Effects of Glutathione on Antioxidant Response Element-Mediated Gene Expression and Apoptosis Elicited by Sulforaphane

Sulforaphane (SFN) and its N-acetyl-L-cysteine (NAC) conjugate are effective inhibitors of tumorigenesis in animal models. These compounds induce the expression of the antioxidant response element (ARE)-related genes and cause apoptosis. We studied the role of reduced glutathione (GSH) in the activations of ARE-mediated gene expression, apoptosis, and the activation of c-Jun NH₂-terminal kinase (JNK) in HepG2-C8 cells. The cellular level of GSH decreased transiently when cells were exposed to SFN and then increased from 4 h, reaching 2.2-fold over control at 24 h. In contrast, SFN-NAC did not change the GSH level substantially during the time of incubation. ARE expression was increased in a dose-dependent manner up to 35 μM SFN and 75 μM SFN-NAC, respectively. The induction of ARE by SFN was 8.6-fold higher than that by SFN-NAC. Pretreatment with L-buthionine sulfoximine increased SFN-induced ARE expression significantly. The decrease in ARE expression at higher concentrations of SFN and SFN-NAC was correlated with accelerated apoptotic cell death, with a dose-dependent activation of caspase 3 activity by SFN. On addition of extracellular GSH within 6 h of treatment with SFN, the effect on ARE expression was blocked almost completely. SFN was able to activate JNK1/2, and that activation was blocked by treatment with exogenous GSH. Taken together, these results suggest that the biological effects of SFN and SFN-NAC on the induction of ARE-related gene expression and apoptosis could be different from each other; however, the different effects on ARE-related gene expression and apoptosis elicited by SFN can be blocked by the addition of GSH.