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系統識別號 U0026-2706201001360500
論文名稱(中文) 三氧化二砷對微膠細胞株細胞毒性之影響
論文名稱(英文) Cytotoxic effect of arsenic trioxide on microglial cell line
校院名稱 成功大學
系所名稱(中) 生命科學系碩博士班
系所名稱(英) Department of Life Sciences
學年度 98
學期 2
出版年 99
研究生(中文) 温詩婷
研究生(英文) Shih-Ting Weng
電子信箱 ice.desert@msa.hinet.net
學號 L5697110
學位類別 碩士
語文別 中文
論文頁數 57頁
口試委員 指導教授-曾淑芬
口試委員-李佳穎
口試委員-侯平君
口試委員-王浩文
中文關鍵字 無機砷  三價砷  三氧化二砷  五價砷  血腦屏障  微膠細胞  中樞神經系統  乙醯基半胱胺酸 
英文關鍵字 inorganic arsenic  arsenite  arsenic trioxide  arsenate  blood-brain-barrier  microglia  central nervous system  N-Acetyl cysteine 
學科別分類
中文摘要 砷 (arsenic) 以不同氧化態 (+3,+5) 且多種複合物形式廣泛分布於環境中,若無機砷經攝食或飲水進入哺乳動物體內,會經由一連串氧化還原及甲基化的代謝過程。許多流行病學研究證實,生活在砷汙染環境下的人們,較容易造成身體不適或生成相關疾病,例如癌症。而砷也被發現會傷害到人類的記憶以及學習行為。近年來更有研究報告指出,三氧化二砷的代謝產物可能會進入血腦屏障 (Blood-Brain Barrier),累積在腦部區域而產生神經毒性。微膠細胞 (Microglia) 在中樞神經系統(CNS)扮演著微環境變化的偵測者及免疫防禦者角色,他們不僅在發育中的CNS協助組織重組及神經細胞的正常生長,也能在CNS受傷後快速地被活化,並進行一連串的發炎反應。因此,想了解砷對於微膠細胞是否會造成影響。首先,以微膠細胞株BV2與單核巨噬細胞株Raw 264.7進行MTT測試,觀察三價及五價砷對細胞的存活率之影響。結果發現,五價砷的細胞毒性較三價砷低,而微膠細胞株則比周邊血液的巨噬細胞對於砷的毒性較為敏感,且三價砷的毒性會隨著劑量升高或暴露時間增長而越大。此外,若處理乙醯基半胱胺酸(N-Acetyl cystein,NAC),一種榖胱胺肽(Glutathione) 的合成先驅物及解毒劑,來觀察細胞的存活率,結果發現NAC能保護微膠細胞免於三價砷所誘發的細胞毒性。另外,我們還使用多種天然甲醇萃取物來檢測其對微膠細胞存活率的影響。結果發現天然萃取物中可能含有對微膠細胞有毒的未知物質。
英文摘要 The arsenic wildly distributes in various forms with different oxidation states (+3,+5) in the environment. In mammalian after ingesting inorganic arsenic (iAs), it goes through a sequential redox and methylation processes. Epidemiological evidence has demonstrated that people exposed to arsenic can be increased at risk for serious illness or diseases, such as cancer. Arsenic is also found to impair human memory and learning behaviors. Evidence has also shown that arsenic trioxide metabolites are able to penetrate blood-brain-barrier (BBB), accumulate in the brain, and induce neurotoxicity. Microglia, the resident macrophages of the central nervous system (CNS), play the important roles in micro-environmental sensor and immune defender in the CNS. These cells are responsible for tissue remodeling and neurite outgrowth in developing CNS. Moreover, microglia are rapidly activated after the CNS injuries, and generate a series of inflammatory reactions. Therefore, we attempted to understand the effect of arsenic on microglia. At first, we examined the cell viability of the immortalized microglial cell line BV2 and leukaemic monocyte macrophage cell line Raw 264.7 by arsenite [As(III)] and arsenate [As(V)]. The results showed that As (V) exerted less cytotoxicity. Moreover, microglia were more susceptible to As(III) and As(V) compared to macrophages. As(III) induced cytotoxicity of microglia is dose- and is time-dependent. Furthermore, we found that N-Acetyl cysteine (NAC), a precursor for the synthesis of glutathione and a detoxifying agent in the body, can rescue microglia from As(III)-induced cytotoxicity. In contrast, we also examined the effect of the tested natural methanol extracts on cell survival and found that there were undefined toxic compounds derived from those natural extracts to CNS macrophages.
論文目次 中文摘要 1
Abstract 2
致謝 3
目錄 4
圖表目錄 6
縮寫表 8
前言 9
一、 中樞神經系統的微膠細胞 9
1. 微膠細胞的來源與分布 9
2. 微膠細胞的功能 9
3. 微膠細胞與神經退化性疾病 10
二、 環境中的砷 10
三、 砷的代謝及累積 11
四、 砷中毒及相關疾病 12
1. 急性中毒 12
2. 慢性砷中毒 13
五、 砷的累積對中樞神經系統的傷害 14
研究目的 16
材料與方法 17
一、 材料 17
二、 實驗方法 17
1. 細胞培養 17
2. 細胞活性分析 (cell viability analysis ) 18
3. 天然甲醇萃取物製備 19
4. 統計分析 19
實驗結果 20
一、 砷對微膠細胞株BV2及巨噬細胞株Raw 264.8的毒性傷害 20
三氧化二砷 (As2O3) 20
砷酸鈉(Na2HAsO4) 21
二、 乙醯基半胱氨酸(N-acetyl-cysteine; NAC)對三氧化二砷 (As2O3)誘發的BV2細胞毒性之影響 21
三、 抗壞血酸 (Ascorbic acid) 與白藜蘆醇 (Resveretrol)對三氧化二砷 (As2O3) 誘發的細胞毒性之影響 22
四、 三十五種天然物對As2O3誘發的BV2細胞毒性之影響 23
討論 26
一、 三價與五價的砷化物對細胞毒性之探討 26
二、 砷對微膠細胞的毒性影響 28
三、 抗氧化劑對砷中毒的保護作用 28
四、 天然物萃取物 (中藥材) 對微膠細胞的影響 31
五、 結論 32
參考文獻 33
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