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目的:探讨5-杂氮-2′-脱氧胞苷(5-Aza-dC)和制霉菌素A(TSA)对人喉癌细胞系中CHFR基因启动子区甲基化与mRNA表达水平的影响。方法:采用荧光定量PCR技术和甲基化特异性PCR技术检测5-Aza-dC和TSA处理前后的Hep-2人喉癌细胞系基因CHFR启动子区甲基化与mRNA表达水平的变化。结果:在对照组Hep-2人喉癌细胞系中CHFR基因启动子区显示DNA甲基化,用5-Aza-dC作用后DNA甲基化程度明显减弱,mRNA表达量上调(1.75±0.21);用TSA作用后DNA甲基化程度和mRNA表达量(1.05±0.13)无明显变化。联合使用5-Aza-dC和TSA后DNA甲基化程度明显减弱,mRNA表达量明显上调(2.15±0.18)。结论:CHFR基因启动子区DNA甲基化在喉癌的发生、发展中是一个频发事件,是导致其mRNA表达下调的主要原因,5-Aza-dC单独应用和联合TSA能够逆转DNA甲基化状态,从而调控其基因表达,为临床使用药物治疗喉癌提供了新思路。
AIM: To investigate the effects of 5-Aza-dC and TSA on the promoter methylation and mRNA expression of CHFR gene in human laryngeal carcinoma cell lines . Methods: The changes of methylation and mRNA expression of CHFR promoter in Hep-2 human laryngeal carcinoma cell lines before and after 5-Aza-dC and TSA treatment were detected by real-time PCR and methylation-specific PCR. Results: The DNA methylation was found in the promoter region of CHFR gene in Hep-2 human laryngeal carcinoma cell line. The DNA methylation was significantly attenuated by 5-Aza-dC and the mRNA level was increased by 1.75 ± 0.21 ; DNA methylation and mRNA expression (1.05 ± 0.13) did not change significantly after treatment with TSA. DNA methylation was significantly weakened after combined use of 5-Aza-dC and TSA, and the mRNA expression was significantly increased (2.15 ± 0.18). Conclusion: DNA methylation in the promoter region of CHFR gene is a frequent event in the development and progression of laryngeal carcinoma, leading to the down-regulation of its mRNA expression. 5-Aza-dC alone and in combination with TSA can reverse DNA methyl And thus regulate its gene expression, providing a new idea for the clinical use of drugs for the treatment of laryngeal cancer.