腺病毒介导IL-1ra基因对小鼠局灶性脑缺血模型中ICAM-1表达抑制作用

Expression of ICAM-1 is Reduced in Permanent Focal Cerebral Ischemic Mouse Brain through Thansfer of Adenoviral mediated IL-1ra Gene

目的 IL 1可诱导内皮细胞粘附分子表达 ,促进缺血部位炎症和局部细胞损伤 ,应用腺病毒介导IL 1ra基因观察其对小鼠局灶性脑缺血后内皮细胞ICAM 1蛋白表达的抑制作用。方法 构建携带外源性基因、复制缺陷的人腺病毒 ,并将其作为载体介导IL 1ra基因、EcoliLacZ基因或以生理盐水分别注射于成年CD 1小鼠右侧脑室。 5d后 ,对实验小鼠分别进行脑血管腔内栓塞术 ,造成持续性中动脉栓塞 ,缺血程度由激光多普勒流量仪确定。采用组织病理切片观察病灶部位白细胞浸润和炎症反应现象 ,免疫组化染色测定血管内皮细胞ICAM 1蛋白表达和用Western印迹进行定量分析。结果 病灶侧皮质区血管内皮细胞ICAM 1蛋白表达量在IL 1ra基因组有明显下降且局部炎症反应轻微 ,与LacZ基因组和生理盐水组比较差异有显著性。ICAM 1蛋白定量分析表明 ,前组比后两组下降 5 0 %左右。但正常侧和病灶侧的脑基质神经节区无明显差异。结论 IL 1ra基因表达蛋白通过抑制IL 1的生物学功能 ,明显下调脑缺血皮质区ICAM 1蛋白表达 ,有助于减轻局部炎症。

Objective Interleukin 1 (IL 1) may increase local inflammation in ischemic site and contribute to ischemic cell damage by induce expression ICAM 1 on the endothelium. The present study was designed to determinate overexpression of Interleukin 1 receptor antagonist(IL 1ra) can reduce the expression of ICAM 1 on the endothelium through transfer of adenoviral mediated IL 1ra gene after permanent focal cerebral ischemic mouse brain.Methods The construction of replication defective recombinant human adenovirus vector was used, in which the human IL 1ra gene or Eschricha Coli β galactoside (LacZ) gene was inserted in the genome of a adenovirus. Adenovirus vector(1×10 9 particle) encoding the IL 1ra gene or the LacZ gene and normal saline were injected into the right lateral cerebral ventricle of adult CD 1 mice, after five days, permanent middle cerebral antery occlusion (MCAO) was achieved for 24h using an intraluminal suture. Cerebral flow was monitored by transcranial laser Doppler flowmetry to verify the occlusion. ICAM 1 protein was quantified using Western blot analysis and localized immunohistochemistry. Inflammatory response of leukocyte infiltration was determined by histopathological examination.Results There were fewer ICAM 1 positive vessels in the ischemic cortex of the Ad.RSVIL 1ra thansfected mice than in the Ad.RSVLacZ thansfected and normal saline treated mice. Western blot analysis showed that ICAM 1 protein decreased about 50% in the Ad.RSVIL 1ra group compared to the other two groups. There were no significant differences in the numbers of positive vessels in the ischemic basal ganglia and contralateral hemisphere among the three groups.Conclusions IL 1ra overexpression can down regulate the expression of ICAM 1 and decressed inflammatory reaction in the ipsilateral cortex in ischemic mouse brain by inhibiting of IL 1 biological function.