Cytotoxicity of 1,2-epoxynaphthalene is correlated with protein binding and in situ glutathione depletion in cytochrome P4501A1 expressing Sf-21 cells
Cytotoxicity of 1,2-epoxynaphthalene is correlated with protein binding and in situ glutathione depletion in cytochrome P4501A1 expressing Sf-21 cells
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DOI：
10.1093/toxsci/53.2.352
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发表时间：
2000-02-01
影响因子：
3.8
通讯作者：
Hammock, BD
中科院分区：
文献类型：
--
作者：
Greene, JF;Zheng, J;Hammock, BD
萘被几种细胞色素P-450（CYP）单加氧酶代谢为1，2-环氧萘。然而，随后的环氧化物与细胞中的大分子的相互作用，以及这些相互作用对细胞损伤的意义，没有得到很好的表征。此外，CYP 1A 1可将萘代谢为1，2-环氧萘，其可被许多异生物质诱导。然而，萘和CYP 1A 1单独之间的原位相互作用，没有其他异生物质代谢酶的影响，还没有被检查。使用能够过表达重组CYP 1A 1的模型真核表达系统，我们发现萘对表达CYP 1A 1的细胞有毒性，剂量为-（LC 50：0.3 mn？）和时间依赖性（LT 50：12 h）方式。萘处理CYP 1A 1表达细胞导致细胞谷胱甘肽（GSH）水平下降47%。用谷胱甘肽乙酯（一种谷胱甘肽类似物）预处理，可保护表达CYP 1A 1的细胞，使其活力为30%，高于单独用萘处理的细胞。细胞毒性与细胞蛋白的共价结合密切相关（r（2）：0.96）。碱性全甲基化技术表明，细胞蛋白质的半胱氨酰-SH基团是环氧化物代谢物的亲核靶点。这些结果表明，在没有其他途径的情况下，萘被CYP 1A 1修饰为1，2-环氧萘，随后结合蛋白质和GSH上的细胞巯基。
Naphthalene is metabolized by several cytochrome P-450 (CYP) monooxygenases to 1,2-epoxynaphthalene. However, the subsequent interactions of the epoxide with macromolecules in the cells, and the significance of these interactions to cellular injury, are not well characterized. Additionally, CYP1A1, which can metabolize naphthalene to 1,2-epoxynaphthalene, may be induced by a number of xenobiotics. Yet, the in situ interaction between naphthalene and CYP1A1 alone, without the influence of other xenobiotic metabolizing enzymes, has not been examined. Using a model eukaryotic expression system capable of over-expressing recombinant CYP1A1, we found that naphthalene was toxic to cells expressing CYP1A1 in a dose- (LC50: 0.3 mn?) and time-dependent (LT50: 12 h) manner. Naphthalene treatment of CYP1A1-expressing cells resulted in a 47% decrease in cellular glutathione (GSH) levels. Pretreatment with ethyl ester GSH, a GSH analog, protected CYP1A1-expressing cells such that viability was 30%, greater than for cells treated with naphthalene alone. Cytotoxicity was strongly correlated (r(2): 0.96) with covalent binding of cellular proteins. Alkaline permethylation techniques showed that cysteinyl-SH groups of cellular proteins are a nucleophilic target of the epoxide metabolite. These results suggest that, in the absence of other pathways, naphthalene is modified by CYP1A1 to 1,2-epoxynaphthalene, which subsequently binds cellular sulfhydryl groups on proteins and GSH.