Babicheva, Aleksandra

Abstract Endothelial-to-mesenchymal transition (EndMT) is a biological process through which lung vascular endothelial cells (ECs) transdifferentiate into mesenchymal-like cells. EndMT has recently been implicated in the development and progression of pulmonary vascular remodeling in pulmonary hypertension (PH); however, its underlying regulatory mechanisms remain incompletely understood. MicroRNAs (miRNAs) are key post-transcriptional regulators of EC gene expression and cellular responses to various stimuli. Notably, microRNA-153 (miR-153) has been shown to directly target SNAI1 to modulate epithelial-to-mesenchymal transition (EMT), a process closely related to EndMT and extensively studied in cancer. Whether miR-153 also participates in EndMT regulation, however, remains unknown. In this study, we demonstrate that 72-hour hypoxic exposure induces SNAI1-mediated EndMT in human lung vascular ECs. Hypoxia also increased cell proliferation and disrupted intercellular junctions, leading to enhanced endothelial permeability. Reduced miR-153 expression was observed in both hypoxia- and TGF-β1-induced EndMT, as well as in ECs isolated from PH patients exhibiting an EndMT phenotype. Similar to hypoxia, TGF-β1 promoted EC permeability. Loss of miR-153 enhanced SNAI1-mediated EndMT, endothelial survival, and permeability under normoxic conditions, whereas miR-153 overexpression attenuated EndMT induced by hypoxia or TGF-β1. However, miR-153 restoration did not completely restore endothelial barrier integrity disrupted by these stimuli. In vitro findings were validated in experimental PH model. In conclusion, miR-153 serves as a critical regulator of EndMT, maintaining endothelial identity and barrier function. Therapeutic delivery of miR-153 may therefore represent a novel strategy to inhibit EndMT and attenuate pulmonary vascular remodeling in PH.

Rationale: Pulmonary hypertension (PH) is a fatal disease characterized by a sustained elevated pulmonary vascular resistance that results in right heart failure. Despite therapeutic advances, no treatment provides disease progression reversion, resulting in poor long-term survival. Identifying new therapeutic targets for PH therefore requires a deeper understanding of its mechanisms. A defining feature of PH is pulmonary vascular remodeling, a result from excessive proliferation, migration, and apoptosis resistance of pulmonary artery smooth muscle cells (PASMC). SNAI1, a transcription factor regulating cell proliferation and differentiation during development and disease, has recently been implicated in endothelial dysfunction and endothelial-to-mesenchymal transition in PH. However, its role within PASMC remains unknown. Here, we aimed to define the contribution of SNAI1 in PASMC to the pathogenesis of PH. Methods: Normal human PASMC were exposed to normoxia (21% O 2 ) or hypoxia (3% O 2 ) for 72 hours. PASMC were also isolated from patients with or without PH. Cell proliferation and migration were assessed by EdU incorporation and wound scratch assays respectively. Mice were exposed to hypoxia (10% O 2 ) or Sugen5416/hypoxia (10% O 2 plus Sugen5416 20mg/kg, i.p., weekly) for 6 weeks. PH phenotype was characterized by right heart catheterization, right ventricle (RV) hypertrophy by Fulton index and pulmonary artery (PA) remodeling by H&E staining of lung tissues. Pharmacological blocker GN-25 was used to inhibit SNAI1 in vitro and in vivo. Results: SNAI1 expression was markedly increased in hypoxic PASMC and in PASMC from patients with PH. Hypoxia also upregulated HIF-1alpha, Vimentin, p-AKT, PCNA at the protein level in PASMC. EdU incorporation and migration rates were significantly higher in hypoxic cells as well as in PASMC isolated from PH patients compared with controls. In vitro, GN-25 (100 nM) abolished hypoxia-induced PASMC proliferation and migration. Right ventricular systolic pressure (RVSP), a surrogate measurement of PA systolic pressure, estimated mean PAP (mPAP), and RV contractility (RV-±dP/dtmax) were significantly increased in both PH models compared with controls. Additionally, Sugen5416/hypoxia-treated mice exhibited higher RVSP, mPAP and RV-±dP/dtmax compared to hypoxia-treated animals. The Fulton index (weight of RV vs. left ventricle and septum), was increased in both PH groups indicating development of RV hypertrophy in PH compared to control. PH mice lost body weight by 5-10%. Additionally, when Fulton index was normalized by body weight, RV hypertrophy was significantly higher in Sugen5416/hypoxia-treated mice compared with hypoxic animals. Lung tissues from PH mice exhibit PA hypertrophy (media remodeling) with some PA partially or completely occluded. Additionally, we detected protein upregulation of Snai1 and Vimentin in lung tissues of PH mice in both groups. Treatment of PH mice with GN-25 (20 mg/kg, daily, for the last 3 weeks under PH stimuli) in reversal experiments attenuated PH phenotype in both animal models by decreasing hemodynamic parameters, right ventricle hypertrophy and pulmonary vascular remodeling. Conclusions: Our findings identify a key role for SMC SNAI1 in the pathogenesis of lung vascular remodeling during hypoxia-induced PH. Inhibition of SNAI1 in PASMC may constitute a new therapeutic strategy for the treatment of PH. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

EndMT has been reported to contribute to the pathogenesis of PAH. In this study, we aimed to determine the role of Ca 2+ signaling in the development of EndMT in human lung vascular endothelial cells. Our data suggest that TGF-β 1 requires store-operated Ca 2+ entry through STIM1/Orai channels to induce SNAI-mediated EndMT. For the first time, we demonstrated that TGF-β 1 -induced EndMT is a Ca 2+ -dependent event, whereas inhibition of STIM1/Orai interaction attenuated EndMT in response to TGF-β 1 .