小鼠脾脏来源树突状细胞的体外扩增培养

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1、小鼠脾脏来源树突状细胞的体外扩增培养：黎辉，曹宇皎，黄军华，刘俊峰【摘要】目的：对体外诱导小鼠脾脏单个核细胞分化成的树突状细胞(DC)的生物学表型及功能进行检测，并与骨髓的DC进行比较。方法：分离小鼠脾脏单个核细胞，在体外培养的条件下通过添加25ng·ml-1的粒单核细胞集落刺激因子和25ng·ml-1的白细胞介素4(IL4)将其诱导为DC，分别从细胞形态、表型以及功能3个方面对其进行检测，并与骨髓的DC进行比较。结果：小鼠脾脏的DC经磷酸脂多糖(LPS)刺激成熟后其表面显示出明显的树枝状突起，高表达CD11c、CD86及MHCⅡ类分子，并具有极强的刺激同种异

2、基因淋巴细胞增殖的能力，其特征与骨髓的DC相差无几。结论：小鼠脾脏单个核细胞能够被诱导为具有正常形态、表型、功能的DC，为DC的功能研究以及进一步制备相应的肿瘤疫苗提供又一可靠。【关键词】脾脏;骨髓;树突状细胞;诱导;小鼠　Dendriticcells(DC)arethemostpotentantigenpresentingcellsthatareresponsibleforprimingnativeTcellsintheimmuneresponse.ImmunotherapyofDC/tumorvaccinehasbeeanimportanttherapeutics

3、trategyagainsttumors[12].Hoitedbytheloarroousespleenmononuclearcellsintodendriticcellsinvitro,andthenparedthebiologicalcharacteristicsofthemarroononuclearcellsderivedDCs.　　1Materialsandmethods　　1.1Materials　　Mainreagents:rebinantmousegranulocytemacrophagecolonystimulatingfactor(GMCS

4、F),rebinantmouseinterleukin4(IL4),rebinantmouseleukemiainhibitoryfactor(LIF,Peprotech),lipopolysaccharide(LPS,Sigma);mitomycinC(MMC)andmethabenzthiazuron(MTT,Duchfo).DCsculturemedium:highglucoseDMEMplus15%fetalcalfserumplus25mg·L-1rmGMCSFandrmIL4.Mice:sixtoeightaleBALB/cand129miceAn

5、imalCenterofGuiyangMedicalCollege.　　1.2Methods　　1.2.1InductionofDCfrommousespleenCellsice.Mononuclearcellsattheinterfacepholytegradient.Theyl-1.Thesuspendedcellsovedandtheadherentcellsedium.LPSetimemousebonemononuclearcellsicroscopedailyandsuspendedcellsicroscopy.　　1.2.3Cellsurfacemarke

6、ranalysisCellsin.Afterbeingetry.　　1.2.4Mixedlymphocytereactions(MLR)CollectedspleenderivedDCl-1MMCasastimulatorfor45minandplatesatvariousratiosfor4d.Atthesametime,ulators.4hbeforeharvesting,cellsg·ml-1),thenaddedeasured.Thestimulationindexula:SI=absorbencyofstimulationgroupabsorbencyof

7、contrast/theabsorbencyofcontrast.　　1.3Statisticalanalysis　　Dataareexpressedas±sdeviation.TheresultsoftheanalysisousespleenderivedDCs　　Atthefirst2-4d,themousespleenmononuclearcellsgreall,thepopulationexpandedrapidlyeandthenthebulkbecamelargergradually.ImmaturespleenderivedDCsalr