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目的:观察肝细胞系L02及肝癌细胞系SMMC-7721中人端粒酶逆转录酶(human telomerase reverse transcriptase,hTERT)的表达与DNA甲基化修饰之间的相关性,观察5-杂氮胞苷(5’-aza-dC)去甲基化对肝细胞系hTERT表达和端粒酶活性的影响。方法:应用甲基化特异性PCR(Methylation-specific PCR,MSP)分析肝细胞系L02和肝癌细胞SMMC-7721中hTERT的甲基化状态,采用RT-PCR和TRAP-ELISA法检测5’-aza-dC干预对培养细胞hTERT mRNA表达及端粒酶活性的影响。结果:肝细胞系L02中hTERT有甲基化修饰,5’-aza-dC去甲基化处理上调hTERT mRNA表达并呈现端粒酶活性升高;肝癌细胞系SMMC-7721中hTERT则没有甲基化修饰,5’-aza-dC去甲基化处理对其hTERT表达和端粒酶活性无影响。结论:肝细胞中DNA甲基化可能参与抑制肝细胞hTERT的表达,hTERT去甲基化可能是肝癌发生发展的重要机制之一。
OBJECTIVE: To observe the correlation between the expression of human telomerase reverse transcriptase (hTERT) and DNA methylation in hepatoma cell line L02 and hepatoma cell line SMMC-7721, Effects of demethylation of glycoside (5’-aza-dC) on hTERT expression and telomerase activity in hepatocyte line. Methods: Methylation-specific PCR (MSP) was used to analyze the methylation status of hTERT in hepatoma cell line L02 and hepatoma cell line SMMC-7721. RT-PCR and TRAP-ELISA were used to detect the 5’-aza Effects of -dC intervention on hTERT mRNA expression and telomerase activity in cultured cells. Results: The hTERT gene was modified in L02 hepatoma cell line. The demethylation of 5’-aza-dC up-regulated the expression of hTERT mRNA and induced the increase of telomerase activity. The hTERT in hepatoma cell line SMMC-7721 did not have methyl Modification, 5’-aza-dC demethylation had no effect on hTERT expression and telomerase activity. Conclusion: DNA methylation in hepatocytes may be involved in the inhibition of hTERT expression in hepatocytes. Demethylation of hTERT may be one of the important mechanisms of hepatocellular carcinoma.