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,WuhanGeneralHospitalofGuangzhouMilitary430070)(2.Departmentofneurology,WuhanUnionHospital,HuaZhongUniversityofScienceandTechnology430022)[Abstract]Objective：ToobservethechangesinmetabolicmoleculeofBV2microgliaafteritisclassicaloralternativeactivated,andjudgethemethodstoidentifytheMlorM2-typepolarizedmicrogliacells.Methods：BV2microgliacellswereclassicalactivatedbyLPSintheconcentrationoflOOng/mlandalternativeactivatedbyIL-4intheconcentrationof20ng/mLfor24hours,thenELISAwasusedtomeasurethelevelsofIL-12,IL-10,TNF-α,andReal-timePCRwasusedtodetectthemRNAexpressionofiNOS,Arg-l,IL-10andIL-12.Flowcytometry(FCM)wasusedtomeasurethelevelsofMMrmembraneprotein•Results：MicrogliapresentedMl-typepolarizationstateafteritwereclassicalactivated,thesecretionofIL-12andTNF-αztheexpressionofiNOSmRNAandIL-12mRNAweresignificantlyincreased,andthedifferenceweresignificantcomparewiththatofinnormalcontrolgroup(P<0.05).MicrogliapresentedM2-typepolarizationstateafteritwerealternativeactivated,andthesecretionofIL-10,theexpressionofArg-lmRNAandIL-lOmRNAweresignificantlyincreased,andthedifferenceweresignificantcomparedwiththatofinnormalcontrolgroup(P<0.05)•ThelevelsofMMrwerealsoincreasedsignificantlymeasuredbyFlowcytometry(FCM).Conclusion：BV2microgliacouldpresentMl-typepolarizationstateafteritwereclassicalactivated,M2-typepolarizationstateafteralternativeactivated.Thetwotypepolarizationcouldbeidentifieddetectedfromthreeaspectsincludinginflammatoryfactors,argininemetabolicmolecularandspecificcellmembraneproteins.【Keywords】microglia,classicalactivation，alternativeactivation，M1/M2typepolarization,lipopolysaccharide,interleukin-4小胶质细胞是中枢神经系统内重要的固奋免疫细胞，也是介导免疫反应的主要效应细胞[1】。0前研究证实小胶质细胞来源于单核吞噬细胞系统，因此能够表现出巨噬细胞的很多生物学特征，比如被各种内源性及外源性病理刺激所激活、具右特异性的巨噬细胞表面抗原等等。巨噬细胞的激活途径主要包括经典激活途径和替代激活途径［2L经典激活的巨噬细胞为Ml型巨噬细胞，主要分泌大量促炎因子参予对病原体清除及疾病的炎性损伤过程，而替代激活的巨噬细胞为M2型巨噬细胞，它主要干预炎症的进展、修复...
