白藜芦醇苷对脑缺血再灌注损伤小鼠血脑屏障的 保护作用研究

目的:观察白藜芦醇苷（PD）对脑缺血再灌注损伤小鼠血脑屏障的保护作用及对紧密连接蛋白的影响。方法:CD-1 小鼠 （102 只）应用线栓法制作短暂性大脑中动脉闭塞（tMCAO）模型。随机将实验动物分成4 组:假手术组（28 只）、缺血再灌注 组（28只）、白藜芦醇苷低剂量组（30 mg/kg，18只），白藜芦醇苷高剂量组（60 mg/kg，28只）。于缺血再灌注后24 h 进行行为学评 分、TTC 染色、干湿重测定、伊文思蓝染色，应用Western 印迹、免疫荧光检测内皮细胞标志物血小板内皮细胞黏附分子-1（CD- 31）、紧密连接蛋白闭锁小带蛋白-1（ZO-1）、咬合蛋白（Occludin）、闭合蛋白-5（Claudin-5）的表达情况。结果:（1）与缺血再灌注 组相比，白藜芦醇苷高剂量组改善了缺血再灌注小鼠行为学评分、减少了脑梗死体积（33.76%± 13.27%）vs.（57.47%±9.53%）， P<0.05]、减轻了缺血再灌注小鼠脑水肿程度（75.62%±7.95%）vs.（85.72%±6.20%），P<0.05]、降低了伊文思蓝渗出量（1.49±0.47） vs.（2.13±0.44），P<0.05]。（2）Western印迹结果显示白藜芦醇苷上调了ZO-1、Occludin、Claudin-5（均P<0.05）的表达水平。（3）免 疫荧光结果显示:白藜芦醇苷组上调了CD-31 在皮层缺血半暗带区的表达，且可见微血管形态相较缺血再灌注组更加光滑、完 整，同时白藜芦醇苷减少了ZO-1、Claudin-5 的破坏。结论:白藜芦醇苷的干预对缺血再灌注损伤小鼠起到了保护血脑屏障的作 用，上调了紧密连接蛋白ZO-1、Occludin、Claudin-5 的表达水平。

Protective effect of polydatin on blood-brain barrier in mice after transient focal cerebral ischemia

Objective:To investigate the protective effect of polydatin（PD）on the blood-brain barrier and tight junction proteins in ischemia-reperfusion mice. Methods:A total of 102 CD-1 mice were subjected to transient middle cerebral artery occlusion model（tMCAO）. The experimental animals were divided into 4 groups randomly:Sham group（n=28），tMCAO group（n=28），PD-L group（polydatin 30 mg/kg，n=18），PD-H group（polydatin 60 mg/kg，n=28）. Neurological deficit scores，2，3，5-triphenyltetrazolium chloride（TTC）， the standard wet-dry method，Evans blue exudation was evaluated 24 h after ischemia-reperfusion. Western blotting and immunofluorescence was applied to detect the expression level of endothelial cell markers platelet endothelial cell adhesion molecule-1（cd-31），tight junction protein atresia Band-1（ZO-1），occludin and claudin-5. Results:（1）Compared with the tMCAO group 24 h after tMCAO， the neurological deficit score was ameliorated，the infarct volume was reduced（33.76%±13.27%）vs.（57.47%±9.53%），P<0.05]， the brain water content was reduced（75.62%±7.95%）vs.（85.72%±6.20%），P<0.05]，the permeability of blood-brain barrier was de creased （1.49±0.47）vs.（2.13±0.44），P<0.05] in PD-H group respectively.（2）Western blotting showed that the level of ZO-1，Occludin， Claudin-5，was up-regulated significantly in the PD group 24 h after operation，compared with tMCAO group（P<0.05，respectively）.（ 3） Immunofluorescence showed that the expression of CD-31 was up-regulated in the cortex ischemic penumbra，and the tube shape of microvascular was smoother and more complete than that of the tMCAO group. Meanwhile，the disruption of ZO-1 and Claudin-5 staining was reduced in the PD group，compared with tMCAO group. Conclusion:Polydatin administration shows the protective effect of blood-brain barrier inmicewith ischemia-reperfusion injury，meanwhile it can up-regulate the expression of ZO-1，Occludin，Claudin-5.