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Metabolism of reactive oxygen species after cerebral ischemia -Study using human SOD1-transgenic mouse-

脑缺血后活性氧的代谢-使用人SOD1-转基因小鼠的研究-

基本信息

批准号：
14571297
负责人：
SUGAWARA Taku
金额：
$ 2.24万
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：
2002
资助国家：
日本
起止时间：
2002 至 2003
项目状态：
已结题

项目摘要

Recent studies have revealed that activation of extracellular signal-regulated kinase (ERK) may contribute to apoptosis, a cell death process involved in oxidative stress. We examined phosphorylation of ERK1/2 and oxidative stress after transient focal cerebral ischemia (FCI) using transgenic (Tg) mice that overexpress copper/zinc superoxide dismutase (SOD1). The mice were subjected to 60min of middle cerebral artery (MCA) occlusion by intraluminal suture blockade followed by 1,4, and 24 hr of reperfusion. Immunohistochemistry and Western blot analysis showed that phospho-ERK1 was markedly increased in the cortex within the MCA territory at 1 hr of reperfusion (p<0.01), followed by a decrease at 24 hr in wild-type mice. Double staining with phospho-ERK1/2 and neuron-specific nuclear protein showed that phospho-ERK1/2 was primarily expressed in neurons. In SOD1 Tg mice, phospho-ERK1/2 was prominently reduced compared with nonischemic controls, shown by immunohistochemistry. Western blot analysis confirmed a significant decrease in phospho-ERK1/2 1 hr after FCI in the ischemic cortex (p<0.005). Apoptotic-related DNA fragmentation was reduced in the ischemic cortex of SOD1 Tg mice compared with wild-type mice using a cell death assay. These results suggest that phosphorylation of ERK1/2 may be involved in apoptosis or cell death after transient FCI and that SOD1 may attenuate apoptotic cell death mediated by the mitogen-activated protein kinase/ERK pathway.

最近的研究表明，细胞外信号调节激酶（ERK）的激活可能导致细胞凋亡，这是一种与氧化应激有关的细胞死亡过程。我们使用过表达铜/锌超氧化物歧化酶（SOD1）的转基因（Tg）小鼠检测了短暂局灶性脑缺血（FCI）后ERK1/2的磷酸化和氧化应激。小鼠经腔内缝合阻断大脑中动脉（MCA） 60分钟后再灌注1、4、24小时。免疫组织化学和Western blot分析显示，在再灌注1小时时，野生型小鼠MCA区域内皮层磷酸化erk1显著升高（p<0.01）， 24小时后下降。phospho-ERK1/2和神经元特异性核蛋白双染色显示，phospho-ERK1/2主要在神经元中表达。免疫组化结果显示，SOD1 Tg小鼠与非缺血对照组相比，磷酸化erk1 /2明显降低。Western blot分析证实缺血皮质FCI后1小时磷酸化- erk1 /2显著降低（p<0.005）。细胞死亡实验表明，与野生型小鼠相比，SOD1 Tg小鼠缺血皮质中凋亡相关的DNA片段减少。这些结果表明，ERK1/2的磷酸化可能参与了短暂FCI后细胞凋亡或死亡，SOD1可能减弱由丝裂原活化蛋白激酶/ERK途径介导的凋亡细胞死亡。