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1、无血清体外培养树突状细胞的研究作者：张怀东，宋振岚，李伟平【摘要】本研究的冃的是建立对外周血来源的树突状细胞(dendriticcell,DC)的无血清培养方案。以含胎牛血清(FCS)、人AB血清的培养液作为对照。分离健康志愿者外周血单个核细胞(PBMNC),在不同培养液中分别加入GM-CSF(100ng/ml)>IL-4(500U/ml)培养6天,再分别加入钙离子载体A23187(100ng/ml)继续培养24小时,于倒置显微镜下观察细胞形态，流式细胞术分析细胞表型，MTT比色法检测各组DC刺激同种异体外周血T细胞增殖的能力和DC刺激的T细胞对K562细胞的杀伤作用。结果表明：
2、无血清培养组培养出典型形态的DC,其表面CD14分子的表达明显减少，CD83、HLA-DR、CDwl23分子的表达明显增高，具有明显的刺激同种异体T细胞增殖的能力和DC刺激的T细胞对K562细胞的杀伤作用。无血清组与两个含血清的对照组比较,组间无显著性差异(P>O・05)o结论：无血清培养液培养的DC与含血清的培养液培养的DC相比，无显著性差异。DC无血清培养具有临床应用前景。【关键词】树突状细胞；无血清培养液；钙离子载体；外周血TnVitroCultivationofDendriticCel1swithSerum-freeMediumAbstractThisstudywa
3、saimedtoinvestigatetheprotocolinvitrotoincubatethedendriticcell(DC)derivedfromperipheralbloodmonocytesusingserum-freemediumX-VIV020.Peripheralbloodmonocytesfromhealthydonorsweretreatedwith1OOng/mlGM-CSFand500U/mlTL~4,respectively.Aftercultivationfor6days,theyweretreatedwithlOOng/mlcalciumiono
4、phoreA23187・Aftercultivationfor24hoursthecellularmorphologywasobservedunderinvertmicroscope,thesurfacemarkerswereanalyzedbyflowcytometry,theproliferationofallogeneticTcel1swasdetectedbyMTTcolorimetry,thespecificcytotoxicityofTcellsprimedwithDCwasexaminedbyMTTassay.Theresultsshowedthatinal1thr
5、eegroupswithserum-free,fetalcalfserum(FCS)andhumanABserummediums,cellsdisplayedcharacteristicmorphologicalfeaturesofDC・SimultaneouslyCDldexpressionwasdecreased,andCD83,HLA-DRandCDwl23expressionwereincreasedonthesecel1s.Tnaddition,DCsculturedwiththesemethodscouldevidentlystiniiilatetheprolifer
6、ationofal1ogeneticTcel1・Ascomparedwiththetwocontrolsofserumcontaininggroups,theculturedcellsintheserum-freegroupsshowedalmostthesameallo-stimulatorycapabilityandcellularmorphologyandsurfacemarkers,andTlymphocytesprimedwiththeculture~derivedDCexhibitedthesimilarkillingactivitytoK562(P>0.05)
7、.ItisconcludedthatthereisnosignificanceinDCnumbers,morphology,epitopeandabilitytostimulatetheproliferationofallogeneticTcellsbetweenDCinducedbyseruin-freeX-VTV020mediumandDCinducedbyserum-containedmedium・DCculturedandinducedbyserum-freemediui
