变构促红细胞生成素对中年雌性脑缺血小鼠白质损伤的作用研究  

Erythropoietin reduces white matter injury after middle cerebral artery occlusion in middle-aged female mice

作　　者：王荣亮 杨振宏 马舒贝[3] 王韬 罗玉敏 焦力群[2] Wang Rongliang;Yang Zhenhong;Ma Shubei;Wang Tao;Luo Yumin;Jiao Liqun(Cerebrovascular Diseases Research Institute,Xuanwu Hospital,Capital Medical University,Beijing 100053,China)

机构地区：[1]首都医科大学宣武医院脑血管病研究室,北京100053 [2]首都医科大学宣武医院神经外科,北京100053 [3]大连市中心医院神经内科

出　　处：《中国脑血管病杂志》2022年第1期24-34,共11页Chinese Journal of Cerebrovascular Diseases

基　　金：国家自然科学基金资助项目(81801149);辽宁省科学技术计划项目(2019-ZD-0884);大连市医学科学研究计划项目(1911005)。

摘　　要：目的研究变构促红细胞生成素(MEPO)对成年雌性脑缺血小鼠白质的保护作用。方法将30只成年雌性C57BL/6J小鼠按随机数字表法完全随机分为3组:假手术组(Sham组),脑缺血模型组(Vehicle组)及脑缺血MEPO治疗组(MEPO组),每组10只。采用大脑中动脉远端血管永久性闭塞的方式制备小鼠脑缺血模型。MEPO组小鼠在大脑中动脉远端血管永久性闭塞即刻通过腹腔注射MEPO(5000 IU/kg),Vehicle组在制模后注入等量的等渗盐水,Sham组只进行血管暴露,不进行闭塞。脑缺血后第2天开始3组小鼠每天以50 mg/kg的剂量腹腔注射5-溴-2′-脱氧尿苷(BrdU),每日1次,持续至术后11 d。术前、术后每日记录各组小鼠的体质量。分别在术前及术后1、3、5、9、12、14 d采用贴条实验对小鼠神经功能进行评估[记录小鼠第1次触碰贴条所用的时间(接触时间)和将贴条撕掉所用的时间(移除时间)]。小鼠脑缺血后第3天,每组处死5只小鼠,2,3,5-三苯基氯化四氮唑(TTC)染色观察脑梗死和脑水肿程度(脑梗死百分比和脑水肿百分比)。脑缺血后第14天处死其余生存的小鼠,免疫荧光染色分别检测脑白质损伤程度[髓鞘碱性蛋白(MBP)/神经丝200(NF200)荧光强度比值]、小胶质细胞表型(采用Iba1标记小胶质细胞,CD16/Iba1标记M1型小胶质细胞,CD206/Iba1标记M2型小胶质细胞)、少突胶质细胞[2′,3′-环腺苷酸-3′-磷酸二酯酶(CNPase)标记成熟少突胶质细胞,BrdU标记新生细胞,两者共染阳性(CNPase+/BrdU+)表示新生的少突胶质细胞]以及CCAAT增强子结合蛋白β(C/EBPβ)在小胶质细胞中的表达情况(以C/EBPβ+/Iba1+表示)。结果(1)死亡:Vehicle组小鼠死亡2只(分别于术后12 d和13 d),MEPO组小鼠死亡1只(术后3 d),Sham组小鼠无死亡。(2)体质量:与Vehicle组比较,MEPO组小鼠体质量虽然有增加趋势,但差异均无统计学意义(均P>0.05)。(3)神经功能:与Vehicle组小鼠比较,MEPO组小鼠在脑缺血Objective To investigate the potential neuroprotective effects of mutant erythropoietin(MEPO)on white matter in female mice after cerebral ischemia.Methods Total 30 adult female C57BL/6J mice were randomly divided into 3 groups according to the random number table method:Sham group,Vehicle group and MEPO treatment group,with 10 mice in each group.The rat model of cerebral ischemia was established by permanent occlusion of the distal middle cerebral artery.Mice in the MEPO group were immediately injected intraperitoneally with MEPO(5000 IU/kg)after permanent occlusion of the distal middle cerebral artery.Mice in the Vehicle group were injected with isotonic saline after modeling,while mice in the Sham group were only exposed of the blood vessels without occlusion.From day 2 after cerebral ischemia,mice in groups 3 were intraperitoneally injected with 5-bromo-2′-deoxyuridine(BrdU)at a dose of 50 mg/kg,once a day,until 11 days after surgery.The body weight of mice in each group was recorded before and daily after operation.Neurological function of mice was evaluated by adhesive removal tests at 1,3,5,9,12 and 14 days after distal middle cerebral artery occlusion(MCAO;the time of touching the strip for the first time[contact time]and the time of removing the strip[removal time]were recorded).Five mice in each group were euthanized at day 3 after distal MCAO.The degree of cerebral infarction and cerebral edema(percentage of cerebral infarction and percentage of cerebral edema)was measured by 2,3,5-triphen yltetrazolium chloride(TTC)staining.Immunofluorescence staining was used to assess the white matter injury(the relative ratio of myelin basic protein vs.neurofilament protein 200 immunostaining intensity),microglial polarization(Iba1 as the marker of microglia,CD16/Iba1 as the marker of M1-microglia,CD206/Iba1 as the marker of M2-microglia),oligodendrocytes(2′,3′-cyclic adenylate-3′-phosphodiesterase[CNPase]marks mature oligodendrocytes and BrdU marks newborn cells,co-positive[CNPase+/BrdU+]indicates newborn

关 键 词：促红细胞生成素 脑缺血 白质损伤 CCAAT增强子结合蛋白β 

分 类 号：R743.3[医药卫生—神经病学与精神病学]