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1、EphB6在小鼠骨髓间充质干细胞成骨分化中的初步研究李松涛1,刘涌1,宋磊1,董世武2,邸宁1,李殿威1,徐源1,周强1(400038重庆,第三军医大学西南医院骨科,全军矫形外科中心)(问题1:同一基因写法请全文前后、图表与文字一致)[摘要]目的探讨EphB6在小鼠骨髓间充质干细胞(bonemarrow-derivedmesenchymalstemcells,BMSC)成骨分化中的作用及其可能机制。方法将BMSC通过双相接种法接种于脱钙骨基质,分为4组:成骨诱导5、14d(A、B组),普通培养5、14d(C、
2、D组),茜素红染色检测钙结节,定量检测各组碱性磷酸酶活性,Runx2、Osterix和EphB6的mRNA表达,采用游离态配体ephrinB1-Fc激活EphB6并重复检测上述指标。结果ALP活性、Runx2和Osterix在A、B组增高,且B组检测到最高的ALP活性[(7.32±2.02)金氏单位/100ml]、Runx2(3.4426±0.2585)倍和Osterix(2.4823±0.3292)倍。EphB6的表达水平在A、B组降低,且在B组最低(0.7543±0.0235)倍。以ephrinB1-Fc
3、激活EphB6后,茜素红染色显示钙结节明显减少,ALP活性显著下降(P<0.05),在高浓度配体作用后ALP活性[(12.20±1.30)金氏单位/100ml]较低浓度配体[(15.40±1.03)金氏单位/100ml]有进一步下降(P=0.008)。配体作用后,BMSC成骨分化中Runx2和Osterix的表达显著下降。结论EphB6通过调控Runx2和Osterix抑制BMSC成骨分化。[关键词]ephrinreceptor;骨髓间充质干细胞;成骨分化;组织工程[中图法分类号][文献标志码]AEffect
4、ofEphB6onosteogenesisofbonemarrowmesenchymalstemcells:apreliminarystudyLiSongtao,LiuYong,SongLei,DongShiwu,DiNing,XuYuan,LiDianwei,ZhouQiang(DepartmentofOrthopaedics,SouthwestHospital,ThirdMilitaryMedicalUniversity,Chongqing400038,China)[Abstract]ObjectiveT
5、odetecttheeffectofEphB6onosteogensisofbonemarrowmesenchymalstemcells(BMSC)anditsmechanism.MethodsBMSCseededdecalcifiedbonematrixviabiphasicseedingweredividedintoosteogenesis5-day(groupA),osteogenesis14-day(groupB),non-induced5-day(groupC),non-induced14-day(
6、groupD).Alizarinredstainingwasperformed.Activityofalkalinephosphatase(ALP),mRNAofRunx2,OsterixandEphB6werequantitated.ThensolubleephrinB1-Fcwasadministratedinallgroupswiththesametestitems.ResultsALP,Runx2andOsterixincreasedingroupAandB.ALP(7.32±2.02King’sUn
7、it/100mL),Runx2(3.4426±0.2585)andOsterix(2.4823±0.3292)reachedthepeakingroupB.EphB6wasdown-regulatedingroupAandB,andwaslowestinB(0.7543±0.0235).AftertheadministrationofsolubleephrinB1-Fc,ALPactivitydecreased(P<0.05),andwaslessinhigh-doseephrinB1(12.20±1.30K
8、ing’sUnit/100mL)comparedwithlow-doseephrinB1(15.40±1.03King’sUnit/100mL),P=0.008.Runx2andOsterixweredown-regulatedinligand-dependentmanner.Alizarinredstainingsuggestedanevidentreductionincalciumnoduswi