TNF drives aberrant BMP signaling to induce endothelial and mesenchymal dysregulation in pulmonary hypertension.

Abstract

The pathobiology of pulmonary hypertension (PH) is complex and multiple cell types contribute to disease pathogenesis. We sought to characterize the molecular crosstalk between endothelial and mesenchymal cells that promote PH in the tumor necrosis factor α-transgenic (TNF-Tg) model of PH. Pulmonary endothelial and mesenchymal cells were isolated from WT and TNF-Tg mice and underwent single-cell RNA sequencing. Data were analyzed using clustering, differential gene expression and pathway analysis, ligand-receptor interaction, transcription factor binding, and RNA velocity assessments. Significantly altered ligand-receptor interactions were confirmed with immunofluorescent staining. TNF-Tg mice had increases in smooth muscle cells and Col14+ fibroblasts, and reductions in general capillary (gCAP) endothelial cells, Col13+ fibroblasts, pericytes, and myofibroblasts. Pathway analysis demonstrated NF-κB-, JAK/STAT-, and interferon-mediated inflammation, endothelial apoptosis, loss of vasodilatory pathways, increased TGF-β signaling, and smooth muscle cell proliferation. Ligand-receptor analysis demonstrated a loss of BMPR2 signaling in TNF-Tg lungs and establishment of a maladaptive BMP signaling cascade, which functional studies revealed stemmed from endothelial NF-κB activation and subsequent endothelial SMAD2/3 signaling. This system highlights a complex set of changes in cellular composition, cell communication, and cell fate driven by TNF signaling that lead to aberrant BMP signaling that is critical for development of PH.