Role of Reactive Oxygen Species in Transforming Growth Factor - beta1 - inuduced Fibronectin Secretion and  alpha - Smooth Muscle Actin Expression in Human Lung Fibroblasts.

Ha, Hunjoo; Yu, Mi Ra; Uh, Soo Taek; Park, Choon Sik; Lee, Hi Bahl

doi:2005.58.3.267

Tuberc Respir Dis > Volume 58(3); 2005 > Article

Original Article

Tuberculosis and Respiratory Diseases 2005;58(3):267-275.
DOI: https://doi.org/10.4046/trd.2005.58.3.267 Published online March 1, 2005.

Role of Reactive Oxygen Species in Transforming Growth Factor - beta1 - inuduced Fibronectin Secretion and alpha - Smooth Muscle Actin Expression in Human Lung Fibroblasts.

Hunjoo Ha, Mi Ra Yu, Soo Taek Uh, Choon Sik Park, Hi Bahl Lee

¹Hyonam Kidney Laboratory, Soon Chun Hyang University, Korea. hblee@hkl.ac.kr
²Ewha Womans University College of Pharmacy, Korea.
³Department of Internal Medicine, Soon Chun Hyang University College of Medicine, Korea.

Abstract

BACKGROUND
The transforming growth factor-beta1 (TGF-beta1) plays a key role in lung fibrosis. However, the mole?cular mechanisms involved in TGF-beta1-induced lung fibrosis are unclear. TGF-beta1 is the key inducer of myofibroblast transdifferentiation via de novo synthesis of alphasmooth muscle actin (alpha-SMA). Since TGF-beta1 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-beta1-induced fibronectin secretion and alpha-SMA expression in human lung fibroblasts, MRC-5 cells. METHODS: Growth arrested and synchronized MRC-5 cells were stimulated with TGF-beta1 (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 alpha-SMA by Western blot analysis. RESULTS: TGF-beta1 increased the level of fibronectin secretion and alpha-SMA expression in MRC-5 cells in a dose- dependent manner. Both NAC (20 and 30 mM) and DPI (1 and 5 microM significantly inhibited TGF-beta1-induced fibronectin and alpha-SMA upregulation. The TGF-beta1-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-beta1-induced fibronectin secretion and alpha-SMA expression in MRC-5 cells, which leads to myofibroblast transdifferentiation and progressive lung fibrosis.

Key Words: Pulmonary fibrosis, Transforming growth factor-beta1, Reactive oxygen species, alpha-Smooth muscle actin, Fibroblast, Myofibroblast, Fibronectin