REGULATION OF FATTY ACID TRANSPORT ACROSS THE MITOCHONDRIAL MEMBRANES IN HUMAN AND RODENT SKELETAL MUSCLE

Abstract

REGULATION OF FATTY ACID TRANSPORT ACROSS THE MITOCHONDRIAL MEMBRANES IN HUMAN AND RODENT SKELETAL MUSCLE Veronic S. Bezaire Advisor: University of Guelph, 2005 Lawrence L. Spriet This thesis is an investigation of the role and regulation of carnitine palmitoyltransferase I (CPTI), fatty acid translocase (FAT/CD36) and uncoupling protein 3 (UCP3) and their impact on fatty acid (FA) transport across the mitochondrial membranes and metabolism in human and rodent skeletal muscle. The regulation of CPTI activity was examined in intermyofibrillar (IMF) and subsarcolemmal (SS) mitochondria isolated from human and rat skeletal muscle. Maximal CPTI activity and sensitivity to inhibitor malonyl-CoA (M-CoA) was similar between IMF and SS mitochondria from both species. Moderate intensity concentrations of exercise-related metabolites calcium, AMP, ADP and inorganic phosphate failed to override M-CoA inhibition in IMF and SS mitochondria. This data suggests that the regulation of FA transport across the mitochondria during moderate intensity exercise remains unclear. Following the recent identification of FAT/CD36 in rat skeletal mitochondria, the presence and role of FAT/CD36 in human skeletal muscle mitochondria was investigated. In vitro treatment of mitochondria with specific FAT/CD36 inhibitor sulfo-N-succimidyloleate (SSO) decreased palmitate oxidation by 95% (P < 0.01) without affecting mitochondrial octanoate oxidation demonstrating the specificity of SSO towards FAT/CD36. Furthermore, treatment of mitochondria with SSO had no effect on maximal and submaximal CPTI activity but did inhibit palmitoylcarnitine oxidation by 92% (P < 0.001). Therefore, it was hypothesized that FAT/CD36 is required for palmitate oxidation and functions downstream of CPTI, possibly in the transfer of palmitoylcarnitine from CPTI to CPTII in the intermembrane space of human skeletal muscle mitochondria. Given the strong link between FA levels and UCP3 expression, the effects of a physiological overexpression of uncoupling protein 3 (UCP3) and ablation of UCP3 on FA transport and oxidation capacity in mouse skeletal muscle were examined. UCP3 overexpression improved serum lipid profile and increased capacity for LCFA uptake (P < 0.05) and oxidation (P < 0.05) resulting in decreased intramuscular triglyceride stores (P < 0.05). High energy phosphagens, coenzyme A and carnitine levels were increased (P < 0.05) with UCP3 overexpression but unchanged with UCP3 ablation. Despite the lack of change with UCP3 ablation, this study supports an important role for UCP3 in FA metabolism.