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A3754 - Reduced BMPR2 Signaling Alters the Expression of Hypertrophy Related Genes in Induced Pluripotent Stem Cell Derived Cardiomyocytes of PAH Patients
Author Block: K. Kuramoto1, M. Boehm1, X. Tian1, S. Dannewitz1, M. Gu2, S. Silin2, T. Seeger3, T. Kitani3, J. C. Wu3, M. Rabinovitch2, E. F. Spiekerkoetter1; 1Pulmonary and Critical Care Medicine, Stanford University School of Medicine, Stanford, CA, United States, 2Department of Pediatrics, Cardiology, Stanford University School of Medicine, Stanford, CA, United States, 3Cardiovascular Institute, Stanford University School of Medicine, Stanford, CA, United States.
Introduction: Right ventricular (RV) failure is the major cause of death in patients with pulmonary arterial hypertension (PAH), yet some patients seem to be able to adjust better to the increased RV afterload by developing adaptive RV hypertrophy. Bone Morphogenetic Protein Receptor 2 (BMPR2) signaling is reduced in familial PAH (FPAH) patients harboring a BMPR2 mutation. And it is those familial PAH patients who present with more severe disease at a younger age, have worse RV function despite similar pulmonary vascular resistance and an increased mortality. We hypothesized that BMPR2 expression and signaling play an important role for cardiomyocyte function and its adaptation to an increased afterload and that reduced BMPR2 signaling might facilitate RV maladaptation.Recent reports showed that endothelial cells (EC) differentiated from induced pluripotent stem cells (iPSC) from idiopathic (I)PAH and FPAH patients mimicked the endothelial dysfunction and altered gene expression of primary pulmonary endothelial cells. It suggested that iPSC-ECs were good surrogates for modeling of a universal EC dysfunction in PAH. To identify inherent potential protective and deleterious pathways for RV adaptation and to elucidate the role of reduced BMPR2 signaling in cardiomyocytes, we assessed the gene expression and cell function of iPSC derived cardiomyocytes from IPAH and FPAH patients compared to healthy controls. Methods and Results: We differentiated cardiomyocytes from three familial PAH iPSCs (FPAH-CM) with a BMPR2 mutation, four IPAH patient iPSCs (IPAH-CM) and five non-PAH donor iPSCs (donor-CM), using iPSCs derived from skin fibroblasts from patients and donors collected through the pulmonary hypertension breakthrough initiative. Cardiomyocyte differentiation was induced by sequential treatment of glycogen synthase kinase 3 beta inhibitor and Wnt inhibitor. We found no significant difference in differentiation efficiency and spontaneous beating motion between F/IPAH-CM and donor-CM, but expression differences in hypertrophy related gene, such as beta- and alpha myosin heavy chain, and ANP in FPAH-CM compared to donor-CM. To verify whether those gene expression changes were related to a BMPR2 signaling defect, we generated BMPR2 gene mutation corrected FPAH-CM by CRISPR technology. Phosphorylation of SMAD1 and ID1 gene expression, important downstream molecules of BMPR2 signaling, were increased by correction of the BMPR2 gene mutation, and the expression of the hypertrophy-related gene was restored. Conclusion: Our results suggest that the inherent decrease in BMPR2 signaling in iPSC-CM from FPAH patients leads to alterations of hypertrophy-related genes and might facilitate failure to adaptation to an increased RV afterload.