Differential effects of oxyntomodulin and GLP-1 on glucose metabolism

Glucagon-like peptide-1 (GLP-1) and oxyntomodulin (OXM) are peptide hormones secreted postprandially from the gut that stimulate insulin secretion in a glucose-dependent manner. OXM activates both the GLP-1 receptor (GLP1R) and the glucagon receptor (GCGR). It has been suggested that OXM acutely modulates glucose metabolism solely through GLP1R agonism. Because OXM activates the GLP1R with lower affinity than GLP-1, we generated a peptide analog (Q→E, OXMQ3E) that does not exhibit glucagon receptor agonist activity but retains the same affinity as OXM for GLP1R. We compared the effects of OXM and OXMQ3E in a glucose tolerance test and, to better characterize the effect on glucose metabolism, we performed controlled infusions of OXM or OXMQ3E during a hyperglycemic clamp performed in wild-type, Glp1r(-/-), and Gcgr(-/-) mice. Our findings show that OXM, but not OXMQ3E, activates the GCGR in vivo. Second, OXM and OXMQ3E improve glucose tolerance following an acute glucose challenge and during a hyperglycemic clamp in mice. Finally, OXM infusion during a glucose clamp reduces the glucose infusion rate (GIR) despite a simultaneous increase in insulin levels in Glp1r(-/-) mice, whereas OXM and OXMQ3E increase GIR to a similar extent in Gcgr(-/-) mice. In conclusion, activation of the GCGR seems to partially attenuate the acute beneficial effects on glucose and contributes to the insulinotropic action of oxyntomodulin.     

GLP-1(7-36)amide-stimulated insulin secretion in rat islets is sodium-dependent

We examined the Na(+)-dependency of the effects of GLP-1(7-36)amide in normal, overnight cultured rat islets. It was found that GLP-1(7-36)amide stimulated insulin secretion, 45Ca(2+)-efflux, and 86Rb(+)-efflux from prelabelled islets. All these effects were abolished by omitting Na+ from the medium and replacing it with N-methyl-glucamine. This suggests that GLP-1(7-36)amide stimulates insulin secretion by depolarizing the beta-cells by increasing their permeability to Na+.     

GLP-1-(9-36) amide reduces blood glucose in anesthetized pigs by a mechanism that does not involve insulin secretion

Glucagon-like peptide 1 (GLP-1) is a potent anti-hyperglycemic hormone currently under investigation for its therapeutic potential. However, due to rapid degradation by dipeptidyl peptidase IV (DPP IV), which limits its metabolic stability and eliminates its insulinotropic activity, it has been impossible to assess its true efficacy in vivo. In chloralose-anesthetized pigs given valine-pyrrolidide (to block endogenous DPP IV activity), the independent effects of GLP-1-(7-36) amide on glucose and insulin responses to intravenous glucose were assessed, and the metabolite generated by DPP IV, GLP-1-(9-36) amide, was investigated for any ability to influence these responses. GLP-1-(7-36) amide enhanced insulin secretion (P < 0.03 vs. vehicle), but GLP-1-(9-36) amide was without effect, either alone or when coinfused with GLP-1-(7-36) amide. In contrast, GLP-1-(9-36) amide did affect glucose responses (P < 0.03). Glucose excursions were greater after saline (121 +/- 17 mmol x l(-1) x min) than after GLP-1-(9-36) amide (73 +/- 19 mmol x l(-1) x min; P < 0.05), GLP-1-(7-36) amide (62 +/- 13 mmol x l(-1) x min; P < 0.02) or GLP-1-(7-36) amide + GLP-1-(9-36) amide (50 +/-13 mmol x l(-1) x min; P < 0.005). Glucose elimination rates were faster after GLP-1-(7-36) amide + (9-36) amide (10.3 +/- 1.2%/min) than after GLP-1-(7-36) amide (7.0 +/- 0.9%/min; P < 0.04), GLP-1-(9-36) amide (6.8 +/- 1.0%/min; P < 0.03), or saline (5.4 +/- 1.2%/min; P < 0.005). Glucagon concentrations were unaffected. These results demonstrate that GLP-1-(9-36) amide neither stimulates insulin secretion nor antagonizes the insulinotropic effect of GLP-1-(7-36) amide in vivo. Moreover, the metabolite itself possesses anti-hyperglycemic effects, supporting the hypothesis that selective DPP IV action is important in glucose homeostasis.     

Iron overload in Hepc1(-/-) mice is not impairing glucose homeostasis

Diabetes Mellitus is found with increasing frequency in iron overload patients with hemochromatosis. In these conditions, the pancreas shows predominant iron overload in acini but also islet beta-cells. We assess glucose homeostasis status in iron-overloaded hepcidin-deficient mice. These mice presented with heavy pancreatic iron deposits but only in the acini. The beta-cell function was found unaffected with a normal production and secretion of insulin. The mutant mice were not diabetic, responded as the control group to glucose and insulin challenges, with no alteration of insulin signalling in the muscle and the liver. These results indicate that, beta-cells iron deposits-induced decreased insulin secretory capacity might be of primary importance to trigger diabetes in hemochromatosic patients.     

Macrophage inhibitory cytokine 1 (MIC-1/GDF15) decreases food intake, body weight and improves glucose tolerance in mice on normal & obesogenic diets

Food intake and body weight are controlled by a variety of central and peripheral factors, but the exact mechanisms behind these processes are still not fully understood. Here we show that that macrophage inhibitory cytokine-1 (MIC-1/GDF15), known to have anorexigenic effects particularly in cancer, provides protection against the development of obesity. Both under a normal chow diet and an obesogenic diet, the transgenic overexpression of MIC-1/GDF15 in mice leads to decreased body weight and fat mass. This lean phenotype was associated with decreased spontaneous but not fasting-induced food intake, on a background of unaltered energy expenditure and reduced physical activity. Importantly, the overexpression of MIC-1/GDF15 improved glucose tolerance, both under normal and high fat-fed conditions. Altogether, this work shows that the molecule MIC-1/GDF15 might be beneficial for the treatment of obesity as well as perturbations in glucose homeostasis.     

Protective effects of GLP-1 analogues exendin-4 and GLP-1(9-36) amide against ischemia-reperfusion injury in rat heart

Glucagon-Like Peptide-1 (GLP-1) is an incretin peptide secreted from intestinal L-cells, whose potent plasma glucose-lowering action has prompted intense efforts to develop GLP-1 receptor-targeting drugs for treatment of diabetic hyperglycemia. More recently, GLP-1 and its analogues have been shown to exert cardiovascular effects in a number of experimental models. Here we tested exendin-4 (Exe-4), a peptide agonist at GLP-1 receptors, and GLP-1(9-36) amide, the primary endogenous metabolite of GLP-1 (both in the concentration range 0.03-3.0 nM), for their protective effects against ischemia-reperfusion injury (IRI) in an isolated rat heart preparation. When administered, the agents were only present for the first 15 min of a 120 min reperfusion period (postconditioning protocol). Exe-4, but not GLP-1(9-36) amide, showed a strong infarct-limiting action (from 33.2% +/-2.7% to 14.5% +/-2.2% of the ischemic area, p<0.05). This infarct size-limiting effect of Exe-4 was abolished by exendin(9-39) (Exe(9-39)), a GLP-1 receptor antagonist. In contrast, both Exe-4 and GLP-1(9-36) amide were able to augment left ventricular performance (left ventricular developed pressure and rate-pressure product) during the last 60 min of reperfusion. These effects were only partially antagonized by Exe(9-39). We suggest that Exe-4, in addition to being currently exploited in treatment of diabetes, may present a suitable candidate for postconditioning trials in clinical settings of IRI. The divergent agonist effects of Exe-4 and GLP-1(9-36), along with correspondingly divergent antagonistic efficacy of Exe(9-39), seem consistent with the presence of more than one type of GLP-1 receptor in this system.     

Concentrated loss of insulin secretion in Wistar rats with normal glucose tolerance

Rats with decreased insulin response and with normal glucose tolerance were concentrated by repeated selective breeding of normal Wistar rats with low insulinogenic index. In general, the mean insulinogenic index of the inbred offsprings showed a tendency to decrease more than their parents generation. Thus mean insulinogenic indices in second (F2), third (F3) and fourth (F4) generations were significantly reduced more than the normal rats without glucose intolerance. Pancreatic islets from the F3 and F4 rats lost partially their ability to release insulin at 20 mM glucose in vitro. It is suggested that a defect responsible for the decreased insulin response in the F2, F3 and F4 rats resulted from a loss of the ability to secrete insulin in each islet, and that this defect was concentrated by repeated selective breeding of normal Wistar rats.     

Reduced GLP-1 and insulin responses and glucose intolerance after gastric glucose in GRP receptor-deleted mice

By applying a newly developed ELISA technique for determining biologically active intact glucagon-like peptide [GLP-1, GLP-1-(7-36)amide] in mouse, plasma baseline GLP-1 in normal NMRI mice was found to be normally distributed (4.5 +/- 0.3 pmol/l; n = 72). In anesthetized mice, gastric glucose (50 or 150 mg) increased plasma GLP-1 levels two- to threefold (P < 0.01). The simultaneous increase in plasma insulin correlated to the 10-min GLP-1 levels (r = 0.36, P < 0.001; n = 12). C57BL/6J mice deleted of the gastrin-releasing peptide (GRP) receptor by genetic targeting had impaired glucose tolerance (P = 0.030) and reduced early (10 min) insulin response (P = 0.044) to gastric glucose compared with wild-type controls. Also, the GLP-1 response to gastric glucose was significantly lower in the GRP receptor-deleted mice than in the controls (P = 0.045). In conclusion, this study has shown that 1) plasma levels of intact GLP-1 increase dose dependently on gastric glucose challenge in correlation with increased insulin levels in mice, and 2) intact GRP receptors are required for normal GLP-1 and insulin responses and glucose tolerance after gastric glucose in mice.     

Effects of oral fructose and glucose on plasma GLP-1 and appetite in normal subjects.

Oral glucose is a potent stimulant of glucagon-like peptide-1 (GLP-1) secretion. The effect of oral fructose on GLP-1 secretion in humans is unknown. The aims of this study were to determine (i) whether oral fructose stimulates GLP-1 secretion and (ii) the comparative effects of oral glucose and fructose on appetite. On 3 separate days, 8 fasting healthy males received, in single-blind randomized order (i) 75 g glucose, (ii) 75 fructose, or (iii) 75 g glucose followed by 75 g fructose I h later. Venous glucose, insulin and GLP-1 were measured. Appetite was assessed by visual analog questionnaires and intake of a buffet meal. Whereas glucose and fructose both increased plasma glucose, insulin and GLP-1 (P < 0.000)] for all), the response to glucose was much greater (P < 0.005 for all). There was no increase in plasma GLP-1 when fructose was given after glucose. There was no difference in food intake after oral glucose or fructose. We conclude that oral fructose (75 g) stimulates GLP-1 (and insulin) secretion, but the response is less than that to 75 g glucose. These observations suggest that neither GLP-1 nor insulin play a major role in the regulation of satiation.     

Insulin secretion-independent effects of GLP-1 on canine liver glucose metabolism do not involve portal vein GLP-1 receptors

Whether glucagon-like peptide (GLP)-1 requires the hepatic portal vein to elicit its insulin secretion-independent effects on glucose disposal in vivo was assessed in conscious dogs using tracer and arteriovenous difference techniques. In study 1, six conscious overnight-fasted dogs underwent oral glucose tolerance testing (OGTT) to determine target GLP-1 concentrations during clamp studies. Peak arterial and portal values during OGTT ranged from 23 to 65 pM and from 46 to 113 pM, respectively. In study 2, we conducted hyperinsulinemic-hyperglycemic clamp experiments consisting of three periods (P1, P2, and P3) during which somatostatin, glucagon, insulin and glucose were infused. The control group received saline, the PePe group received GLP-1 (1 pmol.kg(-1).min(-1)) peripherally, the PePo group received GLP-1 (1 pmol.kg(-1).min(-1)) peripherally (P2) and then intraportally (P3), and the PeHa group received GLP-1 (1 pmol.kg(-1).min(-1)) peripherally (P2) and then through the hepatic artery (P3) to increase the hepatic GLP-1 load to the same extent as in P3 in the PePo group (n = 8 dogs/group). Arterial GLP-1 levels increased similarly in all groups during P2 ( approximately 50 pM), whereas portal GLP-1 levels were significantly increased (2-fold) in the PePo vs. PePe and PeHa groups during P3. During P2, net hepatic glucose uptake (NHGU) increased slightly but not significantly (vs. P1) in all groups. During P3, GLP-1 increased NHGU in the PePo and PeHa groups more than in the control and PePe groups (change of 10.8 +/- 1.3 and 10.6 +/- 1.0 vs. 5.7 +/- 1.0 and 5.4 +/- 0.8 micromol.kg(-1).min(-1), respectively, P < 0.05). In conclusion, physiological GLP-1 levels increase glucose disposal in the liver, and this effect does not involve GLP-1 receptors located in the portal vein.     

Impaired insulin secretion and glucose tolerance in beta cell-selective Ca(v)1.2 Ca2+ channel null mice

Insulin is secreted from pancreatic beta cells in response to an elevation of cytoplasmic Ca(2+) resulting from enhanced Ca(2+) influx through voltage-gated Ca(2+) channels. Mouse beta cells express several types of Ca(2+) channel (L-, R- and possibly P/Q-type). beta cell-selective ablation of the gene encoding the L-type Ca(2+) channel subtype Ca(v)1.2 (betaCa(v)1.2(-/-) mouse) decreased the whole-cell Ca(2+) current by only approximately 45%, but almost abolished first-phase insulin secretion and resulted in systemic glucose intolerance. These effects did not correlate with any major effects on intracellular Ca(2+) handling and glucose-induced electrical activity. However, high-resolution capacitance measurements of exocytosis in single beta cells revealed that the loss of first-phase insulin secretion in the betaCa(v)1.2(-/-) mouse was associated with the disappearance of a rapid component of exocytosis reflecting fusion of secretory granules physically attached to the Ca(v)1.2 channel. Thus, the conduit of Ca(2+) entry determines the ability of the cation to elicit secretion.     

Enhanced GLP-1- and sulfonylurea-induced insulin secretion in islets lacking leptin signaling

We have previously reported that the absence of leptin signaling in β-cells enhances glucose-stimulated insulin secretion and improves glucose tolerance in vivo. To investigate the relevance of β-cell leptin signaling in the context of postprandial or therapeutic insulin secretion, we examined the cross talk between leptin and glucagon-like peptide (GLP)-1 and sulfonylurea actions. Single and size-matched islets isolated from control or pancreas-specific leptin receptor knockout (pancreas-ObR-KO) mice were treated either with GLP-1 or with glibenclamide. Leptin suppressed GLP-1-stimulated intracellular Ca(2+) concentrations ([Ca(2+)](i)) increase that paralleled the decrease in insulin secretion in controls. In contrast, and as expected, the ObR-KO islets were nonresponsive to leptin, and instead, showed a 2.8-fold greater GLP-1-stimulated [Ca(2+)](i) increase and a 1.7-fold greater insulin secretion. Phosphorylation of cAMP-responsive element binding protein was enhanced, and phosphodiesterase enzymatic activity was suppressed in MIN6 β-cells with ObR knockdown compared with controls. The ObR-KO islets also showed significantly higher glibenclamide-induced insulin secretion compared with control islets, whereas [Ca(2+)](i) was similar to the controls. These data support enhanced insulinotropic effects of glucose, GLP-1, and sulfonylureas in the islets lacking leptin signaling with potential therapeutic implications.     

Enhanced glucose tolerance in pancreatic-derived factor (PANDER) knockout C57BL/6 mice

Pancreatic-derived factor (PANDER; also known as FAM3B) is a uniquely structured protein strongly expressed within and secreted from the endocrine pancreas. PANDER has been hypothesized to regulate fasting and fed glucose homeostasis, hepatic lipogenesis and insulin signaling, and to serve a potential role in the onset or progression of type 2 diabetes (T2D). Despite having potentially pivotal pleiotropic roles in glycemic regulation and T2D, there has been limited generation of stable animal models for the investigation of PANDER function, and there are no models on well-established genetic murine backgrounds for T2D. Our aim was to generate an enhanced murine model to further elucidate the biological function of PANDER. Therefore, a pure-bred PANDER knockout C57BL/6 (PANKO-C57) model was created and phenotypically characterized with respect to glycemic regulation and hepatic insulin signaling. The PANKO-C57 model exhibited an enhanced metabolic phenotype, particularly with regard to enhanced glucose tolerance. Male PANKO-C57 mice displayed decreased fasting plasma insulin and C-peptide levels, whereas leptin levels were increased as compared with matched C57BL/6J wild-type mice. Despite similar peripheral insulin sensitivity between both groups, hepatic insulin signaling was significantly increased during fasting conditions, as demonstrated by increased phosphorylation of hepatic PKB/Akt and AMPK, along with mature SREBP-1 expression. Insulin stimulation of PANKO-C57 mice resulted in increased hepatic triglyceride and glycogen content as compared with wild-type C57BL/6 mice. In summary, the PANKO-C57 mouse represents a suitable model for the investigation of PANDER in multiple metabolic states and provides an additional tool to elucidate the biological function and potential role in T2D.KEY WORDS: Pancreatic-derived factor, FAM3B, Knockout model, Glycemic regulation, Hepatic insulin signaling, Glucose tolerance     

GLP-1(7-36)NH2的促胰岛素分泌作用和胞内cAMP浓度改变与胰岛素mRNA表达

目的:观察GLP-1(7-36)NH2的促胰岛素分泌作用,并进一步探讨GLP-1(7-36)NH2促进胰岛素分泌的机制.方法:放射免疫分析和细胞原位杂交的方法.结果:随着GLP-1(7-36)NH2浓度的增加,胰岛素分泌逐渐增加;8-Br-cAMP增加了胰岛素分泌,GLP-1(7-36)NH2则增加了胞内第二信使cAMP的浓度;GLP-1(7-36)NH2可增加胰岛素mRNA的表达.结论:GLP-1(7-36)NH2可促进胰岛素分泌,在一定范围内呈剂量依赖性,其机制与GLP-1(7-36)NH2增加胞内第二信使cAMP和促进胰岛素基因表达有关.     

The effects of postprandial glucose and insulin levels on postprandial endothelial function in subjects with normal glucose tolerance

ABSTRACT: BACKGROUND: Previous studies have demonstrated that postprandial hyperglycemia attenuates brachial artery flow-mediated dilation (FMD) in prediabetic patients, in diabetic patients, and even in normal subjects. We have previously reported that postprandial hyperinsulinemia also attenuates FMD. In the present study we evaluated the relationship between different degrees of postprandial attenuation of FMD induced by postprandial hyperglycemia and hyperinsulinemia and differences in ingested carbohydrate content in non-diabetic individuals. METHODS: Thirty-seven healthy subjects with no family history of diabetes were divided into 3 groups: a 75-g oral glucose loading group (OG group) (n = 14), a test meal group (TM group) (n = 12; 400 kcal, carbohydrate content 40.7 g), and a control group (n = 11). The FMD was measured at preload (FMD0) and at 60 minutes (FMD60) and 120 (FMD120) minutes after loading. Plasma glucose (PG) and immunoreactive insulin (IRI) levels were determined at preload (PG0, IRI0) and at 30 (PG30, IRI30), 60 (PG60, IRI60), and 120 (PG120, IRI120) minutes after loading.ResultPercentage decreases from FMD0 to FMD60 were significantly greater in the TM group ([MINUS SIGN]21.19 % [PLUS-MINUS SIGN] 17.90 %; P < 0.001) and the OG group ([MINUS SIGN]17.59 % [PLUS-MINUS SIGN] 26.64 %) than in the control group (6.46 % [PLUS-MINUS SIGN] 9.17 %; P < 0.01), whereas no significant difference was observed between the TM and OG groups. In contrast, the percentage decrease from FMD0 to FMD120 was significantly greater in the OG group ([MINUS SIGN]18.91 % [PLUS-MINUS SIGN] 16.58 %) than in the control group (6.78 % [PLUS-MINUS SIGN] 11.43 %; P < 0.001) or the TM group (5.22 % [PLUS-MINUS SIGN] 37.22 %; P < 0.05), but no significant difference was observed between the control and TM groups. The FMD60 was significantly correlated with HOMA-IR (r = [MINUS SIGN]0.389; P < 0.05). In contrast, FMD120 was significantly correlated with IRI60 (r = [MINUS SIGN]0.462; P < 0.05) and the AUC of IRI (r = [MINUS SIGN]0.468; P < 0.05). Furthermore, the percentage change from FMD0 to FMD120 was significantly correlated with the CV of PG (r = 0.404; P < 0.05), IRI60 (r = 0.401; p < 0.05) and the AUC of IRI (r = 0.427; P < 0.05). No significant correlation was observed between any other FMDs and glucose metabolic variables. CONCLUSION: Differences in the attenuation of postprandial FMD induced by different postprandial insulin levels may occur a long time postprandially but not shortly after a meal.     

Glucagon-like peptide-1(1-37) can enhance blood glucose homeostasis in mice

Tyrosyl O-sulfation is a common posttranslational derivatization of proteins that may also modify regulatory peptides. Among these are members of the cholecystokinin (CCK)/gastrin family. While sulfation of gastrin peptides is without effect on the bioactivity, O-sulfation is crucial for the cholecystokinetic activity (i.e. gallbladder emptying) of CCK peptides. Accordingly, the purification of CCK as a sulfated peptide was originally monitored by its gallbladder emptying effect. Since then, the dogma has prevailed that CCK peptides are always sulfated. The dogma is correct in a semantic context since the gallbladder expresses only the CCK-A receptor that requires sulfation of the ligand. CCK peptides, however, are also ligands for the CCK-B receptors that do not require ligand sulfation. Consequently, unsulfated CCK peptides may act via CCK-B receptors. Since in vivo occurrence of unsulfated products of proCCK with an intact α-amidated C-terminal tetrapeptide sequence (-Trp-Met-Asp-PheNH(2)) has been reported, it is likely that unsulfated CCK peptides constitute a separate hormone system that acts via CCK-B receptors. This review discusses the occurrence, molecular forms, and possible physiological as well as pathophysiological significance of unsulfated CCK peptides.