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目的探讨Notch1对骨髓源性脂肪祖细胞(BMAPCs)向脂肪细胞分化的影响。方法无菌取SD大鼠股、胫骨骨髓制作细胞悬液,差速贴壁法培养,流式分析检测细胞表面标志表达;实时荧光定量聚合酶链式反应(real-time qPCR)检测脂肪相关基因和Notch信号分子表达。Notch1基因沉默实验分为空载体组和Notch1小RNA干扰组。Western blotting检测干扰效率;STEMPRO Adipogenesis Differentiation Kit行细胞成脂诱导。结果 BMAPCs表达CD34、CD44、CD45及CD90,且表达6种脂肪相关基因及Notch信号分子;成脂诱导促进Notch1 m RNA表达(P<0.01);Notch1小RNA干扰抑制Notch1蛋白表达(P<0.05)。Notch1小RNA干扰组的Cebpa、Lpl和Ppaγ、Slc2a4与Fabp4 m RNA表达均较对照显著降低,而pread1 mRNA表达显著增加(P<0.05~P<0.01);Notch小RNA干扰显著抑制脂肪细胞百分率(P<0.05)。结论 Notch1基因沉默抑制BMAPCs向成熟脂肪细胞分化。
Objective To investigate the effects of Notch1 on adipocyte differentiation of bone marrow derived adipogenic progenitor cells (BMAPCs). Methods SD rats were aseptically harvested from the femoral and tibial bone marrow of the tibia and cultured by differential adherence method. Flow cytometry was used to detect the expression of cell surface markers. Real-time quantitative real-time qPCR was used to detect the expression of fat-related genes And Notch signaling molecule expression. Notch1 gene silencing experiments were divided into empty vector group and Notch1 small RNA interference group. Western blotting was used to detect the interference efficiency; STEMPRO Adipogenesis Differentiation Kit was induced by adipocytes. Results The expression of Notch1 mRNA and Notch1 mRNA in BMAPCs were significantly increased (P <0.01). The expression of Notch1 mRNA and Notch1 mRNA in BMAPCs were significantly decreased (P <0.05) . The expression of Cebpa, Lpl, Ppaγ, Slc2a4 and Fabp4 mRNA in Notch1 small RNA interference group was significantly lower than that in control group, while the expression of pread1 mRNA was significantly increased (P <0.05 ~ P <0.01); Notch siRNA interference significantly inhibited the percentage of adipocytes P <0.05). Conclusion Notch1 gene silencing inhibits the differentiation of BMAPCs into mature adipocytes.