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研究生:王若瑜
研究生(外文):Jo-YuWang
論文名稱:蟲草素與順鉑對口腔癌細胞在細胞凋亡中的抗癌效果
論文名稱(外文):Anticancer Effect of Cordycepin and Cisplatin on Apoptosis in Oral Cavity Cancer Cells
指導教授:黃步敏
指導教授(外文):Bu-Miin Huang
學位類別:碩士
校院名稱:國立成功大學
系所名稱:細胞生物及解剖學研究所
學門:醫藥衛生學門
學類:醫學學類
論文種類:學術論文
論文出版年:2012
畢業學年度:100
語文別:英文
論文頁數:50
中文關鍵詞:順鉑蟲草素細胞凋亡合併處理加乘性口腔癌
外文關鍵詞:Cisplatincordycepinapoptosiscombination treatmentsynergetic effectoral cancer
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檳榔引發的口腔癌在許多領域都很常見,包括在台灣。順鉑 (Cisplatin) 是一種用於治療口腔癌的的基礎的化療藥物。然而近年來許多研究指出順鉑會有抗藥性的產生。蟲草素 (3'-脫氧腺苷) (Cordycepin) 為一從冬蟲夏草中分離出的純化合物,已被證明具有抗腫瘤的特性。最近的研究表示,合併使用化療藥物或許可以對抗抗藥性的產生。因此,我們研究了在不同口腔癌的細胞株 (OC3,OEC-M1和Fadu) 中蟲草素以及順鉑的抗癌作用。結果發現,蟲草素(100 μM),順鉑(300 μM和600 μM)單獨處理,以及合併使用 (蟲草素100 μM合併順鉑300 μM或600 μM)24小時後,在形態學變化下可觀察出三個細胞株皆會傾向細胞凋亡。在細胞存活試驗 (MTT assay)中,所有細胞株隨著蟲草素以及順鉑劑量的增加(蟲草素10 μM至1000 μM,順鉑100 μM至1000 μM)存活率顯著下降 (p〈0.05),同時可觀察出蟲草素(100 μM)與順鉑(300μM和600 μM)合併處理達到加乘的效果。流式細胞儀檢測中,細胞百分比在蟲草素組別中相較於對照組在subG1週期顯著增加了2〜3倍。蟲草素及順鉑(300 μM)合併處理組的subG1期細胞比例顯著增加至20%。在細胞機制研究中,蟲草素及順鉑誘導caspase-8,caspase-9,caspase-3以及PARP的裂解,同時在合併處理組別中可觀察出表現量較單獨處理組別更高。在MAPKs途徑研究中,合併處理會活化JNK,ERK和p38的表現。綜合以上結果可得知,順鉑和蟲草素會藉由活化外在/內在caspase和MAPKs途徑引發人類口腔癌細胞株的細胞凋亡,高度表明了蟲草素及順鉑的組合可能具有潛在的加乘性抗癌效果。

Betel quid-related oral cavity cancer is common in certain areas, including Taiwan. Cisplatin is a platinum-based chemotherapy drug used to treat oral cancer. However, the development of drug-resistance to cisplatin has been reported recently. Cordycepin (3’–deoxyadenosine), a pure compound isolated from Cordyceps sinensis, has been demonstrated to have anti-tumor properties. Recent studies illustrate that combining chemotherapy agents may counteract drug resistance. Hence, we investigated the anticancer effect of cordycepin and cisplatin on different oral cancer cell lines; OC3, OEC-M1 and Fadu. Results demonstrated that cordycepin (100 μM), cisplatin (300 μM and 600 μM) and combined treatments (cordycepin 100 μM with cisplatin 300 μM or 600 μM ) caused cell death morphological changes after 24 hour treatment among 3 cell lines. In viability test, all three cell lines surviving rate significantly decreased as the dosage of cordycepin and cisplatin increased (10 μM to 1000 μM of cordycepin and 100 μM to 1000 μM of cisplatin), and the synergistical effect could be observed in cordycepin (100 μM) with cisplatin (300 μM and 600μM, respectively) treatments. In flow cytometry assay, percentage of subG1 phase cells in cordycepin group significantly increased by 2~3 times than control group. The percentage of subG1 phase cells in cordycepin and cisplatin (300 μM) co-treatment group significantly increased to 20%. In cellular mechanism study, both cordycepin and cisplatin induced cleavage of caspase-8, -9, and -3 and cleavage of poly ADP-ribose polymerase (PARP), and co-treatments induced more expression compared to cisplatin or cordycepin alone treatment among 3 cell lines. In the MAPKs pathway, co-treatments activated more expression of JNK, ERK and p38 than alone treatment among 3 cell lines. In conclusion, cisplatin and cordycepin possess synergistically/addictively apoptotic effect by activating the expression of extrinsic/intrinsic caspase and MAPK pathways in human oral cavity cancer cell lines, which highly suggest that the combination of cordycepin and cisplatin might be a potential anti-cancer drug compared to the single agent chemotherapy.
ABSTRACTS
Chinese abstract …………………………………………i
English abstract …………………………………………ii
ACKNOWLEDGEMENTS …………………………………………iv
TABLE OF CONTENTS …………………………………………v
LIST OF FIGURES …………………………………………vi
INTRODUCTION …………………………………………1
METERIALS AND METHODS
Chemicals …………………………………………5
Cells and cell culture …………………………………………5
MTT cell viability test …………………………………………6
Morphology study …………………………………………7
Flow cytometry analysis …………………………………………7
Immunoblotting Analysis …………………………………………8
Statistics …………………………………………8
RESULTS
Effects of cordycepin and/or cisplatin on cell viability of oral cavity cancer cells…………………………………………10
Effect of cordycepin and/or cisplatin on morphological change in oral cavity cancer cells………………11
The analysis of cell cycle under cordycepin and/or cisplatin influence in oral cavity cancer cells………12
Modulation of the effect on extrinsic and intrinsic pathways of apoptosis………………………………………………………13
Effect on the regulation of MAPK pathway……………………………16
DISCUSSION …………………………………………18
REFERENCES …………………………………………43
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