星状神经节阻滞对大鼠脑缺血再灌注时M1型小胶质细胞活化的影响

Effects of stellate ganglion block on activation of M1 microglia during cerebral ischemia-reperfusion in rats

目的评价星状神经节阻滞(SGB)对大鼠脑缺血再灌注时M1型小胶质细胞活化的影响。方法SPF级健康雄性SD大鼠54只,8~10周龄,体重240~270 g,采用随机数字表法分为3组(n=18):假手术组(Sham组)、脑缺血再灌注组(IR组)和SGB组。Sham组仅手术操作但未插入线栓,IR组采用线栓阻塞大脑中动脉90 min恢复灌注的方法制备大鼠脑缺血再灌注损伤模型,SGB组于再灌注即刻采用颈交感神经干离断术行左侧SGB。于再灌注6、12和24 h时心尖采血,采用ELISA法检测血清TNF-α、IL-6和IL-1β浓度;于再灌注24 h时行神经功能评分后处死大鼠取脑组织,采用HE染色法观察皮质区细胞病理学结果;采用TTC染色法确定脑梗死体积百分比;采用TUNEL法检测皮质区细胞凋亡情况并计算细胞凋亡率;采用Western blot法检测小胶质细胞标志物Iba-1、活化的M1型小胶质细胞标记物CD68的表达。结果与Sham组比较,IR组和SGB组神经功能评分、脑梗死体积百分比、皮质区细胞凋亡率、血清TNF-α、IL-6和IL-1β浓度升高,Iba-1和CD68表达上调(P<0.05);与IR组比较,SGB组神经功能评分、脑梗死体积百分比、皮质区细胞凋亡率、血清TNF-α、IL-6和IL-1β浓度降低,Iba-1和CD68表达下调(P<0.05)。SGB组脑组织病理学损伤较IR组减轻。结论SGB减轻大鼠脑缺血再灌注损伤的机制与抑制M1型小胶质细胞活化有关。

Objective To evaluate the effects of stellate ganglion block(SGB)on the activation of M1 microglia during cerebral ischemia-reperfusion(I/R)in rats.Methods Fifty-four SPF healthy male Sprague-Dawley rats,aged 8-10 weeks,weighing 240-270 g,were divided into 3 groups(n=18 each)using a random number table method:sham operation group(group Sham),cerebral I/R group(group IR)and SGB group.Blood vessels were only exposed,without occlusion in group Sham.Cerebral I/R was induced by occlusion of the middle cerebral artery for 90 min followed by reperfusion in group IR.Cervical sympathetic trunk transaction was performed to induce left SGB immediately after onset of reperfusion in group SGB.Blood samples were collected from the apex of the heart at 6,12 and 24 h of reperfusion for determination of the concentrations of tumor necrosis factor(TNF)-α,interleukin(IL)-6,and IL-1βin the serum(using enzyme-linked immunosorbent assay).The animals were sacrificed after the neurological function was evaluated at 24 of reperfusion,and brain tissues were removed for microscopic examination of the pathological changes in cortex,for determination of percentage of cerebral infarct size(by TTC staining),for assessment of cell apoptosis and apoptosis rate in cortex(by TUNEL),and for determination of the expression of microglial biomarker Iba-1 and activated M1 microglia biomarker CD68(by Western blot).Results Compared with group Sham,the neurological function score,percentage cerebral infarct size,apoptosis rate in cortex,concentrations of TNF-α,IL-6 and IL-1βin the serum,and the expression of Iba-1 and CD68 were significantly increased in IR and SGB groups(P<0.05).Compared with group IR,the neurological function score,percentage cerebral infarct size,apoptosis rate in cortex,concentrations of TNF-α,IL-6 and IL-1βin the serum,and the expression of Iba-1 and CD68 were significantly decreased in group SGB(P<0.05),and the pathological changes of brain tissues were significantly attenuated in group SGB.Conclusion The mechanism by which SGB reduces cerebral I/R injury is related to inhibiting activation of M1 microglia in rats.