大鼠局灶性脑缺血再灌注后内皮细胞对不同缺血时间的耐受性

背景脑缺血后内皮细胞结构和功能的完整性是决定缺血时间窗和出血转化的重要因素.目的动态观察缺血再灌注内皮细胞形态学和超微结构变化,了解内皮细胞对不同缺血时间的耐受性.设计随机对照实验.单位暨南大学第二临床学院神经内科.材料实验于1998-03/1999-03在暨南大学第二临床学院实验动物室完成.选择SD大鼠53只,随机分为9组①假手术组5只.②缺血3 h组6只.③缺血3 h再灌3 h组6只.④缺血6 h组6只.⑤缺血6 h再灌3 h组6只.⑥缺血12 h组6只.⑦缺血12 h再灌3 h组6只.⑧缺血24 h组6只.⑨缺血24 h再灌3 h 6只.方法采用线栓并环扎的方法建立大鼠局灶脑缺血模型.冠状面按A,B,C,D,E 5等分切脑,取C片脑组织TTC染色定位边缘区.取D片常规脱水、透明、包埋、切片,苏木精-伊红染色,光镜观察.取B片缺血周围区和中心区脑组织,经固定包埋,半薄切片,超薄切片,透射电镜下观察.主要观察指标①显微镜下观察不同缺血时间点内皮细胞的变化和出血性梗死发生的时间.②电镜下观察不同缺血时间点内皮细胞的超微结构改变.③电镜下观察紧密连接开放时间.④电镜下观察不同缺血时间点胶质细胞足突层空泡化程度.结果53只大鼠均进入结果分析.光镜下缺血3 h即可见神经毡疏松,小血管周围水肿,缺血12 h再灌3 h可见缺血中心区小动脉破裂出血.电镜下缺血3 h可见毛细血管内皮细胞核肿胀,胞浆内胞饮增加,足突层空泡化呈(+);缺血3 h再灌3 h中心区足突层空泡化呈(++),边缘区呈(+++);缺血6 h再灌3 h可见内皮紧密连接开放,足突层空泡化呈(+++);缺血12 h再灌3 h后胞饮明显减少,线粒体肿胀也少见,但紧密连接开放增加,足突层空泡化呈(+++)～(++++).结论内皮细胞在缺血3 h即可发生明显的结构变化,缺血6 h可见内皮细胞紧密连接开放,缺血12 h后可发生出血性转化,出血转化多发生于再灌注后的缺血中心区.

Endothelial cell endurance to various courses of focal brain ischemia-reperfusional injury in rats

BACKGROUND: Endothelial cell structural and functional integrity is importnat decisive fatcor for ischemic time-window and hemorragic transformation follwing brain ischemic injury.OBJECTIVE: To investiagte the endotheliocyt endurance to various course of ischemic injury basing on dynamical observation of morphological and ultrastructural changes of endotheliocyte during IR injury.DESIGN: Randomized controlled experiment.SETTING:Neurological Internal Department of the Second Affiliated Hospital of Jinan University.MATERIALS: This experiment was carried out in the Animal Experimental Laboratory of the Second Affiliated Hospital of Jinan University from March 1998 to March 1999. Totally 53 SD rats were randomly dihours of 6 rats.METHODS: Thread-bolt occlusion method was used to establish focal brain ischemia model on rats. Brain tissue was evenly cut into five coronary segments: namely A, B, C, D and E, segments C underwent TTC staining for marginal region location. segments D was taken for routine dehydration, transparency, envelop, slice and HE staining,optical microscopic observation. Ischemic surrounding area and central brain tissues was obtained from slice B, fixed and enveloped before cutting into ultrathin slices that was observed under transmission electron microscope.the occurring time of hemorrhagic infarction at different ischemic time cell vacuolization degree in foot process layer at different ischemic time points.RESULTS: Totally 53 rats were enrolled in this experiment and all data was entered into results analysis. Under optical microscope: Neuropil loose and small vascular surrounding edema was observed at ischemia 3 hours.Small arterial broken and hemorrhage occurred at ischemia 12 hours reperfusion 3 hours. Under electron microscope: Capillary endothelial nuclear swelling was observed at ischemia 3 hours, with cytoplasmic pinocytosi increasing and vacuolization in foot process layer appearing+; At ischemia 3 hours reperfusion 3 hours, the foot process layer vacuolization in center area was (++) and (+++) in marginal area; while at ischemia 6 hours reperfusion 3 hours, endothelia tight junction opened and vacuolization in foot process layer was (+++); pinocytosis was found obviously reduced after ischemia 12 hours reperfusion 3 hours, mitochondrial swelling was seldom observed, but tight junction increasingly opened and vacuolization in foot process layer appeared (+++) - (++++).CONCLUSION: Obvious structural changes of endotheliocyte appeared in post-ischemia 3 hours, endotheliocyte tight junction openning was observed at ischemia 6 hours, and hemorrage transformation occurred after ischemia 12 hours, mainly at the post-reperfusional ischemia center.