Pre-clinical and observational studies in ovariectomized (OVX) animals and pre- and post-menopausal women respectively, have suggested the cardio- protective effects of estrogen replacement therapy. However, randomized clinical trials have not confirmed estrogen-mediated cardio-protection. Although uncertainties about the duration and optimal type of estrogen replacement regimen might explain the disparity, other factors that may mask the protective effects of estradiol (E2) on cardiovascular outcome, need scrutiny. Increased ethanol consumption may be one such factor. We examined the effect of E2 supplementation on ethanol consumption in OVX mice and the effect of ethanol consumption on E2-mediated vascular repair, in vivo. OVX mice implanted with E2 pellets consumed significantly more ethanol compared to those receiving placebo pellets. E2-induced increase in ethanol consumption was not affected by the absence of either estrogen receptor (ER)- or ER. Re-endothelialization following carotid artery denudation was repressed and neo-vascularization in ischemic hind limbs was blunted in mice consuming ethanol, despite E2 supplementation. In vitro, ethanol dose-dependently attenuated E2-induced endothelial cell (EC) proliferation and tube formation activity and enhanced EC apoptosis, suggesting that ethanol blocks E2-induced EC survival and function. Taken together our data suggest that increased ethanol consumption following E2 supplementation blunts the beneficial effects of E2 on EC function and that novel approaches to estrogen replacement for cardio-protection may benefit from the control of alcohol consumption.