An IL-2 dependent adult T cell leukemia cell line (SO4) has been established that is sensitive to CD95-mediated apoptosis as well as a subline (R-SO4) that is resistant. Incubating SO4 cells with anti-CD95 IgM mAb caused concentration-dependent cell death. On the contrary, R-SO4 cells did not die even at 1000 ng/ml of anti-CD95 IgM mAb. The levels of CD95 expression on R-SO4 cells were one-third of those on SO4 cells. However a blocking Ab, anti-CD95 IgG mAb, did not induce complete resistance of SO4 cells to anti-CD95 IgM mAb as R-SO4 cells. As CD95 and TNF receptor are similar, and TNF/TNF receptor binding induces oxygen radicals, the involvement of oxidant and antioxidant systems in CD95-mediated apoptosis has been examined. The addition of anti-CD95 IgM mAb resulted in formation of intracellular oxygen radical species in the SO4 cells as measured using 2',5',-dichlorofluorescein as substrate. The oxygen radical production induced DNA damage as determined by formation of 8-hydroxydeoxyguanosine. No increase in the formation of oxygen radicals was observed in R-SO4 cells. Concentrations of the intracellular antioxidant, glutathione, and the key enzyme for its synthesis, gamma-glutamylcysteine synthetase, were 150% increased in R-SO4 cells in comparison with that of SO4 cells. Moreover, glutathione ester decreased the formation of 8-hydroxydeoxyguanosine. These results suggested that apoptosis mediated by CD95 in ATL cells is related to the production of oxygen radical species and cellular antioxidant systems, especially, glutathione synthesis.