Department of Infection, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan 450052, Peoples Republic of China.
Cell Biol Int. 2012 May 1;36(5):427-32. doi: 10.1042/CBI20100851.
Aberrant methylation of promoter CpG islands is a major inactivation mechanism of tumour-related genes that play a crucial role in the progression of silencing in human cancers, including HCC (hepatocellular carcinoma). We have examined the promoter methylation status of five important DNA damage response genes in fresh-frozen HCC tissues and cell lines, as well as the possible correlation between methylation patterns and clinical features of the carcinoma. Promoter methylation status of RASSF1A (Ras association domain family 1), CHFR (checkpoint with forkhead and ring finger domains), GSTP1 (glutathione-S-transferase-pi gene), MGMT [O(6)-methylguanine-DNA methyltransferase] and hMLH1 (human mutL homologue 1) were examined by the MSP (methylation-specific PCR) in 70 HCC tissues and five HCC cell lines. The mRNA expression levels of these genes were measured by RT-PCR (reverse transcription-PCR). Methylation frequencies of these genes tested in HCC were 54 (78%) for RASSF1A, 30 (43%) for CHFR, 26 (38%) for GSTP1 and 22 (32%) for MGMT. No hypermethylation was detected for hMLH1 in any case of HCC or HCC cell lines. Moreover, promoter hypermethylation of RASSF1A, CHFR and GSTP1 in both HepG2 and SNU398 cells, and hypermethylation of MGMT in Huh7 cells, were detected. Treatment of three cell lines with 5Aza-dC (5-aza-20-deoxycytidine) restored or increased the expression of these genes, implicating aberrant DNA methylation in transcriptional silencing. Hypermethylation of RASSF1A and patient age were significantly associated. CHFR methylation status showed a statistically significant correlation with HCC progression. Methylation of the RASSF1A, CHFR, GSTP1 and MGMT genes seem therefore to play an important role in the pathogenesis of HCC. These epigenetic changes may have prognostic importance for patients with HCC.
启动子 CpG 岛的异常甲基化是肿瘤相关基因失活的主要机制，这些基因在人类癌症的沉默进展中起着至关重要的作用，包括 HCC（肝细胞癌）。我们已经检查了五个重要的 DNA 损伤反应基因在新鲜冷冻 HCC 组织和细胞系中的启动子甲基化状态，以及甲基化模式与癌的临床特征之间的可能相关性。通过 MSP（甲基化特异性 PCR）检查了 70 个 HCC 组织和 5 个 HCC 细胞系中 RASSF1A（Ras 相关结构域家族 1）、CHFR（带有叉头和环指结构域的检查点）、GSTP1（谷胱甘肽-S-转移酶-pi 基因）、MGMT[O(6)-甲基鸟嘌呤-DNA 甲基转移酶]和 hMLH1（人类 mutL 同源物 1）的启动子甲基化状态。通过 RT-PCR（逆转录-PCR）测量这些基因的 mRNA 表达水平。在 HCC 中测试的这些基因的甲基化频率为 RASSF1A 为 54（78%），CHFR 为 30（43%），GSTP1 为 26（38%），MGMT 为 22（32%）。在任何 HCC 或 HCC 细胞系中均未检测到 hMLH1 的高甲基化。此外，在 HepG2 和 SNU398 细胞中均检测到 RASSF1A、CHFR 和 GSTP1 的启动子高甲基化，并且在 Huh7 细胞中检测到 MGMT 的高甲基化。用 5Aza-dC（5-氮杂-20-脱氧胞苷）处理三个细胞系可恢复或增加这些基因的表达，表明异常 DNA 甲基化与转录沉默有关。RASSF1A 的高甲基化与患者年龄显著相关。CHFR 甲基化状态与 HCC 进展呈统计学显著相关性。RASSF1A、CHFR、GSTP1 和 MGMT 基因的甲基化因此似乎在 HCC 的发病机制中起着重要作用。这些表观遗传变化可能对 HCC 患者具有预后意义。
