Glutathione depletion inhibits oxidant-induced activation of nuclear factor-kappa B, AP-1, and c-Jun/ATF-2 in cultured guinea-pig gastric epithelial cells

Abstract:

The aim of this study was to reveal the role of intracellular glutathione in the oxidative stress responses of gastric epithelial cells. Metabolic radiolabeling with L-[³⁵S]methionine and analysis of synthesized proteins by gel electrophoresis and fluorography showed that upon exposure to hydrogen peroxide (H₂O₂) or diamide, primary cultures of guinea-pig gastric epithelial cells rapidly induced several undefined proteins, as well as heat shock proteins. When intracellular glutathione was depleted to less than 10% of the control value by treatment with buthionine-[S,R]-sulfoximine, these inductions were completely inhibited. Gel mobility shift assay demonstrated that H₂O₂ and diamide rapidly activated nuclear factor-kappa B (NF-κB), and diamide activated activator protein (AP)-1, and c-Jun/activating transcription factor (ATF)-2, suggesting that the response may be coupled to these reduction-oxidation (redox)-sensitive transcription factors, as well as heat shock transcription factor 1. The activations of NF-κB, AP-1, and c-Jun/ATF-2 by the oxidants did not occur in glutathione-depleted cells. Northern blot analysis showed that glutathione depletion markedly or completely suppressed the diamide-induced expression of c-fos and c-jun mRNAs. These results suggest that intracellular glutathione redox may participate in the initiation of oxidative stress responses; thereby, it plays an important role in gastric mucosal defense.