急性创伤性脑损伤大鼠血脑屏障通透性变化与Claudin-5的相关性研究

Role of Claudin-5 in permeability of blood brain barrier in rats after acute traumatic brain injury

目的研究大鼠急性创伤性颅脑损伤（TBI）后血脑屏障（BBB）通透性的变化和机制。方法雄性Wistar大鼠96只按随机数字表法分为假手术组和TBI组，TBI模型参照Feeney自由落体致伤法制作，假手术组仅行开颅术不行打击致伤。每组按伤后处死时间的不同分为3h、6h、12h、24h、48h、72h6个亚组，每亚组4只。采用干湿重比法测定脑组织含水量，Westernblotting检测脑皮质紧密连接蛋白Claudin．5的表达，伊文思蓝（EB）法检测脑组织BBB通透性变化。结果与假手术组比较，TBI组大鼠创伤后不同时间点脑组织含水量、EB含量均增多。组内比较显示二者创伤后3h开始增多，24h达高峰，随后下降，差异有统计学意义（P〈0．05）；与假手术组比较，TBI组大鼠创伤后6h、12h、24h、48h、72h紧密连接蛋白Claudin-5表达降低。组内比较显示其创伤后6h表达降低，24h达最低值，随后回升，差异有统计学意义（P〈0．05）；大鼠脑Claudin-5蛋白的相对表达水平和脑组织水含量、脑组织EB含量呈负相关关系（r=-0．994，P=-0．000；r=-0．846，P=-0．036）。脑组织含水量和EB含量呈正相关关系（r=0．863，P=-0．027）。结论TBI后大鼠BBB通透性变化和脑损伤程度具时间依赖性，紧密连接蛋白Claudin-5与BBB变化具有一定程度的相关性。

Objective To study the changes of permeability of blood brain barrier （BBB） in rats after acute traumatic brain injury and explore its mechanism. Methods Ninety-six male Wistar rats were randomly divided into sham-operated group （S group） and traumatic brain injury group （TBI group）. Each group was divided into 6 subgroups （3, 6, 12, 24, 48 and 72 h after trauma）. Brain injury animal models were established according to Feeney＇s method. Measurement of Evans Blue （EB） was performed at different time points after injury to measure the changes of BBB permeability. Brain water content was tested by wet-dry weighting method. Expression of tight junction protein Claudin-5 was detected by Western blotting. Results As compared with that in rats of the S group, the brain water content and content of EB in rats of the TBI group were significantly increased at each time point （P〈0.05）; and in TBI subgroups, the brain water content and content of EB were begun to increase 3 h after TBI, reaching its peak level 24 h after TBI. As compared with that in rats of the S group, the expression of Claudin-5 in rats of the TBI group was significantly decreased at 12, 24, 48 and 72 h after TBI （P〈0.05）; and in TBI subgroups, the expression of Claudin-5 was begun to decrease 6 h after TBI, reaching its lowest level 24 h after TBI. Negative correlations were noted between protein expression level of Claudin-5 and both the brain water content and content of EB （r=-0.994, P=-0.000; r=-0.846, P=0.036）; positive correlation was observed between the brain water content and content of EB （r=0.863, P=-0.027）. Conclusion The degree of brain injury and changes of permeability of BBB may be time dependent in rats after acute brain traumatic injury; and Claudin-5 may be correlated with the changes of BBB.