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*
Laboratory of Immunology, Korea Research Institute of Bioscience and Biotechnology, Taejon, Republic of Korea;
Department of Biology, Yonsei University, Seoul, Republic of Korea;
Department of Biological Science, Ewha Womans University, Seoul, Republic of Korea;
§
Biomolecule Research Team, Korea Basic Science Institute, Taejon, Republic of Korea; and
¶
Laboratory of Cell Signaling, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892
Stimulation of human lung fibroblast cells with TGF-ß1 resulted in a transient burst of reactive oxygen species with maximal increase at 5 min after treatment. This reactive oxygen species increase was inhibited by the antioxidant, N-acetyl-L-cysteine (NAC). TGF-ß1 treatment stimulated IL-6 gene expression and protein synthesis in human lung fibroblast cells. Antioxidants including NAC, glutathione, and catalase reduced TGF-ß1-induced IL-6 gene expression, and direct H2O2 treatment induced IL-6 expression in a dose-dependent manner. NAC also reduced TGF-ß1-induced AP-1 binding activity, which is involved in IL-6 gene expression. It has been reported that Ca2+ influx is stimulated by TGF-ß1 treatment. EGTA suppressed TGF-ß1- or H2O2-induced IL-6 expression, and ionomycin increased IL-6 expression, with simultaneously modulating AP-1 activity in the same pattern. PD98059, an inhibitor of mitogen-activated protein kinase (MAPK) kinase/extracellular signal-related kinase kinase 1, suppressed TGF-ß1- or H2O2-induced IL-6 and AP-1 activation. In addition, TGF-ß1 or H2O2 increased MAPK activity which was reduced by EGTA and NAC, suggesting that MAPK is involved in TGF-ß1-induced IL-6 expression. Taken together, these results indicate that TGF-ß1 induces a transient increase of intracellular H2O2 production, which regulates downstream events such as Ca2+ influx, MAPK, and AP-1 activation and IL-6 gene expression.
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