Abstract Background Pulmonary arterial hypertension (PAH) is a progressive disease marked by pulmonary microvascular loss and vascular remodeling. Oxidative stress and apoptosis in pulmonary endothelial cells are central features of endothelial dysfunction in PAH. Carboxylesterase 1 (CES1), an endoplasmic reticulum-associated enzyme, regulates lipid metabolism, cellular detoxification, and redox balance. Previous work demonstrated that CES1 deficiency causes lipotoxicity and lipid peroxidation in human pulmonary microvascular endothelial cells (PMVECs). Transcriptomic analyses (bulk RNA-seq and scRNA-seq) revealed modulation of ferroptosis-related genes following CES1 knockdown in PMVECs and CES1 knockout in mice. We hypothesized that CES1 deficiency induces ferroptosis in PMVECs, contributing to endothelial dysfunction in PAH. Methods PMVECs derived from healthy donors and PAH patients were transfected with siRNA or CES1-expressing plasmids to knockdown or overexpress CES1, respectively. Cells were treated with the ferroptosis inducer erastin (10 µM) or the ferroptosis inhibitor ferrostatin-1 (10 nM). Functional assays included lipid peroxidation, immunofluorescence staining, cell viability and tube formation on Matrigel. Western blot was performed to assess the ferroptosis inducing proteins. Results CES1 knockdown in PMVECs increased lipid peroxidation as seen by BODIPY C11 and cell death, both of which were exacerbated by erastin treatment. CES1-deficient PMVECs also showed impaired tube formation and reduced survival, indicating endothelial dysfunction l. Ferrostatin-1 treatment reduced lipid peroxidation and cell death, while improving tube formation and survival. In PAH-derived PMVECs, CES1 overexpression and ferrostatin-1 treatment both mitigated lipid peroxidation and cell death. Conclusions CES1 deficiency promotes endothelial dysfunction in PAH, potentially via ferroptosis. These findings suggest that targeting ferroptotic pathways may offer a novel therapeutic strategy for restoring endothelial function in PAH. This abstract is funded by: NIH
Agarwal et al. (Fri,) studied this question.