PEG10基因在肝癌组织中高表达的分子机制及其功能的研究  

Molecular mechanism and function research of highly expressed PEG 10 gene in hepatocellular carcinoma

作　　者：林昀[1] 张云丽 胡昕 邓庆 李琦[1] 高勇[1] 滕小梅[3] 韩泽广 黄健[2,4] 

机构地区：[1]同济大学附属东方医院肿瘤科,上海200120 [2]上海市疾病与健康基因组学实验室-省部共建国家重点实验室培育基地,国家人类基因组南方研究中心,上海201203 [3]苏州大学附属第一人民医院心血管外科,江苏苏州215006 [4]生物芯片上海国家工程研究中心,上海201203

出　　处：《肿瘤》2013年第1期28-35,共8页Tumor

基　　金：上海市科委基础研究重点项目(编号:11JC1408800);上海市医学重点专科建设资助项目(编号:ZK2012A26);上海市浦东新区科技发展基金创新资金资助项目(编号:PKJ2010-Y09);上海市浦东新区卫生系统中青年业务骨干培养基金资助项目(编号:PWGG09-1)

摘　　要：目的:探讨印迹基因父系表达基因10(paternally expressed gene10,PEG10)在肝癌组织中高表达的分子机制,及其对肝癌细胞生长的影响。方法:采用半定量RT-PCR法检测30例肝癌患者术后肝癌和癌旁组织中以及15株肝癌细胞株中PEG10基因的表达水平;甲基化特异PCR(methylation specific PCR,MSP)技术检测17例PEG10基因高表达的肝癌和癌旁组织中PEG10基因启动子区域CpG岛甲基化状态的差异,及其与PEG10基因表达水平的关系。采用小干扰RNA(small interfering RNA,siRNA)技术沉默肝癌QGY-7703细胞中PEG10基因的表达,并观察对细胞克隆形成能力的影响。结果:与癌旁组织相比,PEG10基因在67%(20/30)的肝癌组织中表达水平明显提高,在15株肝癌细胞株中均有表达。PEG10基因CpG岛2在29.4%(5/17)肝癌组织中显示有肝癌特异性低甲基化,同时采用亚硫酸氢盐测序验证了PEG10基因CpG岛2在肝癌病例中总体低甲基化的状态。与对照组相比,短发夹状RNA(short hairpin RNA,shRNA)干扰PEG10表达后,肝癌细胞株QGY-7703的克隆数目明显减少(P<0.05)。结论:印迹相关基因PEG10在肝癌中高表达,可能与其启动子区域CpG岛2的低甲基化密切相关,沉默PEG10表达可以抑制肝癌细胞的生长,提示PEG10基因可能是一个新的肝癌治疗的药物靶点。Objective： To study the molecular mechanism of dysregulation of PEGIO （paternally expressed gene 10）, an imprinted gene in HCC （hepatocellular carcinoma）, and to analyze its effect on the growth of hepatocellular carcinoma cells. Methods： The expression level of PEG10 mRNA was detected by semi-quantitative RT-PCR in 30 paired HCC/adjacent non-cancerous tissues and 15 types of liver cancer cell lines. Methylation status of CpG islands at the promoter of PEG10 gene in 1 7 paired HCC/adjacent non- cancerous tissue samples in which the expression of PEGIO gene in cancer tissues was significantly up- regulated as compared with non-cancerous tissues was detected using MSP （methylation specific PCR）. The PEG10 shRNA （short hairpin RNA） was used to silence the expression of PEGIO gene in HCC QGY-7703cells, and the colony-forming ability of QGY-7703 cells was detected by Western blotting and colony formation assay. Results： PEGIO mRNA expression level was significantly up-regulated in 67% （20/30） HCC tissues as compared with adjacent non-cancerous tissues. PEGIO was expressed in 15 types of HCC cell lines. The methylation status of PEG10 gene＇s CpG2 island was significantly decreased in 29.4% （5/17） HCC tissue samples, and the overall decreased methylation status of PEGIO gene＇s CpG2 island in HCC tissues was verified using bisulfite sequencing. Interestingly, silencing the expression of PEG10 in HCC QGY-7703 cells using shRNA targeting PEGIO gene markedly inhibited the colony-forming efficiency as compared with the HCC QGY-7703 cells transfected with negative control shRNA （P 〈 0.05）. Conclusion： PEGIO, an imprinted gene, is up-regulated in HCC, and may be closely related with the decreased methylation status of CpG2 island at the promoter of PEGIO gene. Silencing the expression of PEG10 can inhibit the growth of HCC cells, suggesting that PEGIO gene may play an important role in the hepatocarcinogenesis, and it may be used as a new marker or a potential therapeutic target for HC

关 键 词：肝肿瘤 DNA甲基化 PEG10基因 

分 类 号：R735.35[医药卫生—肿瘤]