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系統識別號 U0026-1607201317513600
論文名稱(中文) 探討CD90於癌細胞遷移所扮演之角色
論文名稱(英文) The role of CD90 in cancer cell migration
校院名稱 成功大學
系所名稱(中) 生物化學暨分子生物學研究所
系所名稱(英) Department of Biochemistry and Molecular Biology
學年度 101
學期 2
出版年 102
研究生(中文) 郭瑞智
研究生(英文) Jui-Chih Kuo
學號 s16004123
學位類別 碩士
語文別 中文
論文頁數 62頁
口試委員 指導教授-賴明德
口試委員-張文粲
口試委員-陳昌熙
中文關鍵字 CD90  腫瘤幹細胞  細胞遷移 
英文關鍵字 CD90  cancer stem cell  migration 
學科別分類
中文摘要 CD90為免疫球蛋白超家族(immunoglobulin superfamily)的一員,是一種藉由醣練鑲嵌於細胞膜上脂筏區(lipid rafts)的醣磷脂醯肌醇錨定醣蛋白(GPI-anchored protein)。過去的研究發現, CD90可作為一個肝癌的腫瘤幹細胞標誌(cancer stem cell maker),並具有參與細胞之間、細胞與基質之間訊息調節的功能,更可藉由整合蛋白(integrin)調控細胞的生理功能,如細胞黏著(cell adhesion)、細胞遷移(migration) 及纖維化(fibrosis)等,但是目前對於 CD90是否會影響肝癌的細胞遷移能力仍然沒有明確的研究報導。因此,在本篇研究中,我們想瞭解 CD90在肝癌中,是否會影響肝癌細胞的遷移能力?並嘗試瞭解 CD90在這過程中是扮演什麼樣的角色?為此,我們使用會穩定表現 CD90的肝癌細胞株,進行傷口癒合爬行能力分析(wound-healing assay)及細胞爬行能力分析(Boyden chamber assay),結果發現於肝癌細胞 Hep3B中表現 CD90時,有增進肝癌細胞 Hep3B之細胞遷移行為的現象。進一步以西方墨點法(western blot assay)來觀察細胞內部分子機制的變化,根據實驗的結果,我們發現於肝癌細胞 Hep3B中表現 CD90時,其磷脂醯肌醇-3激酶(phosphoinositide 3-kinase, PI3K)與粘著斑激酶(focal adhesion kinase, FAK)分子表現量有減少的傾向,另外也發現,於肝癌細胞 Hep3B中表現 CD90時,其鈣黏著素E (E-cadherin)表現量有減少的傾向,而鈣黏著素N (N-cadherin)表現量有增加的傾向。本篇研究發現 CD90在肝癌細胞 Hep3B中會降低細胞粘著的能力,產生表皮-間質轉換 (epithelial-mesenchymal transition, EMT)的現象,並增進肝癌細胞 Hep3B的細胞遷移能力,具有造成癌症轉移的風險。
英文摘要 CD90 is a member of immunoglobulin supergene family, and is a glycosylphosphatidylinositol-anchored glycoprotein, which localizies at cell lipid rafts on the outer surface of the lipid bilayer. Previous studies have suggested that CD90 could be a liver cancer stem cell marker, and it is play an important role in cell-cell and cell-matrix interactions via integrin signaling pathway, and also affects a lot of biological processes, including cellular adhesion, migration and fibrosis, but it is unclear whether CD90 will affect liver cancer migration. In this study, we want to investigate the role of CD90 in cancer cell migration and whether CD90 affect signal transduction in other pathway. For this reason, we analyzed the cell migration of stable cells expressing exogenous CD90 with wound healing assay and Boyden chamber assay. The result showed that ectopic expression of CD90 enhanced cell migration in Hep3B hepatoma cell line. By western bolt assay, we also found that ectopic expression of CD90 decreased the level of phosphoinositide 3-kinase, focal adhesion kinase and E-cadherin, but increased the level of N-cadherin in Hep3B cell. Our results suggested that CD90 facilitated Hep3B hepatoma cell migration and epithelial-mesenchymal transition, reduced ability of cell adhesion, and had high chance to cause cancer metastasis.
論文目次 目錄

緒論 1
一、細胞遷移與癌症轉移 1
二、腫瘤幹細胞於癌症中所扮演之角色 4
三、腫瘤幹細胞與表皮-間質轉換行為 6
四、腫瘤幹細胞與原發性肝癌 7
五、CD90於癌症中所扮演之角色 9
六、研究目標與策略 10

材料與方法 12
一、細胞培養與繼代保存(Cell culture) 12
二、西方墨點法(Western blot assay) 17
三、細胞爬行能力分析(Boyden chamber assay) 25
四、傷口癒合爬行能力分析(Wound-Healing assay) 28

實驗結果 29
一、CD90不影響肝癌細胞 Hep3B、HepG2及卵巢癌細胞 SKOV3之傷口癒合爬行能力 29
二、CD90會影響肝癌細胞 Hep3B及卵巢癌細胞 SKOV3之細胞爬行能力 29
三、CD90會影響肝癌細胞 Hep3B及卵巢癌細胞 SKOV3之細胞遷移相關分子的表現 31
四、CD90會影響肝癌細胞 Hep3B及卵巢癌細胞 SKOV3之 EMT相關分子的表現 32

討論 35

結論 39

參考文獻 40

附圖 46

圖目錄

圖一、CD90不影響肝癌細胞 Hep3B之傷口癒合爬行能力 46
圖二、CD90不影響肝癌細胞 HepG2之傷口癒合爬行能力 48
圖三、CD90不影響卵巢癌細胞 SKOV3之傷口癒合爬行能力 50
圖四、CD90會影響肝癌細胞 Hep3B之細胞爬行能力 52
圖五、CD90不影響肝癌細胞 HepG2之細胞爬行能力 54
圖六、CD90會影響卵巢癌細胞 SKOV3之細胞爬行能力 55
圖七、CD90會影響肝癌細胞 Hep3B之細胞遷移相關分子的表現 57
圖八、CD90不影響肝癌細胞 HepG2之細胞遷移相關分子的表現 58
圖九、CD90會影響卵巢癌細胞 SKOV3之細胞遷移相關分子的表現 59
圖十、CD90會影響肝癌細胞 Hep3B之 EMT相關分子的表現 60
圖十一、CD90不影響肝癌細胞 HepG2之 EMT相關分子的表現 61
圖十二、CD90會影響卵巢癌細胞 SKOV3之 EMT相關分子的表現 62
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