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研究生: 張藝馨
Chang, Yi-Hsin
論文名稱: 白藜蘆醇衍生物Resveratroloside對脂多醣誘導的RAW264.7細胞之抗發炎研究
Study of Anti-inflammatory Effects of Resveratrol Derivative Resveratroloside on Lipopolysaccharide-induced RAW264.7 Cells
指導教授: 張誌益
Chang, Chi-I
吳裕仁
Wu, Yu-Jen
學位類別: 碩士
Master
系所名稱: 農學院 - 生物科技系
Department of Biological Science and Technology
論文出版年: 2022
畢業學年度: 110
語文別: 中文
論文頁數: 37
中文關鍵詞: 抗發炎白藜蘆醇衍生物脂多醣巨噬細胞西方墨點法免疫螢光
外文關鍵詞: anti-inflammatory, resveratrol derivative, LPS, macrophage, Western blot, Immunofluorescence
研究方法: 一氧化氮生成試驗 、 細胞存活率試驗 、 細胞免疫螢光染色 、 西方墨點法
DOI URL: http://doi.org/10.6346/NPUST202200345
相關次數: 點閱:40下載:3
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  • 前人研究顯示白藜蘆醇的生物活性可能具有治療作用,像是抗氧化、抗發炎、預防心血管疾病、抗癌,這些天然活性物質具有開發成藥物的潛力。本研究探討白藜蘆醇的衍生物Resveratroloside於脂多醣(Lipopolysaccharide, LPS)所誘導的巨噬細胞RAW264.7中的抗發炎能力及其機制。經由MTT細胞存活率測試、一氧化氮(Nitric Oxide, NO)生成試驗、西方墨點法(Western blot)、免疫螢光染色(Immunofluorescence, IF)的實驗結果發現Resveratroloside對細胞毒性低且能抑制發炎反應物NO生成,並且抑制發炎相關蛋白質iNOS、COX-2之表現量。研究發現Resveratroloside透過抑制MAPK途徑及NF-κB磷酸化生成及轉位來阻斷發炎反應的進行。這些研究結果說明Resveratroloside的低細胞毒性及抗發炎能力具有作為抗炎藥物開發的潛能。

    Previous studies have shown resveratrol possibly has biological activities for the treatment, such as antioxidant, anti-inflammatory, anti-cancer, cardiovascular disease prevention, and anti-cancer, which is potential in the drug. The purpose of this study was to investigate the anti-inflammatory activity and mechanism of resveratrol derivative Resveratroloside in Lipopolysaccharide (LPS) treated macrophage RAW264.7 cells. The results of MTT cell viability test, Nitric Oxide (NO) determination, Western blot, and Immunofluorescence (IF) showed that Resveratroloside has lower cytotoxicity and it can inhibit the production of nitric oxide, and also inhibit pro-inflammatory enzymes expression of nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), that Resveratroloside reduces the inflammatory responses through inhibiting the MAPK signaling pathway and the phosphorylation and translocation of NF-κB. These data suggested that Resveratroloside has lower cytotoxicity and anti-inflammatory effect, which is potential for anti-inflammatory drug development.

    中文摘要 I
    Abstract II
    謝誌 III
    圖目錄 VI
    壹、 前言 1
    貳、 文獻回顧 3
    一、白藜蘆醇介紹 3
    二、LPS 脂多醣(Lipopolysaccharide) 3
    三、發炎反應(inflammatory response) 4
    四、發炎相關細胞激素 5
    (一) 一氧化氮合成酶家族(nitric oxide synthase family of enzymes) 5
    (二) 環氧合酶(Cyclooxygenase, COX) 6
    五、巨噬細胞(macrophage) 7
    六、NF-κB 途徑 7
    七、MAPK 途徑 8
    參、 材料與方法 11
    一、材料 11
    (一)實驗之化合物 11
    (二)儀器設備 11
    (三)實驗藥品及試劑 11
    (四)培養之細胞株 12
    二、方法 12
    (一)細胞培養基配置 12
    (二)細胞解凍活化 13
    (三)細胞繼代 13
    (四)細胞保存 13
    (五)一氧化氮生成試驗(Nitric Oxide determination) 14
    (六)細胞存活率試驗(MTT assay) 14
    (七)細胞免疫螢光染色 15
    (八)蛋白質萃取 15
    (九)蛋白質定量 16
    (十)聚丙烯醯胺膠體電泳(SDS-PAGE) 16
    (十一)蛋白質轉印 18
    (十二)西方墨點法(Western Blot) 18
    (十三)統計分析 19
    肆、 結果 20
    一、Resveratroloside 對 RAW264.7 巨噬細胞存活率之影響 20
    二、Resveratroloside 對 LPS 誘導的 RAW 264.7 巨噬細胞發炎之 NO
    生成影響 21
    三、Resveratrolosid 對 LPS 誘導的 iNOS、COX-2 蛋白質表現量之影
    響 22
    四、Resveratroloside 對 LPS 誘導的 NF-κB 途徑表現量之影響 23
    五、Resveratroloside 對 LPS 誘導的 MAPK 途徑表現量之影響 26
    伍、討論 28
    陸、結論 31
    柒、參考文獻 32

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