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1、全反式维甲酸增强骨形态蛋白9诱导间充质干细胞成骨分化作用研究黄帆1,刘映孜2,杨秋珺2,周龙洋2,周岐新2,罗进勇3,何百成2,邓忠良(400010重庆,重庆医科大学附属第二医院骨科1;400016重庆,重庆医科大学药理教研室,重庆医科大学检验学院2)[摘要]目的研究全反式维甲酸(ATRA)对骨形态蛋白9(BMP9)诱导间充质干细胞成骨分化的影响及可能机制方法利用组织化学染色检测处理后第5、7天碱性磷酸酶活性,Westernblot方法检测第9、11天骨桥素蛋白表达水平及Smad1/5/8磷酸化水平,茜素红染色检测第14、20天钙盐沉积,萤光素酶报告质粒检测BM
2、PR-Smad信号通路的活化程度。结果ATRA及BMP9均诱导C3H10T1/2细胞碱性磷酸酶活性升高,但ATRA合并BMP9组碱性磷酸酶活性明显强于ATRA或BMP9组;Westernblot结果显示,BMP9合并ATRA组骨桥素蛋白表达水平高于BMP9组;BMP9合并ATRA组钙盐沉积染色也明显于BMP9组。BMP9合并ATRA组BMPR-Smad报告质粒萤光素酶活性明显强于BMP9组(P<0.01),Samd1/5/8磷酸化水平也呈相同变化。结论ATRA能增强BMP9诱导的间充质干细胞成骨分化,其机制可能与ATRA促进Smads信号转录活性有关。[关键词]
3、全反式维甲酸;骨形态蛋白9;干细胞;Smad信号[中图法分类号]R681.1[文献标志码]A[通信作者]邓忠良,电话:(023)63693558,E-mail:deng7586@gmail.comResearchontheeffectofall-transretinoicacidenhancetheosteogenicdifferentiationinducedbyBMP9inmesenchymalstemcellsHuangFan1,LiuYingzi2,YangQiujun2,ZhouLongyang2,ZhouQixin2,LuoJinyong2,HeBa
4、icheng2,DengZhongliang1(1DepartmentofOrthopaedics,theSecondAffiliatedHospitalofChongqingMedicalUniversity,Chongqing,400010;2DepartmentofPharmacology,ChongqingMedicalUniversity,Chongqing,400016,China)[Abstract]ObjectiveToinvestigatetheeffectofall-transretinoicacid(ATRA)onosteogenesisi
5、nducedbybonemorphogeneticprotein9(BMP9)inmesenchymalstemcellandtodissectthepossiblemechanismunderlayingthiseffect.MethodsThisresearchemployedhistochemicalstainingtotestthealkalinephosphatase(ALP)activitieswhentreatedtheC3H10T1/2cellswithBMP9and/orATRAfor5or7days.Then,evaluatedtheexpr
6、essionlevelofosteopontinbyWesternblotonD9andD11,sodidthemineralizationwithAlizarinRedStainingonD14andD20.Finally,analyzedtheactivationofBMP-SmadsignalingwithBMPR-SmadbindingsiteluciferasereporterassayandWesternblottodeterminethephosphorylationlevelofSmad1/5/8.ResultsTheactivitiesofAL
7、PareincreasedbyBMP9and/orATRA,buttheALPactivitiesofBMP9combinewithATRAgrouparemorepronouncedthanthatofBMP9orATRAalone.ATRApotentiatestheosteopontinexpressionandthemineralizationinducedbyBMP9inC3H10T1/2cells.ATRApromotesthephosphorylationlevelofSmad1/5/8andincreasestheluciferaseactivi
8、tyofBMPR-Sma