Effect and mechanism of microRNA-24 on cell proliferation and migration of osteosarcoma cell line U2OS

Abstract

Objective To investigate the effect of microRNA-24 (miR-24) on cell proliferation and migration in osteosarcoma cell line U2OS and its possible mechanism. Methods U2OS cell line with miR-24 over-expression was established by transfecting miR-24 mimic, and then cell proliferation and migration in control group, negative control group and miR-24 over-expression group were detected with real-time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) assay, cell counting kit-8 (CCK-8) assay and Transwell assay, respectively. Epithelial-mesenchymal transition (EMT) progression and activation of nuclear factor-κB (NF-κB) pathway were detected by Western blotting. Results The expressions of miR-24 in the control group, negative control group and miR-24 over-expression group were 1.00±0.00, 1.03±0.08 and 2.46±0.29, with significant difference (F=11.026, P=0.012). Compared with the control group, the expression of miR-24 in human osteosarcoma U2OS cell line was significantly increased after transfecting miR-24 mimic (t=4.604, P=0.009). After over-expression of miR-24 for 24 h, 48 h and 72 h, U2OS cell viabilities were decreased significantly compared with the control group [(3.56±0.27)% vs. (8.63±0.79)%, t=3.896, P=0.016; (20.16±1.09)% vs. (54.77±5.42)%, t=4.813, P=0.008; (45.47±3.16)% vs. (95.52±8.56)%, t=7.173, P=0.002)]. After over-expression of miR-24 for 24 h, the cell migration rates in control group, negative control group and miR-24 over-expression group were (100.00±0.00)%, (99.26±5.85)% and (31.37±2.09)%, respectively, and there was statistically significant difference among the three groups (F=12.175, P=0.009); and compared with control group, cell migration rate was decreased significantly after over-expression of miR-24 (t=3.843, P=0.004). Meanwhile, over-expression of miR-24 up-regulated the expression levels of epithelial cell markers E-cadherin (t=3.852, P=0.018) and β-catenin (t=3.512, P=0.024), while down-regulated the expression levels of mesenchymal cell markers N-cadherin (t=3.832, P=0.018) and vimentin (t=4.058, P=0.012), with a suppressed EMT progress. Other than that, miR-24 over-expression inhibited the expressions of NF-κB (p65) (t=4.813, P=0.008), phosphorylation inhibitor of nuclear factor kappa-B kinase α (p-IKK-α) (t=3.764, P=0.013) and phosphorylation inhibitor of nuclear factor kappa-B kinase complex α (p-IκB-α) (t=4.064, P=0.012), suppressing the activation of NF-κB pathway. Conclusion miR-24 can suppress cell proliferation and migration of osteosarcoma cell line U2OS in vitro, and its mechanism may be related to the suppression of NF-κB activation and EMT progression. It hints that miR-24 may be used as a potential new target for osteosarcoma therapy. Key words: Osteosarcoma; Cell movement; MicroRNA-24