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研究生:詹智宇
研究生(外文):Chan, Chih-Yu
論文名稱:當歸酒精萃取物抑制肺癌細胞轉移與增生
論文名稱(外文):Angelica sinensis ethanol extract inhibit Lewis lung cancer cell metastasis and proliferation
指導教授:吳彰哲
指導教授(外文):Wu, Chang-Jer
口試委員:廖光文詹伊琳張君如
口試委員(外文):Liao, Kuang-WenChan, Yi-LinChang, Chun-Ju
口試日期:2018-07-04
學位類別:碩士
校院名稱:國立臺灣海洋大學
系所名稱:食品科學系
學門:農業科學學門
學類:食品科學類
論文種類:學術論文
論文出版年:2018
畢業學年度:106
語文別:中文
論文頁數:44
中文關鍵詞:當歸酒精萃取物抑制轉移癌症惡病質免疫細胞數量
外文關鍵詞:Angelica sinensis ethanol extractMetastasisCachexiaImmune cell population
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癌症位居全球死因之首,而在臺灣又以肺癌致死率最高。肺癌之所以有極高 的致死率主要是因為初期診斷不易,當患者感受到明顯的不適感時,大多都已是 肺癌晚期或末期,癌症已有轉移擴散的現象,且在癌症後期,患者可能因癌症或 是治療上的副作用,進而產生癌症惡病質的症狀,導致治療效果不佳。近年來, 中西醫合併治療已越來越受到重視,當歸具有抗氧化、調節免疫以及抑制血小板 凝集等功效,本研究探討當歸酒精萃取物 (Angelica sinensis ethanol extract, ASE) 對於抗肺癌轉移與增生之效果。本實驗之樣品為乾燥當歸經研磨成粉後,以純酒 精於室溫下避光浸泡四天,再以減壓濃縮而得。體外試驗結果顯示 ASE 對於肺 癌細胞 (Lewis lung cancer cells, LLC) 具有毒殺效果,且在傷口癒合試驗與遷移 試驗中皆有效抑制 LLC 轉移。體內試驗以 C57BL/6 小鼠以皮下注射之方式建 立動物模型,並以化療藥物順鉑 (Cisplatin) 作為正控制組,結果顯示 ASE 有效 降低腫瘤重量與體積以及抑制癌細胞轉移到小鼠肺臟,而在癌症惡病質,亦觀察 到且 ASE 能減緩因惡病質所導致的體重下降脂肪組織減少以及血清中白蛋白表 達量下降等現象。進一步分析其他生理指標發現 ASE 能增加 T 免疫細胞的數量 與增加生物性抗氧化能力之趨勢,抑制腫瘤增生,且 ASE 未導致腫瘤背負小鼠 肝腎損傷。綜上期許未來能利用當歸酒精萃取物於輔助肺癌治療,以提升肺癌患 者之治療效果。
Lung cancer is a high mortality rate in the world. Because the initial diagnosis of lung cancer is not easy, most of the diagnoses are in the late stage of cancers. Most of the patients have had cancer metastasis, cancer cachexia and other treated side effect lead to poor treatment. Therefore, it is necessary to reveal novel methods against cancer and its complications. This study investigated investigated the therapeutic effects of Angelica sinensis ethanol extract (ASE), against cancer metastasis and cancer related cachexia in vitro and in vivo. In in vitro experiments, ASE was shown to have antioxidation and anti-proliferation in Lewis lung cancer cell (LLC). It also inhibited LLC cancer metastasis. In in vivo experiment, this study used LLC tumor-bearing mice as animal model. As a result, ASE could inhibit tumor growth, cancer cell metastasis, as well as attenuate the body weight decline, and white adipose tissue lose in animal model. Moreover, ASE increased population of cytotoxic T cells, and increased anti-oxidant capacity. In summary, these results revealed that ASE is able to inhibit tumor growth and ameliorate cachectic symptoms via activating immune function and anti-oxidative stress.
壹、前言................................................1
貳、文獻回顧.............................................2
一、癌症之特徵...........................................2
二、肺癌.................................................2
1. 肺癌流行病學..........................................2
2. 肺癌病理學............................................2
3. 肺癌之特徵............................................3
4. 肺癌之分期............................................3
5. 癌症惡病質 (Cancer cachexia)..........................4
6. 肺癌治療現況與困境....................................4
三、當歸及其活性物質......................................4
1. 當歸.................................................5
2. 阿魏酸 (Ferulic acid)................................5
3. 藳本內酯 (ligustilide)................................6
四、研究動機.............................................6
參、實驗流程設計.........................................7
一、當歸酒精萃取.........................................7
二、當歸酒精萃取物分析....................................7
三、體外試驗.............................................8
四、體內試驗.............................................8
肆、實驗材料與方法......................................10
一、實驗材料............................................10
1. 實驗細胞株...........................................10
2. 細胞培養相關試劑.....................................10
3. 當歸來源.............................................10
4. 抗氧化試劑...........................................10
5. 細胞存活率試劑.......................................10
6. 細胞轉移試驗相關試劑..................................10
7. 實驗動物.............................................10
二、儀器設備............................................11
三、實驗方法............................................12
1. 當歸酒精萃取.........................................12
2. 高效液相色譜法分析...................................12
3. 細胞株之保存活化與培養................................12
3.1 細胞株之保存........................................12
3.2 細胞株之活化........................................12
3.3 細胞培養............................................12
4. 細胞毒性測試.........................................13
5. 細胞遷移試驗.........................................13
5.1傷口癒合試驗.........................................13
5.2細胞遷移試驗.........................................13
6. 動物實驗.............................................13
7. 當歸酒精萃取物ASE抗氧化能力試驗.......................14
7.1 DPPH (α,α-diphenyl-β-picrylhydrazyl) 自由基清除能力測定.....................................................14
7.2 總抗氧化能力測定....................................14
7.3 細胞內過氧化氫含量測定...............................14
8. 免疫細胞分析.........................................15
9. 白蛋白分析...........................................15
10. 肝腎損傷分析........................................15
11. 統計分析............................................15
伍、結果................................................16
一、當歸酒精萃取物分析...................................16
1. HPLC分析............................................16
2. 當歸酒精萃取物抗氧化活性之探討.........................16
二、體外試驗............................................16
1. 當歸酒精萃取物對細胞毒性之影響........................16
2. 當歸酒精萃取物對LLC肺癌細胞株抑制轉移之效果............16
三、利用動物實驗探討當歸酒精萃取物對肺癌腫瘤背負小鼠之影響 .......................................................17
1. 當歸酒精萃取物對肺癌腫瘤背負小鼠之影響.................17
2. 當歸酒精萃取物抑制肺癌腫瘤增生.........................17
3. 當歸酒精萃取物抑制肺癌腫瘤轉移.........................18
4. 當歸酒精萃取物減緩肺癌腫瘤背負小鼠癌症惡病質之現象 .......................................................18
5. 當歸酒精萃取物對免疫細胞之調節作用.....................19
6. 當歸酒精萃取物抗氧化能力..............................19
7. 肝腎損傷分析.........................................20
陸、討論................................................21
柒、結論................................................24
捌、參考文獻............................................25
玖、圖表................................................31
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