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. ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Intramural Research (Z01)
Project #
1Z01ES101589-04
Application #
7327217
Study Section
(LPC)
Project Start
Project End
Budget Start
Budget End
Support Year
4
Fiscal Year
2006
Total Cost
Indirect Cost
Name
U.S. National Inst of Environ Hlth Scis
Department
Type
DUNS #
City
State
Country
United States
Zip Code
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