Cytotoxicity of Superoxide and Nitric Oxide (NO)

超氧化物和一氧化氮 (NO) 的细胞毒性

基本信息

批准号：
05045052
负责人：
NAKAZAWA Hiroe
金额：
$ 3.2万
依托单位：
Tokai University
依托单位国家：
日本
项目类别：
Grant-in-Aid for international Scientific Research
财政年份：
1993
资助国家：
日本
起止时间：
1993 至 1995
项目状态：
已结题

项目摘要

1. MEASUREMENT OF SUPEROXIDE,NITRIC OXIDE (NO) and ONOO^-The generation of superoxide, NO and ONOO^- from human polymorphonuclear leukocytes were measured. For superoxide cytochrome c reduction and MCLA chemiluminescence methods were used. For NO,end uprodct of NO,nitrite+nitrate was measured by Griess method after reducing nitrite to nitrate. For ONOO^- we established a stable method by modifying the previously reported HPA-NO_2-HPLC method. Using these methods, the roles of these active molecules in various diseases were investigated and we found that the plausible cytotoxic molecules in polymorphonuclear leukocytes-mediated injury appears to be ONOO^- when the target of leukocytes is cardiac myocytes but superoxide is the major cytotoxic molecule when the target of leukocytes is endothelial cells.2. REAL-TIME MEASUREMENT OF SUPEROXIDE,NO and ONOO^-The real-time measurement of above mentioned molecules from isolated organs were also performed to investigate the cytotoxic mechanisms u … More nderlying the ischemia-reperfusion injury. Among several methods applied, we found that the oxymyoglobin method was applicable for that purpose.3. DEMONSTRATION OF ONOO^--MEDIATED INJURYTo demonstrate ONOO^--mediated tissue damage in various diseases such as liver cirrhosis, stomach ulcer and intestinal cancer, nitrotyrosine which is the reaction product of ONOO^- and tyrosine residues of protein was measured. Plasma and tissue concentration of nitrite+nitrate were also measured. The immunohistological staining using nitrotyrosine monoclonal antibodies was also performed for tissue samples obtained at the time of resection or through biopsy. The nitrotyrosine and nitrite+nitrate concentrations were significantly elevated in ulcer tissue. The increase correlated with severity ulcer. In contrast these levels remained low in the center of cancer and slightly elevated in the marginal portion of cancer. The nitrotyrosine staining method gave some positive staining in diseased portion but the specificity was shown to need further improvement. Less

1.超氧化物、一氧化氮(NO)和ONOO^-的测量对于超氧化物细胞色素c还原，使用MCLA化学发光方法测量人多形核白细胞产生的超氧化物、NO和ONOO^-。 NO，亚硝酸盐+硝酸盐是在将亚硝酸盐还原为硝酸盐后通过格里斯法测量的。对于ONOO^-我们通过修改方法建立了稳定的方法。先前报道的HPA-NO_2-HPLC方法，利用这些方法，研究了这些活性分子在各种疾病中的作用，我们发现，当白细胞的靶标出现时，多形核白细胞介导的损伤中可能的细胞毒性分子是ONOO^-。当白细胞的靶标是内皮细胞时，心肌细胞中超氧化物是主要的细胞毒分子。 2．超氧化物、NO和ONOO^的实时测量。还对来自分离器官的上述分子进行了测量，以研究缺血再灌注损伤的细胞毒性机制。在所应用的几种方法中，我们发现氧合肌红蛋白方法适用于该目的。3。 -介导的损伤为了证明ONOO^-介导的各种疾病中的组织损伤，如肝硬化、胃溃疡和肠癌，硝基酪氨酸是ONOO^-的反应产物还测量了切除时或通过活检获得的组织样品的亚硝酸盐+硝酸盐的血浆和组织浓度。溃疡组织中的这种增加与溃疡的严重程度相关，相反，这些水平在癌症的中心仍然较低，而在癌症的边缘部分则略有升高。硝基酪氨酸染色法在病变部分得到了一些阳性染色，但特异性需要进一步改进。