Lipopolysaccharide Activates Toll-Like Receptor 4 and Prevents Cardiac Fibroblast-to-Myofibroblast Differentiation

Bacterial lipopolysaccharide (LPS) is a known ligand of Toll-like receptor 4 (TLR4) which is expressed in cardiac fibroblasts (CF). Differentiation of CF to cardiac myofibroblasts (CMF) is induced by transforming growth factor-β1 (TGF-β1), increasing alpha-smooth muscle actin (α-SMA) expression. In endothelial cells, an antagonist effect between LPS-induced signaling and canonical TGF-β1 signaling was described; however, it has not been studied whether in CF and CMF the expression of α-SMA induced by TGF-β1 is antagonized by LPS and the mechanism involved. In adult rat CF and CMF, α-SMA, ERK1/2, Akt, NF-κβ, Smad3, and Smad7 protein levels were determined by western blot, TGF-β isoforms by ELISA, and α-SMA stress fibers by immunocytochemistry. CF and CMF secrete the three TGF-β isoforms, and the secretion levels of TGF-β2 was affected by LPS treatment. In CF, LPS treatment decreased the protein levels of α-SMA, and this effect was prevented by TAK-242 (TLR4 inhibitor) and LY294002 (Akt inhibitor), but not by BAY 11-7082 (NF-κβ inhibitor) and PD98059 (ERK1/2 inhibitor). TGF-β1 increased α-SMA protein levels in CF, and LPS prevented partially this effect. In addition, in CMF α-SMA protein levels were decreased by LPS treatment, which was abolished by TAK-242. Finally, in CF LPS decreased the p-Smad3 phosphorylation and increased the Smad7 protein levels. LPS treatment prevents the CF-to-CMF differentiation and reverses the CMF phenotype induced by TGF-β1, through decreasing p-Smad3 and increasing Smad7 protein levels.