Study On The Methylation Level Of MiRNA-34a Promoter And Its Regulatory Effects In Osteosarcoma

Objective To detect the level of miRNA-34 a promoter methylation in osteosarcoma and observe the effect of demethylation on the proliferation of osteosarcoma cells and the expression of miRNA-34 a,and to investigate the regulation of miRNA-34 a promoter methylation in osteosarcoma.Methods Real time quantitative PCR(real-time PCR)was used to detect the difference of mi RNA-34 a expression in osteosarcoma cells(MG-63,SAOS-2)and osteoblasts(hFOB1.19);matrix-assisted laser desorption/ionization–time of flight(MALDI-TOF)was used to detect the difference of miRNA-34 a promoter CpG island methylation level in osteosarcoma cells and osteoblasts,osteosarcoma paraffin tissue and normal bone tissue.Demethylation of osteosarcoma cells with 1% 5-Aza-2'-deoxycytidine,to observe the growth status of osteosarcoma cells,and to detect the expression of miRNA-34 a and the methylation level of mi RNA-34 a promoter in osteosarcoma cells before and after demethylation treatment.Results The expression of miRNA-34 a in MG-63 and SAOS-2 cells was significantly lower than that in hFOB1.19 cells,(P<0.05).The methylation level of miRNA-34 a promoter CpG island in MG-63 and SAOS-2 cells was higher than that in hFOB1.19 cells,(P<0.05).MG-63 and SAOS-2 cells were inhibited by demethylation,the expression of miRNA-34 a was up-regulated(P<0.05),and the methylation level of miRNA-34 a promoter CpG island was decreased(P < 0.05).Conclusion The promoter of mi RNA-34 a is a high methylation level,and the down-regulation of miRNA-34 a expression is related to its promoter methylation in osteosarcoma.The proliferation of osteosarcoma cells after demethylation is limited,suggesting that mi RNA-34 a and DNA methylation may be involved in the proliferation of osteosarcoma development process.