Estrogen induces the Akt-dependent activation of endothelial nitric-oxide synthase in vascular endothelial cells

Although estrogen is known to activate endothelial nitric oxide synthase (eNOS) in the vascular endothelium, the molecular mechanism responsible for this effect remains to be elucidated. In studies of both human umbilical vein endothelial cells (HUVECs) and simian virus 40-transformed rat lung vascular endothelial cells (TRLECs), 17 beta -estradiol (E2), but not 17 alpha -E2, caused acute activation of eNOS that was unaffected by actino-mycin D and was specifically blocked by the pure estrogen receptor antagonist ICI-182,780, Treatment of both TRLECs and HUVECs with 17 beta -E2 stimulated the activation of Akt, and the PI3K inhibitor wortmannin blocked the 17 beta -E2-induced activation of Akt, 17 beta -E2-induced Akt activation was also inhibited by ICI-182,780, but not by actinomycin D, Either treatment with wortmannin or exogenous expression of a dominant negative Akt in TRLECs decreased the 17 beta -E2-induced eNOS activation. Moreover, 17 beta -E2-induced Akt activation actually enhances the phosphorylation of eNOS, 17 beta -E2-induced Akt activation was dependent on both extracellular and intracellular Ca2+. We further examined the 17 beta -E2-induced Akt activity in Chinese hamster ovary (CHO) cells transiently transfected with cDNAs for estrogen receptor cw (ER alpha) or estrogen receptor beta (ER beta), 17 beta -E2 stimulated the activation of Akt in CHO cells expressing ERa but not in CHO cells expressing ERP, Our findings suggest that 17P-E2 induced eNOS activation through an Akt-dependent mechanism, which is mediated by ER alpha via a nongenomic mechanism.