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Signal transduction mechanism of cell death-role of kinase cascade.

细胞死亡的信号转导机制——激酶级联的作用。

基本信息

批准号：
07670706
负责人：
MUTOH Tatsuro
金额：
$ 1.6万
依托单位：
Fukui Medical School
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

项目摘要

To understand molecular mechanism of many neurodegenerative disorders, we should elucidate how neurons and muscle cells die and the signals for cell death in such diseases. Accumulating evidences have suggested that apoptotic cell death can occur in patients' brain from some neurodegenerative disorders. We have been involved in the study of the intracellular signal transduction pathway of apoptotic cell death in terms of intracellular protein kinase casccade. In this project, we tried to develope the model system where cells die through apoptotic mechanism in neuronal culture systems and examine the intracellular signal transduction pathway of apoptotic cell death. For these purposes, we found that L6 myocytes are killed by HMG-CoA reductase inhibitor (HCRI), widely used medicine for the treatment of hypercholesteroraemia, involving the induction of apoptotic mechanism. Hydrophobic derivative of HCRIs, simvastatin kills L myocytes at the concentration of 30 ug/ml and causes internucleo … More somal DNA fragmentation and tyrosine phosphorylation reaction of several cellular proteins. Herbimycin A or genistein, a potent inhibitor of protein tyrosine kinase activity prevented simvastatin-induced apoptotic cell death. In the next step, we tried to identify the protein which got tyrosine phosphorylated in response to simvastatin treatment and succeeded in identifying one of such proteins. Phospholipase C-gamma 1 was found to be tyrosine-phosphorylated in simvastatin-treated cells. Tyrosien phosphorylation of PLC-gamma1 was evident as early as 2 min after addition of simvastatin into culture medium and reached maximum at 10 min after its addition. Heretofore, PLC-gamma1 is activated when it gets tyrosine-phosphorylated, although tyrosine phosphorylation is not the only way for the activation. Therefore, we tried to check whether simvastatin induces PI turnove in simvastatin treated cells. Intracellular concentration of IP3 increased up to 3-fold than the basal level at 10 min after addition of simvastatin. U73122, a potent inhibitor of PLC,clear inhibited simvastatin-induced cell death. These results indicate the special role of tyrosine phosphoryaltion and PLC activity in apoptotic cell death of muscle cells. Less

为了了解许多神经退行性疾病的分子机制，我们应该阐明这些疾病中神经元和肌肉细胞是如何死亡的以及细胞死亡的信号。越来越多的证据表明，某些神经退行性疾病患者的脑组织中可发生凋亡性细胞死亡。我们参与了细胞内蛋白激酶级联的凋亡细胞死亡的细胞内信号转导途径的研究。本研究试图建立神经元培养系统中细胞通过凋亡机制死亡的模型系统，并研究凋亡性细胞死亡的细胞内信号转导途径。为了这些目的，我们发现L 6心肌细胞被HMG-CoA还原酶抑制剂（HCRI）杀死，HCRI是广泛用于治疗高胆固醇血症的药物，涉及诱导凋亡机制。辛伐他汀是HCRIs的疏水衍生物，在30 μ g/ml浓度下杀死L型心肌细胞，并引起核内 ...更多信息体DNA断裂和几种细胞蛋白酪氨酸磷酸化反应。除草霉素A或染料木黄酮，一种有效的蛋白酪氨酸激酶活性抑制剂，阻止辛伐他汀诱导的细胞凋亡。在下一步中，我们试图鉴定在辛伐他汀治疗后酪氨酸磷酸化的蛋白质，并成功地鉴定了这样的蛋白质之一。发现磷脂酶C-γ 1在辛伐他汀处理的细胞中被酪氨酸磷酸化。在培养基中加入辛伐他汀后2 min，PLC-γ 1的酪氨酸磷酸化明显，并在加入辛伐他汀后10 min达到最大值。因此，PLC-γ 1在酪氨酸磷酸化时被激活，尽管酪氨酸磷酸化不是激活的唯一途径。因此，我们试图检查辛伐他汀是否在辛伐他汀处理的细胞中诱导PI翻转。在加入辛伐他汀后10分钟，细胞内IP 3浓度比基础水平增加了3倍。U 73122是一种有效的PLC抑制剂，明显抑制辛伐他汀诱导的细胞死亡。这些结果表明酪氨酸磷酸化和PLC活性在肌细胞凋亡中的特殊作用。少