Thiram-induced cytotoxicity is accompanied by a rapid and drastic oxidation of reduced glutathione with consecutive lipid peroxidation and cell death

Abstract

The toxic effect of thiram, a widely used dithiocarbamate fungicide, was investigated in cultured human skin fibroblasts. Cell survival assays demonstrated that thiram induced a dose-dependent decrease in the viable cell recovery. Thiram exposure resulted in a rapid depletion of intracellular reduced glutathione (GSH) content with a concomitant increase in oxidized glutathione (GSSG) concentration. Alteration of glutathione levels was accompanied by a dose-dependent decrease in the activity of glutathione reductase (GR), a key enzyme for the regeneration of GSH from GSSG. Thiram-exposed cells exhibited increased lipid peroxidation reflected by enhanced thiobarbituric acid reactive substances (TBARS) production, suggesting that GSH depletion and the lower GR activity gave rise to increased oxidative processes. To investigate the role of decreased GSH content in the toxicity of thiram, GSH levels were modulated prior to exposure. Pretreatment of fibroblasts with N-acetyl- l-cysteine (NAC), a GSH biosynthesis precursor, prevented both lipid peroxidation and cell death induced by thiram exposure. In contrast, thiram cytotoxicity was exacerbated by the previous depletion of cellular GSH by l-buthionine-(S,R)-sulfoximine (BSO). Taken together, these results strongly suggest that thiram induces GSH depletion, leading to oxidative stress and finally cell death.