Upregulation of endothelial hypoxia‐inducible factor‐2 alpha in animals with pulmonary hypertension
Authors: Powers Ryan , Babicheva Aleksandra , Jain Pritesh , Chen Jiyuan , Zhao Tengteng , Makino Ayako , Yuan Jason X‐J 
Published: May 2022

Introduction

Idiopathic pulmonary arterial hypertension (IPAH) is a life‐threatening disease with no cure. Pulmonary vascular remodeling due to increased pulmonary artery smooth muscle (PASMC) and lung vascular endothelial cell (LVEC) proliferation is a major cause for the elevated pulmonary vascular resistance (PVR) and pulmonary arterial pressure (PAP) in patients with IPAH. We have previously demonstrated that endothelial hypoxia‐inducible factor‐2α (HIF‐2α) contributes to the development of pulmonary vascular remodeling in LVEC isolated from IPAH patients. Moreover, HIF‐2α is involved in the initiation of endothelial‐to‐mesenchymal transition (EndMT) by upregulating Snai1 and Snai2 (EndMT drivers). However, the exact role of HIF‐2α in the development and progression of pulmonary hypertension (PH) remains elusive. We aimed to investigate whether endothelial HIF‐2α is involved in pulmonary vascular remodeling in animals with hypoxia/sugen‐induced PH (SuHx‐PH) and monocrotaline‐induced PH (MCT‐PH).

Methods

Hypoxia/sugen‐PH (SuHx‐PH) model in mice was induced by exposing animals to the combination of normobaric hypoxia (10% O2) and sugen 5416 (20 mg/kg, i.p., weekly) for 6 weeks. Monocrotaline‐MCT‐PH model in rats was induced by single administration of MCT (60 mg/kg, s.c.) followed by experimental measurements after 28 days. Body weight (BW) was collected from all animals weekly. Hemodynamic parameters including right ventricular systolic pressure (RVSP), mean pulmonary arterial pressure (mPAP), the Fulton index (FI), right ventricular (RV) contractility, heart rate (HR) were measured to establish PH. The pulmonary artery (PA) wall thickness was estimated by H&E staining. The protein expression of Snai1 and HIF‐2α was determined by Western blot and immunocytochemistry. Normal human LVEC was treated with TGF‐β1 for 7 days to induce EndMT. LVEC isolated with patients with IPAH was treated with HIF‐2α inhibitor Compound 2 for 8‐48 hrs.

Results

Exposure of mice to the SuHx (6 weeks) or rats to MCT (4 weeks) resulted in the significant increase of RVSP, mPAP, RV contractility, and FI compared to control animals. We did not detect any changes in HR between corresponded groups. BW was decreased by 7% after the first week of the SuHx treatment in mice. By the end of 6 weeks BW was significantly lower by 12% in SuHx mice compared with normoxic animals. BW in MCT‐treated rats was 5% lower after the first week and at the end of experiment compared to vehicle‐treated animals. Pulmonary vascular remodeling estimated by PA wall thickness was enhanced in both SuHx‐PH mice and MCT‐PH rats. The protein expression of HIF‐2α and Snai1 was higher in lung tissues isolated from SuHx‐PH mice and MCT‐PH rats compared to controls. Moreover, HIF‐2α and Snai1 was activated in normal human LVEC treated with TGF‐β1 (10ng/ml) for 7 days. Additionally, HIF‐2α inhibitor, Compound 2 (30µM) was able to downregulate Snai1 protein expression in LVEC isolated from IPAH patients after 16 hours.

Conclusions

Our results indicate that HIF‐2α is involved in pulmonary vascular remodeling in animals with PH. Our studies support the premise that targeting HIF‐2α would be a novel therapeutic approach for the treatment of IPAH and other forms of PH.