Study on the apoptosis mechanisms in NIH/3T3 cells induced by tetrachlorohydroquinone（TCHQ） and elucidate the role of antioxidants during the process of TCHQ-induced genotoxicity and cytotoxicity

• Main Authors: Yu-Ping Lin, 林裕萍
• Other Authors: Ying-Jan Wang
• Format: Others
• Language: zh-TW
• Published: 2002
• Online Access: http://ndltd.ncl.edu.tw/handle/3q4guh

碩士 === 國立成功大學 === 環境醫學研究所 === 90 === Tetrachlorohydroquinone (TCHQ) has been identified as a major toxic metabolite of the widely used wood preservative pentachlorophenol and also implicated in its genotoxicity. In our preliminary two-stage carcinogenesis experiment, it was proved that the tumor promotion activity of PCP was higher than TCHQ but the toxicity of TCHQ was higher than PCP. We assumed that TCHQ, but not PCP, may cause apoptotic cell death and thus decrease the tumor promotion activity and tumorigenecity. This study was designed to test the hypothesis that TCHQ induced apoptotic cell death in 3T3 cells and to investigate the mechanisms of apoptosis induced by TCHQ. Moreover, we identified some antioxidants with the ability to reduce TCHQ-induced ROS, and defined whether these agents may inhibit cytotoxicity and genotoxicity induced by TCHQ. In the present study, we found that apoptotic cell death can be induced in 3T3 cells treated with 50μM TCHQ for 8 hrs. The generation of ROS and collapse of mitochodrial transmembrane potrntial has been observed to occur in early apoptosis induced by TCHQ, and the process is related to bcl-2 protein expression. The activity of caspase9 and caspase3 subsequently increased with a time- and dose- dependent pattern. Bcl-2 over-expressed 3T3 cells provided marked protection against the cytotoxicity induced by TCHQ. This result suggested that Bcl-2 protein play an important role in the process of apoptosis in 3T3 cells, which also further indicated that apoptotic cell death in 3T3 cells occurs through the mitochondria-mediated pathway. Captopril, penicillamine, dihydrolipoic acid (DHLA) and 2,3-dimercaptosuccinic acid (DMSA) are antioxidants with thiol groups. All of them were found to counteract the TCHQ-induced cytotoxicity. No protective ability was found in lipoic acid, oxidized form of DHLA, indicated that the protection comes from the –SH group which could bond with ROS and reduces the ROS-mediated DNA damage. This study also demonstrates that the comet assay is a feasible method for measuring DNA damage induced by TCHQ. Using this method, we found that captopril provided marked protection against genotocixity induced by TCHQ.