脑缺血/再灌注损伤后大鼠不同时间点神经细胞的动态变化  

Dynamic changes of neuronal cells at different time points following cerebral ischemia-reperfusion injury in rats

作　　者：邹旭欢 兰瑞[2] 付雪琴 王玮玮 王漫漫[1,2] 唐琛 刘双[1,2] 李泓宇 沈晓明 ZOU Xu-huan;LAN Rui;FU Xue-qin;WANG Wei-wei;WANG Man-man;TANG Chen;LIU Shuang;LI Hong-yu;SHEN Xiao-ming(the First Clinical Medical College,Henan University of Chinese Medicine,Zhengzhou 450046,China;Brain Disease Center,the First Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450099,China)

机构地区：[1]河南中医药大学第一临床医学院,河南郑州450046 [2]河南中医药大学第一附属医院脑病中心,河南郑州450099

出　　处：《中国药理学通报》2024年第6期1056-1066,共11页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No 81973618,81503422);河南省中医药科学研究专项课题(No 2022JDZX090)。

摘　　要：目的通过建立脑缺血/再灌注损伤模型,探讨急性CIRI后不同时间点神经细胞的动态变化规律。方法将SPF级雄性SD大鼠随机分为6组,分别为假手术组(Sham)和脑缺血/再灌注损伤(IR)不同时间点组。采用大脑中动脉线栓法(MCAO)建立局灶性脑缺血/再灌注损伤模型,采用Longa评分法评估大鼠神经行为评分,模型制备成功后,常规方法制作石蜡切片,进行TUNEL染色和免疫组织化学染色观察细胞凋亡,采用NeuN抗体进行免疫染色观察神经元细胞存活率,采用免疫组织化学法对星形胶质细胞标志物GFAP、小胶质细胞标志物IBA-1、内皮细胞标志物CD31进行染色,在不同时间点观察各组不同细胞的变化规律。结果假手术组的大鼠神经细胞在不同时间点未见明显变化,在脑缺血/再灌注损伤不同时间点组,细胞凋亡在IR3h被激活,且随着时间的延长,凋亡细胞数量增多且伴随形态学的破坏;NeuN+神经元在IR3h后出现缺血损伤的迹象,细胞形态异常,在24 h时最为严重;GFAP+星形胶质细胞在IR3h后急剧减少,IR6h标记不良星形胶质细胞数增加,到24、48 h星形胶质细胞在梗死区基本消失。IR3h IBA-1+小胶质细胞阳性细胞数减少,IR6h IBA-1+小胶质细胞体积增大,IR12h梗死区小胶质细胞死亡。CD31+内皮细胞在IR3h后在梗死周围皮层和纹状体明显增加,并持续到48 h。结论脑缺血/再灌注损伤后,凋亡细胞的数量随着时间延长而增加,NeuN+神经元细胞在24 h损伤最重;梗死周围皮层GFAP+星形胶质细胞、小胶质细胞随着时间增长而逐渐死亡;CD31+内皮细胞数量在再灌注3 h后在梗死周围皮层和纹状体明显增加,并持续到48 h。Aim To investigate the dynamic changes of neuronal cells at different time points following acute cerebral ischemia-reperfusion injury by establishing a model of brain ischemia-reperfusion injury.Methods Thirty male Sprague-Dawley(SD)rats were randomly divided into six groups:sham group and cerebral ischemia-reperfusion injury(IR)groups at different time points.Focal cerebral ischemia-reperfusion injury model was established using the middle cerebral artery occlusion(MCAO)technique.The Longa scoring method was used to assess neurobehavioral scores in rats.After successful model preparation,routine paraffin sections were made,and TUNEL staining and immunohistochemistry staining with NeuN antibody were performed to observe cell apoptosis and neuronal cell survival,respectively.Immunohistochemistry staining was also performed to investigate the changes in glial fibrillary acidic protein(GFAP)as a marker for astrocytes,ionized calcium-binding adapter molecule 1(IBA-1)as a marker for microglia,and CD31 as a marker for endothelial cells at different time points.Results No significant changes were observed in neuronal cells of the sham group at different time points.In the cerebral ischemia-reperfusion injury groups,cell apoptosis was activated at IR3h and increased in quantity with morphological damage as time progressed.NeuN+neurons showed signs of ischemic injury after IR3h,with abnormal cell morphology.From 12 h,NeuN+neurons decreased in a time-dependent manner and reached their peak severity at 24 h.GFAP+astrocytes decreased significantly after IR3h,while poorly labeled GFAP+astrocytes increased at IR 6 h and almost disappeared in the infarcted area at 24 h and 48 h.The number of IBA-1+microglia-positive cells decreased at IR3h,and their volume increased at IR6h.Microglial cell death was observed in the infarcted area at IR12h.CD31+endothelial cells around the infarcted cortex and striatum increased significantly after IR3h and persisted until 48 h.Conclusions After cerebral ischemia-reperfusion injury,the number o

关 键 词：脑缺血/再灌注 不同时间点 动态变化 凋亡细胞 神经元细胞 星形胶质细胞 小胶质细胞 内皮细胞 

分 类 号：R-332[医药卫生] R322.81R329.25R743.31