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Aug.2009,Volume6,No.8(SerialNo.57)JournalofUS-ChinaMedicalScience,ISSN1548-6648,USA1511221.2625002.400038MSCs30MSCsAd-EGFP124812BrduBrdu124812124812BrduMSCsMSCsTheviabilityandproliferationofallograftedmesenchymalstemcellsintheintervertebraldiscLIHua-zhuang,FANGJun,ZHOUYue,WANGWei-dongAbstract:ObjectiveToobservetheviabilityandproliferationofallograftedmessenchymalstemcellsintheintervertebraldisc.MethodsBonemesenchymalstemcellsisolatedfrom12JapanesewhiterabbitswerelabeledwithAd-EGFPandwereallograftedintolumbarintervertebraldiscofwhichthenucleuspulposushadbeenaspirated.Thelumbarspinewereextracted1,2,4,8,12weeksaftertheoperation,dealedwithBrdu,theviabilityandproliferationofthecellswereassessedwithconsecutivesectionsusingfluorescentmicroscopeandimmunohistochcmistryofBrdu.ResultsFluorescencewasobservedinthespecimensfor1,2,4,8,12weeksaftertheoperationandpositiveimmunohistochemicalstainingforBrduwasobservedinthespecimensfor1,2,4,8,12weeksaftertheoperation.ConclusionAllograftedmesenchymalstemcellscansurviveandproliferateintheintervertebraldisc.ResultsfromourstudyprovideexperimentalevidenceforMSCsasseedingcellsofintervertebraldisctissueengineering.Keywords:intervertebraldisc;marrowmsenchymalstemcells;allograft;viability;proliferation【】1970-MesenchymalStemCellsMSCs16MSCsMSCsMSCs1.1.12301.3kg6MSCs1248121.21.2.1MSCs23mm185ml3000u/ml0.1ml2ml10%FBSDMEM1.073g/mlPercoll900g4℃15minPBS325cm210%100u/ml0.1mg/mlDMEM37°C5%CO24812151.2.2Ad-EGFPMSCs75cm250%70%200pfu/cellAd-EGFP37℃5%CO2410%DMEM24DMEM1.2.3MSCsMSCsAd-EGFP481×106MSCs0.2cm×0.5cm×0.5cmMSCs/37℃5%CO242110mlL2-L3L3-L4L4-L5MSCs/1.2.41248120.3mg/mlBrdu37℃1h5μ)Brdu2.2.1MSCsMSCs2d7d1214d1MSCs24h10d2.2Ad-EGFPAd-EGFP4890%MSCs22.31248123Brdu417114dMSCs×1002Ad-EGFP4890%MSCs×1003Ad-EGFPMSCs12EGFPA12EGFPB184BrdU1A×20012B×2003.MSCs[1]MSCsMSCsMSCs[2]MSCs[3-5]Sakai[6]Melrose[7]MSCs[6][8][9-10]8[11-12]MSCsMSCsBrdusMSCsMSCs[1]CaplanA.I.Mesenchymalstemcells.JOrthopRes,1991,9:641-650.[2]PittengerM.F.,MackayA.M.,BeckS.C.,etal.Multilineagepotentialofadulthumanmesenchymalstemcells.Science,1999,284(5411):143-147.[3]MeinelL.,HofmannS.,KarageorgiouV.,etal.46Engineeringcartilage-liketissueusinghumanmesenchymalstemcellsandsilkproteinscaffolds.BiotechnolBioeng,2004,88(3):79-391.[4]BAIX.,XIAOZ.,PANY.,etal.Cartilage-derivedmorphogeneticprotein-1promotesthedifferentiationofmesenchymalstemcellsintochondrocytes.BiochemBiophysResCommun,2004,325(2):453-460.[5]LIW.J.,TuliR.,OkaforC.,etal.Athree-dimensionalnanofibrousscaffoldforcartilagetissueengineeringusinghumanmesenchymalstemcells.Biomaterials,2005,26(6):599-609.[6]SakaiD.,MochidaJ.,YamamotoY.,etal.TransplantationofmesenchymalstemcellsembeddedinAtelocollagen®geltotheintervertebraldisc:apotentialtherapeuticmodelfordiscdegeneration.Biomaterials,2003,24(20):3531-3541.[7]MelroseJ.,SmithS.,GhoshP.Assessmentofthecellularheterogeneityoftheovineintervertebraldisc:comparisonwithsynovialfibroblastsandarticularchondrocytes.EurSpine,2003,12(1):57-65.[8].[J].2005255316-318.[9]EdelmanE.R.Vasculartissueengineering:Designerarteries.CircRes,1999,85(12):1115-1117.[10]LangerR.,VacantiJ.P.Tissueengineering[J].Science,1993,260(5110):920-926.[11]BrodinH.Pathsofnutritioninarticularcartilageandintervertebraldiscs.ActaOrthopScand,1955,24(3):177-183.[12]OgataK.,WhitesideL.A.1980Volvoawardwinnerinbasicscience.Nutritionalpathwaysoftheintervertebraldisc.Anexperimentalstudyusinghydrogenwashouttechnique.Spine,1981,6(3):211-216.JaneChen