Preparation-characterization-of-bmsc-cell-derived-

Preparationandcharacterizationofbonemarrowmesenchymalstemcell-derivedextracellularmatrixscaffoldsYanXu,1Guang-yueXu,2ChengTang,1BoWei,1XuanPei,3Jian-chaoGui,1Byoung-HyunMin,4,5Cheng-zheJin,1Li-mingWang11DepartmentofOrthopedics,AffiliatedHospitalofNanjingMedicalUniversity(NanjingFirstHospital),Nanjing,China2DepartmentofOrthopedics,NanjingDrumTowerHospital,AffiliatedHospitalofNanjingUniversityMedicalSchool,Nanjing,China3SchoolofBiologicalScienceandMedicalEngineering,SoutheastUniversity,Nanjing,China4CellTherapyCenter,AjouUniversitySchoolofMedicine,Suwon,Gyeonggi,Korea5DepartmentofOrthopaedicSurgery,AjouUniversitySchoolofMedicine,Suwon,Gyeonggi,KoreaReceived9November2013;revised11May2014;accepted28May2014Publishedonline4July2014inWileyOnlineLibrary(wileyonlinelibrary.com).DOI:10.1002/jbm.b.33231Abstract:Theaimofthisstudywastoassessthefeasibilityofcreatingextracellularmatrix(ECM)scaffoldsfrommesenchy-malstemcells.Bonemarrowmesenchymalstemcell(BMSC)-derivedECM(BMSC-dECM)scaffoldswerefabricatedbylyophilizationaftercrosslinking,withoutusingadecellulariza-tionprocess.Acellularporcinechondrocyte-derivedECM(AC-dECM)scaffoldswereusedasacontrol.Thesurfacemorphol-ogy,internalstructure,wateruptakeratio,mechanicalproper-ties,andbiocompatibilityofthescaffolds,aswellastheinvitrobehaviorofcellsgrownonthescaffoldswereexaminedandcomparedbetweenthetwoscaffoldtypes.FortheBMSC-dECMscaffolds,theaverageporesizewas304.46108.2lm,averageporositywas93.3%64.5%,averagecompressivemoduluswas6.861.5kPa,andaveragewateruptakeratioexceeded20.TheBMSC-dECMscaffoldssupportedtheinvitroattachmentandproliferationofcells,withtheseaspectslikelybeingcomparabletothoseoftheAC-dECMscaffolds.ThefindingsofthispreliminarystudyhighlightthepotentialutilityofBMSC-derivedECMscaffoldsforfuturecartilagetissue-engineeringapplications.VC2014WileyPeriodicals,Inc.JBiomedMaterResPartB:ApplBiomater,103B:670–678,2015.KeyWords:autologousmesenchymalstemcells,extracellu-larmatrix,porousscaffold,tissueengineeringHowtocitethisarticle:XuY,XuG-Y,TangC,WeiB,PeiX,GuiJ-C,MinB-H,JinC-Z,WangL-M.2015.Preparationandcharacterizationofbonemarrowmesenchymalstemcell-derivedextracellularmatrixscaffolds.JBiomedMaterResPartB2015:103B:670–678.INTRODUCTIONScaffoldsplayimportantrolesintissuerepairandregenera-tion.1Desirablepropertiesofscaffoldsincludeporosity,bio-compatibility,biodegradability,andadequatemechanicalstrength.2Extracellularmatrix(ECM)-basedscaffoldshaveshowngreatpotentialforconstructingmanycomplextissuesinpreclinicalstudiesandclinicaltrials.3ECMcanbederivedfromavarietyoftissuesandorgans,suchastheskin,4ten-dons,5spinalcord,6andliver.7ECM-basedscaffoldsarecom-posedofacomplexmixtureofmolecules,suchascollagenandglycosaminoglycans,whichmediatethestructuralandbiologicalpropertiesofthescaffold.8,9Theycontainvariousgrowthfactors,includingtransforminggrowthfactor-b(TGF-b),10,11basicfibroblastgrowthfactor,11,12andvascularendothelialgrowthfactor,13whichfurtherpromotethecellu-larbehaviorsneededforoptimaltissuefunctioning.3ThelimitedavailabilityofautologoustissuesourcesisalimitationtoharvestingadequatequantitiesofECM.Usingallogeneicorxenogeneictissuesmayresultinadverseinflam-matoryresponsesorovertimmune-mediatedrejection.14–16Tominimizeorpreventtheseundesirableimmunologicresponses,adecellularizationprocessmustbeperformedtoremovetheantigenicepitopes.3Thethree-dimensionalstruc-tureoftheresultingECMmimicsthatoftheoriginatingtissue.17Commondecellularizationproceduresinvolveacom-binationofphysicalandchemicaltreatments,2,6whichreducethemechanicalandstructuralintegrityoftheECM.18,19Toretainorimproveitsmechanicalproperties,acellularECMmustbesubjectedtochemicalcrosslinking.20Forinstance,the1-ethyl-3-(3-dimethylaminopropyl)carbodiimide(EDC)/N-hydroxysuccinimide(NHS)systemcanbeusedtocrosslinkcollagen-basedmaterialsefficiently,withoutleavingcytotoxicresidues.21,22Moreover,theEDC/NHSsystemiswatersolu-ble,whichfacilitatesremovalofresidualreagents.23Bonemarrow(BM)-derivedmesenchymalstemcells(BMSCs)arecapableofsecretingECMwhenculturedinvitro.Becauseoftheirself-renewalandproliferativeabilities,24,25mesenchymalstemcells(MSCs)theoreticallyrepresentanCorrespondenceto:L.-M.Wang(e-mail:[e-mail])andCZJin(e-mail:[e-mail])Contractgrantsponsor:NationalScienceFoundationofChina;contractgrantnumber:31070861670VC2014WILEYPERIODICALS,INC.unlimitedsourceforECMscaffolds.BMSCscanbeextractedwithoutsacrificingthedonoranimal,andautologousBMSCscanbeobtainedfromhumanpatients.26Theriskofimmuno-logicalrejectionofautologousBMSCsisminimal,26anadvantagethatfurthercontributestothetherapeuticpoten-tialofBMSC-derivedECMscaffolds(BMSC-dECMscaffolds).Wepreviouslydevelopedauniqueproceduretoderiveacellularporcinechondrocyte-derivedECM(AC-dECM)scaf-folds.27,28Thesescaffoldsweremadefromporcinechondro-cytesusingauniqueproceduredevisedinourlaboratoryand,inouropinion,closelymimickedthenaturalcartilageenvironment.Inthisreport,weutilizedadifferentstrategytogenerateBMSC-dECMscaffolds.Specifically,ouraimwastoassessthefeasibilityofcreati