The study of NASH mechanism-in terms of reactive oxygen species and mitochondrial function-

我们在小鼠（Pten KO小鼠）中产生了Pten的肝细胞特异性零突变，并将这些小鼠建立为人类NASH的模型。体外实验，我们将过氧化氢加入到过夜培养的Pten KO肝细胞中。虽然暴露于过氧化氢1h后肝细胞活力下降至20%，但NAC预孵育可将肝细胞活力提高至约80%。虽然过氧化氢暴露1h后肝细胞ROS水平较过氧化氢未暴露组升高约3倍，但NAC预孵育使ROS下降至与过氧化氢未暴露组相同的水平。NAC对线粒体损伤有抑制作用。在体内实验中，为了测试n-乙酰半胱氨酸（NAC）、二十碳五烯酸（EPA）、熊去氧胆酸（UDCA）、改善脂质代谢或抗氧剂对NASH是否有效，我们在断奶后给pten缺陷小鼠服用了70周。在40和70多周时，通过肉眼和显微镜观察到脂肪性肝炎和肝肿瘤的改善。在第10周和第40周进行生化分析，定量分析肝脏中所含的脂质和脂肪酸组成，血清活性氧（ROS）和肝脏中与脂肪生成或消除ROS相关的分子如SREBP1c的表达。Western blot检测ERK和Akt的磷酸化水平。NAC通过降低ROS改善肝炎。EPA和UDCA通过降低SREBP1c的表达改善脂肪变性。此外，他们通过灭活ERK和改变硬脂酸与油酸的比例来减少肝肿瘤的发生。EPA和UDCA也通过消除ROS来减轻肝炎。此外，EPA通过增加AMPK α 1调控SREBP1c的表达。我们得出NAC、EPA、UDCA的作用均与ROS的消除有关，但作用点不同。因此，我们认为这些药物的混合治疗对人类NASH是有效的。少

We generated a hepatocyte-specific null mutation of Pten in mice (Pten KO mice) and established these mice as a model of human NASH.In vitro experiment, we added hydrogen peroxide to overnight cultured Pten KO hepatocytes. Although hepatocytes viability decreased to 20% 1h after exposure to hydrogen peroxide, preincubation of NAC improved hepatocytes viability to about 80%. Although ROS level of hepatocytes increased about 3 times over compared to that of hydrogen peroxide-unexposured group 1h after exposure to hydrogen peroxide, preincubation of NAC decreased ROS to the same level as that of hydrogen peroxide-unexposured group. Moreover mitochondrial injury were suppressed after addition of NAC.In vivo experiments, to test whether N-acetyl-cystein (NAC), eicosapentaenoic acid (EPA), Ursodeoxycholic acid (UDCA), agents for improving lipid metabolism or for anti oxygen are effective for NASH, we have administered them to Pten-deficient mice for 70 weeks just after weaning. At 40 and 70 … More weeks, improvement of steatohepatitis and hepatic tumor were observed by macroscopic and microscopic findings. At 10 and 40 weeks, biochemical analysis, the quantitative analysis of lipids and fatty acids composition contained in the liver, serum reactive oxygen species (ROS) and the hepatic expression of molecules related with lipogenesis or elimination of ROS such as SREBP1c were examined. Moreover phosphorylation of ERK and Akt were performed by Western blot analysis.NAC improved hepatitis by reducing ROS. EPA and UDCA improved steatosis by decreasing expression of SREBP1c. Moreover they reduced onset of hepatic tumor by inactivating ERK and change of the ratio of stearic acid to oleic acid. EPA and UDCA also reduced hepatitis by elimination of ROS. Moreover EPA controls expression of SREBP1c by increasing of AMPK α 1.We concluded the effects of NAC, EPA, UDCA are all related with ROS elimination however the effective points are different. Accordingly, we propose mixed therapy of these agents are effective of human NASH. Less