Role of Reactive Oxygen Species in Transforming Growth Factor - β1 - inuduced Fibronectin Secretion and α - Smooth Muscle Actin Expression in Human Lung Fibroblasts

Background : The transforming growth factor-1 (TGF-1) plays a key role in lung fibrosis. However, the mole cular mechanisms involved in TGF-1-induced lung fibrosis are unclear. TGF-1 is the key inducer of myofibroblast transdifferentiation via de novo synthesis of -smooth muscle actin (-SMA). Since TGF-1 signals through reactive oxygen species (ROS) and ROS have been shown to induce accumulation of extracellular matrix (ECM) in various tissues, this study examined if ROS play a role in TGF-1-induced fibronectin secretion and -SMA expression in human lung fibroblasts, MRC-5 cells. Methods : Growth arrested and synchronized MRC-5 cells were stimulated with TGF-1 (0.2-10 ng/ml) in the presence or absence of N-acetylcysteine (NAC) or diphenyleneiodonium (DPI) for up to 96 hours. Dichlorofluorescein (DCF)-sensitive cellular ROS were measured by FACScan and secreted fibronectin and cellular -SMA by Western blot analysis. Results : TGF-1 increased the level of fibronectin secretion and -SMA expression in MRC-5 cells in a dosedependent manner. Both NAC (20 and 30 mM) and DPI (1 and 5 M) significantly inhibited TGF-1-induced fibronectin and -SMA upregulation. The TGF-1-induced cellular ROS level was also significantly reduced by NAC and DPI. Conclusions : The results suggest that NADPH oxidase-dependent ROS play an important role in TGF-1-induced fibronectin secretion and -SMA expression in MRC-5 cells, which leads to myofibroblast transdifferentiation and progressive lung fibrosis. (