Objective To investigate the effects of hypoxia response element (HRE)-mediated Bcl-2 gene silencing on hypoxia-induced anti-apoptotic pulmonary vascular endothelial cells. Methods The siRNA sequence for Bcl-2 was designed. Bcl-2-shRNA expression vectors carrying HRE were constructed and packaged by lentivirus. Pulmonary vascular endothelial cells infected with recombinant lentivirus were cultured under the normoxic condition (21% O₂) and hypoxic condition (5% O₂), respectively. The infection efficiency was observed by the fluorescent marker after 96 h. The expression of Bcl-2 protein was detected by the Western blotting and the cell apoptosis was detected by the flow cytometry. Results The silencing of Bcl-2 gene significantly down-regulated the mRNA and protein expressions of Bcl-2. The activity of HRE promoter enhanced only in hypoxic environment. The transfection efficiency was about 90% after pulmonary microvascular endothelial cells were infected with recombinant lentivirus for 96 h and the protein level of Bcl-2 decreased and cell apoptosis increased. The apoptosis rat of Lv-HRE-Bcl-2-shRNA group significantly increased because of the regulation of HRE in hypoxic environment. Conclusion HRE acts as an oxygen-sensitive regulatory switch under the hypoxic condition, which can further enhance the effects of Bcl-2-shRNA gene and induce the apoptosis of pulmonary vascular endothelial cells, so as to provide a potential therapeutic target for vascular remodeling of pulmonary arterial hypertension.