mTORC1信号促进前成骨细胞MC3T3-E1的分化成熟  

mTORC1 signaling promotes osteoblast differentiation of preosteoblast MC3T3-E1

作　　者：张婧婧[1] 张楠[2] 吴伟[3] 王鹏[3] 杨景瑞[1] 段巨洪[1] 于海川[1] 

机构地区：[1]河南省新乡医学院医学检验学院和分子诊断与医学检验技术河南省协同创新中心,新乡453003 [2]河南省新乡医学院,新乡453003 [3]河南省新乡医学院第三附属医院,新乡453003

出　　处：《中国免疫学杂志》2016年第11期1573-1577,共5页Chinese Journal of Immunology

基　　金：国家自然科学基金项目(No.31301135)

摘　　要：目的:研究mTORC1信号对前成骨细胞MC3T3-E1向成骨细胞分化成熟的调控作用。方法:通过向MC3T3-E1转染pcDNA3.1-Raptor,对mTORC1信号相关蛋白Raptor进行过表达。向MC3T3-E1转染Raptor siRNA,对mTORC1信号蛋白Raptor进行基因沉默。通过Real-time PCR方法测定Raptor的基因表达,通过蛋白免疫印迹法测定Raptor蛋白水平,并通过茜素红染色检测成骨矿化情况,以测定成骨分化程度。通过Real-time PCR检测成骨分化指标的基因表达。结果:与对照组相比,Raptor过表达组的Raptor mRNA和蛋白水平明显增加;茜素红染色结果显示Raptor过表达组染色更深,说明成骨矿化程度更高;荧光定量PCR结果显示,Raptor过表达组的成骨分化标记基因以及成骨转录因子的表达量均高于对照组。与对照组相比,Raptor siRNA组的Raptor mRNA和蛋白水平明显降低;茜素红染色结果显示Raptor siRNA组染色更浅,说明成骨矿化程度更低;荧光定量PCR结果显示,Raptor siRNA组的成骨分化标记基因以及成骨转录因子的表达量均低于对照组。结论:mTORC1信号促进前成骨细胞MC3T3-E1向成骨细胞分化成熟。Objective： To investigate the regulatory role of mTORC1 signaling on osteoblast differentiation from preosteoblast cell MC3T3-E1. Methods： To overexpress the gene Raptor,which was a critical component of mTORC1 signaling,pc DNA3. 1-Raptor was transfected to preosteoblast cell MC3T3-E1. The Raptor overexpression was confirmed by Real-time PCR and Western blot. For the gene silence of Raptor,Raptor si RNA was transfected to preosteoblast cell MC3T3-E1. Alizarin red staining was used to detect the mineralization of MC3T3-E1. Real-time PCR was used to detect the gene expression of osteoblast specific marker genes. Western blot was used to detect the protein level of Raptor. Results： Compared to the control group,the levels of Raptor m RNA and proteins increased significantly in the Raptor overexpression group; the Alizarin red staining results revealed darker staining in the Raptor overexpression group,suggesting higher level of mineralization; the Real-time PCR results showed that the expression of osteoblast differentiation marker genes and osteoblast transcription factors in the Raptor overexpression group was higher than the control group. Compared to the control group,the levels of Raptor si RNA and proteins decreased significantly in the Raptor si RNA group; the Alizarin red staining results revealed lighter staining in the Raptor si RNA group,indicating lower level of mineralization; the real-time PCR results showed that the expression of osteoblast differentiation marker genes and osteoblast transcription factors in the Raptor si RNA group was lower than the control group. Conclusion： mTORC1 signaling promoted the osteoblast differentiation from preosteoblast cell MC3T3-E1.

关 键 词：mTORC1信号 成骨分化 前成骨细胞 

分 类 号：R329.24[医药卫生—人体解剖和组织胚胎学]