摘要
目的:探讨骨形态发生蛋白2(BMP2)诱导鼠胚胎间充质干细胞C3H10T1/2向成软骨分化的可能分子机制。方法:20μg/mL BMP2诱导C3H10T1/2细胞0,4 h,1,3,6,10,13,15 d后,RT-PCR检测BMP信号通路中关键分子BMPRⅠ,BMPRⅡ,Smad1/5/8的表达,Western blot检测smad蛋白及MAPK信号通路中p38磷酸化水平变化,QRT-PCR检测成软骨标志基因aggrecan,Col2a1以及成软骨相关转录因子FGFR3,Sox9表达水平,同时用Alcine Blue染色,观测C3H10T1/2细胞成软骨分化情况。结果:经BMP2诱导后,C3H10T1/2细胞表现出成软骨分化,smad蛋白及p38磷酸化水平有所上升,同时成软骨标志基因Col2a1以及成软骨相关转录因子FGFR3,Sox9表达水平都有增加。与未诱导组相比,Col2a1,FGFR3,Sox9在3 d的表达与对照组相比差异有统计学意义(P<0.05),FGFR3,Sox9在6 d的表达与对照组相比差异有统计学意义(P<0.05)。结论:在一定培养条件下,BMP2具有诱导C3H10T1/2细胞成软骨分化能力。
Objective： To investigate the possible mechanism by which bone morphogenetic protein2 （BMP2） induces chondrogenie differentiation of murine embryonic mesenchymal stem cell C3H10T1/2. Methods： C3 H10T1/2 cells were induced with 20 μg/ml BMP2 for 0,4 h, 1,3,6,10,13 and 15 d, respectively. RT-PCR was employed to detect BMPRI, BMPRII and Smadl/5/8 （key molecules in the BMP signaling pathway） ,Western blot assay to examine the phosphorylation levels of phosphorylated Smad proteins and p38 （of the MAPK signaling pathway ） , and QRT-PCR to detect the expression levels of chondrogenic marker genes （ aggrecan and Col2al ） and ehondrogenesis related transcription factors （ Sox9 and FGFR3 ）. Furthermore, the C3HIOTI/2 cells were stained with Aleine Blue and studied for status of chondrogenie differentiation. Results： After BMP2 induction,C3H10T1/2 cells showed ehondrogenic differentiation,increased phosphorylation levels of smad protein and p38, and also elevation in expression levels of chondrogenic marker gene Col2al and chondrogenesis related transcription factors Sox9 and FGFR3. The expressions of Col2al, Sox9 and FGFR3 in C3H10T1/2 on day 3 were statistically different from those in controls. Conclusion： Under certain cuhure conditions, BMP2 appears potent in inducing chondrogenic differentiation of C3H10T1/2 cells.
出处
《广州医学院学报》
2012年第2期11-15,共5页
Academic Journal of Guangzhou Medical College
