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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 莊雅惠(Ya-Hui Chuang) | |
dc.contributor.author | Bi-Jhen Syu | en |
dc.contributor.author | 許碧珍 | zh_TW |
dc.date.accessioned | 2021-06-16T02:33:26Z | - |
dc.date.available | 2020-09-25 | |
dc.date.copyright | 2015-09-25 | |
dc.date.issued | 2015 | |
dc.date.submitted | 2015-07-28 | |
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/53919 | - |
dc.description.abstract | 原發性膽道硬化症(Primary biliary cirrhosis;PBC)為膽道受破壞導致膽汁鬱積甚至肝硬化的慢性肝臟自體免疫疾病。先前研究指出PBC病人血清中IFN-γ表現比起健康的人顯著增加,肝臟內亦可測得PDC-E2專一性輔助性T細胞浸潤。在不同PBC動物模式下也證實Th1細胞及IFN-γ參與在PBC病程,但如何作用於PBC尚未清楚。本研究探討Th1細胞和IFN-γ在PBC所扮演的角色及其作用機轉。我們使用先前實驗室建立的2-OA-OVA誘發PBC之小鼠模式,分析Th1細胞和IFN-γ在此PBC小鼠模式之表現,發現PBC小鼠肝臟中IFN-γ基因表現量以及PBC小鼠第四週血清IFN-γ的表現量均明顯升高。以細胞內染色方法分析PBC小鼠肝臟單核細胞分泌細胞激素的表現亦發現分泌IFN-γ的CD4 T細胞(Type I helper T cell;Th1)百分比顯著增加。為了釐清IFN-γ及Th1如何調控PBC免疫反應,我們將攜帶IFN-γ基因的Adeno-asoociated virus(AAV)給予小鼠同時誘發小鼠產生PBC,再測量其PBC疾病程度。結果顯示大量IFN-γ會增加PBC小鼠肝臟內免疫細胞浸潤,包括CD4+ T、CD8+ T、B、NK與NKT細胞以及增加屬於Th1且與組織發炎相關的TNF-α基因的表現。同時發現IFN-γ也會使PBC小鼠肝臟抗原呈獻細胞數增加且其MHC class II表現增強,而且Th1細胞數量也顯著增加。由以上實驗結果暗示Th1參與在PBC病程,且IFN-γ會增加抗原呈獻細胞並加強其MHC class II之表現以及促進更多CD4 T細胞分化成Th1細胞並製造更多IFN-γ以引起更多免疫反應,例如活化CD8+ T、B、NK與NKT細胞,最後使PBC肝臟門脈發炎情況變得更嚴重。 | zh_TW |
dc.description.abstract | Primary biliary cirrhosis (PBC) is a slowly progressive liver-specific autoimmune disease caused by the destruction of small intra-hepatic bile ducts, which leads to cholestasis or even cirrhosis. In patients with PBC, the serum levels of IFN-γ are much higher than healthy people and PDC-E2 specific autoreactive helper T cells could be found in their livers. Previous studies showed that IFN-γ and Th1 cells regulate the immune responses in PBC, but the mechanism remains unclear. By using our previous established 2-OA-OVA immunized mouse model of PBC, we found that liver and serum levels of IFN-γ expression were significantly increased in PBC mice while IL-4 and IL-17 were undetectable. In addition, the percentage of IFN-γ-producing CD4 T cells (Th1 cells) in the liver of PBC mice was significantly increased. To study the effects of Th1 cells and IFN-γ on PBC, we intravenously injected IFN-γ expressing adeno-associated virus (AAV-IFN-γ) to 2-OA-OVA immunized PBC mice and measured the features of PBC in these mice. Results showed that administration of IFN-γ significantly increased liver mononuclear cells including CD4+ T, CD8+ T, B, NK and NKT cells and TNF-α expression. Increased numbers of antigen-presenting cells (APCs) with a higher MHC class II expression were also found in AAV-IFN-γ injected PBC mice. Moreover, the frequency and number of IFN-γ producing Th1 cells were also increased. These results suggest that IFN-γ producing Th1 cells play a role in the pathogenesis of PBC. IFN-γ promotes a higher antigen-presenting capability of APCs and Th1 cell differentiation. Th1 cells produce more IFN-γ to augment immune responses such as activating CD8+ T, B, NK and NKT cells, which then exacerbates portal inflammation of PBC. | en |
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dc.description.tableofcontents | 致謝 i 中文摘要 ii Abstract iii 縮寫對照表 v 目錄 vii 圖目錄 x 第一章 研究背景 1 1.1 原發性膽道硬化症(Primary biliary cirrhosis;PBC) 1 1.1.1 PBC臨床診斷標準 1 1.1.2血清學診斷之PBC自體抗體與自體抗原 1 1.1.3 病理學診斷之PBC病理特徵與疾病分期 2 1.1.4 PBC致病因子 3 1.1.5 PBC治療 4 1.2 自體免疫疾病與Th1及Th17之關係 4 1.3 Interferon-γ(IFN-γ) 5 1.3.1 IFN-γ之分泌/作用細胞以及其生理功能 5 1.3.2 IFN-γ與自體免疫疾病 7 1.4 PBC之免疫反應 8 1.4.1 PBC之先天性免疫反應 8 1.4.2 PBC之後天性免疫反應(臨床證據文獻) 9 1.4.3 PBC之後天性免疫反應(動物模式文獻) 9 1.5 Adeno-associated virus(AAV) 11 1.5.1 AAV-DJ Helper Free Bicistronic Expression System(GFP) 12 1.6 研究目的 12 第二章 實驗材料與方法 13 2.1 實驗用小鼠 13 2.2 2-OA-OVA induced PBC小鼠模式 13 2.3 血清樣品之收集 13 2.4 血清中抗粒線體抗體效價測定 13 2.5 血清細胞激素測定 14 2.6 小鼠肝臟灌流與肝臟單核細胞分離 15 2.7 小鼠淋巴結之細胞分離 15 2.8 分離肝臟單核細胞與淋巴結單核細胞中之CD4+ T細胞 16 2.9 以流式細胞儀分析細胞表面及細胞內抗原 16 2.10 小鼠肝臟病理組織切片之製作 17 2.11 RNA反轉錄成cDNA 17 2.12 以即時定量反轉錄聚合酶連鎖反應偵測特定基因之mRNA表現 18 2.13 AAV製備 18 2.14 AAV之純化 19 2.15 AAV之濃縮 19 2.16 AAV之定量 19 2.17 以AAV施打PBC小鼠 20 2.18 繪圖與統計分析 20 第三章 實驗結果 21 3.1 分析以2-OA-OVA誘發小鼠產生原發性膽道硬化症之細胞激素表現 21 3.1.1分析PBC小鼠血清細胞激素表現量以及肝臟組織之細胞激素基因表現 21 3.1.2以細胞內染色方式偵測PBC小鼠肝臟單核細胞細胞激素之表現 21 3.1.3分析肝臟旁淋巴結之淋巴細胞生成細胞激素的情形 22 3.2探討IFN-γ對於致敏十週之PBC小鼠的免疫反應及影響 22 3.2.1給予AAV-IFN-γ之PBC小鼠於實驗期間,血清中皆可測得IFN-γ的表現 22 3.2.2給予AAV-IFN-γ之十週PBC小鼠的門脈浸潤無明顯差異,不過肝臟TNF-α mRNA的表現有顯著增加 23 3.2.3給予AAV-IFN-γ之十週PBC小鼠肝臟中組織纖維化相關蛋白的基因表現與對照組無差異 23 3.2.4給予AAV-IFN-γ之十週PBC小鼠並無改變AMA,但肝臟淋巴細胞數顯著上升 24 3.2.5給予AAV-IFN-γ之十週PBC小鼠浸潤於肝臟的NK與NKT細胞數顯著上升 24 3.2.6給予AAV-IFN-γ之十週PBC小鼠肝臟中活化的NK細胞百分比顯著上升 24 3.2.7給予AAV-IFN-γ之十週PBC小鼠肝臟記憶性T細胞數產生變化 24 3.3探討IFN-γ對於致敏五週之PBC小鼠的免疫反應 25 3.3.1給予AAV-IFN-γ之五週PBC小鼠肝臟單核細胞與其分群數量皆顯著上升 25 3.3.2給予AAV-IFN-γ之五週PBC小鼠肝臟中活化的NK以及CD8 T細胞百分比顯著上升 25 3.3.3給予AAV-IFN-γ之五週PBC小鼠肝臟記憶性T細胞數目顯著增加 26 3.4探討IFN-γ對PBC作用機轉 26 3.4.1當小鼠大量表現IFN-γ會使肝臟抗原呈獻細胞數目顯著增加且其表現之MHC class II也增加 26 3.4.2 當小鼠大量表現 IFN-γ時,會使肝臟 CD4 T 細胞與 CD8 T 細胞分泌 IFN-γ之百分比顯著上升 26 第四章 結論與討論 28 圖 33 第五章 參考文獻 56 附錄 63 | |
dc.language.iso | zh-TW | |
dc.title | 以小鼠模式探討第一型輔助型T細胞和IFN-γ在原發性膽道硬化症的角色 | zh_TW |
dc.title | The Roles of Type I Helper T cells and IFN-γ in a Mouse Model of Primary Biliary Cirrhosis | en |
dc.type | Thesis | |
dc.date.schoolyear | 103-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 繆希椿,胡忠怡,潘建雄 | |
dc.subject.keyword | 肝臟自體免疫疾病,原發性膽道硬化症,IFN-γ,Th1細胞,CD4 T細胞, | zh_TW |
dc.subject.keyword | Liver autoimmune disease,Primary biliary cirrhosis,IFN-γ,Th1 cells,CD4 T cells, | en |
dc.relation.page | 65 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2015-07-28 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 醫學檢驗暨生物技術學研究所 | zh_TW |
顯示於系所單位: | 醫學檢驗暨生物技術學系 |
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