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研究生:陳韻宜
論文名稱:轉形生長乙型因子誘導基因在人類食道癌細胞中之表現
論文名稱(外文):Expression of Transforming Growth Factor-β-inducible Gene on Human Esophageal Carcinoma Cells
指導教授:翁芬華
學位類別:碩士
校院名稱:國立陽明大學
系所名稱:公共衛生研究所
學門:醫藥衛生學門
學類:公共衛生學類
論文種類:學術論文
論文出版年:2003
畢業學年度:91
語文別:中文
論文頁數:64
中文關鍵詞:食道癌
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食道癌和所有其他的癌症一樣,致病機轉目前並不清楚。且食道癌的預後差,五年活存率也低,因此早期診斷乃是治療食道癌最重要的法則。為尋找與食道癌相關的基因,以利在未來做為食道癌診斷標記和預後指標,我們利用Subtractive hybridization技術篩選與食道癌相關之基因。並利用反轉錄作用─聚合酵素連鎖反應(RT-PCR)與即時定量聚合酵素連鎖反應(Real-time PCR)觀察選出之基因在10對食道癌病人腫瘤組織鄰近的正常食道組織與腫瘤組織中的表現。
其中Hs.69559、Hs.301444、Hs.283714、Hs.118787等四個基因在10對食道癌病人腫瘤組織鄰近的正常食道組織與腫瘤組織中的表現是具有統計學上顯著差異性的。另外Hs.351928這個基因在10對食道癌病人組織中皆是腫瘤組織表現高於腫瘤組織鄰近的正常食道組織。Hs.118787這個基因為轉形生長乙型因子誘導基因(Transforming Growth Factor-β-inducible gene,TGFBI),在10對食道癌病人組織中有9對腫瘤組織表現高於腫瘤組織鄰近的正常食道組織,且表現量皆大於2倍,且在10株食道癌細胞株中TGFBI也有高量的表現,為食道癌病人腫瘤組織鄰近的正常食道組織平均表現量之20倍。
同時以北方墨點法分析發現IGF-1、EGF、IL-6皆會誘導TGFBI的表現。顯示TGFBI並非只受到TGF-β誘導表現,也可能受到其他生長因子的誘導而增加表現量。TGFBI在食道癌細胞中的作用與所扮演的角色值得進一步研究。
目錄
中文摘要
英文摘要
第一章 緒論………………………..………………………………………………1
一、 關於食道癌………………………………………………………………1
二、目前已知與食道癌相關之基因…………………………………………4
三、關於TGFBI……………………………………………………………..6
四、研究目的………………………………………………………………....9
第二章 實驗材料與方法………………………………..…………………….….10
一、細胞培養……………………………………………………….……….10
二、萃取與定量核醣核酸……………………….…………………….…….11
三、反轉錄作用─聚合酵素連鎖反應………..………………….….……...13
四、即時定量聚合酵素連鎖反應…………………………………………...16
五、北方墨點法………………………………………………………………17
第三章 結果………………………..……………………………………………..21
一、以減除雜交法、微陣列分析及National Cancer Institute資料庫中查詢分析比較各種基因在食道癌病人腫瘤組織鄰近的正常食道組織
與腫瘤組織檢體中之表現……………………………………………...21
二、 反轉錄作用─聚合酵素連鎖反應(RT-PCR)與即時定量聚合酵素連鎖反應(Real-time PCR)比較分析選出之基因在食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中的表現…………………………..22
三、以即時定量聚合酵素連鎖反應(Real-time PCR)比較分析選擇之基因在食道癌細胞株中的表現……………………………………………...25
四、以北方墨點法(Northern blot)分析TGFBI這個基因在食道癌細胞株CE81T/VGH經由IGF、EGF、IL-6、TGF-β處理後的表現………..27
第四章 討論…………………………………………………………….………...30
參考資料……………………………………………………………..….………...34
表……………………………………………………………………..….………...40
表一、選擇的基因……………………………………………..….………...40
表二、以反轉錄作用─聚合酵素連鎖反應(RT-PCR)與即時定量聚合酵素連鎖反應(Real-time PCR)比較分析在11對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中各種新基因RNA的表現…42
表三、以反轉錄作用─聚合酵素連鎖反應(RT-PCR)與即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在11對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中各種已知基因RNA的表現…
……………………………………………………………………….43
表四、分析比較以即時定量聚合酵素連鎖反應(Real-time PCR)在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中具有顯著差異性的5個未知與已知基因其在各對組織中之比值…………44
表五、利用減除雜交法(Subtractive hybridization)分析比較以即時定量聚合酵素連鎖反應(Real-time PCR)在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中具有顯著差異性的5個未知與已知基因在食道癌、口腔癌、鼻咽癌與子宮頸癌中之表現………….45
圖……………………………………………………………………..….………...46
圖一、研究方向………………………………………………..….……….…46
圖二、以反轉錄作用─聚合酵素連鎖反應(RT-PCR)分析比較在11對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中各種基因RNA的表現…………………………………………………………47
圖三、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.279789基因RNA的表現………………………………………..48
圖四、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.9343基因RNA的表現……………………………………………………49
圖五、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.158688基因RNA的表現………………………………………..50
圖六、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.351928基因RNA的表現………………………………………..51
圖七、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.69559基因RNA的表現…………………………………………………....52
圖八、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.77665基因RNA的表現.…………………………………………………...53
圖九、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.12702基因RNA的表現.…………………………………………………...54
圖十、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.301444基因RNA的表現………………………………………..55
圖十一、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.75748基因RNA的表現……………………………………….56
圖十二、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.118787(TGFBI)基因RNA的表現…………………………….57
圖十三、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.283714基因RNA的表現……………………………………...58
圖十四、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.325093基因RNA的表現……………………………………...59
圖十五、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.49349基因RNA的表現…………………………………….…60
圖十六、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和腫瘤組織中Hs.350547基因RNA的表現……………………………………...61
圖十七、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和10種食道癌細胞株中Hs.301444基因RNA的表現………………………………62
圖十八、以即時定量聚合酵素連鎖反應(Real-time PCR)分析比較在10對食道癌病人腫瘤組織鄰近的正常食道組織和10種食道癌細胞株中TGFBI基因RNA的表現…………………………………….63
圖十九、以北方墨點法(Northern blot)分析以EGF 100ng/ml、IL-6 100ng/ml、TGF-β 1ng/ml、IGF-1 100ng/ml處理食道癌細胞株(CE81T/VGH)後TGFBI基因RNA的表現…………………...64
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