Cytosokic Acetyl CoA Hydrolase and Induction by Clofibrate

已知降血脂药物氯苯氧异丁酸乙酯（氯贝特）在大鼠和小鼠长期给药后诱导过氧化物酶体增殖并具有致癌性。我们检查了使用该药物治疗长达18个月对大鼠肝脏中胞浆ATP刺激和ADP抑制的乙酰辅酶A水解酶的影响。在雄性donryu白化病大鼠的饮食含有0.5%的氯贝丁酯，酶活性增加到约2和3倍的初始水平每毫克肝蛋白和胞质蛋白，分别为2倍，每毫克DNA在3天，然后保持在这个水平长达18个月。给氯贝酸3个月的大鼠，在一周内，当氯贝丁酯停止给药时，增加的活性恢复到对照水平。酶活性的变化与酶蛋白量的变化一致，酶蛋白量的变化是通过用任何抗体对从大鼠肝胞质溶胶中纯化的乙酰辅酶A水解物进行免疫印迹来测定的。肉眼未检测到肝脏肿瘤 关于我们 在服用氯贝丁酯18个月后进行镜检。然而，我们的研究结果表明，细胞溶质乙酰辅酶A氢化酶可能是这类药物诱导的过氧化物酶体外标记酶。大鼠肝脏中的线粒体外乙酰辅酶A水解酶[EC 3.1.2.1]，被ATP刺激，被ADP抑制，已知具有极强的冷不稳定性。在低温（2-4 ℃）下进行亚细胞分级分离时，大部分酶活性丧失;然而，大部分酶活性可通过在高浓度磷酸钾存在下于37 ℃下复温而恢复。这使我们能够测量冷处理样品的活性。大部分的ATP-刺激和ADP-抑制的acctyl-CoA水解酶活性在大鼠肝脏中检测到的胞质组分和少量的过氧化物酶体组分中检测到。氯贝酯处理后，过氧化物酶体ATP刺激的乙酰辅酶A水解酶活性没有明显增加。然而，细胞质活性大大增加后，氯贝特治疗。每克肝脏中分离的过氧化物酶体组分的活性约为对照组肝脏胞质组分的5%，约为氯贝酯治疗大鼠的2%。尽管过氧化物酶体组分具有相似的核苷酸（ATP和ADP）敏感性和冷不稳定性，但根据用抗大鼠肝胞质中纯化的乙酰辅酶A水解酶的抗体进行的Western印迹分析，过氧化物酶体组分中的酶蛋白迁移到与胞质中乙酰辅酶A水解酶相同的位置。这些结果提示过氧化物酶体酶和胞浆酶可能是同一种酶。少

The hypolipidemic drug ethyl chlorophenoxyisobutyrate (clofibrate) is known to induce peroxisome proliferation and to ve carcinogenic after long term administration to rats and mice. We examined the effects of treatment with this drug for periods of up to 18 months on cytosolic ATP-stimulated and ADP-inhibited acetyl-CoA hydrolase in rat liver. In male donryu albino rats on a diet containing 0.5% clofibrate, the enzyme activity increased to about 2- and 3-fold the initial level permilligram liver protein and cytosolic protein, respectively, and 2-fold per milligram DNA in 3 days, and then remained at this level for up to 18 months. The increased activity in rats receiving clofivrate for 3 months returned to control level within a week when clofibrate was sithdrown. The change in enzyme activity paralleled the change in the amount of enzyme protein determined by immunoblotting with anyibody against purified acetyl-CoA hydrolasa from rat liver cytosol. No liver tumors were detected macro … More scopically after administration of clofibrate for 18 months. However, our results suggest that cytosolic acetyl-CoA hydrelase could be an extraperoxisomal marker enzyme induced by this type of drug.An extramitochondrial acetyl-CoA hydrolase [EC 3.1.2.1] in the rat liver, which is atimulated by ATP and inhibited by ADP, is known to ve extremely cold-labile. During subcellular fractionations at low temperatures (2-4*C), most of the enxyme activity was lost ; however, most could be recovered by rewarming at 37*C in the presence of a high concentration of potassium phosphate. This enabled us to measure the activities of cold-treated samples. The majority of the ATP-stimulated and ADP-imhibited acctyl-Coa hydrolase activity in rat livers was detected in the cytosolic fraction and small amounts were detected in the peroxixomal fraction. The activity of peroxisomal ATP-stimulated acetyl-CoA hydrolasa was not noticeably increased after clofibrate-treatment. However, the cytosolic activity greatly increased after clofibrate treatment. The activity in the isolated peroxisomal fraction per g of liver was about 5% of that in the cytosolic fraction of liver from the control and about 2% in that from clofibrate-treated rats. Vesides having similar nucleotide (ATP and ADP) sensitivity and cold lability, the enzyme protein in the peroxisomal fraction migrated to the same position as the cytosolic acetyl-CoA hydroiase based on Western blot analysis with antibody against purified acetyl-CoA gydrolase from rat liver cytosol. These results suggest that the peroxisomal enzyme and cytosolic enzyme may be the same extity. Less