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目的:探讨miR-21与BMP9之间的关系,明确miR-21在BMP9诱导间充质干细胞成骨分化中的作用。方法:(1)Ad-BMP9感染C3H10T1/2细胞,Real-time-PCR检测miR-21表达。RTPCR检测ALP的表达。(2)MiR-21转染C3H10T1/2细胞,Real-time-PCR检测miR-21和BMP9表达。(3)MiR-21和BMP9-CM处理C3H10 T1/2细胞,ALP活性和染色实验检测C3H10 T1/2细胞早期成骨能力。茜素红S染色实验检测钙盐沉积情况。(4)MiR-21和BMP9-CM处理C3H10T1/2细胞,Real-time-PCR检测成骨分化相关因子ALP,OCN的表达。(5)MiR-21和BMP9-CM处理C3H10T1/2细胞,Western blot检测p-Smad1/5蛋白水平的表达。结果:(1)BMP9暂时降低miR-21的表达。MiR-21也可以暂时降低BMP9的表达。(2)MiR-21可以协同BMP9增强ALP和钙盐沉积。(3)MiR-21协同BMP9增加了p-Smad1/5蛋白水平的表达。结论:MiR-21与BMP9存在相互关系,两者可以互相调节表达。MiR-21可以协同BMP9促进间充质干细胞C3H10T1/2细胞成骨分化,这一过程与增强BMP9/Smad信号的激活程度有关。
AIM: To investigate the relationship between miR-21 and BMP9 and clarify the role of miR-21 in osteogenic differentiation of mesenchymal stem cells induced by BMP9. Methods: (1) C3H10T1 / 2 cells were infected with Ad-BMP9, and miR-21 expression was detected by Real-time-PCR. RTPCR detection of ALP expression. (2) MiR-21 transfected C3H10T1 / 2 cells, Real-time-PCR detection of miR-21 and BMP9 expression. (3) The C3H10 T1 / 2 cells were treated with MiR-21 and BMP9-CM, and the early osteogenic ability of C3H10 T1 / 2 cells was detected by ALP activity and staining. Alizarin red S staining test calcium deposition. (4) C3H10T1 / 2 cells were treated with MiR-21 and BMP9-CM. Real-time-PCR was used to detect the expression of ALP and OCN. (5) C3H10T1 / 2 cells were treated with MiR-21 and BMP9-CM, and the expression of p-Smad1 / 5 protein was detected by Western blot. Results: (1) BMP9 temporarily reduced the expression of miR-21. MiR-21 can also temporarily reduce BMP9 expression. (2) MiR-21 can cooperate with BMP9 to enhance ALP and calcium deposition. (3) MiR-21 cooperates with BMP9 to increase the expression of p-Smad1 / 5 protein. Conclusion: There is a correlation between MiR-21 and BMP9, and the two can regulate each other. MiR-21 can cooperate with BMP9 to promote the osteogenic differentiation of mesenchymal stem cell C3H10T1 / 2 cells, which is related to the enhancement of the activation of BMP9 / Smad signal.