结直肠癌中胰岛素样生长因子结合蛋白-相关蛋白1表达上调与其5＇CpG岛低甲基化的相关性

Hypomethylation of 5＇ CpG island of insulin-like growth factor binding protein-related protein 1 is associated with its overexpression in colocectal cancer

目的探讨胰岛素样生长因子结合蛋白-相关蛋白1（IGFBP-rP1）在结直肠癌中的表达改变并分析其与该基因5＇CpG岛甲基化改变的关系。方法采用半定量逆转录聚合酶链反应（RT-PCR）和甲基化特异性聚合酶链反应（MSP）检测46例结直肠癌组织和配对的正常黏膜中IGFBP-rP1基因的表达水平及5＇CpG岛的甲基化状况，并通过T-A，克隆、测序加以验证。不表达IGFBP-rP1基因的结肠癌细胞株LoVo和SW620经去甲基化药物5-氮-2＇-脱氧胞苷（5-aza-2＇deoxycytidine，5-aza-dC）处理后，分别用RT-PCR和MSP检测IGFBP-rP1表达改变和甲基化状况。结果mRNA水平上，IGFBP-rP1在结直肠癌组织中的表达水平高于配对的正常黏膜（P〈0．01）。在46例结直肠癌组织中，28例（60．9％）可检测到IGFBP-rP1基因5＇CpG岛发生甲基化，而在配对的正常黏膜中，37例（80．4％）可检测到，二者差异有统计学意义（P〈0．05）。IGFBP-rP1基因的表达和甲基化水平之间存在负相关（P〈0．05）。5-aza-dC处理后，两株细胞均出现IGFBP-rP1基因的再表达，MSP法检测证实基因发生了去甲基化。结论IGFBP-rP1基因的表达水平与5＇CpG岛甲基化水平之间存在负相关。DNA甲基化是结直肠癌中IGFBP-rP1表达调控的一种机制。IGFBP-rP1低甲基化引起的基因表达上调可能与结直肠癌的发生发展有关。

Objective To investigate the methylation status of 5＇ CpG island of insulin-like growth factor binding protein-related protein 1 （ IGFBP-rP1 ） in colorectal cancer and its relationship with gene expression and clinicopathologic parameters. Methods Semi-quantitative reverse transcription-PCR （RT- PCR） was used to detect the expression of IGFBP-rP1 in 46 cases of colorectal cancer and their matched normal mucosa. Methylation-specific PCR （MSP） was applied to evaluate the methylation status of 5＇ CpG island of IGFBP-rP1. Colon cancer cell lines LoVo and SW620 were treated with demethylation agent 5-aza- 2＇-deoxycytidine （5-aza-dC）, followed by RT-PCR and MSP detection. Results At the mRNA level, the expression of IGFBP-rP1 was higher in colorectal cancer tissue than that in the matched normal mucosa （P 〈 0. 05）. IGFBP-rP1 was methylated in 28/46 （60. 9% ） cases of colorectal cancer and 37/46 （ 80. 4% ） matched normal mucosa samples （P 〈 0. 05 ）. A negative correlation was found between IGFBP-rP1 expression and its methylation status. The expression of IGFBP-rP1 was restored in LoVo and SW620 after treatment with 5-aza-dC and MSP confirmation of its demethylation status. No relationships was found between the methylation status and clinicopathologic parameters. Conclusions IGFBP-rP1 expression is negatively correlated with its methylation status in colorectal cancer. DNA methylation is one of the mechanisms regulating the expression of IGFBP-rP1. Hypomethylation of IGFBP-rP1 gene with its overexpression plays an important role in the initiation and development of colorectal cancer.