Abstract Intro Pulmonary arterial hypertension (PAH) is a rare disorder characterized by excessive smooth muscle cell (SMC) accumulation and elevated pulmonary artery pressure. Endothelial cells (ECs) can contribute to vascular remodeling through endothelial-to-mesenchymal transition (EndoMT); however, the upstream molecular drivers of this process remain unclear. Vimentin, a cytoskeletal regulator of epithelial-to-mesenchymal transition (EMT), may similarly modulate EndoMT in PAH and contribute to disease development. Methods We established a murine model of severe pulmonary hypertension (PH) by performing extended pneumonectomy (EP, removal of the left lung and right caval lobe) in adult mice and then exposing them to three weeks of hypoxia (10% FiO2) on post-operative day (POD) 7. Hemodynamic assessments for PH were performed on POD 28. SMCs were fate-mapped using Acta2-CreER::R26-mTmG mice. Single-cell RNA-seq (scRNA-seq) was conducted in all conditions (sham, EP, sham/Hx, EP/Hx) using 10x Genomics. A human idiopathic PAH (IPAH) scRNA-seq dataset was analyzed for validation. Vimentin expression was evaluated with immunofluorescence (IF) staining of precision-cut lung slices (PCLSs) from EP/Hx mice and human PAH lung tissues. Results EP/Hx mice developed more severe PH compared to Hx-only controls, demonstrated by elevated right ventricle systolic pressure (RVSP 40.7 vs 33.7mm Hg) and right ventricle hypertrophy (Fulton Index: 42.6 vs 32.8%) (Fig 1A). In EP/Hx SMC fate-mapping lungs, a subset of Sma+/GFP- cells appeared in distal vessels, suggesting a non-SMC source contributing to remodeling. Differential gene expression analysis of EP/Hx scRNA-seq data revealed that capillary ECs exhibited increased expression of SMC markers (Acta2: 3.4-fold; Tagln: 2.3-fold) and Vim (1.1-fold), with a concomitant decrease in EC markers Eng (0.2-fold) and Cdh5 (0.1-fold). Similarly, human IPAH capillary ECs also upregulated TAGLN and VIM. Gene set enrichment analysis revealed upregulation of EMT, myogenesis, and TGF-β signaling in both datasets. Lastly, IF staining confirmed increased Vimentin expression in ECs within remodeled vessels in both EP/Hx mice and human PAH tissues (Fig 1B). Conclusion ECs contribute to vascular remodeling in PH through EndoMT, and our results show Vimentin emerges as a regulator of endothelial plasticity in PH. The EP/Hx mouse model provides a robust platform for investigators to study EndoMT and evaluate potential therapeutic interventions targeting PH. This abstract is funded by: NHLBI
Klouda et al. (Fri,) studied this question.