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[微小RNA-150通过靶向RUNX2基因抑制骨肉瘤细胞增殖]

[MicroRNA-150 inhibits osteosarcoma cell proliferation by targeting RUNX2 gene].

作者信息

Wang Longfei, Wang Weiguo, Li Jinsong, Chen Shijie, Zhan Ruisen

机构信息

Department of Orthopaedic Trauma, Tianjin Hospital, Tianjin 300211, China.

Department of Orthopedics, Third Xiangya Hospital, Central South University, Changsha 410013, China.

出版信息

Zhong Nan Da Xue Xue Bao Yi Xue Ban. 2016 Dec 28;41(12):1285-1290. doi: 10.11817/j.issn.1672-7347.2016.12.006.

Abstract

To investigate the microRNA (miR)-150 expression level in human osteosarcoma cell lines (Saos-2, MG-63) and its function in cell proliferation, and to explore the potential molecular mechanisms. 
 Methods: MiR-150 expression levels in human osteosarcoma cell lines (Saos-2, MG-63) and normal osteoblast cell line (NHOst) were detected by relative quantitative real-time PCR (qRT-PCR). MiR-150 was overexpressed in Saos-2 and MG-63 cells by lentivirus infection. Cell proliferation rates were monitored by MTS assay. RUNX2 and β-actin protein levels were examined by Western blot. Inhibitory effect of miR-150 on binding RUNX2 3'-UTR was detected by Dual-Luciferase assay.
 Results: MiR-150 expression level is lower in human osteosarcoma cell lines (Saos-2, MG-63) compared to the normal osteoblast cell line (NHOst) (0.23±0.02 and 0.32±0.03 vs 1.00±0.02), which showed statistical significance (P<0.01). After lentivirus infection, miR-150 level increased in Saos-2 (P<0.01) and MG-63 cells (P<0.01). Overexpression of miR-150 decreased cell proliferation and RUNX2 protein level in Saos-2 and MG-63 cells. The binding of miR-150 to RUNX2 3'-UTR decreased luciferase activity by 69% in Saos-2 cells (P<0.05) and 59% in MG-63 cells (P<0.05). Administration of exogenous RUNX2 recovered the cell proliferation in miR-150 overexpressed Saos-2 and MG-63 cell lines (P<0.01).
 Conclusion: MiR-150 inhibites proliferation in human osteosarcoma cell lines through binding to RUNX2 3'-UTR, resulting in the reducion of RUNX2 protein level.

摘要

研究人骨肉瘤细胞系(Saos-2、MG-63)中微小RNA(miR)-150的表达水平及其在细胞增殖中的作用,并探讨其潜在的分子机制。方法:采用相对定量实时荧光定量PCR(qRT-PCR)检测人骨肉瘤细胞系(Saos-2、MG-63)和正常成骨细胞系(NHOst)中miR-150的表达水平。通过慢病毒感染在Saos-2和MG-63细胞中过表达miR-150。采用MTS法监测细胞增殖率。通过蛋白质免疫印迹法检测RUNX2和β-肌动蛋白的蛋白水平。采用双荧光素酶报告基因检测法检测miR-150对RUNX2 3'-UTR结合的抑制作用。结果:与正常成骨细胞系(NHOst)相比,人骨肉瘤细胞系(Saos-2、MG-63)中miR-150的表达水平较低(分别为0.23±0.02和0.32±0.03,而NHOst为1.00±0.02),差异具有统计学意义(P<0.01)。慢病毒感染后,Saos-2细胞(P<0.01)和MG-63细胞(P<0.01)中miR-150水平升高。miR-150过表达降低了Saos-2和MG-63细胞的增殖及RUNX2蛋白水平。在Saos-2细胞中,miR-150与RUNX2 3'-UTR的结合使荧光素酶活性降低了69%(P<0.05),在MG-63细胞中降低了59%(P<0.05)。外源性RUNX2的加入恢复了miR-150过表达的Saos-2和MG-63细胞系中的细胞增殖(P<0.01)。结论:miR-150通过与RUNX2 3'-UTR结合抑制人骨肉瘤细胞系的增殖,导致RUNX2蛋白水平降低。

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