主催: 日本毒性学会
Butylated hydroxyanisole (BHA) is a phenolic antioxidant and classified class-2B carcinogen. It is biotransformed to tert-butylhydroquinone (TBHQ), which readily auto-oxidizes to the electrophlic metabolite tert-butylbenzoquinone (TBQ). These metabolites are associated with reactive oxygen species (ROS) generation. Several lines of evidence suggest that TBHQ derived from BHA can activate Nrf2, a transcription factor that is negatively regulated by Keap1 and plays a role in the initial response to chemicals causing oxidative or electrophilic stress. However, the relative contributions of oxidative stress and covalent modification of TBQ, derived from TBHQ, to Nrf2 activation are not well understood. Here, we examined the role of TBQ in Nrf2 activation by using RAW264.7 cells and recombinant Keap1 protein. Exposure of RAW264.7 cells to TBQ activated Nrf2 and up-regulated its downstream proteins responsible for detoxification of TBQ; under these conditions, TBQ produced cellular ROS. However, while pretreatment with catalase conjugated with polyethylene glycol (PEG-CAT) did not affect the TBQ-induced activation of Nrf2, the ROS generation caused by TBQ was abolished by PEG-CAT, suggesting that ROS is not the dominant factor for TBQ-dependent Nrf2 activation. Furthermore, UPLC-MSE analysis with purified Keap1 revealed that TBQ covalently binds to Keap1 through Cys23, Cys151, Cys226 and Cys368. Interestingly, Nrf2 silencing in RAW264.7 cells significantly enhanced TBQ-induced cytotoxicity. Taken together, we conclude that TBQ derived from BHA activates Nrf2 through electrophilic modification of Keap1 rather than ROS formation and that Keap1/Nrf2 system plays a role in the detoxification of TBQ.
Abiko Y. et al., Toxicology and Applied Pharmacology, 255, 32-39, 2011.
