Dose-dependent activation of antiapoptotic and proapoptotic pathways by ethanol treatment in human vascular endothelial cells - Differential involvement of adenosine

Moderate but not heavy drinking has been found to have a protective effect against cardiovascular morbidity. We investigated the effects of ethanol (EtOH) treatment on the cell survival-promoting phosphatidylinositol 3-kinase (PI3K)/Akt pathway in cultured human umbilical vein endothelial cells (HUVEC). Exposure of cells to 2-20 mm EtOH resulted in rapid (< 10 min) induction of Akt phosphorylation that could be prevented by pertussis toxin or the PI3K inhibitors wortmannin and LY294002. Among the downstream effectors of PI3K/Akt, p70S6 kinase, glycogen synthase kinase 3alpha/p, and IKB-alpha were phosphorylated, the latter resulting in 3-fold activation of NF-kappaB. EtOH also activated p44/42 mitogenactivated protein kinase in a PI3K-dependent manner. Low concentrations of EtOH increased endothelial nitric-oxide synthase activity, which could be blocked transfection of HUVEC with dominant-negative Akt, implicating the PI3K/Akt pathway in this effect. The adenosine Al receptor antagonist 1,3-dipopyleyclopentylxanthine prevented the phosphorylation of Akt observed in the presence of EtOH, adenosine, or the Al agonist N-6-cyclopentyladenosine. Incubation of HUVEC with 50100 mm EtOH resulted in mitochondrial permeability transition and caspase-3 activation followed by apoptosis, as documented by DNA fragmentation and TUNEL assays. EtOH-induced apoptosis was unaffected by DPCPX and was potentiated by wortmannin or LY294002. We conclude that treatment with low concentrations of EtOH activates the cell survival promoting PI3K/Akt pathway in endothelial cells by an adenosine receptor-dependent mechanism and activation of the proapoptotic caspase pathway by higher concentrations of EtOH via an adenosine-independent mechanism can mask or counteract such effects.