探討第16型人類乳突狀病毒於子宮頸上皮內贅瘤之CpG甲基化情形

Detection of Human Papillomavirus Type 16 CpG methylation in Cervical intraepithelial neoplasia

賴曉鈴(Hsiao-Ling Lai)

指導教授 ： 李瑞年

共同指導教授 ： 吳明蒼

高雄醫學大學/生命科學院/醫藥暨應用化學研究所/碩士(2006年)

摘要

子宮頸癌(cervical cancer)為全球婦女癌症中佔第二位，其致病主因為感染高危險性人類乳突狀病毒(high-risk human papillomaviruses, HPVs)，如HPV-16, 18, 31, 33等。從病毒感染到癌症發生約需8-10年時間，但多數病人在感染HPV後會自行痊癒，僅少數病人會發展成癌症。對於何種因子會促使感染HPV的細胞從沒有症狀或輕微症狀的子宮頸上皮內贅瘤(Cervical intraepithelial neoplasia, CIN)發展成癌症則目前所得到的資料仍然有限。因此本研究欲從收集之子宮頸抹片細胞檢體中找出與子宮頸癌發生有關的標記(marker)。HPV-16佔子宮頸癌中50%以上高比率，因此本研究特別針對HPV-16進行探討。在已收集530位正常及CIN個案的子宮頸細胞檢體，經由特定HPV 引子: MY09/11及Gp5+/6+進行nested-PCR 發現202位HPV(+)個案，另外再由特定HPV-16 引子進行nested-PCR發現34位HPV-16(+)個案。本研究利用Bisulfite sequencing分析HPV-16(+)個案檢體中HPV long control region(LCR) CpG甲基化模式，本研究分析34檢體中只有15個檢體之LCR可成功放大分析，15位個案中一位為正常，ㄧ位為子宮頸發炎，ㄧ位CIN I，二位CIN II，6位CIN III以及一位子宮頸癌患者，這些個案之LCR均無甲基化情形，因此難以推論LCR甲基化與CIN之相關性。另外本研究同時發現2個案之LCR具有deletion情況，以及2個案之LCR序列具有變異性。具有deletion之個案分別為CIN III以及子宮頸癌患者，LCR序列具有變異性則為CIN II患者。因此未來需利用啟動子活性測試探討此變異與疾病發生之相關性。

關鍵字

人類乳禿狀病毒 ； 甲基化

並列摘要

A subset of HPV types are the causative agent of cervical cancer, and those types are categorized as high-risk types, eg. HPV 16, HPV 18. Infection of the genital tract by HPVs can initially result in cervical intraepithelial neoplasia (CIN), many of those lesions are cleared by the immune system. However, there is some of those lesions are developed to cervical cancer. Until recently, no test can predict whether CIN will develop into cancer or not. In this study, we are trying to find out a biomarker which can predict the CIN prognosis. We have collected 530 samples and which 202 are HPV positive samples. And in those samples we got 34 HPV-16 positive samples and analyzed the HPV-16 long control region (LCR) DNA methylation and deletion state. Only 15 samples of which LCR regions can be amplified and be analyzed, 1 is normal, 2 are CIN II, 6 are CIN III and 1 is cervical cancer. It is hard to conclude the relationship between methylation and CIN develop, because those sample are not methylated. We are discovered 2 samples which have the deletion within LCR and 2 samples are nucleotide variants. The deletion samples are CIN III and cervical cancer. The nucleotide variant samples are CIN II. Further studies are needed to determine the correlation between the promoter activity and the sequence variants and the correlation with the state of disease.

並列關鍵字

HPV ； methylation

參考文獻

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