Retinal neovascularization is the leading cause of visual impairment in diabetic retinopathy, retinopathy of prematurity, and age-related macular degeneration. The extracellular matrix breakdown by metalloproteinase leads to vascular complications in various proliferative retinopathies. A disintegrin and metalloproteinase domain-containing protein 10 (ADAM10) is involved in physiological angiogenesis. However, limited information exists regarding the role of ADAM10 in proliferative retinopathies. In this study, the levels of active ADAM10 were significantly up-regulated in the ischemic retina, and down-regulation or inactivation of ADAM10 significantly inhibited the proliferation, sprouting, migration, and tube formation of human retinal microvascular endothelial cell. Furthermore, the endothelial cell (EC)-specific deletion of ADAM10 (ADAM10(iΔEC)) significantly attenuated vascular leakage, edema, endothelial cell sprouting, and retinal neovascularization in ischemic retinas of mice exposed to oxygen-induced retinopathy. In experiments investigating the mechanisms through which ADAM10 regulated pathologic angiogenesis, ADAM10 regulated ephrin B2 (EfnB2) expression in endothelial cells. Down-regulation of EfnB2 expression influenced human retinal microvascular endothelial cell proliferation, migration, sprouting, and tube formation. In addition, a significant up-regulation of EfnB2 expression in the ischemic retina was detected. EC-specific depletion of ADAM10 significantly reduced EfnB2 expression, suggesting its involvement in ADAM10-regulated retinal neovascularization. The findings demonstrate how EC-specific ADAM10 regulates pathologic retinal neovascularization in the ischemic retina, indicating its significance as a potential therapeutic target for proliferative retinopathies.