Abstract Rationale Vascular remodeling in PAH involves endothelial-to-mesenchymal transition (EndMT), characterized by phenotypic shift, increased proliferation, loss of endothelial markers, and acquisition of mesenchymal markers. Although in vitro models utilize TGF-β1 to induce these changes, there are limited strategies available for phenotypic reversal and functional restoration. Hypothesis Targeting TGF-β1 CDK5 axis, novel reversible CDK5 small molecule inhibitor can effectively restrict EndMT and restore endothelial function. Methods Mouse lung endothelial cells from Tg(TIE2GFP)287Sato/J mice were validated for purity via morphology and endothelial markers. EndMT was induced with TGF-β1 at multiple time points, assessing morphological and mesenchymal marker changes. CDK5 expression was examined in WT and SuHx5 rat lungs. TGF-β1-treated cells were combined with various CDK5 inhibitors, including a novel reversible small molecule, to evaluate CDK5’s role in EndMT. Functional assays included proliferation, angiogenesis, and collagen gel contraction. Single-cell RNA sequencing was performed across transition stages for detailed transcriptomic analysis. Results Elevated CDK5 expression in SuHx5 rat lungs indicates its role in PAH. TGF-β1 induced phenotypic transition, characterized by fibroblast-like morphology, increased proliferation, and upregulation of mesenchymal markers (αSMA, SM22α, Calponin). CDK5 inhibition, especially with a reversible inhibitor, suppressed these changes, restoring endothelial function by reducing mesenchymal markers and enhancing angiogenesis while decreasing collagen contraction. scRNA-seq revealed phenotypic reversal of EndMT through CDK5 inhibition. Conclusion CDK5 is a key regulator of EndMT in PAH. A novel small molecule inhibitor of CDK5 effectively suppresses EndMT and restores endothelial function in TGF-β1-stimulated mLECs, supporting CDK5 inhibition as a potential therapeutic strategy for vascular diseases, including PAH. This abstract is funded by: NIH
Bharti et al. (Fri,) studied this question.