臺灣博碩士論文加值系統

牛樟芝（Antrodia camphorata）屬多孔菌科，只生長於台灣特有牛樟樹，生長緩慢且數量稀少。傳統上樟芝子實體用來治療腹瀉、腹痛、高血壓與肝癌，樟芝子實體與菌絲體萃取物的生物功能過去已被廣泛的研究，包括抗氧化、抗發炎、抗癌，然而有關樟芝成分結構及其生物活性則鮮少被研究。本研究以合成的樟芝菌絲體中馬來酸衍生物camphorataimide B來進行實驗，探討其對乳癌細胞MDA-MB-231的侵犯、轉移作用的影響。首先我們研究camphorataimide B是否可以抑制乳癌細胞增生，以細胞毒性分析（MTT）、細胞聚落形成（colony formation assay），發現camphorataimide B能有效抑制乳癌細胞增長，接著以DAPI染色和DNA斷裂進行分析，結果發現大於10 μM的camphorataimide B會造成乳癌細胞的細胞凋亡現象；也發現camphorataimide B可以誘導MDA-MB-231細胞造成細胞週期停止於G2/M期。再來我們以細胞發散性測定、F-actin 的螢光染色、傷口癒合、Boyden chamber 移動等試驗看到camphorataimide B會抑制乳癌細胞的移動能力。更進一步我們想要了解camphorataimide B會不會也抑制了乳癌細胞的侵犯能力，所以我們先進行貼附能力試驗，再以Boyden chamber 模擬細胞基質進行侵犯試驗，結果顯示camphorataimide B也抑制了乳癌細胞的侵犯作用。再來利用西方墨點法、zymography 的分析方式分析細胞侵犯相關蛋白及蛋白酶的表現，最後發現camphorataimide B是抑制了組蛋白酶（cathepsin D）的表現，同時我們也發現了調控cathepsin D的轉錄因子缺氧誘導因子（hypoxia-inducible factor-1α, HIF-1α）也受到了camphorataimide B的抑制。同時我們利用動物實驗模式研究camphorataimide B對乳癌細胞在裸鼠中肺部移生的影響，結果發現camphorataimide B減少肺部大小、重量和肺部移生現象。

Abstract
Antrodia camphorata, known as“niu-chang-chin”in Taiwan, is a fungus of the family polyporaceae that grows slowly and rare species. Traditionally, the fruit body of Antrodia camphorata used to treat diarrhea, abdominal pain, hypertension, and liver cancer. Many previous studies reported that the extract of fruit body and mycelium of Antrodia camphorata possesses a wide range of biological functions, such as antioxibant, anti-inflammation, anticancer. However, the relationship of structure bioactive component of Antrodia camphorata and bioactivity is not well known. In this studies, we investigated the anticancer potential of synthetic camphora-
taimide B, the maleic derivatives from the mycelium, on MDA-MB-231 breast cancer cells. First, we studied whether camphorataimide B can inhibit breast cancer cells growth by cytotoxicity assay and colony formation assay. The results showed that camphorataimide B can restrain the proliferation of breast cancer cells. In addition, by
DAPI staining and DNA fragmentation assay, the results found camphorataimide B induced apoptosis of MDA-MB-231 B breast cancer cells at concentrations of 10-40 μM. Furthermore, camphorataimide B was able to induce cell cycle arrest in G2/M phase. Then, we performed cell scattering, F-actin fluoresence staining, wound healing, and Boyden chamber migration assay to examine anti-migration of camphora
-taimide B on breast cancer cells. Moreover, we want to know whether camphorataimi
-de B will also inhibit breast cancer cell invasion. We conducted adhesion assay and Boyden chamber invasion assay. We found that camphorataimide B also inhibited breast cancer cells invasion. By weatern blotting analysis, it showed camphora
-taimide B suppressed the expression of cathepsin D and HIF-1α, a transcription factor of cathepsin D. Finally, we used animal models to study camphorataimide B on lung colonization of MDA-MB-231 cells in nude mice. The results showed camphora-
taimide B reduced the size and weigth of lung as well as lung colonization of MDA-MB-231 cells.

目 錄

縮寫表 .................................................... II
中文摘要 ....................................................... 01
英文摘要 ....................................................... 02
壹、 緒論 ...................................................... 04
第一部分 癌症（Cancer） ..................................... 04
第二部分 藥物來源 .......................................... 06
第三部分 癌症轉移（metastasis）............................... 08
貳、 研究背景與動機 ............................................ 14
參、 實驗材料與實驗方法 ........................................ 15
肆、 實驗結果 .................................................. 30
伍、 討論 ...................................................... 40
陸、 參考文獻 .................................................. 43
柒、 實驗結果圖表 .............................................. 48
捌、 附圖 ...................................................... 76

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