Lotlikar P D, Raj H G, Bohm L S, Ho L L, Jhee E C, Tsuji K, Gopalan P
Fels Research Institute, Temple University School of Medicine, Philadelphia, Pennsylvania 19140.
Cancer Res. 1989 Feb 15;49(4):951-7.
The effect of phenobarbital (PB) pretreatment of rats on both hepatic aflatoxin B1 (AFB1)-DNA binding and AFB1-glutathione (AFB1-SG) conjugation have been examined in studies in vivo and in vitro. Male Sprague-Dawley rats fed a commercial diet with 0.1% PB in their drinking water for 1 week had total wet liver weight and microsomal protein content about 27% and 38% higher, respectively, than controls. Hepatic cytochrome P-450 content, microsomal cytochrome P-450 mediated AFB1 binding to exogenous DNA and formation of hydroxy metabolites of AFB1 were also about threefold higher in PB-treated rats and cytosolic reduced glutathione S-transferase activities were about doubled. Microsome-mediated AFB1-DNA binding, when examined at 2 microM and 10 microM levels of AFB1, was inhibited two-to threefold more by cytosols of treated rats whereas AFB1-SG conjugation was two- to threefold higher by cytosols of treated rats. In reconstitution experiments with 2 microM AFB1, with intact nuclei serving as a source of endogenous DNA, addition of microsomes from either group generated a large amount of AFB1-DNA binding (68-105 pmol) and a smaller amount of AFB1-SG conjugate (12-21 pmol). The presence of cytosol from the controls reduced AFB1-DNA binding to a much lesser extent than the cytosol from the treated group whereas AFB1-SG conjugation was much higher with the cytosol from the treated group. These results are in agreement with the studies in vivo. In isolated hepatocytes at 33 nM, 2 microM and 10 microM AFB1 levels, AFB1-DNA binding was decreased 50 to 70% by prior PB-treatment whereas AFB1-SG conjugation was two- to threefold higher in treated compared to control hepatocytes. In hepatocytes, addition of 1 mM diethylmaleate increased DNA binding two- to threefold with a corresponding decrease in AFB1-SG conjugation. Addition of 1 mM styrene oxide caused 5- to 10-fold increases in AFB1-DNA binding at levels of AFB1 of 33 nM and 2 microM; but at 10 microM AFB1, increases in AFB1-DNA binding were two- to threefold. In intact rats, PB treatment reduced hepatic AFB1-DNA binding to 30% of controls with concomitant increase in biliary excretion of AFB1-SG conjugate. It appears that the induced cytosolic GSH S-transferases after PB treatment of rats plays a significant role in inhibiting hepatic AFB1-DNA binding and hepatocarcinogenesis presumably by inactivation of the reactive AFB1-epoxide.(ABSTRACT TRUNCATED AT 400 WORDS)
在体内和体外研究中，均检测了苯巴比妥（PB）预处理大鼠对肝脏黄曲霉毒素B1（AFB1）-DNA结合及AFB1-谷胱甘肽（AFB1-SG）结合的影响。给雄性斯普拉格-道利大鼠饮用含0.1%PB的商业饲料1周，其肝脏总湿重和微粒体蛋白含量分别比对照组高约27%和38%。PB处理组大鼠肝脏细胞色素P-450含量、微粒体细胞色素P-450介导的AFB1与外源DNA的结合以及AFB1羟基代谢产物的形成也约为对照组的三倍，胞质还原型谷胱甘肽S-转移酶活性约增加一倍。当在2 microM和10 microM的AFB1水平检测时，处理组大鼠的胞质溶胶对微粒体介导的AFB1-DNA结合的抑制作用比对照组高2至3倍，而AFB1-SG结合则高2至3倍。在用2 microM AFB1进行的重组实验中，以完整细胞核作为内源性DNA的来源，添加两组中的任何一组微粒体均产生大量的AFB1-DNA结合（68 - 105 pmol）和少量的AFB1-SG结合物（12 - 21 pmol）。对照组胞质溶胶对AFB1-DNA结合的降低程度远小于处理组胞质溶胶，而处理组胞质溶胶的AFB1-SG结合则高得多。这些结果与体内研究一致。在分离的肝细胞中，当AFB1水平为33 nM、2 microM和10 microM时，预先用PB处理可使AFB1-DNA结合减少50%至70%，而处理组肝细胞中的AFB1-SG结合比对照组高2至3倍。在肝细胞中，添加1 mM马来酸二乙酯可使DNA结合增加2至3倍，同时AFB1-SG结合相应减少。添加1 mM环氧苯乙烷可使33 nM和2 microM的AFB1水平下的AFB1-DNA结合增加5至10倍；但在10 microM的AFB1水平下，AFB1-DNA结合增加2至3倍。在完整大鼠中，PB处理可使肝脏AFB1-DNA结合降至对照组的30%，同时胆汁中AFB1-SG结合物的排泄增加。似乎大鼠经PB处理后诱导产生的胞质谷胱甘肽S-转移酶在抑制肝脏AFB1-DNA结合和肝癌发生中起重要作用，可能是通过使活性AFB1-环氧化物失活来实现的。（摘要截短至400字）
