Molecular mechanism of inhaled anesthetic-induced hepatotoxicity

我们在体内和体外研究了吸入麻醉剂引起肝毒性的分子机制。苯巴比妥预处理的大鼠暴露在氟烷低氧环境中，细胞内游离血红素浓度迅速升高，随后肝脏微粒体细胞色素P450(CYP)含量显著降低。血红素加氧酶-1(HO-1)也有明显的诱导作用。氯化血红素预处理不仅能诱导肝脏HO-1，而且几乎完全消除了氟烷所致的肝毒性。这些结果表明，氟烷所致的肝毒性是由于肝脏中作为一种强有力的促氧化剂的游离血红素浓度的增加所致，而HO-1的诱导是对这种变化的重要保护性反应。接下来，我们研究了在低氧条件下，先用苯巴比妥处理的大鼠肝脏中HSP70和HO-1的诱导，然后再暴露于异氟烷或氟烷。氟烷低氧处理…诱导热休克蛋白70的表达More高于异氟醚低氧处理。血清丙氨酸氨基转移酶(ALT)活性与小叶中心坏死程度呈正相关，与HSP70升高的变化相似。相反，HO-1只被氟烷低氧处理诱导，而不被其他处理诱导。这些结果表明，氟烷低氧和异氟烷低氧对肝脏的损伤、HO-1和HSP70的诱导有显著差异。已知异氟醚比氟烷更安全，这可能部分是由于异氟烷的氧化应激较小，热休克蛋白的诱导比氟烷治疗要小。最后，由于吸入麻醉药是由细胞色素P450-2E1(细胞色素P450-2E1)代谢的，而吸入麻醉药是四氯化碳的衍生物，因此我们测定了CCl_4对表达细胞色素P450-2E1的肝细胞系(HLE/2E1)和母细胞系(HLE)的细胞毒作用。此外，还检测了CCl_4对氧化应激的潜在标志物HSP70基因表达的影响。与HLE细胞相比，CCl4作用后HLE/2E1细胞的存活率明显降低。Northern印迹分析显示，经CCl_4处理后，两种细胞HSP70mRNA水平均显著升高，但HLE/2E1细胞HSP70mRNA水平的增加幅度远大于HLE细胞。以上结果提示，在高表达细胞中，氧化应激在CCl_4的细胞毒性增强中起重要作用，高表达的人肝细胞株可能适用于研究挥发性麻醉药的肝毒性。较少

We investigated the molecular mechanism of inhaled anesthetic-induced hepatotoxicity in vivo and in vitro. Exposure of phenobarbital-pretreated rats to halothane-hypoxia caused a rapid increase in cytosolic free heme concentration, which was preceded by a significant decrease in microsomal cytochrome P450 (CYP) content in the liver. There were also marked induction in heme oxygenase-1 (HO-1). hemin pretreatment of these animals not only induced hepatic HO-1 , but also almost completely abrogated the halothane-induced hepatotoxicity. These findings indicate that halothane-induced hepatotoxicity is due to an increase in hepatic free heme concentration that is a potent prooxidant, and HO-1 induction is an important protective response against such changes. Next, we examined the induction of HSP70 and HO-1, in rat livers pretreated with phenobarbital, followed by exposure to isoflurane, or halothane under hypoxic condition. The induction of HSP70 was observed by halothane-hypoxia treatment … More is higher than that by isoflurane-hypoxia treatment. Serum alanine aminotransferase (ALT) activity correlated well with the extent of centrilobular necrosis, showed similar changes with increases in HSP70. In contrast, HO-1 was induced only by halothane-hypoxia treatment, but not by other treatments. These findings demonstrate that there is a significant difference in hepatic injury, HO-1 and HSP70 induction, between halothane-hypoxia and isoflurane-hypoxia. Isoflurane is known to be safer than halothane, which may be in part accounted for by its lesser oxidative stress as assessed by a smaller induction of HSPs than halothane treatment. Finally, since inhaled anesthetics are metabolized by cytochrome P450-2E1 (CYP2E1) and inhaled anesthetics are derivates of carbon tetrachloride, the cytotoxic effects of CCl_4 in a liver cell line expressing CYP2E1 (HLE/2E1) in comparison to those in the mother cell line (HLE) were determined. The effects of CCl_4 on the gene expression of HSP70, a potential marker of oxidative stress, were also examined. The viability of HLE/2E1 cells after exposure to CCl_4 was significantly decreased compared with that of HLE cells. Northern blot analysis revealed that the HSP70 mRNA level was significantly increased after CCl_4 treatment in both cell lines, while the magnitude of its increase was much greater in HLE/2E1 cells than HLE cells. These results suggest that the oxidative stress induced by CYP2E1 plays an important role in the increase in cytotoxicity of CCl_4 in CYP2E1-overexpressing cells and that CYP2E1-overexpressing human liver cell line may be suitable for investigating the hepatotoxicity of volatile anesthetics. Less