Indole-3-carbinol (I-3-C), a normal dietary constituent, has been shown to decrease chemical carcinogenesis in experimental animals. N-nitrosodimethylamine (NDMA) requires metabolic oxidation in order to express its hepatotoxic, mutagenic and carcinogenic potential. Our results show that treatment of mice by gavage with I-3-C decreases the covalent binding of (14C)NDMA metabolites to DNA or protein up to 90% with no change in the rate of NDMA demethylation of cytochrome P-450 levels. We found that ethanol or methylene chloride extracts of livers from mice treated with I-3-C decreased the covalent binding of NDMA metabolites to DNA and protein in liver cell fractions from untreated mice. In order to explain these observations, we developed the hypothesis that treatment of mice with I-3-C gives rise to lipophilic nucleophiles that protect tissue macromolecules from electrophilic attack by a scavenging mechanism. To test this hypothesis, we developed a procedure to assign a relative value of nucleophilicity to chemicals or chemical mixtures, such as tissue extracts. The nucleophilic index value (NIV) for liver tissue extracts is inversely proportional to the rate of covalent binding of MDMA metabolites to DNA and protein, and independent of the rate of NDMA demethylation.
The Specific Aims of this project are: a) to find the I-3-C dose regimen to maximize tissue NIV; b) to correlate NIV with indices of hepatotoxicity and DNA damage; c) to fractionate solvent extracts of livers from I-3-C treated mice in order to purify and identify the active nucleophilic material(s), using the HPLC separation techniques in parallel with liquid scintillation counting and high resolution mass spectrometry compound identification procedures; d0 to establish the mechanism of protection by I-3-C. The objectives of this research plan are a) to establish and validate a model system which uses the NIV of the liver to predict biochemical and functional susceptibility of the liver to damage from the hepatotoxic and caracinogenic chemical NDMA; b) to identify the chemical species and the mechanisms involved in the I-3-C protection from molecular and cellular damage. This project offers a predictive model for tissue susceptibility to, and a molecular basis for dietary protection from certain environmental toxic and mutagenic/carcinogenic insults.
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