The Effect of Methylation Level of microRNA Promoter on the Expression of microRNAs and on the Proliferation,Migration and Invasion of Lung Cancer Cells
Abstract
To study the effect of methylation level of microRNA promoter on the expression of microRNAs (miRNA34a,miRNA34b,miRNA148a,miRNA203a) and on the proliferation,migration and invasion of lung cancer A549 cells. MethodsThe proliferation of A549 cells treated with different concentrations of demethylated drug 5-aza-2′-deoxycytidine (5-Aza-CdR) was measured by CCK8 assay and calculated the inhibitory rate in 24 h,48 h and 72 h,respectively. After 72 h of treatment with 20 μmol/L 5-Aza-CdR,methylation-specific PCR (MSP) was used to detect the methylation level of A549 cells in miRNAs gene promoter regions,and real-time quantitative PCR (real-time PCR) was used to test the expression of miRNAs. The migration abilities of A549 cells treated with 20 μmol/L 5-Aza-CdR in 24 h and 48 h were performed with wound healing assay,while the invasion abilities in 48 h were evaluated by Transwell assay,respectively. ResultsThe proliferation inhibition rate of A549 cells gradually increased with the treatment concentration of 5-Aza-CdR increased and the treatment time prolonged. Compared with the control group,the methylated band of the experimental group was weaker and the unmethylated band was stronger,and the miRNAs gene promoter regions methylation level of the experimental group was lower than that of the control group. The expression level of miRNAs was significantly increased in the experimental group （P<0.05）. The migration and invasion of the experimental group of A549 cells were inhibited compared with the control group （P<0.05）. Conclusion5-Aza-CdR can reverse methylation levels of miRNAs promoter regions and upregulate the expression level of miRNA34a,miRNA34b,miRNA148a,miRNA203a,resulting in significantly inhibiting the proliferation,migration and invasion of lung cancer cells.
Keywords: Lung cancer cell line, A549microRNA, Methylation, Proliferation, Migration Invasion
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