The relationship between N-nitrosodimethylamine metabolism and DNA methylation in isolated rat hepatocytes
| Autor: | Barry I. Gold, Lance Encell, Peter G. Foiles |
|---|---|
| Rok vydání: | 1996 |
| Předmět: |
Male
Cancer Research Biology Methylation Dimethylnitrosamine Rats Sprague-Dawley chemistry.chemical_compound N-Nitrosodimethylamine medicine Animals Deoxyguanosine Cytochrome P450 DNA General Medicine Metabolism Molecular biology Rats medicine.anatomical_structure Liver chemistry Biochemistry Hepatocyte DNA methylation Carcinogens Microsome biology.protein |
| Zdroj: | Carcinogenesis. 17:1127-1134 |
| ISSN: | 1460-2180 0143-3334 |
| DOI: | 10.1093/carcin/17.5.1127 |
| Popis: | The metabolism of N-nitrosodimethylamine (NDMA) and its methylation of DNA were simultaneously determined in hepatocytes isolated from untreated and saline- and pyrazole-treated male Sprague-Dawley rats. Metabolism of NDMA was directly measured by monitoring its disappearance via gas chromatography coupled with a sensitive and specific detector for N-nitrosamines. DNA methylation was determined in the same cells employed in the metabolism studies using a monoclonal antibody-based competitive ELISA procedure specific for O6-methyldeoxyguanosine (6-Me-dG). The apparent Km and Vmax for NDMA metabolism are 61 microM and 56 pmol/min/10(6) cells respectively for hepatocytes isolated from untreated rats. It was found that the addition of pyrazole to the in vitro hepatocyte incubations caused a dose-dependent inhibition of both metabolism and DNA methylation. However, when DNA methylation is expressed as a function of NDMA metabolized, there is no significant difference between hepatocyte incubations without or with pyrazole, with an average value of 79 nmol 6-Me-dG/mol dG/nmol NDMA metabolized. Based on the pyrazole inhibition studies, cytochrome P450IIE1 is responsible for at least 60% of the DNA methylation in rat hepatocytes. In pyrazole-pretreated rats there was an inconsistent increase in NDMA metabolism, but when metabolism was elevated so was DNA methylation. In contrast, microsomes isolated from pyrazole-pretreated rats consistently showed elevated metabolism of NDMA. Based on the simultaneous determination of adduct levels and metabolism, there is approximately 1 6-Me-dG adduct formed/133 000 NDMA molecules metabolized in the uninduced hepatocytes. |
| Databáze: | OpenAIRE |
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