体外血脑屏障模型的建立及功能测定-论文.pdf

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1、卒中与神经疾病2014年8月第21卷第4期·195··论著·体外血脑屏障模型的建立及功能测定陈雪王军王伟骆翔喻志源【摘要】目的利用大鼠原代微血管内皮细胞及星形胶质细胞建立体外血脑屏障模型，并通过跨内皮细胞电阻(Trans-epitheliumelectricalresistant，TEER)方法对血脑屏障模型进行功能测定。方法原代分离纯化SD大鼠脑微血管内皮细胞和星形胶质细胞，用免疫荧光检测内皮细胞标志物vwF，紧密连接蛋白ZO-1，星形胶质细胞标志物GFAP；用微血管内皮细胞和星形胶质细胞在Transwell小室上建立体外血脑屏障模型，观察TEER值的动

2、态变化。结果原代的微血管内皮细胞培养至融合后具有典型的梭形“铺路石”样外观，ⅧF鉴定细胞纯度达到95以上，Zo-1免疫荧光鉴定证实细胞间形成紧密连接；原代培养的星形胶质细胞呈现具有多个突起的典型形态，GFAP鉴定细胞纯度达到95以上；在第10d，单独微血管内皮细胞血脑屏障模型的TEER值为(42±1．41)acm2，内皮细胞和星形胶质细胞共培养血脑屏障模型的TEER值为(65±1．42)flcm2。结论建立了体外血脑屏障模型，通过TEER值测定证明共培养使模型更加完整，更加接近在体血脑屏障模型的特性。【关键词】血脑屏障微血管内皮细胞星形胶质细胞跨内皮细胞电

3、阻【中图分类号】R741【文献标识码】A【文章编号】10O7—0478(2O14)04—0195—04[DOPl10．3969／j．issn．1007—0478．2014．04．001，nIeestablishmentandfunctionclI{lracteristicsofbloodbrainbarriermodelinvitroChenXue，W口ngJun，Wangw，eta1．DepartmentofNeurology，TongjiHospital，TongjiMedicalCollege，HuazhongUniver-sityofScience

4、andTechnology，Wuhan430030[Abstract]ObjectiveToestablishinvitroblood-brainbarriermodelusingmicrovascularendotheliacellsandastrocytesofprimaryrat，andidentifythemodelbymeasuringTEERvalue．MethodsPrimarySDratmicrovascularendothelialcellsandastrocyteswereisolatedandpurified．Theendotheli

5、alcel1markerVWF，tightjunctionproteinsZO-1，theastrocytemarkerGFAPweredetectedusingimmunofluorescencetechniques．Aninvitroblood-brainbarriermodelwasestablishedinTranswellinsertsusingendothelialcellsandastrocytescoculturesystem．WeobservedthedynamicchangeofTEER，studiedtheestablishmenta

6、ndcharacteristicsofthebloodbrainbarrier．ResultsOncefusing，primarymicrovascularendothelialcellspresentedatypicalfusi—forrncobblestoneappearance．Morethan95ofendothelialcellswerevwFpositive．Z()_1confirmedthefotinoftightjunctionsbetweenendothelialcells．Primaryastrocytespresentedatypic

7、almorphologywithmanypocesses．Morethan95％ofastrocyeswereGFAPpositive．TheTEERofmicrovascularendothelialcellalonemodelwas(42±1．41)Qcn12，whiletheTEERofthecoculturemode1was(65±1．42)Qcm2．ConclusionsVitroblood-brainbarriermodelwasestablished．BytheTEERvaluemeasurementweprovethatcocultures

8、ystemofthebloodbrainbarriermodelw