Smurf1拮抗BMP-2对C2C12细胞的诱导成骨作用

目的研究Smad泛素化调节因子1（Smurf1）对基因重组人骨形态发生蛋白2（rhBMP-2）诱导成骨作用的影响。方法通过将质粒pcDEF3-Smurf1临时转染C2C12细胞并应用实时PCR和Western免疫印迹法鉴定其表达Smurf1，空载体pcDNA3．1作为阴性对照；然后用比色法测定细胞碱性磷酸酶活性和实时PCR分析细胞碱性磷酸酶、骨钙素、成骨转录因子Osterix基因表达，观察Smurf1对rhBMP-2诱导C2C12细胞向成骨细胞分化的影响。同时在相差显微镜下观察细胞形态的变化。结果Smurf1在C2C12细胞可以通过临时转染获得很好的表达。尽管在细胞形态上没有明显的差别，但Smurf1转染的C2C12细胞在rhBMP-2作用48h后，细胞碱性磷酸酶活性以及细胞碱性磷酸酶、骨钙素、成骨转录因子Osterix基因表达均较对照组明显下降。结论Smurf1在BMP诱导成骨过程中发挥着负性调节作用。

Smurf1 inhibits osteoblast differentiation of C2C12 induced by BMP-2

Purpose To determine the effect of Smad ubiquitination regulatory factor 1(Smurf1) on osteoblast differentiation of C2C12 induced by recombinant human bone morphorgenetic protein 2(rhBMP-2). Methods C2C12 cells were transiently tansfected by pcDEF3-Smurf1 plasmid and pcDNA3.1 as control.Smurf1 expression was demonstrated by real-time PCR and Western blot analysis.The effect of Smurf1 on osteoblast differentiation of C2C12 induced by rhBMP-2 was checked by alkaline phosphatase activity and gene expression of alkaline phosphatase,osteocalcin and osterix using real-time PCR methods.Morphological changes were also checked under phase-contrast microscope. Results Smurf1 was well expressed in C2C12 cell by transiently transfection.Although no significant cell morphological changes could be seen,Alkaline phosphatase activity and gene expression of alkaline phosphatase,osteocalcin and osterix were significantly decreased in Smurf1 transfected C2C12 cells 48 hours after rhBMP-2 incubation. Conclusions Smurf1 inhibits osteoblast differentiation of C2C12 induced by BMP-2.