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研究生:鍾怡萱
研究生(外文):I-Hsuan Chung
論文名稱:探討鼻咽癌中乳酸對於癌幹細胞的特性所造成的影響
論文名稱(外文):Investigating the Evolution of Cancer Stem Cells Fueled by Lactate in Nasopharyngeal Carcinoma
指導教授:陳燕彰陳燕彰引用關係
指導教授(外文):Yann-Jang Chen
學位類別:碩士
校院名稱:國立陽明大學
系所名稱:生命科學系暨基因體科學研究所
學門:生命科學學門
學類:生物訊息學類
論文種類:學術論文
論文出版年:2015
畢業學年度:103
語文別:英文
論文頁數:55
中文關鍵詞:癌幹細胞乳酸鼻咽癌
外文關鍵詞:Cancer stem cellsLactateWarburg effectReverse WarburgMetabolic reprogram
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癌症幹細胞為腫瘤中一群具自我更新能力的癌細胞,他們被認為是造成癌症復發及轉移的主因。癌細胞被認為能透過與腫瘤所處的微環境的交互作用,進而獲得幹細胞特性。許多研究發現腫瘤中有高濃度的乳酸堆積,並且發現惡性腫瘤中乳酸的堆積和癌症惡性程度的發展有著密切的關係。因此,本研究的目的為探討乳酸在調控癌症幹細胞特性中所扮演的角色以及其可能的機制。
為了瞭解乳酸在已分化的癌細胞中所扮演的角色,我們用乳酸處理鼻咽癌細胞株(TW01 &; HONE1),並且利用及時定量聚合酶連鎖反應去量測幹細胞相關基因及上皮細胞中胚轉化的相關基因的表現量,同時利用免疫螢光染色及西方墨點法去量測這些基因所產生的蛋白質的表現量。另外,我們也利用細胞行為分析實驗來檢測經乳酸處理過後的細胞是否增加癌幹細胞特性。
我們發現乳酸不僅刺激了幹細胞及上皮細胞中胚轉化的相關基因的表現,也造成帶有CD24、CD44表現的鼻咽癌幹細胞族群的增加。我們發現乳酸導致幹細胞特性及上皮細胞中胚轉化的情形會隨著時間的增長而有增加趨勢。另外我們也利用癌症幹細胞相關的行為分析實驗,例如腫瘤聚球分析法、細胞遷移分析法以及側群細胞分析法,這些發現顯示乳酸能使鼻咽癌細胞獲得幹細胞特性,進而具有長期自我更新及對治療有抵抗性的能力。
因此,我們認為乳酸,這種大量在腫瘤中堆積的代謝物,能夠將已分化的癌細胞轉化成癌幹細胞,進而發展成更惡性腫瘤。然而,乳酸導致癌幹細胞特性增加的詳細機制尚不清楚,這將會是我們的下一個需要深入研究的目標。

Cancer stem cells (CSCs) are a subpopulation of tumor cells that possess ability of self-renewal and are considered as the main causes of tumor recurrence and metastasis. Cancer cells may acquire stemness properties from tumor microenvironments. High concentration of lactate accumulated in tumors has been reported in many researches, and the accumulation of lactate in solid tumors may be a pivotal event in the development of malignancies. Thus, our aims in this study are to investigate the role of lactate involving in regulating stemness properties and its possible mechanism.
To understand the role of lactate in differentiated cancer cells, we treated NPC line, TW01 and HONE1 cells with 10 mM lactate to evaluate the expression of stemness and EMT genes by Western blot, RT-qPCR and immunofluorescence staining assay. Furthermore, we utilized behavior assay to examine whether these cells acquire CSC properties under lactate treatment.
We found that lactate treatment not only stimulated expressions of stemness and EMT related genes but also increased the CD24high/CD44high CSC-like subpopulation in TW01 parental cells. We also found the expression of stemness and EMT related genes increased along with duration of lactate treatment. Moreover, through behavior assays, such as tumor sphere assay, migration assay and side population assay, we found that lactate treated cells poses more CSC properties.
Collectively, these findings showed that lactate confers long-term self-renewal and therapeutic resistance in NPC cells. Thus, it implies that lactate, the high concentration metabolite in tumor microenvironment, can develop malignancy of NPC by reprograming differentiated cancer cells into CSCs.


中文摘要 i
Abstract ii
Contents iv
Introduction 1
Cancer stem cells 1
Nasopharyngeal carcinoma 6
Metabolism in cancer cells 8
Specific Aims 12
Materials and methods 13
I. Materials 13
II. Methods 13
Results 20
Lactate rescued the expression of stemness and EMT-related genes which were halted by glucose depletion 20
Knockdown of MCT1 gene abolishes the lactate-induced expression of stemness and EMT-related genes in low-glucose cultured CSCs 20
Lactate escalates the expression of stemness genes in TW01 parental cells 21
Lactate increased CD44high/CD24high CSC-like subpopulation in NPC parental cells 22
The lactate induced stemness and EMT increases in a time-dependent manner 23
Knockdown of the MCT1 gene leads to suppress the expression of lactate-induced genes 24
Lactate increased long-term anti-anoikis ability of cancer cells 24
Lactate treatment would increase migratory ability of cancer cells 25
Lactate treatment would increase side population of cancer cells 25
Knockdown of the ACLY gene lead to suppress the lactate-induced stemness properties 26
Discussion 27
Tables 38
Figures 41


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