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研究生:陳穎儀
研究生(外文):Ying-Yi Chen
論文名稱:牛樟芝乙醇萃取物抑制人類非小細胞肺癌以及人類肝癌細胞遷移相關分子機制
論文名稱(外文):Ethanolic extract of Antrodia cinnamomea inhibitshuman non-small cell lung carcinoma and hepatocellular carcinoma migration and its related mechanisms
指導教授:許明志
學位類別:博士
校院名稱:中國醫藥大學
系所名稱:藥學系博士班
學門:醫藥衛生學門
學類:藥學學類
論文種類:學術論文
論文出版年:2015
畢業學年度:103
語文別:中文
論文頁數:109
中文關鍵詞:牛樟芝蕈類中草藥肺癌肝癌
外文關鍵詞:Antrodia cinnamomeamushroomChinede herballung cancerliver cancer
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牛樟芝(Antrodia cinnamomea; AC)為台灣特有的藥用真菌,寄生於牛樟樹上,是常見的傳統中藥,目前對樟芝的研究已顯示具有抗發炎、抗氧化與抗腫瘤等功效。癌細胞轉移的過程中通常伴隨著細胞外基質(extracellular matrix; ECM) 的分解,而主要扮演分解ECM 的角色為基質金屬蛋白??(Matrix metalloproteinase; MMPs)。本研究先以MTT 測試細胞存活率,在對於細胞較不具毒殺性且存活率皆大於80%以上的濃度下進行實驗。首先在細胞遷移試驗 (wound healing assay) 與細胞移行試驗 (trans well migration assay) 方法中顯示,隨著EEAC 濃度增加,可以抑制肺癌與肝癌細胞株的遷移和移行能力。另外,由西方墨點法結果顯示EEAC 能降低MMP-9 與MMP-2 的蛋白表現,以及MMP-9與MMP-2 的活性。此外,EEAC 也增加基質金屬蛋白?“磻蹌狼IMP-1、TIMP-2 的蛋白表現。進一步探討訊息傳遞,EEAC 可以降低ERK1/2、P38 與JNK1/2 的磷酸化,在肺癌細胞中,結果顯示加了PI3K 抑制劑LY294002、ERK 抑制劑PD98059、p38 抑制劑SB203580及JNK 抑制劑SP600125 可以降低MMP-9 與MMP-2 的蛋白表現,若再加上EEAC 則能更顯著降低MMP-9 與MMP-2 的蛋白表現。因此,本實驗在觀察細胞遷移與移行相關蛋白的表現皆顯示EEAC 具有降低肺癌與肝癌細胞株移行的能力,且可能透過抑制ERK1/2、P38 與
JNK1/2 路徑影響MMP-9 與MMP-2 的表現進而抑制肺癌與肝癌細胞株
的遷移。由以上的實驗結果得知,藉由探討與細胞增生以及遷移相關
的蛋白質表現,EEAC 可以減緩細胞移行,來抑制肺癌與肝癌細胞株
的生長,並且抑制細胞遷移。故牛樟芝是很有潛力可發展成抑制肺癌
VI與肝癌細胞株遷移的抗癌藥物。

Niu-Chang-Chih (Antrodia cinnamomea; AC) is unique to Taiwan medicinal fungi, parasites on the stout camphor tree, is a common traditional Chinese medicine.It has been shown to exhibit anti-inflammatory, antioxidant and anti-tumor effects. Process cancer metastasis is usually accompanied by extracellular matrix (extracellular matrix; ECM) degradation. Matrix metalloproteinase (matrix metalloproteinase; MMPs) plays the crucial role for the degradation of ECM. In this study, first to investigated cell viability by MTT assay. For cancer cells, we chosed non toxic and survival rate more
than 80% concentration for our experimental concentration. In cell migration assay (wound healing assay and trans well migration assay)results showed that EEAC could inhibit cancer cells migration and migration ability of lung and liver cancer cell lines when increasing EEAC concentration. According to western blot results showed that EEAC reduced MMP-9 and MMP-2 protein expression, and activity of MMP-9 and MMP-2. In addition, EEAC also increased matrix metalloproteinase inhibitor
TIMP-1, TIMP-2 protein expression. Further studies showed that EEAC regulated MMP-9 and MMP-2 production via MAPK signaling pathway, as evidenced by the findings that EEAC inhibited the phosphorylation of ERK1/2, P38 and JNK1/2. In lung cancer cells showed that the addition of
PI3K inhibitor (LY294002), ERK inhibitor (PD98059), p38 inhibitors (SB203580) and JNK inhibitor (SP600125) resulted in reduced activities of
MMP-9 and MMP-2 protein expression, and when added with EEAC could significantly reduced the protein expression of MMP-9 and MMP-2. Thus,results showed that EEAC could reduced lung cancer and liver cancer cell line migration, possibly through the inhibition of ERK1 / 2, P38 and JNK1 /2 pathways. These results suggested that EEAC could reduce the migration via MAPK signaling pathways in human lung cancer and liver cancer cells,and may use EEAC as an anti-migration agent in theprevention and treatment of human lung cancer and liver cancer.

總目錄 I
圖目錄 II
表目錄 IV
中文摘要 V
英文摘要 VII
第一章 前言 1
第一節 牛樟芝之文獻考察 1
第二節 肺癌簡介 9
第三節 肝癌簡介 10
第四節 腫瘤侵入與轉移 10
第五節 研究目的12
第二章 研究材料和方法 14
第一節 研究設計 14
第二節 研究材料 14
第三節 實驗方法 26
第三章 結果 34
第四章 討論 44
第五章 結論 50
第六章 參考文獻 51

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