咪康唑与纤维蛋白协同促进神经球分化为成髓鞘样细胞  

Miconazole and fibrin synergistically promote the differentiation of neurospheres into myelinating-like cells

作　　者：崔学文[1] 周焕高 陈平波[1,2] 杨文静 龚爱华[2] 张志坚[2] 陈谦[2] Cui Xuewen;Zhou Huangao;Chen Pingbo;Yang Wenjing;Gong Aihua;Zhang Zhijian;Chen Qian(The Affiliated Hospital;The Medical School, Jiangsu University, Zhenjiang 212013, China)

机构地区：[1]江苏大学附属医院,镇江212013 [2]江苏大学医学院,镇江212013

出　　处：《神经解剖学杂志》2018年第2期158-164,共7页Chinese Journal of Neuroanatomy

基　　金：国家自然科学基金(81501077);镇江市社会发展科技基金(SH2012026);江苏大学高级人才启动基金(15JDG083);江苏省普通高校研究生创新计划项目(SJLX16-0449;SJZZ16-0202)

摘　　要：目的:探讨咪康唑与纤维蛋白能否协同促进神经球分化为成髓鞘样细胞。方法:培养并鉴定胎鼠神经球;通过MTT法确定不同浓度咪康唑对神经球存活的影响;将神经球按培养条件不同分为四组,对照组、咪康唑组(mico组)、纤维蛋白组(fibrin组)及纤维蛋白-咪康唑联用组(fibrin+mico组),培养两周后以免疫荧光染色比较髓磷脂碱性蛋白(MBP)的表达;Western Blot法比较MBP、2’3’环核苷酸磷酸二酯酶(CNPase)、整合素、细胞外调节蛋白激酶及其磷酸化(ERK、p-ERK)的表达。结果:(1)神经球阳性表达巢蛋白(nestin)和CD133:当咪康唑浓度低于或等于1.5μmol/L时,神经球存活在各组之间不存在统计学差异(P>0.05),但2.5μmol/L组显著低于2μmol/L组和1.5μmol/L组(P<0.05)。(2)免疫荧光结果显示:神经球分化所成的成髓鞘样细胞表达MBP阳性,呈多突起形态;fibrin组及mico组MBP相对荧光强度均高于对照组(P<0.05),fibrin+mico组神经球分化为网络状的成髓鞘样细胞团,其相对荧光强度高于其他组(P<0.05)。(3)Western Blot结果显示:fibrin+mico组MBP及CNPase表达显著高于mico组和fibrin组(P<0.05),fibrin+mico组整合素表达及p-ERK/ERK显著高于mico组和fibrin组(P<0.05),同时咪康唑及纤维蛋白对MBP、CNPase、整合素的表达及p-ERK/ERK的效应具有交互作用(P<0.05)。结论:咪康唑与纤维蛋白能协同促进神经球分化为成髓鞘样细胞,其机制可能与ERK磷酸化有关。Objective： To explore the synergism of miconazole and fibrin to promote the differentiation of neurospheres into myelinating-like cells. Methods： Neurospheres were cultured from fetal mice brain and then characterized;MTT assay was used to determine the effects of miconazole of different concentrations on the viability of neurospheres;neurospheres were divided into four groups according to the various culture conditions：（ 1） control group,（ 2） miconazole（ mico group）,（ 3） fibrin group,（ 4） combination of fibrin and miconazole（ fibrin ＋ mico group）. After cultured for two weeks,these cells were subjected to immunofluorescence staining to compare the expression of myelin basic protein（ MBP）,and Western Blot was used to analyze the expressions of MBP,2＇-3＇cyclic nucleotide phosphodiesterase（ CNPase）,integrin,and extracellular signal-regulated kinase and its phosphorylation（ ERK and p-ERK）. Results：（ 1） Neurospheres positively expressed nestin and CD133. The difference of neurospheres viability in the groups treatedwith miconazole lower than 1. 5 μmol/L had no statistical significance（ P〉0. 05）. However,the viability of neurospheres in the 2. 5 μmol/L group was significantly lower than those in the 2 and 1. 5 μmol/L groups（ P〈0. 05）.（ 2） The results of immunofluorescence staining demonstrated that the myelinating-like cells differentiated from neurospheres positively expressed MBP and showed multi-processed morphology. The relative fluorescence intensity of MBP in fibrin group and mico group was significantly higher than that in control group respectively（ P〈0. 05）. The neurospheres in fibin ＋ mico group differentiated to myelinating-like cells which formed networks. The relative fluorescence intensity in fibrin ＋ mico group was higher than other groups（ P〈0. 05）.（ 3） The results of Western Blot showed that the expression of MBP and CNPase in fibrin ＋ mico group was higher than that in fibrin or mico group（ P〈0. 05）;

关 键 词：咪康唑 纤维蛋白 神经球 成髓鞘样细胞 

分 类 号：R651.2[医药卫生—外科学]