依达拉奉通过Caveolin-1/VEGF信号通路促进缺血性卒中大鼠脑血管新生及神经功能修复的作用机制探讨

Exploration on the mechanism of edaravone in promoting cerebral angiogenesis and nerve function repair in ischemic stroke rats through Caveolin-1/VEGF signaling pathway

目的探讨依达拉奉促进缺血性卒中大鼠脑血管新生及神经功能修复的作用机制。方法选择无特定病原(SPF)级SD雄性大鼠60只。随机取12只作为假手术组,剩余48只大鼠通过阻塞大脑中动脉血流构建缺血性卒中模型,并分为缺血性卒中组、依达拉奉组、抑制剂组和依达拉奉+抑制剂组,每组12只。通过Morris水迷宫检测认知功能,通过2,3,5-三苯基氯化四氮唑(TTC)染色检测脑梗死面积。取全脑组织,冷冻切片后通过免疫荧光法观察缺血核心区小凹蛋白-1(Caveolin-1)/CD34、双肾上腺皮质激素(DCX)/5-溴脱氧尿苷(BrdU)蛋白荧光强度。取脑组织缺血侧基底结区组织,通过Western blot法检测Caveolin-1、血管内皮生长因子(VEGF)、脑源性神经营养因子(BDNF)与酪氨酸激酶受体B(TrkB)蛋白表达水平。结果与假手术组相比,缺血性卒中组大鼠神经功能评分增加,逃避潜伏期增加且穿越平台次数减少,Caveolin-1、CD34荧光强度增加,Caveolin-1与VEGF蛋白表达增加,BDNF、TrkB、DCX和BrdU蛋白表达减少,差异有统计学意义(P<0.05)。与缺血性卒中组相比,依达拉奉组大鼠神经功能评分、逃避潜伏期减少,穿越平台次数增加,脑梗死面积减少,Caveolin-1与CD34共表达提高,Caveolin-1、VEGF蛋白表达水平进一步增加,BDNF、TrkB、DCX和BrdU蛋白表达增加,差异有统计学意义(P<0.05)。与依达拉奉组相比,抑制剂组与依达拉奉+抑制剂组大鼠Caveolin-1与VEGF蛋白表达降低,Caveolin-1与CD34共表达减少,脑梗死面积与逃避潜伏期增加,差异有统计学意义(P<0.05)。结论依达拉奉的抗缺血性卒中作用与Caveolin-1/VEGF信号通路激活相关,并通过促进神经血管新生,恢复缺血区血供,减轻大鼠神经功能损伤,提高认知功能。

Objective To explore the mechanism of edaravone in promoting cerebral angiogenesis and nerve function repair in ischemic stroke rats.Methods Sixty specific pathogen free(SPF)male Sprague-Dawley(SD)rats were selected.Among the 60 SD rats,twelve rats were randomly selected as sham operation group,and the other 48 rats were made into an ischemic stroke model by blocking the blood flow in the middle cerebral artery,and the model rats were divided into ischemic stroke group,edaravone group,inhibitor group and edaravone+inhibitor group,with 12 rats in each group.Cognitive function was detected by Morris water maze,and cerebral infarction size was measured by 2,3,5-triphenyltetrazolium chloride(TTC)staining.The whole brain tissues were taken,frozen and sectioned,and the fluorescence intensities of Caveolin-1/CD34 and doublecortin(DCX)/5-bromo-2′-deoxyuridine(BrdU)protein in the ischemic core area were observed by immunofluorescence method.The ischemic basal ganglia area of the brain tissues was taken.The expression levels of Caveolin-1,vascular endothelial growth factor(VEGF),brain-derived neurotrophic factor(BDNF)and tyrosine kinase receptor B(TrkB)were detected by Western blot.Results Compared with those in the sham operation group,the rats in the ischemic stroke group had increased neurological function scores,increased escape latency and decreased number of crossing platforms,increased fluorescence intensities of Caveolin-1 and CD34,increased expressions of Caveolin-1 and VEGF proteins,and decreased expressions of BDNF,TrkB,DCX and BrdU proteins,and the differences were statistically significant(P<0.05).Compared with the ischemic stroke group,the edaravonegroup had reduced neural function scores and escape latency,and increased number of crossing platforms,and decreased cerebral infarction size,and the increased co-expressions of Caveolin-1 and CD34,and further increased expression levels of Caveolin-1 and VEGF,and increased expressions of BDNF,TrkB,DCX and BrdU proteins,and the differences were statistically significant(P<0.05).Compared with those in the edaravone group,the rats in the inhibitor group and the edaravone+inhibitor group had decreased expressions of Caveolin-1 and VEGF proteins,and decreased co-expressions of Caveolin-1 and CD34,and increased cerebral infarction size and escape latency,and the differences were statistically significant(P<0.05).Conclusion The anti-ischemic stroke effect of edaravone is related to the activation of Caveolin-1/VEGF signaling pathway,and edaravone can improve cognitive function by promoting neuroangiogenesis,restoring blood supply in ischemic area and alleviating nerve function injury in rats.