Hepatic Insulin Action: Role of the Pentose Cycle

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Hepatic insulin action: Role of the pentose cycle. Insulin stimulation of PI3-Kinase results in an increase in phosphatidylinositol-3,4,5 tri-phosphate (PIP3) formation, which has direct inhibitory effects on glucose-6-phosphatase (G6Pase) activity, as well as initiating the activation of a signaling effector chain, via Akt activation and GSK-3 inhibition, that stimulates glycolysis, glycogen storage, and pentose phosphate pathway (PPP) flux, and inhibits gluconeogenesis. The control of substrate flux between the hexose phosphate pool and the triose phosphate pool has been recognized as the key mechanism for insulin's regulation of HGP. The hexose phosphate pool and the triose phosphate pool are in equilibrium through the oxidative and non-oxidative branches of the pentose phosphate pathway (PPP). Our 13C mass isotopomer distribution analysis (MIDA) flux experiments indicate that insulin's stimulation of PI3-K coordinates flux between the glycolytic/gluconeogenic pathways and the pentose phosphate pathway (PPP) via PI3-K's inhibition of G6Pase. Insulin can upregulate flux through the non-oxidative limb of the PPP, or the oxidative limb via glucose-6-phosphate dehydrogenase (G6PDH), raising xylulose-5-phosphate (X5P) levels, which can stimulate glycolysis. G6PDH is also the principal source of NADPH, which keeps the cytoplasm in a reduced state. Studies suggest that when the liver cytoplasm is more oxidized, HGP is increased. Hypothesis: Insulin-stimulated PI3-K stimulates PPP flux, and the stimulation of PPP flux enhances the function of insulin action. Specific aims for hypothesis testing: To quantitate the roles, and actions on the flux between the hexose phosphate and the triose phosphate pools, of PI3-K and its signaling effectors/modulators Akt, MMAC, a phosphatidylinositol 3- phosphatase, and GSK-3, via adenoviral mediated overexpression, and selected inhibitors of PI3-K signaling. Specific 13C labeled substrates will be used to determine flux passing to and from the G-6-P pool using mass isotopomer analysis in primary rat hepatocyte culture. These studies may identify areas where defects in hepatic insulin action occur, and where hepatic-based gene therapies can be directed.