The long-term objective of this research is to develop an understanding of the role of DHEAS in glucose regulation and its potential therapeutic role in diabetes mellitus. Many authors have demonstrated a beneficial effect of DHEA administration in rodent diabetes of various etiologies, however, the mechanism of this effect is unclear. The preliminary studies show that DHEAS strongly enhances the insulinotropic effect of glucose, implicating an effect of DHEAS at the pancreatic beta-cell as a basis for its anti-diabetic effects. Furthermore, the investigators show that DHEAS alters beta-cell expression of specific genes, acyl CoA synthetase-2 (ACS-2) and acyl CoA oxidase (ACO), involved in the intracellular metabolism of fatty acids. Beta cell lipid stores and free fatty acid (FFA) concentrations strongly modulate glucose-stimulated insulin secretion and cytoplasmic long-chain fatty acyl CoA (LC CoA) has been postulated to be an important signal for insulin secretion. It is proposed to test the hypothesis that DHEAS enhances glucose-stimulated insulin secretion by initially stimulating ACS-2 activity, resulting in increased cytoplasmic LC-CoA levels. In addition to enhancing glucose-stimulated insulin secretion, the investigators postulate that the elevated LC-CoA, or a product of LC-CoA, activates beta-cell peroxisome proliferator-activated receptors (PPARs) and consequently increases the activity of the lipid oxidation enzyme ACO. The following Specific Aims will be addressed: (1) Determine whether changing ACS-2 expression alters glucose-stimulated insulin section, ACS activity, cytoplasmic LC-CoA levels, PPARalpha activation and ACO expression, in a similar manner to incubation with DHEAS. Experiments will include assessing: i) the effect of DHEAS on ACS-2 mRNA expression, and ii) the effect of ACS-2 over-expression on glucose-stimulated insulin secretion. (2) Determine whether modulation of LC-CoA levels alters glucose-stimulated insulin secretion, PPARalpha activation or ACO expression. Modulation of LC-CoA levels will be achieved by beta-cell incubation with FFA, etomoxir (an inhibitor of carnitine palmitoyl transferase-1) and Triacsin C (an inhibitor of ACS). (3) Determine whether PPARalpha knockout alters the effect of DHEAS on glucose-stimulated insulin secretion, ACS activity and ACO mRNA expression. (4) Assess the effects of DHEAS on ACS activity, glucose-stimulated insulin secretion and ACO mRNA expression in human pancreatic islets.
