MMP14-generated endostatin drives vascular remodeling via Notch3 activation in idiopathic pulmonary arterial hypertension

<bold>Introduction:</bold> Pulmonary arterial hypertension (PAH) is driven by chronic vascular remodeling that leads to extensive changes in the basement membrane (BM). Increased degradation of BM components releases active yet unexplored signaling peptides called matrikines. Here, we investigate the effects of the collagen XVIII-derived matrikine endostatin (ES) in idiopathic PAH. <bold>Methods:</bold> We employed single-cell RNA sequencing and immunoblotting to assess Col18a1 and ES levels in IPAH and control lungs. Mechanistic and functional effects of ES on pulmonary arterial endothelial cells (PAECs) and smooth muscle cells (PASMCs) were analyzed using EdU assays, immunoblots and co-immunoprecipitation (Co-IP). Relevant proteases were studied in PASMC and macrophage co-cultures. <italic>In vivo</italic>, protease inhibition was tested in the Sugen5416/hypoxia (SuHox) rat and mouse model. <bold>Results:</bold> Col18a1 and ES levels were significantly elevated in PAH patients. Recombinant ES reduced the proliferation of PAECs while increasing proliferation, collagen I expression and Notch3 activation in PASMCs. Further Co-IP experiments confirmed ES as a novel non-canonical activator of Notch3. The specific MMP14 inhibitor NSC405020 prevented ES fragmentation, Notch3 activation and ES-induced PASMC proliferation <italic>in vitro</italic>. Application of NSC405020 in SuHox mice and rats significantly reduced right heart hypertrophy, arterial muscularization and tendentially ameliorated RVSP. <bold>Conclusion:</bold> Our results delineate the proteolytic mechanisms and functional receptor interactions of ES. Targeting ES via MMP14 inhibition improved PAH <italic>in vivo</italic> and presents a promising approach for the restauration of a healthy pulmonary vasculature.