Abstract 17443: Wwox is a Critical Regulator of Pulmonary Artery Smooth Muscle Cell Homeostasis

Abstract

Introduction: Pulmonary artery smooth muscle (PASMC) proliferation and apoptosis resistance are characteristic of pulmonary arterial hypertension (PAH). Expression loss of the tumor suppressor WW-domain containing oxidoreductase (WWOX) promotes cell survival and metabolic reprograming in cancer. We hypothesized that loss of WWOX expression in PASMC promotes pulmonary vascular remodeling by similar mechanisms. Methods and Results: mRNA and protein levels of WWOX are decreased in PASMC from patients with PAH, and in rat models of pulmonary hypertension (Sugen/ hypoxia and monocrotaline). PASMC-specific conditional Wwox knockout mice (WWOX-SMC KO) were treated with tamoxifen or vehicle for 5 days and exposed to 10% hypoxia for 4 weeks. WWOX-SMC KO animals developed worse hypoxia induced PH with increased right ventricular systolic pressure, right ventricular hypertrophy and pulmonary vascular remodeling, compared to control animals. WWOX loss via siRNA decreased mRNA and protein levels of BMPRII, upregulated HIF 1alpha expression, increased hPASMC proliferation and migration. RNAseq revealed dysregulation in multiple pathways in WWOX-silenced hPASMCs, including cell-cycle regulation, TGF-β, mTOR and STAT3 signaling. Loss of WWOX showed decreased basal and maximal respiration, ATP production, reduced glycolytic capacity (40% in relation to control) and decreased mRNA expression of glycolytic enzymes (HK2, PFK1, PKM), generating 58% less lactate, and reduced mRNA expression of lactate dehydrogenase A. These cells exhibited increased mRNA and protein expression of glutaminase-1 and citrate lyase and had higher dependency on fatty acid oxidation and glutaminolysis compared to glucose, suggesting that these pathways are used when WWOX is suppressed. We also found mitochondrial hyperpolarization (JC-1 assay), decreased Sirtuin 3 expression, increased expression of acetylated manganese-superoxide dismutase, high production of reactive oxygen species and mitochondrial fission, indicating that loss of WWOX contributes to mitochondrial dysfunction. Conclusion: Loss of WWOX promotes cell cycle progression, proliferation, and metabolic reprogramming in PASMCs. Further studies are ongoing to better understand these effects.