人肝癌中SLIT2基因表达及启动子甲基化分析  

The expression and promoter methylation of SLIT2 gene in human primary hepatocellular carcinomas

作　　者：金洁[1] 游海燕[2] 余艳军[2] 黄明辉[2] 舒惠群[2] 姚根富[2] 杨胜利[1,3] 覃文新[2] 

机构地区：[1]江苏大学生命科学院 [2]上海交通大学医学院-上海市肿瘤研究所癌基因及相关基因国家重点实验室 [3]上海交通大学医学院-上海市肿瘤研究所癌基因及相关基因国家重点实验室上海200032

出　　处：《复旦学报（医学版）》2007年第2期202-206,共5页Fudan University Journal of Medical Sciences

基　　金：国家重点基础研究发展计划(2002CB513104);上海市优秀学科带头人计划(05XD14013)项目

摘　　要：目的研究SLIT2在肝癌组织中表达和沉默的机制。方法32个肝癌病人的癌组织及癌旁肝组织、3例正常肝组织及8个肝癌细胞系样本DNA和RNA抽提后,半定量RT-PCR检测SLIT2的表达。亚硫酸氢钠处理DNA,用MSP和测序法检测SLIT2启动子甲基化状态。结果SLIT2启动子在肝癌组织中的甲基化率为84.3%,在相对应的癌旁肝组织中甲基化率为56.3%。肝癌细胞系的甲基化率为75%。而在正常肝组织中无甲基化。SLIT2启动子甲基化与肝癌的临床病理指标无相关性,但和患者的年龄有关。在用甲基化酶抑制剂5-aza-dC处理细胞后,SLIT2表达缺失的肝癌细胞系恢复了mRNA的表达。结论SLIT2启动子甲基化是肝癌发生过程中的早期事件,可能是SLIT2在肝癌中表达沉默的主要机制。Purpose To study the SLIT2 expression and mechanism of its silencing in hepatocellular carcinoma （HCC）. Methods Genomic DNA and total RNA were extracted from 3 normal liver tissue samples, 32 HCC samples and 8 liver cancer cell lines, respectively. The expression of SLIT2 was evaluated by semi-quantitative RT-PCR. The methylation specific PCR （MSP） and bisulfite DNA sequencing were performed to examine the methylation status of SLIT2 gene promoter. Results Promoter methylation of SLIT2 gene was found in 27 of 32 liver tumor tissues （84.3%） and 18 of 32 noncancerous liver tissues （56.3%） from 32 HCC patients, and 6 of 8 liver cancer cell lines （75%）, whereas SLIT2 methylation was not detected in normal liver tissues from healthy controls. There was no correlation between promoter hypermethylaion and pathological data for primary HCC patients, including tumor size, lymph metastasis, and clinical stage. However, the frequency of methylation in elder patients （≥60 years old） was significantly higher than that in younger patients （〈60 years old） （P〈0.05）. Interestingly, the expression of SLIT2 could be induced by the demethylating agent 5-aza- 2-deoxycytidine in HCC cell lines with the promoter hypermethylation of SLIT2 gene. Conclusions Promoter hypermethylation of SLIT2 gene may occur as an early event in the development of HCC. Methylation is possibly a major mechanism for inactivation of SLIT2 gene in HCC.

关 键 词：肝癌 SLIT2 DNA甲基化 抑癌基因 

分 类 号：R735.7[医药卫生—肿瘤]