Recovery of hyperoxia‐induced DDAH1 suppression with FXR agonist GW4064 in human pulmonary microvascular endothelial cells

Objective Pulmonary hypertension (PH) predicts mortality in patients with bronchopulmonary dysplasia (BPD), and chronic hyperoxia exposure contributes to BPD‐PH disease progression. BPD‐PH is characterized by aberrant pulmonary vascular wall proliferation/remodeling, which occludes the microvascular lumen, increasing pulmonary vascular resistance. Endogenous nitric oxide (NO) production by pulmonary vascular endothelial cells promotes apoptosis of pulmonary vascular endothelial and smooth muscle cells. In vitro studies of human pulmonary microvascular endothelial cells (hPMVEC) have shown that dimethylarginine dimethylaminohydrolase‐1 (DDAH1) promotes NO‐mediated apoptosis. DDAH1 metabolizes the majority of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of endothelial NO synthase. Therefore, our hypothesis is that pharmacologically enhancing DDAH1 expression in hPMVEC will restore endogenous endothelial NO production. Methods hPMVEC were grown at 37 ̊C to ~70% confluence. Farnesoid X receptor (FXR) agonist, GW4064 (1.25 µM) was used to bind FXR element (FXRE) of the DDAH1 promoter to enhance transcription. hPMVEC were pre‐treated with FXR agonist for 24 hours and then exposed to 85%O 2 or 21%O 2 for an additional 24 hours. DDAH1 protein expression was analyzed by Western blot. Groups were compared by 1‐way ANOVA. Results In hPMVEC, hyperoxia exposure resulted in lower DDAH1 protein levels compared to hPMVEC grown in normoxia (N=6 in each group, p=0.003). Treatment with GW4064 prior to exposing to hyperoxia resulted in 4.3‐fold greater DDAH1 protein levels than in vehicle/85%O 2 hPMVEC (N=6, p=0.044), and restored DDAH1 protein levels to 72% of that seen in the vehicle/21%O 2 exposed hPMVEC. Conclusions Chronic hyperoxia suppresses DDAH1 levels, and FXR agonist promotes DDAH1 recovery in hPMVEC. We speculate that promoting endogenous DDAH1 expression may prevent aberrant proliferation in the pulmonary vascular wall and thereby attenuate or prevent the vascular remodeling characteristic of pulmonary hypertension in BPD.