缺锌诱导的DNA异常甲基化在食管鳞状上皮细胞癌变过程中的机理研究

Esophageal squamous cell carcinoma (ESCC) is one of the top list incidence and mortality disease worldwide, especially in China. And most of the ESCC was diagnosed as the late stage due to lack of diagnosis biomarker for early stage disease, leading to an emergent biomarker discovery for early ESCC diagnosis. On the other hand, the combination of unhealthy diets and carcinogen exposures promotes many cancer progressions. For example, zinc deficiency and low doses of NMBA carcinogen significantly produced the progression of ESCC in rat model, through inducing extensive gene expression changes, in which especially those inflammatory related genes, as well as DNA methylation related gene (DNMT3A). In this proposal, we hypothesis that DNA methylation changes in zinc deficiency rat model might be another important mechanism involved in ESCC progression, especially during the initial neoplasm. We proposed to examine the DNA cytosine methylation status in a panel of tumor suppressor genes and zinc deficiency down-regulated genes, as well as global DNA methylation status, through COBRA screening, pyrosequencing quantitative DNA methylation analysis. At the same time, we will examine the related gene expression by real time quantitative PCR and western-blotting analysis. Our aim is to develop a potential DNA methylation based genetic biomarker for ESCC early diagnosis. Our proposal has the potential clinical significance in the early diagnosis of ESCC, especially for those who under the risk of carcinogen exposures and live with unbalance diet.

食管鳞状细胞癌(ESCC)上我国高发病率和高死亡率的癌症之一，并居世界之首。鉴于ESCC缺乏早期临床症状，一旦确诊大都属于癌变晚期，从而使得ESCC早期发病机理以及早期诊断的肿瘤标志物的研发迫切需要。另外饮食习惯和环境致癌因素的相互作用已被证实是各种癌症的重要诱发因素，并日益受到重视。本课题结合微营养因素(锌)和低剂量的致癌物(NMBA)，相对真实的模拟在环境因素和亚健康的饮食习惯的条件下建立适用于ESCC早期癌变与长期癌症发展的机理研究的大鼠动物模型。通过DNA甲基化定性与定量分析，实时定量PCR以及免疫印迹等试验手段，本课题研究旨在发现一些具有异常DNA甲基化的，可以适用于ESCC早期检测及其后期癌变发展过程预测的肿瘤标志基因。本课题的研究成果将对那些具有不健康的饮食习惯并有暴露于低剂量致癌环境的特定人群具有潜在的临床实践意义。

食管鳞状上皮细胞癌（ESCC）在我国属于高发病率和高死亡率的重大肿瘤疾病之一，且居世界首位。ESCC发病一般都属于亚临床症状，这给其早期诊断带来极大困难。因此，了解ESCC的早期发病机制并发现早期诊断分子标志物显得迫切需要。本项目采用缺锌条件下饲养的大鼠作为ESCC的动物模型，对癌变前后和正常对照组的食管鳞状上皮组织中的DNA甲基转移酶活性和挑选出来的12个基因的启动子区域甲基化水平进行定性与定量检测。我们研究结果发现癌变前的食管上皮组织中DNA甲基转移酶活性显著升高，且G0S2 基因启动子区的CpG岛甲基化水平明显高于正常对照组，随着ESCC的进一步发展，G0S2基因的甲基化水平持续增高。意外地是，我们并未发现G0S2基因的高度甲基化下调其mRNA或者蛋白水平的表达。进一步探索G0S2基因表达调控的分子机制发现其上游调控因子PPAR gamma在ESCC肿瘤细胞中与G0S2共同高度表达。我们研究结果提示G0S2 基因的甲基化除了可能可以作为ESCC的早期检测分子标志物外，还启示DNA甲基化在基因表达调控中可能存在着新的作用机制。