ROLE OF OXIDATIVE STRESS IN THE MECHANISM OF HEPATIC DAMAGE

氧化应激在肝损伤机制中的作用

基本信息

批准号：
02454237
负责人：
ISHII Hiromasa
金额：
$ 4.48万
依托单位：
KEIO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1990
资助国家：
日本
起止时间：
1990 至 1991
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-02454237/
关键词：
Oxidative stress Hepatic microcirculation Carbon tetrachloride Ischemic damage Endotoxin Nitric oxide Xanthine oxidase Digital image process NADH自家蛍光 redox state

项目摘要

Oxidative stress in the perfused rat liver was attempted to be analyzed by digital microfluorography using a fluorescent probe for hydroperoxide, dichlorofluorescin (DCFH). DCFH is known to be oxidized by hydorperoxide to dichlorofluorescein (DCF). Hepatic damage was also assessed by propidium iodide (PI), which is known to label the nuclei of non viable cells. Fluorescence of these probes was observed by fluorescence microscope equipped with silicon intensified target camera, and digitally processed by a image processor (ARGUS-200, Hamamatsu photonicus, Japan). Carbon tetrachloride induced oxidative stress was demonstrated in zone 3 by DCF fluorescence, followed by cell damage assessed by PI. These changes were prevented by SKF525A, an inhibitor of cytochrome P450, or promethazine, a radical scavanger. Low flow hypoxia increase DCF fluorescence in zone 2 first, and spread to zone 3. PI fluorescence was observed in zone 3 after increase in DCF fluorescence, suggesting midzonal oxidative stress preceed cell death. These changes were prevented by addition of allopurinol, a xanthine oxidase inhibitor, or PGE1.Rhodamine 123(Rh123), an indicator of mitochondrial energization, was used to assess lipololysaccharide induced liver damage. Addition of LPS to perfusate decreased Rh123 fluorescence, and the change was blocked by LNMA, and in hibitor of nitric oxide synthetase. Mitochondrial dysfunction in hepatocytes induced by LPS was observed only in the presence of Kupffer cell, suggesting that nitric oxide released from activated Kupffer cell mediates hepatic mitochondiral dysfunction in LPS induced liver damage.

尝试使用数字微荧光摄影来分析灌注大鼠肝脏中的氧化应激，并使用荧光氧化物（DCFH）进行荧光探针进行荧光探针。已知DCFH被羟基氧化物氧化为二氯氟乙烯（DCF）。还通过碘化丙啶（PI）评估了肝损伤，该丙二醇丙啶（PI）已知可以标记非活细胞的核。这些探针的荧光是通过配备硅加强目标摄像头的荧光显微镜观察到的，并由图像处理器（Argus-200，Hamamatsu Photonicus，Japan）数字处理。 DCF荧光在第3区证明了四氯化碳诱导的氧化应激，然后通过PI评估的细胞损伤。这些变化是由细胞色素P450的抑制剂SKF525A或根治化的清除剂抑制剂所预防的。低流动性缺氧首先会增加2区2区的DCF荧光，并扩散到3区。在增加DCF荧光后，在区域3中观察到PI荧光，表明区域氧化应激中期预期细胞死亡。这些变化是通过添加别嘌呤醇，黄氨酸氧化酶抑制剂或PGE1.Rhodamine 123（Rh123）的添加（RH123）（线粒体能量的指标）来评估Lipololycarysacachary诱导的肝损伤的。在灌注液中添加LPS会降低RH123荧光，并在LNMA和一氧化氮合成酶的Hibor中阻断了变化。仅在Kupffer细胞的存在下才观察到LPS诱导的肝细胞中的线粒体功能障碍，这表明从活化的kupffer细胞中释放出的一氧化氮会介导LPS在LPS中LPS诱导的肝功能障碍引起的肝功能障碍。