RESUMO
Effects of acetaminophen (AAP) pretreatment on hepatic aflatoxin B1 (AFB(1))-DNA binding, cellular proliferation, and AFB(1)-induced glutathione S-transferase placental form (GST-P) positive hepatocytes and foci have been examined in young male rats and hamsters. Intraperitoneal (i.p.) dosing of 600mg AAP 3h before AFB(1) i.p. injection showed three-fold more AFB(1)-DNA binding in hamsters and 40% less binding in rats. Cell proliferation analyzed by bromodeoxyuridine incorporation was not significant (0.4-0.6%) 24-96h after AAP (600mg) treatment of rats; however, proliferation was stimulated and was maximum (11%) in hamsters at 72h after AAP treatment. Dosing of rats with AFB(1) alone at 0.5 or 2.5mg level gave an appreciable number of GST-P positive minifoci (two to nine cells) with a few foci larger than 100 microm; pretreatment with AAP (300 or 600mg) 48h before 0.5 or 2.5mg AFB(1) had no effect on the number and focal area of foci. In hamsters, 1 or 2mg AFB(1) alone yielded GST-P positive hepatocytes without any minifoci. Pretreatment with AAP (600mg) 48 or 72h before 1 or 2mg AFB(1) produced increases in both GST-P positive hepatocytes and minifoci. Thus, marked changes are observed after AAP pretreatment in hamsters compared to rats.
Assuntos
Acetaminofen/farmacologia , Aflatoxina B1/toxicidade , Analgésicos não Narcóticos/farmacologia , DNA/efeitos dos fármacos , Neoplasias Hepáticas/patologia , Animais , Divisão Celular/efeitos dos fármacos , Cricetinae , DNA/metabolismo , Glutationa/metabolismo , Glutationa Transferase/metabolismo , Neoplasias Hepáticas/induzido quimicamente , Neoplasias Hepáticas/enzimologia , Neoplasias Hepáticas/metabolismo , Masculino , Ratos , Ratos Endogâmicos F344RESUMO
A single i.p. dose of aflatoxin B1 (AFB1) (1.0 and 2.0 mg/kg body wt)-induced hepatocarcinogenesis with phenobarbital as a promoter has been examined in young male Fischer rats. Immunohistochemical method has been employed to detect AFB1-induced glutathione S-transferase placental form (GST-P)-positive hepatic foci observed from 3 week and 10 week to 40-48 week periods. With 2.0 mg AFB1 dosing, the number, area and volume occupied by GST-P-positive hepatic foci increased significantly and progressively from 3 week, 10 week and 48 week periods. In long term studies (40-48 weeks), 1.0 mg and 2.0 mg AFB1 dose levels yielded linear response in area and volume occupied by AFB1-induced hepatic foci. Pretreatment of rats with L-buthionine sulfoximine (BSO), a GSH depleter, at a dose of 4 mmol/kg body wt 4 and 2 h before 1.0 or 2.0 mg AFB1 treatment enhanced the number, area and volume of GST-P-positive hepatic foci, increases being the largest at shorter time periods (3 and 10 weeks) compared to longer time periods (40 and 48 weeks). This report appears to be the first example of an enhanced chemical induced hepatocarcinogenesis in a long term study in any experimental animals species by a GSH depleting agent.
Assuntos
Butionina Sulfoximina/farmacologia , Carcinógenos , Inibidores Enzimáticos/farmacologia , Glutationa Transferase/metabolismo , Neoplasias Hepáticas/induzido quimicamente , Aflatoxina B1 , Animais , Sinergismo Farmacológico , Imuno-Histoquímica , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Masculino , Neoplasias Experimentais/patologia , Fenobarbital , Ratos , Ratos Endogâmicos F344 , Fatores de TempoRESUMO
Several studies have demonstrated that green tea (GT) inhibits various chemically induced cancers in experimental animals. In the present study, effect of GT has been examined on the initiation of aflatoxin B1 (AFB1)-induced hepatocarcinogenesis in the rat. Young male Fischer rats were given AIN-76A diet with or without 0.5% instant GT powder in their drinking water for 2 or 4 weeks. Initiation was examined by hepatic AFB1-DNA binding in vivo, AFB1 metabolism in vitro and by the appearance of AFB1-induced glutathione S-transferase placental form (GST-P)-positive hepatocytes detected by immunohistochemical method. There was no influence of GT feeding on microsome-mediated AFB1 binding to exogenous DNA. However, GT feeding enhanced microsome-mediated formation of non-toxic hydroxylated metabolites of AFB1 by 2-3-fold. Hepatic nuclear AFB1-DNA binding in vivo was significantly inhibited by about 20-30% in animals pretreated with GT: AFB1-induced GST-P positive single hepatocytes were inhibited significantly by 60-70% in rats pretreated with GT. These results suggest that feeding of GT inhibits initiation of AFB1-induced hepatocarcinogenesis in the rat by modulation of AFB1 metabolism, thereby inhibiting AFB1-DNA binding and AFB1-induced GST-P-positive hepatocytes.
Assuntos
Aflatoxina B1/antagonistas & inibidores , Aflatoxina B1/toxicidade , Anticarcinógenos/uso terapêutico , Carcinógenos/toxicidade , Glutationa Transferase/metabolismo , Neoplasias Hepáticas Experimentais/induzido quimicamente , Neoplasias Hepáticas Experimentais/prevenção & controle , Chá , Aflatoxina B1/metabolismo , Animais , Carcinógenos/metabolismo , Adutos de DNA/metabolismo , DNA de Neoplasias/efeitos dos fármacos , DNA de Neoplasias/metabolismo , Indução Enzimática/efeitos dos fármacos , Glutationa Transferase/biossíntese , Neoplasias Hepáticas Experimentais/metabolismo , Masculino , Ratos , Ratos Endogâmicos F344RESUMO
Rat is susceptible whereas hamster is resistant to aflatoxin B1 (AFB1) hepatocarcinogenesis. Effect of cell proliferation on AFB1-induced glutathione S-transferase placental form (GST-P) positive foci has been examined in these two species after a single i.p. dose of AFB1 and phenobarbital (PB) as a promoter in a 3 week period. Bromodeoxyuridine incorporation as a measure of cell proliferation and GST-P hepatic foci were analyzed by immunohistochemical methods. Hepatic cell proliferation was maximum at 24 h after either partial hepatectomy (PH) or CCl4 (4 mmol/kg) pretreatment of rats whereas cell proliferation was maximum at 48 h after PH or CCl4 (1 mmol/kg) treatment of hamsters. Enhanced number of GST-P positive hepatic minifoci (two to nine cells) and foci (>100 microns) and focal area were observed in rats with either AFB1 (0.5 mg/kg) given 24 h after PH or AFB1 (0.5 or 2.5 mg/kg) given 48 h after CCl4 dosing. In hamsters, 1 or 2 mg AFB1 treatment produced only GST-P positive single hepatocytes without presence of any minifoci whereas 3 or 6 mg AFB1 produced minifoci consisting only of doublets. Pretreatment with CCl4 48 or 72 h before 1 mg AFB1 dose level increased GST-P positive single cells and minifoci several fold. PH 24 or 48 h before 1 or 2 mg AFB1 dose level increased minifoci. However, increase in minifoci was higher in PH hamsters at 48 h compared with those at 24 h. These results indicate that even though maximum initiation occurs in both speices when AFB1 is administered at the peak of DNA synthesis, rats are more responsive than hamsters to cellular proliferation in the initiation phase of AFB1-induced hepatocarcinogenesis.
Assuntos
Aflatoxina B1/toxicidade , Carcinógenos/toxicidade , Fígado/citologia , Fígado/enzimologia , Animais , Tetracloreto de Carbono/toxicidade , Divisão Celular/efeitos dos fármacos , Divisão Celular/fisiologia , Cricetinae , Sistema Enzimático do Citocromo P-450/metabolismo , Relação Dose-Resposta a Droga , Glutationa Transferase/metabolismo , Hepatectomia , Isoenzimas/metabolismo , Fígado/efeitos dos fármacos , Neoplasias Hepáticas Experimentais/induzido quimicamente , Neoplasias Hepáticas Experimentais/enzimologia , Masculino , Mesocricetus , Ratos , Ratos Endogâmicos F344 , Especificidade da EspécieRESUMO
Glutathione (GSH) conjugation of microsome-mediated and synthetic aflatoxin B1 (AFB1)-epoxide and styrene oxide has been studied with purified glutathione transferases (GSTs) from mouse and hamster liver cytosols. In hamster, with microsomally activated epoxide, the alpha group of GSTs show about 10-fold more activity than the mu group. With the synthetic AFB1 epoxide, the mu enzymes designated H3B and C show considerable activity although less than alpha, whereas H3A and D demonstrate similar ranges of activity as the alpha group. The pi class of GST could not be assayed due to its absence in the hamster liver. The mouse liver cytosols show 3.6-fold greater activity than hamster cytosol in microsome mediated assay system. The mouse alpha and mu enzymes have similar levels of activity in the microsome mediated system; this activity could not be determined with the pi GST due to shortage of this enzyme. The alpha group has 2- and 5-fold higher activity than mu and pi group of GSTs, respectively, with the synthetic epoxide of AFB1. With styrene oxide, the purified GSTs from hamster liver show total loss of activity whereas in the mouse alpha, mu and pi classes of GSTs have similar range of activity as the cytosol. The role of alpha and mu isozymes of GST in rendering these animals resistant to hepatocarcinogenecity is suggested.
Assuntos
Aflatoxina B1/análogos & derivados , Compostos de Epóxi/metabolismo , Glutationa Transferase/metabolismo , Glutationa/metabolismo , Fígado/enzimologia , Microssomos Hepáticos/metabolismo , Aflatoxina B1/biossíntese , Aflatoxina B1/metabolismo , Animais , Cricetinae , Citosol/enzimologia , Glutationa Transferase/isolamento & purificação , Masculino , Mesocricetus , Camundongos , Camundongos Endogâmicos BALB C , Microssomos Hepáticos/enzimologia , Ratos , Ratos Endogâmicos F344 , Sensibilidade e Especificidade , Especificidade da EspécieRESUMO
The effects of pretreatment of buthionine sulfoximine (BSO) alone or in combination with diethylmaleate (DEM) on glutathione (GSH) levels and aflatoxin B1 (AFB1)-DNA binding have been examined in livers and kidneys of young male Fischer rats and Syrian golden hamsters 2 h after an intraperitoneal injection of [3H]AFB1 (0.4 mg/kg body wt.). Animals were treated with BSO (4 mmol/kg body wt.) alone at 4 h and 2 h or with DEM (3 mmol/kg body wt.) at 4 h and BSO at 2 h before AFB1 injection. Hepatic AFB1-DNA binding was about 29.0 and 6.0 pmol/mg DNA in rats and hamsters, respectively. In rats, BSO increased AFB1-DNA binding by about 40% with a drop in GSH by 70%. Treatment with DEM-BSO increased AFB1-DNA binding by about 80% with a concomitant drop in GSH in both species. In hamsters, BSO increased AFB1-DNA binding by only 10% with a 50% drop in GSH. The kidneys of both species have lower GSH levels and AFB1-DNA binding than their respective liver tissues. The effect of BSO alone or in combination with DEM on both GSH levels and AFB1-DNA binding are comparable even though BSO alone is less effective in both species. The role of modulation of GSH levels on AFB1-DNA binding and hence biological effects of AFB1 in these two species is discussed.