PROBLEM: Preeclampsia is characterized by endothelial activation and excessive inflammation, of which interferon (IFN)-gamma is a potent inducer. Eph receptor B4 (EPHB4) also involved in endothelial activation in inflammation. Their role and relationship in preeclampsia remain unclear. METHOD OF STUDY: Intercellular adhesion molecular (ICAM)-1 was employed as the hallmark of endothelial activation. The serum levels of IFN-gamma and the expression of EPHB4 and ICAM-1 were assessed by ELISA, qRT-PCR and WB, respectively. Primary human umbilical vein endothelial cells (HUVECs) were treated with IFN-gamma of different concentration or for different times to determine the effect of IFN-gamma on EPHB4 and ICAM-1 expression. Overexpression and shRNA constructs, chromatin immunoprecipitation (ChIP) and luciferase assays were conducted to clarify the regulation mechanism of IFN-gamma/STAT1 on EPHB4 resulting in HUVECs activation. Endothelial-trophoblast co-culture model was used to illustrate the role of EPHB4 in the process of activated endothelial cells resisting trophoblast invasion. RESULTS: IFN-gamma, EPHB4 and ICAM-1 expression were elevated in preeclampsia. IFN-gamma induced HUVECs activation through EPHB4 expression. ChIP and luciferase assays revealed that IFN-gamma promoted EPHB4 transcription by STAT-1 binding to EPHB4 promoter. EPHB4 probably involved in resisting trophoblasts displacement by IFN-gamma-activated HUVECs. CONCLUSION: This study uncovered the character of EPHB4-regulating endothelial activation in the pathogenesis of preeclampsia.