Abstract 4373256: BMP9 drives vasoactive gene expression in endothelial cells via non-canonical ALK1-SMAD3 signaling

Abstract

Rationale: The role of BMP9 in pulmonary arterial hypertension (PAH) remains controversial. Loss-of-function GDF2 /BMP9 mutations in heritable PAH suggest its function as a vascular endothelial homeostasis factor, however, modulating BMP9 signaling in experimental pulmonary hypertension (PH) models can yield pathogenic or protective effects. The impact of BMP9 upon intracellular and intercellular angiogenic signaling cascades remains poorly understood. Methods/Results: We analyzed BMP9-mediated transcription in pulmonary microvascular endothelial cells (PMVEC) derived from controls and patients, and in the presence or absence of co-cultured pulmonary artery smooth muscle cells (PASMC). We tested recombinant BMP9, BMP9/BMP10 ligand trap ALK1-Fc, anti-BMP9, and activin/GDF ligand trap ACTRIIA-Fc in hypoxia, SUGEN5416+hypoxia (SU-Hx) and monocrotaline (MCT) experimental PH rodent models. The BMP9-regulated secretome of ECs was examined for potential modulation of PASMC phenotype and function. Inhibition of BMP9 and/or BMP10 was protective whether administered before or after the development of experimental PH, and attenuated experimental PH when administered therapeutically in the SU-Hx model. In PMVEC, BMP9 elicited expression of vasoactive genes that were also elevated in experimental PH and human PAH lungs, including EDN1, CXCL12, IGFBP4, COL18A1, VEGFA, PDGFB , and SERPINE1 , several of which were normalized in lungs of SU-Hx rats with anti-BMP9 treatment. Several of these genes required non-canonical activation of SMAD3 downstream of BMPR2, ALK1, and ENG. Promoter analysis of human ET-1 revealed cooperation of SMAD3 and SMAD1/5 binding elements is required for BMP9-mediated expression of ET-1 . Co-culture models revealed the essential role of BMP9-mediated PMVEC paracrine signaling in modulating PASMC contractile phenotype markers ( CNN1 ; TAGLN ), which was attenuated by anti-CXCL12, CXCR4 antagonist, or anti-BMP9. Conclusions: BMP9 is a central regulator of vasoactive endothelial genes via ALK1-SMAD3 signaling that modulate PASMC phenotype and contribute to experimental PH.