腺相关病毒及慢病毒载体对骨髓间充质干细胞基因转染效率de比较.pdfVIP

中国康复理论与实践 2014~1El2月第20卷第 12期ChinJRehabilTheoryPract，Dec．，2014，Vo1．20，No．12 — 1l17 一 DOI：10．3969／j．issn．1006—9771．2014．12．005 · 基础研究 · 腺相关病毒及慢病毒载体对骨髓间充质干细胞基因转染效率的比较 苏玉金，赵育梅 ，顾漪 摘【要l 目的 比较腺相关病毒 (w)载体和慢病毒(LV)载体在间充质干细胞(MSCs)基因转染中的效率。方法 密度梯度离心 法分离培养MSCs，HE染色、Nestin免疫荧光染色鉴定，BrdU标记观察增殖情况 。分别包装AAV与LV假病毒颗粒，并感染 MSCs，通过B-gal染色和绿色荧光蛋白检测，分别计算两者的转染效率。结果 MSCs成功从骨髓分离。AAV载体介导的基因转染 率为49．1％，LV载体的转染率为91．4％(：0．01)。结论 L、r载体介导的基因转染方法转染效率更高。 I关键词l基因转染；腺相关病毒载体；慢病毒载体；骨髓间充质干细胞 EfficiencyofGeneTransfectionwithAdeno．associatedViralVectororLentiviralVectorinBoneM arrow DerivedMesenchymal Stem Cells SU ，—fin．ZHAO Yu—mei,GU ，】i．BejiingNeurosurgicalInstitutG TiantanHospital,CapitalMedicalUniversity,Bejiing 100050,China Abstract：ObjectiveTocompare2kindsofcommonlyusedviralvectors，adeno-associatedviral(AhV)vectorandlentiviral(LV)vec— torinthegenetransfectionforbonemarrowderivedmesenchymalstemcells(MSCs)．MethodsMSCswereisolatedwithdensitygradient (1ymphocytesseperation)andidentifiedwithHEstainingandimmunocytochemistorystainingforNestin．TheproliferationofBMSCswas detectedwithBrdU labeling．AAV mediatedgenetransfectionwascarriedoutthroughrecombinantAAV-LacZviralparticles．ForLV medi— atedgenetransfection，theLVparticleswereuseddirectly．ThetrnasfectionefficiencywasestimatedwithB—galstainingnadgreenfluores- centproteinunderthefluorescentmicroscoperespectively．ResultsMSCswassuccessfullyisolatedfrom thebonemarrow．HE staining showedthatMSCswaswithbignucleus，1-3nucleoli，andhighnucleocytoplasmicratio．BrdU labelingsuggestedthatMSCswereprolifer— ating．SomeMSCsexpressedNestin．ThegenetransfectionefficiencymediatedwihtAAVvectorwas49．1％．anditwas91．4％withLVvec— torfP0．O1)．ConclusionTheLVvectorismoreefficientongenetransfectionthanAAV vector． Keywords：genetransfection；adeno—associatedviralvector；lentivi