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| 釉基质蛋白诱导MC3T3-E1细胞成骨分化过程中微小核糖核酸的表达 |
| MicroRNA Expression During the Osteogenic Differentiation of MC3T3-E1 Cells by Enamel Matrix Derivative Induction. WANG Yan, WU Tingting, LI Jing, SHI Qiongling, HUANG Hui. Department of Prosthodontics, Ninth People’s Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai 200011 |
| 投稿时间：2014-03-20 修订日期：2014-03-30 |
| DOI： |
| 中文关键词: 微小核糖核酸 成骨分化 MC3T3-E1细胞 釉基质蛋白 |
| 英文关键词:miRNA osteogenic differentiation MC3T3-E1 Cells EMD |
| 基金项目:国家自然科学基金资助项目(81170988)；上海市科学技术委员会基金资助项目(11ZR1420200)。 |
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| 中文摘要: |
| [摘要] 目的：探讨微小核糖核酸(micro ribonucleicacid , miRNA)是否参与到釉基质蛋白(Enamel Matrix Derivative, EMD)诱导的小鼠前成骨细胞系MC3T3-E1细胞成骨分化的过程，并对发生显著变化的miRNA进行分析。方法：将小鼠前成骨细胞系MC3T3-E1用含/不含(对应：EMD刺激组/阴性对照组)EMD的培养液培养0、7、14天，碱性磷酸酶(alkaline phosphatase, ALP)染色检测ALP活性，实时聚合酶链式反应(real-time quantitative polymerase chain reaction, RT-PCR)技术分析成骨分化标记物ALP、骨涎蛋白(bone sialoprotein，BSP)和骨钙素(osteocalcin，OC) 的信使RNA(message RNA, mRNA)水平的表达变化情况。利用基因芯片技术分析miRNA的相对表达。结果：0、7、14天的ALP染色结果显示随着培养时间的增加，阴性对照与EMD刺激组其ALP活性均逐渐增强，EMD刺激组ALP活性增强较阴性对照组更为明显。RT-PCR结果表明，与阴性对照组相比，ALP、BSP、OC的mRNA表达量均显著增加，ALP与OC均于14天时表达量达到峰值，BSP则于7天时处于峰值表达，EMD刺激组MC3T3-E1成骨活性更强；在0、7、14天的培养周期中，11个miRNA表达上调，28个miRNA表达下调，其中miR-335-5p, miR-503已被证实可参与促进骨形成，而miR-30家族 (miR-30a, -30b, -30c 和-30d )则被证实参与抑制成骨。 结论：miRNA参与EMD诱导的MC3T3-E1细胞的成骨分化过程，这一发现可为了解EMD促进成骨分化的机理及临床应用提供指导。
[关键词] 微小核糖核酸 成骨分化 MC3T3-E1细胞 釉基质蛋白 |
| 英文摘要: |
| [Abstract] Objective : In this study, we aimed to identify whether micro ribonucleicacid (microRNA, miRNA) participated in the process of osteogenic differentiation induced by Enamel Matrix Derivative( EMD) and to categorize a full range of miRNAs induced in response to EMD. Methods: The mouse pre-osteoblast cell line MC3T3-E1 was cultured with/without （the EMD stimulating /negative control group）EMD induction(0, 7 and 14 day), and osteogenic differentiation was detected by alkaline phosphatase (ALP) staining and real-time quantitative polymerase chain reaction( RT-PCR) analysis. TaqMan MicroRNA Arrays (0, 7 and 14 day) was used to analyse the relative expression of miRNAs. Results: The results of the ALP staining indicated that EMD could enhance osteogenic differentiation in MC3T3-E1 cells, the results of EMD group are more significant than negative group. The results of RT-PCR showed that when compared to the negative group, the expression of ALP, BSP and OC increased significantly. In addition, both ALP and OC reached their peak levels at 14 days, while BSP reached its peak at 7 days . These findings indicate that EMD enhanced osteogenic differentiation in MC3T3-E1 cells. In the samples collected at 0, 7 and 14 day, 11 miRNAs were upregulated and 28 miRNAs were downregulated significantly during the process of osteogenic differentiation induced by EMD. And miR-335-5p, miR-503 have been confirmed to promote bone formation, while miR-30 family have been confirmed to be negative regulators of osteogenic differentiation. Conclusion: miRNAs were involved in EMD induced MC3T3-E1 osteogenic differentiation. These findings could be used to understand the mechanism of the regulatory process of osteogenic differentiation induced by EMD and guide clinical application.
[Key words] miRNA osteogenic differentiation MC3T3-E1 Cells EMD |
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