Role of Cyt P450-Mediated Oxidation in Chemical-Induced

Cyt P450 介导的氧化在化学诱导的氧化中的作用

• 批准号: 7327217
• 负责人: BURHAN I GHANAYEM
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7327217
• 关键词: Role; Cyt; P450; Mediated; Oxidation

Human exposure to environmental chemicals is considered a contributing factor to increased susceptibility to chemical induced toxicity, genotoxicity, and carcinogenicity. The mechanism(s) of action of these chemicals are not well understood, however, a great number of them are metabolized to epoxide intermediates via CYP2E1. We therefore hypothesized that epoxidation of these chemicals is a pre-requisite for the induction of toxicity, genotoxicity, and carcinogenicity. Using trichloroethylene (TCE) as a model chemical, studies were undertaken to investigate the metabolic and molecular basis for the induction of lung damage by epoxide-forming chemicals. Trichloroethylene (TCE) elicits lung cytotoxicity that selectively damages bronchiolar Clara cells. Current study was undertaken to test the hypothesis that bronchiolar damage is associated with TCE bioactivation by CYP2E1 and CYP2F2 within the Clara cells. Histopathology confirmed dose-dependent Clara cell injury and disintegration of the bronchiolar epithelium in mice treated with TCE doses of 200 to 1000 mg/kg. Immunohistochemical studies, using an antibody that recognizes dichloroacetyl lysine adducts, revealed dose-dependent formation of adducts in the bronchiolar epithelium. The localization of dichloroacetyl adducts in the Clara cells coincided with bronchiolar damage in the lungs of TCE-treated mice. Pretreatment of mice with diallyl sulfone, an inhibitor of CYP2E1 and CYP2F2, abrogated formation of the dichloroacetyl protein adducts and protected against TCE-induced bronchiolar damage. Treatment of wild-type and Cyp2e1-/- mice also induced bronchiolar damage that correlated with the presence of adducts in the bronchiolar epithelium. Immunoblotting, using lung microsomes from TCE-treated mice, showed dose-dependent production of dichloroacetyl adducts that co-migrated with CYP2E1 and CYP2F2. However, TCE treatment resulted in loss of CYP2E1 and CYP2F2 proteins and p-nitrophenol hydroxylation, a catalytic activity associated with both P450 enzymes. The TCE metabolite, chloral hydrate, was formed in incubations of TCE with lung microsomes from CD-1, wild-type and Cyp2e1-/- mice. Levels of chloral hydrate were higher in CD-1 than in either wild-type or Cyp2e1-/- mice; no difference was detected between the wild-type and Cyp2e1-/- mice. These findings indicated that dichloroacetyl adducts formed within the Clara cells and involving CYP2F2 and CYP2E1 are associated with TCE-induced bronchiolar cytotoxicity. Public Health or Environmental Health Significance: Cytochrome P450 2E1 (CYP2E1) is responsible for the bioactivation of a wide variety of environmentally important xenobiotics including 1,3-butadiene, acetaldehyde, acetaminophen, aniline, benzene, carbon tetrachloride, trichloroethylene, dichloroethylene, ethylene glycol, vinyl chloride, acrylonitrile, and nitrosamines. Most of these chemicals undergo oxidative metabolism via the cytochrome P450 enzymes to form epoxide intermediates that are thought to play a role in increased human susceptibilities to various diseases. Epoxides are three membered cyclic ethers and are among the most potent known acute toxicants, mutagens, reproductive toxins, and carcinogens. Environmental chemicals that are metabolized to epoxides share a similar pattern of activity. Further, genetic polymorphisms in CYP2E1 were considered important risk factors in the development of xenobiotic-induced diseases in humans. Once the mechanism of action of epoxides is established, the present approach may be used to predict the effects of chemicals that are metabolized to epoxides. Further, once the role of epoxides in environmentally caused human diseases is established, a more accurate assessment of human risks to epoxide-forming chemicals may be accomplished and the connection between metabolism of chemicals,toxicity, and CYP polymorphisms in humans may be established. CYP2E1 polymorphisms and variability in CYP2E1 activity associated with, for example: diabetes, obesity, starvation, and alcohol consumption, may result in increased metabolic efficiencies leading to differential susceptibilities to the epoxide-forming chemicals in humans. In addition, many of the model chemicals that we use in our research are of interest to the National Toxicology Program and therefore our work will have the additional advantage of complementing the National Toxicology Program mission. Finally, current work suggest that CYP-null mice are an appropriate model for the investigation of the role of oxidative metabolism in the toxicity, mutagenicity, and carcinogenicity of environmental chemicals.

人类对环境化学物质的暴露被认为是增加对化学诱导毒性，遗传毒性和致癌性的易感性的促成因素。这些化学物质的作用机制尚不清楚，但是，其中大量通过CYP2E1代谢为环氧中间体。因此，我们假设这些化学物质的环氧化是诱导毒性，遗传毒性和致癌性的先决条件。使用三氯乙烯（TCE）作为模型化学物质，进行了研究，以研究环氧化学化学物质诱导肺损伤的代谢和分子基础。 三氯乙烯（TCE）引发有选择地损害支气管clara细胞的肺细胞毒性。进行了当前的研究，以检验以下假设：支气管损伤与CYP2E1和CYP2F2在Clara细胞内的TCE生物活化有关。组织病理学证实了用200至1000 mg/kg治疗的小鼠中支气管上皮的剂量依赖性克拉拉细胞损伤和细胞支气管上皮的分解。免疫组织化学研究，使用识别二氯乙酰赖氨酸加合物的抗体，揭示了支气管上皮中加合物的剂量依赖性形成。二氯乙酰加合物在克拉拉细胞中的定位与TCE治疗小鼠肺中支气管损伤相吻合。用Dalyl Sulfone（CYP2E1和CYP2F2的抑制剂）对小鼠进行预处理，废除了二氯乙酰蛋白加合物的形成，并保护了TCE诱导的支气管损伤。野生型和CYP2E1 - / - 小鼠的治疗还诱导了支气管损伤，与支气管上皮中的加合物有关。使用来自TCE处理的小鼠的肺微粒体的免疫印迹，显示出与CYP2E1和CYP2F2共同迁移的二氯乙酰化加合物的剂量依赖性产生。然而，TCE处理导致CYP2E1和CYP2F2蛋白的损失以及p-硝基苯酚羟基化，这是两种P450酶有关的催化活性。 TCE代谢产物氯水合物在与CD-1，野生型和CYP2E1 - / - 小鼠的肺微粒体中孵育中形成。 CD-1的氯氢水合水平高于野生型或CYP2E1 - / - 小鼠的水平；在野生型和CYP2E1 - / - 小鼠之间未检测到差异。这些发现表明，在克拉拉细胞中形成的二氯乙酰基合，涉及CYP2F2和CYP2E1，与TCE诱导的支气管细胞毒性有关。 公共卫生或环境健康意义：细胞色素p450 2e1（CYP2E1）负责对各种各样的环境重要的异种生物学的生物活化，包括1,3-丁二烯，乙醛，乙酰氨基酚，乙酰氨基酚，苯胺，苯胺，苯，苯，碳氯乙烯，三氯乙烯，二氯乙烯，二氯乙烯，二氯乙烯，氯化物，丙烯腈和亚硝胺。这些化学物质中的大多数通过细胞色素P450酶进行氧化代谢，以形成环氧中间体，这些中间体被认为在增加人类对各种疾病的敏感性中发挥作用。环氧化物是三个成员的环状醚，是最有效的急性毒物，诱变剂，生殖毒素和致癌物之一。代谢为环氧化物的环境化学物质具有相似的活性模式。此外，CYP2E1中的遗传多态性被认为是人类异生物诱导疾病发展的重要危险因素。一旦建立了环氧化物的作用机理，目前的方法可用于预测被代谢为环氧化物的化学物质的作用。此外，一旦建立了环境引起的人类疾病的作用，就可以更准确地评估人类对环氧化学物质的风险，并且可以在人类中建立化学物质，毒性和CYP多态性的代谢之间的联系。 CYP2E1多态性和CYP2E1活性的可变性，例如：糖尿病，肥胖，饥饿和饮酒，可能导致代谢效率提高，从而导致人类中环氧化物化学物质的不同敏感性。此外，我们在研究中使用的许多模型化学物质都对国家毒理学计划感兴趣，因此我们的工作将具有补充国家毒理学计划任务的其他优势。最后，目前的工作表明，CYP-NULL小鼠是研究氧化代谢在毒性，诱变性和环境化学物质的致癌性中的作用的合适模型。