膀胱癌TCF21基因甲基化与临床意义

发布时间：2018-04-22 15:25

本文选题：DNA甲基化水平 + 膀胱癌　；参考：《华侨大学》2014年博士论文

【摘要】：目的：膀胱癌是一种常见的恶性肿瘤，全球每年约15万人死亡。迄今仍然没有分子靶向药物被批准用于该疾病治疗。TCF21（Transcription Factor21）是编码转录因子21的基因，也是肿瘤抑制基因。TCF21基因甲基化在头颈部癌、肺癌和肾癌上已有报道，然而在膀胱癌上TCF21基因甲基化表征及其临床意义尚未得到充分的揭示，尤其是TCF21基因甲基化定量检测在膀胱癌中未见报道。本研究探讨TCF21基因甲基化与表达的相互关系，以及TCF21基因甲基化在膀胱癌中的表征，目的在于证明对膀胱癌尿脱落细胞中TCF21基因甲基化定量检测，可以成为膀胱癌诊断和预后的重要手段之一。方法：选择膀胱癌细胞株5637进行体外实验，用去甲基化试剂Decitabine和Zebularine处理膀胱癌细胞株5637，采用焦磷酸测序技术定量检测膀胱癌细胞株5637中TCF21基因甲基化水平，分别采用双标记荧光探针技术和Western-blot方法检测膀胱癌细胞株5637中TCF21mRNA和蛋白水平；采用焦磷酸测序技术定量检测83例膀胱癌组织和19例正常膀胱组织中TCF21基因甲基化水平，采用Mann-Whitney U Test统计方法分析、比较在膀胱癌组织中TCF21基因甲基化水平在患者性别、年龄、吸烟史、肿瘤分级和分期五个临床参数上的差异，采用Spearman Correlation统计方法分析在膀胱癌组织中TCF21基因甲基化水平与患者年龄、肿瘤分级和分期三个临床参数的相关性；采用焦磷酸测序技术定量检测33例膀胱癌尿脱落细胞和10例健康人群尿脱落细胞中TCF21基因甲基化水平，采用Mann-Whitney U Test统计方法分析、比较在膀胱癌尿脱落细胞中TCF21基因甲基化水平在患者性别、年龄、吸烟史、肿瘤分级和分期五个临床参数上的差异，采用Spearman Correlation统计方法分析在膀胱癌尿脱落细胞中TCF21基因甲基化水平与患者年龄、肿瘤分级和分期三个临床参数的相关性，采用受试者工作特征（ROC）曲线方法对比膀胱癌组织和膀胱癌尿脱落细胞中TCF21基因甲基化水平对诊断的潜在能力。结果：本研究发现用去甲基化试剂Decitabine和Zebularine处理膀胱癌细胞株5637后，导致TCF21基因甲基化水平降低，TCF21mRNA和蛋白表达增加；与此同时，在膀胱癌组织中TCF21甲基化水平高于正常膀胱组织，在膀胱癌组织中TCF21甲基化水平高于肾癌组织，患者年龄对在膀胱癌组织中TCF21基因甲基化水平存在差异；在膀胱癌尿脱落细胞中TCF21甲基化水平高于健康人群尿脱落细胞，患者肿瘤分级对在膀胱癌尿脱落细胞中TCF21基因甲基化水平存在差异，患者肿瘤分级与在膀胱癌尿脱落细胞中TCF21基因甲基化水平相关，在膀胱癌组织和膀胱癌尿脱落细胞中TCF21的ROC曲线下面积均高于参考值，但在膀胱癌尿脱落细胞中TCF21特异性高于膀胱癌组织。结论：本研究证明TCF21基因CpG岛的异常甲基化在调节TCF21基因的表达中起重要作用；TCF21基因可以作为是膀胱癌诊断和预后的检测的标志物；对膀胱癌尿脱落细胞中TCF21基因甲基化定量检测，有可能成为膀胱癌诊断和预后的重要手段之一。
[Abstract]:Objective: bladder cancer is a common malignant tumor with about 150 thousand people worldwide each year. There are still no molecular targeting drugs approved for the treatment of.TCF21 (Transcription Factor21), a gene encoding transcription factor 21, and a tumor suppressor gene.TCF21 methylation in the head and neck cancer, lung cancer and kidney cancer. However, the methylation of TCF21 gene and its clinical significance in bladder cancer have not been fully revealed, especially the quantitative methylation detection of TCF21 gene has not been reported in bladder cancer. The relationship between the methylation and expression of the TCF21 gene and the characterization of the methylation of the TCF21 gene in bladder cancer are discussed in this study. Quantitative detection of TCF21 gene methylation in cystadenocarcinoma exfoliated cells can be one of the important means for diagnosis and prognosis of bladder cancer.
Methods: bladder cancer cell line 5637 was selected in vitro, and bladder cancer cell line 5637 was treated with demethylation reagent Decitabine and Zebularine. The methylation level of TCF21 gene in bladder cancer cell line 5637 was detected by pyrosequencing technology. The double labeling fluorescence probe technique and Western-blot method were used to detect bladder cancer. The level of TCF21mRNA and protein in cell line 5637 and the methylation level of TCF21 gene in 83 bladder cancer tissues and 19 normal bladder tissues were quantified by pyrosequencing technology. The Mann-Whitney U Test statistical method was used to compare the TCF21 gene methylation water in the bladder cancer tissue for the patient's sex, age, smoking history and tumor classification. The Spearman Correlation statistical method was used to analyze the correlation between the level of TCF21 gene methylation of the TCF21 gene in the bladder cancer tissues and the three clinical parameters of the tumor classification and staging, and the quantitative detection of urine exfoliation in 33 cases of bladder cancer and 10 healthy people by pyrosequencing. The methylation level of TCF21 gene in the cells was analyzed by Mann-Whitney U Test statistical method, and the difference in the five clinical parameters of the level of methylation of TCF21 gene in the urinary exfoliated cells of bladder cancer was compared with the sex, age, smoking history, tumor classification and staging, and the urine exfoliative cells of bladder cancer were analyzed by Spearman Correlation. The correlation between the level of methylation of the TCF21 gene and the three clinical parameters of the patient's age, tumor classification and staging, and the potential ability of the TCF21 gene methylation in bladder cancer and urinary exfoliative cells to compare the diagnostic potential of the bladder cancer tissue and urinary bladder cancer cells were compared by the method of the subject work feature (ROC) curve.
Results: after treating bladder cancer cell line 5637 with demethylation reagents Decitabine and Zebularine, the methylation level of TCF21 gene was reduced and the expression of TCF21mRNA and protein increased. At the same time, the level of TCF21 methylation in bladder cancer tissues was higher than that of normal bladder tissue, and the level of TCF21 methylation in bladder cancer tissues was higher than that in bladder cancer tissue. The level of methylation of TCF21 gene in bladder cancer tissues is different in the patients with renal carcinoma. The level of TCF21 methylation in urinary exfoliated cells of bladder cancer is higher than that of the healthy people. The level of TCF21 gene methylation in the urinary exfoliative cells of bladder cancer is different in patients with bladder cancer, and the tumor classification and bladder cancer in the patients with bladder cancer are different. The level of TCF21 gene methylation in urine exfoliated cells was higher than that of the reference value in the ROC curve of TCF21 in bladder cancer tissue and bladder cancer urine exfoliative cells, but the specificity of TCF21 in bladder cancer urine exfoliated cells was higher than that of bladder cancer tissue.
Conclusion: This study proves that the abnormal methylation of the TCF21 gene CpG Island plays an important role in the regulation of the expression of TCF21 gene, and the TCF21 gene can be used as a marker for the diagnosis and prognosis of bladder cancer, and the quantitative detection of TCF21 gene in the urine exfoliated cells of bladder cancer can be an important hand for the diagnosis and prognosis of bladder cancer. One of the segments.

【学位授予单位】：华侨大学
【学位级别】：博士
【学位授予年份】：2014
【分类号】：R737.14

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