Stimulation of endothelin-1 gene expression by insulin via phosphoinositide-3 kinase-glycogen synthase kinase-3beta signaling in endothelial cells.

Insulin stimulates secretion of the potent vasoactive and mitogenic peptide endothelin-1 (ET-1) from endothelial cells. We sought to investigate whether phosphoinositide-3 kinase (PI3K)-dependent inactivation of glycogen synthase kinase-3beta (GSK3beta) by insulin leads to elevation of ET-1 gene expression in endothelial cells. Inhibition of GSK3beta activity by LiCl or siRNA technique mimicked insulin action to stimulate ET-1 gene expression. Luciferase reporter assay showed insulin stimulated-elevation of ET-1 promoter activity can be abolished by the PI3K inhibitor Wortmannin, but not by the mitogen activated protein kinase (MAPK) inhibitor PD-98059. To further investigate whether the transcription factor vascular endothelial zinc finger-1 (Vezf1) is involved in ET-1 regulation, site-mutated reporter plasmid was used in luciferase reporter assay. A 2-bp mutation in Vezf1 binding element abolished insulin-stimulated elevation of ET-1 promoter activity. Furthermore, siRNA inhibition of Vezf1 led to decline in the levels of ET-1 mRNA and ET-1 peptides. These observations indicate that PI3K-dependent inactivation of GSK3beta by insulin leads to upregulation of ET-1 gene expression and Vezf1 may be a target for ET-1 regulation by insulin. PI3K-GSK3beta signaling may be responsible for insulin stimulation of ET-1 production associated with insulin resistance and hyperinsulinemia.