Ebola virus (EBOV) causes a highly lethal hemorrhagic fever syndrome in humans and has been associated with mortality rates of up to 91% in Zaire, the most lethal strain. Though the viral envelope glycoprotein (GP) mediates widespread inflammation and cellular damage, these changes have mainly focused on alterations at the protein level, the role of microRNAs (miRNAs) in the molecular pathogenesis underlying this lethal disease is not fully understood. Here, we report that the mi-RNAs hsa-miR-1246, hsa-miR-320a and hsa-miR-196b-5p were induced in human umbilical vein endothelial cells (HUVECs) following expression of EBOV GP. Among the proteins encoded by predicted targets of these miRNAs, the adhesion-related molecules tissue factor pathway inhibitor (TFPI), dystroglycan1 (DAG1) and the caspase 8 and FADD-like apoptosis regulator (CFLAR) were significantly downregulated in EBOV GP-expressing HUVECs. Moreover, inhibition of hsa-miR-1246, hsa-miR-320a and hsa-miR-196b-5p, or overexpression of TFPI, DAG1 and CFLAR rescued the cell viability that was induced by EBOV GP. Our results provide a novel molecular basis for EBOV pathogenesis and may contribute to the development of strategies to protect against future EBOV pandemics.