Open AccessUp‐regulation of micro RNA ‐491‐5p suppresses cell proliferation and promotes apoptosis by targeting FOXP 4 in human osteosarcoma
Author(s) -
Yin Zhixun,
Ding Hongmei,
He Erxing,
Chen Jingchen,
Li Ming
Publication year - 2017
Publication title -
cell proliferation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.647
H-Index - 74
eISSN - 1365-2184
pISSN - 0960-7722
DOI - 10.1111/cpr.12308
Subject(s) - osteosarcoma , microrna , reporter gene , flow cytometry , apoptosis , transfection , cell growth , electrophoretic mobility shift assay , cancer research , cell culture , biology , microbiology and biotechnology , luciferase , cell , gene , gene expression , genetics
Background and objectives Micro RNA s are small non‐coding RNA s involved in pathogenesis and progression of human malignancies. Micro RNA ‐491‐5p (miR‐491‐5p) is down‐regulated in many human cancers where it would serve as a tumour suppressor. However, the role played by miR‐491‐5p in pathogenesis of human osteosarcoma has remained largely unknown. This study has been conducted to examine effects of miR‐491‐5p on migration and proliferation of cells of the SAOS ‐2 and MG 63 osteosarcoma lines, and mechanisms of those effects. Materials and methods Levels of miR‐491‐5p expression in osteosarcoma tissues and in human osteosarcoma cell lines were studied using qualitative real‐time polymerase chain reaction ( qRT ‐ PCR ) methods. Cell viability was detected using the CCK ‐8 and EdU assays, while the transwell assay was used to evaluate migration and invasion. Apoptosis was analysed uing flow cytometry and the Hoechst 33342 nuclear staining method. A dual‐luciferase reporter system was used to confirm the target gene of miR‐491‐5p. The electrophoretic mobility shift assay ( EMSA ) with DIG ‐labelled double‐stranded FOXP 4 oligonucleotides was used to confirm whether or not miR‐491‐5p suppressed FOXP 4 activation. Results Cells of osteosarcoma tissues and cell lines had low levels of miR‐491‐5p expression, but high levels of forkhead‐box P4 ( FOXP 4) expression. Transfection of SAOS ‐2 and MG 63 cells with miR‐491‐5p mimics inhibited expression of FOXP 4 protein, which suppressed cell growth and migration, but induced apoptosis. Dual‐luciferase reporter assays confirmed FOXP 4 as the target gene for miR‐491‐5p. Overexpression of miR‐491‐5p suppressed FOXP 4 activity in SAOS ‐2 and MG 63 cells. Knockdown of FOXP 4 in SAOS ‐2 and MG 63 cells using an RNA i strategy resulted in reduced levels of cell proliferation and migration, but increased levels of apoptosis. Conclusion Our in vitro studies showed that up‐regulation of miR‐491‐5p suppressed proliferation of the human osteosarcoma cells and induced apoptosis by targeting FOXP 4 . These findings suggest that miR‐491‐5p could be further studied as a potential clinical diagnostic or predictive biomarker for human osteosarcoma.