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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 陳俊任(Chun-Jen Chen) | |
| dc.contributor.author | Ping-Hsiang Hunag | en |
| dc.contributor.author | 黃品翔 | zh_TW |
| dc.date.accessioned | 2023-03-19T22:38:37Z | - |
| dc.date.copyright | 2022-08-24 | |
| dc.date.issued | 2022 | |
| dc.date.submitted | 2022-08-18 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/85021 | - |
| dc.description.abstract | 甲型烯醇化酶為多功能的蛋白,其功能不同取決於在細胞中之位置。單核球及巨噬細胞發現,發炎過程中甲型烯醇化酶易位至細胞表面並當做纖溶酶原受體,促進細胞外基質降解和細胞遷移。甲型烯醇化酶也會在發炎過程釋放到細胞外,透過類鐸受體4刺激巨噬細胞活化。在這項研究中我們探討嗜中性球於發炎過程其甲型烯醇化酶是否會易位至細胞表面外,也探討甲型烯醇化酶於發炎反應中對於巨噬細胞活化扮演之角色。以脂多醣體內刺激小鼠嗜中性球顯示甲型烯醇化酶於細胞表面表現上升,並增強離體纖溶酶原活化,這個結果被抗甲型烯醇化酶單株克隆抗體阻斷。當小鼠肺泡巨噬細胞MH-S細胞和人類PMA-THP-1巨噬細胞被脂多醣刺激時,也觀察到同樣的現象。此外,抗甲型烯醇化酶單株克隆還可以分別抑制脂多醣和重組甲型烯醇化酶刺激巨噬細胞之活化。此外,脂多醣刺激巨噬細胞活化被傳明酸所阻斷,傳明酸與纖溶酶原結合併阻止其活化。這些數據表明,在脂多醣刺激巨噬細胞後,甲型烯醇化酶介導之纖溶酶活化可以增強發炎反應。總之,結果顯示嗜中性球與巨噬細胞受到脂多醣刺激,其表面甲型烯醇化酶表現增加,促使這些白血球的遷移與活化。因此,通過抗甲型烯醇化酶單株克隆抗體來阻斷這些現象可能被用於治療敗血症等發炎相關之疾病。 | zh_TW |
| dc.description.abstract | The glycolytic enzyme alpha-enolase (enolase-1, ENO1) is a multifunctional protein with various activities depending on its cellular and extracellular localization. In monocytes/macrophages, ENO1 has been shown to translocate to the cell surface and serve as a plasminogen receptor (PLGR), which facilitates extracellular matrix degradation and cell migration during inflammation. ENO1 can also be released to the extracellular space to stimulate macrophages activation via TLR4 during inflammation. In this study, we investigated whether cell surface translocation of ENO1 also occurs in neutrophils, and we also examined whether ENO1 may modulate the activation of macrophages during inflammation. Neutrophils isolated from LPS-challenged mice showed elevated surface expression of ENO1 and enhanced PLG activation ex vivo, which was effectively blocked by anti-ENO1 mAbs. The same phenomenon was also observed when the murine alveolar macrophage MH-S cells and human PMA-THP-1 macrophages cells were stimulated by LPS. In addition, anti-ENO1 mAbs could also inhibit the activation of macrophages stimulated by LPS and recombinant ENO1, respectively. Furthermore, LPS-triggered macrophage activation was blocked by tranexamic acid (TXA), which binds PLG and prevents its activation. These data indicated that upon LPS stimulation of macrophages, ENO1-mediated activation of plasmin could enhance the inflammatory response. Overall, our results indicate that during LPS-stimulated inflammation, upregulated ENO1 expression on the cell surface of neutrophils and macrophages may facilitate the migration and activation of these leukocytes, hence, blocking these events by inhibited by anti-ENO1 mAbs can potentially be used to suppress inflammation in diseases such as sepsis. | en |
| dc.description.provenance | Made available in DSpace on 2023-03-19T22:38:37Z (GMT). No. of bitstreams: 1 U0001-1808202211155500.pdf: 4207479 bytes, checksum: 1f19cf52213e40172908e3707552a74d (MD5) Previous issue date: 2022 | en |
| dc.description.tableofcontents | 口試委員鑑定書 I 致謝 II 中文摘要 III Abstract IV 縮寫 V 目錄 VII 圖表目錄 XI 第一章 緒論 1 1. 發炎反應 1 1.1. 急性發炎 2 1.2. 慢性發炎 2 2. 先天性免疫細胞 3 2.1. 巨噬細胞 3 2.2. 單核球 5 2.3. 嗜中性球 6 3. 促發炎因子 7 3.1. 腫瘤壞死因子 (Tumor necrosis factor, TNF-α) 7 3.2. 細胞介白素第六因子 (Interleukin-6, IL-6) 7 4. 烯醇化酶 (enolase) 8 5. α-烯醇化酶與發炎反應 9 6. 纖溶酶 (Plasmin) 10 7. 纖溶酶與發炎反應 11 研究動機 13 第二章 材料與方法 14 1. 實驗動物 14 2. 細胞株培養與處理相關試劑 14 2.1. 細胞培養基 14 2.2. 細胞株培養 14 2.3. 細胞及組織用相關試劑 15 3. Anti-ENO1抗體 15 4. 細胞表面ENO1之分析 16 4.1. 小鼠肺部巨噬細胞株 (MH-S) 16 4.2. PMA分化之人類單核球細胞株 (PMA-THP-1) 16 4.3. 小鼠嗜中性球 17 4.4. 人類嗜中性球 18 5. 小鼠嗜中性球製備與純化 19 5.1. 血液細胞處理及純化 19 5.2. 骨髓細胞處理及純化 19 6. 纖溶酶活性之測定 20 7. 烯純化酶重組蛋白製備 21 7.1. 將pTrcHis-ENO1質體送入大腸桿菌 21 7.2. 重組蛋白製備 21 8. 細胞激素測定 22 8.1. MH-S細胞株刺激 22 8.2. PMA-THP-1細胞株刺激 22 8.3. 細胞激素測定 23 9. 即時定量反轉錄聚合酶連鎖反應 (qRT-PCR) 24 9.1. RNA萃取 24 9.2. cDNA合成 24 9.3. 即時定量反轉錄聚合酶連鎖反應 (Quantitative reverse transcription Polymerase Chain Reaction, qRT-PCR) 25 9.4. 基因表現分析 (2-ΔΔCT method) 25 10. 西方墨點法 25 10.1. 蛋白質收取與製備 25 10.2. 十二烷基硫酸鈉聚丙烯酰胺凝膠電泳 (SDS-PAGE) 26 10.3. 西方墨點法 (Western blot) 26 10.4. 蛋白質表現分析 (ImageJ) 27 11. 統計與繪圖軟體分析 27 第三章 研究結果 28 1. 發炎反應中小鼠嗜中性球表面ENO1表現上升並介導PLA活化 28 2. 發炎反應中人類嗜中性球表面ENO1表現 29 3. LPS刺激巨噬細胞表面ENO1之上升 29 4. Anti-ENO1 mAbs於發炎反應中對於巨噬細胞活化之影響 30 4.1. Anti-ENO1 mAbs抑制LPS刺激巨噬細胞之活化 30 4.2. Anti-ENO1 mAbs抑制ENO1刺激巨噬細胞之活化 30 5. 發炎反應中巨噬細胞表面ENO1表現上升並介導PLA活化 31 6. PLA於發炎反應中對於巨噬細胞活化之影響 31 第四章 討論 33 第五章 圖表 39 參考資料 58 | |
| dc.language.iso | zh-TW | |
| dc.subject | 類鐸受體4 | zh_TW |
| dc.subject | 纖溶酶原受體 | zh_TW |
| dc.subject | 甲型烯醇化酶 | zh_TW |
| dc.subject | 嗜中性球 | zh_TW |
| dc.subject | 纖溶酶 | zh_TW |
| dc.subject | 巨噬細胞 | zh_TW |
| dc.subject | TLR4 | en |
| dc.subject | Enolae-1 | en |
| dc.subject | Plasminogen receptor | en |
| dc.subject | Plasmin | en |
| dc.subject | Macrophage | en |
| dc.subject | Neutrophil | en |
| dc.title | 甲型烯醇化酶於發炎反應中調節巨噬細胞與嗜中性球活化之角色 | zh_TW |
| dc.title | The role of alpha-enolase in regulating the activation of macrophages and neutrophils during inflammation | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 110-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 江皓森(Hao-Sen Chiang),徐立中(Li-Chung Hsu) | |
| dc.subject.keyword | 巨噬細胞,嗜中性球,甲型烯醇化酶,類鐸受體4,纖溶酶原受體,纖溶酶, | zh_TW |
| dc.subject.keyword | Enolae-1,Plasminogen receptor,Plasmin,Macrophage,Neutrophil,TLR4, | en |
| dc.relation.page | 75 | |
| dc.identifier.doi | 10.6342/NTU202202541 | |
| dc.rights.note | 同意授權(限校園內公開) | |
| dc.date.accepted | 2022-08-18 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 生化科技學系 | zh_TW |
| dc.date.embargo-lift | 2024-08-18 | - |
| 顯示於系所單位: | 生化科技學系 | |
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