mTORC1信号促进前成骨细胞MC3T3-E1的分化成熟

发布时间：2018-05-05 15:50

本文选题：mTORC信号 + 成骨分化　；参考：《中国免疫学杂志》2016年11期

【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。
[Abstract]:Aim: to study the effect of mTORC1 signal on the differentiation and maturation of preosteoblast MC3T3-E1 into osteoblasts. Methods: mTORC1 signal-associated protein (Raptor) was overexpressed by transfection of pcDNA3.1-Raptor. into MC3T3-E1. Raptor siRNAs were transfected into MC3T3-E1 and the mTORC1 signal protein Raptor was silenced. The gene expression of Raptor was detected by Real-time PCR method, the level of Raptor protein was detected by Western blot, and the osteogenic mineralization was detected by alizarin red staining to determine the degree of osteogenic differentiation. The gene expression of osteogenic differentiation index was detected by Real-time PCR. Results: compared with the control group, the levels of Raptor mRNA and protein were significantly increased in the Raptor overexpression group, and alizarin red staining showed that the Raptor overexpression group was more stained, indicating that the degree of osteogenesis mineralization was higher. The results of fluorescence quantitative PCR showed that the expression of osteogenic differentiation marker gene and osteogenic transcription factor in Raptor overexpression group was higher than that in control group. Compared with the control group, the levels of Raptor mRNA and protein in Raptor siRNA group were significantly lower, and alizarin red staining showed that Raptor siRNA group was lighter, indicating that the degree of osteogenesis mineralization was lower. The results of fluorescence quantitative PCR showed that the expression of osteogenic differentiation marker gene and osteogenic transcription factor in Raptor siRNA group was lower than that in control group. Conclusion the MC3T3-E1 of preosteoblast can be differentiated and matured by the signal of 1: mTORC1.
【作者单位】：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;河南省新乡医学院;河南省新乡医学院第三附属医院;
【基金】：国家自然科学基金项目(No.31301135)
【分类号】：R329.2

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