rhEPO对大鼠颅脑损伤早期脑微血管内皮细胞紧密连接的保护作用

Protective effect of recombinant human erythropoietin on tight junctions of endothelial cells of cerebral microvessels after traumatic brain injury in rats

目的探讨重组人促红细胞生成素(rhEPO)对颅脑损伤早期脑微血管内皮细胞的保护作用及机制。方法将81只SD大鼠按随机数字表法随机分为假手术组(n=9)、颅脑损伤(TBI)组(n=36)和rhEPO治疗组(n=36)。用Marmarou等人的方法制作大鼠颅脑损伤模型,分光光度仪测定各组大鼠脑组织伊文氏蓝(EB)含量,实时荧光定量PCR和免疫组织化学法检测闭合蛋白-5(claudin-5)、咬合蛋白(occludin)和胞质紧密粘连蛋白-1(ZO-1)的表达。结果伤后3h、24h、3d,与假手术组对比,TBI组大鼠脑组织EB含量明显升高(P<0.05),claudin-5、occludin和ZO-1mRNA及其蛋白的表达显著降低(P<0.05);与TBI组对比,rhEPO治疗组大鼠脑组织EB含量显著降低(P<0.05),claudin-5、occludin和ZO-1mRNA及其蛋白的表达显著升高(P<0.05)。结论颅脑损伤早期应用rhEPO,可能通过增加紧密连接相关蛋白claudin-5、occludin和ZO-1的表达而保护脑微血管内皮细胞。

Objective To explore the protective effect of recombinant human erythropoietin （rhEPO） on the tight junctions of the endothelial cells of the cerebral microvessels after the traumatic brain injury in rats. Methods Eight-one adult Sprague-Dawley rats were randomly divided into 3 groups, i.e. sham operation group （n=9）, traumatic brain injury （TBI） group （n=36） and rhEPO treatment group （n=36）. The TBI model was established by Marmarou method. The rhEPO （3 000 IU/kg） in the rhEPO treatment group or equal volume of physiological saline in the sham and TBI groups was intraperitoneally administered immediately after the injury and then once a day until the day of the sacrifice of the animals. Nine rats were sacrified 3, 24, 72 and 168 hours after the TBI in both the TBI and the rhEPO groups. The content of Evans blue （EB） in the cerebral tissue was measured by spectrophotometer, and the expression of Claudin-5, Occludin and zonula occludens-1 （ZO-1） were determined respectively by real-time fluorescence quantitative RT-PCR and immunohistochemistry. Results The content of EB in the cerebral tissue was significantly higher and the expression levels of Claudin-5, Occludin and ZO-1 mRNA were significantly lower in the TBI group than those in the sham operation group （P0.05）. The content of EB in the cerebral tissue was significantly lower and the expression levels of Claudin-5, Occludin and ZO-1 mRNA were significantly higher in the rhEPO treatment group than those in the TBI group （P0.05）. Conclusion It is suggested that the rhEPO has protective effect on the endothelial cells of the cerebral microvessels by increasing the expressions of Claudin-5, Occludin, and ZO-1 related to the tight junction in the rats with TBI.