Nurit Kaiser

Role of Lipid Peroxidation and PPAR-δ in Amplifying Glucose-Stimulated Insulin Secretion

OBJECTIVE Previous studies show that polyunsaturated fatty acids (PUFAs) increase the insulin secretory capacity of pancreatic β-cells. We aimed at identifying PUFA-derived mediators and their cellular targets that are involved in the amplification of insulin release from β-cells preexposed to high glucose levels. RESEARCH DESIGN AND METHODS The content of fatty acids in phospholipids of INS-1E β-cells was determined by lipidomics analysis. High-performance liquid chromatography was used to identify peroxidation products in β-cell cultures. Static and dynamic glucose-stimulated insulin secretion (GSIS) assays were performed on isolated rat islets and/or INS-1E cells. The function of peroxisome proliferator–activated receptor-δ (PPAR-δ) in regulating insulin secretion was investigated using pharmacological agents and gene expression manipulations. RESULTS High glucose activated cPLA2 and, subsequently, the hydrolysis of arachidonic and linoleic acid (AA and LA, respectively) from phospholipids in INS-1E cells. Glucose also increased the level of reactive oxygen species, which promoted the peroxidation of these PUFAs to generate 4-hydroxy-2E-nonenal (4-HNE). The latter mimicked the GSIS-amplifying effect of high glucose preexposure and of the PPAR-δ agonist GW501516 in INS-1E cells and isolated rat islets. These effects were blocked with GSK0660, a selective PPAR-δ antagonist, and the antioxidant N-acetylcysteine or by silencing PPAR-δ expression. High glucose, 4-HNE, and GW501516 also induced luciferase expression in a PPAR-δ–mediated transactivation assay. Cytotoxic effects of 4-HNE were observed only above the physiologically effective concentration range. CONCLUSIONS Elevated glucose levels augment the release of AA and LA from phospholipids and their peroxidation to 4-HNE in β-cells. This molecule is an endogenous ligand for PPAR-δ, which amplifies insulin secretion in β-cells.

Impaired Glucose-Stimulated Insulin Secretion Is Coupled With Exocrine Pancreatic Lesions in the Cohen Diabetic Rat

OBJECTIVE—The Cohen diabetes–sensitive rat develops postprandial hyperglycemia when fed a high-sucrose, copper-poor diet, whereas the Cohen diabetes–resistant rat maintains normoglycemia. The pathophysiological basis of diabetes was studied in the Cohen diabetic rat centering on the interplay between the exocrine and endocrine compartments of the pancreas. RESEARCH DESIGN AND METHODS—Studies used male Cohen diabetes–sensitive and Cohen diabetes–resistant rats fed 1-month high-sucrose, copper-poor diet. Serum insulin and glucose levels were measured during glucose and insulin tolerance tests. The pancreas was evaluated for weight, insulin content, macrophage, and fat infiltration. Glucose-stimulated insulin secretion (GSIS) was determined in isolated perfused pancreas and in islets. RESULTS—Hyperglycemic Cohen diabetes–sensitive rats exhibited reduced pancreatic weight with lipid deposits and interleukin-1β–positive macrophage infiltration in the exocrine pancreas. Islet morphology was preserved, and total pancreatic insulin content did not differ from that of Cohen diabetes–resistant rats. Lipids did not accumulate in skeletal muscle, nor was insulin resistance observed in hyperglycemic Cohen diabetes–sensitive rats. Intravenous glucose-tolerance test revealed markedly elevated glucose levels associated with diminished insulin output. Insulin release was induced in vivo by the non-nutrient secretagogues arginine and tolbutamide, suggesting a selective unresponsiveness to glucose. Decreased GSIS was observed in the isolated perfused pancreas of the hyperglycemic Cohen diabetes–sensitive rat, whereas islets isolated from these rats exhibited glucose-dependent insulin secretion and proinsulin biosynthesis. CONCLUSIONS—The association of the in vivo insulin secretory defect with lipid accumulation and activated macrophage infiltration in the exocrine pancreas suggests that changes in the islet microenvironment are the culprit in the insulin secretory malfunction observed in vivo.

Improvement of ER stress-induced diabetes by stimulating autophagy

Pancreatic β-cell dysfunction is central in diabetes. The diabetic milieu may impair proinsulin folding, leading to β-cell endoplasmic reticulum (ER) stress and apoptosis, and thus a worsening of the diabetes. Autophagy is crucial for the well-being of the β-cell; however, the impact of stimulating autophagy on β-cell adaptation to ER stress is unknown. We studied the crosstalk between ER stress and autophagy in a rodent model of diabetes, called Akita, in which proinsulin gene mutation leads to protein misfolding and β-cell demise. We found that proinsulin misfolding stimulates autophagy and, in symmetry, inhibition of autophagy induces β-cell stress and apoptosis. Under conditions of excessive proinsulin misfolding, stimulation of autophagy by inhibiting MTORC1 alleviates stress and prevents apoptosis. Moreover, treatment of diabetic Akita mice with the MTORC1 inhibitor rapamycin improves diabetes and prevents β-cell apoptosis. Thus, autophagy is a central adaptive mechanism in β-cell stress. Stimulation of autophagy may become a novel therapeutic strategy in diabetes.

Characterization of the Unusual Insulin of Psammomys obesus, a Rodent With Nutrition-Induced NIDDM-Like Syndrome

Psammomys obesus fed a high-calorie diet develops a NIDDM-like syndrome. The use of reverse-phase highperformance liquid chromatography (HPLC) to study Psammomys insulin biosynthesis and release revealed a very delayed elution time for the Psammomys insulin peak appearing near the position of human proinsulin. This unusual peak was initially thought to represent partially processed insulin on the basis of its molecular size and susceptibility to trimming by carboxypeptidase B (CpB). However, the findings of an active carboxypeptidase E (CpE) enzyme and the normal amidated forms of gastrin and cholecystokinin octapeptide (CCK-8) in Psammomys tissues were inconsistent with CpE-related aberrant processing of insulin. Moreover, amino acid sequencing of the delayed peak of Psammomys insulin revealed fully processed insulin with amino acid sequence as predicted by the cDNA. The unique presence of a B-30 phenylalanine residue, resulting in an increased hydrophobicity of the insulin molecule, probably underlies the marked delay in elution time on HPLC. The unusual structure of Psammomys insulin does not appear to contribute to the proinsulinemia observed in diabetic Psammomys since the HPLC-purified molecule did not inhibit PCI and PC2 convertase activities in an in vitro assay.

Chronic exposure to high glucose impairs multiple β-cell functions in cultured human pancreatic islets

Deficient insulin secretion and relative hyperproinsulinemia are characteristic of non–insulin-dependent diabetes. Culture of human islets at high glucose concentrations generates β-cell dysfunctions that mimic those observed in diabetic patients, with depleted islet insulin stores. The increased proinsulin ratio probably results from accelerated discharge of insulin granules, rather than diminished expression of convertase. Insulin mRNA levels are decreased, and the rate of transcription of the human insulin gene is suppressed. In human islets exposed to high glucose for 4 to 9 days, the DNA binding of PDX-1, the key transcriptional factor of the insulin gene, is reduced, as is its mRNA level. These effects are reversed by normalizing the culture glucose concentration. It is proposed that β-cell “glucotoxicity” results from the inability of proinsulin biosynthesis to keep pace with chronic insulin hypersecretion. At least in human β-cells, downregulation of PDX-1 expression by hyperglycemia is the main cause for depletion of the insulin stores.