5-氮杂-2'-脱氧胞苷通过抑制端粒酶活性对肝癌细胞的抗肿瘤作用。
Anti-tumor effect of 5-aza-2'-deoxycytidine by inhibiting telomerase activity in hepatocellular carcinoma cells.
机构信息
Tumor Immunology and Gene Therapy Center, Eastern Hepatobiliary Surgery Hospital, The Second Military Medical University, Shanghai 200438, China.
出版信息
World J Gastroenterol. 2012 May 21;18(19):2334-43. doi: 10.3748/wjg.v18.i19.2334.
AIM
To investigate the effect of the demethylating reagent 5-aza-2'-deoxycitidine (DAC) on telomerase activity in hepatocellular carcinoma (HCC) cell lines, SMMC-7721 and HepG2.
METHODS
The related gene expression in cell lines was examined by real-time reverse transcription-polymerase chain reaction and Western blotting analysis. The telomerase activity was examined by telomeric repeat amplification protocol-enzyme-linked immunosorbent assay and DNA methylation was determined by methylation-specific polymerase chain reaction.
RESULTS
The telomerase activity was significantly reduced in both cell lines treated with DAC, accompanied by downregulation of telomerase reverse transcriptase (hTERT). We also observed the effect of DAC on the methylation status of hTERT promoter and the expression of regulatory genes, such as c-myc, p15, p16, p21, E2F1, and WT1. The methylation status of hTERT promoter could be reversed in SMMC-7721 by DAC, but not in HepG2 cells. However, p16 expression could be reactivated by demethylation of its promoter, and c-Myc expression was repressed in both cell lines. Moreover, DAC could enhance the sensitivity to the chemotherapeutic agents, such as cisplatin, by induction of apoptosis of HCC cells.
CONCLUSION
The DAC exerts its anti-tumor effects in HCC cells by inhibiting the telomerase activity.
目的
研究去甲基化试剂 5-氮杂-2'-脱氧胞苷（DAC）对肝癌细胞系 SMMC-7721 和 HepG2 中端粒酶活性的影响。
方法
通过实时逆转录-聚合酶链反应和 Western blot 分析检测细胞系中的相关基因表达。通过端粒重复扩增协议-酶联免疫吸附试验检测端粒酶活性，通过甲基化特异性聚合酶链反应检测 DNA 甲基化。
结果
DAC 处理的两种细胞系中端粒酶活性均显著降低，同时伴随着端粒酶逆转录酶（hTERT）的下调。我们还观察到 DAC 对 hTERT 启动子甲基化状态和调节基因（如 c-myc、p15、p16、p21、E2F1 和 WT1）表达的影响。DAC 可逆转 SMMC-7721 中 hTERT 启动子的甲基化状态，但在 HepG2 细胞中则不能。然而，p16 启动子的去甲基化可使其重新激活，c-Myc 表达在两种细胞系中均受到抑制。此外，DAC 可通过诱导 HCC 细胞凋亡增强顺铂等化疗药物的敏感性。
结论
DAC 通过抑制端粒酶活性发挥其在肝癌细胞中的抗肿瘤作用。
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