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目的:探讨喉癌Hep-2细胞中组蛋白H3-K9甲基化与DNA甲基化及抑癌基因MGMT表达的关系。方法:应用去甲基化制剂5-氮杂-2′-脱氧胞苷(5-Aza-dC)处理体外培养的Hep-2细胞。应用染色质免疫沉淀技术、甲基化特异性聚合酶链反应和实时定量逆转录聚合酶链反应分析药物作用前后MGMT基因启动子区组蛋白H3-K9甲基化、DNA甲基化和MGMT表达情况。结果:①在Hep-2细胞未经药物干预前,MGMT表现为DNA甲基化,组蛋白H3-K9高甲基化,MGMT低表达。②在5-Aza-dC的作用下,Hep-2细胞中MGMT的组蛋白H3-K9甲基化状态被降低;MGMT的DNA甲基化状态得到了逆转;原来低表达的MGMT表达上调。结论:喉癌细胞系中MGMT启动子区甲基化可能是导致其基因失活的主要原因。DNA甲基化可能导致组蛋白H3-K9高甲基化。应用5-Aza-dC能够通过逆转MGMT的DNA甲基化水平从而降低MGMT组蛋白H3-K9甲基化水平来使抑癌基因表达上调,从而抑制肿瘤的发生和发展。
Objective: To investigate the relationship between histone H3-K9 methylation and DNA methylation and tumor suppressor gene MGMT in Hep-2 cells. Methods: Hep-2 cells cultured in vitro were treated with 5-Aza-2’-deoxycytidine (5-Aza-dC). The histone H3-K9 methylation, DNA methylation and MGMT expression in MGMT gene promoter region were analyzed by chromatin immunoprecipitation, methylation-specific polymerase chain reaction and real-time quantitative reverse transcriptase-polymerase chain reaction Happening. Results: ①MHMT showed DNA methylation, histone H3-K9 hypermethylation and low MGMT expression in Hep-2 cells before drug intervention. ② Under the action of 5-Aza-dC, the histone H3-K9 methylation status of MGMT was decreased in Hep-2 cells; the DNA methylation status of MGMT was reversed; the expression of MGMT was up-regulated. CONCLUSION: Methylation of MGMT promoter region in laryngeal carcinoma cell lines may be the main reason for its inactivation. DNA methylation may result in hypermethylation of histone H3-K9. Application of 5-Aza-dC can inhibit tumorigenesis and progression by reversing the DNA methylation level of MGMT and thus decreasing the methylation level of H3-K9 in MGMT.