微小 RNA-520d-3p 通过降解 Akt1 抑制骨肉瘤进展。

MicroRNA-520d-3p inhibits osteosarcoma progression by degradation of Akt1.

机构信息

Department of Orthopedics, The First People's Hospital of Changzhou, Changzhou, China.

出版信息

Eur Rev Med Pharmacol Sci. 2018 Apr;22(8):2315-2320. doi: 10.26355/eurrev_201804_14821.

DOI:10.26355/eurrev_201804_14821

PMID:29762834

Abstract

OBJECTIVE

To detect the expression of microRNA-520d-3p in osteosarcoma tissue and the function on the osteosarcoma cells proliferation.

PATIENTS AND METHODS

We used qRT-PCR to access microRNA-520d-3p level from 10 cases of osteosarcoma and its adjacent tissues. The osteosarcoma cell lines were screened. The microRNA-520d-3p mimics or inhibitor was transfected into human osteosarcoma cells by liposome method, and the cell proliferation of each group was detected by the CCK8 assay. We used bioinformatics methods to detect and predict the target genes of microRNA-520d-3p. Luciferase reporter assay was utilized to detect the relative luciferase activity between microRNA-520d-3p and Akt1. Meanwhile, after cells were transfected with microRNA-520d-3p mimics, microRNA-520d-3p mimics + OE-Akt1, microRNA-520d-3p inhibitor or microRNA-520d-3p inhibitor + si-Akt1, we detected cell viability using CCK-8 assay, respectively to access the interaction between Akt1 and microRNA-520d-3p.

RESULTS

Lowly expressed microRNA-520d-3p in osteosarcoma tissues was observed in comparison with adjacent tissues. After transfecting with microRNA-520d-3p mimics, the viability of MG63 and U-20S cells decreased, which was higher in cells transfecting microRNA-520d-3p inhibitor. Bioinformatics prediction and dual luciferase reporter assay illustrated that microRNA-520d-3p targeted on Akt1. At the same time, Akt1 expression was higher in osteosarcoma tissues than in adjacent ones, cell proliferation was inhibited after blocking its expression. In addition, after transfected with microRNA-520d-3p mimic, viability of MG63 and U-20S cells decreased, which can be reversed by OE-Akt1. In contrast, the viability of MG63 and U-20S cells increased after transfection with microRNA-520d-3p inhibitor and which were reversed by si-Akt1.

CONCLUSIONS

Lowly expressed microRNA-520d-3p was observed in osteosarcoma; overexpression of microRNA-520d-3p can target Akt1 thus inhibiting proliferation of osteosarcoma cells.

摘要

目的

检测微小 RNA-520d-3p 在骨肉瘤组织中的表达及其对骨肉瘤细胞增殖的功能。

方法

我们使用 qRT-PCR 从 10 例骨肉瘤及其相邻组织中获取微小 RNA-520d-3p 水平。筛选骨肉瘤细胞系。通过脂质体法将微小 RNA-520d-3p 模拟物或抑制剂转染入人骨肉瘤细胞中，通过 CCK8 测定法检测各组细胞的增殖情况。我们使用生物信息学方法检测和预测微小 RNA-520d-3p 的靶基因。荧光素酶报告基因检测微小 RNA-520d-3p 与 Akt1 之间的相对荧光素酶活性。同时，在细胞转染微小 RNA-520d-3p 模拟物后，转染微小 RNA-520d-3p 模拟物+OE-Akt1、微小 RNA-520d-3p 抑制剂或微小 RNA-520d-3p 抑制剂+si-Akt1，我们分别使用 CCK-8 测定法检测细胞活力，以评估 Akt1 和微小 RNA-520d-3p 之间的相互作用。

结果

与相邻组织相比，骨肉瘤组织中低表达微小 RNA-520d-3p。转染微小 RNA-520d-3p 模拟物后，MG63 和 U-20S 细胞的活力降低，而转染微小 RNA-520d-3p 抑制剂的细胞活力升高。生物信息学预测和双荧光素酶报告基因检测表明微小 RNA-520d-3p 靶向 Akt1。同时，骨肉瘤组织中 Akt1 的表达高于相邻组织，阻断其表达后细胞增殖受到抑制。此外，转染微小 RNA-520d-3p 模拟物后，MG63 和 U-20S 细胞的活力降低，OE-Akt1 可逆转这一现象。相反，转染微小 RNA-520d-3p 抑制剂后，MG63 和 U-20S 细胞的活力增加，si-Akt1 可逆转这一现象。