Regulation of GLUT4 activity in normal and insulin resistant states.
Published: 2005
Language: eng
Number of pages: 200
Description: Insulin increases GLUT4 translocation and glucose uptake in adipocytes and muscles. In addition, insulin may increase the activity of GLUT4, possibly via p38MAPK. The objective of this thesis is to dissect the insulin signalling pathway to identify the molecule(s) that regulates GLUT4 activity.Next, we determine whether the insulin signals regulating GLUT4 translocation vis-a-vis glucose uptake can be segregated in insulin resistance. We incubated L6 myotubes in high glucose and insulin for 24 h, and examined the response of the IRS-PI3K-Akt signalling pathway to an acute insulin challenge. We found a >50% reduction in insulin-stimulated IRS-1 tyrosine phosphorylation, PI3-kinase activity, and Akt phosphorylation, accompanied by blunted GLUT4 translocation. Surprisingly, the insulin-stimulated glucose uptake in these cells was comparable to that of the controls, suggesting increased GLUT4 activity. Interestingly, both the protein expression and activity of p38MAPK were enhanced, and treatment with a p38MAPK inhibitor (pyridinylimidazole) reduced glucose uptake to a level that matches the amount of GLUT4 present on cell surface.Since both our previous results and the insulin resistant model implicated p38MAPK in regulation of GLUT4 activity, we used siRNA-mediated gene silencing and expression of dominant negative mutants to determine the role of p38MAPK in glucose uptake. In contrast to previous results using the p38MAPK inhibitor (pyridinylimidazole), reduction of p38MAPK expression and activity by >70% had no effect on insulin-stimulated glucose uptake. These results suggested that GLUT4 activity could be upregulated via a pyridinylimidazole-dependent but p38MAPK independent pathway. The putative pyridinylimidazole target may provide significant insight into factors that optimize insulin-stimulated glucose uptake and maintenance of glucose homeostasis.First, we examined the IR/IRSI-2/PI3-K/Akt pathway to determine whether IRS-1 and IRS-2 have differential contribution to insulin-stimulated glucose uptake vs. GLUT4 translocation. We found that both IRS-1 and IRS-2 regulate insulin-stimulated activation of Akt and p38MAPK. However, a 70% reduction in IRS-1 led to &sim;50% reduction in insulin-stimulated glucose uptake and GLUT4 translocation as well as actin remodelling, while a 75% reduction in IRS-2 had no effect on these biological outcomes. Therefore, IRS-1 but not IRS-2 is the main adaptor molecule that regulates glucose uptake and GLUT4 translocation in muscle cells.
Notes: Thesis (Ph.D.)--University of Toronto, 2005.

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Source: Dissertation Abstracts International, Volume: 66-06, Section: B, page: 2957.