【摘要】 背景与目的对肺癌组织基因组DNA异常甲基化进行筛选与分析,探索肺癌发病的表遗传致癌机制。材料与方法从肺癌和癌旁组织中分别提取基因组DNA,经Mse1(甲基化非敏感性酶)单独消化或Mse1和BstU1(甲基化敏感性酶)联合消化,消化产物用甲基化敏感性内切酶指纹法(PCR-basedtechnique-Methylation-sensitiveRestrictionFingerprinting,MSRF)进行分析,肺癌组织出现异常甲基化基因片段,进一步将异常甲基化DNA片段进行亚克隆和序列测定,再与基因文库中的基因进行类比分析,同时按年龄、性别及吸烟状况分组并分析其与肺癌DNA异常甲基化的关系。结果84.5%(49/58)的肺癌样本出现异常甲基化现象,但肺鳞癌(82.1%)与肺腺癌(88.5%)的异常甲基化率差异无统计学意义(χ2=0.073,P>0.05);在异常甲基化DNA片段中,高甲基化现象占83%,低甲基化为17%,发现2条重要的高甲基化基因片段,它们分别与抑癌基因WT-1(Wilm’stumorsuppressorgene)和细胞周期调控基因CCNC(humancyclinCgene)匹配;经统计学分析,吸烟、年龄和性别与肺癌DNA异常甲基化无关。结论基因组DNA的异常甲基化,特别是高甲基化现象,可能在肺癌发生发展中起重要作用,这可能是肺癌发病的表遗传机制。更多还原

【Abstract】 BACKGROUND & AIM: To study the aberrant DNA methylation of genomic DNA in lung cancer tissues as a possible epigenetic mechanism in the development of lung cancer. MATERIAL AND METHODS: Genomic DNA isolated from lung cancer and its adjacent tissue was restriction digested with Mse1 (methylation non-sensitive) alone or with Mse1 and BstU1 (methylation sensitive). The resulting DNA was analyzed for aberrant methylation using a PCR-based technique- Methylation-sensitive Restriction Fingerprinting (MSRF). Several DNA fragments differentially methylated in the transformed cells compared with the non-transformed cells were identified by MSRF. These fragments were subcloned,sequenced and compared with GenBank. At the same time,the DNA methylation was analyzed according to age,sex and smoking status. RESULTS: As compared with the control tissues,84.5% (49/58) of lung cancer tissues were found to have aberrant DNA methylation,but the methylation rate did not differ significantly among the squamous carcinoma (82.1%),and adenocarcinoma (88.5%) and other cancer types (75.0%)(χ2=0.073,P>0.05). Among the fragments methylated,83% was hypermethylated and 17% was hypomethylated. We have identified the fragments encoding for human cyclin C (CCNC) and Wilms tumor (WT-1). When smoking,age and sex were considered as factors,no significant differences were observed. CONCLUSION: The aberrant DNA methylation,especially hypermethylation,seemed to play an important role in the development of lung cancer. It appeared to be a possible epigenetic mechanism for lung cancer.更多还原