MicroRNA-144 inhibits the proliferation, apoptosis, invasion, and migration of osteosarcoma cell line F5M2

This study is aimed to investigate the role of microRNA-144 (miR-144) in osteosarcoma cell line F5M2 proliferation, apoptosis, invasion, and metastasis. Between 2007 and 2014, 66 cases of osteosarcoma samples in the corresponding adjacent normal tissue samples were selected from surgical resection or biopsy in the Department of Orthopedics, Shengjing Hospital, China Medical University. MiR-144 levels and Ezrin messenger RNA (mRNA) levels in osteosarcoma and the adjacent bone tissues were detected, and clinical and pathological features were analyzed. Exogenous miR-144 was transfected into human osteosarcoma cell lines at two different concentrations (low and high), and the expression levels of miR-144 and Ezrin protein between highly metastatic osteosarcoma cells and lowly metastatic osteosarcoma cells were compared. Real-time polymerase chain reaction (RT-PCR) and Western blot were used for detecting the expression levels of miR-144 or Ezrin protein, respectively. Cell proliferation was measured by methylthiazol tetrazolium (MTT) assay. Cell invasion and migration was evaluated by Transwell assays. Finally, flow cytometry was employed to determine the cell apoptosis. MiR-144 expression in osteosarcoma tissue was significantly lower than that in the surrounding normal bone tissue (P < 0.001), while Ezrin mRNA expression in osteosarcoma tissue was significantly higher than that in the surrounding normal bone tissue (P < 0.001); correlation analysis showed a significant negative correlation between miR-144 and Ezrin mRNA levels (r = 0.982, P < 0.001). MiR-144 and Ezrin mRNA expressions were significantly related with cell metastasis (P < 0.05) but were not related with other clinical factors such as gender, age, tumor location, tumor size, Enneking staging, and Dahlin’s histological classification. The results of RT-PCR showed that the expression level of miR-144 in osteosarcoma cells increased after transfected with exogenous miR-144 mimics, and this increase positively correlated with the transfection concentration of miR-144 mimics. Furthermore, Western blotting revealed a significant and dose-dependent decrease in Ezrin protein levels in F5M2 cells transfected with miR-144. MTT and Transwell assays showed that the invasion and metastasis of F5M2 cells was significantly decreased following exogenous overexpression of miR-144. Our study supports the view that the miR-144 may regulate Ezrin protein expression by inhibiting the invasion and metastasis of osteosarcoma cells.