miR-680可促进Runx2诱导C2C12细胞进行成骨细胞早期分化.doc

  miR-680 可促进 Runx2 诱导 C2C12 细胞进 行成骨细胞早期分化# 耿倩倩，余守和，王红梅，孙奋勇** （暨南大学生命科学技术学院,广州,510632） 摘要：目的:验证 miR680 在 Runx2 诱导 C2C12 细胞进行成骨分化过程中的作用。方法:利用可诱导表达 Runx2 的细胞系 C2C12/Runx2Dox，在 8 个不同时间点通过 Real-time PCR 方法检测 miR-680 的表达情况。 C2C12/Runx2Dox 细胞转染 miR-680 mimic 后利用 Real-time PCR 检测成骨细胞标记基因 Alp 及 OC 表达情 况，并利用钙骨染色法分析 ALP 活性。利用在线工具 miRanda, miRWalk 与 TargetScan 共同预测 miR-680 潜在靶基因。利用 DAVID Bioinformatics Resources 数据库对得到的靶基因进行功能聚类分析。结果与结 论：在 Runx2 诱导 C2C12 进行成骨分化过程中 miR-680 表达显著增加。转染 miR-680 mimic 可上调 Alp 表 达，但对 OC 表达无影响；钙钴染色显示转染 miR-680 mimic 也可增加 ALP 活性。靶基因功能聚类分析结果 表明 miR-680 参与细胞的分化过程。研究结果表明，Runx2 通过上调 miR-680 从而调控相关基因的表达对 成骨分化进行调节，以促进 C2C12 进行成骨细胞的早期分化。 关键词：Runx2；成骨分化；miR-680；C2C12 细胞 中图分类号：Q74 MiR-680 promotes Runx2-induced early osteoblast differentiation of C2C12 cells Geng Qianqian, YU Shouhe, Wang Hongmei, Sun Fenyong (JINAN University,guangzhou 510632) Abstract: To investigate the role of miR-680 during Runx2-induced osteoblast differentiation in C2C12 cells. Methods:We used Tet-on inducible gene expression system and established C2C12/Runx2Dox sub-line with conditional Runx2 expression induced by inducer Doxycyclin(Dox) in mouse myoblast C2C12.Firstly,we studied the miR-680 expression induced by Runx2 at different time points by Real-time PCR. Dox induced C2C12/Runx2Dox to the process of osteogenic differentiation in vitro transfection miR-680.Detected the activity of ALP in osteoblast by alkaline phosphatase (ALP) staining and Real-time PCR.We prediction miRNA-680 target gene by online database miRanda, miRwalk, Targetscan and function classification. Results and conclusion: The expression of miR-680 was significantaly highly in osteoblast differentiation. Transfection miR-680 mimics into cells enhanced the activity of alkaline phosphatase and Real-time PCR results showed that the expression of Alp was significantly increased. Results suggested that miR-680 was highly expressed in C2C12/Runx2Dox to the osteogenic differentiation process and can promote the early di