Abstract:

In previous studies from our laboratory we reported the presence in highly purified liver nuclei, free of contamination with other organelles, of an ethanol metabolizing system (NEMS) able to lead to acetaldehyde and 1-hydroxyethyl free radicals (1HEt). In the present study we tested whether this NEMS is inducible by chronic alcohol administration to rats and whether these nuclei also have increased ability to bioactivate N-nitrosodimethylamine (NDMA). Sprague Dawley male rats (125-150g) were fed with a nutritionally adequate liquid diet containing alcohol to provide 36% of total energy (standard Lieber-De Carli rat diet), for 28 days. Controls received an isocaloric diet without alcohol. Animals were sacrificed, livers were excised and microsomes and purified nuclear fractions were prepared. Both microsomes and nuclei from treated animals had significantly increased ability compared to controls, to biotransform ethanol to acetaldehyde using NADPH as cofactor under an air atmosphere. Both organelles also exhibited significantly increased capacity compared to controls, to bioactivate NDMA to formaldehyde and to reactive metabolites that bind covalently to proteins. Nuclear preparations from control animals were also able to metabolize NDMA to formaldehyde and reactive metabolites. Results indicate that liver nuclei may have a CYP2E1 able to bioactivate both NDMA and EtOH and that these processes are being induced by chronic alcohol drinking. The bioactivation of these xenobiotics to reactive metabolites in the neighborhood of nuclear proteins and DNA might have significant toxicological implications.

Author affiliation: Diaz Gomez, Maria Isabel. Consejo Nacional de Investigaciones Científicas y Técnicas. GP. CITEFA - Centro de Investigaciones Toxicológicas (I); Argentina

Author affiliation: Valles, E.. Consejo Nacional de Investigaciones Científicas y Técnicas. GP. CITEFA - Centro de Investigaciones Toxicológicas (I); Argentina

Author affiliation: Fanelli, Silvia Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. GP. CITEFA - Centro de Investigaciones Toxicológicas (I); Argentina

Author affiliation: Delgado, Aurora Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. GP. CITEFA - Centro de Investigaciones Toxicológicas (I); Argentina

Author affiliation: Castro, Gerardo Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas. GP. CITEFA - Centro de Investigaciones Toxicológicas (I); Argentina

Author affiliation: Castro, Jose Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. GP. CITEFA - Centro de Investigaciones Toxicológicas (I); Argentina

Abstract:

Liver microsomal (mic); nuclei (N) and mitochondria (mit) anaerobically nitroreduce Nifurtimox (Nfx) in the presence of NADPH generating system. Simultaneous formation of small amounts of nitrite was observed in microsomes and nuclei but not in mitochondria. The microsomal nitroreductase activity was enhanced by the presence of flavine-adenine-dinucleotide disodium salt (FAD), was not inhibited by CO and was significantly inhibited by diphenyleneiodonium (DPI). In the microsomal NADPH-dependent fraction nitrite formation was null in the presence of FAD, DPI and under air and was partially inhibited by pure CO. Pure human cytochrome P450 reductase in the presence of NADPH significantly nitroreduced Nfx and produced small amounts of nitrite. The nitroreductive process was significantly enhanced by FAD but the nitrite formation became null. FAD itself was able to chemically nitroreduce Nfx without production of nitrite. NADPH generating system enhanced the FAD nitroreductive effect and led to small production of nitrite. Formation of reactive metabolites and nitric oxide during Nfx metabolism might contribute to its toxicity.

Author affiliation: Montalto, Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. GP. CITEFA - Centro de Investigaciones Toxicológicas (I); Argentina

Author affiliation: Diaz, Edith Graciela. Consejo Nacional de Investigaciones Científicas y Técnicas. GP. CITEFA - Centro de Investigaciones Toxicológicas (I); Argentina

Author affiliation: Castro, Jose Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. GP. CITEFA - Centro de Investigaciones Toxicológicas (I); Argentina