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目的探讨5氮杂脱氧胞苷(5-aza-cdr)对乳腺癌细胞中p16基因mRNA表达的影响。方法分别用5-aza-cdr 5、10和20μmol/L处理乳腺癌细胞MCF-7,未处理的MCF-7细胞作为对照。采用甲基化特异性聚合酶链反应(MSP)对药物处理前后的细胞进行p16基因甲基化检测;绿色荧光染料实时反转录聚合酶链反应(SYBR GreenqRT-PCR)检测p16mRNA表达。结果在未处理的MCF-7细胞中(对照)p16基因呈完全甲基化状态,随着5-aza-cdr浓度增加,甲基化水平逐渐减弱,至5-aza-cdr 20μmol/L时p16基因甲基化状态完全被逆转;p16mRNA表达水平也随着药物剂量的递增逐渐增加,5-aza-cdr 20μmol/L处理组与5、10μmol/L处理组以及未处理组比较差异均有统计学意义(均P<0.01)。结论乳腺癌细胞MCF-7中p16基因的甲基化能被5-aza-cdr逆转,去甲基化后可以促进p16mRNA表达。
Objective To investigate the effect of 5-aza-cdr on p16 mRNA expression in breast cancer cells. Methods MCF-7 breast cancer cells were treated with 5-aza-cdr 5,10 and 20 μmol / L, and untreated MCF-7 cells were used as control. The methylation of p16 gene was detected by methylation-specific polymerase chain reaction (MSP) before and after drug treatment. The p16 mRNA expression was detected by SYBR Green qRT-PCR. Results The p16 gene was completely methylated in untreated MCF-7 cells (control). The methylation level decreased gradually with the increase of 5-aza-cdr concentration and reached the level of p16 at 5-aza-cdr 20μmol / L Gene methylation status was completely reversed; p16mRNA expression levels also increased with the dose of drug gradually increased, 5-aza-cdr 20μmol / L treatment group and 5,10μmol / L treatment group and untreated group differences were statistically significant Significance (all P <0.01). Conclusion Methylation of p16 gene in breast cancer cell MCF-7 can be reversed by 5-aza-cdr, and demethylation can promote p16 mRNA expression.