Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/20083
標題: 台灣肝癌細胞位於第十六號染色體上腫瘤抑制基因群之核酸甲基化剖析
Profiling methylation status of tumor suppressor genes on chromosome 16 in human hepatocellular carcinoma in Taiwan
作者: 林怡君
Lin, Yi-Jiun
關鍵字: 甲基化
methylation
肝癌
腫瘤抑制基因
細胞附著分子
hepatocellular carcinoma
tumor suppressor gene
cadherin
CDH13
H-cadherin
T-cadherin
出版社: 生物醫學研究所
摘要: 肝癌業已成為台灣地區人民的頭號生命殺手,慢性B型肝炎與C型肝炎病毒感染為誘發肝癌發生的主要危險因子。近年許多研究指出,腫瘤抑制基因其基因調節區之甲基化可造成基因的表現異常,且與許多人類癌症的生成是息息相關的,肝癌亦是如此;第十六號染色體異質性消失現象常發生於肝癌檢體組織中,意味著有腫瘤抑制基因分布在此染色體上。本試驗之目的首先重於第十六號染色體上腫瘤抑制基因群TSC2、SOCS1、CDH1、E2F4與CDH13於肝癌基因體中甲基化程度之探討,初步試驗結果顯示,CDH13基因的甲基化比例最高(39.4%),其次為SOSC1與CDH1基因(20.6%),而TSC2與E2F4基因則無甲基化。本研究進一歩探討CDH13基因於肝癌中的mRNA與蛋白質表現情形,結果顯示甲基化之CDH13基因在肝腫瘤組織的mRNA表現皆高於正常肝臟組織;且於腫瘤切片的免疫組織化學分析中,顯示CDH13蛋白於肝實質部細胞與肝竇狀血管系統有大量的表現。在三個肝癌細胞株中,亦發現較高度甲基化的CDH13基因其mRNA表現量較高,且在細胞質與膜蛋白表現上,具高度CDH13基因甲基化之HA22T細胞株相較於基因未甲基化之細胞株(HepG2與Hep3B)明顯有不同之蛋白構型。最後,經由Bisulfite-定序分析與搜尋CDH13基因調節區上可能的轉錄作用因子結合位,顯示含CpG序列之特定轉錄作用因子可能受甲基化修飾作用而影響其對基因的調控能力。
In Taiwan, hepatocellular carcinoma is the leading cause of cancer death. Hepatitis B virus and hepatitis C virus infections are the major contributing factors of hepatocarcinogenesis. Recent studies have reported that aberrant hypermethylation in the CpG-rich promoter regions of many tumor suppressor genes is associated with the lack of gene transcription and the development of hepatocellular carcinoma. Loss of heterozygosity on chromosome 16 is a common genetic alternation in human hepatocarcinomas, indicating the existence of tumor suppressor genes on this chromosome. In this study, we focus on the gene mehtylation studies of hundreds hepatoma biopsies in five selected potential tumor suppressor genes, named TSC2, SOCS1, E2F4, CDH1 and CDH13 genes, which located on human chromosome 16. Aberrant methylation frequencies of this 5 genes detected by MS-PCR in 155 hepatoma pairs were39.4% for CDH13, 20.6% for SOCS1 and CDH1, 0% for TSC2 and E2F4. In mRNA expression assay, CDH13 gene hypermethylated hepatoma cells exhibited higher CDH13 mRNA level than its normal tissue pairs. This phenomenon has been demonstrated in three hepatoma cell lines that HA22T cell line with CDH13 gene hypermethylation obtained highest CDH13 mRNA expression, while HepG2 and Hep3B cell lines with CDH13 gene unmethylation showed no detectable CDH13 mRNA expression. In protein expression assay, CDH13 proteins appeared three different isoforms located on cytosol and membrane and differential display in three hepatoma cell lines. In CDH13 protein immunohistochemistry staining, overexpressed CDH13 protein has been detected in tumor vessels and hepatocytes in different hepatitis viruses-infected hepatoma cells. Finally, we found several transcription regulation factors, such as AP2-α, MZF1-4, n-Myc, and c-ETS, which located on the CpG island of CDH13 promoter region, have been hypermethylated in all 20 CpG sites in HA22T cell line. It may alter the regulation of transcription factors in CDH13 gene expression.
URI: http://hdl.handle.net/11455/20083
Appears in Collections:生物醫學研究所

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