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目的:观察发状分裂相关增强子Hey1在骨形态发生蛋白9(BMP9)诱导的小鼠间充质干细胞(MSCs)C3H10T1/2成骨分化中的作用。方法:包装Hey1、BMP9以及GFP的过表达慢病毒,并分别作用于C3H10T1/2细胞,RT-PCR和Western blot检测Hey1、BMP9以及GFP的慢病毒是否包装成功,碱性磷酸酶(ALP)检测早期成骨指标ALP的变化,茜素红S染色检测晚期成骨指标钙盐沉积,MTT检测Hey1对BMP9调控的C3H10T1/2细胞增值的影响,流式细胞术检测Hey1对BMP9调控的C3H10T1/2细胞周期的影响。结果:Hey1、BMP9以及GFP的慢病毒包装成功;在成骨分化早期,过表达Hey1基因可促进BMP9调控的C3H10T1/2细胞的成骨分化与增殖;在成骨分化晚期,过表达Hey1基因可促进BMP9诱导的C3H10T1/2细胞的成骨分化,并将BMP9调控的C3H10T1/2细胞周期阻滞在G1期。结论:Hey1基因可参与调控BMP9诱导的小鼠C3H10T1/2细胞的早晚期成骨分化,并对细胞的增殖与周期有一定的影响。
Objective: To observe the role of Hey1, a hairpin-split-related enhancer, in the osteogenic differentiation of bone marrow-derived mesenchymal stem cells (MSCs) C3H10T1 / 2 induced by bone morphogenetic protein 9 (BMP9). METHODS: Hey1, BMP9 and GFP overexpressed lentivirus were packaged and transfected into C3H10T1 / 2 cells respectively. The packaging of Hey1, BMP9 and GFP lentivirus was successfully detected by RT-PCR and Western blot. The alkaline phosphatase (ALP) Alizarin Red S staining was used to detect the calcium deposition in advanced osteogenesis index. The effect of Hey1 on the proliferation of C3H10T1 / 2 cells was detected by MTT assay. The effect of Hey1 on the expression of C3H10T1 / 2 Effect of cell cycle. Results: The Hey1, BMP9 and GFP lentiviral packaging was successful. In the early stage of osteogenic differentiation, Hey1 gene overexpression promoted osteogenic differentiation and proliferation of C3H10T1 / 2 cells regulated by BMP9. In the late stage of osteogenic differentiation, Hey1 gene overexpression was Promote osteogenic differentiation of C3H10T1 / 2 cells induced by BMP9, and block the C3H10T1 / 2 cell cycle regulated by BMP9 in G1 phase. Conclusion: Hey1 gene may be involved in the regulation of early and late osteogenic differentiation of mouse C3H10T1 / 2 cells induced by BMP9, and may have some effects on cell proliferation and cycle.