【作者】 张元隆；

【导师】 康德智；

【作者基本信息】 福建医科大学 ， 外科学， 2011， 硕士

【摘要】 目的:建立兔蛛网膜下腔出血脑血管痉挛模型,探讨半胱氨酸天冬氨酸特异性酶切蛋白酶3、8(Caspase 3、8)在蛛网膜下腔出血后不同时间点基底动脉的表达情况及其与脑血管痉挛的关系。方法:1、新西兰大白兔36只,随机分为对照组(n=6)和SAH组(n=30),SAH组再按时间随机分为1、3、5、7、10天5个小组,每个时相点各6只,采用枕大池二次注血的方法建立SAH模型。另取4只兔子备行透射电镜检查,2只对照组,2只实验组(术后第7天)。二次注血后在各时间点采用灌流固定法处死动物,留取基底动脉标本。用光学显微镜观察基底动脉形态学改变,测定横截面积判断有无血管痉挛;用透射电镜观察基底动脉超微结构的形态学改变;2、应用TUNEL法显示血管内皮细胞及平滑肌细胞的凋亡情况,免疫组化方法显示Caspase 3、8在基底动脉中的表达情况。结果:1、实验组与对照组相比,血管壁增厚、内皮皱缩,血管内径和管腔面积明显减少,形态学改变符合脑血管痉挛改变。实验组基底动脉横截面积总体小于对照组,与对照组比较差异显著(P<0.05);透射电镜观察到实验组血管部分内皮细胞脱落,紧密联接打开,内皮间隙扩大;内弹力膜扭曲、不连续;平滑肌细胞变形,胞浆空泡化;而对照组内皮细胞连续,细胞间可见紧密连接,胞质内结构无异常。2、TUNEL法观察到凋亡细胞在实验组第1d出现,第7d凋亡水平达到最高,对照组少见凋亡细胞。免疫组化显示Caspase 3、8在实验组内皮细胞中有大量的表达,与对照组比较有显著性差异(P<0.05)。Caspase3、8在SAH后第1d出现,第3d较第1d增多,第5d和第7d出现强烈表达,第l0 d表达明显减弱。结论:1、采用新西兰大白兔枕大池二次注血法建立SAH后CVS模型是稳定可靠的,能够较好地模拟人类SAH后CVS。2、在兔脑血管痉挛模型的基底动脉中存在细胞凋亡,Caspase 3、8的表达明显增加,具有一定的时间规律性。Caspase 3、8参与了基底动脉内皮细胞凋亡的过程和蛛网膜下腔出血后脑血管痉挛的发生。更多还原

【Abstract】 Objectives:To establish the model of cerebral vasospasm after subarachnoid hemorrhage in rabbits and to study the expression of Caspase 3、8 in the basilar artery after subarachnoid hemorrhage at various time point and the relationship between Caspase 3、8 and cerebral vasospasm .Methods: 1、Thirty six New Zealand rabbits were randomized into 2 groups: control group(n=6) and SAH group (n=30).Then the latter group was randomly divided into 5 groups according to the time:1d,3d,5d,7d,10d (6 per group) . Models of SAH were successfully induced via injecting autologous blood into the cisterna magna twice.Another 4 rabbits were used for the examination of the transmission electron microscope:2 control group and 2 SAH group(D7).The perfusion-fixation was performed for sacrifice of the rabbits in different time after the second blood injection in vivo and the basilar arteries were taken.The morphological changes of the basilar artery were observed under optical microscope and the lumen areas were measured to assess vasospasm in all groups and the ultrastructural changes of BA afte SAH were examined by TEM.2、Immunohistochemistry technique and TUNEL were used to observe the expression of Caspase3、8 and the apoptosis in the basilar artery.Results: 1、Compared with the control group, the basilar arteries in experimental groups demonstrate vascular walls thickening、endothelial cells shrinking, diameters and lumen areas significantly decreasing and the morphological changes according with cerebral vasospasm change.The basilar artery lumen areas were smaller in control group than those in SAH groups and the results were statistically significant(P<0.05). The TEM shows part of the vascular endothelial cells falling off , tight junction opening, endothelial cells gap increasing, the inside elastic membrane distorting and breaking, smooth muscle cells deformation,cytoplasm vacuole changing in experimental groups, while in the control group it reveals the vascular endothelial cells in succession, tight junction undamaged, the structure of the cytoplasm in good condition.2、Apoptosis cells were seen in the SAH groups through TUNEL and they appeared in day 1, reaching the highest in day 7,however,they were rare seen in the control group.There were statistically significant difference about the expression of Caspase3 and Caspase 8 in the basilar arteries between the control group and SAH groups (P<0.05). The expression of Caspase3 and Caspase8 in the basilar arteries appeared in day 1 and augmented in day 3,reaching the highest in day 5 and day 7 and decreasing apparently in day 10 .Conclusion:1、The New Zealand rabbits models that were established by injecting autologous blood into the cisterna magna twice are practical,and are reliable for experimental studies of vasospasm after subarachnoid hemorrhage in human .2、There is apoptosis in the basilar arteries of rabbits from cerebral vasospasm models.The expression of Caspase 3 and Caspase 8 increases apparently and have a certain time regularity . Caspase 3 and Caspase8 play roles in the process of apoptosis of endothelial cells in the basilar artery and in the occurrence of cerebral vasospasm after subarachnoid hemorrhage.更多还原