Enhanced Insulin Secretion Research Articles

Glucose-Dependent Insulinotropic Polypeptide (GIP) Reduces Bone Resorption in Patients With Type 2 Diabetes.

ContextIn healthy individuals, glucose-dependent insulinotropic polypeptide (GIP) enhances insulin secretion and reduces bone resorption by up to 25% estimated by absolute placebo-corrected changes in carboxy-terminal type 1 collagen crosslinks (CTX) during GIP and glucose administration. In patients with type 2 diabetes (T2D), GIP’s insulinotropic effect is impaired and effects on bone may be reduced.ObjectiveTo investigate GIP’s effect on bone biomarkers in patients with T2D.DesignRandomized, double-blinded, crossover study investigating 6 interventions.PatientsTwelve male patients with T2D.InterventionsA primed continuous 90-minute GIP infusion (2 pmol/kg/min) or matching placebo (saline) administered at 3 plasma glucose (PG) levels (i.e., paired days with “insulin-induced hypoglycemia” (PG lowered to 3 mmol/L), “fasting hyperglycemia” (mean PG ~8 mmol/L), or “aggravated hyperglycemia” (mean PG ~12 mmol/L).Main Outcome MeasuresBone biomarkers: CTX, procollagen type 1 N-terminal propeptide (P1NP) and PTH.ResultsOn days with insulin-induced hypoglycemia, CTX was suppressed by up to 40 ± 15% during GIP administration compared with 12 ± 11% during placebo infusion (P < 0.0001). On days with fasting hyperglycemia, CTX was suppressed by up to 36 ± 15% during GIP administration, compared with 0 ± 9% during placebo infusion (P < 0.0001). On days with aggravated hyperglycemia, CTX was suppressed by up to 47 ± 23% during GIP administration compared with 10 ± 9% during placebo infusion (P = 0.0005). At all glycemic levels, P1NP and PTH concentrations were similar between paired days after 90 minutes.ConclusionsShort-term GIP infusions reduce bone resorption by more than one-third (estimated by absolute placebo-corrected CTX reductions) in patients with T2DM, suggesting preserved bone effects of GIP in these patients.PrécisShort-term GIP infusions reduce the bone resorption marker CTX by one-third in patients with type 2 diabetes independent of glycemic levels.

Smarter Modeling to Enable a Smarter Insulin.

> I dream things that never were, and I say “Why not?” > > —George Bernard Shaw Pancreatic β-cells secrete insulin to maintain physiological health. While the glucose level is the principle driver of insulin secretion, β-cells also respond to other regulatory inputs including circulating metabolites (most notably amino acids), regulatory hormones, gut peptides, and neurotransmitters. Insulin secretion increases when glucose levels are high, thereby increasing total body glucose utilization and suppressing endogenous glucose production. Of critical importance, insulin secretion is suppressed when glucose levels are low, which protects against life-threatening hypoglycemia. There have been remarkable advances to enhance efficacy and safety of insulin therapy since the discovery of insulin 100 years ago. Insulin pharmacokinetics has been optimized through both formulation and modification of the peptide itself. Sophisticated mechanical devices can provide continuous insulin delivery and continuous glucose monitoring. Pancreatic islets can be transplanted, and bihormonal pumps more closely mimic normal physiology. Despite technological advances, most insulin-dependent patients do not achieve physiological glucose control. The risk of severe hypoglycemia is particularly troublesome because it limits the ability to safely achieve optimal glycemic control, which in turn makes it harder to protect against the long-term risk of microvascular complications (1,2). To underscore this point, half of the Diabetes Control and Complications Trial (DCCT)/Epidemiology of Diabetes Interventions and Complications (EDIC) cohort experienced severe hypoglycemia (∼40 episodes per 100 patient-years) (3). Furthermore, self-reported severe hypoglycemia is associated with 3.4-fold increased risk of death during a 5-year follow-up period (4). The pursuit of a glucose-responsive insulin (GRI) therapy is an ambitious technical objective. In type 2 diabetes, incretin therapy enhances insulin secretion when glucose levels are high but does not cause hypoglycemia because it does not promote insulin secretion when glucose levels are low. Similarly, GRI technology aims to mimic the ability of β-cells …

Insulin Action And Body Composition In Aged C57bl/6 Mice: A New Model For Obesity

The prevalence of obesity in the United States has increased dramatically over the last three decades resulting in a major public health crisis. Feeding mice a high fat diet is a pervasively used model to study mechanisms of human obesity. However, the rapid weight gain that occurs in high fat fed mice and the extremely high fat content of commercially available experimental rodent diets pose serious limitations of this approach. A more appropriate model to study human obesity might be the aged male C57BL/6 mice. PURPOSE: To determine body composition and insulin action in young (YG, 6 months), aged (AG, 18 months), and very old (VO, 28 months) male C57BL/6 mice. METHODS: Body composition was assessed by an LF50 Body Composition Analyzer (Bruker, Inc). Insulin action was determined by conducting insulin tolerance (IT), glucose tolerance (GT), and 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) tolerance (AT) tests. Data was analyzed by using a 1 x 3 analysis of variance and least significant difference post-hoc test. Statistical significance was set at P ≤ 0.05. RESULTS: Body mass (YG: 30.7±1.1 vs AG: 46.3±1.7 vs VO: 39.1±1.6 g) and fat mass (YG: 5.8±1.0 vs AG: 21.6±1.6 vs VO: 10.2±1.8 g) were significantly higher in AG mice compared to VO and YG mice. Lean mass was significantly higher in VO mice compared to AG and YG mice (YG: 20.4±0.4 vs AG: 19.6±0.6 vs VO: 23.4±0.5 g). The area under the curve (AUC) for the GT test was significantly lower in VO mice compared to YG and AG mice (YG: 59030±2817 vs AG: 54835±3423 vs VO: 33378±2286). The AUC for the IT test curve was similar in YG, AG and VO mice (YG: 11320±214 vs AG: 11804±343 vs VO: 11138±968). Although the AUC for the AT test was similar, the decline in glucose following AICAR injections was significantly less in VO mice compared to AG and YG mice, indicating an impairment in AMPK activity. These data suggest that adiposity increases substantially in 18 month old male C57BL/6 mice, a process that appears to be reversed by 28 months. Further, aging does not appear to cause a deterioration in insulin sensitivity when assessed by an IT test. The lower glucose values observed during the GT test in VO mice is likely due to enhanced insulin secretion. Overall our findings indicate that male C57BL/6 mice may be a valuable model to examine mechanisms of obesity when studied at approximately 18 months of age.

Effects of Y1 receptor agonist on the pancreatic islet of diet-induced obese and diabetic mice

AimsAgonists of the NPY receptor might be potential in protecting pancreatic islets from injury. We aimed to characterize the role of [Leu31, Pro34]-PYY, an NPYR1 agonist, in pancreatic islets of a diet-induced obesity and insulin resistance model. MethodsWe studied long-term high-fat diet intake as a model and selective agonist of the Y1 receptor to explore the pancreatic islet architecture and stereology, and insulin secretion in isolated islets and a whole animal model. Gene and protein expressions were assessed in isolated islets investigating the signaling cascades involved in inflammation, insulin signaling, and secretion. Also, the insulin release potential was studied in vitro. ResultsOur data reveal that an infusion of NPYR1 for 14 days did not change the body mass of mice and eating behavior. NPYR1 did not modify the islet and beta-cell mass but positively impacted the inflammatory process by lowering the expressions of Tnf alpha and If gamma. Besides, NPYR1 restored the insulin signaling and the exocytose pattern by activating the PDX1/STAT3 pathway and improving the leptin signaling cascade. ConclusionsThe findings are compellingly indicating the potential effect of the NPYR1 as a target for improving the insulin resistance condition. As such, the infusion of the NPYR1 agonist would help to enhance insulin secretion by the beta-cell from the PDX1/STAT3 pathway and the improvement of the inflammatory process.

Protein Kinase CK2 Controls CaV2.1-Dependent Calcium Currents and Insulin Release in Pancreatic β-Cells.

The regulation of insulin biosynthesis and secretion in pancreatic β-cells is essential for glucose homeostasis in humans. Previous findings point to the highly conserved, ubiquitously expressed serine/threonine kinase CK2 as having a negative regulatory impact on this regulation. In the cell culture model of rat pancreatic β-cells INS-1, insulin secretion is enhanced after CK2 inhibition. This enhancement is preceded by a rise in the cytosolic Ca2+ concentration. Here, we identified the serine residues S2362 and S2364 of the voltage-dependent calcium channel CaV2.1 as targets of CK2 phosphorylation. Furthermore, co-immunoprecipitation experiments revealed that CaV2.1 binds to CK2 in vitro and in vivo. CaV2.1 knockdown experiments showed that the increase in the intracellular Ca2+ concentration, followed by an enhanced insulin secretion upon CK2 inhibition, is due to a Ca2+ influx through CaV2.1 channels. In summary, our results point to a modulating role of CK2 in the CaV2.1-mediated exocytosis of insulin.

Reducing Glucokinase Activity to Enhance Insulin Secretion: A Counterintuitive Theory to Preserve Cellular Function and Glucose Homeostasis.

Pancreatic beta-cells are the only cells in the body that can synthesize and secrete insulin. Through the process of glucose-stimulated insulin secretion, beta-cells release insulin into circulation, stimulating GLUT4-dependent glucose uptake into peripheral tissue. Insulin is normally secreted in pulses that promote signaling at the liver. Long before type 2 diabetes is diagnosed, beta-cells become oversensitive to glucose, causing impaired pulsatility and overstimulation in fasting levels of glucose. The resulting hypersecretion of insulin can cause poor insulin signaling and clearance at the liver, leading to hyperinsulinemia and insulin resistance. Continued overactivity can eventually lead to beta-cell exhaustion and failure at which point type 2 diabetes begins. To prevent or reverse the negative effects of overstimulation, beta-cell activity can be reduced. Clinical studies have revealed the potential of beta-cell rest to reverse new cases of diabetes, but treatments lack durable benefits. In this perspective, we propose an intervention that reduces overactive glucokinase activity in the beta-cell. Glucokinase is known as the glucose sensor of the beta-cell due to its high control over insulin secretion. Therefore, glycolytic overactivity may be responsible for hyperinsulinemia early in the disease and can be reduced to restore normal stimulus-secretion coupling. We have previously reported that reducing glucokinase activity in prediabetic mouse islets can restore pulsatility and enhance insulin secretion. Building on this counterintuitive finding, we review the importance of pulsatile insulin secretion and highlight how normalizing glucose sensing in the beta cell during prediabetic hyperinsulinemia may restore pulsatility and improve glucose homeostasis.

352-OR: Combination Therapy with Dapagliflozin plus Exenatide on Endogenous Glucose Production: A Mechanism of Action Study

Aim: To examine the mechanisms with which GLP-1 RA plus SGLT2i improve glycaemia versus each agent as monotherapy. Study Design: 21 T2D patients (Age= 51±2; BMI=31.2±0.7; A1C=7.9±0.2%, FPG= 172±8; mean PG OGTT [0-5h] =249±8) received 5-hour double tracer (3-3H-glucose IV/1-14C-glucose orally) OGTT and then were randomized to dapagliflozin (DAPA) 10 mg/d, exenatide (EXEN) 2 mg/wk, dapagliflozin plus exenatide (DAPA/EXEN) for 4 months. Results: Combination DAPA/EXEN therapy produced a completely additive effect to reduce HbA1c and near additive effect to decrease FPG and mean PG during OGTT. The additive effect of DAPA/EXEN resulted from: (1) decreased basal EGP; (2) increased urinary glucose excretion (UGE); (3) decreased rate of oral glucose appearance (RaO); (4) enhanced insulin secretion; (5) increased tissue glucose clearance. Of note, exenatide could not overcome the impaired suppression of EGP by dapagliflozin. Disclosure M. Alatrach: None. C. Agyin: None. J.M. Adams: None. O. Lavrynenko: None. N. Laichuthai: None. E. Cersosimo: None. C.L. Triplitt: Speaker’s Bureau; Self; AstraZeneca, Janssen Pharmaceuticals, Inc., Lilly Diabetes, Xeris Pharmaceuticals, Inc. A. Gastaldelli: Consultant; Self; Boehringer Ingelheim Pharmaceuticals, Inc., Eli Lilly and Company, Gilead Sciences, Inc., Inventiva Pharma, Novo Nordisk Inc. M. Abdul-Ghani: None. R.A. DeFronzo: None.

1811-P: Induction of Hepatic Steatosis by a Methionine Choline Deficient Diet Promotes FGF21-Induced Insulin Secretion Independent of Diabetes

While it is known that patients with nonalcoholic fatty liver disease exhibit enhanced insulin secretion, the factor(s) that contribute to this effect are not fully understood especially in nondiabetic subjects. Our aim was to examine crosstalk between a steatotic liver induced by a diet that is deficient in methionine choline (MCD) and pancreatic islets in the absence of diabetes/insulin resistance. Nine-week old male C57BL/6J mice were fed normal chow (CTL) or the MCD diet for 3 weeks followed by evaluation of glucose homeostasis, liver and pancreas pathology and hepatic gene expression by RNA-sequencing. Three weeks of MCD diet caused simple hepatic steatosis with hepatic triglyceride content 2-fold higher in MCD compared to CTL (n=5; p&amp;lt;0.05). Hepatic FGF-21 mRNA, protein and circulating FGF-21 levels were all significantly increased in MCD (129-fold, 8-fold, 5-fold respectively; n=5; p&amp;lt;0.05). Interestingly, while β-cell mass was not significantly altered between groups, insulin secretion as evaluated by ex-vivo GSIS in freshly isolated islets was significantly increased in MCD (5-fold; p&amp;lt;0.05, n=10). Examination of liver samples from patients with or without steatosis mimicked hepatic MCD data in the mouse model. Thus a significant increase in both mRNA and protein FGF-21 were evident in the steatotic group (3-fold, 5-fold respectively; n=6, p&amp;lt;0.05). Furthermore, 3 days treatment of nondiabetic human islets with recombinant FGF-21 protein led to a significant increase in GSIS compared to control (1.5-fold; p&amp;lt;0.05, n=4). FGF-21 treatment of EndoC-βH1 cells revealed significant activation of AKT and ERK1/2 signaling that likely promoted the enhanced insulin secretion. Taken together, liver steatosis significantly increases circulating FGF-21 to promote enhanced insulin secretion via improved signaling in nondiabetic beta cells. Disclosure T. Kimura: None. D.F. De Jesus: None. K. Fukuda: None. S. Kahraman: None. K. Shibue: None. K. Orime: None. R. Kulkarni: None. Funding National Institutes of Health (R01DK067536, R01DK103215)

1797-P: Effect of LCZ696 and Its Components Sacubitril and Valsartan on Insulin Secretion and Glucose Homeostasis in Mice

In type 2 diabetes, HbA1c decreases upon treatment of heart failure with the angiotensin receptor-neprilysin (NEP) inhibitor LCZ696 (LCZ). We sought to determine whether the components of LCZ, the NEP inhibitor sacubitril (SAC) and angiotensin receptor blocker valsartan (VAL), enhance insulin secretion and if so, their effects are additive when combined in LCZ. C57BL/6 mice fed high fat diet (HFD) received injections of vehicle (VEH) or low-dose streptozotocin (3 × 30 mg/kg; STZ), the latter to induce diabetes. Mice were continued for 8 weeks on HFD alone (CTL) or supplemented with LCZ (114 mg/kg/d), SAC (48 mg/kg/d) or VAL (54 mg/kg/d). All drugs significantly reduced body weight gain in both VEH and STZ groups, with LCZ being more effective than SAC (Table). SAC and VAL, but not LCZ, lowered fed and fasting plasma glucose and increased insulin release during an IV glucose tolerance test (IVGTT) in STZ mice only (Table). In VEH and STZ mice, insulin sensitivity was similar among the 4 treatment groups; however, SAC enhanced glucose disposal during IVGTT (Table). None of the treatments altered pancreatic ß- and α-cell mass or lipolysis (not shown). In sum, SAC and VAL exert beneficial anti-obesity, glycemic and insulinotropic effects when given alone in obese and/or diabetic mice. Additional studies are needed to understand why, when SAC and VAL are combined in LCZ, only anti-obesity effects are observed. Disclosure N. Esser: None. C.R. Schmidt: None. B. Barrow: None. L. Cronic: None. D.J. Hackney: None. S.M. Mongovin: None. R.L. Hull: None. S. Zraika: Research Support; Self; Novartis Pharmaceuticals Corporation. Funding Novartis Pharmaceuticals Corporation (LCZ696BUSNC09T)

324-OR: The Incretin Action of GIP Is Mediated through Both Alpha and Beta Cells

Glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide 1 (GLP-1) are the principal incretins connecting nutrient intake to postprandial insulin secretion. Both GIP and GLP-1 directly stimulate insulin secretion in β-cells through cognate receptors (GIPR; GLP-1R) in a glucose-dependent manner. The GIPR is also expressed on α-cells and GIP stimulates glucagon secretion. α- to β-cell communication, manifest as activation of β-cell GLP-1R by proglucagon peptides (PGDP), dictates the magnitude of the insulin response. We hypothesized that GIPR-stimulation of PGDP enhances insulin secretion through α- to β-cell communication, and that GIPR activity in α-cells is nutrient dependent, similar to β-cells. Mouse islets perifused with GIP or alanine alone doubled their secretion of glucagon, but the combination produced ∼10x glucagon secretion, a synergistic effect that was present at both 2.8 and 10 mM glucose. At 10 mM glucose, GIP plus alanine also had a synergistic effect on insulin secretion. This synergy was abolished by the GLP-1R antagonist exendin-(9-39) indicating that amino acid enhancement of GIP-stimulated insulin secretion is dependent on α- to β-cell communication. In vivo, mice with α-cell deletion of GIPR had normal intraperitoneal and oral glucose tolerance, but impaired glycemic control and reduced insulin secretion in response to a mixed nutrient stimulus, a condition that enhances α-cell activity. Together, these results demonstrate that GIPR activity in α-cells is dependent upon amino acid activation and contributes significantly to insulin secretion through α- to β-cell communication. Disclosure K.M. El: None. M. Capozzi: None. S.M. Gray: None. J.L. Brown: None. F.S. Willard: None. P. Emmerson: None. K. Sloop: None. D. D’Alessio: Advisory Panel; Self; Eli Lilly and Company. Consultant; Self; Intarcia Therapeutics. Research Support; Self; Ansh Labs, Eli Lilly and Company, Merck Sharp &amp; Dohme Corp. Other Relationship; Self; Novo Nordisk A/S. J. Campbell: Research Support; Self; Eli Lilly and Company, Novo Nordisk Inc. Speaker’s Bureau; Self; Merck Sharp &amp; Dohme Corp. Funding American Diabetes Association (1-17-JDF-074 to J.C.)

325-OR: ß-Arrestin 1 Regulates Glucagon-Like Peptide-1 Receptor Mediated Insulin Secretion in a Ligand-Dependent Manner

Incretin-based drugs are widely thought to act via a Gαs/adenylate cyclase (AC)/cAMP pathway in β-cells. This model has been challenged by data demonstrating various glucagon-like peptide 1 receptor (GLP-1R) ligands differentially activate multiple signaling pathways including Gαs, Gq, ERK, and β-arrestins. This ligand-mediated signaling bias implicates an array of intracellular mechanisms that regulate GLP-1R action and raises the possibility GLP-1R agonists can be optimized to enhance insulin secretion. We tested the hypothesis that β-arrestin 1 (βarr1), a critical GPCR effector, is necessary for the full insulinotropic action of GLP-1R agonists. The β-cell βarr1 knockout mice (βarr1βcell-/-) used here have reduced Arrb1 expression in the β-cell, but not α- or Δ-cells; Arrb2 expression was unaffected. Islets isolated from βarr1βcell-/- mice have increased glucose stimulated insulin secretion (GSIS) compared to controls. The enhanced GSIS in the knockout islets was ablated by the GLP-1R antagonist exendin-9. In addition, βarr1βcell-/-islets treated with GLP-1 or exendin 4 (Ex4) have increased insulin release compared to controls. Conversely, βarr1 deletion did not impact the insulinotropic actions of glucagon, which signals through both the GLP-1R and the glucagon receptor (GCGR). Moreover, control and βarr1βcell-/- islets produced similar insulin secretion in response to a GCGR specific agonist that does not signaling through GLP-1R and to glucose-dependent insulinotropic polypeptide. Together, these results indicate that βarr1 in β-cells selectively dampens the insulinotropic actions of the GLP-1R, but not the GCGR or GIP receptor. Thus, there is a specific interaction between the GLP-1R and βarr1 that points to divergent regulation between the insulinotropic class B G-protein coupled receptors in β-cells. This data implies that GLP-1R agonists that bias away from βarr1 may be more effective at stimulating insulin secretion. Disclosure J.D. Douros: None. K. Sloop: None. J. Campbell: Research Support; Self; Eli Lilly and Company, Novo Nordisk Inc. Speaker’s Bureau; Self; Merck Sharp &amp; Dohme Corp. D. D’Alessio: Advisory Panel; Self; Eli Lilly and Company. Consultant; Self; Intarcia Therapeutics. Research Support; Self; Ansh Labs, Eli Lilly and Company, Merck Sharp &amp; Dohme Corp. Other Relationship; Self; Novo Nordisk A/S. Funding 1F32DK115031-01

321-OR: Deletion of 14-3-3ζ in Pancreatic ß-Cells Increase Mitochondrial Activity and Insulin Secretion

Pancreatic β-cells continuously sense blood glucose levels and secrete insulin to maintain normoglycemia. In β-cells, ATP generated by glycolysis promotes the closure of ATP-sensitive K+ channels, thereby increasing intracellular calcium and ultimately insulin secretion. 14-3-3 proteins, and in particular 14-3-3ζ, have been found to regulate ATP synthase and mitochondrial respiration, suggesting that members of this family of scaffold proteins in β-cells may influence glucose-stimulated insulin secretion (GSIS) and glucose homeostasis. To date, we have identified critical contributions of 14-3-3 proteins to GSIS. In mouse and human islets, pan-inhibition of 14-3-3 proteins with cell-permeable inhibitors potentiated ex-vivo GSIS. This was associated with increased mitochondrial function, as oxygen consumption and ATP synthesis rates were significantly enhanced. Moreover, increased ATP production was confirmed with luciferase-based assays. Of the seven isoforms, we previously reported critical metabolic roles of 14-3-3ζ in glucose homeostasis, and to understand its role in β-cells, β-cell-specific knockout mice (Ins1CreThor:14-3-3ζfl/fl, β-KO) were generated. When compared to control mice, no differences in body weights or glucose and insulin tolerance were observed, but β-KO mice displayed significantly enhanced insulin secretion following i.p. glucose (2 g/kg), and ex vivo islet perifusion studies revealed enhanced 2nd-phase secretion of GSIS from β-KO islets. Similar to pan-14-3-3 inhibition, increased mitochondrial activity and ATP synthesis were detected in islets of β-KO mice. In summary, these results demonstrate critical functions of 14-3-3ζ and its related proteins in mitochondrial activity in β-cells and the regulation of GSIS. These data also suggest that 14-3-3ζ inhibition may represent a promising target to enhance pancreatic β-cell function in the context of diabetes. Disclosure Y. Mugabo: None. M. Galipeau: None. J. Tan: None. E. Fadzeyeva: None. R. Grygorczyk: None. G.E. Lim: None. Funding Canadian Institutes of Health Research; Canada Research Chairs Program (PJT-153144)

1906-P: Glucose-Dependent Insulinotropic Polypeptide (GIP) Reduces Bone Resorption in Patients with Type 2 Diabetes

In healthy individuals, GIP enhances insulin secretion and reduces bone resorption As GIP has impaired insulinotropic effects in patients with type 2 diabetes (T2D), we investigated the effect of glucose and GIP on bone resorption in these patients. In a randomized, double-blinded, cross-over study, 12 male patients with T2D underwent six study days with 90-minute GIP or matching placebo (saline) infusions on paired days with plasma glucose (PG) clamped at three levels: Fasting hyperglycemia (mean PG ∼8 mmol/l), aggravated hyperglycemia (mean PG ∼12 mmol/l), or insulin-induced hypoglycemia (mean PG 3-8 mmol/l). Primary outcome measure was CTX (a marker of bone resorption expressed as %-change from baseline ± SD). On days with fasting hyperglycemia, CTX was suppressed by 36±15% during GIP compared to 0±9% during saline infusion (p&amp;lt;0.0001). On days with aggravated hyperglycemia, CTX was suppressed by 47±23% during GIP compared to 10±9% during saline infusion (p=0.0005). On days with insulin-induced hypoglycemia, CTX was suppressed by 40±15% during GIP compared to 12±11% during saline infusion (p&amp;lt;0.0001). In conclusion, Short-term GIP infusions during hyperglycemia reduce bone resorption by more than a third suggesting that GIP has preserved effects on bone in patients with T2D. Disclosure M.B. Christensen: None. A. Lund: Speaker’s Bureau; Self; AstraZeneca, Novo Nordisk A/S, Sanofi. N.R. Jørgensen: None. J.J. Holst: Advisory Panel; Self; AstraZeneca, Merck Sharp &amp; Dohme Corp., Novo Nordisk A/S, Zealand Pharma A/S. Other Relationship; Spouse/Partner; Antag Therapeutics. T. Vilsbøll: Advisory Panel; Self; AstraZeneca, Mundipharma International, Novo Nordisk A/S, Sun Pharmaceutical Industries Ltd. Consultant; Self; Boehringer Ingelheim Pharmaceuticals, Inc., Lilly Diabetes, Medscape, Merck Sharp &amp; Dohme Corp., Sanofi. F.K. Knop: Advisory Panel; Self; AstraZeneca, Merck Sharp &amp; Dohme Corp., Mundipharma International, Novo Nordisk A/S, Sanofi. Consultant; Self; Carmot Therapeutics, Inc., Eli Lilly and Company, Novo Nordisk A/S. Research Support; Self; AstraZeneca, Gubra, Novo Nordisk A/S, Sanofi, Zealand Pharma A/S. Speaker’s Bureau; Self; AstraZeneca, Lupin Pharmaceuticals, Inc., Merck Sharp &amp; Dohme Corp., Norgine B.V., Novo Nordisk A/S.

Glucagon-like peptide 1 and glucose-dependent insulinotropic polypeptide stimulate release of substance P from TRPV1- and TRPA1-expressing sensory nerves.

Glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are released from enteroendocrine cells (EECs) in response to nutrient ingestion and lower blood glucose levels by stimulation of insulin secretion and thus are defined as incretins. GLP-1 receptor (GLP-1R) expression has been identified on enteric neurons that include intrinsic afferent neurons, extrinsic spinal, and vagal sensory afferents but has not been shown to have an incretin effect through these nerves. GLP-1 and GIP enter the mesenteric lymphatic fluid (MLF) after a meal via the interstitial fluid (IF) from local tissue secretion and/or blood capillaries. We tested if MLF could induce diet-dependent intransient increases in intracellular calcium ([Ca2+]i) in cultured sensory neurons. Postprandial rat MLF, collected from the superior mesenteric lymphatic duct, induced a significant twofold higher intransient increase in [Ca2+]i in primary-cultured sensory neurons than MLF from fasted rats. Inhibition of transient receptor potential vanilloid 1 (TRPV1) and TRPV1 and ankyrin 1 cation channels (TRPA1) with ruthenium red eliminated the difference. Substance P (SP) (a peptide that stimulates insulin secretion) sensor cells cocultured with sensory neurons showed both the GLP-1R agonist exendin-4 (Ex-4) and GIP induced transient increases in [Ca2+]i directly coupled to SP secretion in the sensory nerves. Ex-4-induced release of SP required expression of either TRPA1 or TRPV1. These data identify unrecognized actions of GLP-1 and GIP as incretins by acting as neurolymphocrines and suggest a mechanism for sensory nerves to respond to the postprandial state through MLF.NEW & NOTEWORTHY Glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are secreted upon eating to lower blood sugar. GLP-1 and GIP were found to induce the secretion of substance P (SP) from cultured sensory nerves. SP enhances insulin secretion. Mesenteric lymphatic fluid (MLF) also stimulates sensory neurons in a diet-dependent manner. These studies identify new actions of GLP-1 and GIP as incretins and suggest a mechanism for sensory nerves to respond to diet through MLF.

Deciphering genetic signatures by whole exome sequencing in a case of co-prevalence of severe renal hypouricemia and diabetes with impaired insulin secretion

BackgroundRenal hypouricemia (RHUC) is a hereditary disorder where mutations in SLC22A12 gene and SLC2A9 gene cause RHUC type 1 (RHUC1) and RHUC type 2 (RHUC2), respectively. These genes regulate renal tubular reabsorption of urates while there exist other genes counterbalancing the net excretion of urates including ABCG2 and SLC17A1. Urate metabolism is tightly interconnected with glucose metabolism, and SLC2A9 gene may be involved in insulin secretion from pancreatic β-cells. On the other hand, a myriad of genes are responsible for the impaired insulin secretion independently of urate metabolism.Case presentationWe describe a 67 year-old Japanese man who manifested severe hypouricemia (0.7 mg/dl (3.8–7.0 mg/dl), 41.6 μmol/l (226–416 μmol/l)) and diabetes with impaired insulin secretion. His high urinary fractional excretion of urate (65.5%) and low urinary C-peptide excretion (25.7 μg/day) were compatible with the diagnosis of RHUC and impaired insulin secretion, respectively. Considering the fact that metabolic pathways regulating urates and glucose are closely interconnected, we attempted to delineate the genetic basis of the hypouricemia and the insulin secretion defect observed in this patient using whole exome sequencing. Intriguingly, we found homozygous Trp258* mutations in SLC22A12 gene causing RHUC1 while concurrent mutations reported to be associated with hyperuricemia were also discovered including ABCG2 (Gln141Lys) and SLC17A1 (Thr269Ile). SLC2A9, that also facilitates glucose transport, has been implicated to enhance insulin secretion, however, the non-synonymous mutations found in SLC2A9 gene of this patient were not dysfunctional variants. Therefore, we embarked on a search for causal mutations for his impaired insulin secretion, resulting in identification of multiple mutations in HNF1A gene (MODY3) as well as other genes that play roles in pancreatic β-cells. Among them, the Leu80fs in the homeobox gene NKX6.1 was an unreported mutation.ConclusionWe found a case of RHUC1 carrying mutations in SLC22A12 gene accompanied with compensatory mutations associated with hyperuricemia, representing the first report showing coexistence of the mutations with opposed potential to regulate urate concentrations. On the other hand, independent gene mutations may be responsible for his impaired insulin secretion, which contains novel mutations in key genes in the pancreatic β-cell functions that deserve further scrutiny.

LSD1 inhibition yields functional insulin-producing cells from human embryonic stem cells

BackgroundHuman embryonic stem cells represent a potentially unlimited source of insulin-producing cells for diabetes therapy. While tremendous progress has been made in directed differentiation of human embryonic stem cells into IPCs in vitro, the mechanisms controlling its differentiation and function are not fully understood. Previous studies revealed that lysine-specific demethylase 1(LSD1) balanced the self-renewal and differentiation in human induced pluripotent stem cells and human embryonic stem cells. This study aims to explore the role of LSD1 in directed differentiation of human embryonic stem cells into insulin-producing cells.MethodsHuman embryonic stem cell line H9 was induced into insulin-producing cells by a four-step differentiation protocol. Lentivirus transfection was applied to knockdown LSD1 expression. Immunofluorescence assay and flow cytometry were utilized to check differentiation efficiency. Western blot was used to examine signaling pathway proteins and differentiation-associated proteins. Insulin/C-peptide release was assayed by ELISA. Statistical analysis between groups was carried out with one-way ANOVA tests or a student’s t test when appropriate.ResultsInhibition or silencing LSD1 promotes the specification of pancreatic progenitors and finally the commitment of functional insulin-producing β cells; Moreover, inhibition or silencing LSD1 activated ERK signaling and upregulated pancreatic progenitor associated genes, accelerating pre-maturation of pancreatic progenitors, and conferred the NKX6.1+ population with better proliferation ability. IPCs with LSD1 inhibitor tranylcypromine treatment displayed enhanced insulin secretion in response to glucose stimulation.ConclusionsWe identify a novel role of LSD1 inhibition in promoting IPCs differentiation from hESCs, which would be emerged as potential intervention for generation of functional pancreatic β cells to cure diabetes.

Role of Impaired Nutrient and Oxygen Deprivation Signaling and Deficient Autophagic Flux in Diabetic CKD Development: Implications for Understanding the Effects of Sodium-Glucose Cotransporter 2-Inhibitors.

Growing evidence indicates that oxidative and endoplasmic reticular stress, which trigger changes in ion channels and inflammatory pathways that may undermine cellular homeostasis and survival, are critical determinants of injury in the diabetic kidney. Cells are normally able to mitigate these cellular stresses by maintaining high levels of autophagy, an intracellular lysosome-dependent degradative pathway that clears the cytoplasm of dysfunctional organelles. However, the capacity for autophagy in both podocytes and renal tubular cells is markedly impaired in type 2 diabetes, and this deficiency contributes importantly to the intensity of renal injury. The primary drivers of autophagy in states of nutrient and oxygen deprivation-sirtuin-1 (SIRT1), AMP-activated protein kinase (AMPK), and hypoxia-inducible factors (HIF-1α and HIF-2α)-can exert renoprotective effects by promoting autophagic flux and by exerting direct effects on sodium transport and inflammasome activation. Type 2 diabetes is characterized by marked suppression of SIRT1 and AMPK, leading to a diminution in autophagic flux in glomerular podocytes and renal tubules and markedly increasing their susceptibility to renal injury. Importantly, because insulin acts to depress autophagic flux, these derangements in nutrient deprivation signaling are not ameliorated by antihyperglycemic drugs that enhance insulin secretion or signaling. Metformin is an established AMPK agonist that can promote autophagy, but its effects on the course of CKD have been demonstrated only in the experimental setting. In contrast, the effects of sodium-glucose cotransporter-2 (SGLT2) inhibitors may be related primarily to enhanced SIRT1 and HIF-2α signaling; this can explain the effects of SGLT2 inhibitors to promote ketonemia and erythrocytosis and potentially underlies their actions to increase autophagy and mute inflammation in the diabetic kidney. These distinctions may contribute importantly to the consistent benefit of SGLT2 inhibitors to slow the deterioration in glomerular function and reduce the risk of ESKD in large-scale randomized clinical trials of patients with type 2 diabetes.

GLP-1 Analogs and DPP-4 Inhibitors in Type 2 Diabetes Therapy: Review of Head-to-Head Clinical Trials.

The incretin hormones glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are released from enteroendocrine cells in response to the presence of nutrients in the small intestines. These homones facilitate glucose regulation by stimulating insulin secretion in a glucose dependent manner while suppressing glucagon secretion. In patients with type 2 diabetes (T2DM), an impaired insulin response to GLP-1 and GIP contributes to hyperglycemia. Dipeptidyl peptidase-4 (DPP-4) inhibitors block the breakdown of GLP-1 and GIP to increase levels of the active hormones. In clinical trials, DPP-4 inhibitors have a modest impact on glycemic control. They are generally well-tolerated, weight neutral and do not increase the risk of hypoglycemia. GLP-1 receptor agonists (GLP-1 RA) are peptide derivatives of either exendin-4 or human GLP-1 designed to resist the activity of DPP-4 and therefore, have a prolonged half-life. In clinical trials, they have demonstrated superior efficacy to many oral antihyperglycemic drugs, improved weight loss and a low risk of hypoglycemia. However, GI adverse events, particularly nausea, vomiting, and diarrhea are seen. Both DPP-4 inhibitors and GLP-1 RAs have demonstrated safety in robust cardiovascular outcome trials, while several GLP-1 RAs have been shown to significantly reduce the risk of major adverse cardiovascular events in persons with T2DM with pre-existing cardiovascular disease (CVD). Several clinical trials have directly compared the efficacy and safety of DPP-4 inhibitors and GLP-1 RAs. These studies have generally demonstrated that the GLP-1 RA provided superior glycemic control and weight loss relative to the DPP-4 inhibitor. Both treatments were associated with a low and comparable incidence of hypoglycemia, but treatment with GLP-1 RAs were invariably associated with a higher incidence of GI adverse events. A few studies have evaluated switching patients from DPP-4 inhibitors to a GLP-1RA and, as expected, improved glycemic control and weight loss are seen following the switch. According to current clinical guidelines, GLP-1RA and DPP-4 inhibitors are both indicated for the glycemic management of patients with T2DM across the spectrum of disease. GLP-1RA may be preferred over DPP- 4 inhibitors for many patients because of the greater reductions in hemoglobin A1c and weight loss observed in the clinical trials. Among patients with preexisting CVD, GLP-1 receptor agonists with a proven cardiovascular benefit are indicated as add-on to metformin therapy.

Loss of β‐cell Gαz protects against high‐fat diet induced glucose intolerance by preserving incretin responsiveness and enhancing insulin secretion

Cyclic‐adenosine monophosphate (cAMP) is an important secondary messenger in the insulin‐secreting pancreatic β‐cells. Intracellular production of cAMP is augmented by stimulatory heterotrimeric G proteins (Gs) and blocked by inhibitory heterotrimeric G proteins (Gi). Pancreatic expression of the unique Gi family member, Gαz, is restricted to the islet, which contains β‐cells and other endocrine cell types, including glucagon/GLP‐1‐secreting alpha cells and somatostatin‐secreting delta cells. We have previously shown that full body Gαz‐null mice are protected from hyperglycemia in several diabetes model systems. Here, we use a high fat diet (HFD) model of obesity and insulin resistance to explore the impact of loss of beta‐cell Gαz on the type 2 diabetes (T2D) phenotype of male C57BL/6J mice. We hypothesized that loss of beta‐cell Gαz in the context of HFD feeding would enhance insulin secretion, protecting mice from HFD‐induced glucose intolerance. To generate beta‐cell specific Gαz‐null and control mice, we bred Gαz flox/flox mice with rat insulin promoter RIP‐Creherr mice. At 12 weeks of age, transgenic Gαz‐null mice were fed a diet containing 45 kcal% fat or a 10 kcal% fat control diet for 26 weeks. At study end, mice were metabolically phenotyped and sacrificed for whole pancreas and islet phenotyping. Beta‐cell‐specific Gαz loss protected mice from HFD induced glucose intolerance due to a selective effect on islet insulin secretion, particularly in concert with a stable agonist of the Gs‐coupled GLP‐1 receptor. This protective effect was independent of changes in Gαz‐coupled EP3 receptor signaling, as neither EP3 expression nor production of its endogenous ligand, PGE2, were enhanced in islets from HFD‐fed mice. This protective effect was also beta‐cell autonomous, as both full‐body and beta‐cell‐specific Gαz‐null mice had enhanced GLP‐1‐positive alpha‐cells, albeit with no changes in active GLP‐1 secretion. Finally, gene microarray results confirm that Gαz loss and HFD feeding synergize to differentially regulate islet gene expression, specifically up‐regulating secretion‐centric genes, many of which are related to cAMP signaling. This study presents a novel inhibitory mechanism of receptor‐independent G‐protein signaling that may hold promise for developing beta‐cell targeted diabetes therapeutics.

Peripheral cannabinoid CB1 receptors control nutrient‐induced incretin secretion in vivo

Enteroendocrine cells along the intestinal epithelium secrete incretins and related hormones (i.e. GIP, GLP‐1, and CCK‐8) following ingestion of nutrients. Incretins bind and activate their respective receptors on cells located in – but not limited to – the endocrine pancreas, which enhances insulin secretion and maintains a tight window of circulating blood glucose. The ability to adequately regulate circulating glucose levels is lost in patients living with prediabetes and type 2 diabetes. The endocannabinoid system controls food intake and glucose homeostasis, and its activity becomes upregulated in the proximal small‐intestinal epithelium of Western‐diet induced obese mice. Cannabinoid subtype‐1 receptors (CB1Rs) are present in various incretin‐producing cells in the intestinal epithelium (e.g. K, L, I‐cells), yet their function in these cell types are poorly characterized. In the current investigation, we examined the ability for CB1Rs to control incretin secretion in vivo in male C57CL/6N mice. We administered the cannabinoid receptor agonist, WIN 55,212‐2 (WIN, 3 mg per kg), alone and in combination with the peripherally‐restricted neutral CB1R antagonist, AM6545 (10 mg per kg). Incretin secretion was then stimulated via corn oil gavage (0.5 mL 30 minutes after drug treatment) and circulating incretins were quantified by sensitive enzyme‐linked immunosorbent assays from plasma obtained 30 minutes later. Activating CB1Rs with WIN blunted corn‐oil induced secretion of GIP, active GLP‐1 and CCK‐8. Co‐administration of WIN with AM6545 completely reversed the inhibitory actions of WIN on incretin release. Collectively, our results suggest that CB1Rs in the small‐intestinal epithelium control nutrient induced incretin release. The functional consequences of these findings on glucose homeostasis are currently under investigation.Support or Funding InformationAcknowledgementsFunded by the National Institutes of Health, USA (Grant no. DK119498, DK114978, and DA034009)