简介：Rats(Rattusnorvegicus)havemanyadvantagesovermiceinscientificstudies,forexample,theyaremorerelevanttohumaninphysiologicalandpharmacologicalresponses.Therefore,ratsarebroadlyusedinexperimentalstudies.Therecentbreakthroughinthegenerationofratembryonicstemcells(rESCs)opensthedoortoapplicationofgenetargetingtocreatemodelsforthestudyofhumandiseases.Inaddition,theinvitrodifferentiationofrESCsintoderivativesofthreegermlineswillserveasapowerfultoolandresourcefortheinvestigationofmammaliandevelopment,cellfunction,tissuerepair,anddrugdiscovery.However,thedistinctcultureconditionandsignalinhibitor-dependedmaintenanceofrESCsstandasaconsiderablechallengeforitsinvitrodifferentiation.Toaddressit,weinvestigatedwhetherrESCsarecapableofformingterminaldifferentiatedcardiomyocytes.Wefoundthattheembryoidbodies(EBs)-basedmethodusedinmouseESC(mESC)differentiationfailedtoworkinthecultivationofrESCs.WethenmodifiedthedifferentiationprotocolandsuccessfullydevelopedaninvitrodifferentiationsystemtodifferentiaterESCsintothreeembryonicgermlayers.Byusingthismethod,therESCsformspontaneousbeatingcardiomyocyteswiththepropertiessimilartothosederivedfromfetalratheartsandmESCs.Thisuniquecellularsystemwillprovideanewapproachtostudytheearlydevelopmentandcardiacfunctionaswellastoperformpharmacologicaltestandcelltherapystudy(Grants:theStateMajorResearchProgramofChina(2009ZX09503-024,2010CB945603)andCAS(XDA01030000).

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简介：BackgroundOurpreviousstudyshowedthe150mg/mLfetalcardiacsupernatant(FCS)couldinducedifferentiationofBMSCsintocardiomyocye-likecellswithoutcardiomyocytetouch,butdifferentiationefficiencyisnothighenough.Inhibitionofglycogensynthasekinase-3enhancedtheproliferationandsurvivesofstemcells.Wetestedif6-bromoindirubin-3-oxime(BIO,glycogensynthasekinase-3inhibitor)enhancestheeffectsofFCSondifferentiationofBMSCsandexplorethegrowthfactorsinFCS.MethodsBMSCswereisolatedfromthefemurandtibiaoffour-week-oldmaleSprague-Dawleyratsandco-culturedwithFCS(150mg/mL)thatwasmadefromfetalheartsfromnineteen-daypregnantWistarrats.BIOwithdifferentconcentration(0,1,10,and100nM)wasintroducedinculturedishes.Transforminggrowthfactorbeta1(TGF-β1),bonemorphogeneticprotein2(BMP-2)andAktincardiacsupernatantandculturemediumwereassayedwithELISAmethods.ResultsAfterco-culturingwithFCS,beatingmyotubeswereobservedin25.9%BMSCsdishesafter1to2weeks’culture.ThelevelsofTGF-β1andBMP-2inFCSconcentrationswerenomorethanthatinyoungandadultcardiacsupernatant.AllBIOgroupssignificantlyenhancedtheeffectsofFCSondifferentiationofBMSCsintothecardiomyocyte-likecells(1nM,83%;10nM,73%;100nM,100%).AktlevelswerehigherinBMSCsculturalmediumwithFCS.ConclusionsFCScouldinducethedifferentiationofBMSCsintothecardiomyocyte-likecells.TGF-β1andBMP-2mightnotplayaroleinthedifferentiationofBMSCsinducedbyFCS.BIOenhancedtheeffectsofFCSonthedifferentiationofBMSCsintocardiomyocyte-likecells,whichmightinvolvetheAktpathway.