脳虚血後の神経細胞傷害と傷害抑制に関する研究 -アポトーシスおよび非アポトーシス性傷害の検討-

研究发现脑缺血后迟发性神经元死亡对细胞凋亡有一定的影响。Ras是细胞内信号转导因子之一，被报道为细胞凋亡的抑制剂，通过PI 3-激酶和Akt介导。在这项研究中，我们研究了Ras在迟发性神经元细胞死亡中的作用，基于大鼠前脑缺血模型缺血-再灌注后海马细胞凋亡的存在。实验1.方法：采用雄性Wistar大鼠前脑缺血（15分钟）模型。在缺血状态下，暂时阻断双侧颈动脉，随后进行失血性低血压。HE染色和TUNEL染色进行组织化学研究。结果：缺血再灌注3d后，缺血再灌注3d，缺血 ...更多信息 对照组。α-胞衬蛋白的分解产物显著增加。海马神经元细胞HE染色可见空泡化，核固缩，TUNEL染色阳性。实验2方法;在前脑缺血（10分钟）模型中，在缺血前侧脑室注射手霉素A，并在再灌注后3天处死。采用HE染色和TUNEL法对海马组织进行染色，免疫印迹法检测海马组织中α-胞衬蛋白降解产物的含量。结果：Manumycin A注射组海马神经元损伤明显轻于对照组（P < 0. 05），而Manumycin A注射组海马神经元损伤明显轻于对照组（P < 0. 05）。结论前脑缺血后Ras和Bcl-2的降解可能参与了迟发性神经元死亡的凋亡调控，海马神经元死亡可能是非凋亡性的。少

It has been noticed that delayed neuronal death after brain ischemia has a certain effect on apoptosis. Ras, one of the factors in intracellular signal transduction, is reported as an inhibitor of apoptosis, intermediating by PI 3-kinase and Akt. In this study, we have investigated the role of Ras in delayed neuronal cell death, based on the presence of apoptosis in hippocampus after ischemia-reperfusion by the model of rat forebrain ischemia.#Experiment 1.METHODS; we utilized the forebrain ischemia (15 minutes) model of male Wistar rat. As ischemic condition, temporary occlusion of bilateral carotid arteries was followed by desanguative hypotension. Histochemical study was performed by HE staining and TUNEL staining. In addition, the amounts of Ras, PI 3-kinase, Akt, Bcl-2, and breakdown product of a-fodrin were surveyed by immunoblotting method.RESULTS; significant decreases of the amounts of Ras, PI 3-kinase, Akt, and Bcl-2 were shown 3 days after ischemia-reperfusion comparing to c … More ontrol group. Breakdown product of a-fodrin increased significantly. Neuronal cells in hippocampus revealed vacuolization and pyknosis of nucleus in the brain sample by HE staining, stained-positive in the same position by TUNEL staining.#Experiment 2METHODS; in the forebrain ischemia (10 minutes) model, manumycin A was injected intracerebroventricularly before ischemia, and sacrificed 3 days after reperfusion. The hippocampi were stained by HE staining and TUNEL method, the quantities their breakdown products of a-fodrin were measured by immunoblotting. As the control group, physiological saline or 2% DMSO were injected intracerebroventricularly before ischemia-repersion.RESULTS; manumycin A injected group revealed that the hippocampal neurons injured less than the control group in every methods.#CONCLUSIONSThe degradation of both Ras and Bcl-2 in forebrain ischemia might suggest the regulation of apoptosis in delayed neuronal cell death, the neuronal cell death in hippocampus might be non-apoptotic judging from the result of the experiment 2. Less