulin resistance in differentiated adipocytes was induced putational Systems Analysis by Dexamethasone (DXM). Adenovirus overexpression AQP7 (Ad-AQP7) was LEONID E. FRIDLYAND, LOUIS H. PHILIPSON, Chicago, IL constructed and transfected to adipocytes. The expression levels of AQP7 Glucagon, a 29-amino acid hormone secreted from the alpha-cells of the and phosphorylated PKB (p-PKB) were measured and the glycerol release islets of the endocrine pancreas, is critical for blood glucose homeostasis from adipocytes were also tested. We found that AQP7 expression levels largely by promoting liver production of glucose in hypoglycemia. Alpha-cells were gradually up-regulated along with the differentiation phase of 3T3-L1 in islets are electrically excitable and their mechanism of glucose sensing preadipocytes, which was consistent with the expression levels of p-PKB. involves the coupling of electrophysiological, cytoplasmic and mitochondrial Dexamethasone treatment down-regulated the expression levels of AQP7, processes. We performed the computational systems analysis of regulation p-PKB and the glycerol contents in adipocytes. Overexpression of AQP7 by of glucagon secretion including metabolic processes, electrical activity and transfecting Ad-AQP7 to DXM-induced insulin resistant adipocytes could Ca2+ dynamics. The mathematical model of alpha-cell sensitivity to glucose attenuate the glycerol secretion of adipocytes which was consistent with through the changes of metabolic parameters is based on a previous model the restoration of p-PKB expression levels. Our fi ndings indicated that AQP7 for pancreatic beta-cells. We also formulated a Hodgkin-Huxley-type ionic down-regulated in adipocytes with insulin resistance and overexpression of model for action potentials in alpha-cells that incorporates Ca2+, K+, Na+ and AQP7 contributed to improve insulin resistance in adipocytes which might Cl- currents. The metabolic and ionic models are coupled to the equations de- correlated with improved phosphorylation of PKB. scribing intracellular Ca2+ homeostasis and glucagon secretion that depends Supported by: Zhejiang Provincial Natural Science Foundation of China (081586) on an activation of specifi c voltage-gated Ca2+ channels. This general model simulates the behavior of alpha-cell action potentials and glucagon secre- 1878-P tion under a wide range of experimental conditions, including block of action Accuracy and Sensitivity of Commercially Available Methods for potential fi ring at increased glucose levels or due to channel activation or the Measurement of Human Glucagon Like Peptide 1 (GLP-1) blockade. Paracrine and endocrine regulation of glucagon secretion were MONIKA J. BAK, JENS PEDERSEN, FILIP KNOP, NILS B. JORGENSEN, BOLETTE modeled following their effects on corresponding alpha-cell ion channels. HARTMANN, LARS O. DRAGSTED, JENS J. HOLST, Copenhagen, Denmark, Hel- We analyzed also the reasons of dysfunction of action potential fi ring and lerup, Denmark, Hvidovre, Denmark glucagon secretion in alpha-cells, and attempt to determine the pharmaco- During the last 3 decades numerous articles about GLP-1 secretion have logical targets for improving regulation of glucagon secretion in type 1 and been published. However, it is often diffi cult to interpret the study results 2 diabetes. This computational systems analysis provides an understanding since little information about assay reliability is generally provided, particu-

of the complex processes of glucose, paracrine and endocrine regulation of larly regarding commercial assays. At least six different isoforms of GLP-1 are Obesity glucagon secretion in pancreatic alpha-cells. found in the circulation, namely (1-, 7-, and 9- forms +/- C-terminal amidation). POSTERS Amidation predominates in humans but is variable in experimental animals.

1876-P Only GLP-1 7-36-amide and GLP-1 7-37 isoforms are biologically active but are Integrated Physiology/ Xenin-25 (Xen) Delays Gastric Emptying and Reduces Postprandial degraded extremely rapidly by dipeptidyl peptidase DPP-IV to the metabolites Glucose Levels in Humans With Normal (NGT) or Impaired Glucose 9-36-amide and 9-37. Also, the concentrations of both active and total GLP- Tolerance (IGT) and Type 2 Diabetes Mellitus (T2DM) 1 in the peripheral circulation are low, ranging between 0-15 pM for active SARA CHOWDHURY, DOMINIC N. REEDS, SONGYAN WANG, HUNG D. TRAN, and 5-50 pM for total GLP-1. The aim of the current study was to analyze the JUDIT DUNAI, DAVID ROMETO, ERIN LACINY, BRUCE W. PATTERSON, KENNETH specifi city and sensitivity of 7 commercial assays for GLP-1.Synthetic replicas S. POLONSKY, BURTON M. WICE, St. Louis, MO, Chicago, IL of the 6 different isoforms of GLP-1 were obtained, structurally verifi ed and Xen is a neurotensin-related peptide produced with glucose-dependent in- quantifi ed by Amino Acid Analysis (QAAA). Buffer and plasma was spiked with sulinotropic polypeptide (GIP) by a subset of enteroendocrine cells. We have concentrations of isoforms ranging from 2.5 pM to 300 pM.None of the com- published that Xen plus GIP, but not Xen alone, increases insulin release in mercial assays detect all 6 GLP-1 isoforms in the concentrations applied here. mice via a cholinergic relay in the periphery. Animal studies have shown that ELISA kit from USCN LIFE could not detect GLP-1 in spiked buffer. MSD ELISA

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measures GLP-1 7-36amide but reacts 30-50 % with the other isoforms, sen- 1881-P sitivity for 7-36 is <2,5pmol/l. DRG ELISA exclusively measures active GLP-1 Chronic Mild Stress Alters Circadian Expressions of Molecular isoforms with sensitivity for 7-36 <2,5pmol/l. Millipore and Alpco kits measure Clock Genes in the Liver only amidated GLP-1 isoforms, for Millipore sensitivity is >5pmol/l and for Alp- KEI TAKAHASHI, TETSUYA YAMADA, YOSHITOMO OKA, HIDEKI KATAGIRI, Sen- co <2,5pmol/l. Phoenix ELISA: mostly amidated forms, however, inconstistent dai, Miyagi, Japan results, suggestive of matrix effect.Our results shows that GLP-1 assays from Chronic stress is well known to affect metabolic regulation. However, different suppliers have markedly different specifi cities and sensitivities. molecular mechanisms interconnecting stress response systems and meta- Supported by: NovoNordisk Foundation Center for Basic Metabolic Research bolic regulations are not fully elucidated. Various physiological processes, including glucose/lipid metabolism, are regulated by the circadian clock, 1879-P and dysregulation of core clock genes reportedly leads to metabolic dis- Combination of Dipeptidyl Peptidase-IV Inhibitor, Vildagliptin and orders. Glucocorticoids, which act as end-effectors of the hypothalamus- Angiotensin-II Receptor Blocker, Varsartan Ameliorates Both Insu- pituitary-adrenal (HPA) axis, entrain the circadian rhythms of peripheral lin Insuffi ciency and Insulin Resistance in Mouse Models of Type 2 organs, including the liver, by phase-shifting the expressions of core clock Diabetes genes. Therefore, we examined whether chronic stress affects the circa- KATSUTOSHI MIYAGAWA, TATSUYA KONDO, RINA MATSUYAMA, RIEKO GOTO, dian expressions of core clock genes and metabolism-related genes in the JUNJI KAWASHIMA, HIROYUKI MOTOSHIMA, EIICHI ARAKI, Kumamoto, Japan liver using the chronic mild stress (CMS) procedure. In BALB/c mice, CMS DPP-4 inhibitors improve pancreatic `-cell function by suppression of GLP- elevated and phase-shifted serum corticosterone levels, indicating overac- 1 degradation, and angiotensin-II receptor blockers reduce incidence of type tivation of the HPA axis. The rhythmic expressions of core clock genes, such 2 diabetes (T2DM). We hypothesize that the combination of these two drugs as Clock, Npas2, Bmal1, Per1 and Cry1, were altered in the liver, while they acts in a synergistic manner on islet function and insulin sensitivity.To test were completely preserved in the hypothalamic suprachiasmatic nuculeus this hypothesis, phlorizin (PHZ), Vildagliptin (ViL: 1 mg/kg/day) and/or Val- (SCN), suggesting that the SCN, where the central circadian pacemaker is sartan (VaL: 10 mg/kg/day) were administered to high-fat diet (HFD) induced located, is not involved in alterations in hepatic core clock gene expres- diabetic or db/db mice for 8 weeks. Islet morphology, biochemical markers sions. In addition, circadian patterns of glucose and lipid metabolism-related and insulin signaling were investigated.BW and food intake were both un- genes, such as peroxisome proliferator activated receptor(Ppar)_, Ppara-1, altered by ViL and/or VaL treatment. Fasting and random fed blood glucose Ppara-coactivator-1_ and phosphoenolepyruvate carboxykinase, were also levels were signifi cantly decreased in PHZ, ViL and ViL+VaL treatments in disturbed by CMS. In contrast, in C57BL/6 mice, the same CMS procedure both HFD and db/db mice. Most effective suppression of random fed blood altered neither serum corticosterone levels nor rhythmic expressions of he- glucose was observed in ViL+VaL treated db/db mice. Upon i.p.GTT, ViL+VaL patic core clock genes and metabolism-related genes. Thus, chronic stress treatment group showed most profound suppression of glucose excursion in can interfere with the circadian expressions of both core clock genes and both mice. ViL+VaL treatment displayed signifi cant increase of insulin secre- metabolism-related genes in the liver possibly involving HPA axis overacti- tion in db/db mice upon glucose challenge. In addition, ViL+VaL treatment vation. This mechanism might contribute to metabolic disorders in stressful showed improvement of insulin sensitivity in db/db mice.CRP, TNF-_, IL-6 modern societies. and MCP-1 were all signifi cantly decreased, and adiponectin was highest in ViL+VaL group. ViL+VaL show improved insulin signaling and attenuated ER 1882-P stress in liver. ViL+VaL treatment group increased insulin contents in db/db A Reappraisal of Antagonists for the Human Glucose-Dependent islets. Furthermore, immunohistochemical analysis of the pancreas revealed Insulinotropic Polypeptide (GIP) Receptor that the combination treatment resulted in an increased expression of insu- MIKKEL CHRISTENSEN, LAERKE S. HANSEN, METTE M. ROSENKILDE, JENS J. lin, PDX1, and maintenance of normal `/_-cell distribution.In conclusion, the HOLST, FILIP K. KNOP, Hellerup, Denmark, Copenhagen, Denmark combination therapy of ViL and VaL improves both pancreatic `-cell function The gut hormone GIP is involved in the regulation of glucose and lipid and insulin sensitivity in mice models of T2DM. This combination therapy homeostasis. Elimination of GIP signaling has been proposed as a treat- may be useful in subjects with T2DM and hypertension. ment of obesity and diabetes, and the effects of several peptide-based GIP receptor antagonists have been evaluated in preclinical models.We aimed 1880-P to evaluate the pharmacological properties in vitro of previously described Secretion of Glucagon-Like Peptide-1 in Patients With Type 2 Dia- GIP receptor antagonists GIP[6-30], GIP[7-30], Pro3GIP and GIP[3-42], and betes Mellitus—Systematic Review and Meta-Analysis of Clinical the glucagon-like peptide-1 receptor antagonist exendin[9-39] in COS-7 Studies cells transfected with the human GIP receptor.Antagonistic properties: The SALVATORE CALANNA, LISE L. GLUUD, JENS J. HOLST, TINA VILSBØLL, FILIP K. effect on GIP-induced cAMP accumulation was measured for all ligands. KNOP, Hellerup, Denmark, Catania, Italy, Copenhagen, Denmark The dose-response curves for GIP was right-shifted dose-dependently and Glucagon-like peptide-1 (GLP-1) exerts multiple actions (e.g. insulinotro- the potency of GIP reduced 8.6 folds with 100 nM GIP[7-30], 21 folds with pic and glucagonostatic effects). Some studies suggest that patients with 100 nM Pro3GIP, and most effectively >100 folds with 100 nM GIP[3-42]. type 2 diabetes mellitus (T2DM) are characterized by reduced secretion of Exendin[9-39] had no impact on the activity of GIP. Importantly, only partial GLP-1 possibly contributing to the hyperglycemic state of these patients. inhibition (10-25% reduction in activity) was achieved with GIP[6-30], GIP[7- We aimed to evaluate GLP-1 secretion in T2DM patients and healthy control 30] and Pro3GIP depending on the activating GIP concentration. Receptor subjects during oral glucose and/or meal test.We performed a systematic activation: GIP[6-30], GIP[7-30], GIP[3-42] and exendin[9-39] had no intrin- review with meta-analyses of clinical studies retrieved by electronic (Co- sic agonistic properties on GIP receptor activity. Pro3GIP acted as a partial chrane Library, Medline, Embase and Web of Science) and manual searches agonist with an effi cacy of 40% as compared to native GIP and a potency (until November 2011). Two reviewers independently extracted data from of 18.3 nM. Affi nity determination: All ligands were tested in competition 125 125 15 trials fulfi lling the inclusion criteria. Subgroup, sensitivity and regression with radio-labeled I-GIP. GIP[7-30] displaced I-GIP with an IC50 of 77 sequential analyses were performed.Meta-analysis of GLP-1 responses dur- nM, whereas Pro3GIP and GIP[3-42] had higher affi nities with IC50 values of ing 22 different stimulation tests in a total of 287 T2DM patients (49% men; 5.3 and 1.1 nM, respectively.In conclusion, of the previously described GIP 2 Obesity age (mean[range]): 57[43-63] years; body mass index (BMI): 31[23-45] kg/m ; receptor antagonists only GIP[3-42] exhibits potent antagonistic properties

POSTERS HbA1c: 7.1[5.2-9.2]%) and 230 controls (53% men; age: 54[37-61] years; BMI: towards the human GIP receptor in vitro, whereas GIP[6-30], GIP[7-30] and 29[22-45] kg/m2; HbA1c: 5.3[4.1-5.9]%) found that the two groups exhibited Pro3GIP display lower potencies. Importantly, substantial intrinsic activity

Integrated Physiology/ similar responses of total GLP-1 evaluated from peak plasma concentrations was observed for Pro3GIP; categorizing this peptide as a partial human GIP (weighted mean difference [95% confi dence interval]: 0.80[-3.48;5.07] pM), receptor agonist with 40% effi cacy as compared to native GIP. absolute total area under curve (AUC) (51[-369;470] pM×min), total AUC per min (0.27[-1.77;2.31] pM), incremental AUC (-126[-466;213] pM×min), and in- 1883-P cremental AUC per min (-0.43[-2.44;1.59] pM). Also, the two groups exhibited Xenin-25 Increases Cytosolic Free Calcium Levels and Acetylcho- similar responses when considering mixed meal tests and OGTTs separately. line Release from a Subset of Myenteric Neurons Meta-regression analyses showed no independent effects of age, BMI, SHENG ZHANG, KRZYSZTOF HYRC, SONGYAN WANG, BURTON M. WICE, St. fasting plasma glucose or HbA1c.The present study constitutes the hitherto Louis, MO most extensive systematic review of GLP-1 secretory data from clinical stud- Xenin-25 (Xen) is a 25 amino acid neurotensin-related peptide produced ies and provides evidence that T2DM patients exhibit normal GLP-1 secre- along with glucose-dependent insulinotropic polypeptide (GIP) by a subset tion in response to oral glucose and meal tests. of intestinal K cells. We have published that Xen, with GIP but not alone, in-

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A482 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES directly increases insulin release in mice via a CNS-independent cholinergic in amounts equal to the baseline plasma glucose specifi c activity. Saline relay in the periphery. We also showed a similar relay operates in humans (S), GLP-1-(9,36)amide at 1.2pmol/kg/min (G), or Exendin-(9,39) at 30pmol/ without type 2 diabetes. This relay may be mediated by myenteric neurons kg/min (Ex30) or at 300pmol/kg/min (Ex300) were infused in random order (MENs) since others have shown that a subset of MENs directly connecting on separate study days. Endogenous Glucose Production (EGP) and glucose islets to the stomach/proximal small intestine (where K cells reside) regu- disappearance (Rd) were calculated using Steele non-steady-state equa- lates islet function. This study examines the effects of Xen on MENs.MENs tions. Peak (9.1±0.4 vs. 9.3±0.3 vs. 9.4±0.4 vs. 8.7±0.4 mmol/l, S, G, Ex30 and isolated from guinea pig proximal intestine myenteric plexi were cultured Ex300 respectively, p = 0.24) and integrated glucose (1894±31 vs. 1832±35 for 1 week with cytosine arabinoside to kill proliferating non-neuronal cells. vs. 1839±45 vs. 1870±35 mmol/l per 6hr, p = 0.84) concentrations did not Remaining MENs exhibited typical neuron-like morphology with extensive differ between study days. No effect on insulin and glucagon secretion was outgrowths emanating from cell bodies. MENs were loaded with FURA2- observed. There was no difference in nadir (2.6±0.9 vs. 2.1±0.6 vs. 2.0±0.6 AM. Increased intracellular free Ca2+ levels ([Ca2+]i) were measured by im- vs. 2.0±0.6 μmol/kg/min, p = 0.28) and integrated EGP. Peak (62±4 vs. 60±5 aging to assess activation of MENs. ATP/UTP, forskolin, and KCl increased vs. 61±4 vs. 57±3 μmol/kg/min, p = 0.68) and integrated Rd did not differ [Ca2+]i in 99.6% (1838/1845), 98% (49/50), and 92% (68/74), respectively between study days. We conclude that, in situations where glucose is of the MENs imaged. Thus, nearly all MENs are functional and activated by delivered intravenously Exendin-(9,39) and GLP-1-(9,36)amide do not alter purinergic receptor signaling, cAMP, and direct depolarization, respectively. glucose metabolism, `-cell and _-cell secretion in non-diabetic subjects. In contrast, [Ca2+]i increased in only 12.7% (114/1134) of MENs treated with This suggests that the effects of Exendin-(9,39) on glucose metabolism are Xen whereas GIP alone, or with Xen, had no effect. Following pre-treatment indirect and arise by competitive inhibition of GLP-1 action. with glyburide to inhibit KATP channels, 23 % (133/578) of the MENs re- sponded to Xen. SR48692, EGTA, and diazoxide blocked the effects of Xen 1886-P indicating Xen activation requires neurotensin receptor-1, extracellular Ca2+, 30 Days of Resveratrol Supplementation Does Not Affect Postpran- and KATP channels, respectively. Xen also increased acetylcholine release dial Incretin Hormones Responses, but Suppresses Postprandial from MENs. Thus, Xen activates only a subset of MENs in the proximal intes- Glucagon tine. Since priming of MENs with glyburide, but not GIP, amplifi ed the effects FILIP K. KNOP, ELLEN KONINGS, SILVIE TIMMERS, PATRICK SCHRAUWEN, JENS of Xen, other peptides may augment the effects of Xen on MENs in vivo. J. HOLST, ELLEN E. BLAAK, Hellerup, Denmark, Maastricht, The Netherlands, Supported by: NIH/NIDDK (R01DK088126) and The Blum Kovler Foundation Wageningen, The Netherlands, Copenhagen, Denmark Resveratrol, a natural polyphenolic compound produced by various plants 1884-P (e.g. red grapes) and found in red wine, has glucose-lowering effects in hu- Central and Peripheral Estrogen Differentially Ameliorates Im- mans and rodent models of obesity and/or diabetes. The mechanisms behind paired Glucose Metabolism in Postmenopausal Obese Mice Model these effects remain elusive, but have been suggested to include resvera- RIKA YONEZAWA, TSUTOMU WADA, MAYUKO MORITA, KANAE SAWAKAWA, trol-induced secretion of the gut incretin hormone glucagon-like peptide-1 YOKO ISHII, MASAKIYO SASAHARA, HIROSHI TSUNEKI, TOSHIYASU SASAOKA, (GLP-1). We aimed to investigate the effect of resveratrol on postprandial in- SHIGERU SAITO, Toyama, Japan cretin hormone and glucagon responses in obese human subjects.Postpran- Age-related loss of ovarian function promotes adiposity and insulin re- dial plasma responses of the incretin hormones glucose-dependent insulino- sistance in women. Estrogen (E2) directly enhances insulin sensitivity and tropic polypeptide (GIP) and GLP-1 (total hormone responses) and glucagon suppresses lipogenesis in peripheral tissues. Recently, the central actions of were evaluated in 10 obese men (subjects characteristics (mean±SEM): age: E2 in the regulation of energy homeostasis are becoming clearer; however, 52±2 years; body mass index: 32±1 kg/m2, fasting plasma glucose: 5.5±0.1 the functional relevance and degree of contribution of the central versus mM) who had been given dietary supplement of resveratrol (150 mg/day) or peripheral actions of E2 are currently unknown. Therefore, we prepared placebo for 30 days in a randomized, double-blind, crossover design with a 4 and analyzed estrogen function in four groups of mice. 1) Control: sham- week wash-out period. In the end of each intervention period a standardized operated mice fed a regular diet, 2) OVX-HF: ovariectomized (OVX) mice fed 4h-meal test (2,600 kJ; 9.8 g of protein, 50.7 g of carbohydrate and 42.3 g a 60% high-fat diet (HF), 3) E2-SC: OVX-HF mice subcutaneously treated with of fat) was performed.Resveratrol supplementation reduced fasting plasma E2, and 4) E2-ICV: OVX-HF mice treated with E2 intracerebroventricularly glucose, but had no impact on fasting levels or postprandial responses of (ICV). OVX-HF mice showed increased body weight with both visceral and GIP (11.2±2.1 vs. 11.8±2.2 pM, P=0.87; 17.0±2.2 vs. 14.8±1.6 mM×min, P=0.20) subcutaneous fat volume enlargement, glucose intolerance, and insulin re- or GLP-1 (15.4±1.0 vs. 15.2±0.9 pM, P=0.84; 5.6±0.4 vs. 5.7±0.3 mM×min, sistance. Both E2-SC and E2-ICV equally ameliorated these abnormalities, P=0.73). However, resveratrol supplementation signifi cantly suppressed whereas mRNAs expression profi les in peripheral tissues were different. postprandial glucagon responses (4.4±0.4 vs. 3.9±0.4 mM×min, P=0.01) with- Peripherally administered E2 decreased the expression of TNF_, lipoprotein out affecting fasting glucagon levels (15.2±2.2 vs. 14.5±1.5 pM, P=0.56).Our lipase, and fatty acid synthase in the white adipose tissue (WAT) of OVX-HF, data suggest that 30 days of resveratrol supplementation does not affect although the size of adipocytes and number of CD11c-positive macrophages fasting or postprandial incretin hormone plasma levels in obese humans, but in perigonadal fat in OVX-HF were similarly reduced by both E2 treatments. suppresses postprandial glucagon responses; the latter likely contributing In contrast, centrally administered E2 increased hormone sensitive lipase importantly to the antihyperglycemic effect of resveratrol. in WAT, decreased the hepatic expression of gluconeogenic enzymes, and Supported by: Top Institute Food and Nutrition elevated core body temperature and energy expenditure with marked up- regulation of uncoupling proteins in the brown adipose tissue. These results 1887-P suggest that central and peripheral actions of E2 regulate insulin sensitivity Altered Transient Receptor Potential Vanilloid 1 Activity on Barore- and glucose metabolism via different mechanisms, and their coordinated ef- ceptors in Streptozotocin-Diabetic Rats fects may be important to prevent the development of obesity and insulin QI ZHANG, YINGYING TAN, FANRONG YAO, Xianyang, China, Fargo, ND resistance in postmenopausal women. It has been well documented that diabetes mellitus is associated with autonomic neuropathy including dysfunction of arterial barorefl ex. The 1885-P mechanisms underlying diabetes-related barorefl ex dysfunction remain

The Effect of Exendin-(9,39) and GLP-1-(9,36)amide on Glucose Me- poorly understood. The transient receptor potential vanilloid 1 (TRPV1) is ex- Obesity tabolism and Islet Secretion in Non-Diabetic Subjects pressed on both somatic and visceral sensory neurons. Here, we examined POSTERS MATHENI SATHANANTHAN, PAULA D. GIESLER, JEANETTE LAUGEN, ALAN R. the TRPV1 function in the barorefl ex of streptozotocin-induced diabetic rats.

ZINSMEISTER, ROBERT A. RIZZA, ADRIAN VELLA, Rochester, MN Aortic baroreceptor neurons in nodose ganglion were retrograde-labeled by Integrated Physiology/ The effect of endogenous GLP-1 secretion is often studied by using Exen- a transported fl uorescent dye, Dil. Immunofl uorescence labeling revealed din-(9,39) as a competitive antagonist. However, in light of the recent sug- that the percentage of TRPV1-positive neurons was 50.2 ± 5.0% in control gestion that Exendin-(9,39) has direct effects on glucose metabolism, the rats and 38.2 ± 1.9% in diabetic rats, respectively. This reduction in TRPV1- observed effects of this compound may not be exclusively due to inhibition positive neurons in baroreceptor neurons in diabetic rats was signifi cant (n of GLP-1. Moreover, Exendin-(9,39) has direct effects on gastric accommoda- = 11, p < 0.01). Streptozotocin-induced diabetic rats showed the signifi cantly tion and emptying, altering the rate of systemic appearance of glucose. To blunted barorefl ex, and the decreased maximum gain of barorefl ex function examine whether Exendin-(9,39) and GLP-1-(9,36)amide have direct effects in the high blood pressure range. Using the whole-cell patch clamp technique, on insulin secretion and glucose metabolism 8 non-diabetic subjects were we found that the inward current activated by the application of capsaicin studied on 4 occasions. Glucose was infused to mimic the systemic appear- (1 μM) was signifi cantly smaller in baroreceptor neurons from diabetic rats ance of glucose after a meal. All infused glucose contained [3-3H]-glucose compared with controls. The mean peak density of capsaicin-induced cur-

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A483 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

rents was 145.7 ± 24.7 pA/pF (n = 16) in diabetic neurons and 269.3 ± 31.8 pA/ n=5), indicating that the stimulation ofACTH secretion was under hypotha- pF (n =15) in controls, respectively. The duration of inward current was de- lamic control via AVP.Thus, out data indicate that the brain senses a fall in creased 51% in diabetic rats compared with the control group. These evoked plasma FFA levelto activate the HPA axis, which would increase lipolysis currents were completely blocked by the capsaicin antagonist capsazepine. and restore plasma FFAlevels. These fi ndings suggest theexistence of FFA Furthermore, capsaicin-induced desensitization of TRPV1 was up-regulated, counterregulatory responses and brain control of circulatingFFA levels. whereas TRPV1 re-sensitization was down-regulated in baroreceptor neu- rons of diabetic rats. These fi ndings provide important new information that 1890-P reduced expression and activity of TRPV1 are involved in the attenuation A Specifi c Type of Diabetes Induced by Intermittent Hypoxia in a of baroreceptor neuron excitability, which subsequently contributes to the Mouse Model of Sleep Apnea impairment of barorefl ex in diabetes state. JAN POLAK, LARISSA A. SHIMODA, LUCIANO F. DRAGER, VSEVOLOD Y. PO- Supported by: National Natural Science Foundation of China (No. 31171101) LOTSKY, NARESH M. PUNJABI, Baltimore, MD Obstructive sleep apnea (OSA) is associated with increased risk of dia- 1888-P betes; however, the underlying mechanisms remain unclear. We speculate Basal Hyperglucagonemia Is Linked to Impaired Glucagon Counter- that recurrent upper airway collapse during sleep with ensuing intermittent regulation (GCR) in Type 1 Diabetes hypoxia (IH) has a causal effect. In this study, we delineated the effects of LEON FARHY, ALICE CHAN, MARC BRETON, STACEY ANDERSON, BORIS KO- IH on parameters of glucose homeostasis, liver and `-cell function. Addition- VATCHEV, ANTHONY MCCALL, Charlottesville, VA ally, we investigated outcomes after IH cessation.C57BL6/J mice (10/group) GCR protects against hypoglycemia, but is impaired in type 1 diabetes were exposed to: (1) 14 days of IH; (2) 14 days of intermittent air (IA; control); (T1DM). Model-based analysis of in vivo animal data predicts that the GCR or (3) 7 days of IH followed by 7 days of IA (Recovery group). IH was admin- defects are linked to basal hyperglucagonemia. To test this hypothesis we istered by decreasing FiO2 from 20.9% to 6%, 60 times/hr from 9 AM to 9 studied the relationship between basal glucagon (BasG) and the GCR re- PM. Intraperitoneal glucose and insulin tolerance tests, plasma sampling, sponse to hypoglycemia in 29 hyperinsulinemic clamps in T1DM patients. and hepatocyte isolation were performed after exposures. HOMA-IR and Glucose levels were stabilized in euglycemia and then lowered to 50 mg/ HOMA-` indices were calculated and statistical signifi cance considered as dL. BasG was measured before the descent into hypoglycemia and at 10 p < 0.05 (ANOVA).IH increased fasting glycemia (89±5 versus 149±5 mg/dL) min intervals after glucose reached 70 mg/dL to assess the response to hy- and worsened glucose tolerance (Fig 1A). Lower sensitivity to exogenous in- poglycemia (GCR). GCR was estimated by the percent relative increase in sulin (Fig. 1B) and an 8.2-fold increase in HOMA-IR was observed in IH versus glucagon (RIG) over the basal value: RIG=100*[Maximal response - BasG]/ IA group. Additionally, hepatocyte glucose output was 55% higher in IH than BasG. The observed relationship between BasG and RIG were analyzed with IA mice. Despite diminished insulin sensitivity, HOMA-` and peak insulin our mathematical model of GCR, which unifi es interactions between islet levels after glucose challenge were not up-regulated. In the Recovery group, peptides and glucose, and reproduces the normal GCR axis and its impair- glucose tolerance and insulin sensitivity partially improved, while HOMA-` ment in T1DM. The model was used to identify a GCR control mechanism dropped by 45% and peak insulin levels decreased by 41%.In conclusion, consistent with the observed relationship between BasG and RIG. We found IH induced insulin resistance, glucose intolerance, `-cell dysfunction and in- that BasG levels were 40.5 ± 21.9 pg/mL and RIG was 1.2 ± 1.1 (mean ± creased liver glucose output. `-cell function deteriorated after IH cessation. standard deviation). Higher BasG was associated with lower GCR response and, in particular, RIG correlated negatively with BasG (r = -0.74, p < 0.0001). Consistent with these results was a model of GCR in which the secretion of glucagon has two components. The fi rst is under (auto) feedback control and drives a pulsatile GCR and the second is feedback independent (basal secre- tion) and its increase suppresses the GCR. This model explains the negative relationships between BasG and GCR during a simulated 2-fold increase in BasG as observed experimentally and predicts that further increase in BasG will block the GCR response. The results supports the hypothesis that basal hyperglucagonemia contributes to the GCR impairment in T1DM. They also confi rm the predictive power of our model and the need to test clinically its prediction that glucagon inhibitors may enhance GCR in T1DM. Supported by: RO1 DK082805, RO1 DK51562, UVa GCRC M01 RR 000847

1889-P A Fall in Plasma FFA Level Activates the Hypothalamic-Pituitary- Adrenal (HPA) Axis Independent of Plasma Glucose YOUNG TAEK OH, KI-SOOK OH, INSUG KANG, JANG YOUN, Los Angeles, CA, Seoul, Republic of Korea The brain plays a crucial rolein energy homeostasis. When the bloodglu- cose level falls, the brain senses it and activates various mechanisms tore- store it. It is unknown whether thebrain also responds to a fall in plasma FFA level to activate mechanisms for its restoration. Nicotinic acid (NA) infusion in rats for 1 h caused85% decreases in plasma FFA levels, accompaniedby increases in plasma ACTH and corticosterone (CO) levels (P<0.05; n=8). We tested if theseeffects were mediated by plasma FFA using two independen- tapproaches. First, we tested if the NA effects are reproduced by lowering

Obesity plasma FFAwith other antilipolytic agents: insulin and the adenosine recep-

POSTERS tor agonist SDZ WAG 994. These agents decreased plasma FFA in timecours- es similar to that of NA (with plasma glucose clamped) and increased plasma

Integrated Physiology/ ACTH and CO (P<0.05;n=5).Second, we tested if preventing a fall in plasma FFA levels with Intralipidinfusion abolishes the NA effects.Intralipid blunted the increases in plasma ACTH and CO during NAinfusion (P<0.05; n=5). Thus, the falls in plasma FFA levels were responsible forthe ACTH and CO respons- es. We also tested if the effect of reduced plasma FFA to increase ACTH involved the hypothalamusthat stimulates ACTH secretion via corticotro- Supported by: NIH (90039163, RC1HL099952, R01HL075078, R01HL80105, phin-releasing hormone (CRH) andarginine vasopressin (AVP). Forinhibition ASMF90047490) of these controls, we employed the CRH receptor antagonist antalarmin and the AVP receptor antagonist SSR149415. Antalarmin decreased basal ACTH levels but not the NA-induced increases. In contrast, SSR149415 had no ef- fect on basalACTH levels but blocked the NA-induced increases (P<0.05;

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A484 INTEGRATED PHYSIOLOGY—OTHERCATEGORY HORMONES

1891-P 1893-P Acute Dipeptidyl Peptidase 4 Inhibition has no Impact on the In- Characterization of a GPR41 Antagonist that Enhances cAMP Levels cretin Effect, Glucose Tolerance or Gastric Emptying in Healthy and Stimulates GLP-1 Release Subjects ANGELA K. NEVINS, KEITH OTTO, KELLY L. WILBUR, RICHARD W. ZINK, DAVID NICOLAI A. RHEE, SIGNE H. ØSTOFT, TINA VILSBØLL, FILIP K. KNOP, Hellerup, B. WAINSCOTT, DONALYN SCHEUNER, ISABEL C. GONZALEZ, CHAHRZAD MON- Denmark TROSE-RAFIZADEH, ANNE MILLER-REIFEL-MILLER, Indianapolis, IN Inhibitors of the enzyme dipeptidyl peptidase 4 (DPP-4), which under nor- Pharmacological modulation of G-protein coupled receptors (GPCRs) on en- mal circumstances degrades and thereby inactivates the incretin hormones teroendocrine cells is emerging as a potential therapeutic strategy for treat- glucose-dependent insulinotropic polypeptide and glucagon-like peptide-1, ment of type 2 diabetes through increased release of gut peptides that con- are thought to exert their antidiabetic effect by increasing the incretin ef- trol gastric emptying, appetite, bodyweight, insulin secretion, and beta cell fect. We aimed to describe the impact of DPP-4 inhibition on the incretin viability. The enteric hormone, glucagon-like peptide-1 (GLP-1) is one such gut effect, glucose tolerance and gastrointestinally-mediated glucose disposal peptide or incretin that potentiates the effect of glucose on insulin secretion (GIGD) in healthy subjects in order to elucidate 1) the mode of action of DPP-4 and improves glycaemia. Currently, novel GLP-1 based strategies are aimed at inhibition and 2) the physiological mechanisms underlying the incretin effect. increasing endogenous GLP-1 release from enteric L-cells. In pursuit of a novel Ten healthy subjects (age: 40±5 years (mean±SEM); body mass index: 24±3 approach to increase secretion of glucagon like peptide 1 (GLP-1) from enteric 2 kg/m , fasting plasma glucose: 5.1±0.2 mM; HbA1c: 5.3±0.1%) were subject- L cells, we hypothesized that antagonism of G-protein coupled receptor 41 ed to 50 g-oral glucose tolerance test (OGTT) (with 1.5 g acetaminophen) and (GPR41), a Gi/Go G-protein coupled receptor, could increase cyclic adenosine isoglycemic iv glucose infusion on separate days with and without (random- monophosphate (cAMP) levels and promote GLP-1 secretion. CHO-K1 cells ized) preceding administration of the DPP-4 inhibitor sitagliptin (100 mg the expressing cloned human GPR41 receptor were derived and an assay was night before and 100 mg in the morning of each experimental day). Incretin developed utilizing propionate as an agonist that reduced NKH477-stimulated effect and GIGD was calculated from beta cell secretory responses and the cAMP formation. Compound GPR41 Antagonist 1 inhibited propionate reduc- amounts of glucose utilized, respectively, during the two glucose administra- tion of cAMP formation with an IC50 of 2.5μM and a maximum inhibition tion forms.Isoglycemia was obtained with and without DPP-4 inhibition in all of 50%. Ongoing analysis of GPR41 Antagonist 1 in GTPaS assays will be subjects. No signifi cant impact of DPP-4 inhibition on fasting plasma glucose reported to complete the characterization of antagonist activity. This com- (5.1±0.1 vs. 4.9±0.1 mM, P=0.3), glucose tolerance (evaluated as area under pound elicited GLP-1 release from STC-1 cells with an EC50 of 7.7μM. Stud- the curve: 151±35 vs. 137±26 mM×min, P=0.7) or peak plasma glucose during ies performed in GPR41 knockdown cells indicate that the observed incretin OGTT (8.5±0.4 vs. 8.1±0.3 mM, P=0.3) was observed. Neither incretin effect secretion was specifi c and receptor mediated. These data suggest that an- (40±9 vs. 40±7%, P=1.0) or GIGD changed following DPP-4 inhibition (52±4 tagonism of GPR41 stimulates an increase in cAMP levels leading to GLP-1 vs. 56±5%, P=0.4). Also, gastric emptying during OGTT remained unchanged. secretion and suggest that development of a GPR41 antagonist could yield an These results suggest that acute inhibition of DPP-4 (and, in consequence, effective GLP-1 secretagogue for the treatment of type 2 diabetes. elevation of active incretin hormone levels) does not affect the incretin ef- fect, GIGD, glucose tolerance or gastric emptying in healthy subjects. 1894-P Elevated Mitochondrial Biogenesis in Skeletal Muscle and Brown 1892-P Adipose Tissue is Associated With Androgen-Induced Body Weight Normal Gastric Emptying in Patients With Maturity Onset Diabetes Loss of the Young TARO USUI, KAZUO KAJITA, KEI FUJIOKA, ICHIRO MORI, TAKAYUKI HANA- SIGNE H. ØSTOFT, JONATAN I. BAGGER, TORBEN HANSEN, OLUF PEDERSEN, MOTO, TAKAHIDE IKEDA, HIDEYUKI OKADA, NORIKO TAKAHASHI, KOHICHIRO JENS J. HOLST, FILIP K. KNOP, TINA VILSBØLL, Hellerup, Denmark, Copenhagen, TAGUCHI, YOSHIHIKO KITADA, YOSHIHIRO UNO, HIROYUKI MORITA, TATUO Denmark ISHIZUKA, Gifu, Japan Maturity onset diabetes of the young (MODY) is a clinically and genetical- We have reported previously that dehydroepiandrosterone (DHEA) ad- ly heterogeneous subgroup of non-autoimmune diabetes constituting about ministration reduces adiposity in association with the suppression of PPARa 1-2% of all diabetes. Gastric emptying (GE) is important for the regulation expression level. Recently, we found that treatment with 0.4% DHEA or tes- of postprandial plasma glucose (PG) excursions in healthy subjects. Data on tosterone (T) containing food equally decreased body weight and fat weight GE in MODY patients are absent. In the present study we measured GE (by without affecting food consumption in rats. Treatment with both hormones the acetaminophen method) in patients with MODY3 (hepatocyte nuclear decreased adipocytes size and triglyceride content in skeletal muscle and factor 1_ gene mutations), MODY2 (mutations in the glucokinase gene) and liver. Moreover, incubation with DHEA or T similarly decreased PPARa, lipo- in a group of matched control subjects.Ten MODY3 patients (age: 31±3 years protein lipase and adiponectin mRNA levels in 3T3-L1 adipocytes. However, (mean±SEM); body mass index (BMI): 24±1 kg/m2; fasting plasma glucose SREBP-1 and fatty acid synthase mRNA levels were not infl uenced with these (FPG): 8.4±0.8 mM; HbA1c: 7.0±0.3%), 9 MODY2 patients (age: 43±5 years; hormones. We could not fi nd the evidence of increased lipolysis, such as 2 BMI: 24±2 kg/m ; FPG: 7.3±0.3 mM: HbA1c: 6.7±0.2%) and 10 control sub- raised glycerol release, both in vivo and in vitro. These results suggest that an- 2 jects (age: 40±5 years; BMI: 24±1 kg/m ; FPG: 5.1±0.2 mM; HbA1c: 5.3±0.1%) drogen-induced a decrease in fat mass cannot be explained by the increased were examined on 2 separate occasions: 4h 50 g-oral glucose tolerance test lipolysis and suppressed triglyceride production in adipocyte. Therefore, pos- (OGTT) and standardized liquid meal test (525 kcal: 65 g carbohydrate, 20 g sible mechanism is suggested to be an increase in energy expenditure. We fat, 21 g protein) -both days 1.5 g acetaminophen was added for the evalua- examined the effect of both hormones on heat production with mitochondria, tion of GE.Peak PG levels were higher in all 3 groups during OGTT compared since little is known concerning with the effect of androgen on mitochon- to the meal test. During both the OGTT and meal test MODY 3 and MODY drial biogenesis, especially in skeletal muscle. Treatment with DHEA and T 2 patients had higher PG peak levels than the control subjects (Table). No elevated the protein level of PPARa coactivator 1_ (PGC1_) and mitochon- differences in GE (according to time-to-peak plasma acetaminophen) were drial protein ATP5B levels in brown adipose tissue (BAT) and skeletal muscle, observed between the 3 groups during OGTT or meal test (Table). Prolonged but not white adipose tissue and liver. In addition, treatment with DHEA and GE rates were seen in all groups during meal test compared to OGTT. Con- T increased the expression level of PGC1_ mRNA in C2C12 myotube. Both

clusively our study indicates that the postprandial hyperglycemia in MODY hormones also increased mitochondria staining levels with Rhodamine 123. Obesity

patients is not explained by differences in GE rates. These results indicate that each treatment with DHEA or T increases the ex- POSTERS pression level of PGC1_ and mitochondrial biogenesis in skeletal muscle and Gastric emptying Plasma glucose

BAT, followed by elevated heat production and reduced adiposity. Integrated Physiology/ OGTT Meal t test (P) OGTT Meal t test (P) Time to peak (acetaminophen) Peak values 1895-P (min±SEM) (mM) Correlation between Decrease in Serum Soluble Insulin Receptor MODY3 67±8 115±8 0.004 MODY3 17±2 14±2 0.008 Ectodomain Levels and Improvement of Glycemic Control in Pa- MODY2 83±6 116±15 <0.001 MODY2 13±1 10±1 <0.001 tients With Diabetes Control 82±9 117±11 0.018 Control 9±0 7±0 <0.001 MARI INOUE, AYAKO ISHIBASHI, HIROSHI TAKATA, YOSHIO TERADA, SHIMPEI FUJIMOTO, Nankoku, Japan t test (P)NSNS t test (P) <0.001 <0.001 In patients with diabetes, the serum level of soluble insulin receptor ect- odomain (sIR) is higher compared to that in healthy subjects, and is positively correlated with blood glucose and A1c levels. However, the relation between

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A485 OBESITY—ANIMALCATEGORY

serum sIR levels and clinical factors is not suffi ciently analyzed. In the pres- to VSG. To test this, we performed VSG or sham surgery in DIO FXR knockout ent study, 32 Japanese patients with diabetes [15 men and 17 women, age: mice (KO) and wild-type (WT) littermate controls. We found that the effect 55 ± 14 years, BMI:26.1 ± 5.1(mean ± SD) ] admitted to Kochi Medical School of the surgery on food intake (p< 0.01), body weight (p< 0.05), and glucose Hospital for glycemic control were recruited. Blood samples were obtained tolerance (p< 0.01) depended on genotype. Among WT mice, weight loss at baseline and after 2 week treatment during admission. Serum sIR concen- was initially achieved by reductions in caloric intake. However food intake trations were measured using an enzyme-linked immunosorbent assay kit. increased over time, so that by 6 weeks daily food intake of VSG mice was Serum sIR levels were signifi cantly correlated with fasting plasma glucose not different than sham controls. The critical distinction, relative to dieting (r=0.559, p<0.001), A1c (r=0.607, p<0.001) levels at baseline, but were not alone, is that VSG animals did not overeat at later time points to compensate correlated with age, duration of diabetes, BMI, IRI, homeostasis model as- for the initial caloric defi cit and, as a result, weight-loss was maintained. At sessment for insulin resistance (HOMA-IR), or lipid levels. During admission, 8 weeks post-surgery, WT VSG mice had consumed 16% fewer total calories serum sIR levels decreased signifi cantly (from 2.1 ± 0.8 to 1.5 ± 0.5 ng/ml, (p< 0.001) and weighed 22% less (p< 0.001) than WT shams. Among KO mice, p<0.0001) and reached plateau within a week, showing that serum sIR lev- VSG also elicited initial weight loss and acute reduction in caloric intake. In els change more rapidly than glycoalbumin and A1c levels do. Improvement contrast to WT mice however, food intake recovered quickly and, beginning of mean preprandial glycemic levels during admission was correlated with at 4 weeks KO VSG mice ate 15% more than KO shams. As a result, by 8 decrease of serum sIR levels during admission (r=0.510, p<0.01). Thus, serum weeks KO VSG mice had fully recovered the weight loss, and consumed the sIR levels may be a suitable clinical marker for glycemic control in diabetic same number of total calories as KO shams. With respect to glucose toler- patients. ance, we observed a 35% reduction in the area under the glucose concentra- tion: time curve (AUC, p< 0.001) in WT VSG vs. WT shams. In contrast, there 1896-P was no difference in AUC between KO VSG and KO shams. Together, these The GLP-1 Receptor Agonist Liraglutide Attenuates Atherosclerotic data support that bile acid signaling via FXR contributes to the benefi cial Lesion Development and Potentially Enhances Plaque Stability in effects of this bariatric surgery. an ApoE-/- Mouse Model RICHARD W. SIMPSON, TRACEY GASPARI, IRESHA WELUNGODA, ROBERT E. & 1898-P WIDDOP, LOTTE B. KNUDSEN, ANTHONY E. DEAR, Melbourne, Australia, Bags- Effects of Bariatric Surgery on the Intestinal and Plasma Levels of værd, Denmark Endogenous Cannabinoids The once daily GLP-1 receptor agonist, liraglutide, has been approved as a SILVANA OBICI, ISABELLE MATIAS, SONIA LIPP, DARLEEN SANDOVAL, RANDY new treatment for type 2 diabetes. Liraglutide improves glycaemic control, SEELEY, DANIELA COTA, Cincinnati, OH, Bordeaux, France lowers body weight and is the subject of clinical trials to evaluate effects Activation of the endocannabinoid system (ECS) is implicated in the on cardiovascular disease.We have previously demonstrated liraglutide- pathogenesis of obesity as it promotes energy intake and storage. To de- mediated attenuation of induced plasminogen activator inhibitor type 1 and termine whether weight loss with bariatric surgery decreases the ECS tone, vascular cell adhesion molecule expression in human vascular endothelial we measured circulating and intestinal levels of 2 major endocannabinoids, cells in vitro and signifi cant improvement in endothelial function in vivo in the anandamide (AEA) and 2 arachidonyl Glycerol (2-AG), in two bariatric surgery ApoE-/- mouse model. The current study aimed to determine the in vivo effect rodent models of dietary obesity induced with high-fat diet: Roux-en-Y-Gas- of liraglutide on atherosclerotic plaque formation and stability in the ApoE- tric Bypass (RYGB) and Vertical Sleeve Gastrectomy (VSG). At 3 months from /- mouse model.In vivo experiments utilized established disease (1) or early surgery, RYGB decreased body weight of obese male Long Evan rats as com- onset disease (2) protocols-:(1) 18 week old ApoE-/- mice on a high fat diet pared to sham-operated controls (575±40 and 767 ± 80 g in RYGB vs Sham, for 12 weeks were treated with either saline (vehicle), liraglutide (300μg/ respectively, P<0.01 n=4). The levels of AEA were not decreased by RYGB ei- kg twice daily, s.c) or liraglutide + exendin-9 (150pmol/kg/min s.c., osmotic ther in plasma or throughout the intestine. Similarly, 2-AG was not changed mini-pump) for 4 weeks or (2) 17 week old ApoE-/- mice on a normal chow in plasma and ileum. By contrast, RYGB signifi cantly decreased 2-AG levels diet were commenced on a high fat diet and treatment regime identical to in duodenum, jejunum and liver. VSG in obese mice signifi cantly reduced BW protocol 1 for 4 weeks. Vascular reactivity studies identifi ed improvement (37.9±1.6 and 43.9 ± 1.2 g in VSG vs Sham, respectively, P<0.05 n=6-7) and fat in endothelial function in liraglutide treated mice from protocol 1. Oil red O mass (12.2±1.3 and 18.3± VSG vs Sham, respectively, P<0.05). Unlike RYGB, staining and intima to media ratio (IMR) analysis of atherosclerotic plaques VSG did not change the intestinal levels of AEA or 2-AG in any portion of the from the aorta and brachiocephalic artery (BCA) identifi ed a signifi cant re- intestine. However, VSG resulted in a signifi cant decrease in plasma AEA duction in staining and IMR in liraglutide treated mice from protocol 2. A and 2-AG.These results suggest that a potential mechanism for the weight trend towards increased vascular smooth muscle cell content of BCA ath- reducing effi cacy of RYGB and VSG is the reduction of endocannabinoids erosclerotic lesions in liraglutide-treated mice from protocol 2 was identifi ed levels, with a tissutal response that depends on the type of bariatric surgery. suggesting a plaque stabilising effect associated with liraglutide treatment. The study of the ECS in bariatric surgery may unveil tissue-specifi c role of Together these results suggest a potential therapeutic benefi t of GLP-1 re- endocannabinoids that may be exploited for the treatment of obesity. ceptor activation by liraglutide in the attenuation of early atherogenesis and Supported by: Ethicon stabilisation of existing atherosclerotic disease. & 1899-P Sleeve Gastrectomy is Effective in Mice Lacking the Glucagon-Like Peptide-1 Receptor HILARY E. WILSON-PEREZ, DIEGO PEREZ-TILVE, KAREN K. RYAN, DARLEEN A. OBESITY—ANIMAL SANDOVAL, ADAM P. CHAMBERS, DANIEL DRUCKER, RANDY J. SEELEY, Cincin- nati, OH, Toronto, ON, Canada Glucagon-Like Peptide-1 (GLP-1) is a peptide hormone that is released Guided Audio Tour: Cutting the Gut—The Effect of Bariatric Surgery (Post- from the gut in response to nutrient ingestion, and has a range of metabolic ers 1897-P to 1904-P), see page 17. effects including enhancing insulin secretion and decreasing food intake.

Obesity Postprandial GLP-1 secretion is greatly enhanced following some bariatric

POSTERS & 1897-P procedures, including Vertical Sleeve Gastrectomy (VSG), and has been hy- The Bile Acid Receptor FXR is Necessary for Metabolic Improve- pothesized to be at least partially responsible for the improvements in glu-

Integrated Physiology/ ments Associated With Bariatric Surgery cose homeostasis and other metabolic outcomes following these surgeries. KAREN K. RYAN, CHRISTOFFER CLEMMENSEN, HILARY E. WILSON-PEREZ, APRIL We tested the hypothesis that GLP-1 action is necessary for the effects of M. HALLER, ANDRYI MYRONOVYCH, ROHIT KOHLI, DARLEEN A. SANDOVAL, VSG by performing VSG surgery in mice that lack the GLP-1 receptor (GLP- STEPHEN C. WOODS, RANDY J. SEELEY, Cincinnati, OH, Copenhagen, Denmark 1rKO), and examining their metabolic and behavioral outcomes. We found In addition to facilitating digestion, bile acids function as signaling mol- that VSG-operated GLP-1rKO mice responded similarly to wild-type controls ecules involved in the regulation of metabolism. Both clinical and rodent in terms of body weight and body fat loss, food intake reduction, meal pat- studies fi nd that bile acid metabolism is altered after bariatric surgery. Con- terns and food selection. VSG improved glucose tolerance in both wild-type sistent with this, we fi nd that vertical sleeve gastrectomy (VSG) leads to a and GLP-1rKO mice, overcoming the baseline glucose intolerance of the GLP- 33% increase in plasma bile acids in mice (p< 0.05). Bile acids bind to and ac- 1rKO animals. We conclude that GLP-1 action is not essential for the meta- tivate the nuclear receptor FXR, to regulate the gene expression. We hypoth- bolic improvements induced by VSG surgery. esized that FXR signaling contributes to metabolic improvements pursuant

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A486 OBESITY—ANIMALCATEGORY

& 1900-P & 1902-P Changes in Cholesterol Absorption and Synthesis and Intestinal and The Foregut Bypass Prevents Beta-Cell Failure in Obese Mice Hepatic Gene Expression after Roux-en-Y Gastric Bypass in Obese DENGPING YIN, WENWEI YAN, RUI XU, LIANLI MA, NORA KAYTON, PHILLIP Rats WILLIAMS, ALVIN C. POWERS, DAVID H. WASSERMAN, NAJI N. ABUMRAD, ZHIBO AN, ERIC P. SMITH, RON BITNER, SUSANNA HOFMANN, HELENA Nashville, TN GYLLING, HILARY WILSON-PEREZ, RANDY J. SEELEY, DAVID A. D’ALESSIO, Cin- Pancreatic beta-cell compensation for insulin resistance in obesity re- cinnati, OH, Kuopio, Finland sults in hypersecretion of insulin and increased beta-cell mass. Eventually, Roux-en-Y gastric bypass (RYGB) leads to sustained weight loss and rapid insulin resistance and obesity lead to beta-cell failure and diabetes in some resolution of metabolic disorders. The mechanism whereby RYGB leads to individuals. Gastric bypass surgery performed for the treatment of obesity an improvement in lipid metabolism remains unknown. This study sought resolves obesity-related hyperglycemia in up to 75% of patients. To deter- to determine the effect of RYGB on lipid metabolism and examine potential mine if gastric bypass surgery prevents the progression to diabetes, we per- molecular mechanisms of changes in plasma cholesterol. Two cohorts of formed Roux-en-Y gastric bypass (RYGB) in pre-diabetic (6 wk) and diabetic RYGB or sham animals were studied; postprandial blood and tissue samples (12 wk) BLKS db/db (BKS-db) mice and in pre-diabetic (6 wk) and diabetic were taken at 6 weeks in the fi rst and 6 months in the second. 1 week prior (12 wk) New Zealand Obese (NZO) mice. 100% of pre-diabetic BKS-db mice to the sacrifi ce, measures of total fat absorption and oral fat tolerance were (10/10) and 50% of pre-diabetic NZO mice (4/8) developed diabetes with conducted. Plasma markers of cholesterol absorption (campesterol, sitos- chow feeding by 14 weeks of age. High-fat diet (HFD) induced diabetes in terol, avenasterol, and cholestanol) and cholesterol synthesis (cholestenol, 100% of pre-diabetic NZO mice (n = 8). A signifi cant reduction of insulin+ desmosterol, squalene, and lathosterol) were measured by gas-liquid chro- islet cell area was noted in diabetic (16 wk) BKS-db mice, compared to young matography. Intestinal mRNA expression of genes related to cholesterol (6 wk) pre-diabetic BKS-db mice. In pre-diabetic BKS-db mice (n = 8), RYGB absorption, ATP-binding cassette A1 (AbcA1), apolipoprotein A-IV (ApoA-IV), improved glucose tolerance independent of weight loss or changes in energy apolipoprotein E (ApoE), and scavenger receptor-B1 (ScarB1), and genes in expenditure. In pre-diabetic BKS-db mice, RYGB increased plasma insulin the cholesterol synthesis, Squalene synthase (SS), HMG-CoA reductase and adiponectin, reduced beta-cell apoptosis and partially prevented beta- (Hmgcr) and HMG-CoA synthase (Hmgcs), were determined. Obese rats had cell loss, compared to mice with sham surgery and pair-feeding. RYGB also improved lipid metabolism after RYGB at 6 weeks following the surgery, and prevented diabetes in pre-diabetic NZO mice fed with HFD (n = 6). However, this was sustained at 6 months. Relative to sham, the markers of cholesterol RYGB did not reverse diabetes in diabetic BKS-db (n = 4) and NZO (n = 5) mice. absorption and synthesis were reduced by 21-43% and 31-54% in RYGB at 6 The results demonstrate that RYGB preserves beta-cell mass and prevents weeks, and 22-66% and 17-47% at 6 months, respectively. At 6 weeks, intes- diabetes if performed in the pre-diabetic stage. tinal mRNA expression of AbcA1, ApoA-IV, ApoE, ScarB1 and SS were sig- Supported by: NIH nifi cantly reduced in RYGB, while hmgcr and hmgcs were increased. Hepatic hmgcr and hmgcs were reduced. At 6 months, the expression of AbcA1, SS, & 1903-P hmgcr and hmgcs were reduced in the jejunum and ileum, and hepatic hmgcr Glucagon-Like Peptide-1 Receptor Agonism Predicts Roux-en-Y was increased. Our data demonstrated the robust and sustained reduction Gastric Bypass Metabolic Benefi ts in Rats of cholesterol absorption and synthesis following RYGB and mechanisms DIEGO PEREZ-TILVE, NICKKI OTTAWAY, JENNA HOLLAND, CHRISTINE RAVER, may involve altered gene expression in the liver and small intestine. ERIN BARTLEY, JOSE BERGER, MOUHAMADOUL TOURE, TIMO D. MUELLER, Supported by: NIH (R01 DK57900), EES Research Grant PAUL T. PFLUGER, RANDY J. SEELEY, DAVE D’ALESSIO, MATTHIAS TSCHOP, KIRK M. HABEGGER, Cincinnati, OH, Munich, Germany & 1901-P The sedentary and dietary life style, which has evolved during the last de- Cognitive Improvements Following Bariatric Surgery cades, has been paralleled by an increase in the incidence of metabolic dis- BERNADETTE E. GRAYSON, MAUREEN F. FITZGERALD, ANDREW P. HAKALA- turbances such as dyslipidemia, obesity and type 2 diabetes. The morbidity FINCH, VERONICA M. FERRIS, JENNIFER D. SCHURDAK, JAMES B. CHAMBERS, and mortality associated with these disease states necessitates novel and STEPHEN C. WOODS, RANDY J. SEELEY, TERRY L. DAVIDSON, STEPHEN C. effective treatments. Currently, bariatric interventions are the most effec- BENOIT, Cincinnati, OH tive therapies to stimulate weight loss and treat obesity-related comorbidi- Various components of Metabolic Syndrome are signifi cant risk factors ties. A common feature in many of these procedures is a hypersecretion of for cognitive decline in humans. With the rising prevalence of obesity and Glucagon-like peptide 1 (GLP-1). While the therapeutic implications of GLP-1 diabetes in the US, non-age related cognitive decline is a serious health con- are well known, we hypothesized that response to GLP-1 prior to interven- cern. Sustained body weight loss ameliorates much comorbidity associated tion may be predictive of outcome after Roux-en-Y gastric bypass (RYGB). with Metabolic Syndrome. Furthermore, chronic calorie restriction improves To test this hypothesis, 200 adult, male Long-evans rats were screened to age-related cognitive decline. Currently pharmacologic treatments result establish response to GLP-1 receptor agonism. Populations of responders in small transient reductions in body-weight. Several bariatric surgeries, and non-responders (30 of each) were identifi ed based on body weight, food however, produce sustained body-weight reduction in obese individuals. An intake and glycemic regulation. Ex4 treatment stimulated an 8.5±0.46% important unanswered question is whether surgically-induced weight-loss body weight decrease in responders, and a 2.1±0.25% decrease in non- restores or improves cognitive function to levels observed in caloric restric- responders. Ex4-stimulated body weight loss was associated with altered tion.To assess this question, we maintained adult Long Evans male rats on a food intake (10.6±3.3 vs 15.0±3.3 g/day). Rats responsive to Ex4 stimulated high-fat diet (HFD) (41% fat) for 7 weeks prior to surgery. Animals received weight-loss also displayed lower fasting glucose and greater glucose toler- either sham surgery (SHAM), vertical sleeve gastrectomy (VSG) or roux-en-Y ance. RYGB was conducted in groups of responders and non-responders, gastric bypass (RYGB) and were fed HFD ad libitum. A control group received while a small group with intermediate response served as sham controls. sham surgery and was pair-fed (PF) to the average daily intake of the VSG Following RYGB, responders and non-responders showed similar BW loss, and RYGB groups. Four weeks after surgery, cogntive function was as- but responders displayed enhanced glucose tolerance. While both groups sessed. Reference (RM) and working (WM) memory function were assessed displayed hypersecretion of total plasma GLP-1 in response to a mixed-meal in a radial arm maze task. VSG, RYGB and PF animals exhibited improved challenge, the responder group was characterized by increased active GLP-1 RM and WM in comparison to SHAM controls. RYGB animals exhibited and plasma insulin. Taken together, these data suggest that prior GLP1-R Obesity POSTERS improved acquisition of the fi xed platform in the Morris Water Maze task. sensitivity predicts glucose tolerance after RYGB, which is associated with RYGB animals also had improved performance in comparison to VSG, PF and increased levels of active GLP-1 and insulin levels in rats identifi ed as re- SHAM animals in a 4-arm spontaneous alternation task which assessed spa- sponders. Integrated Physiology/ tial WM.Collectively, these data suggest that RYGB, and to a lesser extent VSG, cause improvements similar to those observed in caloric restriction in memory tasks. Differences in performance in RYGB and VSG animals may be attributable to the altered milieu of circulating hormones which have unique patterns and levels of release following these surgeries. The impact of circu- lating gut hormones on cognition is a topic of ongoing research. Supported by: NIH (1F32HD68103-01A)

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A487 OBESITY—ANIMALCATEGORY

& 1904-P & 1906-P Glucagon-Like Peptide-1 Receptor Agonism, but not Cannabanoid Inducing Young Subcutaneous Adipose Tissue Formation Improves Receptor-Type 1 Antagonism, Improves Adjustable Gastrict Band- Glucose Tolerance and Insulin Sensitivity in Obese Mice ing in Rats QIQI LU, MINGMING LI, YU ZOU, TONG CAO, Singapore, Singapore KIRK M. HABEGGER, HENRIETTE KIRCHNER, CHUN-XIA YI, JENNA HOLLAND, Development of insulin resistance in obesity is tightly related to the ex- NICKKI OTTAWAY, ERIN BARTLEY, PAUL T. PFLUGER, DIEGO PEREZ-TILVE, DAVE pansion of unhealthy adipose tissue (AT), which is characterized by insuf- D’ALESSIO, RANDY SEELEY, MATTHIAS H. TSCHOP, Cincinnati, OH, Munich, Ger- fi cient angiogenesis, hypertrophy of adipocytes and chronic infl ammation. In many this study, we intend to investigate whether inducing young subcutaneous Several bariatric operations are currently used to treat obesity and its re- AT (SAT) formation is able to alleviate the deleterious effects caused by un- lated comorbidities. Among these interventions, adjustable gastric banding healthy AT expansion. To generate young SAT, we injected the adipogenesis (AGB) and Roux en-Y gastric bypass (RYGB) surgery are the most commonly cocktail (mixture of Matrigel and gelatin microspheres control-releasing ba- used. AGB provides a safer, less invasive alternative to RYGB; however, it sic fi broblast growth factor) around the subcutaneous fat depots of C57BL6/J is far less effective and weight regain after surgery is a considerable prob- mice fed high fat diet (60% fat) for 16 weeks. 8 weeks after injection, mice lem. Nonetheless, AGB is signifi cantly more effective than the few medica- from treatment group gained about 70% more SAT (new + original) than the tions now available for weight loss. Glucagon-like peptide (GLP)-1 receptor sham-operated control group. Histology analysis and real-time PCR showed agonists, such as exenatide (Ex4) and liraglutide, are useful to lower blood that the newly formed SAT had the typical AT morphology and similar adi- glucose and also cause reduced food intake and weight loss. As with AGB, pogenesis genes expression level as the original SAT. Notably, in the treat- GLP1-R agonism is also far less effective than RYGB. We hypothesized that ment group, both new and original SAT had signifi cantly smaller adipocyte the gap between RYGB and less invasive treatments could be closed through size and expressed much higher level of pre-adipocyte marker (Pref-1) than combination therapy. We tested this hypothesis by stimulating body weight control group, suggesting that the pool of pre-adipocytes in SAT had been loss via combined AGB and Ex4 therapy in diet-induce obese rats. Adult, largely expanded and more young adipocytes with smaller cell size were male, DIO Long-Evans rats received either an AGB, or a sham surgery. In generated. With more young SAT formation, both systemic glucose toler- sham and AGB rats, 6 days of Ex4 treatment induced a similar decrease in ance and insulin sensitivity were markedly improved, which were indicated BW compared to vehicle controls. Subsequent band infl ation, at volumes by much faster glucose clearance in the glucose tolerance test (P<0.01) and that were sub-threshold for decreased food intake, did not lead to further signifi cantly lower HOMA-IR index (P<0.01). Moreover, serum analysis also body weight loss in vehicle treated rats, but synergistically enhanced Ex4 showed that HMW adiponectin level (but not NEFA level) was signifi cantly treatment. Reversal of treatment order (infl ation before Ex4) stimulated higher in the treatment group. In conclusion, this study demonstrated that similar trends in body weight loss. Interestingly, cannabinoid receptor-type 1 inducing young SAT formation in obese mice is able to markedly improve (CB1) antagonism, which induced similar decreases in food intake and body glucose tolerance and insulin sensitivity, which suggests rejuvenating SAT weight, was ineffective as a synergist to AGB. Immunohistochemical analy- by tissue engineering technique may be developed as an adjuvant method sis of c-fos in the hindbrains of rats after band infl ation uncovered activation for type 2 diabetes therapy. of neurons in the nucleus solitary tract (NTS). Together these data suggest that AGB induces activity in the NTS of the hindbrain and that AGB-stimulat- & 1907-P ed weight loss can be enhanced by the addition of GLP1-R agonism, but not Maternal High Fat Diet During Gestation and Lactation Induced by the antagonism of CB1 receptors. Adipose Tissue Infl ammation, Liver Steatosis and Deterioration of Pancreatic Ƶ Cell Function With a Gender Difference in Mature Off- spring in Mice Guided Audio Tour: Obesity and Infl ammation (Posters 1905-P to 1912-P), HISASHI YOKOMIZO, TOYOSHI INOGUCHI, NORIYUKI SONODA, YASUTAKA MAE- see page 13. DA, EIICHI HIRATA, TOMOAKI INOUE, RYOICHI TAKAYANAGI, Fukuoka, Japan Intrauterine environment may have a signifi cant infl uence on the health of & 1905-P postnatal offspring. It is reported that undernutrition in dams during preg- Effects of Frequent Weight Cycling on Longevity in C57BL/6 and nancy produces mature offspring who are at a high risk of developing obesity 129S6 Mice and diabetes. Here we examined whether maternal high fat diet (HFD) may NORA KLÖTING, MATTHIAS KERN, MICHAEL STUMVOLL, MATTHIAS BLÜHER, have an effect on glucose metabolism and islet function in mature offspring. Leipzig, Germany Female and male C57/BL6J mice were mating and fed either a control diet The majority of obese patients can not maintain hypo caloric diets over a (CD) or HFD from conception to weaning, and then offsprings were fed CD or longer period of time. As a consequence weight cycling is a frequent phe- HFD from 6 to 20 weeks age, resulting in 4 groups: CD-CD, CD-HFD, HFD-CD nomenon in long term treatment of obesity. Long-term studies on weight and HFD-HFD. Body weight in male and female offspring of maternal HFD cycling and intentional weight loss suggest that non-pharmacological and was greater than in offspring of maternal CD at birth and from 4 to 6 weeks, non-surgical treatment of obesity might entail elevated mortality rates. while there was a signifi cant difference only between HFD-HFD and CD- Therefore, we tested the hypothesis that weight cycling reduces life span HFD in female from 6 to 20 weeks. At both 14 and 20 weeks, intraperitoneal in comparison to both permanent hyper caloric and normal nutrition in two glucose tolerance test and insulin tolerance test showed that maternal HFD different mouse strains.In female 129S6/SvEvTac (N=90) and C57BL/6NTac induced glucose intolerance and insulin resistance in male and female off- (N=90) the effects of a continuously caloric restriction, a life-long high fat springs. Liver triacylglycerol content, white adipose tissue (WAT) mass and diet (Sniff, E15772-34) and weight cycling, which was realized with a 4 week mRNA levels of TNF-_, IL-6, MCP-1, F4/80, and CD11 in WAT were increased period change between both extremes of diet, on longevity have been de- in maternal HFD in both male and female. In contrast, insulin secretion, in- termined. Body weight was recorded weekly. Intraperitoneal insulin toler- sulin contents and mRNA levels of PDX-1 in isolated islets were higher in ance (ITT) test was performed at the age of 24 and 51 weeks and the HbA1c maternal HFD in female, while they were lower in maternal HFD in male. was analyzed at the age of one year. Basal metabolic rate was measured To investigate the mechanism underlying a gender difference in pancreatic with metabolic chambers (TSE, Bad Homburg, Germany).The average life ex- ` cell function, oxidative stress in islets was evaluated by immunostaining Obesity pectancy for weight cycling animals (129S6/SvEvTac average age at death: of 8-hydroxy-deoxyguanosine (8-OHdG). Staining intensities of 8-OHdG was POSTERS 21.5±5.0 month and C57BL/6NTac 21.5±3.5 month) was comparable with the increased in maternal HFD only in male. In conclusion, maternal HFD induced controls under chow diet (129S6/SvEvTac average age at death: 23.5±5.0 insulin resistance and deterioration of ` cell function with a gender differ- Integrated Physiology/ month and C57BL/6NTac 21.1±4 month). The permanent high caloric diet ence in mature offspring, accompanied by the progression of adipose tissue group had a 3.5 month shorter life span than the two other groups (129S6/ infl ammation and liver steatosis. SvEvTac average age at death: 20.5±5.0 month and C57BL/6NTac 17.1±4.8 month). 129S6/SvEvTac mice lived 1.5 months longer than C57BL/6NTac mice (P<0.001). In addition, signifi cant differences between the dietary regi- mens were found for measures of insulin sensitivity (ITT), glucose metabo- lism (HbA1c) and energy expenditure.Taken together, frequent weight cycling signifi cantly improved longevity compared to a lifelong high fat diet. There was no difference in the life span of mice undergoing weight cycling and permanent chow diet. Supported by: NGFNplus (Grant No. 01GI0827), BMBF, FKZ: 01EO1001 (N.K.)

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A488 OBESITY—ANIMALCATEGORY

sociate with insulin resistance. However, it remains unclear whether low & 1908-P grade chronic infl ammation coupled with a western style diet can enhance Myeloid Cell-Specifi c Deletion of SIRT1 Impairs Glucose Metabo- the risk for insulin resistance. We sought to investigate whether a high fat lism With Enhanced Infl ammatory Response to Hypoxia diet (HFD) metabolic challenge during sustained, but mild activation of NF-kB AKIKO TAKIKAWA, ISAO USUI, SATOKO SENDA, SHIHO FUJISAKA, SHINPEI is capable of amplifying insulin resistance in mice. Mild activation of NF- HATTORI, YUKIKO KANATANI, KOICHI TSUNEYAMA, AMINUDDIN AMINUDDIN, kB was globally facilitated through a heterozygous knockout of the inhibi- ARSHAD MEHMOOD, TOSHIYASU SASAOKA, HISASHI MORI, KAZUYUKI TOBE, tory subunit, I-kB, (HET) as compared to a wild type control (WT). Both WT Toyama, Japan and HET were placed on either a normal chow (NC) or high fat diet (HFD) Chronic infl ammation is a basic pathophysiology of insulin resistance. for 19 weeks total. Body weights were signifi cantly increased in both WT- Adipose tissue macrophages (ATMs) play important roles in this infl amma- HFD (81%) and HET-HFD (87%) after 19 weeks of HFD compared to week 0. tory process. The role of SIRT1 in the metabolic regulation hasbeen reported However, there was no difference in total body weight (WT-HFD 46±2g vs in several metabolic tissues, but not fully clarifi ed in ATMs. To investigate HET-HFD 49±0.7g; p=0.31), fat (WT-HFD 17.5±1.3g vs HET-HFD 18.7±0.38g; whether SIRT1 in myeloid cells play any metabolic roles, we originally gener- p=0.44), or lean (WT-HFD 25.1±0.9g vs HET-HFD 26.1±0.49g; p=0.35) mass ated myeloid cell-specifi c SIRT1 knockout mice (KO mice). During six week between WT-HFD and HET-HFD groups at the end of 19 weeks. Plasma serum normal chow diet (NC) or high fat diet (HFD) treatment, body weight and amyloid A-3 (SAA-3) was increased in HET-HFD (501±62 ng/ml) compared food intake were not altered between control and KO mice. After six week to WT-HFD (302±11) (p=0.02) suggesting enhanced infl ammatory stimuli in HFD treatment, glucose concentration in intraperitoneal glucose tolerance HET. Glucose tolerance tests (GTT) were performed after 5, 9, and 16 weeks test was signifi cantly elevated in KO mice. The increased expressions of M1 of HFD. Impaired glucose tolerance was fi rst observed following 5 weeks macrophage-related or infl ammatory cytokine-related genes were observed of HFD (AUC: WT-NC 419±77 vs WT-HFD 1302±138; p<0.01) (AUC: HET-NC in epididymal fat tissues, which may be associated with the worsening of 565±48 vs HET-HFD 1484±139; p<0.01), but was not amplifi ed in HET-HFD glucose tolerance in KO mice. To further clarify the mechanisms for the in- compared to WT-HFD at the 3 time points chosen (p>0.05). Plasma insulin fl ammatory phenotype of KO mice, we next examined the role of hypoxia ob- levels were comparable between WT-HFD and HET-HFD at week 5 GTT and served in obese fat tissues. The expressions of hypoxia-related genes were at the time of sacrifi ce (WT-HFD 7.6±0.8 ng/ml vs HET-HFD 9.2±0.5 ng/ml; enhanced in epididymal fat tissues of KO mice compared to those in control p=0.12). Thus, introduction of a HFD to a pre-existing infl ammatory condition mice. In fl ow cytometry analysis and immunohistochemistry, the uptake of does not appear to exacerbate the onset of insulin resistance. pimonidazole, a hypoxic probe, into M1 ATMs was increased in KO mice. Supported by: DK043748 DK078188 DK059637 DK020593 Furthermore, the expressions of some infl ammatory genes in response to in vitro hypoxic stimuli were increased in bone marrow-derived macrophages & 1911-P from KO mice. These results suggest that SIRT1-deleted M1 ATMs are not Insulin Resistance Is Ameliorated by TNF-Related Apoptosis Induc- only hypoxic but also have the enhanced infl ammatory response to hypoxia ing Ligand in High-Fat Diet Induced Obese Mice observed in obese fat tissues. In conclusion, SIRT1 in myeloid cells is in- MI-KYOUNG PARK, SO-YOUNG PARK, CHANG WON LEE, SO YOUNG PARK, SONG volved in the improvement of glucose metabolism at least via suppressing YEE HAN, DUK KYU KIM, HYE-JEONG LEE, Busan, Republic of Korea infl ammatory response to hypoxia in epididymal fat tissue of HFD-fed mice. TNF-related apoptosis inducing ligand (TRAIL) is a member of TNF family of cytokines, which exist either as type II membrane or as a soluble protein. & Although the well-characterized activity of TRAIL is represented by its anti- 1909-P cancer activity, little is known regarding the effects of TRAIL on metabolic Obesity in Pregnancy Programs the Oocyte and Early Embryo pathways. To address this point, we studied in vivo effects of TRAIL in high- ISAAC SASSON, REBECCA A. SIMMONS, Philadelphia, PA fat diet induced obese mice. C57BL/6 Mice were classifi ed into two groups Maternal obesity is associated with abnormal fetal growth and an in- and fed with the normal (ND) and high-fat diet (HFD; protein 14.3 %, carbo- creased risk of metabolic abnormalities in the offspring. However, it is not hydrate 20.1 %, fat 64.4 % (kcal %)) for up to 22 weeks. All mice of the high- known if the critical window of exposure occurs prior to (oocyte) or during in- fat diet group developed obesity, hyperglycemia and hyperlipidemia. Seven utero development. Oocyte quality was signifi cantly poorer in diet induced days after adenoviral-mediated hTRAIL (Ad.hTRAIL) and control GFP (Ad.GFP) obese (DIO) dams compared to controls. Expression arrays in DIO oocytes delivery to each groups, mice were killed and samples were collected and showed differential expression of genes regulating pathways related to analyzed. The expression of ad.hTRAIL was determined by western blotting chromatin remodeling, RNA processing, mitochondrial metabolism, and with liver tissue. Ad.hTRAIL-infected mice did not show any changes of food embryonic growth. Two-cell embryos were generated by mating DIO dams intake and body weight. However, Ad.hTRAIL-infected mice presented sig- (mean=35.9g) to wild-type males (GFP-) and control dams (mean=20.6g) to nifi cant reduction of blood glucose level and plasma triacylglyceride, total males expressing GFP under a ubiquitous promoter (GFP+). Similar numbers cholesterol and free fatty acid. In addition, glucose tolerance test showed of each type of embryo were transferred together into either a DIO or control signifi cant improvement in Ad.hTRAIL-infected mice compared with Ad.GFP- pseudo-pregnant recipient. When delivered from a control recipient, DIO- infected mice. Insulin resistance induced by high-fat diet was examined by derived pups had lower fetal and placental (e12.5), and neonatal weights decreased phosphorylation of Akt. However, the decreased phosphorylation than siblings derived from a control-dam (p<0.01). Pre-implantation exposure of Akt due to high-fat diet was increased in Ad.hTRAIL-infected mice. Taken to a high fat diet had no effect on adult weight in female progeny; however, together, overespression of TRAIL alters the signaling pathways and the me- animals born to a DIO carrier were signifi cantly larger then animals delivered tabolism in high fat fed state. These fi ndings suggest that TRAIL might have to a control carrier regardless of their pre-implantation exposure (p<0.01). a novel function for metabolism and insulin signaling pathways. Progeny of the DIO carrier displayed impaired glucose tolerance (IGT) when compared to progeny of the control carrier (p<0.05). When carried by a DIO recipient, the control-derived progeny displayed a signifi cantly greater IGT & 1912-P then the DIO-derived siblings (p<0.05). Exposure of the oocyte to obesity High Fat Feeding Initiates Insulin Resistance Syndrome Inducing affects the resulting embryo such that progeny display decreased fetal and Neuronal Pathways in the Ventromedial Hypothalamus (VMH) neonatal weight. By contrast, adult weight in female progeny is impacted by SHUQIN LUO, MICHAEL EZROKHI, YELENA TRUBITSYNA, ANTHONY H. CIN- maternal weight during pregnancy rather than pre-implantation embryonic COTTA, Tiverton, RI Obesity exposure. Exposure of the post-implantation embryo to maternal obesity re- Administrationof free fatty acids (FFA) to fuel sensing neurons within the POSTERS sults in IGT. This IGT is exacerbated when there is a mismatch between the hypothalamusresults in neuroendocrine events leading to enhanced hepatic pre-implantation and post-implantation exposures. insulinsensitivity, yet high fat feeding induces insulin resistance (IR). Among Integrated Physiology/ a wide variety of animal models of IR,increased VMH norepinephrine (NE) and serotonin (S) activities are present and increasingthe levels of these & 1910-P neurotransmitters in normal animals by their exogenousadministration at Pre-existing Chronic Infl ammation does not Amplify High Fat Feed- this site produces insulin resistance syndrome (IRS),including loss of the nor- ing Induced Insulin Resistance mal hypothalamicfuel sensing response to FFA. Wetherefore investigated TYLER MORRIS, TAMMY M. LUNDBLAD, WHITNEY BARHAM, FIONA E. YULL, whether high fat diet may induce an increase in VMH NEand S activities, TIMOTHY S. BLACKWELL, OWEN P. MCGUINNESS, Nashville, TN thereby potentiating IRS. Rats were fed either a high fat diet(60% saturated The NF-kB signaling axis has been implicated with the insulin resistance fat) or regular chow diet (RCD) for 6 weeks. High fat fed ratswere divided into of obesity. Other chronic infl ammatory conditions, such as COPD and rheu- diet sensitive obese (DSO) and diet resistant (DR) groups basedon weight matoid arthritis, also include activation of this signaling axis and often as- gain (N = 6-8/group). In vivo VMH microdialysate samples werecollected ev-

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A489 OBESITY—ANIMALCATEGORY

ery 2 hours over 24 hours. Sampleswere analyzed by HPLC for metabolites of 1915-P S: 5-hydroxy-indoleacetic acid(5-HIAA), NE: NE and 3-methoxy-4-hydroxy- GLP-1-Derived Pentapeptide GLP-1(32-36)amide Attenuates the De- phenylglycol (MHPG), and dopamine:homovanillic acid (HVA). Plasma sam- velopment of Obesity, Diabetes, Hepatic Steatosis and Increases pleswere analyzed for glucose and insulin levels. DSO rats were heavier Energy Expenditure in Diet-Induced Obese Mice than either RCD or DRrats (374±5.5 g vs 289±5.7 g or 304±5.9 g, respectively, EVA TOMAS, VIOLETA STANOJEVIC, REBECCA LAUDONE, JOEL F. HABENER, P<0.05). DSO ratsbut not DR rats exhibited an increase in plasma insulin Boston, MA and HOMA-IR relative toRCD rats (by 299% and 325%, respectively; P<0.05). Type 2 diabetes is associated with hyperinsulinemia and insulin resis- DSO rats exhibitedelevated levels of VMH MHPG, NE, and 5-HIAA compared tance leading to elevated hepatic glucose production, hyperglycemia, and to RCD rats (by 38, 63,and 48%, respectively; P<0.001) and DR rats (by 32, hyperlipidemia. Infusions of the C-terminal pentapeptide LVKGRamide, GLP-1 56, and 32%,respectively; P<0.001). However, theDSO rats had decreased (32-36)amide, derived from glucagon-like peptide-1 (GLP-1), in high fat diet- VMH HVA levels relative to RCD rats (by 48%) and DR rats(by 33%) (P<0.001). induced obese mice for sixteen weeks curtailed the rate of weight gain as These fi ndingsindicate that increased VMH NE and S activity, known to in- early as fi ve weeks. At the end of the sixteen week infusion, body weights duce IRS in normalanimals, may be an integral neuronal pathway by which of mice infused with GLP-1(32-36)amide were decreased by 50% compared high fat feeding inducesthe insulin resistance syndrome. to vehicle control that correlated with a 40% decrease in fat mass with no signifi cant difference in lean mass. Indirect calorimetric studies showed that 1913-P although mice infused with GLP-1(32-36)amide exhibited lower cumulative Defi ciency of PTP1B in Leptin Receptor-Expressing Cells Leads to food consumption, the rate of oxygen consumption was signifi cantly higher Decreased Body Weight and Adiposity compared to vehicle control throughout the light and dark cycles, fi ndings RYAN C. TSOU, DEREK E. ZIMMER, BART C. DE JONGHE, KENDRA K. BENCE, consistent with an increase in energy expenditure. These metabolic effects Philadelphia, PA were not associated with changes in physical activity. Moreover, the infu- Protein tyrosine phosphatase 1B (PTP1B) is a ubiquitously expressed ty- sion of GLP-1(32-36)amide for sixteen weeks in high fat-fed mice attenuated rosine phosphatase, implicated in the negative regulation of leptin and in- the development of diabetes since both plasma glucose and insulin were sulin signaling. Whole body, brain- and POMC neuron-specifi c PTP1B-/- mice, decreased close to values obtained in mice fed a control. Livers obtained possess a lean metabolic phenotype attributed at least partially to improved from peptide-treated mice showed less steatosis that correlated with a hypothalamic leptin sensitivity. Interestingly, mice lacking both leptin and 65% decrease in triglyceride accumulation, equivalent to triglyceride levels PTP1B (ob/ob:PTP1B-/-) show an intermediate body weight phenotype com- in control mice fed a low fat diet. These fi ndings suggest biological actions pared to mice lacking leptin only (ob/ob), suggesting that PTP1B may have and a potential role for the C-terminal pentapeptide, GLP-1(32-36)amide, in important leptin-independent metabolic effects. In this study, we generated the improvement of obesity-related diabetes, insulin resistance and hepatic leptin receptor (Lepr) expressing cell-specifi c PTP1B-/- mice and compared steatosis. The pentapeptide might prove to be useful for the treatment of them with whole body PTP1B-/- mice and Lepr-Cre only wild type controls insulin resistance and metabolic syndrome. (WT) in an array of metabolic phenotyping measures. Lepr-specifi c PTP1B-/- mice (Lepr-PTP1B-/-) have signifi cantly reduced body weight when maintained 1916-P on high-fat diet (HFD) compared to Lepr-Cre only WT controls. Interestingly, The Peripheral Amino Acid Profi le Alterations in Obese Zucker Rats the reduction in body weight of Lepr-PTP1B-/- mice on HFD was comparable Affect Amino Acid Levels in Cerebrospinal Fluid to that of whole body PTP1B-/- mice only after 8 weeks of age, suggesting KENJI NAGAO, HIROKO JINZU, SACHISE ETO, MAKOTO BANNAI, YASUSHI NO- that PTP1B activity outside of leptin receptor-expressing cells may play a role GUCHI, TSUYOSHI KOBAYASHI, Kawasaki, Japan in early weight gain. Adiposity was decreased in Lepr-PTP1B-/- and whole In obese subjects, it has been reported that the blood amino acid levels body PTP1B-/- mice relative to controls on HFD. Lepr-PTP1B-/- mice showed (aminograms) are altered compared with age- and sex-matched controls. enhanced glucose tolerance compared to WT controls, but as expected, were Although such peripheral alterations could affect the set point of central not as glucose tolerant as whole-body PTP1B-/- mice on HFD. Similar to the amino acid concentrations, little is known about precise changes in cere- whole body PTP1B-/- mice, Lepr-PTP1B-/- mice also displayed greater leptin brospinal fl uid (CSF) amino acid levels due to its technical diffi culties. Since sensitivity compared to controls. Overall these results in large part demon- central amino acids play signifi cant roles both as signaling molecules and strate that PTP1B defi ciency in Lepr-expressing cells underlies the metabolic as precursors for several neurotransmitters, we hypothesized that the al- phenotypes seen in whole body and brain-specifi c PTP1B-defi cient models, terations in CSF aminograms were related with obesity-related psychotic however subtle phenotypic differences between Lepr-PTP1B-/- and whole disorders. We used ten week old, male obese and lean Zucker rats as animal body PTP1B-/- mice suggest that PTP1B independent of leptin signaling may models and the CSF and blood were obtained. A sensitive analysis of amino also contribute to energy homeostasis regulation. acids using high-performance liquid chromatography/electrospray ionization Supported by: NIH (R01 DK082417, F31 NS074684) tandem mass spectrometry (LC/ESI-MS/MS) method was employed to ana- lyze each amino acid level, and biochemical parameters were examined. The 1914-P results showed that 1) The plasma aminogram trends in obese Zucker rats Diverse Effects of Hypothalamic PTEN and Akt on Food Intake and were similar to obese human; the levels of valine, leucine and isoleucine Insulin Sensitivity in Rats were highly elevated, while glycine level was decreased compared with TAKASHI SUMITA, HIRAKU ONO, KOUICHI INUKAI, HIDEKI KATAGIRI, TO- lean counterparts. 2) The CSF levels of valine, leucine and isoleucine were MOICHIRO ASANO, SHIGEHIRO KATAYAMA, TAKUYA AWATA, Saitama, Japan, elevated as well, while glycine level was decreased in obese Zucker rats. 3) Miyagi, Japan, Hiroshima, Japan The CSF levels of glutamate, tryptophan, histidine and tyrosine, which play Phosphatidylinositol 3,4,5-triphosphate (PIP3) and Akt are important signal role as neurotransmitters or precursors for neurotransmitters, were not sig- transducers in canonical target tissues of insulin, but their functions in the hy- nifi cantly different between these two groups. These results indicate that pothalamus are not yet well established. We investigated how modifi cations some amino acids, such as glycine, in CSF are affected by peripheral amino- of these signaling molecules infl uence energy balance and glucose metabo- gram changes in obese, diabetic model rats, while other neurotransmitter- lism, using adenoviral interventions in the mediobasal hypothalamus (MBH). related amino acid levels were strictly maintained in CSF. Persons with

Obesity Sprague-Dawley rats were injected in the MBH with adenoviral vectors ex- schizophrenia and other psychotic disorders including sleep disorder have a

POSTERS pressing various PTEN/Akt mutants, then weight gain, food intake and insulin high prevalence of obesity. Decrease in CSF glycine level might be one of the sensitivity were measured. We found that expression of dominant-negative contributors to these symptoms.

Integrated Physiology/ PTEN in the MBH reduced food intake and weight gain in chow-fed rats, while constitutively-active PTEN tended to induce the opposite effects. Interest- 1917-P ingly, these effects of PTEN mutants in the MBH on food intake were blunted Leptin is Required for the Effect of High Fat Diet to Increase Wheel when rats were fed a high-fat diet. However, rats with dominant-negative Running in Mice PTEN in the MBH showed higher insulin sensitivity even when fed a high- TRACY S. TYLEE, KARL KAIYALA, MICHAEL W. SCHWARTZ, BRENT E. WISSE, fat diet. Moreover, expression of constitutively-active Akt in the MBH also Seattle, WA induced higher insulin sensitivity. From these results, we conclude that PIP3 Increases in energy expenditure due to increased spontaneous physical in the MBH suppresses food intake and weight gain, while these effects are activity (SPA) or autonomic mechanisms via leptin signaling confer protec- blunted by over-nutrition. PIP3 and Akt in the MBH improve insulin sensitivity, tion against obesity and diabetes, but whether SPA is itself regulated by lep- independently of their effects on food intake and weight. tin is unknown. In ob/ob mice, leptin defi ciency is associated with reduced SPA suggesting a role for leptin in this behavior. To test this hypothesis,

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A490 OBESITY—ANIMALCATEGORY we investigated whether wheel running (WR), a form of SPA, is altered by box O1 (FoxO1). Deacetylation of PGC1_ by SIRT1 correlated with upregula- changes of plasma leptin levels in mice.We fi rst investigated whether physi- tion of genes controlling mitochondrial biogenesis and improvement of mi- ological leptin replacement increases WR by infusing ob/ob mice with either tochondrial function in the liver and brown adipose tissue. This promoted leptin (100 ng/hr; n = 8) or vehicle (n = 7) via subcutaneous mini-pumps over mitochondrial fatty acid oxidation, reduced lipid accumulation in the liver 3 days. As predicted, leptin replacement increased WR distance compared and prevented HFD-induced obesity in mice. Moreover, SIRT1-deacetylated to baseline (from 322±149 m to 888±346 m, p = 0.04) by increasing total WR FoxO1 was accompanied by decreased expression of pseudokinase tribble3 time (67±16 min to 138±29 min, p = 0.005) but not WR speed (2.3±0.7 m/sec (TRB3) and reduced the association between TRB3 and Akt, which enhanced to 2.6±0.7 m/sec; p = 0.56). We next investigated whether consuming a insulin sensitivity and improved glucose metabolism. These data suggest high fat diet (HFD), which acutely increases leptin levels, increases SPA in that leucine supplementation-triggered SIRT1-dependent deacetylation of C57BL/6 wild type (WT) mice. Relative to chow baseline, 3 d of exposure to PGC1_ and FoxO1 may contribute to attenuation of HFD-induced insulin re- HFD increased plasma leptin levels from 0.6 ng/ml to 1.2 ng/ml, and resulted sistance, metabolic abnormalities and obesity in mice. in increased WR distance (8150±920 m to 11624±821 m; p = 0.001) and run Supported by: NIH, AHA, OCAST time (524±37 min to 625±24 min; p = 0.01), but no change in speed. To deter- mine if leptin is required for the effect of HFD to increase WR, ob/ob mice 1920-P (n = 8) were similarly exposed to 3 d HFD. Contrary to what was observed in A Small Dose of Metformin Suppresses the Tumor Death Ratio WT mice, HFD exposure led to a decrease in total WR distance (469±175 m Accelerated by a High-Calorie Diet in Ras H2 Transgenic Mice: A to 169±80 m, p = 0.03) in ob/ob mice.We report that 1) reduced SPA in ob/ob Model for Evaluating and Designing Tumor Prevention or Regres- mice is mediated in part by leptin defi ciency; 2) physiological leptin replace- sion Therapies ment in ob/ob mice increases run time but not WR speed; 3) HFD in WT SHUICHI OTABE, NOBUHIKO WADA, XIAOHONG YUAN, TOMOKA FUKUTANI, mice also increases WR time without affecting speed, and 4) leptin may be KAYO TANAKA, TSUYOSHI OHKI, SATOMI KAKINO, YAYOI KURITA, TOSHIHIKO required for this response to HFD, since it is not observed in leptin-defi cient HASHINAGA, HITOMI NAKAYAMA, YUJI TAJIRI, KENTARO YAMADA, Kurume, mice. Together, these data implicate leptin as a key regulator of SPA, an ef- Japan fect that may confer protection against excess weight gain and diabetes. Our objective was to investigate whether moderate metformin intake Supported by: Endocrine Fellows Foundation suppresses the rate of tumor death under a high-calorie diet that induces overweight and glucose intolerance.Male rasH2 transgenic mice at the age 1918-P of 8 weeks, a model for evaluating and designing tumor-prevention or re- ABCG1 and Adipose Tissue Lipid Mobilization during Caloric Re- gression therapies, were divided into three groups of equal number: one was striction fed regular chow of 352 kcal/100 g; another was fed a high-calorie diet of HAO WEI, TIMOTHY S. MCMILLEN, ELIZABETH J. TARLING, PETER A. EDWARDS, 592 kcal/100 g ad libitum; the third was fed the same high-calorie diet with RENEE C. LEBOEUF, Seattle, WA, Los Angeles, CA metformin added to the drinking water from initiation of the experimental Weight loss is a main remedy to resolve the increasing problems of obe- diets to death. Metformin was administered at a dose of 10 mg/kg body sity. Our goal is to understand how cholesterol is mobilized from adipose weight per day. Bodyweights of all the mice were measured every week. tissue (AT) during weight loss, given that AT contains a large reserve of Glucose tolerance and insulin sensitivity were evaluated at the ages of 30 unesterifi ed cholesterol. We applied caloric restriction (CR) to male C57BLKs and 40 weeks, respectively.The rasH2 transgenic mice fed a high calorie diet leptin receptor defi cient mice (db/db), with ad libitum (AL) fed db/db and with/without metformin were signifi cantly heavier than those fed regular AL fed wild-type (WT) lean mice as controls to study AT cholesterol. CR (4 chow after the age of 10 weeks (p<0.0001). The mice fed a high-calorie diet weeks) resulted in substantial body weight loss, decreased total cholesterol without metformin had a clearly shorter average life span (n=15; 58.1±22.3 (TC) and triacylglycerol (TG) levels (61% and 72%, respectively) in epididymal weeks) than those fed regular chow (n=16; 76.5±17.6 weeks); p<0.05, the white adipose tissue (EWAT), and marked increases in AT leukocyte (ATL) ac- log-rank test. At autopsy, multiple occurrences of cancers in such organs cumulation. Protein and mRNA levels for lipid metabolism genes were quan- as the lung, liver, and digestive organs were found in both groups. When tifi ed in EWAT using Western blotting and qRT-PCR. The major alteration treated with metformin, the rasH2 transgenic mice fed a high-calorie diet was for ABCG1: mRNA and protein levels increased 5-fold in AL-db/db versus (n=16) showed an amelioration effect of impaired glucose tolerance. All the AL-WT mice, and were further elevated by 3-4-fold in CR-db/db versus AL- mice with metformin survived to the point of 48 weeks. This demonstrates db/db mice, suggesting a role for ABCG1 in AT lipid metabolism. ABCG1 in that metformin is associated with signifi cantly reduced mortality in mice fed AT was primarily expressed in F4/80+ leukocytes and was undetectable in a high-calorie diet (p=0.02).With a high-calorie diet, a constitutional predis- adipocytes. As expected, ABCG1 levels increased (>3 fold) in cholesterol- position to tumors may be enhanced, and the high incidence of tumor death loaded peritoneal macrophages (PMACs). We are currently using ABCG1 may be suppressed, by a small dose of orally administered metformin. transgenic mice to test the hypothesis that ABCG1 is essential for effi cient lipid/sterol mobilization in EWAT during CR. Bone marrow transplantation 1921-P studies are also underway to compare obesity and weight loss between In Vivo Flux Rates of Leucine Metabolism in Obesity WT mice receiving BM from Abcg1-/- versus WT mice. We hypothesize that PENGXIANG SHE, KRISTINE OLSON, CHARLES H. LANG, CHRISTOPHER J. ABCG1 defi ciency will lead to excessive accumulation of cholesterol in ATLs LYNCH, Hershey, PA during weight loss, consequently altering AT infl ammation patterns and ul- Plasma branched-chain amino acids (BCAA, including leucine, isoleucine timately, fat mobilization. and valine) are elevated in animal models and patients of obesity. Elevations Supported by: NIH (RO1 HL055362) in circulating BCAA and related metabolites are now considered as a “meta- bolic signature” for obesity, insulin resistance and glucose intolerance, and 1919-P highly prognostic of late-onset of type 2 diabetes. However, the mechanisms Activation of SIRT1 Is Associated With Leucine Supplementation- underlying elevated BCAA and the role of BCAA in the pathogenesis of obe- Improved Mitochondrial Biogenesis, Insulin Sensitivity, and Energy sity and diabetes remain to be determined. We and others have reported Metabolism in High Fat Diet-Induced Obese Mice a decreased enzymatic capacity of BCAA catabolism in liver and adipose

HONGLIANG LI, CHAOYONG HE, ZHONGLIN XIE, Oklahoma City, OK tissue but not in muscle of rodent models of obesity and diabetes. A primed- Obesity

14 14 POSTERS Leucine supplementation has been shown to prevent high fat diet (HFD)- continuous infusion of NaH CO3 and [1- C]leucine was used to measure fl ux induced obesity, hyperglycemia, and dyslipidemia in animal models, but the rates of leucine metabolism in postprandial Zucker obese rats. Flux rates

underlying mechanisms are not fully understood. Recent studies suggest that were calculated using the specifi c activity of plasma alpha-ketoisocaproate Integrated Physiology/ activation of Sirtuin1 (SIRT1) is an important mechanism to improve energy (KIC). Obese rats at 11-weeks of age had increased body weight (wt), fat and metabolism. The aim of the study was to determine whether leucine supple- lean mass but slightly decreased gastrocnemius wt, compared to lean rats mentation prevents HFD-induced obesity, insulin resistance, and metabolic (n=10). Their plasma concentrations of leucine and KIC were increased 46% dysregulation through regulation of SIRT1. To accomplish this goal, male and 102%, respectively. While whole-body leucine turnover and proteolysis C57BL/6J mice were fed normal diet or HFD, supplemented with or with- per kg of body wt were unaltered, these parameters based on lean mass out leucine. After 2 months of treatment, alterations of SIRT1 activation, were 35% higher in obese than lean rats. In obese rats, rates of whole-body insulin signaling, and energy metabolism were analyzed in the liver, brown leucine oxidation were 22% and 59% higher when corrected for body wt and adipose tissue, and skeletal muscles. Leucine supplementation increased lean mass, respectively. The rate of leucine incorporation into protein was SIRT1 expression and NAD+ production along with decreased acetylation 65% higher in gastrocnemius but tended to be lower (21%, P < 0.07) in liver of peroxisome proliferator-activator-a coactivator-1_ (PGC1_) and forkhead of obese rats, compared to lean rats. The discrepancy between elevated

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A491 OBESITY—ANIMALCATEGORY

whole-body leucine oxidation and declines in BCAA catabolic enzymes in fat 1924-P and liver of obese animals could imply elevated BCAA oxidation in skeletal Exchange Protein Directly Activated by cAMP 1 Is Involved in Ƶ3- muscle. Increased protein synthesis but decreased muscle mass also implied Adrenergic Induction of Brown Adipose Tissue in White Adipose an elevated turnover of muscle protein. Our data suggest a tissue-specifi c Tissue alteration in protein and leucine metabolism in obesity. YINGXIAN CHEN, ANDREW C. TAI, ALAN KAI, AIMIN XU, KAREN S. LAM, SIDNEY TAM, STEPHEN S. CHUNG, SOOKJA K. CHUNG, Hong Kong, China, Zhuhai, China 1922-P Exchange protein directly activated by cAMP (Epac1, Epac2a and Epac2b) Mitochondrial DNA Damage, Oxidative Stress and Mitochondrial were identifi ed as cAMP-regulated guanine nucleotide exchange factors. Dysfunction are Associated With Endoplasmic Stress and Au- Previously Epac1-defi cient mice were shown to have slightly heavier body tophagy in Skeletal Muscle and Liver in High Fat Diet Induced-In- weight, higher respiratory exchange ratio, and develop more severe high- sulin Resistance Mice fat-diet-induced obesity and hyperglycemia than the wild type mice, sug- LARYSA YUZEFOVYCH, SUSAN LEDOUX, GLENN WILSON, LYUDMILA RACHEK, gesting the role of Epac1 in energy expenditure and lipid metabolism. The Mobile, AL `3-adrenergic signaling in white adipocytes is reported to be important in Consumption of high fat diet (HFD) leads to obesity and thus to insulin lipid metabolism and energy homeostasis by induction of brown adipose tis- resistance which represents a major risk factor for metabolic syndrome and sue (BAT) in white adipose tissue (WAT). To investigate the role of Epac1 in type 2 diabetes. Recent studies showed a link between lipid accumulation `3-adrenergic induction of BAT in WAT, a `3-adrenergic receptor agonist in non-adipose tissues, such as skeletal muscle and liver, and insulin resis- (CL 316,243, CL) or saline was administrated to wild type (Epac1+/+) and ho- tance. Although the mechanisms responsible for insulin resistance in those mozygous Epac1 knockout (Epac1-/-) mice. After CL (1mg/1kg body weight/ tissues are different, oxidative stress and mitochondrial dysfunction have day, i.p.) treatment for 10 days, peri-uterine WAT exhibited a BAT-like phe- been implicated in the disease process, however, the underlying mecha- notype with smaller eosinophilic adipocytes with multilocular lipid droplets nisms are still unknown. Among the potential targets is mitochondrial DNA in both Epac1+/+ and Epac1-/- mice. Interestingly, WAT of CL-treated Epac1- (mtDNA), which is highly specialized and encodes for proteins essential for /- mice showed less typical BAT morphology compared to that of CL-treated energy metabolism. The aim of this study was to determine whether a HFD Epac1+/+ mice. In addition, CL-induced up-regulation of uncoupled protein 1 causes mitochondrial DNA (mtDNA) damage and whether this damage is (UCP1) seen in WAT of Epac1+/+ mice was not observed in Epac1-/- by im- associated with mitochondrial dysfunction, oxidative stress, and induction munocytochemical, Western blot and real-time qPCR analyses. Inductions of markers of endoplasmic reticulum (ER) stress and autophagy in skeletal of other genes shown to be critical for thermogenesis (Cidea, PGC1_) and muscle and liver in a mouse model of obesity-induced insulin resistance. adipocyte differentiation (CEBP _ and PPAR a) with CL treatment were also Six week old C57BL/6J male mice were fed either a HFD (60% fat) or nor- not observed in WAT of CL-treated Epac1-/- mice by real-time qPCR analysis. mal chow (NC) (10% fat) for 16 weeks. HFD fed mice gain signifi cantly more Concomitantly, Epac1 expression was increased in WAT of Epac1+/+ mice weight, had higher glucose, insulin, free fatty acids and triglycerides levels; after CL treatment, whereas no Epac1 and no compensation of Epac2 ex- impaired glucose tolerance and were insulin resistant. HFD-induced insulin pressions were observed in Epac1-/- WAT. Taken together, Epac1 plays an resistance correlated with increased mtDNA damage, oxidative stress and important role in `3-adrenergic induction of BAT in WAT. mitochondrial dysfunction in skeletal muscle and liver. Furthermore,a HFD Supported by: Hong Kong Research Council GRF Grant leads to activation of ER stress and autophagy in skeletal muscle and liver. In addition, HFD reduced protein content of insulin signaling, and increased 1925-P apoptosis in skeletal muscle. This study provides new evidence that HFD- Impact of Adipose Tissue Hypoxia on Macrophage Polarization in induced mtDNA damage correlates with mitochondrial dysfunction and in- Diet-Induced Obese Mice creased oxidative stress in skeletal muscle and liver, which is associated SHIHO FUJISAKA, ISAO USUI, MASASHI IKUTANI, AMINUDDIN AMINUDDIN, with induction of markers of ER stress and autophagy. SATOKO SENDA, AKIKO TAKIKAWA, MANABU ISHIKI, YUKIKO KOSHIMIZU, Supported by: NIDDK RO1DK 073808 (L.R.) KOICHI TSUNEYAMA, YOSHINORI NAGAI, KIYOSHI TAKATSU, KAZUYUKI TOBE, Toyama, Japan 1923-P Obesity is recognized as a condition of chronic low grade infl ammation Intrahepatic Lipid, Not Total Abdominal Fat or Muscle Fat, is a in adipose tissues associated with an increased number of infi ltrating mac- Marker for Insulin Resistance in the Elderly Female Monkey rophages. Adipose tissue macrophages (ATMs) are composed of at least two MICHAEL P. CHU, BETHANY J. KLOPFENSTEIN, HENRYK F. URBANSKI, WILLIAM phenotypes i.e., classically activated M1 macrophages and alternatively acti- D. ROONEY, STEVEN G. KOHAMA, CHRISTINE KRISKY, JONATHAN Q. PURNELL, vated M2 macrophages. We have reported that M1 and M2 ATMs are clearly Portland, OR, Beaverton, OR separated by fl ow cytometry and they have quite different gene expression Aging is associated with an increased risk of visceral adiposity and diabe- pattern (Diabetes, 58: 2574-2582, 2009).Recently, we found that the expres- tes in women. Visceral adiposity is also linked with an increased accumula- sions of hypoxia-related genes in M1ATMs were higher than those in M2 tion of ectopic fat, impaired glucose tolerance, and risk for diabetes. How- ATMs in epididymal adipose tissue of obese mice. Adipose tissue hypoxia ever, little is known of the effect of body composition on glucose metabolism in obese animals has been proposed as a possible initiator of ATMs infi ltra- in the aging female non-human primate (NHP). To address this question, we tion. The aim of this study is to clarify the relationship between hypoxia studied these variables in old female monkeys (Macaca mulatta, n = 19, age and ATM polarization in diet induced obese mice.To assess the hypoxic con- range 25-30 yrs). Each animal underwent magnetic resonance imaging and 1H ditions in adipose tissue, mice were intraperitoneally injected with 60mg/ spectroscopy to quantify total abdominal fat, visceral fat (VF), subcutaneous kg pimonidazole, a chemical probe for hypoxia, 30 min prior to sacrifi ce of fat (SQF) area, extramyocellular lipid (EMCL), intramyocellular lipid (IMCL), the mice. Immunoblotting with anti-pimonidazole antibody revealed that the intrahepatic lipid (IHL) content, and DEXA scan for whole body composition. hypoxic signals in epididymal adipose tissue were enhanced in high fat-fed A subgroup (n=11) were fasted and IVGTT glucose and insulin levels were mice. Flow cytometryanalysis demonstrated that the uptake of pimonida- zole into M1 ATMs was more than that into M2 ATMs. Furthermore, ATMs, measured for determination of HOMAIR.Unlike human females, the monkeys had roughly twice as much VF then SQF (median 4390 mm2 vs. 2260 mm2, P = which showed strong signals against pimonidazole in fl ow cytometry, also

Obesity <0.001). Total fat mass by DEXA correlated with SQF (P = 0.002) and IHL (P = expressed strong signals of proinfl ammatory cytokines such as TNF_, IL-6

POSTERS and IL-1 . To know the direct effect of hypoxia on macrophage polarization, 0.02). SQF correlated with HOMAIR (P = 0.013), but not after adjustment ` we cultured the bone marrow-derived macrophages or the stromal vascular for fat mass. IHL demonstrated the strongest correlation with HOMAIR (P =

Integrated Physiology/ 0.001) and remained signifi cant after adjustment for fat mass. VF, IMCL, and fractions of epididymal adipose tissue in hypoxic chamber with 1% oxygen for 24 hours. In these cells, the expression of M1 markers such as CD11c, IL-6 EMCL did not correlate with HOMAIR.Despite a greater amount of VF com- pared to SQF, VF did not associate with markers of insulin resistance (IR) and IL-1` were upregulated by the hypoxic treatment. These result suggest in the older female monkey. IHL content had the strongest relationship to that obesity-induced hypoxia in adipose tissue is involved in M1polarization of ATMs. HOMAIR and was also related to post-glucose challenge levels of insulin. Therefore, IHL, not total abdominal fat or muscle fat, is a marker for IR in the fasting state in elderly female monkeys.

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A492 OBESITY—ANIMALCATEGORY

1926-P C57BL6 mice fed standard chow (SC) or high-fat diet (HF), KK and KKAy mice Distribution of Adipose Tissue Macrophages in Epididymal Fat Pads fed ad-libitum (KK-AL and Ay-AL) or fed reduced amounts of SC (caloric re- of Obese Mice striction (CR); Ay-CR) were studied by RT-PCR, western blot and FRET analy- TAKESHI SHIMADA, YASUHIKO YAMAMOTO, HYUEK JONG LEE, NOZOMU sis. Pharmacological inhibitors for JNK (SP600125; SP) and PKC (Go6983; Go) KAMEI, JONGSOON LEE, STEVEN E. SHOELSON, Boston, MA, Wakayama, Japan, were used to investigate the signaling pathways. Compared with C57BL6 Kanazawa, Japan, Tokyo, Japan mice fed SC, mice fed HF developed obesity and showed higher expression Obesity-induced infl ammation is associated with the recruitment of adi- and activity of TACE. Ay-AL mice were heavier and showed higher expres- pose tissue macrophages (ATMs), and ATM number correlates with degrees sion and activity of TACE in VAT compared with those of KK-AL mice. HF- of insulin resistance. In lean mice ATMs are predominately found individu- fed C57BL6 and Ay-AL mice showed elevated phosphorylation of JNK and ally, but in obesity ATMs surround isolated adipocytes to form crown-like PKC in VAT compared with the controls. Ay-CR mice had less body weight, structures (CLSs). During histological assessments of isolated ATM and CLS lower glucose and insulin levels, and showed lower expression and activ- frequencies, we noted apparent differences in regional distributions, most ity of TACE in VAT compared with Ay-AL mice, which were associated with obviously for CLSs. We hypothesized that CLSs represent dead adipocytes reduced phosphorylation of the kinases. In in-vitro study using 3T3-L1 adipo- that were surrounded by ATMs in a typical tissue repair and remodeling cytes, TACE expression and activity increased 2~3-fold either by TNF-_ or type mechanism, and that adipocyte death might vary across the fat pad LPS stimulation, and pretreatment with SP or Go partially suppressed these and thus occur with regional specifi city. Epididymal (Epi) fat pads were iso- events. Intraperitoneal injection of SP strongly reduced the TACE activity lated after 4 or 8 weeks feeding of high fat diet (HFD) vs normal chow (NCD). in VAT, and suppressed both JNK and PKC signaling in Ay-AL mice. These The fat pads were divided into 3 regions from cranial to caudal we refer to results indicate that expression and activity of TACE in VAT are up-regulated as tip, body and base. These were analyzed using immunohistochemistry, during the development of obesity at least in part via activation of JNK and fl ow cytometry and quantitative PCR. HFD progressively increased fat pad PKC, and benefi cial effects of CR on obesity-induced insulin resistance might mass, ATM number, and severity of insulin resistance. Histomorphometry be mediated in part by inhibition of TACE activity in VAT. and fl ow cytometry consistently yielded over 2-fold greater CLSs in the tip regions than the base. ATM marker genes (CD11b, F4/80 and CD68) were 1929-P also 2- to 4-fold greater in the tip. Numbers of CD11c positive ATMs, which Increased Adipose Tissue Growth Hormone Activity in Insulin Sen- are selectively found in CLSs, were also 2- to 3-fold greater in tip regions. sitive, High Fat Diet-Fed Skeletal Muscle-Specifi c Growth Hormone After 8 weeks HFD, both M1 (TNF_, MCP1) and M2 (Mgl1, Arg-1 and TGF-`) Receptor Knockout Mice markers as well as markers for remodeling were greater in the tip region. ARCHANA VIJAYAKUMAR, YINGJIE WU, NICHOLAS J. BUFFIN, HUI SUN, SHO- These fi ndings show that adipocyte death and the macrophage response to SHANA YAKAR, DEREK LEROITH, New York, NY sterile AT infl ammation occurs more frequently in the tip regions of Epi fat We have previously shown that loss of growth hormone receptor (GHR) pads. The tip regions are also where the greatest degrees of adipose tissue signaling in mouse skeletal muscle in the mGHRKO mice protects them from expansion and contraction occur during the induction of obesity and weight high-fat diet (HFD)-induced obesity and insulin resistance. Interestingly, the loss, respectively. Our fi ndings therefore demonstrate that the macrophages HFD-fed mGHRKO mice do not accumulate fat mass despite having similar in CLSs are involved in AT remodeling and that this occurs to the greatest food intake as the control mice. The predominant effects of growth hormone degree in the tip regions of epididymal fat pads. (GH) in the adipose tissue are to stimulate lipolysis by activating hormone sensitive lipase (HSL), and to suppress triglyceride (TG) uptake by inhibit- 1927-P ing lipoprotein lipase (LPL) activity thereby reducing fat accumulation. Thus, Deletion of Prolyl Carboxypeptidase Attenuates the Metabolic we hypothesized that loss of muscle GHR signaling results in a compensa- Effects of Diet-Induced Obesity tory increase in adipose tissue GH action. Accordingly, we found a fi ve-fold JIN KWON JEONG, GIUSEPPINA MATTACE-RASO, ROSARIA MELI, SABRINA increase in ghr mRNA expression and a two-fold increase in GHR protein DIANO, New Haven, CT, Naples, Italy content in the epidydimal fat pad of the HFD-fed mGHRKO mice. The HFD-fed Alpha-melanocyte-stimulating hormone (_-MSH) is a critical regulator of mGHRKO mice had a greater extent of HSL phosphorylation at Ser660, and energy metabolism. Prolyl carboxypeptidase (PRCP) is an enzyme respon- Ser563 residues under basal conditions which are markers of HSL activation. sible for its degradation and inactivation. PRCP null mice (PRCPgt/gt) showed Furthermore, in response to 12h or 24h fasting, a condition that increases GH elevated level of brain _-MSH, reduced food intake and a leaner pheno- secretion and is also associated with increased adipose tissue HSL activ- type compared to wild type controls. In addition, they are protected against ity, the mGHRKO mice demonstrated signifi cantly elevated FFA levels. When diet-induced obesity. Here we show that PRCPgt/gt animals have improved challenged with an oral fat tolerance test which is an indirect measure of LPL metabolic parameters compared to wild type controls both under a standard activity, the mGHRKO mice demonstrated an enhanced peak suggestive of chow diet (SD) as well as on high fat diet (HFD). Both on SD and HFD, PRCPgt/ slower disposal of TG into peripheral tissues, the adipose tissue being pre- gt mice showed a leaner phenotype due to decreased fat mass, increased en- dominant of these. Thus, our data suggest that in light of negligible muscle ergy expenditure and locomotor activity. They also showed improved insulin GHR signaling, there is an increase in adipose tissue GH action by as yet sensitivity and glucose tolerance compared to WT controls, and a signifi cant unknown mechanisms. GH action in the adipose tissue has been previously reduction in fasting glucose levels. These improvements occur before chang- attributed to insulin resistance; however, an interesting implication of our es in body weight and composition were evident. In support of a reduced study is that increased GH action in the adipose tissue, in the absence of gluconeogenesis, liver PEPCK and G6Pase mRNA levels were signifi cantly muscle GHR signaling, under obese conditions may in fact be benefi cial to reduced in PRCPgt/gt compared to WT mice. A signifi cant decrease in liver overall insulin sensitivity. weight and hepatic triglycerides were also observed in PRCPgt/gt compared to WT mice exposed to both diets. All together, our data suggest that PRCP 1930-P is an important regulator of energy and glucose homeostasis as its deletion Neuropeptide Y Regulation of Adipose Tissue Macrophages signifi cantly improves metabolic parameters in mice exposed to both stan- KANAKADURGA SINGER, DAVID L. MORRIS, KELSIE OATMAN, JENNIFER DEL- dard chow and HF diets. PROPOSTO, CAREY N. LUMENG, Ann Arbor, MI

Supported by: NIH/NIDDK (084065) Chronic stress is associated with obesity and insulin resistance partially Obesity

via accumulation of adipose tissue macrophages (ATMs), but the mecha- POSTERS 1928-P nism is unclear. Neuropeptide Y (NPY) is induced during chronic stress,

Obesity Up-regulates TNF-ƴ Converting Enzyme (TACE) in Visceral modifi es leukocyte function and may link stress to obesity and infl amma- Integrated Physiology/ Adipose Tissues via JNK and PKC Activation tion. The specifi c role of NPY in ATM function is unknown. In this study, SHUJI KAWASAKI, HIROYUKI MOTOSHIMA, SATOKO HANATANI, YUKI TAKAKI, we hypothesize that NPY infl uences metabolism by modifying macrophage MOTOYUKI IGATA, TAKESHI MATSUMURA, TATSUYA KONDO, TAKAFUMI SE- activation state. Visceral fat depots from db/db, lean and obese male C57 NOKUCHI, TAKESHI NISHIKAWA, EIICHI ARAKI, Kumamoto, Japan mice were separated into stromal-vascular fraction (SVF) and adipocytes Inhibition of TNF-_ converting enzyme (TACE), which involves in produc- by collagenase digestion. After fl uorescent activated cell sorting (FACS) tion of TNF-_, resulted in improvement in glucose and insulin levels in a for ATMs (CD11b+F4/80+). Analysis of Npy expression by RT-PCR showed diabetic mouse model, suggesting a crucial role of TACE activity in insulin increased expression in ATMs in obese mice (p=0.035) and in db/db mice resistance. However, the tissue responsible for TACE activation is largely (p=0.006) compared to lean animals. Production of NPY by bone marrow unknown. In this study, we investigated the impact of visceral adipose tissue derived dendritic cells (BMDC) and SVF cells was confi rmed by ELISA. To (VAT) as a target of TACE activation. The expression and activity of TACE in study the effects of peripheral NPY in fat, NPY and NPY scrambled peptides

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A493 OBESITY—ANIMALCATEGORY

(60 ug/kg) were injected intraperitoneally in lean mice for 5-10 days. NPY with lowered PEDF secretion by 45.64 and 44.64% respectively in two IR administration led to a relative decrease in total ATMs (p=0.07) driven by a cell models, as well as downregulated PEDF and upregulated p-AMPK pro- decrease in CD11b+CD11c+ M1 ATMs (p=0.04) and a decrease in circulating tein levels. With compound C treatment, the inhibitive effect of metformin classical Ly6chi monocytes (p=0.045). Analysis of visceral fat gene expres- on PEDF secretion and protein expression had been attenuated in both cell sion demonstrated decreased expression of M1 genes (iNOS, Il6, MHCII) in models.In summary, during IR improvement, metformin inhibits expression fat. Histologic examination showed increased adipocyte size, and further and secretion of PEDF in liver and fat, which might through promoting AMPK metabolic examination revealed no changes in glucose metabolism. In vitro, phosphorylation. BMDC from lean animals were treated with NPY receptor antagonists, Supported by: The National Natural Science Foundation of China (No. 81170751) which increased CD11c+ expression. These results suggest that ATMs are a regulated source of NPY production. NPY administration to lean mice has 1933-P anti-infl ammatory effects on M1 ATMs and circulating classical monocytes Panaxatriol Activates Insulin Signaling Pathway in Skeletal Muscle suggesting a regulatory role for NPY in adipose tissue infl ammation and and Improves Insulin Resistance in KKAy Mice stress responses. KUMIKO KITAMURA, YUSUKE TAKAMURA, TAKU IWAMOTO, HIRONOBU SUG- Supported by: Endocrine Fellows Foundation, Pediatric Endocrine Society IOKA, TARO KOBAYASHI, TOHRU KOMADA, YASUKO MANABE, HIDEAKI IWASA- KI, MOTOYASU OHDERA, NOBUHARU FUJII, TOHRU FUSHIKI, Odawara, Japan, 1931-P Hachioji, Japan, Ibaraki, Japan, Kyoto, Japan Glutamate Release in the Mediobasal Hypothalamus Correlates Previously we reported that panaxatriol (PT, a dammarane-type triterpene With Feeding Behavior and is Increased upon Exposure to an Obe- contained in Panax notoginseng), elicits an anti-diabetic effect by oral ad- sogenic Diet ministration (ADA, 109-LB, 2011). In this study, we further explored mecha- STEPHAN J. GUYENET, MILES E. MATSEN, GREGORY J. MORTON, MICHAEL W. nism of the effect. Genetically obese KKAy mice, (4 weeks old) were fed SCHWARTZ, Seattle, WA a high-fat diet without or with 0.12% PT for 7 days. Insulin (0.8U/kg) was Recent work demonstrates that in rodents, purifi ed high-fat diet (HFD) injected in mice of the both groups after 15 hours fasting and sequential feeding rapidly induces infl ammation (within 24 hr) and markers of neuron in- blood glucose was measured. Insulin sensitivity of PT intake group was jury (within 7d) and reactive gliosis in the arcuate nucleus of the mediobasal signifi cantly higher than that of control group. In addition, a hyperinsuline- hypothalamus (MBH), an important brain area for the control of energy bal- mic-euglycemic clamp study showed that PT intake improves whole body ance and adiposity. To explain this phenomenon, we hypothesized that the insulin resistance of the KKAy mice (Fig.1). Phospholylation of muscle Akt switch to HFD triggers excessive glutamate release and associated hyper- in PT group was signifi cantly higher than that in control group, indicating activation of MBH circuits, leading to excitotoxicity. As a fi rst test of this that PT intake diminished progression of muscle insulin resistance. Further- hypothesis, we asked whether feeding stimulates MBH glutamate release more, addition of PT in cell culture media for L6 myotubes showed increase by implanting a glutamate-sensitive probe (Pinnacle) with a one second tem- in glucose consumption in media. Moreover, the increase was suppressed poral resolution unilaterally into the MBH of free-living rats. Our preliminary by addition of an Akt1/2 kinase inhibitor (Fig.2). These results suggest results show that feeding (but not drinking) behavior correlated tightly with that one of the anti-diabetic mechanisms of PT intake is enhancement of MBH glutamate release. Moreover, when chow-fed animals were acutely glucose transport through insulin signaling pathway in skeletal muscle. exposed to HFD compared to chow at dark-cycle onset, marked increases were observed in both the maximal glutamate increment at the onset of a feeding bout (271 uM vs. 18.9 uM) and the total glutamate response (range 21-2,225 uM vs. 4.5-293 uM) during the fi rst 30 s of feeding. Comparison of mean values showed that the glutamate response was consistently 4-7 fold higher during HFD compared to chow feeding within individual animals. This response was not observed in all animals, and variability in glutamate release between individuals appears to be explained by subtle differences in probe placement.Although preliminary, these fi ndings collectively suggest that feeding triggers glutamate release in the MBH and that this release is markedly increased when animals consume an obesogenic HFD. As such, they offer a plausible mechanism for the rapid-onset infl ammation and neu- ronal injury induced by HFD feeding, a phenomenon causally implicated in obesity pathogenesis.

1932-P Metformin Inhibits Expression and Secretion of PEDF in Liver and Fat via Promoting AMPK Phosphorylation SHUMIN YANG, QIFU LI, QINGFENG CHENG, WEI XIA, LI ZHONG, MEI LUO, Chongqing, China Pigment epithelium-derived factor (PEDF) as an adipokine has been shown to induce insulin resistance (IR). The anti-diabetic drug, metformin, can im- prove IR, although its mechanism is unclear. This study aimed to investigate the effect of metformin on the expression and secretion of PEDF in liver and fat during IR improvement.SD rats were fed normal diet (NC group) or high-fat diet for 15 weeks, then to be gavaged saline (IR group) or metformin 400mg/ kg/d (MET group) for 4 weeks. Hyperinsulinemic-euglycemic clamp test was

Obesity performed. IR models of 3T3-L1 and HepG2 cell were established by in vitro

POSTERS hyperinsulinemia and validated by 2-(N-(7-nitrobenz-2-oxa-1, 3-diazol-4-yl) amino)-2-deoxyglucose (2-NBDG) uptake; 0.1mM metformin and/or 40μM

Integrated Physiology/ compound C (an AMPK inhibitor) were used for the intervention.Compared with the NC group, the IR group showed decreased glucose infusion rate, accompanied with increased serum PEDF levels (1.77±0.14 vs 2.16±0.09 μg/ ml, p<0.05) which negatively correlated with GIR (r=-0.79, p<0.05). The MET group revealed signifi cantly improved IR, concomitant with downregulated PEDF and upregulated p-AMPK protein levels in liver, white adipose tissue, brown adipose tissue and muscle. Treated with 5*10-6mM insulin for 24h, 3T3-L1 adipocytes and HepG2 cells showed decreased 2-NBDG uptake, ac- companied with increased PEDF secretion by 2.10 and 2.12 folds respectively in two cell lines, as well as upregulated PEDF and downregulated p-AMPK protein levels. Whereas metformin increased 2-NBDG uptake, accompanied

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A494 OBESITY—ANIMALCATEGORY

1934-P 1936-P Effects of Dietary Obesity on Hepatic SREBP-1c, ChREBP and AMPK Supplementation With Cholic Acid Reduces Body Weight, but Pro- in Rats duces Hepatotoxicity in Young Mice Fed a High-Fat Diet KATHLEEN V. AXEN, MARIANNA A. HARPER, YU-FU KUO, STEVEN D. CLARKE, HUIJUAN MA, ELVIRA ISGANAITIS, MICHAEL CHEN, WEN KONG, ARISTIDES JEAN GRASSMAN, KENNETH AXEN, Brooklyn, NY LYTRAS, MARY-ELIZABETH PATTI, Boston, MA Hepatic lipogenesis has been implicated in the development of fatty liver Beyond their well-known role in lipid and cholesterol metabolism, bile ac- and insulin resistance in obesity. We investigated the expression of SREBP- ids (BAs) have been identifi ed as regulators of glucose homeostasis and en- 1c and ChREBP, two regulatory transcription factors for lipogenesis, and of ergy expenditure. Dietary supplementation with cholic acid has been shown their main targets, Acetyl CoA Carboxylase (ACC) and Fatty Acid Synthase to protect high fat diet-fed mice against obesity. However, these studies (FAS), in dietary obese rats under both fasted and glucose-stimulated con- were performed in adult mice and utilized a relatively high dose. We asked ditions. We also investigated the expression and activity of AMP Kinase whether lower-dose bile acid supplementation might reduce risk of obesity (AMPK), which inhibits lipogenesis and promotes fat oxidation.Male Sprague and glucose intolerance in young mice. Male C57BL/6 mice, age 3 weeks, Dawley rats consumed a high-fat (HF: 55% of kcal as fat, 30% SFA, N=8) or were treated with high-fat diet, with or without bile acid supplementation, low-fat (LF: 15% fat, N=8) diet ad lib. Compared with LF rats, HF rats at wk for 4 weeks: (1) High fat diet (HFD, 45% fat), (2) HFD supplemented with 10 had higher body fat, plasma leptin, and hepatic lipid (p<0.01), as well as 0.1% w/w cholic acid (0.1 CA), (3) HFD with 0.25% w/w cholic acid (0.25 CA), impaired plasma glucose responses to an ip glucose tolerance test (p<0.01). and (4) HFD with 0.25% w/w ursodeoxycholic acid (UDCA). Food intake did Liver samples obtained after a 16 h fast or 3h after an oral glucose load (2g/ not differ, but body weight was lower in 0.25 CA (9.1% decrease vs. HFD, kg) were analyzed by: 1) qPCR for expression of SREBP-1c, ACC, FAS, and p=0.03). Blood glucose (6 hour fast) was reduced in UDCA-treated mice (222 AMPK and 2) immunoblotting for content and distribution of ChREBP (nuclear ±11 vs. 266 ± 16 mg/dl in HFD, p=0.042, with similar trends in the 0.25 CA vs cytoplasmic) and AMPK activity. In the fasting state, hepatic SREBP-1c group (p=0.08). Insulin levels did not differ between groups. Animals were expression was higher in HF than LF rats. Fasting failed to stimulate AMPK sacrifi ced at 4 weeks for tissue and blood analysis. Liver weight (% of body activity in both HF and LF rats, although it did so in young, lean rats.The weight) was increased in both cholic acid groups (p<0.05 vs. HFD). Moreover, oral glucose load: 1) stimulated SREBP-1c and its targets ACC and FAS in LF serum ALT levels were higher in 0.25 CA groups as compared to HFD (7.6±1.3 but not HF rats; 2) increased ChREBP content in LF (p<0.01) but not HF rats, vs. 4.0±0.4 U/L, p=0.03). In parallel, liver histology demonstrated increased although ChREBP distribution was similar between groups; and 3) stimulated lipid accumulation in cholic acid-treated mice vs. HFD control. Liver weight, AMPK expression in HF but not LF rats. These results suggest that obesity ALT, or histology did not differ in UDCA-treated group vs. HFD control. In produced by a high-fat diet leads to: 1) a predisposition to hepatic lipogene- conclusion, cholic acid supplementation reduced weight gain in young mice sis in the fasting state, due to enhanced expression of SREBP-1c and a failure fed a high fat diet, but also produced mild hepatotoxicity. By contrast, UDCA of AMPK to inhibit lipogenesis; and 2) metabolic infl exibility in response to supplementation reduced glucose, independent of weight gain, without he- glucose, due to impaired augmentation of SREBP-1c, ACC, and FAS expres- patotoxicity. While BA supplementation is an intriguing avenue for treat- sion, and of ChREBP content. Taken together, these effects would promote ment of metabolic disease, caution must be exercised when designing stud- accumulation of hepatic lipid and compromise the liver’s ability to respond ies as certain bile acid species may result in hepatotoxicity. to a carbohydrate load by synthesizing fat. Supported by: NIH (1SC3GM086298) 1937-P Macrophage Lipoprotein Lipase Modulates the Development of Ath- 1935-P erosclerosis, but not Obesity Manganese Tetrakis Benzoic Acid Porphyrin Reduces Adiposity MANABU TAKAHASHI, HIROAKI YAGYU, FUMIKO TAZOE, SHUICHI NAGASHIMA, and Improves Insulin Sensitivity in Mice Fed a High Fat Diet TAICHI OSHIRO, KENTA OKADA, JUN-ICHI OSUGA, IRA J. GOLDBERG, SHUN JONATHAN R. BRESTOFF, TIMOTHY BRODSKY, AARON SHEPPARD, LILY NG, ISHIBASHI, Shimotsuke City, Japan, New York, NY MARIA DISANTO-ROSE, ELENA STANSKY, LEILA FUSSELL, THOMAS H. REYN- Atherosclerosis and obesity are associated with chronic infl ammation. OLDS, Saratoga Springs, NY In addition, both processes are associated with and likely driven by mac- Metallic porphyrins appear to promote weight loss and stimulate mito- rophage infi ltration and activation within tissues. Lipoprotein lipase (LpL), chondrial biogenesis, in part, by inducing the heme oxygenase-1 system. The which is mainly synthesized by muscle and adipose tissue, plays a crucial present study determined the effects of manganese tetrakis benzoic acid role in plasma lipoprotein metabolism by hydrolyzing triglycerides in VLDL porphyrin (MnTBAP) on insulin sensitivity, adiposity, and the expression of and chylomicrons. Furthermore, LpL is also synthesized by macrophages and HO-1 and PGC-1_. C57B6 mice were fed a high fat diet (HFD) for six months. macrophage LpL is thought to promote atherosclerosis. However, the role During the last 30 days of the dietary intervention mice received daily intra- of LpL in adipose macrophage biology has not been explored. In this study, peritoneal injections of MnTBAP or vehicle. Insulin sensitivity, as assessed we generated a macrophage-specifi c LpL knockout (MLpLKO) mouse using by an insulin-assisted glucose tolerance test, improved signifi cantly in mice cre/loxP gene targeting, and determined the role of macrophage LpL in ath- treated with MnTBAP compared to vehicle (Area Under Curve: 9,516±1,217 v. erosclerosis and obesity. Loss of LpL in macrophages did not alter plasma 12,264±549; p=0.02). Mice treated with MnTBAP exhibited signifi cant reduc- LpL activity or plasma lipoproteins. MLpLKO mice gained weight similarly tions in body weight (40.1±2.1 vs. 52.8±2.2 g; p < 0.01), subcutaneous white to control mice on a 10-week high fat diet. Peritoneal macrophages from adipose tissue weight (1.75±0.14 v. 3.11±0.17 g; p<0.01), and epididymal apolipoprotein E (ApoE) knockout mice lacking macrophage LpL (MLpLKO/ white adipose tissue weight (EWAT, 1.47±0.13 v. 2. 54±0.11 g; p<0.01) when ApoEKO) had decreased cholesteryl ester formation by 51% (p<0.01) com- compared to mice treated with vehicle. The reductions in adiposity were not pared to ApoEKO mice, when incubated with ApoE-defi cient `-VLDL. VLDL related to changes in caloric intake (22.16±4.1 v.18.24±2.0 kcal/day; p=0.24) loading did not increase triglyceride accumulation in MLpLKO/ApoEKO mac- indicating that MnTBAP enhances metabolism. HO-1 and PGC-1_ expression rophages. CD36 and carnitine palmitoyltransferase-1 levels in MLpLKO/ were signifi cantly higher in EWAT of mice treated with MnTBAP compared to ApoEKO macrophages were suppressed by 45% (p<0.01) and 61% (p<0.01), mice treated with vehicle (HO-1: 9.60±0.19 v. 3.46 ±1.46; PGC-1_: 2.10±0.29 respectively, as compared to ApoEKO macrophages. Although both MLpLKO/ v.1.15±0.15). Since carbon monoxide (CO) is a product of HO-1 activity and ApoEKO and ApoEKO mice developed comparable hypercholesterolemia on

stimulates mitochondrial biogenesis, we assessed the effect of six weeks a 12-week Western-type diet, aortic fatty streak lesion size was signifi - Obesity

of CO treatment (250 ppm, 1 hour/day). CO treatment did not result in an im- cantly less in MLpLKO/ApoEKO mice. In adipose tissue, despite the similar POSTERS provement in insulin sensitivity or a reduction in body weight, indicating that expression levels of macrophage markers, CD68 and F4/80, LpL expression

an increase in endogenously produced CO did not likely mediate the reduc- was not lower in MLpLKO/ApoEKO compared to ApoEKO mice. Additionally Integrated Physiology/ tion in adiposity in MnTBAP treated mice. These fi nding demonstrate that TNF-_ and MCP-1 expressions were not different between the two groups. MnTBAP treatment enhances insulin sensitivity and decreases adiposity in In conclusion, macrophage LpL plays an important role in the development of mice by a mechanism that may be independent of HO-1 expression. atherosclerosis, but not obesity. Supported by: NIH (1R15DK090722-01) Supported by: Ministry of Education, Takeda Science Foundation, Jichi Medical University

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A495 OBESITY—ANIMALCATEGORY

1938-P 24 hours was markedly decreased in AgRP-ROCK1 defi cient mice compared Attenuation of Cytokine Signaling Rescues Diet-Induced Obesity with control mice. The majority of the decrease occurred during the dark and Insulin Resistance in Lymphocyte-Defi cient Rag2 Null Mice cycle. Taken together, these data suggest that activation ROCK1 in AgRP RANDALL H. FRIEDLINE, YONGJIN LEE, DAE YOUNG JUNG, RITA BORTELL, HWI neurons is required for the homeostatic regulation of energy expenditure JIN KO, SEZIN DAGDEVIREN, EUNJUNG LEE, VIVIAN M. BENOIT, PAYAL R. PATEL, and adiposity, suggesting an important role for ROCK1 in modulating energy NICHOLAS TSITSILIANOS, XIAODI HU, YOSHIHIRO AZUMA, HSUN-FAN WANG, metabolism on AgRP neurons. UMBER SHAFIQ, YOJIN PARK, MICHELLE B. MEYERS, DALE GREINER, JASON K. KIM, Worcester, MA 1940-P Insulinresistance is a hallmark of obesity and type 2 diabetes. Alterations Exploratory Factor Analysis of the Cardiometabolic Syndrome in in immunecell types or infl ammatory processes affect energy balance and Rhesus Monkey (Macaca Mulatta) Identifi es a Covariance Struc- glucosehomeostasis. Here we examined the metabolic effects of high-fat ture Consistent With Findings in Humans diet (HFD) inlymphocyte-defi cient Rag2 null mice (Rag2) and mice with com- PAUL B. HIGGINS, JENNIFER D. NEWCOMB, YI-XIN (JIM) WANG, BARBARA C. binedablation of Rag2 and the IL-2 receptor common gamma (aC) chain (Rag2. HANSEN, Kannapolis, NC, Taicang, China, Tampa, FL aC), which mediates intracellular cytokinesignaling. A 2-hr hyperinsulinemic- Factor analysis has been used to study the covariance structure among euglycemic clampwith 3H-glucose and 14C-2-deoxyglucose was performed components of the cardiometabolic syndrome (CMS) in efforts to better toassess insulin sensitivity in awake wild-type(WT), Rag2, and Rag2.aC mice understand its pathophysiology. Common CMS Factors have beendescribed (n=8-15/group). On chow diet,Rag2.aCmice were leaner than Rag2 and WT across diverse human populations. CMS Factors have not yet been identi- mice and more insulin sensitive withsignifi cant increases in whole body glu- fi ed in non-human primates (NHPs); determining the degree of CMS Factor cose turnover (Fig. 1; *P<0.05 vs. WT). During 6 wks of HFD,Rag2 mice be- concordance between humans and NHPs would by extension yield further came markedly more obese than WT mice, whereas Rag2.aC mice were highly insight into the extent of etiologic commonality. Toward this end, we report resistant todiet-induced obesity (Fig. 2). Obese Rag2 and WT mice developed fi ndings from a Factor Analysis on CMS variables in non-diabetic rhesus ma- severe insulinresistance in the liver as insulin failed to suppress hepatic glu- caques (17.1±5.5yrs, 13.9±4.2Kg, n=54) at various stages of CMS, maintained coseproduction during clamps (Fig. 3). In contrast, Rag2.aC mice remained in a controlled laboratory setting. Concentrations of overnight fasted plasma highly insulin sensitive inthe liver after HFD. Insulin-stimulated whole body glucose (FPG), insulin (FI), triglyceride, and HDL-cholesterol were determined. glucose turnover was reducedin HFD-fed Rag2 mice, but remained normal Glucose clearance rate (Kg) and acute insulin response (AIR) to glucose were in HFD-fed Rag2.aC mice (Fig. 4). Overall, these results showthat disrupting determined using an intravenous glucose tolerance test. Exploratory Prin- the cytokine network improves insulin sensitivity and preventsdiet-induced cipal Components Factor Analysis employing Varimax rotation was used to obesity. Our fi ndings further implicate a criticalrole of cytokine signaling in identify factors underlying the clustering of these variables. Two Factors regulating obesity and insulin resistance. emerged (Kaiser Myer Olkin value=0.70) with eigenvalues of 3.2 and 1.5, respectively. Factor 1 explained 48.5% of the covariation (highly signifi cant loadings from FPG, FI, Kg, and AIR) and Factor 2 explained 31.3% of cova- riation (signifi cant loadings from HDL-cholesterol, triglyceride, FI, and Kg). Consistent with studies in humans, we identifi ed the presence of statisti- cally independent glycemia- and dyslipidemia-related factors underlying the structure of the CMS in rhesus macaques. Furthermore and in keeping with clinical fi ndings, we have documented the loading of FI and Kg on both Fac- tors. Our results indicate that a major part of the latent covariance structure of the CMS in monkeys is similar to that of humans. Supported by: NIH (HHSN263200800022C, NO1-AG-3-1012)

1941-P High Fat Diet-Induced Changes in Hepatic Protein Abundance in Mice MOULUN LUO, APRIL E. MENGOS, TIANNA M. STUBBLEFIELD, LAWRENCE J. MANDARINO, Scottsdale, AZ Nonalcoholic fatty liver disease (NAFLD) is associated with obesity, insu- lin resistance, and type 2 diabetes. The purpose of this study was to identify novel proteins and pathways that contribute to the pathogenesis and com- Supported by: NIH (R01-DK080756) plications of NAFLD. C57BL/6J male mice were fed a 60% (HFD) or 10% (LFD) high or low fat diet. HFD induced obesity, hepatic steatosis and insulin resis- tance (glucose clamps, glucose infusion rate: LFD 50.5 ± 6.4 vs. HFD 14.2 ± 1939-P 9.5 mg/(kg.min); n = 12). Liver proteins analyzed by mass spectrometry-based Rho-Kinase Regulates Adiposity by Controlling Energy Expenditure proteomics showed numerous hepatic proteins were altered in abundance and Locomotor Activity in AgRP Neurons by 60% HFD. Nine down-regulated and nine up-regulated proteins were se- HU HUANG, DONG KONG, TIEMIN LIU, IL JI PAEK, BRADFORD B. LOWELL, YOUNG lected for detailed analysis based on the criteria of 1.5-fold difference and BUM KIM, Boston, MA consistency across replicates. Proteins that decreased in abundance were Normal leptin signaling is essential for the maintenance of body weight acyl-coA desaturase-I (SCD-1), acetyl-CoA carboxylase (ACC), fatty acid syn- and energy homeostasis. In the hypothalamus, the POMC- and AgRP/NPY- thase (FAS), pyruvate kinase isozymes R/L (PKLR), NADP-dependent malic producing neurons act as key mediators of leptin action, but the understand- enzyme (ME-1), ATP-citrate synthase (ACL), ketohexokinase (KHK), long- ing of the intracellular transducers of leptin receptor signaling is incomplete. chain-fatty acid-CoA ligase-5 (ACSL-5) and carbamoyl-phosphate synthase-I

Obesity To determine the role of Rho-kinase 1 (ROCK1) in the hypothalamic control (CPS-1). Those that increased were KIAA0564, apolipoprotein A-I (apoA-1),

POSTERS of body-weight homeostasis, mice lacking ROCK1 in AgRP neurons (AgRP- ornithine aminotransferase (OAT), multidrug resistance protein 2 (MRP-2), ROCK1) were studied. Defi ciency of ROCK1 in AgRP neurons caused a mod- liver carboxylesterase-I (CES-1), aminopeptidase N (APN), fatty aldehyde

Integrated Physiology/ est increase in body weight (Male, control mice 33.2 ±1.3g, AgRP-ROCK1 dehydrogenase (FALDH), major urinary protein 2 (MUP-2) and KIAA0664. defi cient mice 37.0±1.3g, p<0.01). This occurred independent of hyperphagia. KIAA0564 and KIAA0664 are uncharacterized, novel proteins. Decreased Consistent with this, MRI analysis indicated that fat mass in AgRP-ROCK1 abundance of normally highly abundant proteins like FAS and CPS-1 was con- defi cient mice was signifi cantly increased by ~50% with a modest decease in fi rmed by Coomassie Blue staining after bands were identifi ed by MS/MS, lean mass, compared with control mice. Adiposity in AgRP-ROCK1 defi cient and immunoblot analysis confi rmed increased abundance of KIAA0664 after mice is accompanied by increased plasma leptin and insulin levels. These 60% HFD. In conclusion, NAFLD is characterized by changes in abundance of effects are mainly due to dysregulated energy metabolism, as evidenced proteins related to cell injury, infl ammation, and lipid metabolism. Two novel, by the fact that oxygen consumption was greatly reduced in AgRP-ROCK1 uncharacterized proteins, KIAA0564 and KIAA0664, may provide insight into defi cient mice compared with control mice (control mice 5124±261 ml/kg/ the pathogenesis of NAFLD induced by lipid oversupply. hr, AgRP-ROCK1 defi cient mice 4561±272 ml/kg/hr, p<0.01). However, heat Supported by: NIH (R01DK47936 and R01DK66483) production was unaltered in these mice. Interestingly, locomotor activity for

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A496 OBESITY—ANIMALCATEGORY

1942-P prevention of obesity. However AIM stimulates infl ammation of adipocytes Role of Opioid Receptors in Regulating Feeding Behavior of Mice simultaneously, which fi nally leaded to insulin resistance. CTLA-4 (Cytotoxic HIROKO IKEDA, CHRISMAWAN ARDIANTO, TAKAHIRO OHASHI, MEGUMI ASA- T-Lymphocyte Antigen), expressed on the surface of Helper T cells, binds to TO, HIROSHI NAGASE, JUNZO KAMEI, Tokyo, Japan CD80/86 (macrophage) and transmits an inhibitory signal to T cells. We here Controlling feeding behavior is one of the effective ways to treat obe- show the benefi cial effect of CTLA4 on weight gain, insulin resistance, and sity. Many substances and neural transmissions are involved in controlling glucose intorelance in high fat diet (HFD) mice, in parallel with the up regula- feeding behavior, and one of the key players is opioid system. Opioid sys- tion of AIM and the down regulation of infl ammatory cytokine / chemokine tem plays an important role in motivation for not only addictive drugs but in the visceral fat. Six-week-old male C57Bl/6 mice were fed HFD or normal also palatable foods. However, the detailed mechanism of opioid system in chow diet (NCD) for 6 weeks, and divided into the CTLA4 (5mg/Kg; two times feeding behavior is unknown. Thus, the present study examined the role of a week) or saline administrated group (n = 10; respectively). The body weights opioid system in feeding behavior of mice. All experiments were performed in the group of HFD + CTLA4 were signifi cantly lower than the group of HFD using male ICR mice after food deprivation for 16 hours. The non-selective + Saline from only a week later of beginning the treatment. Both insulin and (naloxone, 3 mg/kg), mu (beta-funaltrexamine, 10 mg/kg), delta (naltrindole, glucose tolerance test showed signifi cant amelioration by CTLA4. The levels 3 mg/kg) and kappa (norbinaltorphimine, 20 mg/kg) opioid receptor antago- of mRNA for AIM were signifi cantly increased in the group of HFD + CTLA4, nists signifi cantly decreased food intake. The mu (morphine, 1 mg/kg), delta compared to other 3 groups. Furthermore, we evaluated mRNA for M1 / M2 (KNT 127, 1 mg/kg) and kappa (U50,488H, 3 mg/kg) opioid receptor agonists, macrophage markers and infl ammatory cytokine /chemokine. We confi rmed which dose is ineffective on locomotor activity, did not signifi cantly alter the shift of macrophage from M1 to M2 by CTLA4 in the HFD group, and food intake. Previous reports indicate that the stimulations of mu- and delta- reduction of infl ammatory cytokine / chemokine comparable to therevel of opioid receptors enhance motivational effects whereas the stimulation of the control group. In conclusion, CTLA4 regulates the expression of AIM and kappa-opioid receptors inhibits motivation. However, our results indicate infl ammatory cytokine / chemokine. Eventually, CTLA4 leads to prevention that the effect of kappa-opioid receptor blockade is the same as the effects of diet-induced obese and insulin resistance with shifting the polarization of of mu- and delta-opioid receptor blockade. Thus, it is possible that addition- macrophage. This immunotherapy may evolve the novel therapeutic target al effect of opioid systems rather than the motivational effect may control molecule for manipulating metabolic systems. feeding behavior. Since the hypothalamus is known to regulate energy ho- meostasis, we examined the changes of opioid system in the hypothalamus. The expression levels of mRNA for endogeneous opioids, proopiomelanocor- & 1945-P tin, proenkephalin and prodynorphin, in the hypothalamus were signifi cantly Triglyceride-Rich Lipoprotein Metabolism in the Central Nervous decreased after food deprivation. These results indicate that endogenous System Regulates Brain Lipid Sensing and Plays an Essential Role opioids in the hypothalamus are inhibited during fasting and that inhibition in Body Weight Regulation of increased opioids during feeding decreases feeding behavior. HONG WANG, CHARLES W. REHRER III, MATTHEW D. TAUSSIG, AMBER A. PALUMBO, KALYANI G. BHARADWAJ, IRA J. GOLDBERG, ROBERT H. ECKEL, Au- 1943-P rora, CO, New York, NY Enhanced Expression of Nesfatin/Nucleobindin-2 (NUCB2) in White Lipoprotein lipase (LPL) is rate-limiting in the hydrolysis of triglycerides Adipose Tissue of Ventromedial Hypothalamus (VMH)-Lesioned Rats (TG) and uptake of fatty acids from circulating dietary lipids. We have recent- AYA OHSAKI, HIROYUKI SHIMIZU, NORIKO ISHIZUKA, YUKO SUZUKI, SHUJI IN- ly created mice with neuron-specifi c LPL defi ciency (NEXLPL-/-) that have OUE, MASATOMO MORI, Maebashi, Japan, Midori, Japan reduced uptake of TG-rich lipoprotein-derived fatty acids and decreased Circulating concentrations of anorexigenic protein, nesfatin-1, are nega- levels of n-3 long chain polyunsaturated fatty acids (n-3 PUFAs) in the hypo- tively correlated with body mass index in humans. Nesfatin/Nucleobindin-2 thalamus. Of great interest these mice become obese on chow by 16 wk (Cell (NUCB2), a precursor of nesfatin-1 is widely expressed in the body. The exact Metabolism, 2011). We have now conducted experiments in 10 wk old mice to mechanism by which circulating nesfatin-1 concentration is regulated in vivo test if dietary supplementation of n-3 PUFAs prevents obesity. After 40 wk has been still unknown. In the present study, we investigated the infl uence on a synthetic high carbohydrate diet (HC, 70% CHO, 10% fat, n-6:n-3 ratio of the modulation of autonomic nerve activity balance by ventromedial hy- of 7:1) w/o n-3 PUFA supplement (n-6:n-3 ratio 1:3), NEXLPL-/- mice have an pothalamic (VMH) lesion on nesfatin/nucleobindin-2 (NUCB2) production in average of 35% fat mass vs. 27% in WT mice on both diets. After 24 wk on various peripheral tissues of Sprague-Dawley rats. Nesfatin/NUCB2 is most synthetic high fat diets (HF, 45% fat) w/o n-3 PUFA supplement, both NEX- potently expressed in pancreas and liver, and moderately in subcutaneous LPL-/- and WT mice gain less body weight (38.0 g vs. 39.1 g on HF/n-3 PUFA; white adipose tissue (SWAT), mesenteric white adipose tissue (MWAT) and 50.1 g vs. 47.6 g on HF diet; respectively). This reduction in body weight for interscapular brown adipose tissue (iBAT), but its expression was very weak both groups on the HF/n-3 PUFA diet is likely an effect of larger amounts of in skeletal muscle. VMH lesion caused a rapid increase of body weight and n-3 PUFA on palatability. Noticeably, HF feeding enhanced the body weight SWAT, MWAT weight accompanied with normoglycemic hyperinsulinemia, and fat mass gain in NEXLPL-/- mice much less than in WT mice, rendering but did not affect nesfatin/NUCB2 expression in pancreas, liver and skeletal similar body compositions at the end of the HF feeding (42.6% vs. 42.0% muscle. In contrast, nesfatin/NUCB2 expression was signifi cantly increased fat mass, respectively). Taken together, these data suggest that when LPL in both SWAT and MWAT of VMH-lesioned rats. However, no differences is defi cient in neurons, n-3 PUFA supplementation in the diet cannot pre- were observed on nesfatin/NUCB2 expression in the iBAT between VMH- vent obesity; moreover, NEXLPL-/- mice become insensitive to the amount lesioned and sham-operated animals. Continuous, peripheral delivery of of fat in the diet and fatty acid composition. Overall, results from the feeding carbachol (180 μg/kg body weight/hour) for 5 days did not affect nesfatin/ studies further support our hypothesis that LPL regulates TG-rich lipoprotein NUCB2 expression in the SWAT. In contrast, chemical sympathectomy with metabolism, providing important signaling molecule(s) such as n-3 PUFAs 6-hydroxy dopamine (50 mg/kg body weight) signifi cantly increased nesfa- that are involved in lipid sensing in the CNS, and plays an essential role in tin/NUCB2 expression in the SWAT with the reduction of body weight gain. regulation of energy balance and body weight. These data indicated that the modulation of autonomic nerve activity bal- Supported by: NIH (K-12 BIRCWH, NIH-NIDDK R01) ance by VMH lesion increases nesfatin/NUCB2 expression in white adipose

tissue of rats, perhaps via the inhibition of sympathetic nerve activity. Obesity Supported by: Japanese Ministry of Education, Culture, Sports, Science, and 1946-P POSTERS Technology Long-Acting PYY and GLP-1 Agonism in Combination Synergisti- cally Normalizes Weight, Glucose, Metabolic and Metabolomic Integrated Physiology/ 1944-P Parameters in Diet Induced Obese Mice CTLA4 Prevents Diet-Induced Obese and Insulin Resistance by MARK PAULIK, BRUCE HAMILTON, JOYCE BOUCHERON, BOB WIARD, ANDREA Upregulation of Apoptosis Inhibitor of Macrophage in Visceral Adi- STROUP, KAMAL AL-BARAZANJI, LUCY HOLT, CHRIS HERRING, MAKDA ME- pose Tissue BRAHTU, CHANDI ELANGBAM, SHARON WENG, SHANE ROLLER, REBECCA MASAKAZU FUJII, TOYOSHI INOGUCHI, NAONOBU SUGIYAMA, NORIYUKI HODGE, ANDREW YOUNG, LAURENT JESPERS, PAUL FELDMAN, Research Tri- SONODA, RYOICHI TAKAYANAGI, Fukuoka, Japan angle Park, NC, Cambridge, United Kingdom Obesity induces insulin resistance accompanied by low grade infl am- TM PYY(3-36) and exendin-4 conjugated to AlbudAbs (Albumin domain anti- mation of adipose tissue. Finally, it develops the increased risk of type 2 bodies) to extend half-life were tested in combination for weight loss in diet- diabetes, furthermore its associated complications.Recently it has reported induced obese (DIO) C57BL/6 mice. Combined administration of the PYY and that apoptosis inhibitor of macrophage (AIM) induced lipolysis resulting in exendin-4 peptides every other day at ED50 doses resulted in weight loss of

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A497 OBESITY—ANIMALCATEGORY

28% vs. vehicle after 28 days, far exceeding the sum of effects of PYY and 1948-P exendin-4 when administered alone (4.6 and 8.8%, sum = 13.4%). Cumulative Potential Role of the Intestinal Lipid Messenger OEA in the Effects food intake was reduced 35% with combination treatment. Weight loss with of Sleeve Gastrectomy on Food Intake and Preference combination treatment was similar to that in pair-fed controls out to day 14 HILARY E. WILSON-PEREZ, DANIELA COTA, ISABELLE MATIAS, RANDY SEELEY, (~22%), consistent with anorectic activity of the combination being the domi- SILVANA OBICI, Cincinnati, OH, Bordeaux, France nant driver of weight loss. After day 14, combination treatment invoked ~5% Oleoylethanolamide (OEA) is a diet-derived intestinal lipid messenger that more weight loss than in pair-fed animals. An increase in energy expenditure promotes satiety by acting on peripheral sensors located in the gut. We have in the combination group was confi rmed by elevated O2 consumption. Fur- found that the weight-reducing bariatric procedure Vertical Sleeve Gast- ther, combination therapy reverted multiple biomarkers (e.g. glucose, HbA1c, rectomy (VSG) increases OEA levels in the duodenum (0.48+- 0.02 versus cholesterol, triglycerides, alanine aminotransferase, aspartate aminotrans- 0.75+- 0.09, sham and VSG mice respectively N=6-7 p<0.05) in high-fat diet- ferase) back to lean control values after 28 days. Metabolomic profi ling of induced obese mice, without signifi cant changes in ileum, jejunum or plasma the serum showed a pattern of changes with combination treatment distinct levels. We have previously published that VSG results in a marked weight- from those observed with individual peptide treatments, with a major ef- independent reduction in the secretion of intestinal triglycerides. Thus, the fect on energy metabolism and especially fatty acid beta-oxidation; these increased levels of duodenal OEA occur despite a lower dietary supply of changes were not seen in pair-fed controls. fatty acid precursors.OEA has been shown to increase meal frequency, and has been suggested to play a role in fat intake. We tested the effect of VSG on food intake, food preference and meal pattern. VSG mice have sig- nifi cantly reduced body weight and fat mass, but display daily calorie intake similar to sham operated mice following a transient hypophagia in the fi rst 3 post-operative weeks. Despite normal daily calorie intake, meal frequency was increased in VSG compared to Sham mice. Using a macronutrient selec- tion paradigm to examine food choice, we found that VSG-operated mice decrease their fat intake and increase their carbohydrate intake compared to sham-operated controls.The changes in food intake, meal pattern and food preferences in VSG mice reproduce several of the anorectic effects of OEA. Taken together these results point to OEA as a potential intestinal mediator of the VSG-induced effects on food intake and food preference.

1949-P GM3 Synthase Depletion Ameliorates Glucose Tolerance in Obesity Related Type 2 Diabetes Mice SUSUMU IMAZU, MASAKI IIZUKA, MASAKAZU NAGAFUKU, JIN-ICHI IN- OKUCHI, JEFFREY ENCINAS, Kobe, Japan, Sendai, Japan 1947-P Ganglioside GM3, a sialic acid-containing glycosphingolipid, is abundantly localized in the plasma membrane, especially in caveolae and glycolipid- Molecular Insights of Improved Cholesterol Metabolism Following enriched membrane microdomains (GEMs). It has been reported that insulin Vertical Sleeve Gastrectomy in Diet Induced Obese Rats resistance induced by TNF_ in 3T3-L1 cells is accompanied by an increase in ZHIBO AN, ERIC P. SMITH, RON BITNER, DREW KAISER, HELENA GYLLING, HI- GM3 synthase (GM3S) expression and that plasma GM3 levels are elevated LARY WILSON-PEREZ, RANDY J. SEELEY, DAVID A. D’ALESSIO, Cincinnati, OH, in diabetic subjects. Insulin receptors (IRs) are normally resident in caveloae Kuopio, Finland in association with caveolin-1 (Cav-1). Increased GM3, however, causes dis- Vertical sleeve gastrectomy (VSG), a new bariatric surgical procedure in sociation of IR from Cav1, shifting IRs from caveolae to GEMs and suppress- which the majority of the stomach is removed while leaving the intestine ing insulin signaling. Control of GM3 production may therefore be a potential intact, is gaining in popularity. This study was designed to determine the therapy for insulin resistance and diabetes.The aim of this study was to ex- effect of VSG on lipid metabolism, and to exam the molecular mechanisms amine the effects of loss of GM3S on metabolic control in a type 2 diabetes involved. Blood samples and tissue biopsies were obtained at 6 months after (T2D) mouse model. For this purpose, we generated GM3S defi cient mice on the VSG or sham surgery (n=7 per group) following fasting-refeeding. Total KKAy and KK backgrounds: GM3S(-/-)KKAy and GM3S(-/-)KK, respectively. fat absorption and oral fat tolerance were measured 1 week before sacrifi ce. KKAy mice develop spontaneous symptoms of T2D and express higher levels Plasma markers of cholesterol absorption (campesterol, sitosterol, avenas- of GM3S mRNA in adipose tissue than KK mice. GM3S(-/-)KKAy mice, on the terol, and cholestanol) and synthesis (cholestenol, desmosterol, squalene, other hand, showed better metabolic parameters. Body weight (BW) gain in and lathosterol) were measured by gas-liquid chromatography. mRNA ex- GM3S(-/-)KKAy mice was slower compared to KKAy, reaching 40.7 ± 2.3 g at pression levels of genes related to cholesterol absorption, including AbcA1, 18-weeks, versus 47.3 ± 1.9 g in KKAy (P<0.01), unlike a known report that the apoA-IV, apoE, and SCARB1, cholesterol synthesis, including HMG-CoA BW in GM3S(-/-)C57BL/6 diet induced obese (DIO) mice was not signifi cant reductase, HMG-CoA synthase, mevalonate kinase and squalene synthase, difference from that in wild type DIO mice. This suppression of BW gain and genes pertaining to bile metabolism, including AbcG5, AbcG8, Cyp7a1 may be a result of their reduced food intake, which at 18-weeks was 5.2 ± and Cyp7b1 were determined. VSG improved fat tolerance, while fat absorp- 0.6 g/day, versus 7.2 ± 0.8 g/day in KKAy (P<0.001). Glucose tolerance was tion and cholesterol absorption was unchanged at 6 months following VSG. also dramatically improved in GM3S(-/-)KKAy, to a level similar to that in KK Plasma markers of cholesterol synthesis were reduced by VSG by 14-34% vs. mice. Furthermore, while KKAy mice show plasma insulin levels that are half sham, and mRNA expression of genes related to cholesterol synthesis was of those in KK, those in GM3S(-/-)KKAy were similar to KK, suggesting that signifi cantly reduced. The increased expression of hepatic AbcG5, AbcG8 pancreatic islets are preserved by GM3S defi ciency. These data suggest that indicated activated biliary secretion of liver cholesterol, and the increased GM3S plays a key role in the development of obese-related T2D and can be a expression of Cyp7a1, Cyp7b1 indicated stimulated hepatic bile synthesis.

Obesity target for therapeutic approaches. Our data suggest that the improved cholesterol metabolism after VSG is re- POSTERS lated to reduced intestinal cholesterol synthesis, enhanced hepatic bile pro- duction and increased hepato-biliary cholesterol transport, thus providing 1950-P Integrated Physiology/ a novel mechanistic explanation for the improved cholesterol metabolism Excess Fatty Acid Supply Alters White Adipose Progenitor Cells following VSG. Proliferation and Differentiation in Subcutaneous Fat Depot Supported by: NIH (R01 DK57900), EES Research Grant DONYA MOHEBALI, YONG HWAN HAN, SHAOBO PEI, SEYRAN SABER, SIHEM BOUDINA, Salt Lake City, UT Obesity is considered a growing epidemic in the developed countries and is increasing at an alarming rate due to excess caloric intake and reduced physical activity. White adipose tissue (WAT) adapts to nutrient supply by increasing both adipocyte size and number and it has been shown recently that adipose progenitor cell expansion can occur in adults. However, it is still unknown whether dietary lipids regulate the proliferation and differentiation of WAT progenitor cells. Using an in-vitro approach, we isolated WAT pro-

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A498 OBESITY—ANIMALCATEGORY genitors from subcutaneous or visceral fat depots of C57B6 mice using fl ow (20%) than that of age-matched established diet-induced-obesity (eDIO) cytometry and incubated them in the presence of 250 μM palmitate or BSA mice fed with a high fat diet (HFD) (N=6/group, male C57 mice). Referencing control for 6 or 24 hours. In parallel, male C57B6 mice were fed either high to the methyl hydrogen peak, the average numbers of double bonds were fat diet (HFD) consisting of 45% fat or low fat diet, containing 10% fat, for 5 determined to be 1.27±0.03 and 1.02±0.05 (p<0.004) per FA chain for the weeks and WAT progenitors from subcutaneous and visceral fat depots were lean and DIO mice respectively. Whole body fat 13C-NMR spectra obtained isolated and cultured. Differentiation, proliferation and cell cycle distribution in mice also showed a higher unsaturated fat content globally in low fat were determined. Both palmitate and high fat feeding reduced WAT progeni- fed mice than that of high fat. In conclusions, we validated in vivo methods tor’s proliferation that was more pronounced in thesubcutaneous versus the for quantifying level of unsaturated body fat locally and globally in oil stan- visceral fat depot. Interestingly, palmitate treatment increased G2/M cell cy- dards. Using these methods, we assessed unsaturated body fat fractions cle arrest whereas HFD blocked the progression to S-phasein the subcutane- in C57 mice under different dietary conditions. It demonstrated that chow ous depot only. Furthermore, both palmitate treatment and HFD reduced adi- fed mice maintain a signifi cant high level of unsaturated body fat level in pose progenitor cells differentiation in the subcutaneous depot as evidenced visceral adipose tissue and whole body. And, unsaturated fraction of body by the reduction in oil red o staining and a blunted induction of differentiation fat can be modifi ed with a high saturated fat diet. genes. Finally, palmitate treatment was associated with an early increase in superoxide generation in the progenitor cells of subcutaneous fat depot. 1953-P Together these results suggest that excess fatty acid supply specifi cally alter RAGE Defi ciency in Bone Marrow-Derived Cells Imparts Partial the proliferation and differentiation of WAT progenitor in the subcutaneous Protection against High Fat Diet-Induced Insulin Resistance and depot via mechanisms associated with increased oxidative stress. Obesity Supported by: AHA and NIH FEI SONG, ROSA ROSARIO, RAVICHANDRAN RAMASAMY, ANTHONY FERRAN- TE, ANN MARIE SCHMIDT, New York, NY 1951-P We demonstrated that global deletion of the receptor for AGE (RAGE) A Natural Compound Protects against Diet-Induced Metabolic Dis- is protective against high fat diet (HFD)-induced obesity (DIO) and insu- order by Activation of AMPK and by Modulation of PGC-1ƴ Regu- lin resistance. RAGE is expressed in infl ammatory cells and plays roles in lated Mitochondrial Respiration and Hepatic Gluconeogenesis macrophage activation. After three months on HFD, RAGE defi cient mice PASHA APONTES, XISONG LI, RADHIKA H. MUZUMDAR, LINDA A. JELICKS, demonstrated signifi cantly less CD11c-expressing macrophages in adipose BHAVAPRIYA VAITHEESVARAN, IRWIN J. KURLAND, ANTHONY A. SAUVE, YUL- tissue vs. littermate mice fed HFD. To determine if bone marrow RAGE ex- ING CHI, Bronx, NY, New York, NY pression contributes to DIO, we performed lethal irradiation of wild-type Obesity has become epidemic in the developed nations of the world. It (WT) mice followed by transplantation of RAGE null vs. WT bone marrow. produces a cluster of interrelated pathological conditions including diabetes, One month after lethal irradiation/bone marrow transplantation, mice were liver steatosis, cardiovascular disorders and kidney dysfunctions. Although fed low fat diet chow (LFD) vs. high fat (60% calories by fat) diet (HFD) (8-9 genetics can predispose individuals to obesity, a combination of sedentary mice/group). In mice fed LFD, introduction of RAGE defi cient bone mar- lifestyle and excessive calorie intake are the major contributors. Diffi cul- row had no impact on body weight or glucose tolerance at 3 weeks or 3 ty in management of lifestyle and diet suggests that strategies that can months (Fig. 1-4). After 3 weeks HFD, mice receiving RAGE defi cient bone stimulate selective nutrient utilization and limit glucose production could be marrow displayed signifi cantly better glucose tolerance vs. recipients of effi cacious approaches to treating obesity associated metabolic disorders. WT bone marrow (Fig. 2) which was not sustained at 3 months HFD (Fig. We screened a small library of natural compounds and identifi ed YCOB01 4). Although there were no differences in body weight of HFD-fed mice that could reduce high fat diet (HFD)-induced obesity in mice. Calorimetry at 3 weeks, at 3 months, mice receiving RAGE null bone marrow weighed revealed that YCOB01 increased locomotor activity and enhanced energy ex- signifi cantly less than mice receiving WT bone marrow (Figs. 1&3). These penditure (EE). Using implanted osmotic pumps, we determined that YCOB01 results suggest that the protection of global RAGE defi ciency in preven- reduced hepatic glucose production and recycling. Increased EE and reduced tion of DIO and insulin resistance is accounted for, in part, by RAGE ex- hepatic gluconeogenesis resulted in signifi cantly improved glucose homeo- pression in bone marrow. These studies will be extended to determine if stasis. YCOB01 reversed HFD caused hyperglycemia and glucose intoler- the impact of bone marrow RAGE expression on body weight is sustained. ance. A euglycemic clamp study determined that YCOB01 increased glucose infusion rate in HFD-fed mice by 7-fold, a consequence of increased insulin sensitivity. Mechanistically, YCOB01 induced phosphorylation of AMPK in muscle and liver tissues of treated mice. YCOB01 activation of AMPK was also found in cultured C2C12 cells and hepatocytes. In muscle tissues and C2C12 cells YCOB01 induced PGC-1_ and mitochondrial respiration genes. Conversely, in hepatic tissues and cells, the compound suppressed PGC-1_ and gluconeogenesis genes. In summary, YCOB01 prevents HFD induced obesity and corrects glucose homeostasis by activating the energy sensor, AMPK, and differentially modulating PGC-1_ in muscle (activation) and in liver (suppression). Supported by: AHA

1952-P In Vivo Evaluation of Saturated and Unsaturated Body Fat Contents Locally and Globally HAIYING LIU, DAN ZHOU, CHUNLIAN ZHANG, Rahway, NJ The Objectives of the study are: 1) to evaluate the feasibility of non-inva-

sively measuring saturated and unsaturated body fat contents locally and Obesity globally in vivo; 2) to investigate whether diet infl uences the level of unsatu- POSTERS rated fatty acids (FA) in body fat. The saturated and unsaturated fractions 1954-P

of local and whole body fat were measured using magnetic resonance spec- Anti-Obesity Effects of KR-66195, a Synthetic DPP-IV Inhibitor, in Integrated Physiology/ troscopy (MRS) in two groups of mice fed with either a low or high fat diet. Diet-Induced Obese Mice and Obese-Diabetic ob/ob Mice The fat composition measurements of adipose tissue and whole body were EUN YOUNG LEE, YEON WOOK KIM, SUN OK SONG, HANNA SEOK, EUN YOUNG done using proton and carbon-13 NMR respectively at 9.4 or 11-Tesla under CHOI, BYUNG-WAN LEE, EUN SEOK KANG, CHUL WOO AHN, BONG SOO CHA, gas anesthesia (1.0% isofl uorane) and the resulting MR spectra revealed HYUN CHUL LEE, Seoul, Republic of Korea peaks corresponding to both unsaturated and saturated FA components. A variety of dipeptidyl peptidase (DPP)-IV inhibitors improve glycemic Spectral peaks from hydrogen or carbon attached to carbons with double control in patients with type 2 diabetes by increasing plasma level of active bonds were identifi ed and quantifi ed against a reference peak. The meth- glucagon like peptide (GLP)-1. However, unlike GLP-1 analogues, DPP-IV in- ods were validated using oil standards whose saturated, mono and poly- hibitors failed to inhibit body weight gain. In this study, we found KR-66195, unsaturated fat compositions are known. The unsaturated fraction of body a new DPP-IV inhibitor, could prevent weight gain as well as improve gly- triglyceride (TG) in chow-fed lean mice was found to be signifi cantly higher cemic control in both diet-induced obese (DIO) C57BL/6 mice and obese-

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A499 OBESITY—ANIMALCATEGORY

diabetic ob/ob mice. In DIO mice, intraperitoneal administration of KR-66195 Importantly, at this early time point, there was a transient increase in fed at 10mg/kg/day for 8 weeks resulted in increase in the plasma level of (147±2.6 vs. 122±1.9 mg/dl, p<0.01) and fasting hyperglycemia (122±6.1 vs. GLP-1 and improvement of glucose tolerance. This treatment also reduced 95.8±7.0 mg/dl, p<0.05), together with a marginal increase in weight gain, body weight gain (5.38±1.89g vs. 12.08±1.36g), epididymal fat accumulation mild hyperinsulinemia, glucose intolerance and impaired insulin secretion. (0.86±0.01g vs. 2.30±0.07g) and food intake (2.41±0.09g vs. 2.79±0.11g). To By 2 weeks, hyperglycemia (136±1.96 mg/dl vs. baseline) and levels of tnf-_ investigate the mechanisms of the anti-obesity effects of the KR-66195, we and Il-1` (44% and 68% respectively) were reduced in HFD mice with a sig- performed a pair-feeding study in DIO mice. Mice were housed in metabolic nifi cant up-regulation of islet growth, survival, insulin signaling and glucose cages for 4 weeks. The pair-fed group showed a 10% decrease in body heat sensing genes. At a later time point of 8 weeks, despite worsening of fasting (kcal/kg/hr) with no differences in body weight and activity. These results hyperglycemia and enhanced levels of tnf-_ in the hypothalamus (3.6-fold, revealed that the KR-66195 caused weight loss in DIO mice by modulating p<0.01) and epididymal fat (17-fold, p<0.05) that were coincident with severe food intake and energy expenditure. In ob/ob mice, the same administration insulin resistance, `-cell compensatory responses were maintained in the of KR-66195 for 3 weeks resulted in the comparable effects. In conclusion, HFD mice. These results show that hypothalamic infl ammation is transient the KR-66195 might be further developed as therapeutic drug to treat obe- and manifests early during HFD, well before whole body insulin resistance sity as well as diabetes. and BCM increases suggesting an important role in the induction of the `- cell adaptation process. 1955-P Serum FetuinA Levels Related With Microalbuminuria in Diet-in- 1957-P duced Obese Rats Effects of Resveratrol on Neuroinfl ammation and Memory Defi cit in YANYAN LI, XIAODONG SUN, YERONG YU, Chengdu, China High Fat Diet-Induced Obesity Mice FetuinA, an abundant serum protein predominantly synthesized and se- BYEONG TAK JEON, SOO KYOUNG KIM, TAE SIK JUNG, DONG HOON LEE, HYUN creted by liver, is deemed to repress the production of adiponectin which JOON KIM, SANG SOO KANG, GYEONG JAE CHO, WAN SUNG CHOI, GU SEOB has a strong inverse correlation with microalbuminuria (MAU) in obese sub- ROH, Jinju, Republic of Korea jects. We hypothesized that the elevated serum fetuinA levels in obesity Obesity and Type 2 diabetes mellitus (T2DM) are associated with chronic may related with increased urine albumin excretion,and reducing fetuinA infl ammation and are considered to risk factors for neurodegeneration. Obe- secretion may have renal protective effects by elevating adiponectin levels. sity-induced peripheral infl ammation and oxidative stress may be closely as- Male Wistar rats 4 weeks in age were randomly divided into three groups sociated with neuroinfl ammation in the central nervous system. Resveratrol and fed with normal chow (control group), high-fat chow (obesity group) or (RES) has antioxidant effect and improves insulin sensitivity. In the pres- high-fat chow plus fenofi brate (fenofi brate group). After 24 weeks feeding, ent study, we tested the hypothesis that an AMP-activated protein kinase the obese rats showed elevated serum free fatty acids (FFAs) concentra- activator, RES, which is known to exert potent anti-infl ammatory effects, tion, higher liver triglyceride (LTG) content, and increased ALT and AST levels would attenuate neuroinfl ammation and improve memory defi cit in high-fat when compared with controls. Serum fetuinA was increased and adiponec- diet (HFD)-fed mice. C57BL/6J mice were fed a HFD or a HFD supplemented tin was decreased in obese rats in comparison to controls. Fenofi brate in- with RES for 20 weeks. Metabolic parameters in serum were evaluated, and tervention reduced the levels of serum FFAs, LTG, ALT/ AST, and fetuinA, western blot analysis and immunohistochemistry in the peripheral organs while increased adiponectin levels. Urinary albumin/creatinine ratio (ACR) and hippocampus were completed. In addition, we used the Morris water was increased in obese rats, which was decreased after fenofi brate inter- maze test to study the role of RES on memory function in HFD-treated mice. vention (table 1). Correlation analysis showed ACR had a positive correlation RES treatment reduced hepatic steatosis, macrophage infi ltration, and in- with FFAs, LTG ,ALT, AST, and fetuinA(r=0.839, 0.715, 0.727, 0.742, 0.602, re- sulin resistance in HFD-fed mice. In the hippocampus of HFD-fed mice, the spectively, P<0.01) and had a negative correlation with adiponectin(r=-0.635, protein levels of tumor necrosis factor-_ and Iba1 expression were reduced P<0.01). We conclude that higher circulating fetuinA concentrations may in- by RES treatment. The phosphorylation of insulin receptor, adenosine mono- crease MAU in diet-induced obese rats and the mechanism may related with phosphate-activated protein kinase (AMPK), acetyl-CoA carboxylase (ACC), repressing adiponectin production. and glycogen synthase kinase (GSK)-3` in the hippocampus of HFD-fed mice Table 1. Serum parameters and urine ACR in three groups were not reduced by RES treatment. In addition, dietary RES reduced HFD- induced phosphorylation of Tau in the hippocampus. Choline acetyltrans- ferase was unchanged and the phosphorylation of tau was increased in the Groups Control Obesity Fenofi brate P value hippocampus of HFD-fed mice upon RES treatment. In particular, we found FFAs (mmol/L) 0.37±0.12 1.45±0.33 0.78±0.11 <0.001 that RES signifi cantly improved memory defi cit in HFD-fed mice. These data LTG (μmol/g) 110.9±49.46 258.8±127.6 192.4±77.9 <0.05 suggest that chronic dietary RES exerts as anti-infl ammatory effects and provides adjunctive therapy for type 2 diabetes and neurodegenerative dis- ALT (IU/L) 48.5±13.7 83.6±39.5 51.5±12.6 <0.05 eases such as Alzheimer’s disease. AST (IU/L) 124±32.4 222±24.3 108±13.8 <0.001 Supported by: NRF (No. 2011-0006196) FetuinA (μg/L) 86.2±18.0 130.9±22.4 87.9±27.7 <0.01 Adiponectin (μg/L) 272.3±56.7 93.7±47.5 228.7±104.4 <0.01 1958-P ACR (mg/g) 19.6±7.8 54.5±23.3 20.7±6.7 <0.01 Genetic Ablation of Cytokine Signaling Prevents Diet-Induced Obe- sity and Insulin Resistance in Lymphocyte-Defi cient NOD-SCID Mice Supported by: National Nature Science Foundation of China (Grant number DAE YOUNG JUNG, RITA BORTELL, HWI JIN KO, RANDALL H. FRIEDLINE, 81070675). YONGJIN LEE, PAYAL R. PATEL, SEZIN DAGDEVIREN, HSUN-FAN WANG, NICHO- LAS TSITSILIANOS, UMBER SHAFIQ, YOSHIHIRO AZUMA, XIAODI HU, GREGORY 1956-P WU, ADEOLA O. OTUYELU, LEONARD D. SHULTZ, DALE L. GREINER, JASON K. Hypothalamic Infl ammation Precedes Beta-Cell Adaptation during KIM, Worcester, MA, Bar Harbor, ME Short-Term High Fat Diet Nonobese diabetic (NOD) mice and lymphocyte-defi cient mice with se-

Obesity MINA PESHAVARIA, DHANANJAY GUPTA, THOMAS L. JETTON, JACK L. LEAHY, vere combined immunodefi ciency (SCID) mutation are widely used for auto-

POSTERS Colchester, VT immune type 1 diabetes and islet graft studies. While their immunological In diet-induced obesity, `-cells adapt to a progressive increase in pe- profi les are well characterized, their metabolic phenotypes are poorly un-

Integrated Physiology/ ripheral insulin resistance by hypersecretion of insulin and enhanced `-cell derstood. Here we examined the effects of high-fat diet (HFD) on insulin mass (BCM). Hypothalamic infl ammation, in response to high fat diet (HFD) sensitivity in normoglycemic NOD and NOD-SCID mice. During 4 wks of HFD, in rodents also promotes insulin and leptin resistance in addition to weight NOD and NOD-SCID mice became obese with less whole body fat mass in gain. However, the role of hypothalamic infl ammation with respect to `-cell NOD-SCID mice (Fig. 1). A hyperinsulinemic-euglycemicclamp study was per- adaptation remains unclear. In the present study, we evaluated the effect formed in awake mice (n=6-8/group) and showed that both groups of mice of short (1, 2 week) and long-term (8 week) HFD (60% energy from fat) on were insulin sensitive on chow diet. Following HFD, NOD and NOD-SCID hypothalamic infl ammation and `-cell adaptation in C57Bl/6 mice. Within mice developed insulin resistance with signifi cant reductions in glucose 1 week, proinfl ammatory gene expression levels of tnf-_ (1.53-fold, p<0.01) infusion rates (Fig. 2; *P<0.05vs. Chow Diet). This was due to 20~40% reduc- and Il-1` (2.14-fold, p<0.05) were increased in HFD mice. In contrast, lev- tions in whole body glucose turnover and hepatic insulin action (Fig. 3 & 4). els of islet genes associated with `-cell adaptation (ins1, pdx1, ppar-a) and Thus, NOD and NOD-SCID mice developed diet-induced obesity and insulin insulin secretion (Slc2a2, glp1r Gck) remained comparable to chow mice. resistance. Furthermore, we examined the effects of HFD in lymphocyte-

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A500 OBESITY—HUMANCATEGORY free NOD-SCID mice with deletion of the IL2 receptor common gamma (aC) chain (NSG), which mediates cytokine signaling. During 4 wks of HFD, NSG & 1960-P mice were highly resistant to diet-induced obesity (Fig. 1), and remained Effects of Free Fatty Acid Availability on Adipose Tissue Fatty Acid more insulin sensitive than NOD and NOD-SCID mice. Overall, our fi ndings Storage indicate that attenuating cytokine signaling rescues NOD-SCID mice from MANPREET S. MUNDI, CHRISTINA KOUTSARI, MICHAEL D. JENSEN, Rochester, diet-induced obesity and further implicate an important role of cytokine net- MN work in obesity-associated insulin resistance. Women have greater free fatty acid (FFA) fl ux than men and greater di- rect FFA storage into upper (UBSQ) and lower body subcutaneous (LBSQ) fat. Greater FFA storage rates in women might be related to greater FFA fl ux or more body fat than men. We tested these hypotheses by measuring direct adipose tissue FFA storage rates in obese (n=8) and non-obese men (n=8) under conditions of increased FFA availability using a lipid emulsion infu- sion to create FFA fl ux levels seen in 22 postabsorptive non-obese women studied using a similar protocol but without a lipid emulsion infusion. Direct FFA storage (μmol · kg fat-1 · min-1) in UBSQ and LBSQ fat was assessed us- ing stable isotope tracer infusions (both [U-13C] oleate and [U-13C]linoleate) to measure FFA rate of disappearance and an IV [1-14C]oleate bolus/timed biopsy approach to measure adipose FFA storage. Fractional FFA storage into the UBSQ and LBSQ depots was multiplied by FFA disappearance to cal- culate regional storage rates. Body composition was assessed using a DXA and abdominal CT scan. FFA fl ux was similar in all three groups by design, however, FFA storage in women was greater in both UBSQ and LBSQ adi- pose tissue when compared to obese men. FFA storage in women was also greater when compared to non-obese men in LBSQ with a trend towards being greater in UBSQ adipose tissue (p=0.07). There was no difference in FFA storage per kg fat in either depot between obese and non-obese men. Supported by: NIH R01-DK080756 In conclusion, the greater FFA adipose tissue storage rates found in women cannot be explained by greater FFA availability or more body fat, suggesting inherent adipocyte differences. Table 1. FFA fl ux and storage rates in subcutaneous adipose tissue depots OBESITY—HUMAN Obese Men Non-Obese Non-Obese Men Women Guided Audio Tour: Human Obesity (Posters 1959-P to 1966-P), see page n=8 n=8 n=22 13. FFA fl ux 423 ± 85 456 ± 145 395 ± 118 FFA Concentration 565 ± 133* 530 ± 145 423 ± 98 & 1959-P UBSQ FFA storage 0.67 ± 0.21* 1.02 ± 0.43 1.58 ± 0.73 Regulation of Thrombospondin (TSP-1) in Adipocytes and Mac- LBSQ FFA storage 0.74 ± 0.55* 1.01 ± 0.32** 1.59 ± 0.56 rophages by Coculture and Docosahexaenoic Acid (DHA) Values are means ± SD; fl ux rates are μmol · min-1 and direct FFA storage rates BRIAN S. FINLIN, BEIBEI ZHU, CATHERINE P. STARNES, CHARLOTTE A. PETER- are μmol · kg fat-1 · min-1.UBSQ, upper-body subcutaneous; LBSQ, lower-body SON, PHILIP A. KERN, Lexington, KY Adipose tissue dysfunction with obesity includes both infl ammation and subcutaneous; * P <0.01 vs. non-obese women. **P<0.05 vs. non-obese women. fi brosis. In recent work from our lab, obese adipose tissue was associated Supported by: NIH with an increase in both infl ammatory (M1) and anti-infl ammatory (M2) macrophages, which contribute to fi brosis. Our objective was to understand & 1961-P whether fatty acids, including the t-3 fatty acid DHA, affect fi brosis path- Low Relative Skeletal Muscle Mass is Independently Associated ways during coculture of macrophages and adipocytesHuman adipocytes With Arterial Stiffness in Patients With Type 2 Diabetes were prepared from stem cells, along with human THP-1 macrophages that YOUNGJU CHOI, SOO-KYNG KIM, SEOK W. PARK, CHUL S. KIM, HEE Y. CHO, were differentiated to either M1or M2 phenotypes. The coculture of adi- SUN-HA JEE, EUN-JIG LEE, KAP B. HUH, Seoul, Republic of Korea, Seongnam, pocytes with M1 or M2c macrophages induced the expression of TSP-1, a Republic of Korea, Anyang, Republic of Korea key regulator of fi brosis and angiogenesis. Coculture induced TSP-1 gene ex- Diabetic patients tend to have age-related skeletal muscle loss (sarcope- pression in both adipocytes (5.5-fold, P<.05) and M1 or M2c macrophages (> nia), which is associated with an increased risk of adverse health outcomes. 30-fold, P<.05) and induced TSP-1 protein secretion into the media (>10 fold, Basal leg blood fl ow declines with aging, thus arterial stiffness may have an P<.05). DHA treatment during the coculture potently inhibited both the mac- independent role on sarcopenia. In this study, we investigated whether skel- rophage TSP-1 mRNA level (85% inhibition, P<.05) and the level of TSP-1 pro- etal muscle mass was associated with arterial stiffness in type 2 diabetic tein secreted into the media (>70% inhibition, P<.05). TSP-1 activates TGF-`, subjects. We enrolled 3595 patients (mean age 57±12 yr, men 46%) with and CTGF, a target of TGF-`, displayed a similar expression pattern to TSP-1. type 2 diabetes, who visited Huh’s Diabetes Center from 2004 to 2009. The We explored the mechanism of inhibition of TSP1 in M2c macrophages and skeletal muscle mass was estimated from bioimpedance analysis measure- found that DHA did not affect the differentiation state. However, IL-10 ex- ments and expressed as skeletal muscle mass index (SMI = skeletal muscle pression in M2c macrophages displayed a similar pattern to TSP-1: induced mass/body mass x 100). We measured brachial-ankle PWV(baPWV) using by coculture and inhibited by DHA. Since IL-10 has been shown to regulate an automatic device, VP-1000(Colin Co. Ltd, Komaki, Japan). The SMI was Obesity

TSP-1 in other cell types, we reduced IL-10 expression with siRNA in the M2c negatively correlated with age (r=-0.286, p<0.01), total cholesterol (-r=0.115, POSTERS cells and found that this caused TSP-1 to be reduced in response to adipo- p<0.01), systolic blood pressure (r=-0.223, p<0.01), waist circumference cyte coculture by 60% (P<.05).Thus, these data show that DHA reduced TSP1

(-r=0.055, p<0.05), mean PWV (-r=0.227, p<0.01), and positively correlated Integrated Physiology/ in adipocytes and macrophages, likely through a decrease in IL-10, resulting with HDL-cholesterol (r=0.074, p<0.01). In multiple regression model, the in decreased TGF-` signaling. These results suggest that supplementation SMI was independently associated with mean PWV after adjusting for age, with dietary t-3 fatty acids could have an unappreciated benefi t of reducing smoking history, glucose, triglyceride, HDL cholesterol, waist circumference fi brosis in human adipose tissue. and systolic blood pressure in men (`=-0.084, p<0.05). But the independent Supported by: R01 DK80327 association was not observed in women.In conclusion, the low relative skeletal mass indicative of sarcopenia is associated with arterial stiffness in male type 2 diabetic patients. Interventions leading to skeletal muscle hypertrophy such as resistance training and/or amino acid supplementation may need to be tested in diabetic patients with high baPWV.

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A501 OBESITY—HUMANCATEGORY

& 1962-P & 1964-P Copy Number Variation at a Chromosome 1 CNV Hotspot Strongly No Change in Insulin Sensitivity 2 Months After Sleeve Gastrectomy Contributes to the Risk of Obesity or Low-Calorie Diet in Obese Individuals JULIA SARAH EL-SAYED MOUSTAFA, PETROS TAKOUSIS, FRANCESCO PESCE, CHARMAINE TAM, LEANNE REDMAN, FRANK GREENWAY, ERIC RAVUSSIN, AMÉLIE BONNEFOND, JOHANNA C. ANDERSSON-ASSARSSON, MASHAEL Baton Rouge, LA NEDHAM ALSHAFAI, LENA M. CARLSSON, FRANCOIS PATTOU, TIMOTHY D. Sleeve gastrectomy has evolved to become a primary bariatric procedure. SPECTOR, PHILIPPE FROGUEL, MARIO FALCHI, London, United Kingdom, Lille, Despite this, few studies have described the effects of this procedure on France, Gothenburg, Sweden insulin sensitivity.Seventeen morbidly obese subjects (BMI:49.5±9.9kg/m2) Although the heritability of obesity is estimated at up to 70%, much of were observed at baseline and 7 weeks after sleeve gastrectomy (SG) (n=9, the genetic variance remains unaccounted for, despite recent GWAS fi nd- 7 females, 42±13y) or an 800kcal/d low-calorie diet (LCD) (n=8, 7 females, ings. A proportion of the heritability of obesity may be accounted for by 48±2y). We examined changes in insulin sensitivity (glucose disposal rate) multiallelic copy-number variants (CNVs), whose effects have not been thor- with a two-step hyperinsulinemic clamp (glucose concentration=120mg/dL) oughly investigated to date.We developed famCNV to carry out association and glucose and insulin responsiveness to a standardized breakfast (400kcal). between CNVs and quantitative traits in families directly using the continu- After 2 months, SG subjects had lost 10% of their body weight (143±25 to ous distribution of the intensity signals from GWAS arrays. We used the 130±31kg, P=0.11) compared to 6% weight loss (137±33 to 128±36kg, P=0.04) program to identify copy-number variants in dosage sensitive genes infl u- in the LCD group. Both fasting glucose and insulin levels were signifi cantly encing transcriptional levels in subcutaneous adipose tissue from a Swed- decreased in SG (Glucose: 113±13 to 91±7mg/dL, P=0.003; Insulin:22±13 to ish family sample (N = 342) ascertained for obesity. Integration of these 11±6μU/mL, P=0.007) and LCD (Glucose:119±35 to 109±31mg/dL, P=0.02; data with obesity phenotypes identifi ed a CNV hotspot on chromosome 1 Insulin: 29±27 to 24±25μU/mL, P=0.008) groups. However, there were no strongly involved in obesity susceptibility, and this fi nding was replicated signifi cant changes in glucose disposal rate adjusted for metabolic body size in fi ve different cohorts including a total of 6,000 subjects. Low versus high (fat-free mass +17.7kg) in either SG or LCD subjects. The insulin responsive- copy-number carriers showed high risk of obesity (OR = 8.9; 95% confi dence ness to the breakfast (6Insulin0min-30min /6Glucose0min-30min) was however de- interval = 4.77-16.59; P = 6.3 x 10-12). The number of gene copies at this locus creased in the SG (P=0.05), but not LCD (P=0.67) group. Insulin secretion, as was subsequently determined by qPCR for 1,300 subjects for validation and measured by HOMA-B, was unchanged by SG (149.5 ± 82.7 to 150.3 ± 81.7%; in-depth characterisation. We also measured levels of the protein encoded P=0.24) or LCD (168.2 ±95.8 to 160.9 ±145.5%; P=0.49).Our results show that by this gene in blood from 119 morbid obese subjects, and showed signifi - peripheral and hepatic insulin sensitivity or `-cell function is unchanged 2 cant inverse association between circulating protein levels and body mass months after sleeve gastrectomy or LCD, despite markedly decreased glu- index (P = 6.0 × 10-3).The CNV at this locus may explain more than 17% of cose and insulin levels.1. Confl ict of Interest: None2. Funding: Research re- the heritability of obesity, thus confi rming the power of integrative genetic lated to this abstract was funded by Ethicon-Endo Surgery. Inc approaches for the identifi cation of novel frequent genomic variants strongly Supported by: Ethicon Surgery contributing to metabolic diseases. & 1965-P & 1963-P Exenatide is Associated With Weight Loss and an Increase in FGF21 Ethnic Differences in Weight Loss and Diabetes Resolution After Levels in Obese, Non-Diabetic Women Bariatric Surgery: a Meta-Analysis JODY DUSHAY, EMILIE MITTEN, ELEFTHERIA MARATOS FLIER, Boston, MA WANDA ADMIRAAL, FUNDA CELIK, JOOST HOEKSTRA, VICTOR GERDES, FRITS The mechanisms of weight loss with glucagon-like peptide-1 receptor HOLLEMAN, Amsterdam, The Netherlands (GLP-1R) agonists are not fully understood but may include changes in en- It has been postulated that the effectiveness of bariatric surgery varies ergy expenditure (EE). We investigated the effect of exenatide on weight between ethnic groups. However, data concerning this topic are inconclu- loss and metabolic parameters in obese, non-diabetic women in a ran- sive, as most studies included few patients from minority groups or did not domized, double-blind, placebo-controlled crossover study. Two 16-week fi nd statistically signifi cant differences. We conducted a meta-analysis to treatment periods were separated by a 3-week washout. Resting EE (REE) determine a difference in percentage of excess weight loss (%EWL) 1 year was measured using indirect calorimetry. Data were analyzed with a least after bariatric surgery in people from African (AF) and Caucasian (CA) de- squares means procedure. Exenatide treatment led to an average of 2.49 scent. We also studied ethnic differences in diabetes mellitus (DM) resolu- ± 0.66 kg weight loss, while no change in weight was seen with placebo tion.We performed a MEDLINE and EMBASE search for studies reporting (p = 0.01). Interestingly, subjects who lost weight with exenatide did not %EWL and/or DM resolution after bariatric surgery and including both AF show the expected reduction in REE. This suggests that preservation of REE and CA. The 646 obtained publications were reviewed. We included 14 stud- contributes to exenatide-induced weight loss. In rodents, administration of ies (1087 AF and 2714 CA); all provided data on %EWL and 3 on DM reso- fi broblast growth factor 21 (FGF21) causes weight loss and increased EE. lution. We extracted surgery type, %EWL and DM resolution 1 year after The effect of GLP-1 agonism on FGF21 levels in humans is not known. There- surgery. After analyzing %EWL for any surgery type, we performed suba- fore, we looked at serum FGF21 levels in this study population. Serum FGF21 nalyses for malabsorptive and restrictive surgery.The overall mean %EWL levels were measured using a human ELISA (R&D Systems) and analyzed difference between AF and CA was -8.36 (-10.79, -5.93) signifi cantly in favor with paired t-tests. With exenatide treatment, FGF21 levels increased sig- of CA (Figure 1). Results were similar in malabsorptive (-8.39 (-11.38, -5.40)) nifi cantly from 259 to 368 pg/mL (p = 0.01). With placebo treatment, FGF21 and restrictive (-8.46 (-12.95, -3.97)) surgery. DM resolution was higher in AF levels showed no signifi cant change (254 pg/mL at baseline and 226 pg/mL than in CA (OR 1.73 (95% CI 1.01-2.95).In %EWL terms, bariatric surgery is at the end of the treatment period, p = 0.30). To better understand the effect more effective in CA than in AF, regardless of procedure type. Our fi nding of of weight loss with exenatide versus placebo on FGF21 levels, we selected a higher DM resolution in AF, while based on limited data, suggests that DM subset of subjects who lost weight with placebo and compared the change in AF may be more determined by insulin resistance than in CA. Further stud- in FGF21 levels after * 2.5% weight loss with either treatment. In subjects ies need to clarify exact mechanisms behind ethnic differences in bariatric who lost * 2.5% weight with exenatide, FGF21 levels increased signifi cantly surgery effectiveness. from 255 to 438 pg/mL (p = 0.003). In subjects who lost a similar amount of Obesity weight with placebo, FGF21 levels decreased from 294 to 219 pg/mL (p = POSTERS 0.059). In conclusion, exenatide maintains REE in the setting of weight loss. This effect may be mediated by an increase in FGF21. Integrated Physiology/ Supported by: Amylin Pharmaceuticals

& 1966-P BMI and Development of Depression in Patients With Newly Diag- nosed Diabetes FRITHA MORRISON, MARIA SHUBINA, ALEXANDER TURCHIN, Boston, MA Both diabetes and obesity have been reported to increase the risk of de- pression. Depression co-morbid with diabetes is associated with increased mortality and morbidity. However, little is known about the relationship between BMI and development of depression among patients newly diag-

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A502 OBESITY—HUMANCATEGORY nosed with diabetes.This retrospective cohort study of 3,232 patients with (T2D: 12.5±2.8, 15.4±3.2 p=0.17, 24.6±4.4 p=0.02; NGT: 18.3±3.7, 15.5±1.7 newly diagnosed diabetes aimed to determine whether abnormal BMI is as- p=0.24, 28.2±4.8 g/kgffm/min/pmol/L p=0.01) at which time weight loss was sociated with development of depression. Patients had at least two years of 14.2±1.1% (p<0.01) in T2D and 19.4±2.3% (p<0.01) in NGT.Decreased hepatic follow-up with primary care practices affi liated with two teaching hospitals glucose production was observed 1 wk after RYGB in T2D explaining the ear- between 1/1/2000 and 1/1/2010.Comparing patients with low (< 20 kg/m2) ly reduction in fasting glucose. Peripheral insulin sensitivity did not improve vs. normal (*20 and < 25 kg/m2)vs. elevated (* 25 kg/m2) BMI, incidence prior to weight loss but was evident at 3 mo postoperatively. RYGB may have of depression decreased from 6.9% to 2.6% to 1.8%, respectively (Figure). differential effects on tissue insulin sensitivity with early improvements in Incidence of depression among patients with low BMI was higher than in hepatic insulin sensitivity and later weight-loss dependent improvements in either normal (p = 0.21) or elevated (p = 0.059) BMI groups. In multivari- peripheral insulin sensitivity. able analysis, using patients with low BMI as a reference, hazard ratio for development of depression was 0.42 (95% CI: 0.085 - 2.04, p = 0.28) and 0.24 & 1968-P (95% CI: 0.058 - 1.03, p = 0.054) for patients with normal and elevated BMI, Does Roux-en Y Gastric Bypass Surgery Have Independent Effects respectively.These fi ndings suggest that newly diagnosed patients with dia- on Ƶ-Cell Function and Insulin Sensitivity after Marked Weight betes with low BMI may be at an increased risk for depression. Contrary to Loss? other published studies, no increase in risk could be demonstrated for pa- DAVID BRADLEY, CATERINA CONTE, J. CHRISTOPHER EAGON, ESTEBAN VA- tients with elevated BMI. Further prospective research is needed to confi rm RELA, BETTINA MITTENDORFER, AMALIA GASTADELLI, BRUCE PATTERSON, these results and identify the pathophysiological mechanisms underlying SAMUEL KLEIN, St. Louis, MO, Pisa, Italy the increased risk of depression in patients with low BMI. Bariatric surgery often results in resolution of type 2 diabetes (T2D) and procedures that divert nutrients away from the upper gastrointestinal (UGI) tract are more effective than those that simply restrict stomach size. We tested the hypothesis that bariatric surgery that involves bypass of the UGI tract has important weight loss-independent effects on the two major factors involved in the pathogenesis of T2D (`-cell dysfunction and insulin resistance). We evaluated insulin sensitivity by using the hyperinsulinemic- euglycemic clamp procedure (HECP) and `-cell function by calculating the disposition index (DI) (insulin sensitivity x insulin response to mixed meal ingestion by using the minimal model) in obese subjects before laparoscopic adjustable gastric banding (LAGB; n=10; BMI: 47±3 kg/m2) or roux-en-Y gas- tric bypass (RYGB; n=10; BMI: 46±2 kg/m2), and after 20% weight loss. The stimulation of glucose disposal during insulin infusion increased after weight loss in both the LAGB (from 122±20 to 297±15%; p<0.001) and RYGB (from 136±37% to 305±59%; p<0.001) groups, but the increase was not different between groups (p=0.83). Total insulin secretion in response to the meal decreased after weight loss in both the LAGB (from 46.9 ± 8.1 to 37.2 ± 7.6 103 pmol/L per 360 min p<0.001) and RYGB (from 51.7 ± 14.8 to 39.0 ± 6.2 103 pmol/L per 360 min; p<0.001) groups, but there was no difference between groups (p=0.53). Weight loss caused a 30% increase in DI values in both the LAGB (from 1851±253 to 2604±318 μmol/kg FFM/min per 360 min; p<0.05) and the RYGB (from 1816±410 to 2543±292 μmol/kgFFM/min per 360 min; p<0.05) groups, but there was no difference between groups (p=0.72). These data demonstrate that weight loss itself is responsible for the therapeutic effects of RYGB surgery on `-cell function and insulin sensitivity in non- Supported by: 1R18HS017030, 1RC1LM010460 and DAREF diabetic obese adults, and is likely the major contributor for the therapeutic effect of RYGB on T2D. Values are mean±SEM. Supported by: NIH (DK37948, DK56341, UL1 RR024992, UL1 RR-00954), Ethicon Guided Audio Tour: Adipocyte Physiology (Posters 1967-P to 1974-P), see EndoSurgery, Inc. page 17. & 1969-P & 1967-P Resolution of Metabolic Syndrome, Diabetes Mellitus, and Hyper- Hepatic and Peripheral Insulin Sensitivity 1 Week and 3 Months tension Through Gastric Banding; Effect of Age and of Duration of after Roux-en-Y Gastric Bypass Surgery in Subjects With Type 2 Obesity Diabetes and Normal Glucose Tolerance ALBERTO MORABITO, ANTONIO E. PONTIROLI, MICHELE PAGANELLI, ALES- KIRSTINE N. BOJSEN-MØLLER, CARSTEN DIRKSEN, SIV H. JACOBSEN, NILS SANDRO SAIBENE, LUCA BUSETTO, Milano, Italy, Padova, Italy B. JØRGENSEN, ANNETTE K. SERUP, DORTE L. HANSEN, DORTE WORM, JENS Bariatric surgery leads to resolution of some co-morbidities like hyperten- J. HOLST, ERIK A. RICHTER, STEN MADSBAD, Hvidovre, Denmark, Copenhagen, sion and diabetes mellitus; isolated reports indicate that response to bariat- Denmark ric surgery is lower in older patients. Aim of this study was to evaluate the Roux-en-Y gastric bypass (RYGB) improves glucose metabolism in type role of age and of duration of obesity on frequency of co-morbidities in mor- 2 diabetes (T2D) within days after surgery but whether early changes in bid obesity, as well as on resolution of co-morbidities. 837 consecutive sub- insulin sensitivity are part of the physiological mechanism is not known. jects undergoing gastric banding (187 men/650 women, aged 18-65 years), 2 We studied 6 obese subjects with T2D (BMI: 39.0±2.6 kg/m (mean ± sem), with known duration of obesity (5-55 years), were considered for this study; HbA1c: 7.1±0.5%) and 7 subjects with normal glucose tolerance (NGT) (BMI: they were divided into quartiles of age and of obesity duration. Presence Obesity 2 40.2±1.0 kg/m , HbA1c: 5.3±0.1%) before, 1 week (wk) after and 3 months of co-morbidities (diabetes, hyertension, metabolic syndrome), metabolic POSTERS 2 (mo) after RYGB using 4 hours hyperinsulinemic (40 mU/m /min) euglycemic variables (cholesterol, HDL-cholesterol, triglycerides, blood glucose), clini- (5.5 mmol/L) clamp combined with infusion of [6,6-2H ]-glucose to assess 2 cal variables (body weight, blood pressure, fat distribution), and weight loss Integrated Physiology/ hepatic glucose production (HGP). Fat free mass (ffm) was assessed by during 24 months were considered. Surgery improved all clinical and meta- whole body DEXA scan.Fasting glucose levels decreased in both groups at bolic parameters and led to frequent resolution of hypertension (134/312 1 wk and remained lower after 3 mo (T2D: 8.1±0.9, 6.7±0.5 p=0.05, 5.9±0.4 cases), diabetes (141/187 cases), metabolic syndrome (173/245 cases). Older p=0.08; NGT: 5.1±0.1, 4.7±0.2 p=0.02, 4.5±0.1 mmol/L p<0.01). HOMA-IR was subjects had a higher frequency of all co-morbidities, and lost less weight signifi cantly reduced in both groups at 1 wk and 3 mo (T2D: 4.9±1.3, 3.7±1.1 than younger subjects; however, diabetes and hypertension disappeared to p=0.02, 1.7±0.4 p=0.06; NGT: 2.9±0.5, 1.9±0.2 p=0.04, 0.9±0.2 p=0.02). the same extent in subjects of different ages, while metabolic syndrome Basal HGP decreased 1 wk after RYGB in T2D (3.7±0.4, 3.0±0.2 mg/kgffm/ disappeared more in older subjects, accompanied by a greater decrease of min p=0.05) whereas no signifi cant change was observed in NGT (3.2±0.3, blood glucose. Obesity duration affected frequency of co-morbidities, not 2.8±0.1 mg/kgffm/min p=0.25). Peripheral insulin sensitivity measured by response to surgery; at logistic regression, obesity duration had a moderate Mffm/I was unchanged 1 wk after RYGB in both groups but improved at 3 mo effect, independent from age, on frequency of diabetes. Older subjects, in

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A503 OBESITY—HUMANCATEGORY

spite of lower weight loss, have a similar response to younger subjects to Table 1. Factors that impact insulin discontinuation after bariatric surgery in bariatric surgery. Age and duration of obesity should not be regarded as early and late stage limits to bariatric surgery. Early (1 month) P value Late (12 months) P value Supported by: Ministero dell’Istruzione dell’Università e della Ricerca (Italy) Odds Ratio Odds Ratio (95% CI) (95.0% C.I.) & 1970-P Sex (%Male) 1.02 (0.88-1.19) NS 0.85 (0.75-0.96) 0.009 Effect of Weight Loss on Sympatho-Vagal Balance in Patients With Race (%Caucasian) 0.94 (0.83-1.10) NS 0.95 (0.85-1.05) NS Grade-3 Obesity; Restrictive Surgery Versus Hypocaloric Diet ANTONIO E. PONTIROLI, CLAUDIA MERLOTTI, ANNAMARIA VERONELLI, PAOLO Procedure (%RYGB) 2.21 (1.88-2.61) <.001 1.57 (1.34-1.84) <.001 TERRANOVA, FRANCESCA FRIGÈ, MARCO CESARANO, FEDERICO LOMBARDI, Age (years) 0.98 (0.98-1.00) <.001 0.98 (0.98-0.99) <.001 Milano, Italy Preoperative BMI (kg/m2) 1.00 (0.99-1.01) NS 1.01 (1.00-1.02) 0.033 Few and mostly uncontrolled studies have indicated that weight loss im- EBWL at 1 month (%) 1.03 (1.02-1.04) <.001 NA NA proves heart rate variability (HRV) in grade-3 obesity. Aim of the study was to compare surgery and hypocaloric diet on clinical and metabolic variables, EBWL at 12 months (%) NA NA 1.02 (1.02-1.03) <.001 and on selected indices of HRV, that estimates sympatho-vagal interaction, in grade-3 obesity. 24 obese patients (body mass index, BMI 46.1±0.89 kg/ 2 m ) underwent surgery (n = 12, gastric banding or intragastric balloon) or & 1972-P received diet (n = 12, 1,200 kg/day). Clinical [BMI, systolic (SBP) and diastolic Longitudinal Changes in Gene Expression in Human Subcutaneous blood pressure (DBP)], metabolic variables [glucose, cholesterol, HDL- and and Omental Fat Following Bariatric Surgery LDL-cholesterol, triglycerides, AST and ALT transaminases], and parameters PATRIC S. LUNDBERG, CRISTOPHER J. LYNCH, JERRY L. NADLER, ANCA D. DO- of HRV [time: R-R intervals and their standard deviation, SDNN; frequency: BRIAN, ROBERT N. COONEY, Norfolk, VA, Hershey, PA LF/HF ratio, and QT interval on 24-h Holter ECG recordings], were measured Bariatric surgery has long-term benefi cial effects on the metabolic and at baseline and after 6 months. The two groups were identical at baseline. cardiovascular phenotype. Visceral omental fat (OM) is traditionally regard- BMI (-7.5±1.03, kg/m2, mean±SE,), glucose (-24.1±7.73 mg/dl), SBP (-16.7±6.41 ed as more pro-infl ammatory compared to the subcutaneous (SC) depot. The mmHg) and DBP (-6.2±2.47 mmHg) decreased in surgery patients (always p aim of this project was to determine changes in gene expression in SC and < 0.05), not in controls. At 6 months, SDNN increased in surgery patients OM adipose tissue in paired, longitudinal samples collected at a median time (24.9±10.73 ms, p < 0.05), whereas the LF/HF ratio diminished (2.9±1.84 ver- of 1.2 years between bariatric and subsequent corrective surgeries. Gene sus 4.9±2.78; p = 0.01) in comparison to controls. Decrease of BMI correlated expression changes in 8 obese, type 2 diabetic subjects were determined with SBP and DBP decrease (p < 0.05); DBP decrease correlated with HR using the human genome U133A2.0 array. Results were analyzed by Ingenu- decrease (p < 0.05) and QT decrease (p < 0.05), and HR decrease correlated ity Pathway Analysis software package. Of the top 15 signaling pathways with QT decrease (p < 0.001).We conclude that weight loss is associated with most signifi cantly changed (p-value<0.01), 9 were common for both OM and improvement of glucose metabolism, of blood pressure, and with changes in SC depots, including 4 of the top 5 pathways. Specifi c changes in OM path- time and frequency domain parameters of HRV; this suggests a more physi- ways included tissue remodeling (integrin signaling) and metabolic response ological autonomic control, with increase in parasympathetic and reduction (AMPK), while in SC included cancer signaling related to EMT. Interestingly in sympathetic modulation of cardiovascular function. the HIF1_ canonical signaling was changed more in the OM depot. Real- Supported by: Ministero dell’Istruzione dell’Università e della Ricerca (Italy) time PCR analysis confi rmed that HIF1_ expression was reduced (p=0.03) in the OM but not SC fat following bariatric surgery. None of the top 10 most & 1971-P up-regulated molecules were shared by the OM and SC depots. However, Changes in Insulin Dependence after Bariatric Surgery of the top 5 transcriptional regulators, SMAD1 and WT1 were present in ALI ARDESTANI, DAVID RHODES, ALI TAVAKKOLIZADEH, Boston, MA both depots, while STAT5b, HAND1 and NR0B1 were specifi c for the OM Bariatric surgery, including gastric bypass surgery (RYGB) and laparo- and Rxr, MEF2C and SRF for the SC. Interestingly, Runx3 was decreased in scopic adjustable gastric banding (LAGB), leads to signifi cant improvements both depots, by 2.9 fold (Z score -2.251) in SC and by 0.4 fold (Z score -2.053) in Type 2 Diabetes Mellitus (T2DM) and has recently been incorporated in in OM. Also, some of the target molecules such as CD4, CCR7, IL8, Jag1 the treatment algorithm of most diabetic societies. Although insulin de- were also signifi cantly decreased following bariatric surgery. Collectively pendent patients have lower success rates with surgery, few studies have data indicates a pattern of reduction in several components of infl ammatory evaluated this in a large cohort of patients. Using a prospectively collected pathways in both depots and a predominance of change in the ones involved national database (Bariatric Outcomes Longitudinal Database), we identi- in tissue remodeling and hypoxia in the OM. Signifi cant differences between fi ed113,638 adult patients who had undergone bariatric surgery, 10% had depots in changes of individual genes suggest depot-specifi c regulatory re- insulin-dependent T2DM, from which we analyzed 5,225 patients who had sponses following bariatric surgery. at least one year of follow up. At 12 months 62% of RYGB and 34% LAGB patients were off insulin (Figure 1). Regression models were developed to identify factors that contribute to insulin independence at 1 and 12-months & 1973-P postoperatively (Table 1). At both time points, RYGB was a strong predictor DPP4 in Morbidly Obese Patients: Increase in Diabetic Subjects, of insulin independence. For LAGB, insulin independence appeared linked to Correlation With Markers for Fatty Liver and Decrease after Weight weight loss, while for RYGB the effect appeared to be weight independent Loss Induced by Bariatric Surgery in early stages. With increasing utility of bariatric procedures for treatment HENRIKE SELL, JOAN TORDJMAN, PIERRE BEDOSSA, JEAN-LUC BOUILLOT, of T2DM, RYGB appears to offer a greater chance of insulin independence JÜRGEN ECKEL, KARINE CLÉMENT, CHRISTINE POITOU, Düsseldorf, Germany, (over 60% at 1 year), and may be the procedure of choice. Paris, France, Clichy, France, Boulogne-Billancourt, France Dipeptidyl peptidase 4 (DPP4) is a novel adipokine. Larger adipocytes from obese subjects release more DPP4 paralleled by increased circulating DPP4 levels. Serum DPP4 is associates with BMI and the metabolic syndrome Obesity

POSTERS in obese subjects in vivo. This study investigated DPP4 serum levels in 53 morbidly obese patients in association with metabolic diseases (type 2 dia- betes, non-alcoholic fatty liver disease) and after surgery-induced weight Integrated Physiology/ loss. Patients were examined at baseline before bariatric surgery as well as 3, 6 and 12 months after surgery. Furthermore, liver biopsies were taken at baseline to analyze the presence/degree of liver pathology.DPP4 was signifi - cantly elevated in morbidly obese patients with type 2 diabetes compared to patients without diabetes independently from BMI (664±39 ng/ml vs 534±34 ng/ml, p=0.015). Preoperatively, serum DPP4 correlated positively with trig- lycerides and liver parameters (AST, ALT and gammaGT). Baseline DPP4 was elevated in patients with a activity score for NAFLD *3 compared to pa- tients with a score <3 (757±75 ng/ml versus 504±60 ng/ml, p=0.018). DPP4 was also elevated in subjects with azonal/panacinar location of steatosis

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A504 OBESITY—HUMANCATEGORY compared to steatosis located around central veins (725±60 ng/ml versus between B and C. Total calories were signifi cantly reduced by 41% as were 449±66 ng/ml, p=0.007). During the observation period of 12 months, DPP4 fat grams (F = 13.1, p =.008, power = .87). Increasing pre-meal dopamine decreased signifi cantly from 589±27 ng/ml at baseline to 530±21 ng/ml after may promote reduced calorie and fat intake in obese women independent 3 months, 538±27 ng/ml after 6 months and 467±18 ng/ml after 12 months. of BED status. The decrease of DPP4 over 12 months after surgery was related to the de- Supported by: NIDDK crease in blood glucose and HOMA-IR.DPP4 concentrations are elevated in morbidly obese patients correlating with type 2 diabetes and markers of & 1976-P liver pathology. DPP4 plasma levels decrease after surgery-induced weight Association Between the Serotonin Transporter in the Infundibular loss in parallel to improved insulin sensitivity. This study suggests that DPP4 Nucleus of the Hypothalamus and Body Mass Index: a Post-Mortem might be a marker of obesity-associated metabolic diseases which improve Study after gastric bypass. KARIN E. KOOPMAN, ANNEKE ALKEMADE, ANKE J. BORGERS, ERIC FLIERS, Supported by: German Research Council (DFG) MIREILLE J. SERLIE, SUSANNE E. LA FLEUR, Amsterdam, The Netherlands Obesity is one of the major risk factors for the development of type 2 dia- & 1974-P betes. An association between the serotonergic system and body mass index Roux-en-Y Gastric Bypass vs. Sleeve Gasterctomy for Obese Pa- (BMI) has recently become evident. Both brain imaging- and genetic studies tients With Type 2 Diabetes: A Randomized Trial in human subjects have shown associations between serotonin transporters KAREN J. HERSHKOP, ANDREI KEIDAR, CHAYA SCHWEIGER, LIOR HECHT, RAM (SERT) and BMI. From rodent studies, it is clear that the serotonin system WEISS, Jerusalem, Israel within the arcuate nucleus of the hypothalamus (infundibular nucleus in Bariatric surgery is gaining acceptance as “metabolic surgery” for obese humans) plays an important role in body weight regulation. However, the patients type 2 diabetes (T2DM). The optimal surgery and the mechanism of neuroanatomical resolution of in vivo SERT imaging does not allow for the action of these procedures are unknown.We performed a randomized trial distinction between separate hypothalamic nuclei in humans. In the present of Roux-en-Y gastric bypass (RYGB) vs. Sleeve Gastrectomy (SG) for obese study we aimed to investigate the relationship between SERT expression patients with T2DM and followed glucose tolerance and body composition and BMI in post-mortem samples of human hypothalamic infundibular nucle- at 3 and 12 months. Non diabetic obese patients undergoing the same proce- us (IFN).We divided hypothalami of n=22 nondiabetic subjects in two groups, dures were used as a reference group for assessment of the effects on indi- i.e., BMI < 25 kg/m2 (n=13, median 20.3 kg/m2 (range 15.2 - 25)) and BMI > ces of insulin secretion and action.Twenty nine of 30 obese diabetic patients 25 kg/m2 (n=9, median 32.0 kg/m2 (range 26.0 - 41.0)). Median age was 75 (13RYGB/16SG) and 17 of 19 obese non diabetic patients (7RYGB/10SG) years (range 50 - 92) and 76 years (range 26 - 100) in the two groups, respec- completed the protocol. Weight loss (~30-35 kg reduction) and changes in tively. There was no obvious difference in causes of death or co-morbidities. lean body mass and fat mass were similar between the groups. Hba1C levels Serial coronal sections (6μm) were cut over the entire rostro-caudal axis dropped signifi cantly and equally from 7.6+0.3 and 8.3+0.6 % at baseline to of the hypothalamus and every 100th section containing the IFN was used 6.3+0.09 and 6.1+0.2% at 2 months in the RYGB and SG groups respectively. for immunocytochemical staining. We performed computer assisted density Whole body insulin sensitivity doubled by 12 months in all participants. The measurements of SERT immunoreactivity as an estimate of relative SERT acute insulin response improved equally in diabetic patients. The oral dis- protein expressionIn the group with a BMI < 25 kg/m2, median SERT density position index improved signifi cantly in diabetic patients (from 2.2±2.8 and was 16.2 arbitrary units (a.u.) (range 6.5 - 35.8), while in subjects with a BMI 1.65±2.2 at baseline to 5.7±3.1 and 6.1±3.5 at 12 months in the RYGB and SG > 25 kg/m2 median SERT density was 11.3 a.u. (range 3.3 - 16.9), which was groups respectively, p<0.001 for both) yet remained well below the baseline signifi cantly lower (p=0.046, Mann Whitney U).These fi ndings suggest that disposition index of non diabetic patients.RYGB and SG have comparable hypothalamic SERT expression is negatively associated with BMI. However, clinical effects on diabetes resolution and anthropometric parameters dur- the pathophysiological mechanism behind this observation remains to be ing 12 months follow up. Diabetes resolution is impressive yet the disposi- investigated. tion index of patients remains far from normal. Supported by: EFSD & 1977-P Physiological Decrements in Glucose Levels Are Associated With Hunger and Deactivation of the Pre-Frontal Cortex Guided Audio Tour: Brain and Obesity (Posters 1975-P to 1979-P), see page RENATA BELFORT DE AGUIAR, SARITA NAIK, DONGJU SEO, CHERYL M. LACADIE, 17. TODD CONSTABLE, RAJITA SINHA, ROBERT SHERWIN, New Haven, CT Hypoglycemia is associated with increased hunger, but whether small & 1975-P physiological changes in plasma glucose (PG) levels infl uence food-seeking Effect of Dopamine Agonist on Food Reinforcement and Selection in behavior is unclear. To address this question we evaluated 20 healthy sub- Obese Women With and Without Binge Eating Disorder (BED) jects (mean age 29±7, A1c 5.4±0.3, BMI 24±2) as they underwent a 2mU/ JENNIFER A. NASSER, CARLA WOLPER, ALLAN GELIEBTER, RICHARD N. ROSEN- kg.min hyperinsulinemic euglycemic clamp (goal=PG ~95mg/dl) while view- THAL, PASCAL KOLAK, SAMI A. HASHIM, Philadelphia, PA, New York, NY ing food and non-food pictures combined with functional MRI. Hunger rat- Obese binge eaters (B) increase their operant responding for food after ings were collected at baseline and 2h. An age and BMI matched control consumption of a satiating palatable preload compared to a fasting con- group of 10 healthy subjects was studied using a 0.9% saline infusion in- dition while obese non-binge eaters (C) decrease their operant responding stead of a clamp.During the clamp, PG initially rose from 85±11 at baseline to under the same condition. To test if this sensitization response is due to 93±13 mg/dL after 10min. During the next 10 to 60 min interval PG showed low dopamine tone, we hypothesized that operant responding for food in some variability (85±11 to 99±11 mg/dL): PG decreased In 10 subjects by the fed state would decrease after dosing with methylphenidate (Ritalin), a 14±10 mg/dL (G.Fall group) and increased (G.Increase group) in 10 subjects dopamine agonist. Ten obese subjects (5 B, 5 C) completed one session of an by 12±12 mg/dL. Subsequently, PG stabilized at 95±8 mg/dL in both groups ad libitum meal of snack foods (no Ritalin), and three sessions of computer until the end. Hunger ratings signifi cantly increased from 2±2 at baseline operant task performance 1 hour after Ritalin dosing. One of three doses of to 5±3 at the end of the study (p<0.01). Change in hunger ratings during

Ritalin (0, 20, 40 mg) was tested each session. Subjects consumed 400 kcal the clamp was associated with the drop in glucose between 10 and 60mins Obesity of a liquid meal 2 hours prior to a session, and 600 kcal of the same liquid (r=-0.469, p=0.032). Hunger was associated with increased activation in POSTERS meal during the session, 30 minutes before the ad libitum snack food meal the insula, thalamus and striatum. The G.Fall compared to the G.Increase

or operant task performance. The computer generated progressive ratio group showed deactivation of the ventral medial prefrontal cortex (PFC). Integrated Physiology/ consisted of ten, 1 minute trials, in which subjects chose between food or The changes in glucose were not associated with hormone levels (gluca- non-food gift categories, completing an increasing number of responses by gon, cortisol or ghrelin). Brain activation was not different between the two pressing on the designated computer key. The main outcome measure was groups.Small decrements in PG within the normal range provoked increased food break point (FBP; the highest ratio of computer responses completed hunger, activation of the striatal-limbic system, and deactivation of the PFC when choosing the food category). Additional outcome measures were total (cognitive control) when individuals were exposed to visual food cues. These calories and grams of macronutrients of foods obtained in the operant task. data suggest that the capacity of the brain to control desire for food and Data were analyzed by repeated measures ANOVA with BED status as the reward-motivation pathways is impaired by small decrements in circulating between subjects factor. B consumed 55% (p =.028) more calories than C in glucose and might contribute to inappropriate over-eating and weight gain the ad lib snack meal. Dosing with 20 mg and 40 mg Ritalin produced a 41% in healthy individuals. decrease in FBP (F = 6.5, p = .038, power = .59) but no signifi cant difference Supported by: R37-20495, UL1-DE019586, PL1-DA024859

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A505 OBESITY—HUMANCATEGORY

1980-P & 1978-P Gastric Bypass Surgery Modifi es Ethanol Consumption Smoking Diminishes Motivation-Reward Neural Responses to Food JON F. DAVIS, JENNIFER D. SCHURDAK, IRWIN J. MAGRISSO, JORAM MUL, Cues Independent of Body Weight BERNADETTE GRAYSON, PAUL T. PFLUGER, MATHIAS H. TSCHOEP, RANDY J. ANIA M. JASTREBOFF, RAJITA SINHA, CHERYL LACADIE, ROBERT S. SHERWIN, SEELEY, STEPHEN C. BENOIT, Cincinnati, OH MARC N. POTENZA, New Haven, CT Roux en Y gastric bypass (RYGB) surgery is an effective weight loss strat- Co-occurrence of smoking and obesity is synergistically associated with egy employed to treat obesity and associated complications. Importantly, increased risk of cardiovascular disease and cancer. Smoking status is as- the RYGB procedure has been reported to modify alcohol intake. Prior stud- sociated with relatively lower body weight, yet why this occurs has not been ies report increased alcohol intake in RYGB patients following surgery; a fully elucidated. Both smoking and obesity have been shown to affect neural more recent study concluded that despite no observable change in intake responses in reward-motivation pathways. We matched smokers and non- following surgery, bariatric patients have a higher lifetime prevalence of smokers based on body mass index (BMI), age, gender, and education and alcohol misuse. Thus, it is unclear if RYGB surgery is capable of altering al- examined whether smoking status altered neural activation in motivation- cohol intake. The present work examined the hypothesis that RYGB surgery reward and emotion-regulating regions of the brain in response to brief mediates alcohol intake and reward in individuals that frequently consume guided-imagery of favorite foods. Twenty-three smokers and 23 non-smok- alcohol. To do this, alcohol intake was examined in human bariatric patients ers participated in six trials of exposure to personalized food-cue, stress, (n=6,165) who self-reported alcohol intake prior to receiving the RYGB pro- and neutral-relaxing situations using a validated, functional MRI (fMRI) cedure. In addition we utilized a rodent model of RYGB and examined ethanol paradigm. In group contrast brain maps of neural responses during exposure consumption and ethanol reward in male alcohol preferring (P) rats which to the food-cue condition, smokers as compared to non-smokers exhibited are selectively bred to consume large volumes of ethanol. Patients that decreased activation in corticolimbic-striatal regions including the caudate, self-reported occasional-frequent alcohol use reported decreased usage putamen, insula, and thalamus. In condition contrast brain maps of food-cues following RYGB surgery. Moreover, the RYGB procedure decreased ethanol vs. neutral-relaxing condition, non-smokers exhibited increased activation in intake and the reinforcing properties of ethanol in P rats. Notably, the at- the inferior frontal gyrus (IFG), middle temporal gyrus (MTG), putamen, cau- tenuating effect of RYGB surgery on ethanol consumption was associated date, insula, amygdala, and hippocampus; smokers demonstrated increased with ethanol-induced excursions in the gut hormone glucagon like peptide-1 activation in the IFG and MTG but did not exhibit increased activation in stri- (GLP-1). Pharmacological manipulation of GLP-1 signaling modulated ethanol atal or limbic regions (p<.05 whole brain corrected). No statistically signifi - consumption in P rats. In addition, pharmacological replacement of the gut cant differences were found between smokers and non-smokers with regard hormone ghrelin restored drinking behavior in P rats following RYGB. These to insulin resistance (HOMA-IR), fasting insulin, glucose, leptin, and cortisol data indicate that baseline alcohol consumption may explain the variety of levels, or measures of subjective food craving. These fi ndings suggest that outcomes reported in patients that undergo RYGB. Moreover, this work de- smokers demonstrate diminished neural responses to food cues irrespective tails the neuroendocrine signaling mechanisms which modify consummatory of insulin resistance or BMI status. Differences in neural response to food behaviors following RYGB surgery cues in smokers may contribute to their relatively lower BMI and perhaps to Supported by: Ethicon EndoSurgery, Inc. the weight gain seen with attempts to quit smoking. Supported by: RL1AA017539, R37-DK 20495, UL1-DE019586, UL1-RR024139 1981-P Incident Type 2 Diabetes is Related to Visceral Adiposity and Liver Fat Assessed by CT: Three Year Prospective Data from the INSPIRE & 1979-P ME IAA Study Differential Effect of Glucose Ingestion on the Cerebral Processing JEAN-PIERRE DESPRÉS, STEVEN M. HAFFNER, BEVERLEY BALKAU, CHRISTINE of Food Stimuli in Lean and Overweight MASSIEN, JESSICA SMITH, ANNE-LAURE BOREL, JULIE-ANNE NAZARE, NATA- MARTIN HENI, CAROLINE KETTERER, RALF VEIT, STEPHANIE KULLMANN, MAR- LIE ALMÉRAS, ROBERT ROSS, FOR THE INSPIRE ME IAA INVESTIGATORS, Que- TINA GUTHOFF, HARALD STAIGER, FAUSTO MACHICAO, HANS-ULRICH HÄRING, bec City, QC, Canada, Houston, TX, Villejuif, France, Paris, France, Kingston, ON, HUBERT PREISSL, ANDREAS FRITSCHE, Tübingen, Germany, Little Rock, AR Canada Prevalence of type 2 diabetes and obesity is rising worldwide. Since eat- Although there are cross-sectional data providing evidence that visceral ing behavior plays an important role in both disorders, understanding how adiposity and liver fat accumulation are associated with an altered cardio- food intake is regulated is of great importance. Aims of this study were to metabolic risk profi le, few prospective studies have quantifi ed the respec- investigate how the processing of food cues is infl uenced by glucose in- tive contributions of these ectopic fat depots to the incidence of type 2 gestion compared to water in lean and overweight and to study potential diabetes (T2D). The INternational Study of Prediction of Intra-Abdominal hypothalamic differences regarding variation in the obesity-risk gene FTO. Adiposity and its Relationships with CardioMEtabolic Risk/Intra-Abdominal Twelve lean (6 female, BMI 21.2±1.1 kg/m², age 22.9±2.1 years) and 12 over- Adiposity (INSPIRE ME IAA) evaluated cardiometabolic risk factors, includ- weight subjects (6 female, BMI 30.5±1.8 kg/m², age 24.7±2.4 years) geno- ing visceral adipose tissue (VAT), abdominal subcutaneous adipose tissue typed for FTO SNP rs8050136 participated. Functional magnetic resonance (SAT) and liver attenuation by computed tomography (inversely related to imaging (fMRI) during stimulation with food pictures was performed before, liver fat) at baseline in 2157 non diabetic patients from 29 countries in Asia, 30, and 120 min after ingesting a 75 gram glucose solution on one day and Europe, Latin America and North America, aged 39 to 71. One ninety nine an equal amount of water on another day in randomized order (challenge). incident T2D cases (109 men, 10.2% ; 90 women, 8.3%) were identifi ed dur- We detected a signifi cant main effect of weight group (lean versus over- ing the 3-yr follow-up.Hazards ratios (95% CI) for incident diabetes adjusted weight) in the left putamen and increased activity after glucose intake in for age, ethnicity, recruiting physician specialty. the hypothalamus independent of weight group. There were interactions between weight group and challenge in the left lingual gyrus and the right MEN WOMEN anterior cingulate cortex (p<0.001). For these areas, post-hoc tests revealed BMI* 30 vs <25 kg/m2 2.36 [ 1.28 , 4.34 ] p=0.006 3.74[ 2.01 , 6.95 ] p<0.001 signifi cant differences between water and glucose only in lean but not in VAT (tertile 3 vs 1) 4.12[ 2.20 , 7.72 ] p<0.001 4.87[ 2.31 , 10.30 ] p<0.001 overweight subjects, 30 min after glucose ingestion. We furthermore found Obesity signifi cant differences between FTO genotypes in the hypothalamus 30 min SAT (tertile 3 vs 1) 1.56[ 0.91 , 2.68 ] ns 1.95[ 1.02 , 3.71 ] p=0.043 POSTERS post glucose load with reduced activity in non-risk allele carriers.Our results Liver attenuation (tertile 3 vs 1) 0.50[ 0.30 , 0.84 ] p<0.008 0.22[ 0.12 , 0.42 ] p<0.001 show that glucose ingestion has specifi c effects on the processing of food IFG/IGT vs NGT 4.73[ 2.72 , 8.21 ] p<0.001 5.26 [ 3.10 , 8.92 ] p<0.001 Integrated Physiology/ stimuli in brain areas important for eating behavior and energy balance. This could contribute to the postprandial termination of food intake. The detected Overall, results were stronger for VAT and liver attenuation than for SAT effects may be critical to prevent overfeeding and are dependent on weight or BMI and trends were stronger in women than in men. This prospective and FTO genotype. Our fi ndings provide new insights in the pathogenesis study provides evidence that visceral adiposity and liver fat assessed by of obesity and might help to understand how genetic variation infl uences computed tomography are key predictors of T2D risk in both men and body weight. women. Supported by: sanofi -aventis

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A506 OBESITY—HUMANCATEGORY

1982-P 1984-P Circulating Sex Hormone Binding Globulin Predicts Fatty Liver and Impact of FTO Genotypes on Weight and Body Mass Index in Poly- Insulin Resistance in Children and Adolescents cystic Ovary Syndrome: A Systematic Review and Meta-Analysis KATARZYNA LINDER, FABIAN SPRINGER, JÜRGEN MACHANN, FRITZ SCHICK, PIOTR WOJCIECHOWSKI, ANNA LIPOWSKA, PRZEMYSLAW RYS, KATHRYN ANDREAS FRITSCHE, HANS-ULRICH HÄRING, STEFAN EHEHALT, GERHARD G. EWENS, STEVEN FRANKS, SUSAN TAN, ELISABETH LERCHBAUM, JOSEF BINDER, NORBERT STEFAN, Tübingen, Germany VCELAK, MAREK STRACZKOWSKI, RICARDO AZZIZ, THOMAS BARBER, ANKE In adults sex hormone-binding globulin (SHBG) levels decrease when fat HINNEY, BARBARA OBERMAYER-PIETSCH, PETRA LUKASOVA, BELA BEND- accumulates in the liver and SHBG is involved in the pathogenesis of type LOVA, IRINA KOWALSKA, MARK O. GOODARZI, JEROME F. STRAUSS III, MARK 2 diabetes. In the present study we investigated whether circulating SHBG MCCARTHY, MACIEJ T. MALECKI, Krakow, Poland, Atlanta, GA, Philadelphia, PA, may be a predictor of fatty liver and insulin resistance already in children and London, United Kingdom, Essen, Germany, Graz, Austria, Prague, Czech Republic, adolescents.Total body- and visceral fat were precisely measured by mag- Bialystok, Poland, Augusta, GA, Coventry, United Kingdom, Los Angeles, CA, Rich- netic resonance (MR) tomography and liver fat by 1H-MR spectroscopy in 38 mond, VA, Oxford, United Kingdom children and adolescents. They were matched to 38 adults who underwent The FTO gene single nucleotide polymorphism rs9939609 has been as- measurements by the same MR techniques by BMI. The BMI of the chil- sociated with body weight and related traits as well as with type 2 diabetes. dren and adolescents was calculated by following their individual percentile Recently, several small studies have suggested a greater than expected curves to the age of eighteen.In children and adolescents fasting circulat- effect of FTO variants on weight in polycystic ovary syndrome (PCOS). We ing SHBG was not different between boys and girls (means±SE; 19.3±2.9 aimed to examine the impact of the FTO genotypes on weight and BMI in and 22.5±4.1, p=0.53), while in adults circulating SHBG was lower in males PCOS women by combining relevant studies in a meta-analysis. A system- compared to females (20.9±2.2 vs. 34.4±5.2, p=0.007). In children and ado- atic search of major medical databases (PubMed, EMBASE, Cochrane’s lescents circulating SHBG strongly decreased with increasing age (r=-0.49, CENTRAL) was conducted through April 2011. Six studies describing seven p=0.0015), while no correlation with age was found in adults (r=0.21, p=0.2). distinct cohorts of PCOS women were retrieved. Mean weight and BMI were After adjustment for age and gender, a strong correlation of circulating calculated for each genotype and pairwise meta-analyses were performed SHBG was found with liver fat content (r=0.44, p=0.0079) in children and in in order to compare groups. A total of 2359 PCOS women were included adolescents, as well as in adults (r=0.40, p=0.017). Furthermore, circulating in the study; of these 710 were rs9939609 TT homozygotes, 1166 had an SHBG correlated negatively with the HOMA-IR, both in children and adoles- AT genotype, and 483 were AA homozygotes. AA homozygotes weighed on cents (r=-0.35, p=0.034) and in adults (r=-0.31, p=0.068). Furthermore, both, average 9.00 kg more than the TT homozygote group [95% CI 6.31, 11.68]. AA in the group of children and adolescents as well as in the group of adults homozygosity increased BMI by 3.05 kg/m2 as compared to TT homozygotes circulating SHBG was found to strongly predict fatty liver (area under the [95% CI 2.14, 3.96]. Meta-regression analysis demonstrated that a single ROC curve: 0.74 for children and 0.73 for adults).In conclusion, our data sug- A allele impacted body weight and BMI only in the non-obese cohorts. In gest that while differences in the relationships of circulating SHBG with age conclusion, the difference in weight and BMI in PCOS patients between FTO and gender exist between the groups of children and adolescents and adults, rs9939609 AA and TT genotypes in this analysis were around three times circulating SHBG may represent a strong predictor of nonalcoholic fatty liver greater than values reported in the large population based studies. Our re- both, in children and adolescents and in adults. sults suggest that the metabolic context or specifi c polygenic background of PCOS modifi es the infl uence of FTO on weight and BMI. 1983-P Evaluating the Feasibility and Impact of an Internet-Based Lifestyle 1985-P Modifi cation Program in a Clinical Setting Weight Loss (WL) and Change in Antidiabetic Medications Over 2 DIANA SHERIFALI, GAYA AMIRTHAVASAR, ANKA BROZIC, HERTZEL GERSTEIN, Years With Controlled-Release Phentermine/Topiramate (PHEN/ Hamilton, ON, Canada, Cambridge, ON, Canada TPM CR) Lifestyle programs that incorporate physical activity, nutrition and be- ROBERT F. KUSHNER, BARBARA TROUPIN, Chicago, IL, Mountain View, CA haviour modifi cation have been shown to promote weight loss and improve Type 2 diabetes mellitus (T2DM) management focuses on glycemic con- glycemic control. The Virtual Lifestyle Management (VLM) Program is a trol via WL, lifestyle interventions, and medication. Signifi cant WL has been web-based lifestyle program based on the Diabetes Prevention Program, demonstrated in subjects with T2DM receiving PHEN/TPM CR vs placebo and offers education and coaching to facilitate lifestyle change. The pur- (PBO) over 56 weeks. Longer-term effects of PHEN/TPM CR on WL and T2DM pose of this pilot study was to assess the impact of the VLM program in a management were evaluated through 108 weeks in SEQUEL, a blinded ex- diabetes clinic setting. Using an observational design, 79 individuals were tension of CONQUER, with randomization to lifestyle intervention plus PBO recruited to the VLM program for 1 year, in addition to their care from a (n=227), PHEN 7.5mg/TPM CR 46mg (7.5/46; n=153), or PHEN 15mg/TPM CR diabetes clinic. Inclusion criteria were: a) adults >40 yrs of age; b) had type 92mg (15/92; n=295 ) maintained. Metformin monotherapy or diet/exercise 2 diabetes; c) a Body Mass Index (BMI) >30; d) able to read/write English; was permitted at enrolment; subsequent addition of oral hypoglycemics and e) had internet access. Participants were assessed at baseline, 6 and 12 was allowed if control was inadequate. This post hoc analysis in subjects months for the following outcomes: a) glycated hemoglobin (A1C); b) lipids; with T2DM at baseline (PBO, n=55; 7.5/46, n=26; 15/92, n=64) assessed WL, c) weight; d) BMI; and e) body fat %.Eighty people were screened for the change in HbA1c, and net change in concomitant antidiabetic medication study, 79 consented and 66 individuals (84%) logged onto the system >2 dose (percent of subjects increasing minus decreasing dose), including medi- times. At baseline, the mean age of participants was 57.9 yrs of age and cations added post-baseline. Baseline metformin use was as follows: PBO, 52% were female. Changes to study outcomes from baseline to 6 months n=29; 7.5/46, n=12; 15/92, n=38. At Week 108, least-squares mean percent and baseline to 12 months are presented in the table below. This pilot study WL (ITT-LOCF) was signifi cantly greater in subjects receiving PHEN/TPM CR demonstrates that an internet-based lifestyle modifi cation program (VLM) vs PBO (P<.0001): 2.0%, 9.0%, and 9.0% for PBO, 7.5/46, and 15/92, respec- can improve outcomes in a clinical setting. The study fi ndings are applicable tively. PHEN/TPM CR also improved HbA1c: 0.0%, -0.4%, and -0.2% for PBO, to any diabetes clinic setting seeking to extend their current lifestyle modifi - 7.5/46, and 15/92, respectively (P=NS vs PBO). Furthermore, 25.5% of PBO cation services and capabilities. subjects had a net increase in antidiabetic medication dose, while 7.5/46

subjects had no net change in dose and 3.1% of 15/92 subjects had a net Obesity Baseline, 6 and 12 month mean differences for study outcomes (*p<0.05) decrease (P=.0005 vs PBO, both comparisons). Percentages of subjects in POSTERS Outcomes N Baseline - 6 N Baseline - 12 the overall safety population discontinuing the study due to adverse events MnMean Diff MnMean Diff

were 3.1%, 4.6%, and 4.4% for placebo, 7.5/46, and 15/92, respectively. Integrated Physiology/ (95% CI) (95% CI) Subjects treated with PHEN/TPM CR showed greater WL and improved A1C 48 -0.4 (-0.1,-0.6)* 49 -0.3 (-0.1,-0.5)* diabetes control despite requiring less concomitant antidiabetic medication, TC 45 -0.29 (-0.0,-0.6)* 38 -0.04 (-0.3,0.2) representing a potential added benefi t of PHEN/TPM CR for obese patients TC/HDL 45 -0.4 (-0.1,-0.6)* 38 -0.2 (-0.01,-0.4)* with T2DM. Supported by: Vivus, Inc. Wt (lbs) 50 -7.4 (-4.2,-20.5)* 45 -8.6 (-3.7,-13.6)* BMI 47 -1.2 (-0.7,-1.7)* 43 -1.5 (-0.7,-2.3)* Body Fat% 43 -1.7 (-0.9,-2.5)* 41 -1.8 (-0.9,-2.7)*

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1986-P 1987-P Increased Mortality in Normal Weight People With Metabolic Syn- Determinants of Visceral Fat Storage in Visceral Adipose Tissue in drome Compared With Metabolically Healthy Obese Individuals: Healthy Men During a 2-Month High-Fat Overfeeding The South-West Seoul (SWS) Study MAUD ALLIGIER, LAURE GABERT, EMMANUELLE MEUGNIER, DEBARD CYRILLE, HO CHEOL HONG, JAE HEE AHN, MYONG JIN CHO, NAM HOON KIM, HAE YOON STÉPHANIE LAMBERT, MONIQUE SOTHIER, BÉATRICE MORIO, HUBERT VIDAL, CHOI, SAE JEONG YANG, YOON JUNG KIM, JOO HYUNG KIM, CHAI RYOUNG MARTINE LAVILLE, Oullins, France, Clermont-Ferrand, France EUN, HYE JIN YOO, HEE YOUNG KIM, JI A. SEO, NAN HEE KIM, SIN GON KIM, Obesity is associated with a cluster of metabolic abnormalities which have SEI HYUN BAIK, DONG SEOP CHOI, KYUNG MOOK CHOI, TAE NYUN KIM, Seoul, been related to an increase in visceral fat. Mechanisms leading to the increase Republic of Korea, Busan, Republic of Korea in visceral fat are still unknown. Overfeeding experiment is a unique tool to The purpose of this study was to investigate the impact of body mass study early mechanism. Anthropometry, abdominal fat mass distribution as- index (BMI) and metabolic syndrome (MetS) on all-cause, cardiovascular sessed by MRI, and fasting metabolic parameters were measured in 44 male and cancer mortality in elderly Korean men and women. A total of 2,373 subjects before (D0) and after overfeeding, (HF-OF) (D56). The HF-OF effect on elderly people aged over 60 years were studied using follow-up data of the the metabolic fate of postprandial lipids was studied during a lipid labelled test South-West Seoul (SWS) Study, a prospective cohort study during a median meal, the regulation of the expression of genes coding for the key enzymes follow-up of 9 years. Participants were classifi ed by presence (n = 751) or and regulators of lipid metabolism in abdominal subcutaneous adipose tissue absence (n = 1,619) of MetS and by BMI (<23 kg/m2 = normal weight, n = (ASAT) biopsies. To characterize the determinants of visceral fat storage, asso- 716; 23 to 24.9 kg/m2 = overweight, n = 616; *25 kg/m2 = obese, n = 1,038). ciations between changes in gene expression, anthropometric and metabolic In age- and gender-adjusted death rate ratios from all-cause and cardiovas- parameters and changes in abdominal fat distribution were studied.HF-OF cular disease, individuals without MetS showed a U-shaped association increased body weight of 2.5kg (p=0.001) and fat mass (p=0.001). Visceral adi- with the BMI, whereas individuals with MetS exhibited a reverse J-shaped pose tissue volume (VAT) increased but with large inter-individual differences pattern. In Cox proportional-hazard models adjusting for confounding fac- (from -29% to +104%). Labelled lipid concentration in plasma non-esterifed tors, overweight subjects without MetS had the lowest risk of all-cause and fatty acids fraction was increased at D56, suggesting a higher spill-over rate cardiovascular mortality. Metabolically healthy but obese (MHO) subjects and was positively correlated with the changes in VAT (r=0.728, p=0.0055). (obese individuals without MetS) showed slightly increased risk of all-cause Unlike subjects with a low VAT gain, subjects with a high VAT gain presented a mortality (HR: 1.23, 95% CI: 0.87-1.74) and cardiovascular mortality (HR: weaker induction of several genes as DGAT2, SREBP1c and CIDEA, suggesting 1.92, 95% CI: 1.01-3.65). Metabolically obese but normal weight (MONW) a potential impaired stimulation of lipid storage. Initial low level of CPT1 ap- subjects (normal weight individuals with MetS) exhibited highest risk of all- pears as a predictor of VAT gain.Defect in ASAT appears to be involved in the cause mortality (HR: 2.41, 95% CI: 1.57-3.70) and cardiovascular mortality development of VAT in our model of experimental HF-OF. (HR: 3.63, 95% CI: 1.70-7.77). In contrast to MHO, elderly individuals with MONW phenotype exhibited increased all-cause and cardiovascular mortal- 1988-P ity during 9 years of follow-up. Intestinal Methane and Hydrogen Production on Breath Testing is Predictive of Higher BMI and Percent Body Fat in a Large Scale Pro- spective Study RUCHI MATHUR, MERIDYTHE AMICHAI, KATHLEEN S. CHUA, CHRISTOPHER CHANG, GILLIAN M. BARLOW, MARK PIMENTEL, Los Angeles, CA Increasing evidence supports a role for gut microfl ora, particularly ar- chaea, in weight gain. We found the ratio of Methanobrevibacter smithii to other organisms is an important determinant of body mass in a rat model. M. smithii is also the dominant methanogen in the human gut, and we found that methane production is associated with a 6.8kg/m2 higher BMI in obese hu- man subjects. Here we examined if methane is associated with greater BMI in an unselected population.Subjects were 792 consecutive adults referred for breath testing, and were not selected based on BMI. BMI, percent body fat, demographic variables, medications, metabolic disorders and breath test results were recorded. Four groups were identifi ed: Normal (N, N=343); Positive for Hydrogen (>20ppm rise by 90 minutes) (H, N=320); Positive for Methane (>3 ppm) (M, N=101); and Positive for both Methane and Hydrogen (M+H, N=28). Multivariate analysis was performed controlling for confound- ing variables associated with higher or lower BMI, including age and gender. Presence of M+H was most associated with higher BMI (see fi gure) and per- cent body fat compared to each other group (P)0.01) and to all other groups combined.We conclude that a combination of methane and hydrogen on breath testing is associated with signifi cantly greater BMI and percent body fat. Since methanogens require hydrogen to maximize methane production, the two may be co-dependent in the association with greater body weight. It is possible that M. smithii along with its hydrogen producing syntroph plays an important role in the development of obesity. Obesity POSTERS Integrated Physiology/

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1989-P fore we studied expression profi les of both pro- and anti-infl ammatory mark- The Association Between Body Mass Index and Hospitalization for ers in three different adipose tissue compartments (omental, mesenteric Heart Failure in 83,021 Patients With Type 2 Diabetes: A Population- and subcutaneous) in morbidly obese women undergoing bariatric surgery. Based Study from the Swedish National Diabetes Registry We compared the expression profi les with lean controls undergoing elec- SVENJA GLOGNER, ANNIKA ROSENGREN, MARITA OLSSON, SOFFIA GUDB- tive cholecystectomy. We determined insulin sensitivity and lipolysis using JÖRNSDOTTIR, ANN-MARIE SVENSSON, MARCUS LIND, Trollhättan, Sweden, a hyperinsulinemic euglycemic clamp with stable isotopes.We included 16 Gothenburg, Sweden obese (mean BMI 45 [39-51] kg/m2) and 6 lean women (BMI 22 [20-24] kg/m2). Studies are sparse to which extent low weight, overweight, and different The expression of both pro- and anti-infl ammatory markers was increased in degrees of obesity contribute to the risk of hospitalization for heart failure all adipose tissue compartments compared to the lean controls except for (HF) in patients with type 2 diabetes.The Swedish National Diabetes Reg- CD11c. The mesenteric compartment showed the most prominent increase ister (NDR) was linked with the national hospital discharge and mortality in the expression profi les. Lipolysis rates and FFA during hyperinsulinemia registries. We studied patients included in the NDR during 1998-2003 and correlated positively with the expression of osteopontin in all adipose tissue followed them until hospitalization for a primary or secondary diagnosis of compartments. Peripheral insulin sensitivity was not correlated to adipose HF or death or until 31 December 2009.In total 83,021 patients with type 2 tissue infl ammation, but correlated negatively with FFA.We show that both, diabetes (mean age 65.8 years, mean BMI 28.9 kg/m2) were included. During pro- and anti- infl ammatory markers are markedly present in all adipose tis- a median follow-up of 7.2 years, 10,969 patients (13.2 %) were hospitalized sue compartments of obese women compared to lean controls. Osteopontin with HF. In Cox regression analyses adjusting for age, sex, HbA1c, antidi- might play an important role in the higher lipolysis rate found in obesity. abetic medications, systolic and diastolic blood pressure, antihypertensive medications, diabetes duration, smoking, myocardial infarction, ischemic 1992-P heart disease, valve disease and atrial fi brillation, there was a signifi cant The Relationship between Overweight and Obesity With Hospital- association between BMI and hospitalization for HF (p<0.001). By catego- ization for Heart Failure in 20,985 Patients With Type 1 Diabetes: A ries of BMI, with BMI 20-25 kg/m2 as reference, hazard ratios (HR) for pa- Population-Based Study from the Swedish National Diabetes Reg- tients during follow-up were 1.15 (95% CI 0.97-1.35) for a mean BMI of <20.0 istry (NDR) kg/m2, 1.01 (95% CI 0.95-1.07) for BMI 25.0-<27.5 kg/m2, 1.18 (95% CI 1.11- DANIEL VESTBERG, ANNIKA ROSENGREN, MARITA OLSSON, SOFFIA GUDB- 1.25) for BMI 27.5-<30.0 kg/m2, 1.47 (95% CI 1.38-1.55) for BMI 30-<35 JÖRNSDOTTIR, ANN-MARI SVENSSON, MARCUS LIND, Trollhättan, Sweden, kg/m2, 2.06 (95% CI 1.90-2.23) for BMI 35-<40 kg/m2 and 3.23 (95% CI 2.88- Gothenburg, Sweden 3.61) for BMI 40 kg/m2 or higher.In conclusion, the risk of hospitalization for Hospitalization for heart failure (HF) is more common in patients with type HF in patients with type 2 diabetes is increased by 10%-25% in patients 1 diabetes than in the general population. The infl uence of body weight on with BMI 27.5-<30.0 kg/m2, approximately 50% at BMI 30-<35 kg/m2 and risk of HF in patients with type 1 diabetes has not been well studied.We 2-3 times higher at levels at or above 35 kg/m2. linked the Swedish National Diabetes Register (NDR) with data from the Swedish national hospital discharge register. Patients included in the NDR 1990-P during 1998-2003 and initially free of HF were followed until hospitalisation Depot-Specifi c Expression of Caveolin-1 in Human Adipose Tissue for HF or death or until 31 December 2009.In total 20,985 type 1 diabetic and their Relationship With Obesity and Insulin Resistance patients were studied (mean age 38.6 years; mean BMI 25.0 kg/m2 ). During LI XIAOFENG, LU HONGYUN, ZENG LONGYI, Guangzhou, China, Zhuhai, China median follow-up of 9.0 years 635 patients were hospitalized with HF as a This study aimed to explore the depot-specifi c mRNA and protein expres- primary or secondary diagnosis.The unadjusted incidence rates of hospital- sion of Caveolin-1 in human subcutaneous and omental adipose tissue, and ization for HF per 1000 person-years in different BMI categories were 3.85 then investigate their relationship with obesity and insulin resistance. Forty- (95% CI 2.69-5.50) for BMI35 kg/m2 .Cox regression adjusting for age, sex, one subjects with normal glucose regulation who underwent a selective diabetes duration, smoking, HbA1c, systolic and diastolic BP, microalbuminu- abdominal surgery were recruited in the study. Among them, 29 cases had ria and baseline and intercurrent co-morbidities (including myocardial infarc- normal body mass index(BMI) and 12 cases had BMI*24kg/m2 . The levels of tion) showed a signifi cant relationship between BMI and hospitalisation for fasting insulin(FINS) were measured by chemiluminescence ELISA kit. Fasting heart failure (p<0.001). With BMI 20-25kg/m2 as reference, the hazard ratio plasma glucose was tested by glucose oxidase mehtod and the home model (HR) for patients with BMI<20kg/m2 was 1.16 (95% CI 0.79-1.70), 1.02 (95% insulin resistance index(HOMA-IR) was calculated. Real-time PCR and west- CI 0.85-1.22) for BMI 25-30 kg/m2 , 1.75 (95% CI 1.36-2.24) for BMI 30-35 kg/ ern blot were employed to assess the relative mRNA and protein expression m2 and 3.43 (95% CI 2.28-5.18) for BMI >35kg/m2.In conclusion, obesity, in of Caveolin-1 in subcutaneous and omental adipose tissues.The mRNA and particular severe obesity, is a strong risk factor for hospitalisation for heart protein expression of Caveolin-1 from different fat depots were compared failure in patients with type 1 diabetes, whereas overweight and low body and their correlations with BMI and HOMA-IR were analyzed. We found that weight show no similar relations. FINS and HOMA-IR were signifi cantly higher in overweight and obesity group than those in the normal BMI group [FINS: (8.82±3.79)mU/L vs (6.43±4.38) 1993-P mU/L, P<0.05; HOMA-1R: 1.9l±0.85 vs 1.364±0.72, P<0.05]. The normal BMI group subjects had the higher expression levels of caveolin-1 mRNA and pro- tein expression in omental adipose tissue than overweight subjects(mRNA: WITHDRAWN 2.84±0.86 vs 1.02±0.36, P<0.01; protein expression: 1.68±0.67 vs 0.73±0.29, P<0.05), whereas those differences in subcutaneous adipose tissue was not signifi cant (P>0.05). The expressions of caveolin-1 mRNA were signifi cantly negatively correlated with BMI, waist circumstance, triglyceride and HOMA- IR in the omental adipose tissue (r=-0.441, -0.615, -0.539, -0.688,p<0.05), but not in subcutaneous adipose tissue. This study indicated that there is depot-specifi c expression of caveolin-1 in human subcutaneous and omental

adipose tissues.A low expression of caveolin-1 in omental adipose tissue may Obesity

contribute to the pathogenesis of obesity and IR. POSTERS Supported by: National Natural Science Foundation of China Integrated Physiology/ 1991-P The Expression of Pro- and Anti-Infl ammatory Markers in Adipose Tissue in Relation to Metabolic Fluxes in Obese Women BARBARA A. DE WEIJER, M. VAN EIJK, M.T. ACKERMANS, I.M. JANSSEN, F. BE- RENDS, K. KAASJAGER, A. VAN DE LAAR, A.P. HOUDIJK, E. FLIERS, M.J. SERLIE, Amsterdam, The Netherlands, Arnhem, The Netherlands, Alkmaar, The Netherlands Infi ltration and activation of adipose tissue macrophages contributes to low grade infl ammation and insulin resistance in obesity. Whether the expression profi les of infl ammatory markers in all adominal adipose tissue compartments correlate with metabolic fl uxes in humans is unclear.There-

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1996-P Changes in Plain Water Intakes and Long-Term Weight Gain in Women AN PAN, TAO HAO, DARIUSH MOZAFFARIAN, WALTER C. WILLETT, FRANK B. HU, Boston, MA 1994-P Plain water has been widely recommended to replace caloric beverages Liver Fat Not Visceral Fat Drives Insulin Resistance and Dyslipi- especially sugar-sweetened beverage (SSB) and fruit juices, which have demia Among Obese Hispanic Youth been associated with an increased risk of obesity and weight gain. In this TANYA L. ALDERETE, CLAUDIA M. TOLEDO-CORRAL, MARC J. WEIGENSBERG, study, we aimed to investigate changes in plain water intakes in relationto MICHAEL I. GORAN, Los Angeles, CA long-term weight changes, and estimate the effects of substituting plain The purpose of this study was to examine the separate and independent water for SSB and fruit juices in two large cohorts. We followed 43431 effects of liver fat and visceral adipose tissue on metabolic outcomes in women aged 40-65 in Nurses’ Health Study (NHS) from 1986 to 2006, and obese Hispanics.One hundred and eighty-fi ve Hispanics aged 8-25 years 44889 women aged 27-44 in NHS II from 1991 to 2007, without obesity and were examined. Visceral (VAT) and subcutaneous (SAT) adipose tissue vol- chronic diseases at baseline. Dietary intakes including various beverages ume and hepatic fat fraction (HFF) were measured by MRI. Total fat mass were assessed every 4 years using validated food frequency questionnaires. was assessed by DEXA. Insulin sensitivity (SI), acute insulin response (AIR), Body weight was self-reported every 2 years. Multivariate linear regression and fasting insulin were determined by FSIVGTT with minimal modeling. with robust variance and accounting for within-person repeated measures Triglycerides (TAG), low-density lipoprotein (LDL)-cholesterol, and high-den- were used to evaluate the association between changes in lifestyle behav- sity (HDL)-cholesterol were measured from fasting blood. Participants were iors (including beverage intakes) and weight changes within 4-year intervals. dichotomized on two factors: 1) high or low liver fat as defi ned by a hepatic The baseline median intake level of plain water was 2.5 cups/d, the mean fat fraction above or below 5.5% and 2) high or low VAT relative to SAT as changes in plain water intakes were 0.14 cup/d (5th to 95th percentile, -1.00 defi ned by positive or negative residuals from the regression between VAT to 1.50) for NHS women and 0.01 cup/d (-1.02 to 0.88) for NHS II women, and SAT. Two-way ANCOVA was used to examine the effect of high/low and the mean weight gains during each 4-year periods were 1.09 kg and liver fat and high/low VAT on metabolic outcomes. A prior signifi cance level 2.14 kg in NHS and NHS II, respectively. After adjustment for age, baseline was P<0.05. Covariates include age, sex, and total fat mass.Elevated liver BMI, and changes in other lifestyle behaviors (foods and beverages, smoking fat was associated with an 18% higher fasting insulin level (P<0.001), 46% habits, exercise, alcohol, sleep times, TV watching), we found that increased lower SI (P=0.01), 5% higher AIR (P<0.03), 3% higher TAG (P=0.02), and 8% plain water intake was associated with signifi cantly less weight gain (-0.12 higher VLDL-cholesterol (P<0.001) compared to low liver fat. High VAT was kg, 95% CI, -0.13 to -0.11, for 1 cup/d increase in water intake within each associated with a 7% higher fasting insulin level (P=0.04) and a 3% higher 4-year period). We estimated that replacement of 1 serving/d of SSB and AIR (P<0.03) compared to those with low VAT. High VAT had no effects on fruit juices by 1 cup/d of water was associated with 0.45 kg and 0.30 kg SI, TAG, or VLDL-cholesterol. VAT signifi cantly modifi ed the effect of liver fat less weight gain over 4-year period, respectively. In conclusion, our results on HDL-cholesterol (P=0.05). Among those with low VAT, high liver fat was suggest that increasing plain water intake at the expenses of SSB and fruit associated with a 4% lower HDL-cholesterol (P<0.01) while liver fat had no juices is associated with lower long-term weight gain. effect among those with high VAT (P=0.54).These results provide evidence that elevated liver fat, not VAT, is the main ectopic fat depot driving de- 1997-P creased insulin sensitivity and dyslipidemia among obese Hispanics. These Low Level of IL6 Are Predictor of Development of Hepatic Steatosis results suggest that high liver fat, in the context of lower VAT, may be more during a 2 Month High Fat Overfeeding in Healthy Men detrimental to metabolic health. MAUD ALLIGIER, EMMANUELLE MEUGNIER, DEBARD CYRILLE, STÉPHANIE Supported by: P60MD002254, R01DK059211, NIH NCMHD P60 MD00254, Atkins LAMBERT, BÉATRICE MORIO, HUBERT VIDAL, MARTINE LAVILLE, Oullins, France, Foundation Clermont-Ferrand, France Obesity is associated with a cluster of metabolic abnormalities, including 1995-P increase in hepatic lipid content. The mechanisms leading to lipid accumula- The Relationship between Thyroid Function, Insulin Resistance, In- tion in the liver are poorly understood. We studied the early mechanisms of fl ammation, Visceral Obesity and Brown Adipose Tissue Detected weight gain and lipid deposition in liver in humans during controlled over- by FDG-PET/CT: A Prospective, Matched Case-Control Study feeding with a lipid rich diet (excess of 760 Kcal/day).Abdominal fat mass HYE JIN YOO, JAE HEE AHN, HAE YOON CHOI, HO CHEOL HONG, NAM HOON repartition and hepatic lipid content were assessed by MRI in 44 healthy KIM, YOON JUNG KIM, JOO HYUNG KIM, CHAI RYOUNG EUN, SAE JEONG male subjects before (D0) and after overfeeding (D56). Metabolic and in- YANG, HEE YOUNG KIM, JI A. SEO, NAN HEE KIM, SIN GON KIM, SEI HYUN BAIK, fl ammatory (IL6, CRP, TNF_) markers were measured in plasmaBody weight DONG SEOP CHOI, KYUNG MOOK CHOI, Seoul, Republic of Korea (+2.5kg, p=0.001) and total fat mass (p=0.001) increased during overfeed- Recent studies have shown that adult human possesses brown adipose ing. Mild insulin resistance (+0.15, p=0.05) occurred after the diet. Visceral tissue (BAT), which might have an active role in regulating energy expen- and subcutaneous fat increased at D56. Hepatic lipid deposition displayed diture and adiposity. In this prospective, matched case-control study, we large inter-individual differences (from -21% to +70%). Increase hepatic lipid evaluated whether the presence of BAT are associated with thyroid func- storage correlated with initial plasma levels of IL6 (r= -0.5, p=0.001). No re- tion, insulin resistance, infl ammation and visceral obesity in adult humans. lationship was found with other measured parameters.Our results suggest We enrolled 40 BAT positive and 40 BAT negative subjects matched in age, that low level of IL6 could be a risk marker for the development of hepatic gender, use of levothyroxine or `-blocker medication and season/year per- steatosis during overfeeding in healthy subjects. Because obesity is gener- forming 18F-FDG positron emission tomography/computed tomography (FDG- ally related to increased levels of IL6, this observation is surprising. However PET/CT). All the follow-up measurements were performed after median 23.0 IL6 knockout mice displayed hepatosteatosis and patients treated with IL6 (16.0, 31.0) months.Among total of 4,736 consecutive FDG-PET/CT scans in receptor blocker showed fasting hypertriglyceridemia, suggesting lipid me- 4,736 subjects, positive scans were seen in 146 subjects (3.1%). The BAT de- tabolism defect in liver.

Obesity tected group and matched undetected group did not show signifi cant differ- POSTERS ence in waist circumference and other components of metabolic syndrome 1998-P (all p >0.05). Furthermore, visceral fat area (VFA) and subcutaneous fat area Generation of Human Adipose Tissue Foam Cells Correlates With

Integrated Physiology/ measured using abdominal CT were also not different (p = 0.985 and p = Clinical Parameters Associated With the Metabolic Syndrome 0.544,respectively). When stratifi ed by VFA tertile, BAT positive rate was HAGIT SHAPIRO, RUTHY SHAKO-LEVI, TANYA TARNOVSCKI, TAL PACHT, BORIS not different among VFA tertile groups (p = 0.964). Furthermore, freeT4 (p = KURSHTEIN, NAVA BASHAN, MATTHIAS BLÜHER, ASSAF RUDICH, Beer-Sheva, 0.982), TSH (p = 0.964), homeostasis model assessment of insulin resistance Israel, Leipzig, Germany (HOMA-IR) (p = 0.172) and high-sensitivity C-reactive protein levels (hsCRP) Infl ammation is one mechanism proposed to link obesity with diabetes. (p = 0.693) were not signifi cantly different between groups.The present Macrophages accumulation in adipose tissues is proportional to obesity and study do not support that the presence of BAT detected by FDG-PET/CT in suggested to contribute to insulin resistance and diabetes. We discovered thermoneutral condition is associated with thyroid function, insulin resis- in histological sections of human fat biopsies a sub-group of macrophages tance, infl ammation and visceral obesity in adult women. loaded with lipids which resemble atherosclerotic foam cells. Here we in- vestigated whether adipose tissue foam cells (ATFC) correlate with clinical parameters associated with the metabolic syndrome. ATFC accumulated in

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A510 OBESITY—HUMANCATEGORY crown-like structures surrounding the adipocytes in omental fat biopsies of tant (IR) based on AUCINS below or above the median, there were no differ- obese patients. Omental ATFC signifi cantly increased with BMI (spearman ences in demographic or physical parameters; but the plasma cholesterol, correlation, p: 0.0181, n=25). Among obese non-diabetic persons, increased LDL, VLDL, AUCINS and AUCFFA were all higher (p < 0.05) and serum Klotho number of ATFC were found in omental depots with intra-abdominal (central) was lower (p = 0.019) in the IR group (Table 1). Our data suggests that lower fat distribution compared with subcutaneous adiposity (3.4 ±0.4 fold, n=11 serum concentration of Klotho could be a marker of impaired lipid and glu- and 12 for central and subcutaneous adiposity, respectively). Importantly in cose metabolism commonly observed in obese humans. Future studies are another group of patients, elevated number of ATFC were detected in diabet- required to establish a potential mechanistic effect of circulating Klotho on ic compared with non-diabetic patients (2.5 ±0.3 fold, n=11 and 15, respec- cardiovascular risk factors in obesity. tively). Using fl ow cytometry to quantify ATFC, we gated on CD45+, CD14+, CD64+ cells and analyzed their lipid content by Bodipy staining for neutral lipids. ATFC were more abundant in obese males and signifi cantly increased in omental compared with subcutaneous adipose tissues (2.6 ±0.3 fold in mean lipid content, p: 0.012, n=18 and 14, respectively).In conclusion, we discovered in human adipose tissues formation of foam cells in crown-like structures surrounding the adipocytes. Foam cells were abundant in omental adipose tissues and correlated with males, BMI, intra-abdominal adiposity and diabetes, which are clinical parameters associated with the metabolic syndrome. We propose that the foam cells participate in clearance of lipids from dead adipocytes accumulated in response to nutritional overload in obesity and its linked morbidities.

1999-P TNF-ƴ and IL-6 Induces Caspase and Inhibits BCL2 in Human Adi- pose Tissue: A Potential Relationship With Obesity and the Onset of Insulin Resistance FRANCISCO TINAHONES, LETICIA COÍN, MORA MURRI, WILFREDO OLIVA OLI- VERA, MARIA DOLORES MAYAS TORRES, EDUARDO GARCÍA FUENTES, NURIA BARBARROJA, RICARDO GÓMEZ HUELGAS, MARÍA M. MALAGÓN, RAJAA EL BEKAY, Málaga, Spain, Córdoba, Spain Cell death is proposed to regulate adipose growth and development, which determines the onset of obesity and associated insulin resistance (IR). The aim of our study was to analyze the relationship between obesity, adipose tissue (AT) apoptosis, infl ammation status and insulin signaling.The expression levels of initiator (CASP9, CASP8) and effector (CASP3, CASP7) caspases as well as the anti-apoptotic BCL2 and infl ammatory makers were assessed in visceral (VAT) and subcutaneous (SAT) human AT from patients with different degrees of obesity and lacking IR or diabetes. To mimic the infl ammatory state evoked by weight gain, we analyzed AT explants from healthy lean subjects that were cultured in the presence of TNF-_ or IL-6 and the mRNA expression levels of apoptotic mediators and insulin-signaling pathway components. Gene expression data from cytokine-treated explants were then compared to basal mRNA expression levels in AT samples ob- tained from morbidly obese subjects.SAT and VAT exhibited increased ex- pression levels of CASP3, CASP7, CASP8 and CASP9, and decreased mRNA expression of BCL2 with weight gain. These changes were accompanied by increased infl ammatory cytokine mRNA levels and markers of macrophage infi ltration. In obese subjects, activation of CASP3/7 and down-regulation of BCL2 correlated with the expression levels of IRS 1 and 2. Moreover, gene expression levels of caspases, BCL2, p21, p53, IRS-1, IRS-2, GLUT4, PTP1B and LAR in TNF-_ or IL-6-treated explants from healthy lean subjects were comparable to those found in freshly isolated AT samples from obese sub- jects.We show that weight gain is associated with CASP3/7 activation and repression of BCL2 expression in AT, in a process that is most likely driven Supported by: NIH/NIDDK (072158) by the infl ammatory conditions associated with obesity. Furthermore, our results support the view that this pro-apoptotic state correlated with insulin 2001-P signaling and may contribute to the development of IR. Retinol Binding Protein 4 (RBP4) Expression in Adipose Tissue and Supported by: Spanish Ministry of Health (FIS) and Consejería de Innovación Liver in Relation to Metabolic Fluxes in Obese Women BARBARA A. DE WEIJER, M. KILICARSLAN, H. CAKIR, M. VAN EIJK, M. ACKER- MANS, I. JANSSEN, F. BERENDS, A. VAN DER LAAR, A.P. HOUDIJK, E. FLIERS, 2000-P B.B. KAHN, M.J. SERLIE, Amsterdam, The Netherlands, Arnhem, The Netherlands, Circulating Klotho and Cardiovascular Risk Factors in Obese Hu- Alkmaar, The Netherlands, Boston, MA Obesity

mans RBP4 has been proposed to be an adipokine that is involved in obesity- POSTERS DEMIDMAA TUVDENDORJ, MANISHA CHANDALIA, NICOLA ABATE, Galveston, induced insulin resistance. RBP4 is primarily synthesized in hepatocytes, TX but also in other sites including adipocytes. RBP4 mRNA is increased in Lower circulating Klotho has been proposed to associate with higher visceral and subcutaneous adipose tissue of obese subjects. The relation- Integrated Physiology/ likelihood of having cardiovascular disease. However, whether it may affect ship between RBP4 expression in different adipose tissue compartments lipid and glucose metabolism in humans is not known. To clarify this question and lipolysis, and RBP4 expression levels in liver of obese subjects without we measured serum Klotho concentration in 25 obese normoglycemic adults liver disease are not known.We studied RBP4 expression in 3 adipose tissue (age: 47±15 years, BMI: 35±6 kg/m2, fasting glucose: 97±8 mg/dl; Mean±SD). compartments (omental, mesenteric and subcutaneous) and liver in obese During OGTT, the area under curve for fatty acid (AUCFFA, a measure of adi- women undergoing bariatric surgery. We compared expression profi les with pose tissue insulin resistance) and for insulin (AUCINS, a measure of periph- lean controls undergoing elective cholecystectomy. In the obese group, we eral insulin resistance) were obtained. Lower serum Klotho concentration assessed endogenous glucose production (EGP) and lipolysis using stable 2 signifi cantly associated with higher AUCFFA (r = 0.32, p = 0.002) (Figure 1). isotopes 2 weeks before surgery.We included 16 Caucasian women (mean When we compared two groups, defi ned as insulin sensitive (IS) and resis- BMI 45 [39-51] kg/m2) and 6 lean normal glucose tolerant women (mean BMI

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A511 OBESITY—HUMANCATEGORY

22 [20-24] kg/m2).RBP4 expression in liver was higher in obese versus lean LDLc, HDLc, triglycerides, AST, ALT, and the presence of hepatic steatosis by women (fi g.1) but did not correlate with EGP (mean 13.6 (10.3-18.1) μmol/ ultrasound were compared between two groups. The prevalence of MS was kgFFM/min). RBP4 expression in subcutaneous (p=0.01) and mesenteric fat 83% (88/106). Women with MS were older 47.3±11.8 vs. 37.2 ±11.0 years, (p=0.058) was higher in obese versus lean women but did not correlate with p=0.01). The BMI and the waist was not different between groups (34.8 ± fasting lipolysis rates (mean 3.0 (1.9-5.0) μmol/kg.min) or plasma FFA (mean 6.9 vs.35.6 ± 8.0 Kg/m2 and 106.8 ± 13.0 vs. 107 ± 17.0 cm), however, the 0.81 (0.66-1.13) mmol/L).In conclusion, obesity is associated with higher waist/hip ratio was higher (0.93 ± 0,.6 vs. 0.90 ± 0.06, p=0.04), and the hip RBP4 expression in liver in women.The increased levels of RBP4 in adipose circumference lower in the MS group (114 ± 12 vs. 121 ± 17 cm (p=0.04). Cen- tissue are not correlated with the lipolytic fl ux in obese women. tral obesity was the most frequent component of MS (94.4%), followed by high blood pressure (77.8%), hyperglycemia (44.4%), high triglycerides levels (32.6%) and low HDL (30.6%). Except one patient with hepatic steatosis had no MS. The Tg/HDL ratio was higher in patients with MS (3.46 ± 2.2 vs. 1.99± 0.6 mg/dL, p=0.007). Using MS as endpoint, the prevalence ratio was 19,5; CI(95%)=(2,4-152,8) for hyperglycemia; 4,55; CI(95%)=(1,5-13,6) for high blood pressure, 1,3; IC (95%)=(1,1-1,5) for high triglycerides levels. The best variables to predict MS. The best variables to predict MS using ROC curve was blood glucose (AUC = 0.69; p = 0.002), triglycerides (AUC = 0.709; p = 0.014), PAS (AUC = 0.724; p=0,009) and PAD =0.741; p=0,005). In conclusion, in this population, the waist/hip ratio was more important than waist cir- cumference or BMI to predict MS. The highest hip circumference in women without MS suggests a protective effect of hip fat. Elevated blood glucose was the best component to predict MS. An elevated Tg/HDL ratio and he- patic steatosis were also strongly associated with MS. Supported by: FAPESB

2004-P Both Lifetime Maximum BMI and BMI Change Since Age 20 Years to Maximum Were Strongly Associated With having Undiagnosed Diabetes in Japanese Women: Toranomon Hospital Health Manage- 2002-P ment Center Study Inverse Relationship between Serum Osteocalcin Level and Vis- SAKIKO YOSHIZAWA, YORIKO HEIANZA, YASUJI ARASE, HIROSHI TSUJI, KA- ceral Fat Area in Chinese Men ZUMI SAITO, SHIUN DONG HSIEH, SATORU KODAMA, NOBUHIRO YAMADA, DOU JIANXIN, MA XIAOJING, HAO YAPING, WANG FEIFEI, YANG RONG, TANG SHIGEKO HARA, HIROHITO SONE, Mito, Japan, Tokyo, Japan JUNLING, XIAO YUNFENG, BAO YUQIAN, JIA WEIPING, Shanghai, China Although lifetime maximum BMI and BMI change since age 20 years were Osteocalcin has been reported to play an important role in regulating suggested to be useful to identify individuals with undiagnosed diabetes, glucose and fat metabolism. We investigated the relationship between the usefulness of these and other parameters on lifetime BMI change have serum osteocalcin level and visceral fat area (VFA) in Chinese men.Serum not been directly compared. Cross-sectionally investigated were 5201 Japa- osteocalcin concentration was measured by electrochemiluminescence im- nese women aged 21-87 years without a self-reported history of clinician- munoassay in 511 men (aged 36 to 65 years). Study subjects were divided diagnosed diabetes. We estimated the probability of having undiagnosed into four groups according to quartiles of serum osteocalcin level. VFA and diabetes (diabetes indicated by fasting plasma glucose level *7.0 mmol/L subcutaneous fat area (SFA) were quantifi ed by magnetic resonance imag- and/or HbA1c *6.5%) for the following obesity indices: current BMI, BMI at ing. The association between serum osteocalcin and VFA and other clinical age 20 years (BMI20y), lifetime maximum BMI (BMImax), BMI change since parameters including components of metabolic syndrome (MS) was exam- age 20 years to maximum (BMI620y-max), and BMI change since age 20 ined. The MS was diagnosed by the 2007 Joint Committee for Developing years to current (BMI620y-cur). Prevalence of undiagnosed diabetes was Chinese Guidelines on prevention and treatment of dyslipidemia defi nition. 1.7% among all subjects. After adjustment for age and lifestyle factors, odds With the increment of serum osteocalcin there was a decreasing trend in ratios (ORs) for undiagnosed diabetes for each 1-SD increment in current body mass index (BMI), %fat, waist circumference, VFA, diastolic blood pres- BMI and BMImax were 1.67 (95% CI: 1.41-1.98) and 1.40 (1.19-1.65), respec- sure, fasting and 2-h glucose (all p for trend <0.05), as well as the frequency tively. However, BMI20y was not signifi cantly associated with undiagnosed of the MS (p<0.01). Compared with subjects in the lowest quartile group diabetes. When we assessed BMI change since age 20 years, ORs for each (osteocalcin<12.75 ng/ml), those in the third (osteocalcin16.17-19.80ng/ml) 1-SD increment in BMI620y-max and BMI620y-cur were 1.61 (1.38-1.88) and the highest quartile (osteocalcin*19.80ng/ml) groups had signifi cantly and 1.56 (1.29-1.89), respectively. As to the combination of BMImax and lower VFA (85.03 (25th-75th, 49.28-124.54) vs 106.33 (66.61-137.48), 85.28 BMI620y-max in identifying undiagnosed diabetes, individuals with BMI- (46.51-119.98) vs 106.33 (66.61-137.48) cm2, respectively, both p<0.05). Se- max *24.2 kg/m2 and BMI620y-max *3.9kg/m2 had a markedly higher OR rum osteocalcin level was inversely correlated with fasting and 2-h glucose, of 3.00 (1.94-4.64) compared with other individuals. Our results showed that hemoglobin A1c, total cholesterol and VFA after adjusted for age and BMI both BMImax and BMI620y-max were most strongly associated with having (all p<0.05 ). Multiple stepwise regression analysis showed that VFA and 2-h undiagnosed diabetes among those investigated. This combination could be glucose were independently associated with serum osteocalcin level after a valid marker to identify diabetic individuals who were not aware of their adjustment with age, %fat, SFA, hemoglobin A1c, triglycerides, low density condition. lipoprotein cholesterin, HOMA-B, HOMA-IR, current smoker (all p<0.01). Serum osteocalcin level was inversely correlated with VFA as well as with 2005-P components of the MS in Chinese men. Platelet Activation and Adhesion in Morbidly Obese Patients Un- Supported by: The National Natural Science Funds of China (81170788) Obesity dergoing Weight Loss Induced by Roux-en-Y-Bypass

POSTERS STEPHAN BLECHINGER, RICHARD KALTENBOECK, MIRIAM SATLER, MARIE EL- HENICKY, CLEMENS HOEBAUS, FLORIAN HOELLERL, JOHANNA M. BRIX, HANS 2003-P Integrated Physiology/ P. KOPP, GUNTRAM SCHERNTHANER, GERIT H. SCHERNTHANER, Wien, Austria Hip Circumference: A Protective Factor to Metabolic Syndrome in Morbidly obese (MO) patients are at increased risk for cardiovascular dis- Overweight Women? ease, events and death. 30% of the added risk still needs to be explained. MARIA D. LIMA, ANA M. LADEIA, LUCAS OLIVIERI, PAULO GOES, ARMÊNIO Apart from an elevated endogenous thrombin potential in MO, we hypothe- GUIMARÃES, Salvador, Brazil size that platelets exhibit more activation and adhesion molecules.We inves- Metabolic syndrome (MS) is a cluster of obesity, abnormal glucose, dys- tigated 18 MO patients (15 female) before and after weight loss (observation: lipidemia and hypertension. Several studies associated MS with increased 17.5±2.7 months). We obtained expression of platelet activation and adhe- cardiovascular disease. This sectional study aimed to compare the meta- sion molecules CD31, CD36, CD42b, CD49b, CD49f, CD62P, CD63 and CD154 bolic profi le between overweight women with and without metabolic syn- by multi-colour fl ow cytometry of freshly isolated platelet rich plasma. A drome . In 196 overweight women (BMI *25 Kg/m2), waist circumference, p-value <0.05 was considered signifi cant. To investigate changes of mol- hip, waist/hip ratio, BMI, blood pressure, fasting blood glucose, cholesterol, ecules during weight loss and their associations, deltas were calculated.We

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A512 OBESITY—HUMANCATEGORY are fi rst to report a reduction of adhesion molecules CD31 and CD49b: mean lipids reported to be implicated in insulin resistance. The purpose of this fl uorescence intensity: CD31: 157±29 vs 145±28, p=0.007; CD49b: 198±80 vs study was to examine the effects of gastric bypass surgery (GPS) induced 151±52, p=0.009. CD42b, CD62p and CD154 show a trend. CD36, CD49f and weight-loss, both with and without exercise training, on insulin sensitiv- CD63 are not different. Preoperative CD31 is associated with fasting Insulin ity and muscle sphingolipid and diacylglycerol (DAG) content. 1-3 months (Beta=0.45), after weight loss with LDL-C (B=0.42), 2-hour Insulin (B=0.57). following GPS, subjects were randomized to 6 months of either moderate Expression of CD49b is associated with triglyceride (B=0.44), fasting Insulin structured exercise (EX; n=18) or weight-loss-only control (CON; n=14). Insu- (B=-0.48), HOMA-IR (B=-0.44), fi brinogen (B=0.46) preoperative and weight lin sensitivity was determined by intravenous glucose tolerance test (IVGTT) (B=-0.47), BMI (B=-0.55), CRP (B=-0.40) postoperative. Delta CD31 was asso- and minimal modeling, and percutaneous biopsies of the vastus lateralis ciated with 1-hour glucose (B=0.55) and delta CD49b with HDL-C (B=-0.43), were obtained, before and after the 6-month interventions. Sphingolipid triglyceride (B=0.48), fasting Insulin (B:-0.59), HOMA-IR (B:-0.60), fi brinogen and DAG content of muscle were determined by high-performance liquid (B:0.46).Although sheded surface molecules associated with infl ammation chromatography-tandem mass spectrometry. Body composition was deter- are reduced after weight loss, their origin seem not to be platelets. We can mined by DEXA. Both groups lost a similar amount of weight (-23.5±1.5 kg) only report a marked reduction of two adhesion related molecules (CD31, and body fat (-20.6±1.08 kg). The addition of exercise resulted in signifi cantly CD49b), but failed to reach signifi cance for infl ammatory molecules (CD62P, greater improvements in the insulin sensitivity index (SI) compared to CON CD63, CD154). Whether the previously reported sheded molecules have their (+3.09±0.68 vs.+1.06±0.58, P=0.037) Total muscle sphingolipid content was origin in the endothelium or maybe even in the liver needs to be elucidated. reduced (P<0.01) to a similar extent in CON (-1.86±0.65 pmol/mg) and EX (-1.80±0.50pmol/mg). Total unsaturated and saturated muscle ceramide con- 2006-P tent was also lowered to a similar extent in EX and CON. Total muscle DAG Association of the Obesity/Type 2 Diabetes-Related Genetic Vari- content was not signifi cantly altered with weight loss or exercise. In con- ants With Visceral Fat Accumulation in Obese, Overweight and clusion, regular exercise performed during extensive weight loss following Lean Subjects gastric bypass surgery results in greater improvements in insulin sensitivity ADAM KRETOWSKI, NATALIA WAWRUSIEWICZ-KURYLONEK, ANNA CITKO, JO- compared to weight loss alone. These improvements in insulin sensitivity, ANNA GOSCIK, EDYTA ADAMSKA, KATARZYNA MALISZEWSKA, MARIA GÓR- however, did not appear to be correlated with changes in either intramyocel- SKA, Bialystok, Poland lular ceramides or DAG. It has been recently suggested the genetic predisposition to obesity Supported by: NIH/NIDDK (DK078192-01A1) to B.H.G. leads to increased risk of type 2 diabetes by its obesity predisposing ef- fect. Visceral fat accumulation has an important role in increasing the risk 2008-P of developing metabolic disorders, such as type 2 diabetes, dyslipidemia and Beloranib Improves Cardiometabolic Risk and Reduces Fat Mass in CVD, but data concerning the genetic background of body fat distribution Women Completing Two Phase I Trials are limited. The aim of our study was to analyze whether there are spe- THOMAS E. HUGHES, JAMES E. VATH, JOE PROIETTO, JOANNE MARJASON, cifi c genetic factors that infl uence the risk of visceral fat accumulation. We DENNIS D. KIM, Cambridge, MA, Heidelberg, Australia, Herston, Australia genotyped 64 SNPs within 40 genes associated with obesity and/or type Beloranib is a fumagillin class methionine aminopeptidase 2 inhibitor that 2 diabetes (identifi ed by GWAS ) in 468 overweight/obese patients and reduces food intake and causes long term weight loss in obese animals by 245 healthy volunteers with normal weight (302 men and 411 women), who increasing triglyceride mobilization and ketogenesis. The effects of twice underwent anthropometry (BMI, WHR) and body composition analysis: weekly intravenous beloranib on weight loss, cardiometabolic risk fac- body fat percentage, visceral (VAT) and subcutaneous abdominal adipose tors, metabolic syndrome, and body composition were evaluated in obese tissue (SAT) by multi-frequency bio-impedance method. The association of women completing at least 25 days of treatment (7 doses) from 2 Phase I visceral fat content and VAT/SAT ratio with the distribution of genotypes studies. Data are presented for completers treated with 0.9 mg/m2, 3 or 6 of the following SNPs: MC4R rs1350341 (p=005; p=0,015) (Figure 1.), MC4R mg beloranib (beloranib N=17) or placebo (N=11). A total of 34 subjects were rs17782313 (p=0,035, p=0,002), PPARG rs1801282 (p=0,02, p=0,0019); BDNF enrolled into these treatment arms. Baseline characteristics of completers rs6265 (p=0,034, p=0,006) and SH2B1 rs7498665 (p=0,042, p=0,0038) has (mean±SD): 93% Caucasian, 49.2±8.9 years, 102.6±12.5 kg bodyweight, and been found in the studied subjects. Our results suggest that there are VAT- 39.1±3.7 BMI. Weight loss was greater with beloranib (mean±SE: 4.3±0.4 kg; specifi c genetic factors that infl uence the risk of visceral fat accumulation. p<0.001, paired t test for change from baseline) than with placebo (-0.6±0.5 The identifi ed genetic variants may help to understand the mechanisms of kg, NS). Beloranib-treated subjects also showed improvements in cardiomet- body fat distribution and serve as early biomarkers of increased risk of meta- abolic risk factors including reduced triglycerides (mean±SE: -34.4±8.8 mg/ bolic disorders development in overweight/obese patients. dL), LDL cholesterol (-34.1±8.0 mg/dL), waist circumference (3.9±0.9 cm), and diastolic blood pressure (-4.5±2.1 bpm; all p<0.05, paired t test for change from baseline), whereas there was no change with placebo. There was a trend for beloranib to be associated with a reduction in the percentage of subjects with metabolic syndrome that did not reach statistical signifi cance. Body composition, measured in one study, (N=9 beloranib 3 and 6 mg, N=8 placebo) indicated expected reduction in fat mass with beloranib. The most common AEs were nausea, infusion site injury and headache. Most were of mild/moderate intensity and tended to be self-limiting. Among women com- pleting at least 25 days of treatment, intravenous beloranib was associated with weight loss and corresponding reductions in many cardiometabolic risk factors.

2009-P Glucose and Cholesterol Metabolism after Bariatric Surgery in

Grade-3 Obesity: Differences Between Malabsorptive and Restric- Obesity

tive Surgery POSTERS ALBERTO BENETTI, MARINA DEL PUPPO, ANDREA CROSIGNANI, ANNAMARIA Supported by: National Science Center Grant (M.G.) VERONELLI, ENZO MASCI, LUCIA MIELE, GIANCARLO MICHELETTO, ANTONIO E. Integrated Physiology/ PONTIROLI, Milano, Italy 2007-P Malabsorptive bariatric surgery (biliopancreatic diversion and biliointes- Effects of Bariatric Surgery-Induced Weight-Loss With or Without tinal by-pass) reduces serum cholesterol levels more than restrictive sur- Exercise on Insulin Sensitivity and Muscle Sphingolipid Content gery (gastric banding), and this is thought to be due to the greater weight PAUL M. COEN, CHARLES J. TANNER, NICOLE L. HELBLING, GABRIEL S. DUBIS, loss. Our aim was to evaluate glucose metabolism [blood glucose and serum MAJA STEFANOVIC-RACIC, GEORGE M. EID, JOSEPH A. HOUMARD, BRET H. insulin levels, and HOMA index], and cholesterol metabolism [absorption: GOODPASTER, Pittsburgh, PA, Greenville, NC serum campesterol and sitosterol levels; synthesis: serum lathosterol lev- The effects of regular exercise on insulin sensitivity following bariatric els; catabolism: rate of appearance of serum 7-alpha-OH and serum 27-OH- surgery are unknown. Moreover, it is not known whether these improve- cholesterol after infusions of deuterated 7-alpha- and 27-hydroxycholesterol ments in insulin sensitivity are associated with changes in intramyocellular in sequence] in grade-3-obesity subjects undergoing bilio-intestinal by-pass

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A513 OBESITY—HUMANCATEGORY

(n=6) and gastric banding (n=8). Evaluations were performed before and 6 quantitative meta-analysis of 3 cohort studies indicated that a 100 g/day in- months after surgical procedures. Subjects had similar values at baseline. cremental increase in terms of milk intake was associated with a 2.1% [95%CI, Weight loss was similar in the two groups of subjects; blood glucose, insulin 0.7-3.5%] reduction in future MetS risk. This meta-analysis suggests that daily levels, and HOMA decreased in a similar way; in contrast, serum cholesterol dairy intake lowers the risk of future MetS, although excessive intake of dairy and LDL-cholesterol levels, together with serum sitosterol and campesterol products leading to high-energy intake should be avoided. levels, decreased, and lathosterol levels increased only in bilio-intestinal by-pass subjects, not in gastric banding subjects. A signifi cant increase in 7-alpha-OH- cholesterol appearance occurred only in by-pass surgery. These data indicate that malabsorptive surgery specifi cally affects cholesterol lev- els, independently of weight loss, and independently of glucose metabolism and of insulin resistance; a decreased sterol absorption leads to decreased cholesterol and LDL-cholesterol levels, accompanied by enhanced choles- terol synthesis, and by enhanced cholesterol catabolism. Malabsorptive sur- gery seems to be indicated in grade-3 obese subjects with raised cholesterol levels. Supported by: Ministero dell’Istruzione dell’Università e della Ricerca (Italy)

2010-P Vitamin D Defi ciency as a Potential Risk Factor for Insulin Resis- tance in Healthy Women TATIANA KARONOVA, ELENA GRINEVA, ELENA MICHEEVA, OLGA BELYAEVA, ELENA TSVETKOVA, ELENA BAZHENOVA, OLGA GALKINA, EUGENE SHLYAKHTO, St. Petersburg, Russian Federation Some studies suggest that serum 25(OH) vitamin D level could be as- sociated with fat mass quantity and glucose metabolism parameters. However the data seems to be contradictory. We examined the interlinks between serum 25(OH)D with body mass and fat mass indices, glucose as well as insulin levels, HOMA-IR, and ISI (0,120) in healthy women.We ex- amined 320 healthy late reproductive age women 40 to 52 years (mean age 46.1±4.5). The mean waist circumference was 92.4±1.0 cm, waist-to-hip ratio 2012-P (WHR) - 0.86±0.01, body mass index (BMI) was 30.2±6.1 kg/m2. We used Glycemic Control and Hypoglycemia Prevalence in Japanese Par- DEX in 134 women to determine fat mass and calculate fat mass index ticipants With Type 2 Diabetes According to BMI: A Sub-Analysis (FMI). These parameters were 42.5±0.6% and 12.7±0.3 kg/m2 accordingly. of the ALOHA Data Serum 25(OH)D and insulin levels were determined by ELISA, plasma glu- MASATO ODAWARA, EDWARD WANG, JAY LIN, KARIM ADMANE, TAKAYUKI cose (PG) levels - by standard biochemistry. Insulin resistance was evaluated ITO, TAKASHI KADOWAKI, Tokyo, Japan, Bridgewater, NJ, Flemington, NJ, Paris, by HOMA-IR and insulin sensitivity index (ISI -0,120).Fasting PG level was France 5.9±0.1 mMol/L, insulin level 11.5±0.6 (mIU/ml), HOMA-IR - 3.2±0.1 and ISI The aim of this analysis was to determine the reduction levels of A1C and (0,120) - 7.78±0.27. Serum 25(OH)D level was from 19.4 to 134.1 nMol/L (mean prevalence of hypoglycemia among Japanese people with type 2 diabetes 52.9±22.7). Vitamin D insuffi ciency and defi ciency (lower than 75 nMol/L) (T2DM) according to body mass index (BMI).Patients with T2DM who required was revealed in 86.8% of healthy women.Correlation analysis did not show insulin with oral antidiabetic drugs (OADs) and met the inclusion criteria were correlation between 25(OH)D level and glucose metabolism parameters in observed for 24 weeks in a real-life setting. Baseline and endpoint A1C, fast- total population, however we found inversed correlations between serum ing blood glucose (FPG), postprandial plasma glucose (PPG), body weight, and 25(OH)D and fasting insulin level (r=-0.26, p<0.01), and PG and insulin levels BMI were measured.BMI was *25 kg/m2 (mean [SD] 27.2 [1.4] kg/m2) in 1410 2 hours after GTT (r=-0.31, p<0.01 and r=-0.4, p<0.01 accordingly) and ISI- patients and <25 kg/m2 (21.8 [2.2] kg/m2) in 2496 patients. A1C levels were re- 0,120 (r=0.28, p<0.01) in obese patients. We found that Vitamin D level lower duced signifi cantly in both BMI groups (P < 0.001), and there was a signifi cant than 50 nMol/L was associated with obesity (OR 2.1; 95%) and diabetes difference in A1C between groups (*25 kg/m2: mean -1.27 (1.27)%; <25 kg/m2: type 2 (OR 1.67; 95%).Our results showed that vitamin D defi ciency is highly mean -1.50 (1.22)%; P < 0.001). A signifi cantly greater percentage of patients prevalent in healthy women population and low Vitamin D level is correlated with a low BMI achieved a target A1C of <7% (*25 kg/m2: 13.22%; <25 kg/m2: with high fat mass, PG level and decreased tissues insulin sensitivity. Hence 18.33%; P < 0.001) and the incidence of severe hypoglycemia was low in both vitamin D insuffi ciency might possibly be a risk factor for obesity and insulin groups (*25 kg/m2: 0.07%; <25 kg/m2: 0.12%; P = 0.64). A signifi cantly lower resistance development leading to diabetes type 2. dose of insulin glargine was used in the low BMI group (*25 kg/m2: mean 10.31 [7.03] U/day); <25 kg/m2: mean 8.71 [5.27] U/day); P < 0.001). The insulin 2011-P glargine with OADs regimen resulted in a signifi cant reduction in FBG and PPG Relationship between Dairy Intake and Risk of Metabolic Syn- levels and an increase in body weight and BMI in patients with a low BMI. drome: A Meta-Analysis In general, patients with a high BMI had a greater occurrence of prior com- CHIKA HORIKAWA, SATORU KODAMA, YORIKO HEIANZA, AYUMI SUGAWARA, plications.The insulin glargine with OADs regimen resulted in signifi cant A1C MIHO MAKI, SAKIKO YOSHIZAWA, YOKO YACHI, KAZUMI SAITO, HIROHITO reductions with a low rate of severe hypoglycemia and weight gain in both SONE, Mito, Japan BMI groups. However, lower reductions in A1C are seen in non-overweight Inconsistent fi ndings exist regarding the relationship between daily intake T2DM Japanese patients compared to overweight patients. This study dem- of dairy products and risk of metabolic syndrome (MetS). This meta-analysis onstrates that patients may have received a suboptimal dose of glargine, and this was more evident in the subgroup of overweight patients. Obesity aimed to clarify this relationship. A MEDLINE- and EMBASE-based literature

POSTERS search was conducted for cross-sectional or cohort studies that examined Supported by: sanofi -aventis the association between dairy product intake and MetS risk using databases.

Integrated Physiology/ Studies were limited to those that assessed dairy products derived from at 2013-P least 2 of the 3 major dairy products: milk, cheese, and yogurt. The odds or Association of Insulin Resistance and Calcium Mediators in Obese risk ratio (OR/RR) for highest vs. lowest dairy intake group in each study was Individuals Following Gastric Bypass pooled with a random-effects model. For quantitative assessment, the dose of RAELENE E. MASER, M. JAMES LENHARD, MICHAEL B. PETERS, Newark, DE dairy product intake was converted into milk intake in gram units for studies Actions of parathyroid hormone (PTH) in calcium homeostasis are well that allowed qualifi cation of dairy intake. The pooled analysis was repeated for defi ned. There is evidence of an association between insulin resistance and the OR/RR for incremental increases in dairy intake. Eleven eligible (7 cross- PTH as cultures of adipocytes have shown that PTH inhibits insulin signaling. sectional and 4 cohort) studies were analyzed. Qualitatively, high dairy intake We investigated the association of homeostasis model assessment-insulin did not have a signifi cant association with MetS risk (OR (95% confi dence resistance (HOMA-IR) and PTH in obese persons before and 6-months after interval [95%CI]), 0.83 [0.65-1.07])) in cross-sectional studies, but had an in- Roux-en-Y gastric bypass. Subjects (n=32, 91% women, 25% type 2 diabetes) verse association in cohort studies (RR [95%CI], 0.65 [0.44-0.96]) (Figure). The were aged 38±11 (mean±SD) yrs with BMI 51±11 kg/m2 at baseline. For pur-

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A514 OBESITY—HUMANCATEGORY poses of analysis, subjects with a Vitamin D<25 ng/mL, calcium>10.3 mg/dL, nisms for such improvements are not well-defi ned. Ghrelin is an orexigenic or creatinine>1.3 mg/dL were excluded. Thus 19 and 8 subjects were excluded hormone produced predominately by P/D1 cells of the fundic stomach that at baseline and follow-up, respectively. At baseline, HOMA-IR and PTH were regulates food intake and energy balance. Recent studies indicate that acyl not associated (r =-0.14; p=0.65). Post surgery 575% were taking 400+ mg ghrelin (AG) induces insulin resistance and desacyl ghrelin (DAG) may coun- of calcium/day. Table 1 shows a strong correlation of PTH and HOMA-IR at teract the negative effects of AG on insulin sensitivity. AG has an octanoyl follow-up. Table 2 compares those in the lowest tertile for PTH with those in group at Ser-3 and is a ligand for GHS-R1a, while DAG has no known recep- the highest two tertiles at follow-up. PTH levels following surgically-induced tor. We hypothesize that a decrease in AG and an altered ratio of AG to DAG weight loss are associated with HOMA-IR, independent of vitamin D and occurs after RYGB and contributes to early improvements in insulin sensitiv- BMI. PTH appears to function beyond that of calcium homeostasis. These ity.To test this hypothesis, fasting blood samples were collected from 16 results suggest that PTH levels infl uence insulin-stimulated glucose uptake. patients on the day of RYGB surgery and at the postoperative visit to the Table 1. Correlation of PTH with HOMA-IR, BMI, and Calcium Mediators at surgical weight loss clinic (7-10 days later) for quantifi cation of AG, DAG, Follow-up (n=24) insulin, and glucose levels. A serine protease inhibitor, Pefabloc SC, was added to the blood sample for AG measurement to ensure stability of the HOMA-IR BMI Vitamin D Calcium (kg/m2) (ng/mL) (mg/dL) octanoyl group. Plasma insulin and glucose levels were measured to esti- mate insulin sensitivity by HOMA-IR. Within the fi rst 10 days, patients lost PTH (pg/mL) r=0.57 r=0.33 r=-0.05 r=-0.45 5.7 ± 0.3 kg of body weight (p = .001) and HOMA-IR decreased from 3.0 ± p-value p=0.005 p=0.12 p=0.82 p=0.03 0.3 to 2.4 ± 0.3 (p =.034). Plasma levels of AG decreased from 198 ± 36 to 63 ± 11 pg/ml (p < .0001) and DAG from 132 ± 17 to 81 ± 10 pg/ml (p = 0.001) after surgery. Further, there was a decrease in the proportion of AG relative Table 2. Lowest vs. Highest Two Tertiles Combined Based on PTH at Follow-up to total ghrelin after RYGB from 55 ± 4% to 42 ± 4% (p = .001), resulting in a Lowest Tertile Highest Two Tertiles p-value shift from predominately AG before RYGB to predominately DAG after RYGB. (n=8) (n=16) This decrease in the form of ghrelin known to exert the hormone’s appetite HOMA-IR 0.76±0.27 1.14±0.38 =0.02 stimulating and insulin desensitizing effects may contribute to the rapid im- BMI (kg/m2) 34±5 35±8 =0.80 provement in satiety and insulin sensitivity following RYGB. Supported by: DK091748, DRTC-SRTP T35 DK007383 Vitamin D (ng/mL) 37±8 37±8 =0.90 Calcium (mg/dL) 9.7±0.4 9.2±0.6 =0.03 Supported by: American Society for Metabolic & Bariatric Surgery 2016-P Translating the Diabetes Prevention Program (DPP) into Primary Care: A Randomized Trial VERONICA YANK, LAN XIAO, RANDALL S. STAFFORD, LISA GOLDMAN ROSAS, 2014-P SANDRA R. WILSON, JUN MA, Palo Alto, CA Validation of a Fat Accumulation Index Derived from Self-Reported Translating the Diabetes Prevention Program (DPP) into Primary Care: A Behaviors in the Pathobiology of Prediabetes in A Biracial Cohort Randomized Trial. We randomized 241 overweight/obese adults with in- (POP-ABC) Study creased cardiometabolic risk from a primary care clinic to 1) usual care, 2) ANDREW B. BOUCHER, SOTONTE EBENIBO, CHIMAROKE EDEOGA, SAMUEL a self-directed DPP-based intervention on 12 lifestyle topics delivered by DAGOGO-JACK, Memphis, TN DVD after a single 2-hour in-person group session, or 3) a coach-led DPP- Standardized dietary and physical activity questionnaires are the main- based intervention delivered during 12 weekly 2-hour group sessions with stay of behavioral assessment in clinical research. We determined the in- the same topics and guided exercises. Intervention groups had online access teractions of dietary and exercise habits with measures of body size and to a lifestyle coach and public self-management portal for 12 more months. cardiometabolic risk in a biracial cohort of 300 nondiabetic adult offspring Of the participants, 54% were men, mean (SD) age was 52.9 (10.6) yrs, and (159 black, 141 white) of type 2 diabetes (T2DM) parents. The subjects are baseline mean weight and BMI were 93.8 (17.7) kg and 32.0 (5.4) kg/m2. At 15 enrolled in POP-ABC, a longitudinal study of incident dysglycemia among months, the mean (SE) weight change from baseline was -2.4 (0.9) kg in the normoglycemic offspring of T2DM parents. Their mean (+ SD) age was 45 + usual care (P=0.004 vs. baseline), -4.5 (0.9) kg in the self-directed (P=0.015 10.2 y, BMI 30.0 + 6.88 kg/m2, and fasting glucose was 92.1+ 6.79 mg/dl. vs. usual care), and -6.3 (0.9) kg in the coach-led groups (P<0.001 vs. usual At baseline, each subject had a physical examination and 75-g OGTT, com- care, P=0.032 vs. self-directed) (Figure). The percentage of participants who pleted the 20-item Food Habits Questionnaire (FHQ) and the Modifi able lost * 5% of initial weight was 35.6% in the usual care group vs. 60.7% Activity Questionnaire (MAQ), and had fasting lipids and body composition (P=0.004) in the self-directed and 66.6% (P<0.001) in the coach-led groups. analyses. To integrate dietary and exercise behavior, we derived a Fat Ac- The median (IQR) number of online coach contacts was 31 (30, 32) in the cumulation Index (FAI): FAI = [10/MAQ] x [mean FHQ score] for each subject. self-directed and 19 (18, 22) in the coach-led groups.While the more labor- The mean FHQ, MAQ (MET-hrs/week), and FAI scores were then evaluated intensive coach-led intervention resulted in signifi cantly greater changes, a against objective measures of adiposity and lipid profi le.The FHQ and MAQ majority of participants in both scalable DPP-based lifestyle interventions scores individually interacted weakly with BMI (r=0.14, P=0.02) and waist achieved clinically important weight loss. Both interventions demonstrated circumference (r = 0.15, P= 0.03), but not with measured body fat. In contrast, the potential for translation and substantial health impact in primary care. the FAI showed robust interactions with total body fat (P=0.0014), trunk fat (P=0.0011), systolic BP (P=0.02) and triglycerides (P=0.0008), in addition to BMI and waist. The FAI also signifi cantly predicted aggregation of metabolic syndrome markers (P=0.0002). These results were consistent across race/ ethnicity. FAI score was higher in African Americans than Caucasians (10.9 + 1.51 vs. 5.67 + 1.02, P=0.0042), and accurately predicted ethnic differences in BMI and body fat. Thus, the FAI, a derivative of self-reported diet and

exercise behavioral, is a reliable predictor of measured adiposity and cardio- Obesity metabolic risk in African Americans and Caucasians. POSTERS Supported by: NIH, NIDDK Integrated Physiology/ 2015-P Altered acyl to desacyl Ghrelin Ratio Early after Roux-en-Y Gastric Bypass Surgery ANNA GARCIA, JOSEPH ANTOUN, RONALD H. CLEMENTS, BRANDON WIL- LIAMS, PAMELA MARKS-SHULMAN, NAJI N. ABUMRAD, ROBYN A. TAMBOLI, Nashville, TN Patients undergoing Roux-en-Y gastric bypass (RYGB) surgery demon- strate increased insulin sensitivity within 2-7 days of the procedure, before substantial weight loss has occurred. Additionally, ~30% of patients dis- continue their anti-diabetes medication upon hospital discharge. Mecha-

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A515 OBESITY—HUMANCATEGORY

2017-P prebiotic effect of a moderate intake of red wine polyphenols on the growth Baseline But Not Post-Interventional Resting Energy Expenditure of selected bacterial groups in the gut microbiota capable of producing host (REE) is Associated With Larger Body Weight Reduction after Lap- health benefi ts.Ten healthy male volunteers underwent a randomized, cross- aroscopic Adjustable Gastric Banding (LAGB) in Morbidly Obese over, controlled intervention study. After a washout period, all the subjects Subjects received red wine, the equivalent amount of de-alcoholized red wine or gin ALESSANDRO SAIBENE, MONICA MARCHI, VALENTINA DE COSMI, MARINA for 20 days each. Total fecal DNA was submitted to PCR-denaturing gradient SCAVINI, PAOLA VEDANI, ELISABETTA DEVECCHI, GUIDO LATTUADA, FRANC- gel electrophoresis and real-time quantitative PCR to monitor and quantify ESCA RAGOGNA, MICHELE PAGANELLI, EMANUELE BOSI, GIANFRANCO FERLA, changes in fecal microbiota. Several biochemical markers were measured. ANTONIO E. PONTIROLI, GIANLUCA PERSEGHIN, Milano, Italy The dominant bacterial composition did not remain constant over the differ- LAGB is recognized as a valuable therapeutic option in the treatment of ent intake periods. Compared to baseline the daily consumption of red wine morbid obesity. The outcome may show a large subject-to-subject variability polyphenol for 4 weeks signifi cantly increased the number of Enterococcus, and to test the hypothesis whether the response to LAGB may be related Prevotella, Bacteroides, Bifi dobacterium, Bacteroides uniformis, Eggerthella with whole body energy expenditure we studied a cohort of 35 obese sub- lenta, and Clostridium coccoides-Eubacterium rectale group, but signifi cantly jects (30F/5M) with age=38±12 ys, BMI=42±6 kg/m2, body fat=56±11 kg, reduced the clostridia number (P<0.05). In parallel, systolic and diastolic (48±6% of body mass) and measured their REE and body composition us- BP, triglycerides, total cholesterol, HDL cholesterol, and C-reactive protein ing indirect calorimetry and bioelectric impedance analyser (BIA) before and concentrations were signifi cantly reduced (P<0.05). Moreover, changes in 6 months after LAGB. The weight loss was 13±5% (range: 2-30%) and in cholesterol and C-reactive protein concentrations were linked to changes in parallel an improvement was detected for HbA1c (p=0.02), HDL-cholesterol the bifi dobacteria number.This is the fi rst study showing that red wine con- (p=0.02), ALT (p=0.01), but not for serum total cholesterol (p=0.22), triglycer- sumption can signifi cantly potentiate the growth of selected gut microbiota ides (p=0.23), uric acid (p=0.92), creatinine (p=0.07), fi brinogen (p=0.28) and in humans, which suggests the possible prebiotic benefi ts associated with TSH (p=0.23). REE dropped from 2017±446 to 1717±368 Kcal/die (14±11%; the inclusion of red wine polyphenols in the diet. p<0.00001) meanwhile the respiratory quotient did not change (from Supported by: CICYT-AGL 2006-14228-C03-02 (Spanish Ministry of Education 0.82±0.09 to 0.81±0.06; p=0.77). The cohort was segregated into two sub- and Science) groups: those who had a body weight reduction ) (poor response) or > 13% (good response). The pre-surgery anthropometric and metabolic features of 2020-P these two subgroups were not different with the exception of lower serum Weight Loss (WL) by Degree of Comorbidity as Assessed by the fi brinogen (356±78 vs 436±21 mg/dL; p=0.02) and a greater REE (35.4±3.6 Edmonton Obesity Staging System (EOSS) in Subjects Receiving vs. 32.7±3.7 kcal/kg fat-free mass; P=0.02) in those with good response than Controlled-Release Phentermine/Topiramate (PHEN/TPM CR) in those with poor response. In contrast the post-intervention drop in REE ARYA M. SHARMA, RAJ S. PADWAL, WESLEY W. DAY, Edmonton, AB, Canada, was not different (17±13% vs. 11±9%; p=0.26) in good vs. poor response. Mountain View, CA In conclusion, we demonstrated that in morbidly obese subjects undergo- Body mass index is a standard metric for categorizing degree of over- ing LAGB a larger body weight reduction was associated with higher pre- weight/obesity; however, it does not incorporate weight-related comor- interventional REE but no difference in the decline of REE post-intervention bidities that affect morbidity/mortality. EOSS is a novel measure (validated was detected between patients with good or poor response. against NHANES data) that quantifi es obesity-related morbidity based on the Supported by: Angela Musazzi and Mario Stellato Family presence/extent of comorbidities, with higher EOSS scores associated with increased mortality. In the Phase 3, double-blind, 56-week CONQUER study 2018-P in overweight/obese subjects with *2 weight-related comorbidities, PHEN/ Acute and Chronic Effects of Biliopancreatic Diversion With Duo- TPM CR demonstrated signifi cant WL vs placebo (PBO). Subjects were ran- denal Switch on Plasma Visfatin and Apelin Levels domized to receive PBO, PHEN 7.5mg/TPM CR 46mg (7.5/46), or PHEN 15mg/ SARAH-MAUDE CARON-CANTIN, Quebec, QC, Canada TPM CR 92mg (15/92). In this post hoc analysis, WL was assessed according Visfatin is an adipokine that has been linked to obesity, infl ammation and to subjects’ baseline EOSS category. EOSS was scored as 0=no apparent risk to insulin-mimetic effects. Apelin is an adipokine that has positive inotropic factors (n=1), 1=weight-related subclinical risk factors (n=97), 2=established effects on the cardiac system and may be related to infl ammatory molecules. weight-related chronic disease (n=2170), or 3=established end-organ damage Direct regulation of apelin expression by insulin was reported. Responses of (n=180) . At Week 56, PHEN/TPM CR treatment led to signifi cant weight re- plasma visfatin and apelin levels after bariatric surgery are controversial. The duction vs PBO across all baseline EOSS categories (P<.05; Table). Common objective of this study was to evaluate the acute and chronic impacts of a adverse events were dry mouth, constipation, and paraesthesia. Overweight/ biliopancreatic diversion with duodenal switch (BPD-DS) surgery in patients obese subjects with established weight-related comorbidities achieved sig- with severe obesity to better understand the regulation of visfatin and apelin. nifi cantly greater WL when treated with PHEN/TPM CR vs PBO, regardless of In this study, patients who underwent a BDP-DS (BPD-DS group) were com- baseline EOSS score. EOSS may potentially identify patients with the great- pared to a severely obese group (control group).Anthropometric measures and est need for treatment intervention, and PHEN/TPM CR appears to reduce blood tests were performed before, at day 1 and 5 and at 6 and 12 months af- weight irrespective of obesity severity or comorbidities. ter surgery in BDP-DS group. For the control group, the tests were performed at baseline and at 6 and 12 months.Forty-two subjects in BPD-DS group and 14 in the control group were included. As expected, one year after surgery, there was a reduction in body weight (136.0±24.3 vs. 87.4±16.8 kg) and body mass index (49.9±7.3 vs. 31.1±5.6 kg/m2) in the BPD-DS group (both p<0.001). Plasma visfatin levels only decreased at day 1 (19.55±8.10 vs. 16.8±6.55 ng/ ml p=0.019). Plasma apelin levels decreased at day 1 and 5 (608.07±329.43 Supported by: Vivus, Inc. vs. 538.32±257.85 vs. 511.71±246.67 pg/ml;p=0.008 and 0.015 respectively). In the control group, there was no change in any parameter.There is an acute 2021-P

Obesity decrement in plasma visfatin and apelin levels after BPD-DS but in the long- Differential Effects of GBP and SG on GIP and GLP-1

POSTERS term, weight lost had no effect. The acute effect might be due to a decrease AHMED YOUSSEIF, EFTHIMIA KARRA, MOHAMED ELKALAAWY, EMMA DAYKIN, in production of the two adipokines or an increase in their clearance. Both ALICE GAZET, JACQUELINE DOYLE, HELEN KINGETT, MAAN HASAN, CAROLYN

Integrated Physiology/ may rather be modulated by infl ammation than weight loss. DEACON, ANDREW JENKINSON, MAJID HASHEMI, MARCO ADAMO, NICHO- LAS FINER, JENS JUUL HOLST, RACHEL BATTERHAM, London, United Kingdom, 2019-P Copenhagen, Denmark Infl uence of Red Wine Polyphenols and Ethanol on the Gut Micro- Foregut exclusion is a key theory postulated to mediate the diabetes biota Ecology amelioration post-bariatric surgery. Gastric bypass (GBP) and sleeve gast- MARIA ISABEL QUEIPO-ORTUÑO, MARIA BOTO, FERNANDO CARDONA, CRIS- rectomy (SG) entail distinct gastrointestinal manipulations; foregut exclu- TINA ANDRES-LACUEVA, FRANCISCO J. TINAHONES, Malaga, Spain, Barcelona, sion and hindgut overstimulation post-RYGBP; stomach reduction with intact Spain foregut post-SG.In this prospective parallel group study we compared the Few studies have investigated the impact of dietary polyphenols on the effects of GBP and SG on glucose homeostasis, active glucagon-like pep- complex human gut microbiota and these focused mainly on single polyphe- tide-1 (GLP-1) and glucose insulinotropic peptide (GIP) [total GIP (t-GIP); intact nol molecules and selected bacterial populations. To evaluate the possible GIP (i-GIP)] in the fasted state and in response to a liquid test-meal preop-

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A516 OBESITY—HUMANCATEGORY eratively, at 6 and 12 weeks (w) post-surgery; in 10 non diabetic females 2023-P post-GBP (mean age: 46.5±1.4 years, Body Mass Index (BMI): 45.1±1.6kg/m2) Increased Central Adiposity in Hispanic Children Exposed to Gesta- and 10 age and adiposity-matched non diabetic females post-SG (mean age: tional Diabetes In Utero 44.3±2.4 years, BMI: 45.7±1.56 kg/m2).Both operations resulted in compa- KATHLEEN A. PAGE, ANA ROMERO, ISABEL ENRIQUEZ, VICTORIA CHIRIKIAN, rable excess weight-loss at 6w (GBP=27.9±3.4%, SG= 26.2±4.3%) and 12w THOMAS A. BUCHANAN, ANNY XIANG, Los Angeles, CA, Pasadena, CA (GBP=40.7±3.4%, SG=34.4 ± 4.4%). There was no difference in postopera- Obesity and Type 2 diabetes (T2D) rates are rising dramatically, and peo- tive changes in glucose, insulin levels and homeostatic model for insulin re- ple of Hispanic origin are disproportionately affected. Evidence suggests sistance (HOMA-IR) between the two groups. GLP-1 signifi cantly increased that intrauterine exposure to diabetes increases future risk for obesity and after GBP and SG (GLP-1 Area Under the Curve (AUC): GBP p<0.0001; SG T2D. Pima Indian children who were exposed to diabetes in utero have a p<0.01 at 6 and 12-weeks); yet this effect was greater post-GBP (p=0.05 at 10-fold increased risk of developing obesity and T2D. Animal studies show 6w; p=<0.005 at 12w). Post-GBP t-GIP decreased (TGIP AUC p<0.0001at 6 w; that intrauterine exposure to diabetes leads to malprogramming of fetal and 12 w; p<0.001) whilst post-SG there was no change in t-GIP. i-GIP evalu- tissues, including the hypothalamus and pancreatic islets. No studies have ated at 12 weeks, mirrored t-GIP changes in either group. GIP reduction was investigated the effects of intrauterine exposure to gestational diabetes signifi cantly greater post-GBP vs. SG at both visits (p<0.03).The comparable (GDM) on adiposity specifi cally in the Hispanic population. We tested the hy- glycaemia improvements between the two groups argue against a dominant pothesis that intrauterine exposure to GDM results in increased body mass role for foregut exclusion in the short-term glycaemia amelioration observed index (BMI) and increased abdominal fat in Hispanic children. Participants post-bariatric surgery. However, the dissociated incretin response between are the index offspring of probands who participated in our established ge- GBP and SG, with signifi cantly greater GLP-1 increases and GIP decreases netic cohort of Hispanic women with documented GDM and age-, ethnicity-, post-GBP vs. SG might predict superior long-term weight loss and glycaemia BMI-, and parity-matched women with documented normal glucose levels amelioration post-GBP compared to SG. in pregnancy. We measured height, weight, waist and hip circumferences in 46 offspring (35 GDM-exposed: 25 male;10 female; age 8.6±2 yrs and 11 2022-P non-exposed: 6 male;5 female; age 8.3±2 yrs). BMI, BMI z-score, waist and Diabetes Prevention in Subjects With Prediabetes Using Controlled- hip circumferences and waist to hip ratio were compared between GDM- Release Phentermine/Topiramate (PHEN/TPM CR) Over 2 Years exposed and non-exposed children after adjusting for age, gender, maternal W.T. GARVEY, CHARLES H. BOWDEN, Birmingham, AL, Mountain View, CA pre-pregnancy weight and maternal weight gain during pregnancy. GDM- Diet-induced weight loss (WL) in subjects with Prediabetes (fasting glu- exposed offspring had signifi cantly greater BMI (20.8±1.3 vs. 16.1±1.4 kg/ cose *100 to )125 mg/dL or 2-hour oral glucose tolerance test *140 to )199 m2, p=0.03), BMI z-score (0.9±0.4 vs. -0.3±0.4, p=0.04), waist circumfer- mg/dL) reduces progression to type 2 diabetes mellitus (T2DM). The effi cacy ence (71.2± 2.8 vs. 61.7±3.1 cm, p=0.04), and waist:hip ratio (0.92±0.02 vs. and safety of PHEN/TPM CR was assessed over 108 weeks in 675 obese 0.85±0.02, p=0.02) compared to non-exposed offspring. Hip circumference subjects with *2 weight-related comorbidities randomized to placebo (PBO), did not differ signifi cantly between GDM-exposed and non-exposed groups PHEN 7.5 mg/TPM CR 46 mg (7.5/46), or PHEN 15 mg/TPM CR 92 mg (15/92), (78.7±2.4 vs. 72.9±2.6, respectively, p=0.17). These results suggest a role of along with lifestyle intervention. WL, HbA1c, and progression to T2DM at intrauterine exposure to GDM as a risk factor for increased central adiposity Week 108 were assessed in all subjects with Prediabetes at baseline (Week in Hispanic children. 0; n=316) and in subjects >65 years with Prediabetes. In all subjects with Supported by: K23 DK092702 Prediabetes, least-squares mean percent WL was signifi cantly greater with both doses of PHEN/TPM CR vs PBO at Week 108 (P<.0001; ITT-LOCF): -2.2%, 2024-P -11.1%, and -12.7% for PBO (n=103), 7.5/46 (n=83), and 15/92 (n=130), respec- Weight Loss for Patients With Type 2 Diabetes after Sustained Low- tively. PHEN/TPM CR signifi cantly lowered HbA1c levels vs PBO (P<.01): Intensity Lifestyle Intervention and Intensifi ed Medical Treatment 0.08%, -0.01%, and -0.08% for PBO, 7.5/46, and 15/92, respectively, and FINN E. VON EYBEN, Orkanger, Norway markedly reduced annualized incidence rates of T2DM by 49% and 89% in This study of patients with type 2 diabetes aimed to evaluate whether subjects receiving 7.5/46 and 15/92, respectively, compared with PBO (Fig- obesity was associated with metabolic risk factors and the medical treat- ure). Among subjects >65 years, the magnitude and direction of effi cacy out- ment. This was a 4 years follow-up of 186 patients (115 men and 71 women) comes were comparable between the overall population with Prediabetes. treated at a Norwegian hospital. During follow-up for median 11.5 months, Common adverse events in the Prediabetes SEQUEL safety population were dose of statins, metformin, and insulin was increased signifi cantly (P <0.001, constipation, dry mouth, and paraesthesia. In summary, PHEN/TPM CR led to t-test), as was the number of antihypertensive drugs (P <0.001, r2 test). Pa- signifi cant WL sustained over 2 years, improved HbA1c, and greatly reduced tients underwent a sustained low-intensity lifestyle intervention and had a progression to T2DM in obese subjects with Prediabetes. signifi cant weight loss during the follow-up (P = 0.030, t-test). At the end of the study, 154 patients were abdominally obese patients and treated with a higher number of antihypertensive drugs (P = 0.022), and a higher dose of statins (P = 0.007), and of insulin (IU/kg/d, P = 0.006) than 32 abdominally lean patients, and they had also higher triglycerides (P = 0.007), and lower HDL cholesterol (P = 0.010). In multiple linear regression analysis, BMI - but not waist circumference - was signifi cantly associated with dose of insulin (IU/kg/d), and waist circumference - but not BMI - was signifi cantly associ- ated with number of antihypertensive drugs and dose of statins (mg/d), and the levels of triglyceride and HDL cholesterol. Patients with type 2 diabetes may gain from a combination of lifestyle intervention and intensive medical treatment.

2025-P

Acceptability and Feasibility of Remote Patient Monitoring for Dia- Obesity betes Prevention POSTERS MELANIE I. STUCKEY, SHEREE SHAPIRO, KRISTIN J. SABOURIN, ROBERT J. PE-

TRELLA, London, ON, Canada Integrated Physiology/ Obesity and metabolic syndrome (MS) increase the risk of developing type 2 diabetes (T2D) and are more poorly managed in rural than urban areas. The purpose of this study was to examine feasibility and acceptability of remote patient monitoring (RPM) technologies to manage MS in a rural population. Participants with at least 2 MS risk factors (n=62; aged 56.8±10.0y) reported to the clinic at 0 (V0), 12 (V1), 24 (V2) and 52 (V3) weeks. A technology sat- Supported by: Vivus, Inc. isfaction survey was completed at V3. Participants received a Blackberry® (BB), Bluetooth® enabled blood pressure (BP) and blood glucose (BG) moni- tors and a pedometer (PED). BP and BG were measured thrice weekly and PED was logged on the BB daily. Real-time measurements were sent to a

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secure online database monitored by researchers. Thresholds were set and 2027-P out-of-range measures triggered an alarm sent to the physician’s BB. Alarms Adiponectin, Weight Change and the Reduction of Interleukin-6 resulted in 1 T2D and 1 hypertension diagnosis. RPM guided 1 modifi cation Levels in Men and Women With Type 2 Diabetes: Findings from Look of BG and 11 modifi cations of BP medications. Participant compliance to AHEAD RPM was initially high with 92±17% of BP, 90±18% of BG and 84±21% of PED L. MARIA BELALCAZAR, WEI LANG, STEVEN M. HAFFNER, RON C. HOOGEVEEN, readings completed between V0 and V1. Compliance dropped between V1 DAWN C. SCHWENKE, F. XAVIER PI-SUNYER, ANDREA M. KRISKA, RUSSELL and V2 (BP: 87±20%; BG: 87±20%; PED: 77±28%) and V2 and V3 (BP: 78±27%; P. TRACY, CHRISTIE M. BALLANTYNE, THE LOOK AHEAD RESEARCH GROUP, BG: 77±28%; PED: 63±35%). The survey showed that 74% of participants Galveston, TX, Winston-Salem, NC, San Antonio, TX, Houston, TX, Phoenix, AZ, New were immediately comfortable using the BP monitor, 57% the BG monitor, York, NY, Pittsburgh, PA, Colchester, VT 88% the PED and 52% the BB. Participants were comfortable using all tech- Adipose tissue is a major source of circulating interleukin-6 (IL-6), a pro- nology by 2 week with 2 exceptions: 1 was not comfortable with the BG infl ammatory cytokine linked with increased cardiovascular risk. The effects monitor for 1 month and 1 was not comfortable with the BB for > 3 months. of weight loss on IL-6 remain unclear. We investigated the effects of a 1-year Using a likert scale from 1-5 (strongly disagree - strongly agree) it was noted intensive lifestyle intervention for weight loss (ILI) on IL-6 levels in obese that RPM helped participants stay motivated to exercise (3.8±1.2), connect type 2 diabetic men and women and determined if changes in weight and/ to healthcare providers (3.8±1.1), increase personal awareness (4.3±0.9), and or in adiponectin, an anti-infl ammatory adipokine and a marker of adipose feel secure (3.7±1.2) and that RPM was not time consuming (1.9±1.1) and did tissue function, were associated with IL-6 change.We examined a subset of not lose novelty (2.1±1.1). Thus, RPM is a feasible and acceptable interven- 1,382 participants (591 men and 791 women) who had adiponectin and IL-6 tion for T2D prevention in an obese rural population. levels at baseline and 1-year in Look AHEAD, a randomized trial evaluating Supported by: CIHR Grant #: CCT-83029, Heart & Stroke Foundation Grant #: PG- whether ILI reduces cardiovascular events in obese type 2 diabetic persons 07-0364 when compared to usual care (Diabetes Support and Education, DSE). Mean age and BMI of participants were 57.1 years and 36.4 kg/m2. Mean weight 2026-P decreased 10.2 and 7.6 kg; median adiponectin levels increased 23.6 and Mortality Risks Associated With Adiposity Above Percentiles 25 6.2% from baseline with ILI, in men and women, respectively. IL-6 levels and 75 (median [interquartile range]) at baseline were 1.9 (1.0, 2.8) and 3.3 (1.5, 3.4) HENRY S. KAHN, KAI M. BULLARD, LAWRENCE E. BARKER, GIUSEPPINA IM- pg/mL in men and women, respectively. Changes in IL-6 levels were -0.50 PERATORE, Atlanta, GA (-1.10,0.13) and -0.66 (-1.70, 0.10) pg/mL with ILI and -0.20 (-0.80, 0.40) and We explored categorical prediction of all-cause mortality at quartile -0.28 (-0.97, 0.50) with DSE, in men and women (p<0.0001 for ILI vs DSE).ILI boundaries p25 (midrange vs quartile 1) or p75 (quartile 4 vs midrange) for signifi cantly reduced IL-6 levels in men and women. In women, adiponectin body mass index (BMI) and 4 abdominal adiposity indicators (waist circum- change was associated with IL-6 change, independently of weight change. ference [WC], waist-to-height ratio [WHtR], waist-to-hip ratio [WHR] and Future studies on the effects of gender and adipose tissue function on the waist-to-thigh ratio [WTR]). The baseline exam included 11,437 adults aged regulation of chronic infl ammation are needed. 18-64 in the 1988-1994 National Health and Nutrition Examination Survey with mortality follow-up through 2006. We also calculated the “lipid ac- cumulation product” (LAP; [WC enlargement*fasting triglycerides]) for the fasting subsample (n=6,890). We estimated sex-specifi c mortality risks for each indicator with proportional-hazard models adjusted for age, black an- cestry, tobacco exposure, and socioeconomic position.The cohort had 1,081 deaths. Among adiposity indicators treated as continuous variables, we found little variation in linear risk prediction (adjusted hazard ratios [aHRs] per 1 SD increase 1.2-1.4 for men, 1.3-1.5 for women). Categorical aHRs are in the Table. Supported by: NIDDK, NHLBI and other Look AHEAD sponsors Categorical adjusted mortality hazard ratios [95% CI] at the boundaries of adiposity midrange Boundary at p25 Boundary at p75 2028-P (Midrange versus Quartile 1) (Quartile 4 versus Midrange) Effects of Lorcaserin on Body Composition in the BLOSSOM Study of Obese and Overweight Patients Men Women Indicator Men Women BRIAN RAETHER, JACEK KROLIKOWSKI, MATILDE SANCHEZ, WILLIAM R. SHA- 0.78 [0.56-1.09] 1.24 [0.87-1.77] BMI 1.54 [1.18-2.01] 1.54 [1.18-2.00] NAHAN, CHRISTEN M. ANDERSON, San Diego, CA

0.85 [0.63-1.15] 1.50 [1.06-2.13] WC 1.54 [1.18-2.02] 1.64 [1.30-2.07] Lorcaserin (Lor) is an investigational selective 5-HT2C agonist for weight 0.91 [0.63-1.31] 1.39 [0.98-1.99] WHtR 1.70 [1.31-2.19] 1.65 [1.26-2.17] management. The randomized, placebo controlled BLOSSOM study used dual emission x-ray absorptiometry (DXA) in a subset of 189 patients. The 1.03 [0.75-1.42] 1.23 [0.71-2.14] WHR 1.23 [0.94-1.61] 1.80 [1.42-2.27] goal was to determine the contributions of fat and lean body mass loss to 1.38 [0.98-1.93] 1.20 [0.79-1.82] WTR 1.13 [0.86-1.49] 1.72 [1.28-2.31] body weight loss achieved over 1 year.Obese and overweight adults took 1.03 [0.72-1.49] 1.26 [0.75-2.15] LAP 1.11 [0.66-1.85] 1.48 [0.90-2.43] Lor 10 mg once daily (Lor QD), Lor twice daily (Lor BID), or placebo (Pbo); all received behavior modifi cation counseling. DXA scans were performed at At p25 women’s risk with increased WC was elevated for non-blacks (aHR baseline, 6 mo and 1 yr. Body fat content and lean mass were analyzed by 1.6 [1.1-2.5]) but not for blacks (aHR 0.8; interaction p=0.04). At p75 increased sex and by magnitude of weight loss.Lor BID was associated with 9.9% fat LAP conferred elevated risk for tobacco-exposed persons (aHR>1.6), but not mass loss compared with 4.6% with Pbo in the overall population. Weight for non-exposed (aHR<1.0; interaction p<0.05). A WC or WHtR above p75 loss in all groups was predominantly loss of fat mass, however Lor BID in- conferred elevated risk for persons without diabetes (aHR>1.5), but not for creased loss of fat mass relative to Pbo at all levels of body weight loss, diabetic persons (aHR 0.7-1.1).Adiposity’s categorical association with mortal- Obesity reaching 18% among women who lost *5% body weight. Men in the Pbo ity depends on the indicator employed, the midrange boundary of interest, POSTERS group had greater decreases in body fat than did women, while fat mass sex, ancestry, and clinical characteristics including diabetes status. decreased similarly in men and women treated with Lor.In summary, in pa-

Integrated Physiology/ tients receiving diet and exercise counseling, loss of fat mass was greater with Lor than with Pbo.

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the highest postprandial hypertriglyceridemia showed a higher increase Pbo Lor QD Lor BID in lipopolysaccharide (LPS) levels in serum and the chylomicron fraction Change from Baseline to Wk 52 (ITT/LOCF) after the fat overload. Postprandial chylomicron LPS levels correlated posi- n Mean n Mean n Mean tively with the difference between postprandial triglyceridesand baseline (sem) (sem) (sem) triglycerides. The main variables contributing to serum LPS levels after fat Weight (kg) All 1539 -2.9 (0.2) 771 -4.7 (0.2)* 1560 -5.8 (0.16)* overload were baseline and postprandial triglyceride levels but not glucose or insulin resistance. In addition, superoxide dismutase activity decreased Women 1205 -2.6 (0.2) 630 -4.4 (0.3) 1258 -5.8 (.2) signifi cantly after the fat overload.Postprandial LPS increase after a fat Men 334 -3.9 (0.4) 141 -6.0 (0.7) 302 -5.6 (0.4) overload is related to postprandial hypertriglyceridemia but not to degree of Weight (% change) All 1539 -2.8 (0.1) 771 -4.8 (0.2)* 1560 -5.9 (0.2)* insulin resistance in morbidly obese patients. Fat mass (% change) All 69 -4.6 (1.1) 35 -6.1 (2.0) 85 -9.9 (1.4)* Supported by: ISCIII PS09/00997, FIS 08/1655 and CP07/00095, SAS 08/325 and 10/0696 Women 53 -3.4 (1.1) 31 -6.0 (2.2) 77 -9.9 (1.5) Men 16 -8.5 (2.7) 4 -6.6 (4.7) 8 -10.4 (5.2) 2031-P Lean mass (% change) All 69 -0.3 (0.4) 35 -2.0 (0.6) 85 -1.9 (0.5)** Women 53 -0.2 (0.5) 31 -2.1 (0.7) 77 -2.0 (0.5) WITHDRAWN Men 16 -0.6 (0.6) 4 -0.7 (0.5) 8 -0.8 (1.0) Fat mass (% change in Women with weight loss) *3% weight loss 21 -9.7 (1.8) 17 -12.7 (3.2) 55 -14.6 (1.6) *4% weight loss 18 -10.8 (2.0) 15 -14.5 (3.3) 49 -16.4 (1.6) *5% weight loss 15 -11.9 (2.3) 14 -15.3 (3.5) 43 -18.0 (1.7) *p<0.005; **p<0.05 vs. Pbo

2029-P Suppression of Ghrelin after Gastric Bypass Surgery is Associated With Weight Loss in Extremely Obese Patients With T2DM AASHISH SAMAT, STEVE MALIN, HAZEL HUANG, PHILIP SCHAUER, JOHN KIR- WAN, SANGEETA KASHYAP, Cleveland, OH Suppression of Ghrelin after Gastric Bypass Surgery is Associated with Weight Loss in Extremely Obese Patients with T2DMWeight loss following gastric bypass surgery (GBS) is associated with dramatic changes in glucose metabolism. We postulated that restored ghrelin responses following GBS are linked to improved glucose metabolism and improved insulin sensitivity, and weight loss.Cumulative ghrelin concentrations were determined during a mixed meal tolerance test (MMTT) as the average of the fasting, 30 and 60 min value in 9 extremely obese subjects with T2DM (age 42±6 y, 5F/4M, BMI 47.6±1 kg/m2, HbA1c 7.9±1%) at pre-surgery, and 1, and 12 months (m) post-GBS. We also assessed changes in body weight, glucose tolerance, in- sulin sensitivity (Matsuda Index), and insulin secretion based on c-peptide kinetics.Body weight was reduced by ~10%, fasting glucose nearly normal- ized (100±6 vs142±11 mg/dl, P<0.05) and insulin sensitivity improved 2-fold 2032-P at 1 m post-GBS. Mean ghrelin levels during the MMTT were reduced at 1 Investigation of Criteria for Diagnosis of Metabolic Syndrome by m (231±40 vs 326±52 pg/ml, P=0.01), and were similar to baseline at 12 m Using a Home Body Composition Meter (354±42 pg/ml, P=0.48). At pre-surgery, cumulative ghrelin levels strongly YUJI WADA, HIROYUKI MORITA, KAZUO KAJITA, TATSUO ISHIZUKA, Gifu, Japan correlated with insulin secretion (r=0.77, P=0.02) and trended towards an More than 100 cm2 of visceral fat area is defi ned as visceral fat accumula- inverse correlation with fasting glucose (r= -0.62, P=0.07). At 1 m, the incre- tion in Japanese criteria for diagnosis of metabolic syndrome. Waist circum- mental change in ghrelin levels was inversely correlated with the change ference is defi ned as an essential component in the criteria, and the cutoff in weight loss (r= -0.71, P=0.03) and BMI (r= -0.62, P=0.08). There were no value corresponding to 100 cm2 of visceral fat area is 85 cm for men and 90 statistically signifi cant correlations between ghrelin and insulin sensitivity, cm for women. On the other hand, more than 30 kg/m2 of body mass index insulin secretion, or glycemia at 1 or 12 m.These preliminary data suggest (BMI) is defi ned as visceral fat accumulation in criteria for WHO and NCEP- that ghrelin suppression may be linked to anatomical resection of the gas- ATPIII, and their cutoff values of waist circumference are defi ned as more tric fundus leading to altered appetite and/or nutrient sensing after surgery. than 30 kg/m2 of BMI. To reevaluate the cutoff values of waist circumference The absence of statistical correlation between ghrelin, and glucose and/or in those criteria, we analyzed a relationship between waist circumference insulin action does not completely rule out a potential cause-effect role for and visceral fat area or BMI in 515 persons (246 men and 269 women, aged ghrelin in glucose homeostasis. 30 to 93 years) who visited our clinic for physical check-up between 2004 and 2010. The visceral fat area was measured by a home body composition meter 2030-P (HBCM:TANITA Inner Scan BC-522). A signifi cant correlation (r=0.83) between Endotoxin Increase after Fat Overload Is Related to Postprandial the visceral fat area measured by HBCM and CT scan has been reported. The 2 Hypertriglyceridemia in Morbidly Obese Patients waist circumference corresponding to 100 cm of visceral fat area was 77 Obesity

FERNANDO CARDONA, MARIA ISABEL QUEIPO-ORTUÑO, MERCEDES CLEM- cm for men and 103 cm for women, respectively. The lower cutoff value of POSTERS ENTE-POSTIGO, MARIA BOTO, CRISTINA ANDRES-LACUEVA, FRANCISCO J. waist circumference in men than in women was concordant with Japanese

TINAHONES, Malaga, Spain, Barcelona, Spain criteria. The regression lines between waist circumference and visceral fat Integrated Physiology/ The low-grade infl ammation observed in obesity has been associated area for men and women never crossed, and waist circumference for men with a high-fat diet, though this relation is not fully understood. Bacterial was constantly lower than that for women, regardless of any cutoff values endotoxin, which is produced by gut microbiota, may be the linking factor. of visceral fat area. In the correlation between waist circumference and BMI, However, this has not been confi rmed in obese patients. To analyze post- waist circumference was lower in men than in women at 25 kg/m2 of BMI but prandial endotoxemia in morbidly obese patients after a fatoverload and not at 30 kg/m2. The regression lines for men and women crossed at 94 cm for to determine its relationship with postprandial hypertriglyceridemia and waist circumference and 25.3 kg/m2 for BMI. One of the reasons why the cut- insulin resistance.The endotoxin levels were determined in serum and in the off value of waist circumference is lower in men than in women in Japanese chylomicron fraction at baseline and 3h after a fat overload in 40 morbidly criteria and higher in those of WHO and NCEP-ATPIII is as follows: visceral fat obese patients and their levels related with the degree of insulin resistance accumulation is defi ned by visceral fat area in Japanese criteria in contrast to and postprandial hypertriglyceridemia. The morbidly obese patients with by BMI in WHO and NCEP-ATPIII ones.

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A519 OBESITY—HUMANCATEGORY

2033-P 2035-P Plasminogen Activator Inhibitor-1 (PAI-1) and C-Reactive Protein Human Adipocyte Stem Cells Express Transient Receptor Potential (CRP) Comprise Integral Components of the Metabolic Syndrome Melastatin 4 Channel MARK M. SMITS, PIER WOUDSTRA, KRISTINA M. UTZSCHNEIDER, JENNY TRAN DOAN NGOC TRAN, LI-JUN YANG, HAI WANG, MARXA L. FIGUEIREDO, TONG, FERNANDO GERCHMAN, MIRJAM FAULENBACH, DARCY B. CARR, KATH- OLGA ZOLOCHEVSKA, HENRIQUE CHENG, Baton Rouge, LA, Gainesville, FL, RYN ASTON-MOURNEY, ALAN CHAIT, ROBERT H. KNOPP, JAMES B. MEIGS, ED- Galveston, TX WARD J. BOYKO, STEVEN E. KAHN, Seattle, WA, Boston, MA The Transient Receptor Potential Melastatin 4 protein (TRPM4) is a Ca2+ The metabolic syndrome (MetS) is a clustering of factors that predict the activated non-selective cation channel member of the TRP family of ion development of type 2 diabetes and cardiovascular disease, but the underly- channels. TRPM4 allows Na+ entry into cells leading to depolarization. In ing pathophysiology remains unclear. Infl ammation has been proposed as a non-excitable cells (e.g. human adipocyte stem cells (hASCs)), it inhibits Ca2+ possible etiologic mechanism underlying MetS. We used confi rmatory factor entry via store-operated Ca2+ channels.During the differentiation process, analysis (CFA) to test the hypothesis that adipocytokines are associated with Ca2+ oscillations are commonly observed and linked to the expression and the risk factor cluster that characterizes MetS in a cohort of 134 nondiabetic activity of fatty acid synthase (FAS) and peroxisome proliferator activated subjects for whom direct measurements of insulin sensitivity and visceral receptor a (PPARa). However, TRPM4’s role in hASCs has not been investi- fat were available. Insulin sensitivity (SI) was quantifi ed using the minimal gated.Our objective was to identify and characterize TRPM4 in hASCs by model from intravenous glucose tolerance tests and intra-abdominal fat (IAF) RT-PCR and patch-clamp and to test their differentiation capability into area by CT scan. Fasting plasma HDL, triglycerides (TG), monocyte chemo- adipocytes. We also examined the response to agonist-induced increases attractant protein-1 (MCP-1), serum amyloid A (SAA), tumor necrosis factor-a in intracellular Ca2+ signals.First, we identifi ed TRPM4 gene expression in (TNFa), adiponectin, resistin, leptin, interleukin-6 (IL-6), C-reactive protein hASCs, human and rat adipose tissue. Second, exposure of hASCs to adipo- (CRP) and plasminogen activator inhibitor-1 (PAI-1) were measured. The basic genic induction medium resulted in lipid droplet accumulation as indicated model representing the metabolic syndrome included four factors: IAF, SI, by Oil Red O staining. Third, electrophysiological recordings with increasing 2+ TG/HDL and mean arterial pressure and demonstrated excellent goodness- [Ca ]i revealed TRPM4-like currents in a dose-dependent manner. We also of-fi t using CFA. Using multivariate analysis, MCP-1, SAA and TNFa were demonstrated the voltage dependency of TRPM4 where positive potentials not independently associated with any of the MetS factors (p-value >0.05). increased and negative potentials inhibited its activity. Replacement of Adiponectin, resistin, leptin and IL-6 were associated with at least one of extracellular Na+ by N-Methyl-D-Glucamine (NMDG) inhibited TRPM4 cur- the risk factors, but when added to the basic model as separate fi fth factors rents, which confi rms Na+ permeability. Lastly, stimulation of hASCs with 2+ they decreased the goodness-of-fi t. PAI-1 was associated with all MetS fac- histamine, a Gq-protein coupled receptor agonist, increased [Ca ]i in a dose- tors, while CRP was associated with the lipid and obesity factors. Addition dependent manner.We identifi ed TRPM4 in hASCs and demonstrated that of both CRP and PAI-1 improved the goodness-of-fi t compared to the basic the channel is functionally active. These stem cells were also capable of dif- model. In conclusion, our data suggest that infl ammation (indicated by PAI-1 ferentiating into adipocytes and responding to agonist stimulus during Ca2+ and CRP) is an integral feature of the MetS and could be included as fi fth dis- imaging analysis.The signifi cance of this study is that it reports for the fi rst order in the criteria. Interestingly, adiponectin did not emerge as an integral time TRPM4 in hASCs. The ability of the channel to control Ca2+ signaling feature of the syndrome. could have a signifi cant impact on hASC differentiation and adipogenesis.

2034-P 2036-P The Gerodiab Cohort: Triglyceride-to-HDL Cholesterol Ratio is As- EGFR-Mediated Activation of STAT3-Anorexigenic Pathway by Es- sociated With Renal Dysfunction, Waist Circumference, BMI and trogen in the Hypothalamus HbA1c in Older Type 2 Diabetic Patients HYE JIN WANG, EUN SEOK KANG, OBIN KWON, EUN YOUNG CHOI, BYUNG JEAN-PIERRE LE FLOCH, BERNARD BAUDUCEAU, CHRISTIANE VERNY, JEAN WAN LEE, BONG SOO CHA, HYUN CHUL LEE, CHUL HOON KIM, Seoul, Republic DOUCET, SFD-SFGG INTERGROUP, AND THE GERODIAB GROUP, Villecresnes, of Korea, Boston, MA France, Saint-Mandé, France, Paris, France, Rouen, France Estrogen is an anorectic hormone which can regulate synaptic plasticity Triglyceride-to-HDL Cholesterol ratio (TG/HDL) has recently been report- in Proopiomelanocortin (POMC) neurons of hypothalamus, thereby affecting ed as a marker of insulin resistance in diabetic patients. The observational energy balance. Signal transducer and activator of transcription 3 (STAT3) Gerodiab study aims to describe over 5 years the mortality and morbidity in phosphorylation by estrogen in the brain is implicated in producing anorectic 987 type 2 diabetic patients 70 year-old and above, and their associated fac- effect in mice. However, the underlying molecular mechanism of how estro- tors. The present analyzes deal with data at inclusion, and looked for factors gen activates STAT3 in the hypothalamus has remained elusive. GPR30 is a G associated with TG/HDL. Data (mean±SD) was analyzed using linear correla- protein-coupled receptor and recently identifi ed as an estrogen-binding pro- tions and ANOVA.TG/HDL was available in 872 out of the 987 patients of the tein.In this study, we revealed a novel mechanism of STAT3 activation that cohort, 420 men and 452 women, aged 77±5 years, with duration of diabetes involves GPR30. Estrogen did not activate STAT3 in HeLa cells, while HeLa 18±12 years, HbA1c 7.6±1.3% and BMI 29.7±5.2 kg/m². On average, Triglyc- cells transfected with GPR30 showed increased STAT3 Y705 phosphoryla- eride, HDL-Cholesterol and TG/HDL were 137.3±72.5 mg/dl, 51.5±17.0 mg/ tion with estrogen treatment. Estrogen also induced the activation of STAT3 dl and 3.10±2.35, respectively. Higher TG/HDL values were associated with in primary cultured hypothalamic neurons, which was blocked by G-15, a higher BMI values (<30 kg/m²: 2.83±2.36; [30-40[ kg/m²: 3.48±2.35; *40 kg/ selective GPR30 inhibitor. Interestingly, epidermal growth factor (EGF) re- m²: 3.12±1.75; P<0.001), higher waist circumferences (cut-off points 0.80m ceptor inhibitors prevented the activation of STAT3 by estrogen and G-1, a women, 0.94m men: 1.81±0.28 vs 3.24±0.08; P<0.001) and higher waist to hip selective GPR30 activator, in both HeLa cells transfected with GPR30 and ratios (P<0.01) but not with gender (men 3.15±0.12 vs women 3.08±0.11; NS). primary-cultured hypothalamic neurons. These provide a novel mechanism They were associated with lower MDRD levels(<30mL/min: 4.07±0.46; [30- of STAT3 activation by estrogen in the hypothalamus through GPR30-EGF 60[mL/min: 3.85±0.14; *60 mL/min: 2.70±0.10; P<0.001), with higher HbA1c receptor transactivation. levels (<6%: 2.21±0.37; [6-7.5%[: 2.99±0.1; *7.5%: 3.35±0.12; P<0.001), but not with fasting glucose levels. In stepwise multiple regression, TG/HDL 2037-P

Obesity was explained by MDRD, waist circumference, HbA1c, and BMI, succes- Obesity Related Comorbidities 2 years Following Bariatric Surgery POSTERS sively (R²=0.099; P<0.001).In older diabetic patients, renal dysfunction, large in a Group of 215 German Subjects waist circumference, high BMI and poor glycemic control are associated JENS ABERLE, STEFAN WOLTER, FRANZISKA REINING, OLIVER MANN, NINA

Integrated Physiology/ with elevated TG/HDL, a marker of insulin resistance. SAUER, Hamburg, Germany Background and Methods: Bariatric surgery has shown to effectively re- duce body weight and the prevalence of obesity associated comorbidities. However, especially data concerning diabetes is still confl icting. Also the benefi t of surgical therapy varies strongly depending on individual patient characteristics. Therefore we intended to investigate a heterogeneous group of obese patients (n=215) undergoing bariatric surgery in terms of related comorbidities and subgroup benefi ts .Results: After one year mean reduction of body weight was 45±17 kg (BMI 15.4±5.1). High density cho- lesterol (HDL) increased on average by 8.0±9.2 mg/dl (p<0.05), low density cholesterol (LDL)-Levels were reduced by 14.5 ± 27.3 mg/dl (p<0.05). The

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A520 OBESITY—HUMANCATEGORY amount of patients who suffered from sleep apnea reduced from 29.6% to observed to be a fi nding of signifi cant interest relating to pathophysiology. 5.2 % after two years, those suffering from arthropathia from 83% to 20.7 Thus, compared to obese NAA with T2DM, VNA have a higher adipose bur- % and 11.1% respectively after one and two years (p<0,05). The prevalence den and subsequent increased risk of cardiometabolic disease. of hypertension declined from 65.4% to 34.7% after 1 year (p< 0.05). If albu- minuria was initially above 20 mg, available data demonstrated an average decrease of 126.9±214.3 mg without reaching statistical signifi cance.Diabe- tes parameters and showed a u-shaped curve with initial amelioration but deterioration of values from visit 3 on. However 85% of patients taking an- tidiabetics reduced the amount of oral antidiabetics and 100% the injected amount of insulin.Subgroup analysis demonstrated a greater overall benefi t regarding comorbidities for women or patients below 40 years (except for sleep apnea). Reduction of HbA1c and loss of insulin was seen to a greater extent in patients with BMI > 50 kg/m2.Conclusion: Bariatric surgery is an effective treatment of obesity which leads to a signifi cantly lower BMI and has an immense impact on the improvement of comorbidities. Regarding the Supported by: LSUHSC School of Medicine effect on diabetes close follow-up should be provided in order to optimise therapy after bariatric surgery. 2040-P Body Adiposity Index as a New Measure of Obesity and a Predictor 2038-P for Metabolic Derangement in Young Korean Women Impact of Bariatric Surgery on Mid-Thigh/Abdominal Computed To- HYE JIN LEE, JEE-YOUNG OH, YOUNG SUN HONG, YEON-AH SUNG, HYEWON mography Scans and Insulin Resistance CHUNG, Seoul, Republic of Korea AUDREY AUCLAIR, JULIE MARTIN, NADINE BONNEVILLE, FREDERIC-SIMON Obesity is a major public health problem because many metabolic abnor- HOULD, SIMON MARCEAU, PAUL POIRIER, Quebec, QC, Canada malities are associated with obesity. Consequently, assessment of metabol- Bariatric surgery is an effi cient long time treatment for weight loss in se- ic obesity is very important. Body mass index (BMI) is most commonly used vere obesity. The purpose of the study was to assess the impact of weight to diagnose the obesity, but has several limitations to differentiate lean loss and changes in insulin resistance following biliopancreatic diversion from fat mass. A new measure, the body adiposity index (BAI), has recently with duodenal switch (BPD-DS) procedure on adiposity and skeletal muscle. been proposed to provide valid estimates of body fat percentage. We aimed Fasting plasma glucose and insulin, anthropometric measurements as well to compare BMI and BAI for recognizing the fatness and association with as mid-thigh and abdominal computed tomography (CT) were all performed metabolic abnormalities.Among 2,950 young women (age 25±5yrs), weight, at baseline and at 12 months in severely obese subjects who underwent height, and hip circumference were measured, and BMI, BAI (Hip circumfer- BPD-DS (n=20) and in a severely obese control group (n=10). At baseline, ence (cm)/height (m)1.5-18) were calculated. Bioelectric impedance analysis the two groups were similar for age, sex, weight, body mass index (BMI) was used to evaluate the body fat contents. Glucose tolerance status was and mid-thigh/abdominal CT composition. At 12 months, a decreased in assessed by 75 g OGTT, and lipid profi les were measured. glycemia (-21.8±19.4%), insulinemia (-74.6±23.7%) and insulin resistance Table. Correlation coeffi cients of BMI, BAI, adiposity indexes and metabolic (HOMA) (-79.4±22.7%) was observed in the BPD-DS group; all p)0.001. parameters Regarding body composition, there were signifi cant decrements in weight (-37.4±7.1%), BMI (-37.6±7.2%), mid-thigh CT composition; deep adipose tis- Fat Fat Fasting Post-load Fasting Post-load Total tri glycerides HDL mass percent glucose 2h glucose insulin 2h insulin cholesterol cholesterol sue (-54.5±18.1%), subcutaneous adipose tissue (-50.8±17.0%), fat infi ltrated muscle (-32.5±12.5%) and non-fat infi ltrated muscle (-10.3±7.6%) and vis- BMI 0.935 0.791 0.257 0.333 0.485 0.463 0.133 0.374 -0.315 ceral adipose tissue (-61.6±17.4%); all p<0.001. A decrease in insulin resis- (0.000) (0.000) (0.000) (0.000) (0.000) (0.000) (0.000) (0.000) (0.000) tance was associated with reduction of thigh deep adipose tissue (r=0.468, BAI 0.735 0.748 0.196 0.270 0.370 0.378 0.141 0.294 -0.263 p=0.038) and visceral adipose tissue (r=0.681, p=0.001), but not with the (0.000) (0.000) (0.000) (0.000) (0.000) (0.000) (0.000) (0.000) (0.000) reduction of fat infi ltrated muscle (r=0.521, p=0.062). Mobilization of the ( ): P value ROC analysis showed that BMI was a better diagnostic test to thigh ectopic fat namely deep adipose tissue and fat infi ltrated muscle was predict the body fat percent (*35.0%, AUCs 0.908 vs. 0.868, P<0.001). In young related to visceral adipose tissue diminution (deep adipose tissue r=0.732, Korean women, current BMI-based classifi cations for obesity may be superior fat infi ltrated muscle r=0.625; both p<0.05). There was no change in all these for diagnosing body fatness or predicting metabolic dysfunction. parameters in the control group. In conclusion, BPD-DS surgery had major impact on thigh ectopic fat/visceral adiposity which is related to decrement in insulin resistance. 2041-P Lifestyle Interventions Reduce Risk Factors for Metabolic Syn- drome and Improve Early Insulin Responses to Glucose in Subjects 2039-P With an Impaired Early Insulin Response Ectopic Adipose Distribution In Normal Glucose Tolerant, Impaired TAKESHI NISHIKAWA, KIMINORI SADA, TAKESHI YAMASHIRO, DAISUKE KUKI- Glucose Tolerant, and Type 2 Diabetic Vietnamese Americans DOME, SATOSHI ISAMI, HIROYUKI MOTOSHIMA, TAKESHI MATSUMURA, EIICHI LAN CHI T. LUU, ROBERT DUBIN, TIMOTHY ALLERTON, GABRIEL I. UWAIFO, WIL- ARAKI, Kumamoto, Japan LIAM CEFALU, New Orleans, LA The benefi ts of lifestyle interventions on metabolic syndrome (MS) could Asian Americans are at greater risk for type 2 diabetes mellitus (T2DM) be accompanied by improvements in insulin sensitivity. However, it is un- and exhibit higher degrees of total body fat at a given BMI versus non-Asian clear whether lifestyle interventions have benefi ts on MS or insulin secre- Americans (NAA).This study evaluated adiposity and “ectopic” fat deposi- tion in subjects with impaired insulin secretion rather than insulin resistance. tion in a cohort of normal glucose tolerant (NGT), impaired glucose toler- Therefore, we evaluated whether the early insulin response could infl uence ant (IGT) and T2DM Vietnamese Americans (VNA). Thirty VNA subjects (15 the effi cacy of lifestyle interventions. Subjects with at least one component

NGT, 8 IGT, and 7 T2DM) completed dual energy x-ray absorptiometry to of MS and who participated in a group-based lifestyle intervention study Obesity assess percent body fat (%BF). Computer tomography (CT) was performed (Tabaruzaka study) were included in this study. Subjects were divided into POSTERS to measure visceral (VAT) and adiposity subfacial (AS, thigh) and hepatic three groups according to baseline insulinogenic indices, a marker of the fat deposition. Cohort demographics included 46.7% females with a mean early insulin response to glucose, of < 0.4, 0.4-0.8 and * 0.8 for Groups A, B Integrated Physiology/ age of 48.5+13.3 years; mean BMI was 24.7+3.1 kg/m2 and mean A1C was and C, respectively. Two-hour plasma glucose (2h-PG) and the mean number 6.1+0.7%. BMI, %BF, fat-free mass (FFM), VAT, and AS variables did not dif- of MS components decreased signifi cantly, while HDL-cholesterol increased fer signifi cantly between NGT, IGT, and T2DM subgroups. Hepatic fat (as signifi cantly after the intervention compared with baseline levels in all three measured by liver/spleen CT attenuation) was signifi cantly increased in groups. HOMA-IR was signifi cantly decreased and whole-body insulin sen- both IGT and T2DM versus NGT (p<0.0005). Compared to a cohort of obese sitivity index was signifi cantly increased in Group C; neither factor changed T2DM African American and Caucasians, VNA demonstrated signifi cantly signifi cantly in Group A. On the other hand, the insulinogenic index was higher degrees of adiposity at lower BMIs (see table).In this cohort of Viet- signifi cantly increased by 87.5 % in Group A and signifi cantly decreased by namese American, progression to IGT and T2DM appears to be associated 30.4% in Group C. HOMA-` increased signifi cantly in all three groups (79.9, with greater adiposity when corrected for BMI as compared to obese NAA 37.2, or 31.2 % in Group A, B, or C, respectively). Stepwise multiple regres- with T2DM. Interestingly, increased “ectopic” fat deposition in the liver was sion analysis showed that the baseline insulinogenic index was signifi cantly

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A521 ISLET BIOLOGY—APOPTOSISCATEGORY

associated with 2h-PG and urinary 8-hydroxydeoxyguanosine, a marker of results point to a possible context-dependent pro-survival role for SLITs in oxidative stress. These results suggested that group-based lifestyle inter- the adult `-cells. Since diabetes results from a defi ciency in functional `-cell ventions could reduce risk factors for MS in subjects with insulin resistance mass, these studies could potentially contribute to the development of novel and those with impaired insulin secretion. The benefi ts of these interven- therapies to improve `-cell survival. tions in subjects with impaired early insulin responses might be mediated by Supported by: JDRF, SCN, MSFHR, CIHR, NSERC improvements in the early insulin response to glucose. Supported by: Japan Society for the Promotion of Science & 2044-P Gata Transcription Factors Promote Endoplasmic Reticulum Integ- rity and Pancreatic Ƶ-cell Survival ISLET BIOLOGY—APOPTOSIS DANIEL J. SARTORI, CHRISTOPHER J. WILBUR, SIMON LONG, CHANGHONG LI, WILLIAM T. PU, JAKE A. KUSHNER, Philadelphia, PA, Boston, MA, Houston, TX Despite decades of research pancreatic `-cell survival remains poorly Guided Audio Tour: Islet Survival and Demise (Posters 2042-P to 2049-P), understood. Gata transcription factors broadly regulate embryonic develop- see page 17. ment and infl uence survival of several cell types. Gata4 and Gata6, although suggested to be expressed in adult `-cells, have no known role. We sought & 2042-P to explore the role of Gata factors in adult `-cell survival. To study the role Pancreatic Complement Activation in Pancreata of Patients With, of Gata4 in adult `-cells, we derived `-cell inducible and whole-body in- or at, Increased Risk for Type 1 Diabetes ducible Gata4 knockout mice. `-cell inducible Gata6 knockout as well as PATRICK A. ROWE, BYRON P. CROKER, CLIVE WASSERFALL, MARTHA CAMP- Ddit3 (CHOP) null mice were also derived, along with compound mutants. BELL-THOMPSON, DESMOND SCHATZ, MARK ATKINSON, Gainesville, FL Acute Gata4 or Gata6 gene deletion was induced followed by microscopic A role for autoantibody-mediated complement activation in type 1 diabe- and gene expression studies. Gata4 and Gata6 were detected at low lev- tes (T1D) pathogenesis has been diffi cult to demonstrate due to the scarcity els in primary adult `-cells. Loss of Gata4 led to rapid `-cell death and of suitable pancreatic material for analysis, and the reliance on peripheral ultrastructural abnormalities consistent with models of ER stress. Several blood (complement-fi xing islet cell autoantibodies) which may not refl ect stress response signals, especially CHOP, a mediator of ER stress-induced what is actually occurring in the ` cell. Pancreatic tissue collected through apoptosis, were up-regulated in Gata4 and Gata6 knockouts, confi rming that the Network for Pancreatic Organ donors with Diabetes (nPOD) effort was Gata factors infl uence ER homeostasis. Simultaneous deletion of Gata4 and used to quantify, by immunohistochemical techniques, deposition of the CHOP partially restored `-cell survival. In contrast to `-cell knockout mice, complement activation product C4d in patients with T1D (n=11; 1 to 23 yr du- whole-body inducible Gata4 knockouts failed to exhibit analogous induction ration), those without diabetes (n=11), without diabetes but at risk (i.e., posi- of stress response signals in other Gata4-expressing tissues such as heart tive for one or more islet autoantibodies (mIAA, IA-2, GAD65, ZnT8; n=10)), and lung. Indeed, distinct Gata transcriptional targets were differentially and type 2 diabetes (T2D; n=8). Immunostaining was quantifi ed by image expressed in islets compared to other Gata-enriched tissues. We conclude analysis software, which was substantiated by C4d ranking conducted by that Gata transcription factors promote endoplasmic reticulum integrity and a board certifi ed pathologist blinded to the donor groups. Regardless of do- survival of pancreatic `-cells in a tissue-specifi c manner. nor group or staining density, C4d immunoreactivity was primarily restricted Supported by: 1R01AG040110, 1R01DK081469, NIH P30 DK079638 to blood vessels. C4d density (% C4d+ pixels per section) was signifi cantly higher on pancreatic sections from T1D patients compared to subjects with- & 2045-P out diabetes (T1D: 25.2±4.1%, no diabetes:2.1±0.5%; mean±SE, p<0.001). No RAGE Contributes to Pancreatic Ƶ-Cell Failure in Type 2 Diabetes signifi cant differences were found between autoantibody-positive (AAB+) DONG HAN, YASUHIKO YAMAMOTO, KOICHI TSUNEYAMA, SEIICHI MUNESUE, and autoantibody-negative (AAB-) nondiabetic subjects (AAB+:2.8±0.7%, HIROSHI YAMAMOTO, Kanazawa, Japan, Toyama, Japan AAB-:2.1±0.5%). C4d density among patients with T2D tended to be higher Objective: Lipotoxicity and glucotoxicity, which are exerted by free fatty (11.7± 1.8%, NS) than in subjects without diabetes, but signifi cantly lower acids (FFA) and prolonged hyperglycemia, have been implicated in pancre- than patients with T1D (p<0.01). C4d density was not correlated with the atic `-cell failure in diabetes. Receptor for advanced glycation end-products presence of one or more islet autoantibodies. Our results demonstrate, for (RAGE) and toll-like receptors (TLRs) 2 and 4 could mediate those danger the fi rst time, that complement activation within pancreatic tissue from T1D signals as pattern-recognition receptors (PRRs) upon expression on `-cells. patients is increased. Further studies will investigate its mechanistic sig- In this study, we examined whether PRRs, especially RAGE, would contribute nifi cance. to `-cell failure in type 2 diabetes. Methods & Results: Pancreatic islets Supported by: JDRF of Langerhans were isolated from diet-induced obesity (DIO), ob/ob, db/db, RAGE-null (RAGE-/-), and the control (WT) mice, dispersed into single cells, & 2043-P and assayed for insulin, RAGE and TLR2 and 4 by fl ow cytometry (BD FACS Paracrine Signaling Loops in Adult Pancreatic Islets: SLIT-ROBO Aria II). RAGE expression was detected in insulin-positive `-cells from ob/ Signaling Modulates Adult Beta-Cell Survival ob and db/db mice, but not from WT, DIO and RAGE-/- mice, suggesting a YU HSUAN CAROL YANG, JAMES D. JOHNSON, Vancouver, BC, Canada possible relationship between inadequate leptin receptor signaling and Adult pancreatic islets contain multiple cell types that secrete well charac- RAGE expression. In vitro, the expressions of RAGE and TLR2 were induced terized hormones. Although it is becoming increasingly apparent that islets by palmitate in MIN6 cell, a mouse `-cell line, in a dose-dependent manner. release and respond to more secreted factors than previously thought, sys- RAGE-/- db/db mice were found to be protected from reduced `-cell mass and tematic analyses are lacking. We have previously compiled a list of secreted from impaired glucose tolerance when compared with RAGE+/+ db/db mice. factors and receptors that are expressed in adult mouse or human islets and Summary: RAGE was expressed on pancreatic `-cells of diabetic ob/ob and discovered novel paracrine signaling loops within the islets. This led to our db/db mice. FFA elevation with concomitant AGE formation during prolonged focus on the pro-survival role of axon guidance factors during neuronal devel- hyperglycemic exposure may cause `-cell damage through RAGE induction opment, including the Netrin family of ligands and their receptors neogenin in type 2 diabetes. and UNC5A-D. We also examined the SLIT family and their ROBO receptors. In addition axon guidance, SLIT-ROBO signaling has also been implicated in & 2046-P the regulation of cell migration, angiogenesis and cell death during develop- Role of Activating Transcriptional Factor 3 on the Induction of Type ment of lung, kidney, liver and mammary glands. Immunostaining of adult 2 Diabetes Via the Impairment of Glucose Metabolism in Chronic islets revealed SLIT-2 expression exclusively in `-cells, while ROBO1 and Ethanol-Fed Mice ROBO2 receptors were detected in `-cells and _-cells. Exposing `-cells to JI YEON KIM, JEONG SUK KANG, KEUN JAE PARK, JEONG EUN KIM, DO HEE recombinant SLIT-2 under hyperglycemic conditions decreased thapsigargin KIM, JIHYUN SONG, WON HO KIM, Chungbuk, Republic of Korea induced cell death and down-regulated cleaved caspase-3 and CHOP protein We have recently reported that chronic ethanol consumption induced levels. This was correlated with the decrease in Ask1-p38MAPK pathway pancreatic `-cell dysfunction and apoptosis through glucokinase nitration activation and Xbp1 expression. SLITs are known to trigger repulsive axon and its downregulation. Also, we demonstrated that ATF3 downregulates POSTERS guidance signals by regulating intracellular Ca2+ levels. In -cells, recombi- Islet Biology/ ` GCK in ethanol-fed mice, results in pancreatic -cell dysfunction and apop- 2+ ` Insulin Secretion nant SLIT-1 and SLIT-2 treatments could deplete ER Ca levels and increase tosis, but the exact molecular mechanisms regulated by ATF3 is still not fully 2+ cytosolic Ca oscillations under 15 mmol/L glucose conditions, suggesting understood. Here, we investigated the effects of ATF3 and its downstream 2+ that Ca is involved in SLIT induced downstream signaling. Together, our regulators on pancreatic `-cell dysfunction through alteration of glucose

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A522