Targeting Endothelial ENO1 (Alpha-Enolase) -PI3K-Akt-mTOR Axis Alleviates Hypoxic Pulmonary Hypertension

It has been shown that glycolytic protein ENO1 (alpha-enolase) contributes to the pathogenesis of pulmonary hypertension through acting smooth muscle cells; however, the roles of ENO1-caused endothelial and mitochondrial dysfunctions in Group 3 pulmonary hypertension remain unexplored.

PCR array and RNA sequencing were used to screen and decipher the differential gene expression by hypoxia-treated human pulmonary artery endothelial cells. Techniques of small-interfering RNA, specific inhibitor and plasmids carrying gene of ENO1, interventions with specific inhibitor and AAV-ENO1 delivery were employed to explore the role of ENO1 in hypoxic pulmonary hypertension in vitro and in vivo, respectively. Assays for cell proliferation, angiogenesis, and adhesion were employed to analyze cell behaviors, while seahorse analysis was used to measure mitochondrial function of human pulmonary artery endothelial cells.

PCR array data showed that ENO1 expression increased in human pulmonary artery endothelial cells exposed to hypoxia, as well as in lung tissues from patients with chronic obstructive lung disease-associated pulmonary hypertension and murine model of hypoxic pulmonary hypertension. Inhibition of ENO1 restored the hypoxia-induced endothelial dysfunction, including excessive proliferation, angiogenesis, and adhesion, while overexpression of ENO1 promotes these disorders of human pulmonary artery endothelial cells. RNA-seq showed that ENO1 targets mitochondrion-related genes and PI3K-Akt signaling pathway, which were validated in vitro and in vivo. Mice treated with ENO1 inhibitor exhibited ameliorated pulmonary hypertension and improved right ventricular failure induced by hypoxia. A reversal effect was observed in mice exposed to hypoxia and inhaled adeno-associated virus overexpressing ENO1.

These results indicate that hypoxic pulmonary hypertension is associated with an increased level of ENO1 and that targeting ENO1 might reduce experimental hypoxic pulmonary hypertension by improving endothelial and mitochondrial dysfunction via PI3K-Akt-mTOR signaling pathway.