鼻咽癌中PMS2基因表达失活的甲基化调控机制  

Inactivation of PMS2 gene by promoter methylation in nasopharyngeal carcinoma

作　　者：倪海峰[1] 江波[1] 周珍[1] 李勇[1] 袁晓阳[1] 曹晓林[1] 黄光武[2] 

机构地区：[1]杭州市第一人民医院耳鼻咽喉头颈外科,310006 [2]广西医科大学第一附属医院耳鼻咽喉头颈外科,南宁530021

出　　处：《中华肿瘤杂志》2016年第11期812-817,共6页Chinese Journal of Oncology

基　　金：基金项目：浙江省医药卫生科技一般项目（2011KYA127）;浙江省医药卫生骨干平台项目（2012RCB038）

摘　　要：目的探讨鼻咽癌中PMS2基因表达失活的甲基化调控机制。方法收集54例鼻咽癌组织、16例正常鼻咽上皮组织样本和5个鼻咽癌细胞株（CNE1、CNE2、TWO3、HNE1、HONE1）、1个正常鼻咽部上皮细胞株（NP69）。采用半定量逆转录PCR检测PMS2 mRNA的表达，采用免疫组化法检测PMS2蛋白的表达，采用甲基化特异性PCR检测PMS2基因启动子甲基化状态。应用甲基转移酶抑制剂5－氮－2－脱氧胞苷处理CNE1和CNE2细胞，半定量逆转录PCR法检测处理前后PMS2 mRNA表达变化，分析鼻咽癌中PMS2基因甲基化及去甲基化对PMS2 mRNA表达的影响，分析PMS2 mRNA和蛋白表达与鼻咽癌临床病理特征的关系。结果5个鼻咽癌细胞株（CNE1、CNE2、HNE1、TW03、HONE1）中均可检测到PMS2基因甲基化，而正常鼻咽部上皮细胞（NP69）中未检测到PMS2基因甲基化。鼻咽癌组织中PMS2基因甲基化发生率为63.0%（34/54），正常鼻咽上皮组织中未检测到PMS2基因甲基化，差异有统计学意义（P〈0.001）。与正常鼻咽上皮组织比较，鼻咽癌组织中PMS2 mRNA和蛋白表达水平明显下调（P〈0.001）。PMS2基因甲基化的鼻咽癌组织中PMS2 mRNA和蛋白表达水平明显低于非甲基化组织（P〈0.001）。经5－氮－2－脱氧胞苷处理后的CNE1和CNE2细胞，可以重新恢复PMS2 mRNA表达。PMS2 mRNA和蛋白表达与鼻咽癌患者临床病理特征无关（均P〉0.05）。结论PMS2基因参与鼻咽癌的发生、发展，PMS2基因是启动子甲基化失活的鼻咽癌侯选肿瘤抑制基因。ObjectiveTo investigate the inactivation of PMS2 gene mediated by promoter methylation and its regulatory mechanism in nasopharyngeal carcinoma （NPC）.MethodsFifty-four NPC tissues, 16 normal nasopharyngeal epithelia （NNE）, 5 NPC cell lines （CNE1, CNE2, TWO3, HNE1 and HONE1） and 1 normal nasopharyngeal epithelial cell line （NP69） were collected.Methylation-specific PCR （MSP） was used to detect the PMS2 promoter methylation, semi-quantitative reverse transcription PCR （qRT-PCR） was applied to determine its mRNA expression, and immunohistochemistry （IHC） was used to detect the protein expression of PMS2. The expressions of PMS2 mRNA in CNE1 and CNE2 cells before and after treated with methyltransferase inhibitor 5-aza-2-deoxycytidine were analyzed by qRT-PCR. The impact of methylation and demethylation on the mRNA expression of PMS2, and the association of mRNA and protein expression of PMS2 with clinicopathological features of nasopharyngeal cancer were analyzed.ResultsMethylation of PMS2 gene was detected in all of the five NPC cell lines, but not in normal nasopharyngeal epithelial NP69 cells. The methylation rate of PMS2 gene in NPC tissues was 63% （34/54）, significantly higher than that of the normal nasopharyngeal epithelia （0/16, P〈0.001）. The expression levels of PMS2 mRNA and protein were significantly down-regulated in the 54 NPC tissues when compared with those in the 16 NNE tissues （P〈0.001）, and were also significantly lower in the 34 methylated NPC tissues than those in the 20 unmethylated NPC tissues （P〈0.001）. After treatment with 5-aza-2-deoxycytidine, the expression of PMS2 mRNA was restored in the CNE1 and CNE2 cells.However, the expressions of PMS2 mRNA and protein were not significantly correlated with patients′ age, gender, TNM stage, histopathologic type or lymph node metastasis （P〉0.05 for all）.ConclusionsPromoter methylation-mediated inactivation of PMS2 gene participates in carcinogenesis and development of NPC. PMS2 may be a candida

关 键 词：鼻咽肿瘤 DNA甲基化 PMS2 基因表达 病理学 临床 

分 类 号：R739.63[医药卫生—肿瘤]