Aims/hypothesis: The gut hormone glucagonlike peptide-1 (GLP-1) decreases beta cell apoptosis in a protein kinase B (PKB)-dependent fashion, and increases islet cell mass and function in vivo. In contrast, cytokines induce beta cell apoptosis, leading to decreased islet mass and type 1 diabetes. In the present study we used rat INS-1E beta cells and primary rat islet cells to examine the potential role of PKB as a mediator of the effect of GLP-1 on cytokine-induced apoptosis. Methods: Cell viability was determined by MTTassay, and apoptosis and necrosis by Hoechst 33342-propidium iodide staining. Immunoblot analysis was used to detect changes in protein expression, including active (phosphorylated) and total PKB, phosphorylated and total glycogen synthase kinase-3β, activated caspase-3 and inducible nitric oxide synthase. Reactive oxygen species were determined by 1,7-dichlorofluorescein (DCF) analysis, and mutant forms of PKB were introduced into cells using adenoviral vectors. Results: Incubation of INS-1E cells with cytokines (IL-1β, TNF-α and interferon-γ; 10-50 ng/ml) for 18 h significantly decreased cell viability (by 44%, p<0.001), cell proliferation (by 80%, p<0.001), and activation of PKB (by 67%, p<0.001). Pre-treatment with exendin-4 (10 −7 mol/l), a long-acting GLP-1 receptor agonist, partially protected the cells against cytokine-induced toxicity (p<0.01) in association with a reduction in cytokine-induced inhibition of PKB phosphorylation (p<0.05). Exendin-4 pre-treatment did not change cell proliferation. Cytokine treatment increased apoptosis (by 156%, p<0.05) and necrosis (from undetectable to 2.6% of cells). These increases were both reduced by pre-treatment with exendin-4 (p<0.05-0.01). Furthermore, cytokine-induced apoptosis and necrosis were significantly increased in cells infected with kinase-dead PKB (p<0.05), and the protective effect of exendin-4 on both parameters was fully abolished in these cells. Similar changes were observed in primary islet cells. In parallel with these changes, exendin-4 decreased the cytokine-induced activation of caspase-3 (by 46%, p<0.05), and decreased levels of inducible nitric oxide synthase (by 71%, p<0.05) and reactive oxygen species (by 27%, p< 0.05). Conclusions/interpretation: The results of our study indicate that GLP-1 plays a protective role against cytokineinduced apoptosis and necrosis in beta cells through a PKB-dependent signalling pathway.

To determine whether glucagon-like peptide (GLP)-1 increases insulin sensitivity in addition to stimulating insulin secretion, we studied totally depancreatized dogs to eliminate GLP-1's incretin effect. Somatostatin was infused (0.8 microg x kg(-1) x min(-1)) to inhibit extrapancreatic glucagon in dogs, and basal glucagon was restored by intraportal infusion (0.65 ng x kg(-1) x min(-1)). To simulate the residual intraportal insulin secretion in type 2 diabetes, basal intraportal insulin infusion was given to obtain plasma glucose concentrations of approximately 10 mmol/l. Glucose was clamped at this level for the remainder of the experiment, which included peripheral insulin infusion (high dose, 5.4 pmol x kg(-1) x min(-1), or low dose, 0.75 pmol x kg(-1) x min(-1)) with or without GLP-1(7-36) amide (1.5 pmol x kg(-1) x min(-1)). Glucose production and utilization were measured with 3-[3H]glucose, using radiolabeled glucose infusates. In 12 paired experiments with six dogs at the high insulin dose, GLP-1 infusion resulted in higher glucose requirements than saline (60.9+/-11.0 vs. 43.6+/-8.3 micromol x kg(-1) x min(-1), P< 0.001), because of greater glucose utilization (72.6+/-11.0 vs. 56.8+/-9.7 micromol x kg(-1) x min(-1), P<0.001), whereas the suppression of glucose production was not affected by GLP-1. Free fatty acids (FFAs) were significantly lower with GLP-1 than saline (375.3+/-103.0 vs. 524.4+/-101.1 micromol/l, P<0.01), as was glycerol (77.9+/-17.5 vs. 125.6+/-51.8 micromol/l, P<0.05). GLP-1 receptor gene expression was found using reverse transcriptase-polymerase chain reaction of poly(A)-selected RNA in muscle and adipose tissue, but not in liver. Low levels of GLP-1 receptor gene expression were also found in adipose tissue using Northern blotting. In 10 paired experiments with five dogs at the low insulin dose, GLP-1 infusion did not affect glucose utilization or FFA and glycerol suppression when compared with saline, suggesting that GLP-1's effect on insulin action was dependent on the insulin dose. In conclusion, in depancreatized dogs, GLP-1 potentiates insulin-stimulated glucose utilization, an effect that might be contributed in part by GLP-1 potentiation of insulin's antilipolytic action.

The intestinal glucagon-like peptides, GLP-1 and GLP-2, are important regulators of nutrient intake, digestion, absorption and metabolism. Extensive studies have shown that the co-secretion of GLP-1 and GLP-2 is regulated by a variety of dietary nutrient, neural and endocrine inputs. Furthermore, secretion of these peptides is altered in a number of pathological conditions, including type 2 diabetes and obesity. The purpose of this review is to discuss the regulation of GLP-1 and GLP-2 secretion in both health and disease.

Glucagon-like peptide-1 (GLP-1) released from the intestine is a potent stimulator of glucose-dependent insulin secretion. To elucidate the factors regulating GLP-1 secretion, we have studied the enteroendocrine GLUTag cell line. GLP-1 secretion was stimulated in a dosedependent fashion by activation of protein kinase A or C with forskolin or phorbol 12,13-dibutyrate, respectively (by 2.3 Ϯ 0.5-fold at 100 M and 4.3 Ϯ 0.6-fold at 0.3 M, respectively; P Ͻ 0.01-0.001). Of the regulatory peptides tested, only glucose-dependent insulinotropic peptide stimulated the release of GLP-1 (by 2.3 Ϯ 0.2-fold at 0.1 M; P Ͻ 0.001); glucagon was without effect, and paradoxically, the inhibitory neuropeptide somatostatin-14 increased secretion slightly (by 1.6 Ϯ 0.3-fold at 0.01 M; P Ͻ 0.05). In tests of several neurotransmitters, only the cholinergic agonists carbachol and bethanechol stimulated peptide secretion in a dose-dependent fashion (by 2.3 Ϯ 0.5-and 1.7 Ϯ 0.3-fold at 1000 M; P Ͻ 0.05-0.001); the ␤-adrenergic agonist isoproterenol and the chloride channel inhibitor ␥-aminobutyric acid did not affect release of GLP-1. Long chain monounsaturated fatty acids (18:1), but not saturated fatty acids (16:0), also stimulated the release of GLP-1 (by 1.7 Ϯ 0.1-fold at 150 M; P Ͻ 0.001). Consistent with the presence of a cAMP response element in the proglucagon gene, activation of the protein kinase A-dependent pathway with forskolin increased proglucagon messenger RNA transcript levels by 2-fold (P Ͻ 0.05); glucose-dependent insulinotropic peptide and phorbol 12,13-dibutyrate were without effect. Therefore, by comparison with results obtained using primary L cell cultures or in vivo models, GLUTag cells appear to respond appropriately to the regulatory mechanisms controlling intestinal GLP-1 secretion. (Endocrinology 139: 4108 -4114, 1998) G LUCAGON-LIKE peptide-1 (GLP-1) is a potent stimulator of glucose-dependent insulin secretion. Administration of GLP-1 to patients with type II diabetes normalizes both fasting and postprandial glycemia (1-13), not only through stimulation of insulin release, but also through concomitant inhibition of glucagon secretion (5, 9, 11, 13) and gastric motility (5, 14) and, possibly, enhancement of insulin sensitivity (8,15,16). GLP-1 is normally synthesized and secreted by the intestinal L cell (17-21); thus, stimulation of endogenous secretion represents an alternative approach to increasing levels of GLP-1 in type II diabetes. It is therefore essential that the factors regulating GLP-1 release from the L cell be elucidated.A number of in vitro cell culture systems have been developed as models of the intestinal L cell, each of which has both advantages and drawbacks. For example, fetal rat intestinal cell (FRIC) cultures are heterogeneous in their cell population, although they have proven to be an excellent model of the rat L cell otherwise, releasing GLP-1 in response to a wide variety of different signal transduction pathways and extracellular mediators (18, 20 -24). FRIC cells hav...