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背景:大鼠急性脑缺血再灌注损伤后有细胞凋亡及凋亡相关基因的表达。目的:观察神经节苷脂对大鼠脑缺血再灌注损伤后细胞凋亡的影响。设计:随机对照动物实验。单位:吉林大学中日联谊医院神经内科。材料:实验于2002-04在吉林大学中日联谊医院动物实验室完成。健康雄性Wistar大鼠48只,鼠龄三四个月,体质量(220±50)g。大鼠随机分为缺血再灌注组和缺血再灌注+给药组(缺血前30min腹腔注射神经节甘脂GM-1),24只/组。其中又根据再灌注时间不同,每组分别分为3个时相点,即3h、6h和24h点,每个时相点8只大鼠。方法:①建立大鼠全脑缺血再灌注模型。②应用二苯胺法测定大鼠脑缺血后3h、6h、24h脑组织DNA裂解率变化,取大脑皮质100mg加0.9mL裂解液制成10%匀浆,加入离心管中,反复冻融,收集上清和沉淀,DNA裂解率=上清吸收值/(上清吸收值+沉淀吸收值)。③免疫组织化学方法观察蛋白激酶Cδ表达,以及神经节苷脂给药后的变化。主要观察指标:大鼠脑皮质DNA裂解率的变化及蛋白激酶Cδ表达强度的变化。结果:实验过程中盐水对照组再灌注6h时死亡1只,麻醉过量死亡,再灌注24h时死亡2只,分别予以补充。随着再灌注时间的延长,DNA裂解率变化明显增加,24h达高峰,蛋白激酶Cδ表达于再灌注6h达高峰,以后逐渐呈下降趋势;神经节苷脂组相应时间点的DNA裂解率及蛋白激酶Cδ表达明显减少。结论:神经节苷脂能抑制脑缺血再灌注后细胞凋亡的发生,并减少蛋白激酶Cδ的表达,对脑缺血再灌注损伤具有明显的保护作用。
BACKGROUND: Apoptosis and expression of apoptosis-related genes after acute cerebral ischemia-reperfusion injury in rats. Objective: To observe the effect of gangliosides on cell apoptosis after cerebral ischemia-reperfusion injury in rats. Design: Randomized controlled animal experiments. Unit: Sino-Japanese Friendship Hospital, Jilin University, Department of Neurology. Materials: The experiment was performed in Animal Laboratory of Sino-Japanese Friendship Hospital, Jilin University from April to April in 2002. Forty-eight healthy male Wistar rats were aged three to four months with a body weight of (220 ± 50) g. Rats were randomly divided into ischemia-reperfusion group and ischemia-reperfusion + administration group (intraperitoneal injection of ganglioglycan GM-1 30min before ischemia), 24 rats / group. Among them, according to different reperfusion time, each group was divided into three time points, namely 3h, 6h and 24h, each time point 8 rats. Methods: ① Establish a rat model of global cerebral ischemia-reperfusion. ② The diphenylamine method was used to determine the changes of DNA cleavage in brain tissue at 3h, 6h, 24h after cerebral ischemia. The cerebral cortex was treated with 100mL of 0.9% lysate and 10% homogenate, centrifuged and centrifuged repeatedly to freeze Supernatant and pellet, DNA cleavage rate = supernatant absorbance / (supernatant absorbance + sediment absorbance). ③ Immunohistochemistry was used to observe the expression of protein kinase Cδ, and the change of ganglioside administration. MAIN OUTCOME MEASURES: Changes of DNA catabolism in rat cerebral cortex and changes of protein kinase Cδ expression intensity. Results: During the experiment, one saline control group died at 6 hours after reperfusion, died of excessive anesthesia, and died at 24 hours after reperfusion. With the extension of reperfusion time, the change of DNA cleavage rate increased significantly, reaching the peak at 24h and the peak of protein kinase Cδ reached its peak at 6h after reperfusion, and then decreased gradually. The rate of DNA cleavage and protein cleavage at the corresponding time point Kinase Cδ expression was significantly reduced. CONCLUSION: Gangliosides can inhibit apoptosis after cerebral ischemia-reperfusion and decrease the expression of protein kinase Cδ, and have a significant protective effect on cerebral ischemia-reperfusion injury.