EFFECTS OF GLUTATHIONE DEPLETION ON CADMIUM-INDUCED METALLOTHIONEIN SYNTHESIS, CYTOTOXICITY, AND PROTOONCOGENE EXPRESSION IN CULTURED RAT MYOBLASTS

Cadmium (Cd) is a highly toxic metal and a known carcinogen. Although the carcinogenic mechanism of action is unknown, Cd will induce transc riptional activation of c-myc and c-jun. We have previously found that the extent of Cd-induced oncogene expression is limited by the presen ce of cellular metallothionein (MT) in rat L6 myoblasts. Glutathione ( GSH) is thought to play an important role in protection against Cd bef ore the onset of MT synthesis. Thus, this study examined the effects o f GSH depletion on Cd-induced MT synthesis, cytotoxicity, and proto-on cogene expression in rat L6 myoblasts after pretreatment with L-buthio nine sulfoximine (BSO), a potent inhibitor gamma-glutamylcysteine synt hetase, which effectively depletes GSH. Exposure of L6 cells to BSO (5 or 25 mu M) resulted in a dose-dependent decrease in cellular GSH lev els, GSH depletion had no effect on Cd- or zinc-induced MT synthesis. Although the depletion of GSH was not itself cytotoxic in L6 cells, BS O pretreatment, particularly at the higher dose (25 mu M), resulted in a dose-dependent increase in the sensitivity to Cd cytotoxicity, as a ssessed by a tetrazolium-based dye (MTT) assay. Low levels of Cd (1 mu M) slightly increased the expression of both c-myc and c-jun as asses sed by increases in gene-specific mRNA levels, in accordance with prev ious studies. GSH depletion (5 mu M BSO) likewise caused an increase i n expression of c-myc and c-jun. However, combined GSH depletion and C d exposure decreased levels of c-myc and c-jun transcription well belo w control levels. These results suggest that increased cytotoxicity re sulting from exposure to Cd after BSO depletion of cellular GSH abroga tes the oncogene activation observed after either treatment alone. Thu s proto-oncogene expression induced by Cd appears to be dependent on t he absence of overt Cd-induced cytotoxicity.