脐血间充质干细胞分离,培养与鉴定

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1、脐血间充质干细胞分离、培养与鉴定李启明1程天民1李宁2*通信作者粟永萍1冉新泽1(1.第三军医大学全军复合伤研究所，创伤、烧伤与复合伤国家重点实验室2.第三军医大学新桥医院,高压氧中心重庆400038)摘要：目的研究脐带血(HUCB)中含有的间允质干细胞(MSCs)体外分离、培养条件，探讨其作为种子细胞用于下一步实验研究的稳定性与实用性。方法无菌条件下取正常足生产的脐带血，经CPDA-1复合抗凝剂抗凝，然后经过去红细胞处理，再用淋巴细胞分离液分离脐血的单个核细胞，DMEM/F12培养基进行培养和纯化获得贴壁细胞层，采用定向诱导分化方法检测鉴定获得的MSCs结果来源于脐血的单个核细胞经体外培养

2、贴壁后出现形态学上可见间充质样的细胞，问充质样细胞为类似成纤维样的细胞形态，但偏向半月形弯曲状，经过成肌、成脂肪、成神经细胞诱导分化成功后确定为间充质干细胞。结论脐血来源的间充质干细胞能够在体外分离、培养出有用的进一步实验所需要的种子细胞。关键词：脐带血；间充质干细胞；细胞培养；诱导分化鉴定中图法分类号：文献标识码：THEISOLATIONCULTUREANDIDENTIFICATIONOFHUCAMSCsLIQi-ming,CHENTian-minLIning,SUYong-ping,RUANxin-ze(1.StateKeyLaboratoryofTrauma,BurnsandCombi

3、nedInjury,InstituteofCombinedInjury,2.HyperbaricoxygenCenterXinqiaoHospital,ThirdMilitaryMedicalUniversity,Chongqing,400038,China)Abstract:ObjectiveToinvestigatetheisolation，cultivation,differentiationandidentificationofhumanumbilicalcordblood(HUCB)mesenchymalstemcells(MSCs)invitro，andtoobservethef

4、easibilityofusingMSCsasseedcellsinexperimentalfurthermore．MethodsHUCBwerecollectedfromfulltermchildbirthscheduledforCesareansectionandvaginadeliveries,a11specimenswasobtainedsterilelywithPreservative-freeCPDA-1compoundnatriumcitricum.Afterlyseredbloodcell,thecordbloodmononuclearcellwasisolatedbyhum

5、anlymphocyteseparatingmedium.culturewithdulbecco'smodifiedeagle'smedium(DMEM/F12)mediumtoproduceadherentlayerandidentificationwithorientationdifferentiationresults.ResultsTheUCB-derivedmononuclearcells，whensetinculture，gaverisetoadherentcells,whichexhibitedeitherafibroblast-1ikebutcrescent-shapedor

6、bending-shapedmorphologyormesenchymal-likephenotype.ItcanbeaffirmedastheMSCswhichcharacterizedwithmulti-differentiatingpotentialityafterthemyogenic、lipogenicandneurogenicdifferentiation.ConclusionMSCsinHUCBcanbeisolatedandcultivatedinvitro，whichcouldberegardedasanalternativesourceofMSCsforfurtherex

7、perimentaltests。Keywords:umbilicalcordBlood；mesenchymalstemcells；cultureinvitro；differentiationandidentification目前认为，成体干细胞实际上主要是指在不同组织中的间充质干细胞,(MSCs)，它是广泛存在生物体尤其骨髓内的一类干细胞，以前被称为基质干细胞，它最初是由Fnedemteinr发现的易于贴附于