Nrf2基因敲除对小鼠蛛网膜下腔出血后小胶质细胞/巨噬细胞活化的影响  

Nrf2 deficiency promotes microglia/macrophage activation after subarachnoid hemorrhage in mice

作　　者：李桃[1] 王汉东[1] 丁宇[1] 何进[1] 丁可[1] 陆新宇[1] 徐建国[2] 韦武亭 

机构地区：[1]南京大学医学院附属金陵医院(南京军区南京总医院)神经外科 [2]南京大学医学院附属金陵医院(南京军区南京总医院)麻醉科,南京210002

出　　处：《医学研究生学报》2015年第1期11-15,共5页Journal of Medical Postgraduates

基　　金：国家自然科学基金(81070974);江苏省医学重点学科基金(XK201129)

摘　　要：目的脑内小胶质细胞/巨噬细胞(microglia/macrophage,M/M)在蛛网膜下腔出血(subarachnoid hemorrhage,SAH)脑组织损伤和修复中具有重要作用。文中研究小鼠SAH后M/M活化及Nrf2基因敲除对M/M活化的影响。方法野生型ICR小鼠70只,来源于ICR的Nrf2基因敲除小鼠35只,采用小鼠视交叉注射自体血建立SAH模型。假手术组行同样抽血、钻孔及置针,但不行注血。实验分为4组,野生型假手术组、野生型SAH组、基因敲除假手术组、基因敲除SAH组。SAH术后第1、3、5天取标本,用Western blot法和免疫组化法检测M/M特异性蛋白Iba1的表达;同时Western blot检测SAH后第3天Iba1蛋白表达,免疫荧光染色检测SAH后第3天CD16/32+Iba1+细胞数量。结果 Western blot检测结果显示野生型假手术组、野生型SAH组第1、3、5天Iba1蛋白表达灰度值分别为0.491±0.039、0.657±0.069、0.930±0.046和0.926±0.046,而Iba1蛋白免疫组织化学染色平均光密度值为0.412±0.122、0.625±0.135、0.963±0.213和0.978±0.224,野生型SAH组第1、3、5天Iba1蛋白表达均较野生型假手术组明显增加(P<0.05),但野生型SAH组第3天与第5天Iba1表达差异无统计学意义(P>0.05);Western blot结果示野生型假手术组、基因敲除假手术组、野生型SAH组、基因敲除SAH组SAH后第3天Iba1蛋白表达灰度值分别为1.157±0.080、1.162±0.073、1.580±0.171和1.913±0.220,CD16/32+Iba1+细胞数量分别为0、0、30.200±6.300和42.800±6.260(个/HP),SAH后第3天基因敲除SAH组Iba1蛋白表达较野生型SAH组增加(P<0.05),且基因敲除SAH组CD16/32+Iba1+细胞数较野生型SAH组明显增多(P<0.05)。结论小鼠SAH后M/M活化增加,Nrf2基因敲除促进了SAH后M/M的活化和M1极化。Objective Subarachnoid hemorrhage （ SAH） is a devastating disease with high fatality and morbidity and micro-glia/macrophage （ M/M） plays a vital role in SAH brain injury with complicated pathophysiological mechanism .This study was to ob-serve the effect of Nrf2 deficiency on M/M activation and M1 polarization after subarachnoid hemorrhage in mice . Meth ods We col-lected 70 wild-type （ WT） ICR mice and 35 Nrf2-knockout （ KO） mice to establish the SAH model by injecting fresh autologous blood into pre-chiasmatic cistern.WT mice were arranged into four groups： sham operation group, post operative day 1 （POD1） group, POD3 group and POD5 group.Then WT mice and Nrf2 Nrf2-knockout mice were divided into sham operation WT group , sham opera-tion KO group, SAH WT group and SAH KO group.Western blotting （WB） and immunohistochemistry （IHC） were applied to observe the activation and proliferation of M/M after SAH on WT mice .Difference in activation and M 1 polarization were observed by detecting Iba1 expression in WB and CD 16/32 ＋Iba1 ＋cells in immunofluorescence between WT and KO mice . Results Gray scale values of Iba1 expression by WB in WT mice are 0.491 ±0.039, 0.657 ± 0.069, 0.930 ±0.046 and 0.926 ±0.046;average optical intensity values of Iba1 expression by IHC in WT mice are 0.412 ±0.122, 0.625 ±0.135, 0.963 ±0.213 and 0.978 ±0.224.The data indica-ted that Iba1 expression increased in SAH KO group in comparison to SAH WT group on 1, 3, 5 day after SAH （P〈0.05）.Moreover, Nrf2 deficiency promoted the activation and polarization of M /M by increased Iba1 protein expression and CD16/32 ＋Iba1 ＋cells after SAH （ P〈0.05）. Conclusion SAH induces M/M activation and proliferation in mice, and Nrf2 deficiency promotes the activa-tion, proliferation and M1 polarization after SAH .

关 键 词：蛛网膜下腔出血 NRF2 小胶质细胞/巨噬细胞 活化 极化 

分 类 号：R743[医药卫生—神经病学与精神病学]