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

NASH机制研究-活性氧和线粒体功能-

基本信息

批准号：
18590714
负责人：
OHSHIMA Shigetoshi
金额：
$ 2.57万
依托单位：
Akita University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18590714/
关键词：
NASH Pten gene reactive oxygen speies steatosis inflammation therapeutic agent Pten遺伝ミトコンドリア

项目摘要

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

我们在小鼠(Pten KO小鼠)中产生了肝细胞特异性的Pten零突变，并建立了这些小鼠作为人NASH的模型。在体外实验中，我们向过夜培养的Pten KO肝细胞中加入了过氧化氢。虽然肝细胞存活率在暴露于过氧化氢后1h下降到20%，但预先孵育NAC可使肝细胞存活率提高到约80%。虽然过氧化氢暴露1h后，肝细胞内ROS水平较过氧化氢对照组升高约3倍，但预先孵育NAC可使ROS下降至与过氧化氢对照组相同的水平。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)、改善脂代谢或抗氧化的药物对NASH是否有效，我们在断奶后立即给予Pten缺陷小鼠70周。在40和70…随着治疗时间的延长，脂肪性肝炎和肝肿瘤的大体及光镜下表现均有改善。分别于10周和40周进行生化分析、肝脏脂类和脂肪酸组成的定量分析、血清活性氧(ROS)测定以及肝脏SREBP1c等与ROS生成或清除相关的分子的表达。此外，通过Western印迹分析ERK和Akt的磷酸化。NAC通过降低ROS而改善肝炎。EPA和UDCA通过减少SREBP1c的表达改善脂肪变性。此外，它们还通过失活ERK和改变硬脂酸/油酸的比例来减少肝肿瘤的发生。EPA和UDCA还通过消除ROS来减少肝炎。此外，EPA还通过增加AMPKα1的表达来调控SREBP1c的表达。结论：NAC、EPA、UDCA的作用均与ROS的清除有关，只是作用点不同。因此，我们认为这些药物的联合治疗对人类NASH是有效的。较少