THE INFLUENCE OF MYOCARDIN EFFECTOR GENES ON THE TGF-BETA SMAD3 SIGNALING PATHWAY IN HUMAN PLEURAL MESOTHELIAL CELLS

Pleural Fibrosis (PF) involves excessive extracellular matrix (ECM) deposition, mainly by myofibroblasts, leading to pleural thickening and impaired lung function. Pleural mesothelial cells (PMCs) contribute via mesothelial and mesenchymal transition (MesoMT), triggered by cytokines like TGF-β. Our lab identified Myocardin (MyoCD), a transcriptional co-activator of cardiac and smooth muscle, is a master regulator of pleural fibrosis through interactions with serum response factor (SRF) and Smad2/3 transcription factors which initiate fibrosis signaling. RNA sequencing and qPCR analysis revealed that silencing MyoCD decreased the expression of CTNNB1, MDFI, and Tspan2. MDFI emerged as a potent candidate, as this gene affecting the expression of Fibrosis markers. Silencing MDFI reduced α-SMA, PAI-1, CNN-1 and MyoCD expression, while increasing the FN-1 and COL-1 mRNA levels. Protein analysis revealed increased expression of COL-1, FN-1, along with enhanced secretion of COL-1, FN-1 and PAI-1, with minimal intracellular changes. These findings suggest that MDFI knockdown influences gene expression and affects the secretion of ECM proteins. Immunocytochemistry revealed enhanced stress fiber formation. These findings suggest that MDFI regulates TGF-β-induced myofibroblast differentiation, through cytoskeletal reorganization in HPMCs via the MyoCD/Smad3 signaling axis, contributing to the pathogenesis of pleural fibrosis.