Glutathione regulates transforming growth factor-[beta]-stimulated collagen production in fibroblasts

Transforming growth factor-[beta] (TGF-[beta]) is a potent fibrogenic cytokine. The molecular mechanism underlying TGF-[beta] fibrogenesis, however, has not been completely elucidated. In this study, we showed that TGF[beta] decreased the intracellular GSH content in murine embryo fibroblasts (NIH 3T3), which was followed by an increase in collagen I mRNA content and collagen protein production. Prevention of GSH depletion with N-acetylcysteine (NAC), GSH, or GSH ester abrogated TGF-[beta]-stimulated collagen production, whereas a decrease in intracellular GSH content with L-buthionine-S,R-sulfoximine, an inhibitor of de novo GSH synthesis, enhanced TGF-[beta]-stimulated collagen production. These results suggest that GSH depletion induced by TGF-[beta] may mediate TGF-[beta]-stimulated collagen production. In addition, we showed that TGF-[beta] stimulated superoxide production and increased release of [H.sub.2][O.sub.2] from the cells, whereas GSH ester decreased basal and TGF-[beta] + glucose oxidase-stimulated [H.sub.2][O.sub.2] release. H202, exogenously added or continuously generated by glucose oxidase, enhanced TGF-[beta]-stimulated collagen production, whereas suppression of superoxide production by diphenyliodonium, an NAD(P)H oxidase inhibitor, blocked TGF-[beta]-stimulated collagen production. These data further suggest that reactive oxygen species are involved in TGF-[beta]-stimulated collagen production and that the effect of GSH depletion on TGF-[beta]-stimulated collagen production may be mediated by facilitating reactive oxygen species signaling. glutathione ester; [[sup.3]H]proline incorporation; collagen I mRNA; reactive oxygen species