bluuebunny/crossref_metadata_2025 huggingface dataset:
19-OR: Controlled-Release Mitochondrial Protonophore (CRMP) Reverses Hypertriglyceridemia and Hepatic Steatosis in Dysmetabolic Nonhuman Primates
Authors: GOEDEKE LEIGH , ROMERAL VALLE MONTALVO , BUTRICO GINA MARIE , KAHN MARIO , DUFOUR SYLVIE , ZHANG XIAN-MAN , CLINE GARY , PETERSEN KITT , CHNG KEEFE , SHULMAN GERALD I.
Published: June 2019

NAFLD is estimated to affect up to one third of the general population and is a major predisposing factor in the pathogenesis of hepatic insulin resistance, NASH, and T2D. While weight loss is highly effective at reversing NAFLD, insulin resistance and T2D, it is difficult to sustain and new therapies are required. Our lab has recently developed a controlled-release mitochondrial protonophore (CRMP) that is functionally liver-targeted and promotes oxidation of hepatic triglycerides by promoting a subtle sustained increase in hepatic mitochondrial inefficiency. While we have previously demonstrated that CRMP safely reverses hypertriglyceridemia, fatty liver, and hepatic inflammation/fibrosis in diet-induced rodent models of obesity, there remains a critical need to assess the safety and efficacy in a model highly relevant to humans. Here, we evaluated the impact of CRMP treatment on hepatic mitochondrial oxidation and the reversal of hypertriglyceridemia, NAFLD, and insulin resistance in a spontaneous nonhuman primate model of NAFLD and the metabolic syndrome. Using Positional Isotopomer NMR Tracer Analysis (PINTA), we demonstrate that acute CRMP treatment (single dose of 5 mg/kg) significantly increases rates of hepatic mitochondrial fat oxidation by 40% (P<0.05). Importantly, CRMP treatment (5 mg/kg/day x 6 weeks) reduced hepatic and plasma triglycerides by 20% (both P=0.05) independently of changes in body weight, food intake, body temperature or adverse reactions. Furthermore, administration of CRMP significantly reduced endogenous glucose production by 18% (P<0.05), which could be attributed to a ~20% reduction in hepatic acetyl-CoA content (as assessed by whole-body βOHB turnover) and pyruvate carboxylase flux.

Conclusion: These studies provide important proof-of-concept data to support the development of novel liver-targeted mitochondrial uncouplers for the treatment of NAFLD/NASH and T2D in humans.

Disclosure

L. Goedeke: None. V. Montalvo Romeral: None. G. Butrico: None. M. Kahn: None. S. Dufour: None. X. Zhang: None. G. Cline: None. K. Petersen: Advisory Panel; Spouse/Partner; Aegerion Pharmaceuticals, AstraZeneca, AstraZeneca, Janssen Research & Development, Merck & Co., Inc., Novo Nordisk A/S. Consultant; Spouse/Partner; Gilead Sciences, Inc. Consultant; Self; Merck & Co., Inc. Consultant; Spouse/Partner; Nimbus Discoverey. Consultant; Self; Novo Nordisk A/S. K. Chng: None. G.I. Shulman: Advisory Panel; Self; AstraZeneca, Janssen Research & Development, Merck & Co., Inc. Advisory Panel; Spouse/Partner; Merck & Co., Inc. Board Member; Self; Novo Nordisk A/S. Consultant; Self; Aegerion Pharmaceuticals, IMetabolic BioPharma Corporation, Longitude Capital, Nimbus Discovery, Inc., Staten Biotechnology B.V.

Funding

National Institutes of Health
190-OR: Weight-Loss after Exercise Increases Non-Insulin-Dependent Glucose Clearance in Adults with Metabolic Syndrome
Authors: HARI ADITHYA , AXELROD CHRISTOPHER L. , MEY JACOB T. , KIRWAN JOHN P.
Published: June 2019

Fasting plasma glucose is a composite measure of hepatic glucose production (HGP) and non-insulin-dependent (basal) tissue glucose clearance rate (bGCR). A rise in fasting plasma glucose, within the nondiabetic range, is purported to be regulated by decreased bGCR rather than increased HGP. However, the role of bGCR in improving metabolic syndrome (MetS) is not yet fully understood. We sought to explore the novel impact of weight loss induced by exercise on bGCR in individuals with MetS. Fourteen sedentary adults with MetS (9 females, 68 years, 35.2 kg/m2) participated in a 12-week moderate intensity exercise program (~65% VO2max, 1 hr/day, 5 days/week). MetS was defined using ATP III guidelines. Insulin resistance (IR) was estimated using a hyperinsulinemic (40 mU/m2/min) euglycemic (90 mg/dL) clamp. Rate of glucose disappearance (Rd) was determined using [6,6-2H2]-glucose during fasting and insulin stimulation. GCR was calculated as Rd relative to the prevailing glucose concentration. At baseline, bGCR correlated with IR and adiponectin (r=0.69 and 0.68, P<0.01). Following exercise training and relative to pre-intervention: (i) body weight, VO2max and IR improved (6.6, 17 and 7.7%, respectively; P<0.05); (ii) cardiometabolic risk factors decreased (Triglycerides: 30%; Total Cholesterol: 15%; LDL: 15%; sBP: 11% and dBP: 14.5%; all P<0.05); (iii) fasting glucose and insulin concentration decreased (6% and 32%, P<0.05), and bGCR increased (~1.5 fold, P<0.01). The increase in bGCR correlated positively with changes in body weight, fat mass, suppression of HGP during the clamp, and serum triglycerides (r=0.5, 0.49, 0.61 and 0.62, respectively; P<0.05). Weight loss following aerobic exercise training breaks the vicious cycle of increased insulin resistance and secretion by increasing the contribution of bGCR to fasting glycemia. This evidences the unique and multifactorial benefits of exercise concomitant with weight loss in mediating cardiometabolic risk.

Disclosure

A. Hari: None. C.L. Axelrod: None. J.T. Mey: None. J.P. Kirwan: None.

Funding

National Institutes of Health; National Center for Research Resources
191-LB: Artesunate Treatment Improves Hyperglycemia in a Rodent Model of Gestational Glucose Intolerance without Increasing Beta-Cell Mass
Authors: SZLAPINSKI SANDRA , HILL DAVID J.
Published: June 2019

Gestational diabetes (GDM) has been associated with a failure to adequately increase maternal β-cell mass (BCM) during pregnancy. The anti-malarial drugs, artemisinins, have been shown to improve glucose homeostasis in rodent models of diabetes by increasing BCM. Our objective was to reverse BCM deficiency and impaired glucose tolerance in a mouse model of GDM using the artemisinin artesunate.

In order to produce glucose intolerance in pregnancy, female mice (F0) were fed a low protein (8%, LP) diet during gestation and lactation, and offspring weaned onto control (20%, C) diet. At 6-10 weeks age female offspring (F1) were time-mated and given C diet throughout pregnancy. Artesunate was administered via drinking water from gestational day (GD) 0.5-6.5. We previously found glucose intolerance, mimicking aspects of GDM, in LP-exposed offspring at GD18 when compared to animals fed a C diet. Thus, glucose tolerance tests (GTT) were performed at GD18 or for non-pregnant age-matched animals. Subsequently, pancreata were removed for fluorescence immunohistochemistry to assess histology.

Artesunate-treated LP females had a lower area under the curve during a GTT (389.9±52.3mmol/L*min) compared to non-treated LP females at GD18 (726.6±56.9mmol/L*min, p<0.01). Furthermore, artesunate-treated LP females had improved glucose homeostasis at 5min (9.1 vs. 18.9mmol/L), 15min (11.6 vs. 20.9mmol/L) and 30min (10.2 vs. 19.2mmol/L) during the GTT compared to non-treated LP females at GD18 (p<0.05). There were no significant differences in BCM, alpha-cell mass, or mean islet size between LP and artesunate-treated LP females at GD18.

The results show that artesunate-treated LP pregnant females had significantly improved glucose homeostasis compared to untreated LP mice at GD18. However, the improved glucose tolerance was not associated with increased BCM or changes in endocrine pancreas morphology.

Disclosure

S. Szlapinski: None. D.J. Hill: None.

Funding

St. Joseph's Health Care Foundation; Ontario Graduate Scholarship
1914-P: A Plant-Based Meal Increases Gastrointestinal Hormones and Satiety More Than an Energy- and Macronutrient-Matched Standard Meal in T2D, Obese, and Healthy Participants: A Three-Group Randomized Crossover Study
Authors: VELEBA JIRI , KLEMENTOVA MARTA , BELINOVA LENKA , HALUZIK MARTIN , PAVLOVICOVA RENATA , HILL MARTIN , PELIKANOVA TEREZIE , KAHLEOVA HANA
Published: June 2019

Objective: Gastrointestinal hormones play a key role in glucose metabolism and satiety. We examined the acute effect of meal composition on these hormones in three population groups.

Research Design and Methods: A randomized crossover design was used to test the effects of two energy- and macronutrient-matched meals: a processed-meat (M-meal) and a vegan (V-meal) on gastrointestinal hormones, and satiety in participants with type 2 diabetes (T2D, n=20), obese participants (O, n=20), and healthy controls (H, n=20). Plasma concentrations of glucagon-like peptide -1 (GLP-1), amylin, and peptide YY (PYY) were determined at 0, 30, 60, 120 and 180 minutes. Satiety was assessed using a visual analogue scale. Repeated-measures ANOVA was used for statistical analysis.

Results: An increase in postprandial secretion of GLP-1 was observed after the V-meal in T2D (by 30.5%; 95% CI 21.2 to 40.7%; p<0.001) and H (by 15.8%; 95% CI 8.6 to 23.5%; p=0.01). An increase in postprandial plasma concentrations of amylin was observed in all groups after the V-meal: by 15.7% in T2D (95% CI 11.8 to 19.6%; p<0.001); by 11.5% in O (95% CI 7.8 to 15.3%; p=0.03); and by 13.8% in H (95% CI 8.4 to 19.5%; p<0.001). An increase in postprandial values of PYY after the V-meal was significant only in H (by 18.9%; 95% CI 7.5 to 31.3%; p=0.03). The participants in all groups reported greater satiety after the V-meal: by 9.0% in T2D (95% CI 4.4 to 13.6%; p=0.004); by 18.7% in O (95% CI 12.8 to 24.6%; p<0.001); and by 25.0% in H (95% CI 18.2 to 31.7%; p<0.001).

Conclusions: Our results indicate there is an increase in gut hormones and satiety, following consumption of a single plant-based meal when compared with an energy- and macronutrient-matched standard meat meal.

Disclosure

J. Veleba: None. M. Klementova: None. L. Belinova: None. M. Haluzik: None. R. Pavlovicova: None. M. Hill: None. T. Pelikanova: None. H. Kahleova: None.

Funding

Ministry of Health of the Czech Republic (AZV15-27338A); Institute for Clinical and Experimental Medicine (MZCR00023001)
1929-P: Role of miR379 in Promoting Skeletal Muscle Atrophy in Type 1 Diabetes (T1D)
Authors: TUNDUGURU RAGADEEPTHI , KATO MITSUO , ABDOLLAHI MARYAM , LANTING LINDA L. , NATARAJAN RAMA
Published: June 2019

T1D can lead to skeletal muscle atrophy which decreases quality of life and increases mortality. Non-coding RNAs such as microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) are increasingly implicated in various pathological conditions. Recently, our lab demonstrated a unique megacluster of ∼ 40 miRNAs including miR379 and their host transcript lnc-MGC (a lncRNA) could induce early features of diabetic kidney disease. Based on new data showing that the expression of miR379 is increased in the atrophied skeletal muscle of T1D mice, in the current study, we investigated the in vivo role of miR379 in promoting skeletal muscle atrophy in vivo in T1D. We used miR379 knockout (KO) mice generated by CRISPR-Cas9 technology. C57BL/6 (WT) and miR379 KO male mice were injected with streptozotocin (STZ 50mg/kg) for 5 consecutive days to induce T1D. miR379 target genes and muscle atrophy markers in the skeletal muscle were analyzed using qPCR. Body composition analysis was performed using Echo-MRI. Fasting blood glucose levels one week after diabetes development, were significantly lower in in miR379 KO mice compared to WT-mice. Moreover, miR379 KO mice showed significant protection from T1D-associated loss in body weight, muscle and fat mass relative to WT mice. Toward interrogating the underlying mechanism, the expression of atrophy-associated genes and miR379 target genes were assessed in skeletal muscle samples. In WT mice, T1D increased the expression of muscle atrophy-associated genes (myostatin and Atrogin1), and decreased FIS1 and EDEM3, miR379 target proteins involved in regulation of mitochondrial health and ER stress respectively. Intriguingly, miR379 KO mice depicted significant protection from these T1D-induced changes. These results suggest miR-379 promotes skeletal muscle atrophy in T1D via upregulating genes involved in muscle wasting and downregulating genes involved in mitochondrial health, thus uncovering novel non-coding RNA dependent mechanisms of skeletal muscle dysfunction in T1D.

Disclosure

R. Tunduguru: None. M. Kato: None. M. Abdollahi: None. L.L. Lanting: None. R. Natarajan: None.

Funding

National Institutes of Health
193-OR: Glycemic Control via Glucagon Modulation by a Novel Brown Adipose Peptide Hormone
Authors: PISTON DAVID W. , USTIONE ALESSANDRO , GUNAWARDANA SUBHADRA C. , HUGHES JING
Published: June 2019

Subcutaneous transplants of embryonic brown adipose tissue (BAT) lead to euglycemia without insulin in mouse models of T1D [Diabetes 2012 61:674]. Euglycemia is accompanied by normalized plasma glucagon levels, restored healthy white adipose tissue, reduced inflammatory cytokines, and increased anti-inflammatory adipokines. Elevated IGF-1 was shown to play a role in successful transplants, putatively by direct stimulation of the insulin receptor [Am. J. Physiol. Endocrinol. Metab. 2015 308:E1043]. However, the mechanism underlying this effect remained unknown. We hypothesized that a secreted product from the transplanted BAT communicates with the recipient allowing a new metabolic equilibrium in the absence of insulin. We tested this using buffer conditioned by cultured immortalized BAT cells. We injected this buffer into diabetic NOD mice once daily for 7 consecutive days. In 5 of the 9 animals studied, blood glucose levels dropped back to the normal range (∼150 mg/dl) for at least 2 months with one animal remaining euglycemic for 5 months with no further intervention. As in embryonic BAT transplants, conditioned buffer injections led to a rapid decrease in plasma glucagon. We found that applying this conditioned buffer to isolated mouse and human islets lowers glucagon secretion directly (without significant changes in insulin or somatostatin). To purify and identify the active BAT-secreted compound, we established a multi-step fractionation protocol using size exclusion, ion exchange, and hydrophobicity columns to concentrate the active compound ∼10 million-fold over the initial BAT-conditioned buffer. Protease, denaturation, and spectroscopy experiments revealed a modified peptide of ∼1600 Da. Initial screening of GPCR targets [Nat Struct Mol Biol. 2015 22:362] revealed binding to the μ-opioid receptor, which is expressed preferentially in α-cells within the islet of Langerhans. Thus, these data suggest a novel path towards clinical antagonism of glucagon secretion.

Disclosure

D.W. Piston: Consultant; Self; Pfizer Inc. A. Ustione: None. S.C. Gunawardana: None. J. Hughes: None.

Funding

National Institutes of Health (DK098659); The Leona M. and Harry B. Helmsley Charitable Trust
1935-P: Decoupling between ATP Utilization and Replenishment in Insulin Resistant Skeletal Muscle
Authors: VASQUEZ JUAN A. , SOLIS-HERRERA CAROLINA , TRIPATHY DEVJIT , ABDUL-GHANI MUHAMMAD , DEFRONZO RALPH A. , CLARKE GEOFFREY D.
Published: June 2019

For optimal energy utilization, depletion and recovery of phosphocreatine (PCr) during exercise must be tightly coupled.

Aim: To measure metabolites in vastus lateralis (VL) muscle of normal glucose tolerant (NGT) and type 2 diabetes mellitus (T2DM) subjects by phosphorus-31 magnetic resonance spectroscopy (31P-MRS) during and following exercise. Matsuda index (MI) of insulin sensitivity, fasting plasma glucose (FPG), intramyocellular lipid concentration [IMCL], and mitochondrial density (MitoD) were also measured.

Methods: 12 NGT (age=50±11y; A1c=5.6±0.3%; BMI=28.5±4.1kg/m-2) and 10 T2DM (age=53±10y; A1c=7.5±0.8%; BMI=30.4±4.6kg/m-2) subjects received an OGTT to determine MI. [IMCL] in VL was quantified by 1H-MRS. In 15 subjects (7 T2DM, 8 NGT) MitD was determined by electron microscopy.

Results: The PCr recovery time constant (k) was lower in T2DM (0.01±0.004 vs. 0.02±0.01 s-1; p=0.05). Baseline [PCr] (25.3±3.0 vs. 28.5±3.3 mM; p=0.03) and average [PCr] over the last minute of exercise (14.5±4.7 vs. 20.9±4.7 mM; p<0.05) were lower in T2DM. PCr percent decrease, (%Δ), was negatively correlated with PCr recovery halftime, t (ρ=0.77; p<0.05). The initial rate of PCr depletion and initial rate of PCr recovery were correlated in NGT (r=0.83; p<0.05) but not in T2DM subjects (r=0.30; p=0.39). FPG negatively correlated with baseline [PCr] (ρ=-0.63; p<0.05) and MI negatively correlated with %Δ (ρ=-0.50; p<0.05) in all subjects. [IMCL] was increased in T2DM (6.9±1.2 vs. 9.1±1.4; p<0.05) and correlated negatively with t (ρ=0.47; p=0.03). MitoD in T2DM subjects correlated with k (r=0.76; p<0.05).

Conclusion: These data suggest a disruption in the regulation of energy utilization and production of ATP and PCr, which may be due to reduced mitochondrial density and/or altered substrate dynamics in T2DM skeletal muscle.

Disclosure

J.A. Vasquez: None. C. Solis-Herrera: Advisory Panel; Self; Sanofi. D. Tripathy: None. M. Abdul-Ghani: None. R.A. DeFronzo: Advisory Panel; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Elcelyx Therapeutics, Inc., Intarcia Therapeutics, Inc., Janssen Pharmaceuticals, Inc., Novo Nordisk Inc. Research Support; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Janssen Pharmaceuticals, Inc., Merck & Co., Inc. Speaker's Bureau; Self; AstraZeneca, Novo Nordisk Inc. G.D. Clarke: None.

Funding

National Institutes of Health (DK-24092-34 to R.A.D), (K25-DK-089012 to G.D.C.)
1949-P: Elevated FGF-21 during Circadian Misalignment in Healthy Humans
Authors: BROUSSARD JOSIANE L. , MORTON SARAH J. , MARKAN KATHLEEN , MCHILL ANDREW W. , HIGGINS JANINE , MELANSON EDWARD , WRIGHT KENNETH
Published: June 2019

Circadian misalignment—sleeping during the biological day and eating at a time when the internal circadian clock promotes sleep—is a risk factor for obesity and diabetes, implicating circadian misalignment as a metabolic stressor. Fibroblast growth factor 21 (FGF-21) is a hepatokine secreted in response to stress and found to be elevated in human obesity and diabetes. Activation of peroxisome proliferator-activator receptor alpha by free fatty acids (FFA), can induce the expression of FGF-21 expression. It has been previously reported that circadian misalignment results in elevated fasting FFA. However, the impact of circadian misalignment on FGF-21 has not been explored.

Fourteen healthy adults (6M; aged 26.4±1.2y, BMI 22.7±0.5 kg/m2; mean±SD) participated in a simulated shift-work protocol using a consecutive study design to examine the impact of circadian misalignment on diabetes- and obesity-related risk factors. Participants were provided with a 3d energy balance diet prior to admission and during the 5d inpatient study. Identical meals were consumed each day to minimize the effects of dietary intake on metabolic hormones. Blood was sampled every 1-2 hours for 24 hours during circadian alignment and circadian misalignment and assayed for melatonin and FGF-21.

The melatonin rhythm was used as a marker of the master circadian clock and was unchanged during simulated shift-work, indicating circadian misalignment. Circadian misalignment led to elevated FGF-21 during scheduled wake (+123±41% mean±SEM; p<0.01) and across 24h (+36±16%; p<0.05) relative to circadian alignment.

Circadian misalignment triggers a stress response that includes an increase in circulating FGF-21. The physiological implications of acute elevations in FGF-21 are unknown, and may contribute to metabolic dysregulation associated with circadian misalignment.

Disclosure

J.L. Broussard: Advisory Panel; Self; National Institutes of Health. Research Support; Self; National Institute of Diabetes and Digestive and Kidney Diseases, Sleep Research Society, Society in Science-The Branco Weiss Fellowship. S.J. Morton: None. K. Markan: None. A.W. McHill: None. J. Higgins: None. E. Melanson: None. K. Wright: Advisory Panel; Self; National Institutes of Health, Torvec, Inc. Research Support; Self; National Institutes of Health, Office of Naval Research, Pacific-12 Conference, SomaLogic, Inc. Other Relationship; Self; Circadian Therapeutics, Inc., Kellogg Company, Torvec, Inc.

Funding

National Institutes of Health (UL1RR025780, P30DK048520); Sleep Research Society Foundation; Branco Weiss Fellowship Society in Science (K01DK110138, R21DK092624)
1954-P: Combined Treatment with Exendin-4 and Metformin Attenuates Breast Cancer Growth
Authors: FUJIMURA YUKI , NOMIYAMA TAKASHI , IWAYA CHIKAYO , KAWANAMI TAKAKO , HAMAGUCHI YURIKO , TANAKA TOMOKO , YANASE TOSHIHIKO
Published: June 2019

Cancer is a major cause of death in patients with diabetes. Incretin therapy has received much attention, because of its tissue protective effects beyond glycemic control. We have previously reported anti-prostate cancer effect of Ex-4 and metformin (Diabetes 2014, PLOS ONE 2015). In addition, we have recently reported that Ex-4 attenuates breast cancer growth (Endocrinology 2017). Then, we next examined the effect of combined treatment of Ex-4 with Metformin in breast cancer. In human breast cancer cell lines, MCF-7, MDA-MB-231 and KPL-1 cells, Ex-4 or metformin reduced cell number in a dose-dependent manner. Further, combined treatment of Metformin and Ex-4 additively attenuated the growth curve progression of breast cancer cells. In BrdU assay, Ex-4 or Metformin significantly decreased breast cancer cell proliferation, and further reduction of BrdU incorporation was observed by combined treatment with Ex-4 and Metformin, suggesting that Ex-4 and Metformin additively decreased DNA synthesis in breast cancer cells. Although apoptotic cells were not observed in Ex-4 treated breast cancer cells, apoptotic cells were clearly detected in metformin treated breast cancer cells in TUNEL assay. Further, the experiments using xenograft model mice transplanting MCF-7 cells into athymic mice revealed that Ex-4 and metformin additively attenuated breast cancer growth in vivo also. These data suggest that Ex-4 and metformin attenuates breast cancer cell proliferation and metformin induces apoptosis in breast cancer cells and combined treatment with Ex-4 and metformin could be an optional therapy to inhibit breast cancer progression.

Disclosure

Y. Fujimura: None. T. Nomiyama: None. C. Iwaya: None. T. Kawanami: None. Y. Hamaguchi: None. T. Tanaka: None. T. Yanase: None.
1955-P: GLP-1R Activation Attenuates Prostate Cancer Growth via Inhibiting Cell Cycle Progression
Authors: SHIGEOKA TORU , NOMIYAMA TAKASHI , KAWANAMI TAKAKO , HAMAGUCHI YURIKO , TANAKA TOMOKO , IRIE SHINICHIRO , NABESHIMA KAZUKI , TANAKA MASATOSHI , YANASE TOSHIHIKO
Published: June 2019

Incretin therapy is one of the most popular treatment for type 2 diabetes. We have previously reported anti-prostate cancer effect of GLP-1R agonist Exendin-4(Ex-4) (Diabetes 2014, PLOS ONE 2015), and LEADER trial suggested that GLP-1R agonist could reduce prostate cancer. In our previous report, the attenuation of prostate cancer cell proliferation was depending on GLP-1R expression, and Ex-4 did not attenuate cell proliferation in ALVA-41, a prostate cancer cell without intrinsic GLP-1R expression. Then, we next examined anti-prostate cancer effect in ALVA-41 forced expressed GLP-1R using lentivirus vector (ALVA-41-GLP-1R). We confirmed abundant GLP-1R expression in ALVA-41-GLP-1R using RT-PCR and immunohistochemistry, and intracellular cAMP level was increased by Ex-4 in ALVA-41-GLP-1R. Ex-4 significantly decreased the proliferation of ALVA-41-GLP-1R in a dose dependent manner, but not in ALVA-41-control. BrdU assay revealed that DNA synthesis was attenuated in ALVA-41-GLP-1R, and G1 to S phase entry was inhibited in cell cycle distribution in flow cytometry analysis. Further, p27kip protein was increased and Skp2, ubiquitin ligase for p27kip, expression was decreased in ALVA-41-GLP-1R treated by Ex-4. In vivo experiments using xenograft model mice transplanting ALVA-41-GLP-1R revealed that Ex-4 decreased prostate cancer growth via activation of GLP-1R forced expressed in ALVA41. Further, GLP-1R expression level in human prostate cancer tissues was inversely associated with Gleason grading system of human prostate cancer. These data suggested that forced expressed GLP-1R attenuated prostate cancer cell proliferation via inhibiting cell cycle progression in vivo and in vitro, and GLP-1R activation could be one of optional therapy for prostate cancer.

Disclosure

T. Shigeoka: None. T. Nomiyama: None. T. Kawanami: None. Y. Hamaguchi: None. T. Tanaka: None. S. Irie: None. K. Nabeshima: None. M. Tanaka: None. T. Yanase: None.

Funding

Japan Society for the Promotion of Science
1960-P: Comparative Effects of TNFα and IL-6 Inhibitors on Glucose and Lipid Metabolism in Patients with Rheumatoid Arthritis (RA)
Authors: ISHIZUKA TATSUO , FUJIOKA KEI , MORI ICHIRO , TAKEDA TOMOFUMI , INUI SAORI
Published: June 2019

Association between TNFα and IL-6 and glucose tolerance has been reported. However, it is not clear that TNFα and IL-6 affect on glucose tolerance. Recently, biologic agents such as TNF and IL-6 inhibitors have been appeared in the treatment of RA. We have examined comparative effects of TNF αand IL-6 inhibitors on glucose tolerance and lipid metabolism in patients with rheumatoid arthritis. Registered 589 patients with RA have been screened by HbA1c level (more than 5.6%), treatment with methotrexate (MTX) and treatment with biologic agents with or without MTX. We have excluded the patients with RA who are treated with insulin, and selected 30 cases of RA patients who were able to follow-up for 12 months (M). Patients with RA were divided to 3 groups by the treatments as follows: TNF inhibitors with MTX (TNF+MTX), IL-6 with and without MTX (IL-6±MTX) and MTX alone. Body weight, HbA1c, LDL-C, HDL-C, LDL-C/HDLC(L/H) and disease activity score (DAS) were measured before and after 6 and 12 M. There were no differences of DAS between each group before and after 6 and 12 M. DAS in each group was a significantly decreased compared with before (P < 0.05-0.001). HbA1c levels in TNF+MTX were not significantly different (before 6.08%, 6 M 5.92%, 12M 5.91%), but those in MTX alone and IL-6±MTX were significantly improved (before 6.42% and 6.2%, 6M 5.87% and 5.78%, P< 0.05, 12M 5.82%, P< 0.05 and 5.81%, P< 0.01), respectively. There were no significant differences of body weight between each group. HDL-C and LDL-C in IL-6±MTX were significantly (P< 0.01) increased (before 58.2 mg/dl and 123.2 mg/dl, 6M 71.7 mg/dl and 148.5 mg/dl, 12M 63.4 mg/dl and 144.2 mg/dl), respectively. L/H in TNF+MTX was significantly (P<0.05) decreased.

In conclusion, effect of glucose tolerance in IL-6 inhibitors was better than TNF inhibitors in RA patients with glucose intolerance. However, TNF inhibitor in lipid metabolism was better than IL-6 inhibitors.

Disclosure

T. Ishizuka: None. K. Fujioka: None. I. Mori: None. T. Takeda: None. S. Inui: None.
1964-P: Type 1 Diabetes: Sclerostin Resistance Improves Glucose Metabolism and Protects Bone Mass
Authors: LEANZA GIULIA , ALIPPE YAEL , LEE SEUNG-YON , STROLLO ROCKY , POZZILLI PAOLO , CIVITELLI ROBERTO , NAPOLI NICOLA
Published: June 2019

Type 1 diabetes (T1D) is characterized by metabolic abnormalities and bone fragility. In humans and preclinical models of T1D, serum markers of bone formation are decreased suggesting reduced bone turnover rates. Moreover, circulating levels of the Wnt inhibitor sclerostin (Scl) are higher in T1D patients. To test the role of Scl in T1D bone disease, we introduced the Scl-resistant Lrp5A214V mutation, associated with high bone mass (HBM), in mice carrying the Ins2Akita mutation (Akita), which causes hypoinsulinemia and hyperglycemia at age 4-5 weeks. Although both Akita and Akita/HBM mutants developed diabetes (non-fasting blood glucose ≥300 mg/dl), they showed a very different onset timing. At 6 weeks, only 30% of Akita/HBM mice developed hyperglycemia, compared to 90% of Akita mice (n=10), with the majority of Akita/HBM mice becoming hyperglycemic by 12 weeks. At 6 weeks, Akita mice showed a significant impairment in glucose tolerance in an intraperitoneal glucose tolerance test (ipGTT) relative to mild glucose intolerance demonstrated in Akita/HBM mice (p<0.05 for difference in AUC; n=5-7). No differences in serum insulin, glucagon and C-peptide were found between Akita/HBM and Akita mice at time 0 and 30 minutes during the ipGTT (n=2-6). Importantly however, Akita/HBM showed a significant improvement in insulin sensitivity in an ip insulin tolerance test relative to Akita mice at 7 weeks of age, which persisted until 30 weeks (p<0.05 for difference in AUC; n= 4-7). Strikingly, bone mass was 18% higher in Akita/HBM relative to Akita littermates at 12 weeks (p<0.001; n=3-6), a difference that persisted up until at least 18 weeks of age, despite the prolonged hyperglycemia. Together, these results suggest that Scl resistance may improve peripheral glucose metabolism and slow down the onset of T1D, although it does not prevent it. Even though we have not yet measured bone strength, Wnt stimulation protects bone mass and may delay the onset of metabolic abnormalities seen in T1D.

Disclosure

G. Leanza: None. Y. Alippe: None. S. Lee: None. R. Strollo: None. P. Pozzilli: None. R. Civitelli: Stock/Shareholder; Self; Amgen Inc., Eli Lilly and Company, Merck & Co., Inc. N. Napoli: None.
1966-P: Manipulating Postprandial Bile Acid Concentrations: Effect on GLP-1 Secretion after Roux-en-Y Gastric Bypass
Authors: SVANE MARIA S. , JONSSON ISABELLA , KRISTIANSEN VIGGO BJERREGAARD , KUHRE RUNE E. , WEWER ALBRECHTSEN NICOLAI JACOB , HOLST JENS J. , MADSBAD STEN , BOJSEN-MOLLER KIRSTINE N.
Published: June 2019

Enhanced glucagon-like peptide-1 (GLP-1) secretion is important for improved glycemic control after Roux-en-Y gastric bypass (RYGB) and higher concentrations of circulating bile acids (BA) have been suggested to be involved. The BA chenodeoxycholic acid (CDCA) stimulates GLP-1 secretion after RYGB in the absence of nutrients. In this study we investigated the effects of oral intake of CDCA or the BA sequestrant colesevelam (COL) on postprandial GLP-1 secretion in RYGB operated subjects. We hypothesized that CDCA would increase secretion and that COL would decrease secretion by reducing re-absorption of endogenous BA. Twelve participants (BMI 31±2 (mean±SEM) kg/m2, age 43±3 years, time from RYGB 4.3±0.5 years, weight loss after RYGB 34±5 kg) were studied in a single-blinded, randomised study with four experimental days: 1) A mixed meal consisting of solid/semisolid components (1523 kJ, 53E% carb, 33E% fat, 14E% prot) 2) Mixed meal added 1250 mg CDCA 3) Mixed meal added COL 3750 mg 4) Mixed meal with COL administered both as 3750 mg tablets the evening before the experiment and as 3750 mg added to the meal. Oral intake of CDCA increased GLP-1 significantly compared with mixed meal alone (p=0.03), whereas neither single (p=0.74) nor double (p=0.28) dosage of COL affected GLP-1 secretion (Incremental AUC (iAUC) GLP-1: Meal: 2792±288 pmol x min, CDCA: 4362±618, COL: 2703±330, COLx2: 3328±380). Plasma glucose and C-peptide (iAUC) decreased after CDCA but both were unchanged after COL administration vs. meal alone. Beta-cell function (AUC C-peptide/AUC plasma glucose) was equal between all study days.

In summary, administration of the exogenous BA CDCA potentiated mixed-meal stimulated GLP-1 secretion after RYGB but did not affect beta-cell function. Administration of the BA sequestrant COL did not affect GLP-1 secretion, glucose concentrations or beta-cell function, suggesting a limited role for endogenous BA for regulation of postprandial glucose metabolism after RYGB.

Disclosure

M.S. Svane: None. I. Jonsson: None. V. Kristiansen: None. R.E. Kuhre: None. N.J. Wewer Albrechtsen: Research Support; Self; Mercodia, Novo Nordisk A/S. Speaker's Bureau; Self; Merck Sharp & Dohme Corp. J.J. Holst: Advisory Panel; Self; Novo Nordisk A/S. S. Madsbad: None. K.N. Bojsen-Moller: None.

Funding

European Research Council (695069)
1967-P: Mild Physiologic Hyperglycemia Impairs Insulin-Mediated Suppression of Plasma Glucagon in Healthy Normal Glucose Tolerant Subjects
Authors: MEROVCI AURORA , CHEN XI , DEFRONZO RALPH A. , TRIPATHY DEVJIT
Published: June 2019

Hyperglycemia adversely affects skeletal muscle and hepatic insulin sensitivity (glucotoxicity). However, the effect of prolonged hyperglycemia on glucose and insulin-mediated suppression of glucagon is not known. The aim of the present study was to evaluate effect of a chronic (72 hours) physiologic increase (+45 mg/dl) in plasma glucose concentration on the suppression of plasma glucagon concentration in healthy NGT individuals: 12 without family history of T2DM (FH-) (9M/3F, age = 50± 4 years, BMI = 27 ± 1 kg/m2) and 8 with FH of T2DM (FH+) (4M/4F, age = 48±2, BMI = 26±1 kg/m2). Subjects received an OGTT and 2-step hyperglycemic (+125 and +300 mg/dl) clamp (duration of each step = 80 minutes) before and after 72-hour glucose infusion. Plasma insulin and C-peptide concentrations were obtained every 2-5 minutes during each hyperglycemic clamp step and plasma glucagon concentrations were measured every 10 minutes. The ratio of insulin/glucagon was measured and used as an index of insulin-medicated suppression of plasma glucagon. FPG concentration increased from 97±4 to 140±4 mg/dl during the 72-hour glucose infusion. Following chronic glucose infusion, plasma insulin levels were significantly higher during the basal state and during each hyperglycemic clamp step (by 59% and 78%) during the 0-80 and 80-160 min time periods, respectively. There was no difference in plasma glucagon levels following chronic glucose infusion. However, the plasma insulin/glucagon ratio was significantly higher during the fasting state (by 76%) and during the first (by 128%) and second (by 178%) hyperglycemic clamp steps. There was no difference in the effect of chronic glucose infusion on glucagon secretion between FH+ and FH-subjects. These results demonstrate that sustained physiologic hyperglycemia for 72 hours (i.e., glucotoxicity) impairs insulin-mediated suppression of glucagon, and could contribute to fasting and post-prandial hyperglycemia in T2DM patients.

Disclosure

A. Merovci: None. X. Chen: None. R.A. DeFronzo: Advisory Panel; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Elcelyx Therapeutics, Inc., Intarcia Therapeutics, Inc., Janssen Pharmaceuticals, Inc., Novo Nordisk Inc. Research Support; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Janssen Pharmaceuticals, Inc., Merck & Co., Inc. Speaker's Bureau; Self; AstraZeneca, Novo Nordisk Inc. D. Tripathy: None.

Funding

National Institutes of Health
1969-P: Time-Based Internalized Characterization of Exendin-4-Based Glucagon-Like Peptide-1 Receptor-Targeting Imaging Probe
Authors: MURAKAMI TAKAAKI , FUJIMOTO HIROYUKI , FUJITA NAOTAKA , HAMAMATSU KEITA , FUJIKURA JUNJI , INAGAKI NOBUYA
Published: June 2019

Aims: For the purpose of visualization and quantification of β cells, glucagon-like peptide 1 receptor (GLP-1R)-targeting imaging including single photon emission computed and positron emission tomography (SPECT/PET) has emerged recently. It is well known that internalization of the probes can affect visualization of regions of interest on SPECT/PET. However, the internalization and its effect on β-cell visualization of GLP-1R-targeting probes have not been fully examined. Therefore, we investigated time-based internalized activities of an exendin-4 (Ex4)-based probe, which was developed for clinical GLP-1R-targeting imaging.

Method: [18F]FB(ePEG12)12-Ex4 (18F-Ex4) was synthesized. Rat insulinoma cell line, INS-1 and Human Embryonic Kidney cells 293 expressing human GLP-1R (HEK_GLP-1R) were used as GLP-1R-positive cells. These cells were incubated with 18F-Ex4 (1.48 MBq) for different periods of time (0-120 minutes). Whole cell (WC) and internalized (IC) radioisotope activities were measured by the gamma counter.

Results: WC per incubated probe (IP) activities showed rapid rise within 30 minutes of incubation time and further gradual increase up to 120 minutes in INS-1 and HEK_GLP-1R cells, whereas wild type HEK293 showed no significant WC activity. The WC/IP differences between INS-1 and HEK_GLP-1R cells were significant at the 15-minutes mark (p=0.04) but reduced to insignificant after 30 minutes, although the two cell lines showed significantly different levels of GLP-1R mRNA expressions. IC/IP increased rapidly within 30 minutes and gradually up to 120 minutes. The IC/WC at the 120-minutes mark were significantly higher than those at the 30-minutes mark in both cell lines.

Conclusion: The 18F-Ex4 probe showed rapid uptakes and internalization in GLP-1R-expressing cell lines, which indicates that dual or delayed phases of GLP-1R-targeting imaging may be beneficial. In addition, a time-based hampering effect of GLP-1R-expression levels for β-cell visualization can be suggested.

Disclosure

T. Murakami: None. H. Fujimoto: None. N. Fujita: None. K. Hamamatsu: None. J. Fujikura: None. N. Inagaki: Research Support; Self; Astellas Pharma Inc., Boehringer Ingelheim Pharmaceuticals, Inc., Daiichi Sankyo Company, Limited, Japan Tobacco Inc., Kyowa Hakko Kirin Co., Ltd., Merck Sharp & Dohme Corp., Mitsubishi Tanabe Pharma Corporation, Novartis Pharmaceuticals Corporation, Ono Pharmaceutical Co., Ltd., Sanofi, Sumitomo Dainippon Pharma Co., Ltd., Takeda Pharmaceutical Company Limited.
1971-P: SGLT1-Mediated Glucose Absorption Is Important for Incretin Hormone Secretion after Gastric Bypass Surgery
Authors: MARTINUSSEN CHRISTOFFER , VEEDFALD SIMON , BOJSEN-MOLLER KIRSTINE N. , DIRKSEN CARSTEN , SVANE MARIA S. , KRISTIANSEN VIGGO BJERREGAARD , HOLST JENS J. , MADSBAD STEN
Published: June 2019

Postprandial GLP-1 secretion is greatly enhanced after Roux-en-Y gastric bypass (RYGB) and seems important for the reduced food intake and improved insulin secretion postoperatively. Understanding the mechanisms of gut hormone release after RYGB could aid the discovery of new drugs for the treatment of diabetes and obesity. Glucose is particularly potent for stimulating the release of GLP-1 as well as GIP compared with protein and fat in RYGB operated patients. It is unclear how glucose triggers gut hormone secretion in humans, but animal studies suggest a crucial role of sodium-glucose transporter 1 (SGLT1).

Our aim was to investigate secretion of GLP-1 and GIP in response to an oral glucose load (50 g) on two separate days with or without acute pretreatment of 600 mg of the dual SGLT1/SGLT2 inhibitor canagliflozin (CANA) in 10 RYGB operated patients with no history of diabetes (age 47 ± 5 years, BMI 36 ± 7 kg/m2, HbA1c 35 ± 4 mmol/mol, time from surgery 6 ± 1 years, mean ± SD).

CANA lowered peak glucose (11.8 vs. 8.4 mmol/l, p<0.01) and tended to decrease glucose concentrations during the 4-hour test period (AUC placebo: 1453, CANA: 1343 min mmol/l, p=0.08). CANA inhibited the early rise in GLP-1 secretion with a 28% reduction in peak concentration (p=0.03, 95% CI [-46%, -2.9%]), but a prolonged secretion was seen at later timepoints and overall GLP-1 secretion was comparable (AUC above basal 6067 vs. 7273 min pmol/l, p=0.23). GIP secretion was diminished by CANA (peak by 57%, p>0.01, and AUC above basal by 28%, p=0.01, respectively). In addition, CANA reduced the postprandial rise in lactate concentrations and pulse rate and caused marked glucosuria (0.2 vs. 11 g).

SGLT1/2 inhibition by canagliflozin during glucose ingestion reduced glycemic excursions and diminished early release of GLP-1 and GIP in RYGB operated patients. The results support that glucose absorption via SGLT1 is important for incretin hormone secretion after gastric bypass.

Disclosure

C. Martinussen: None. S. Veedfald: None. K.N. Bojsen-Moller: None. C. Dirksen: None. M.S. Svane: None. V. Kristiansen: None. J.J. Holst: Advisory Panel; Self; Novo Nordisk A/S. S. Madsbad: None.

Funding

European Union Horizon 2020 Research and Innovation Program (695069-Bypass Without Surgery); Danish Diabetes Academy; Novo Nordisk Foundation (NNF15OC0017188); Research Foundation for Health Research of the Capital Region of Denmark; Hvidovre Hospital
1977-P: Methylglyoxal May Contribute to Bone Loss via Differential Effects on Osteoblast and Osteoclasts
Authors: YUE KEVIN , YAN LEE FIONA CHEUK
Published: June 2019

Purpose of the Study: Diabetes mellitus is characterized by chronic hyperglycemia and diabetic complications. Methylglyoxal (MG) is a reactive advanced glycation end-product precursor in which abnormal accumulation is found in the serum of diabetic patients. In clinical studies, they are found to have higher risk of bone fracture. Yet, the underlying mechanism has not been elucidated. Bone remodeling, depending on the interaction between osteoblast (OB) and osteoclast (OC), is to maintain bone structural integrity, such to resorb aged cells while replacing with newly formed cells and to support the metabolic function of bones. Both OB and OC were found to be severely affected by MG. Hence the effect of MG leading to subsequent bone loss was investigated.

Methods: In this study, the effects of MG on bone in both animal and cell models were investigated. SD rats were treated with MG (50mM, 10mL/kg) for 8 weeks. Proximal end of tibia were collected for analysis. The bone mineral density (BMD) and mRNA expression of bone biomarkers were measured using micro-CT and RT-PCR respectively.

Results: Using micro-CT scanning, MG-treated group has lower trabecular BMD (318.1 ± 10.9 mg HA/ccm) compared with control group (383.2 ± 17.7 mg HA/ccm). Using RT-PCR, it was found there were decreased expressions in OB biomarkers including Bone Gamma-Carboxyglutamate Protein (0.718 fold), Collagen I (0.25 fold); while increasing RANKL/OPG (9.46 fold) and Sclerostin (3.51 fold) after MG treatment, indicating the inhibition of bone formation. Besides, increased expressions in OC biomarkers Cathepsin K (1.51 fold), Osteoclast-associated receptor (1.90 fold), Receptor activator of nuclear factor kappa B (1.34 fold) and Tartrate-resistant acid phosphatase (2.57 fold) were observed indicating activation of OC.

Conclusions: MG activates OC and inhibits OB, leading to lower BMD and more porous bone architecture. Therefore, MG may contribute to the bone loss through imbalanced bone remodeling in both OC and OB.

Disclosure

K. Yue: None. F. Lee: None.
1985-P: Inhibition of Adipocyte-Derived Serotonin Protects from High-Fat Diet-Induced Obesity by Increasing Energy Expenditure
Authors: SHONG KO EUN , CHOI WONGUN , KIM HAIL
Published: June 2019

Serotonin (5-HT) is a monoamine that has various functions in both neuronal and non-neuronal systems. 5-HT has been known as an anorexigenic neurotransmitter in central nervous system. However, accumulating evidences suggest that peripheral 5-HT may also affect organismal energy homeostasis. Here we demonstrate the role of adipocyte-derived 5-HT in the regulation of systemic energy metabolism. The expression of tryptophan hydroxylase-1 (Tph1), the rate-limiting enzyme for 5-HT synthesis, and tissue 5-HT level were increased in white adipose tissue (WAT) when mice were fed with high fat diet. Pharmacological inhibition of 5-HT synthesis using Tph inhibitor suppressed the lipogenesis in epididymal WAT, induced beige adipogenesis in inguinal WAT and activated the adaptive thermogenesis in brown adipose tissue (BAT). Adipocyte-specific Tph1 KO (Tph1 FKO) mice exhibited similar phenotypes, suggesting the localized effects of 5-HT on adipose tissues. Tph1 FKO mice showed decreased body fat mass and increased energy expenditure, resulting in improved glucose tolerance. Ucp1 expression increased in the Tph1 FKO iWAT. These data suggest important roles of adipocyte-derived 5-HT in controlling energy homeostasis.

Disclosure

K. Shong: None. W. Choi: None. H. Kim: None.
2000-P: Intracellular Metabolism-Dependent PDGF-B Induction in Inflammatory Macrophages Contributes to Adipose Tissue Expansion with Obesity
Authors: WADA TSUTOMU , ONOGI YASUHIRO , OKEKAWA AKIRA , MATSUZAWA TAKATOSHI , WATANABE ERI , KOMATSU GO , TSUNEKI HIROSHI , SASAOKA TOSHIYASU
Published: June 2019

White adipose tissue (WAT) expands with angiogenesis. We have demonstrated that adipose-tissue-macrophages (ATMs) highly express platelet-derived growth factor (PDGF)-B in obesity, and exposure of PDGF-B stimulates detachment of pericytes (PCs) from vessels. Since vessel-attached PCs inhibit endothelial cell growth, the detachment restarts angiogenesis. In the present study, we aimed to clarify the significance of ATMs on adipose angiogenesis. Intraperitoneal injection of liposome-encapsulated clodronate (Clod) for 6 weeks achieved depletion of macrophages and decreased expression of Pdgfb mRNA in WAT of HFD-fed mice. Clod-treated mice showed smaller WAT with vessels well covered with PCs compared to control mice. We next investigated underlying mechanism of Pdgfb induction. LPS but not IL-4 stimulated Pdgfb expression in peritoneal macrophages. In RAW 264.7 macrophages, Pdgfb expression induced by high glucose plus LPS stimulation was completely blocked by pretreatment with 2-deoxyglucose (2-DG), a hexokinase inhibitor or heptelidic acid, a GAPDH inhibitor, whereas it was not affected by treatment with 6-aminonicotinamide, an inhibitor of glucose-6-phosphate dehydrogenase acting as the rate-limiting enzyme of the pentose phosphate pathway. In addition, high glucose plus LPS induced phosphorylations of S6 kinase, ERK, and JNK but not p38 MAP kinase. These phosphorylations were significantly attenuated by treatment with 2DG. Importantly, enhanced expression of Pdgfb by high glucose plus LPS was effectively suppressed by inhibitor or knockdown of Erk but not by rapamycin. The expression was also attenuated by inhibition of NFκb pathway.

In conclusion, hyperglycemia and chronic inflammation reprogram metabolic pathways toward glycolysis, and subsequent activation of NFκB plays a pivotal role in the production of PDGF-B in ATMs during obesity development.

Disclosure

T. Wada: None. Y. Onogi: None. A. Okekawa: None. T. Matsuzawa: None. E. Watanabe: None. G. Komatsu: None. H. Tsuneki: None. T. Sasaoka: None.

Funding

Japan Society for the Promotion of Science (JP18K08469)