Effects and mechanism of microRNA-154-5p targeting E2F5 on bioactivity of osteo-sarcoma cells
Objective To investigate the effect of microRNA-154-5p(miR-154-5p)targeting E2F5 on the prolifer-ation and apoptosis of osteosarcoma cells and its molecular mechanism.Methods After miR-154-5p transfection of osteo-sarcoma MG63 cells,the changes of cell proliferation were detected by CCK-8,the changes of cell migration and invasion ability were evaluated by transwell cell migration assay,the level of apoptosis was detected by flow cytometry.Western blot was used to analyze the changes in the expression of B-cell lymphoma protein 2-associated X protein(Bax),Cleaved caspase-3 and B-cell lymphoma protein 2(Bcl-2)markers.Bioinformatics and dual luciferase reporter genes were used to analyze the potential target genes of miR-154-5p.Results The CCK-8 assay results showed that the absorbance values of the blank group,the negative control group and the miR-154-5p simulated group after 72 hours of intervention were 1.179±0.026,1.161±0.024 and 0.502±0.015,respectively,the difference was statistically significant(F=2 533.000,P＜0.001).The Transwell cell migration experiment results showed that the number of migrating MG63 cells in the blank group,the negative control group and the miR-154-5p simulated group after 72 hours of intervention were 79.80±8.53,76.20±6.38 and 49.60±3.05,respectively,the difference was statistically significant(F=33.260,P＜0.001).The flow cytometry detection results showed that the apoptosis rates of MG63 cells in the blank group,the negative control group and the miR-154-5p simulated group after 72 hours of intervention were(4.56±1.04)％,(4.25±1.02)％and(18.12±1.90)％,respectively,the difference was statistically significant(F=99.400,P＜0.001).Western blot further detected that the expression of Bax,Cleaved caspase 3,and Bcl-2 in the blank group,the negative control group and the miR-154-5p simulated group after 72 hours of intervention was significantly increased(Bax:0.124±0.010,0.128±0.013,0.364±0.018;Cleaved caspase 3:0.099±0.020,0.112±0.033,0.513±0.024;Bcl-2:0.672±0.015,0.675±0.017,0.245±0.028;F=188.200,P＜0.001;F=159.200,P＜0.001;F=281.600,P＜0.001).Bioin-formatics and dual luciferase reporter gene assay results showed that E2F5 was a potential target gene of miR-154-5p.Conclusion miR-154-5p can promote osteosarcoma apoptosis and inhibit cell proliferation by inhibiting E2F5.
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