Preeclampsia is a hypertensive complication in pregnancy, closely related to endothelial dysfunction. Endothelial progenitor cells (EPCs) have the capacity for endothelial repair. Both Ephrin-B2 and Dll4/Notch pathway play critical roles in various steps of angiogenesis. In addition, there is an up-regulation of ephrin-B2 expression consequent to Dll4/Notch activation in endothelial cells (ECs). However, the roles of ephrin-B2 and Dll4/Notch signaling on EPCs, as well as the relationship between them, have not been completely characterized.We analyzed expression of ephrin-B2 in the EPCs and placenta from preeclampsia and normal pregnancy. Then up-regulation and down-regulation strategies were employed to detect the effects of ephrin-B2 on EPC proliferation, differentiation, migration and HUVEC-tube formation. The effects of Dll4/Notch signaling on EPCs' functions were determined by repeating the assays in the presence of Dll4 or DAPT as agonists or antagonists of Notch signaling, respectively.Ephrin-B2 expression increased notably in preeclampsia EPCs and placenta, compared with controls. Up-regulation of ephrin-B2 impaired EPCs' proliferation, differentiation, migration and HUVEC-tube formation capabilities. In contrast, down-regulation of ephrin-B2 in EPCs, resulted in the opposite effects. In addition, activation of Dll4/Notch signaling led to increased expression of ephrin-B2 and subsequent inhibition of EPCs activity.Ephrin-B2, a downstream of Dll4/Notch signaling pathway, might be act as an inhibitor of EPC-mediated vasculogenesis in vitro, as well as a potential target in the effort to promote angiogenesis of patients with preeclampsia.