Crotonaldehyde is widespread in the human environment and is a representative enal (Alpha,Beta-unsaturated aldehyde). Enals are relatively reactive compounds which readily undergo the nucleophilic Michael addition reactions. They form conjugates with the thiol group in protein and deplete glutathione in vivo. Crotonaldehyde has been shown to be mutagenic and to modify DNA readily by forming 1,N2-cyclic guanine adducts under physiological conditions. Recently, we have demonstrated for the first time that crotonaldehyde given in drinking water to F344 rats induces liver tumors. The mechanism(s) by which crotonaldehyde induces liver tumorigenesis may be related to its ability to form 1,N2-cyclic guanine adducts in hepatocellular DNA. In this study, the biochemical mechanism(s) of crotonaldehyde tumorigenicity in relationship to DNA adduct formation will be investigated. Various adducts of deoxyguanosine, guanosine and deoxyguanosine-3'-monophosphate with crotonaldehyde and its epoxide will be synthesized as markers. These markers will be used in studying the formation of DNA adducts in rats treated with crotonaldehyde. The three currently most sensitive methods of detection will be used: 1. immunoassay; 2. 32P-postlabeling assay and 3. 3H-postlabeling method. The effect of crotonaldehyde on hepatic glutathione levels and effects of varying glutathione contents on levels of crotonaldehyde-DNA adducts will also be assessed. This research will be further extended to study the ability of 4-hydroxynonenal, a crotonaldehyde homolog, in forming similar DNA adducts. 4-Hydroxynonenal is a major product derived from lipid peroxidation to which humans may be exposed endogenously. The results of this study will provide a basis for assessing its potential tumorigenic effects. Finally, a bioassay for the tumorigenicity of crotonaldehyde using a more defined dose regimen will be performed to better delineate the dose-response relationship of crotonaldehyde tumorigenicity. The results of this program will provide a scientific basis in assessing the possible role of exogenous and endogenous enals in tumorigenesis.
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