Our preliminary studies suggest the hypothesis that an arachidonate 12-lipoxygenase product (e.g. 12-HPETE) participates in insulin secretion since glucose stimulates synthesis of 12-lipoxygenase products by isolated pancreatic islets and 12-lipoxygenase inhibitors suppress glucose-induced insulin secretion. This hypothesis will be further evaluated by examining 1) the time course of rat islet 12-HETE and prostaglandin biosynthesis compared to the biphasic secretion of insulin after glucose stimulation; 2) the insulin secretagogue properties and islet biosynthesis of recently described metabolites of 12-HPETE; 3) the influence islet insulin secretion in response to exogenous 12-lipoxygenase products); 4) the influence of lipoxygenase inhibitors on glucose-induced insulin secretion in isolated human pancreatic islets and 5) the biosynthesis of 12-HETE by human islets in response to glucose. Exogenous addition of the 12-lipoxygenase product 12-HETE only partially reverses the suppression of glucose-induced insulin secretion from isolated islets by lipoxygenase inhibitors. I will examine the possibilities that stimulation of insulin secretion can be more readily demonstrated with: 1) enantiopure rather than racemic 12-HETE and 12-HPETE and 2) carboxylic acid esters of 12-HETE and 12-HPETE rather than the free carboxylate anions (which may not readily enter cells). To explore a possible mechanism for effects of 12-lipoxygenase products, the influence of these compounds on calcium flux from digitonin- permeabilized islets will also be examined. Arachidonate for 12-HPETE synthesis may be derived in part from phosphatidyl inositol-4,5-biphosphate with attendant generation of diacyclglycerol (DG) and inositol-1,4, 5-tris- phosphate (IP3). To evaluate the hypothesis that the coordinated release and degradation of free arachidonate, its metabolites, and of DG and IP3 regulate the phasic nature of glucose-induced insulin secretion, mass spectrometric methods will be developed for quantitation of AA, DG, and IP3 which are similar to those already developed for measurement of 12-HETE. These methods will then be applied to study the time course of 12-HETE, AA, DG, and IP3 appearance in relationship to insulin secretion after glucose stimulation.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Modified Research Career Development Award (K04)
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Metabolism Study Section (MET)
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Washington University
Schools of Medicine
Saint Louis
United States
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Ramanadham, S; Gross, R; Turk, J (1992) Arachidonic acid induces an increase in the cytosolic calcium concentration in single pancreatic islet beta cells. Biochem Biophys Res Commun 184:647-53
Turk, J; Mueller, M; Bohrer, A et al. (1992) Arachidonic acid metabolism in isolated pancreatic islets. VI. Carbohydrate insulin secretagogues must be metabolized to induce eicosanoid release. Biochim Biophys Acta 1125:280-91
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Holtzman, M J; Ferdman, B; Bohrer, A et al. (1991) Synthesis of the 1-O-hexadecyl molecular species of platelet-activating factor by airway epithelial and vascular endothelial cells. Biochem Biophys Res Commun 177:357-64
Mangino, M J; Anderson, C B; Murphy, M K et al. (1991) Renal allograft platelet activating factor synthesis during acute cellular rejection. J Lipid Mediat 4:69-81
Wolf, B A; Pasquale, S M; Turk, J (1991) Free fatty acid accumulation in secretagogue-stimulated pancreatic islets and effects of arachidonate on depolarization-induced insulin secretion. Biochemistry 30:6372-9
Easom, R A; Landt, M; Colca, J R et al. (1990) Effects of insulin secretagogues on protein kinase C-catalyzed phosphorylation of an endogenous substrate in isolated pancreatic islets. J Biol Chem 265:14938-46

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