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1、BV2小胶质细胞经典激活及替代激活的诱导及鉴定吴非1,2罗涛2郭远瑾2黎红华1梅元武2(1.广州军区武汉总医院神经内科湖北武汉430070)(2.华中科技大学协和医院祌经内科湖北武汉430022)【摘要】目的:观察BV2小胶质细胞经典激活及替代激活后代谢分子的变化,判断鉴别Ml及M2极化型小胶质细胞的方法。方法:小胶质细胞以100ng/ml的LPS及20ng/mL的IL-4分别经典激活及替代激活24小时,以ELISA方法检测IL-12、IL-10、TNF-α的分泌量,以实时荧光定量聚合酶链反应检测iNOS、Arg-1、IL-10、IL-12的mRNA
2、表达,以流式细胞术检测细胞膜MMr蛋白表达。结果:小胶质细胞被LPS经典激活后呈Ml型极化,IL-12、TNF-α分泌量、iNOSmRNA、IL-12mRNA表达量明显升高,与生理对照组比较有统计学差异(p<0.05)。小胶质细胞被IL-4替代激活后呈M2极化,IL-10分泌量、Arg-lmRNA及IL-lOmRNA表达量显著升高,流式细胞术检测MMr蛋白荧光强度较生理对照组显著上升(p<0.05)。结论:小胶质细胞的能够被LPS经典激活呈Ml型极化,IL-4替代激活呈M2型极化,M1/M2极化状态的小胶质细胞能够通过检测炎性相关因子、精
3、氨酸代谢分子及细胞膜特异性蛋白三个方面得到鉴别。【关键词】小胶质细胞;经典激活;替代激活;M1/M2型极化;脂多糖;白介素-4【中图分类号】R741【文献标识码】B【文章编号】1003-5028(2015)8-0651-02TheIdentificationofClassicalorAlternativeActivationonBV2MicrogliaWuFeil,2,LuoTao2,GuoYuan-jing2,LiHong-Huai,MeiYuan-wu2(1Departmentofneurology,WuhanGeneralHospitalofGuangzh
4、ouMilitary430070)(2.Departmentofneurology,WuhanUnionHospital,HuaZhongUniversityofScienceandTechnology430022)[Abstract]Objective:ToobservethechangesinmetabolicmoleculeofBV2microgliaafteritisclassicaloralternativeactivated,andjudgethemethodstoidentifytheMlorM2-typepolarizedmicrogliacel
5、ls.Methods:BV2microgliacellswereclassicalactivatedbyLPSintheconcentrationoflOOng/mlandalternativeactivatedbyIL-4intheconcentrationof20ng/mLfor24hours,thenELISAwasusedtomeasurethelevelsofIL-12,IL-10,TNF-α,andReal-timePCRwasusedtodetectthemRNAexpressionofiNOS,Arg-l,IL-10andIL-12.
6、Flowcytometry(FCM)wasusedtomeasurethelevelsofMMrmembraneprotein•Results:MicrogliapresentedMl-typepolarizationstateafteritwereclassicalactivated,thesecretionofIL-12andTNF-αztheexpressionofiNOSmRNAandIL-12mRNAweresignificantlyincreased,andthedifferenceweresignificantcomparewithth
7、atofinnormalcontrolgroup(P<0.05).MicrogliapresentedM2-typepolarizationstateafteritwerealternativeactivated,andthesecretionofIL-10,theexpressionofArg-lmRNAandIL-lOmRNAweresignificantlyincreased,andthedifferenceweresignificantcomparedwiththatofinnormalcontrolgroup(P<0.05)•Theleve
8、lsofMMrwerea