RESUMEN
A V79 Chinese hamster cell line stably expressing human cytochrome P450 1A1 (CYP1A1) was obtained by chromosomal integration of the human CYP1A1 cDNA under the control of the SV40 early promoter. Chromosomal integration was verified by Southern analysis, and effective transcription of the human CYP1A1 cDNA was demonstrated by Northern analysis. The CYP1A1 cDNA-encoded protein was characterized by Western analysis using anti-rat CYP1A1. Intracellular association of CYP1A1 with the endoplasmic reticulum could be visualized by in situ immunofluorescence. Crude cell lysates of the V79 derived cell line was able to catalyze 7-ethoxyresorufin-O-deethylation (EROD) with an activity of about 50 pmol min-1 mg-1 total protein, and an aryl hydrocarbon hydroxylase activity (AHH) of 25 pmol min-1 mg-1. CYP1A1 dependent cytotoxicity, measured by neutral red uptake, and genotoxicity, determined by the frequency of micronucleus formation, of benzo[a]pyrene (B[a]P) and trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene (B[a]P-7,8-diol) could be demonstrated at substrate concentrations as low as 10 nM. Thus, this cell line presents a sensitive tool for studying CYP1A1 mediated metabolism of polycyclic aromatic hydrocarbons (PAH). B[a]P and the purified (+)- and (-)-enantiomers of B[a]P-7,8-diol were compared for their mutagenicity. The (-)-enantiomer was found to be 3-5-fold more mutagenic than the (+)-enantiomer.
Asunto(s)
Benzo(a)pireno/metabolismo , Sistema Enzimático del Citocromo P-450/biosíntesis , ADN Complementario/biosíntesis , Animales , Benzo(a)pireno/toxicidad , Biotransformación , Línea Celular , Cricetinae , Cricetulus , ADN Complementario/efectos de los fármacos , Expresión Génica , Vectores Genéticos , Humanos , Hígado/química , Hígado/efectos de los fármacos , Masculino , Pruebas de Micronúcleos , Ratas , TransfecciónRESUMEN
The microsomal Na+-K+-ATPase of rat brain was inhibited by mercury chloride and methyl mercury. The IC50 was 6.5 X 10(-7) M for mercury chloride and 3.5 X 10(-6) M for methyl mercury. The inhibition was of a non-competitive type with respect to ATP. The non-ionic detergent Lubrol potentiated the inhibitory effect of both mercurials. It is concluded that Lubrol removes the bulk lipids present outside the catalytic center of the enzyme. Consequently, the enzyme will become more sensitive to the inhibition by both mercurials.
Asunto(s)
Encéfalo/enzimología , Cloruro de Mercurio/toxicidad , Compuestos de Metilmercurio/toxicidad , Microsomas/enzimología , Polietilenglicoles/farmacología , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidores , Animales , Encéfalo/ultraestructura , Técnicas In Vitro , Cinética , Lípidos/fisiología , Masculino , Ratas , Sinaptosomas/enzimologíaRESUMEN
Mouse brain synaptosomal Na+/K+-ATPase was inhibited by 23%, 18 hrs after a single intraperitoneal dose of 40 mg lindane (dissolved in olive oil) per kg. The inhibition was of a non-competitive type with respect to ATP. Pretreatment with lindane also potentiated the inhibitory effect of ethanol on this enzyme. It is suggested to consider this interaction at the synaptosomal level when evaluating the anaesthetic effect of ethanol in contaminated persons. Although the synaptosomal Na+/K+-ATPase was inhibited after pretreatment with lindane in vivo, neither lindane nor its metabolites were present in the synaptosomal fraction when determining the subcellular distribution of U-[14C]-lindane in the brain. These results raise some questions regarding the current opinion that lindane exerts some of its central effects through binding to the synaptosomal membrane.
Asunto(s)
Encéfalo/enzimología , Hexaclorociclohexano/farmacología , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidores , Sinaptosomas/enzimología , Animales , Biotransformación , Encéfalo/metabolismo , Encéfalo/ultraestructura , Etanol/farmacología , Hexaclorociclohexano/metabolismo , Inyecciones Intraperitoneales , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
The effect of manganese on free polysomal protein synthesis of immature rat brain (3 weeks old) has been determined after 1, 2, 3, and 4 weeks of daily intake of 55 micrograms manganese/ml of drinking water. The protein synthesis was inhibited up to 35% during the first 3 weeks and returned toward the control level during the fourth week of treatment. Cross-incubation experiments with polysomes and pH 5 enzyme fractions indicated that the inhibition of protein synthesis is due to alteration of the pH 5 enzyme fraction. Furthermore, cerebral t-RNA content was reduced by 20% during the first 3 weeks and also returned to the control level after 4 weeks. The data suggest that the previously reported retardation in learning and memory of manganese treated immature rats may partly be due to alteration of cerebral RNA and protein synthesis. It was also evident that an adaptation mechanism to the observed effect of manganese developes after three weeks of daily intake of 55 micrograms manganese/ml.