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研究生:王秀琦
研究生(外文):Shiu-Chi Wang
論文名稱:鞣花酸對於脂多醣體(LPS)誘導之小鼠巨噬細胞的抗發炎作用研究
論文名稱(外文):The anti-inflammatory action of ellagic acid in LPS-induced macrophage cell line RAW264.7
指導教授:蒙美津蒙美津引用關係邱駿紘邱駿紘引用關係
指導教授(外文):Mei-Chin MongChun-Hung Chiu
學位類別:碩士
校院名稱:亞洲大學
系所名稱:生物科技學系碩士班
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
論文出版年:2007
畢業學年度:95
語文別:中文
論文頁數:76
中文關鍵詞:鞣花酸發炎反應誘導型一氧化氮合成酶
外文關鍵詞:Ellagic acidInflammationiNOS
相關次數:
  • 被引用被引用:6
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  • 下載下載:104
  • 收藏至我的研究室書目清單書目收藏:5
發炎現象是身體對付組織損傷、感染或刺激物的方式,其可被非特異性及特異性反應所啟動。而發炎亦為一種可將傷害侷限在局部區域的局部反應,可導致紅、腫、熱、痛。發炎級聯(cascades)的有益作用及發炎對組織的長期性潛在傷害之間存在著微妙的平衡性。假如發炎現象沒有被控制或消退,將會導致諸如慢性氣喘、風濕性關節炎、心血管疾病及特定癌症等疾病的發展。
在多種與發炎相關的病狀中,一氧化氮會顯著地增加。一氧化氮具有包括血管擴張、神經傳導及細胞毒性等多重生物活性。一氧化氮的自由基特性及與過氧化物的高度反應性,使得一氧化氮成為有效的促氧化物質,因而可誘發氧化傷害對多種胞內標的物的潛在性害處。由於一氧化氮的生成控制對人類健康很重要,很多研究團隊因而聚焦在探討一氧化氮由保護轉為有害作用上的機制。
鞣花酸是一種天然存在的植物多酚類,文獻報導顯示鞣花酸對多種致癌物有抗癌及抗致突變的特性。此外,在體外或體內實驗中,鞣花酸都有抗氧化的功效。為了延伸目前對鞣花酸的瞭解,本研究探討鞣花酸對小鼠巨噬細胞株RAW264.7 經脂多醣誘發後之一氧化氮、腫瘤壞死因子-α(TNF-α)及介白素-6(IL-6)的生成,並闡述可能的相關機轉。
研究結果顯示,隨著鞣花酸劑量的增加,能明顯降低因脂多醣(LPS)誘發的一氧化氮產生量。而且, TNF-α 和 IL-6 的產生量也被鞣花酸所抑制。與脂多醣單獨處理比較,鞣花酸的存在可使誘導型一氧化氮合成酶(iNOS)的蛋白質及 mRNA 表現均明顯下降。鞣花酸的添加也會增加IκB 及NF-κB 的表現量。綜合以上結果可知,鞣花酸或許可應用在巨噬細胞調控之發炎性疾病的控制上。
Inflammation is the body’s way of dealing with tissue damage, infection or irritants and can be initiated by non-specific and specific immune reaction. Inflammation is a local response which limits damage caused by injury to a local site and results in redness, swelling, heat and pain. There is a fine balance between the beneficial effects of inflammation cascades and their potential for long-term tissue destruction. If they are not controlled or resolved, inflammation cascades can lead to the development of diseases such as chronic asthma, rheumatoid arthritis, cardiovascular disease and certain types of cancer.
During many inflammation-associated pathologies, nitric oxide (NO) production increases significantly. NO has a variety of biological activities including vasodilation, neurotransmission and cytotoxicity. The free radical nature of NO and the high reactivity with superoxide renders NO a potent pro-oxidant molecule which is able to induce oxidative damage and potentially harmful toward cellular targets. Owing to the importance of the control of NO production for human health, the attention of many research groups has been focused on the mechanisms responsible of the reversal of NO effects from protective to deleterious.
Ellagic acid (EA) is a naturally occurring plant polyphenol. EA has been shown to possess numerous anticarcinogenic and antimutagenic properties towards a variety of different carcinogens. In addition, EA also has potent antioxidant properties in vivo and in vitro. Expanding upon from current understanding, we examined the effects of EA on lipopolysaccharide (LPS)-induced of nitric oxide (NO), tumor necrosis factor-α(TNF-α)and interleukin-6(IL-6)productions using macrophage RAW264.7 cells and revealed the possible mechanisms underlying.
The results indicate that EA inhibited LPS-induced NO production in a dose-dependent manner. Furthermore, EA also decreased the levels of IL-6 and TNF-α. In the presence of EA, inducible nitric oxide synthase(iNOS)protein and mRNA levels were declined significantly as compared with LPS induction alone. Both IκB and NF-κB expressions were also increased with the supplementation of EA. These findings implicate that EA can be applied in the regulation of macrophage-mediated inflammatory diseases.
中文摘要............................................................5
英文摘要............................................................6
第一章 前言..........................................................8
第二章 文獻回顧......................................................10
第一節 發炎反應與巨噬細胞...........................................10
第二節 脂多醣......................................................13
第三節 一氧化氮與誘導型一氧化氮合成酶.................................15
第四節 腫瘤壞死因子-α...............................................17
第五節 介白素-6....................................................17
第六節 Nuclear factor-kappa B(NF-κB).............................18
第七節 鞣花酸......................................................20
第三章 研究目的......................................................23
第四章 實驗材料與方法.................................................24
第一節 實驗材料....................................................24
第二節 實驗步驟....................................................27
第五章 結果與討論....................................................36
第一節 結果.........................................................36
第二節 討論.........................................................41
第六章 結論..........................................................44
參考文獻............................................................45

圖目錄
圖一、發炎組織中活化的巨噬細胞釋放數種生長因子,對骨髓產生單核球、巨噬細胞及顆
粒性白血球的回饋控制.............................................12
圖二、脂多醣的結構....................................................14
圖三、NF-κB 活化路徑.................................................19
圖四、鞣花酸結構式....................................................20
圖五、MTT 經由琥珀酸去氫酶作用後,還原成Formazan........................29
圖六、NO2-與Sulfanilamide及Naphthylethylenediamine dihydrochloride
反應生成粉紅色偶氮化合物.........................................30
圖七、MTT 分析中Formazan 吸光值與細胞數目之相關性.......................53
圖八、不同劑量的DMSO 對RAW264.7 細胞之細胞毒性分析......................54
圖九、不同濃度的鞣花酸對RAW264.7 細胞之細胞毒性分析......................55
圖十、不同濃度的鞣花酸對RAW264.7 細胞之一氧化氮產生量的影響...............56
圖十一、不同濃度的脂多醣對RAW264.7 細胞之細胞毒性分析....................57
圖十二、不同濃度的脂多醣對RAW264.7 細胞之一氧化氮產生量的影響.............58
圖十三、不同濃度鞣花酸對脂多醣所誘導之一氧化氮產生量的影響.................59
圖十四、鞣花酸抑制脂多醣誘導之一氧化氮的生成.............................60
圖十五、鞣花酸抑制脂多醣誘導之IL-6 的生成...............................61
圖十六、鞣花酸抑制脂多醣誘導之TNF-α 的生成..............................62
圖十七、鞣花酸抑制脂多醣誘導之TNF-α、IL-6 和iNOS 之mRNA表現量............63
圖十八、鞣花酸對RAW264.7 細胞於30 分鐘時對IκB、NF-κB 和 iNOS 之蛋白質表現之
影響.........................................................64
圖十九、鞣花酸對 RAW264.7 細胞之24 小時, IκB、NF-κB 和 iNOS 之蛋白質表現之
影響.........................................................66

表目錄
表一、MTT 分析中Formazan 吸光值與細胞數目之相關性.......................68
表二、不同劑量的DMSO 對RAW264.7 細胞之細胞毒性分析......................68
表三、不同濃度的鞣花酸對RAW264.7 細胞之細胞毒性分析......................69
表四、不同濃度的鞣花酸對RAW264.7 細胞之一氧化氮產生量的影響...............69
表五、不同濃度的脂多醣對RAW264.7 細胞之細胞毒性分析......................70
表六、不同濃度的脂多醣對RAW264.7 細胞之一氧化氮產生量的影響...............70
表七、不同濃度鞣花酸對脂多醣所誘導之一氧化氮產生量的影響..................71
表八、鞣花酸抑制脂多醣誘導之一氧化氮的生成..............................71
表九、鞣花酸抑制脂多醣誘導之IL-6 的生成................................72
表十、鞣花酸抑制脂多醣誘導之TNF-α 的生成...............................72
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