Mitochondrial functional state in clonal pancreatic beta-cells exposed to free fatty acids.

Excessive free fatty acid (FFA) exposure represents a potentially important diabetogenic condition that can impair insulin secretion from pancreatic beta-cells. Because mitochondrial oxidative phosphorylation is a main link between glucose metabolism and insulin secretion, in the present work we investigated the effects of the FFA oleate (OE) on mitochondrial function in the clonal pancreatic beta-cell line, MIN6. Both the long term (72 h) and short term (immediately after application) impact of OE exposure on beta-cells was investigated. After 72 h of exposure to OE (0.4 mm, 0.5% bovine serum albumin) cells were washed and permeabilized, and mitochondrial function (respiration, phosphorylation, membrane potential formation, production of reactive oxygen species) was measured in the absence or presence of OE. MIN6 cells exposed to OE for 72 h showed impaired glucose-stimulated insulin secretion and decreased cellular ATP. Mitochondria in OE-exposed cells retained normal functional characteristics in FFA-free medium; however, they were significantly more sensitive to the acute uncoupling effect of OE treatment. The mitochondria of OE-exposed cells displayed increased depolarization caused by acute OE treatment, which is attributable to the elevation in the FFA-transporting function of uncoupling protein 2 and the dicarboxylate carrier. These cells also had an increased production of reactive oxygen species in complex I of the mitochondrial respiratory chain that could be activated by FFA. A high level of reduction of respiratory complex I augmented acute FFA-induced uncoupling in a way compatible with activation of mitochondrial uncoupling protein by intramitochondrial superoxide. A stronger augmentation was observed in OE-exposed cells. Together, these events may underlie FFA-induced depression of the ATP/ADP ratio in beta-cells, which accounts for the defective glucose-stimulated insulin secretion associated with lipotoxicity.