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系統識別號 U0026-0812200911173664
論文名稱(中文) 周邊血液單核球細胞在腸病毒71型感染中之探討
論文名稱(英文) The study of enterovirus 71 (EV71) infection on peripheral mononuclear cells (PBMC)
校院名稱 成功大學
系所名稱(中) 微生物及免疫學研究所
系所名稱(英) Department of Microbiology & Immunology
學年度 92
學期 2
出版年 93
研究生(中文) 蘇玲瑤
研究生(英文) Ling-Yao Su
學號 s4691410
學位類別 碩士
語文別 中文
論文頁數 54頁
口試委員 指導教授-黎煥耀
口試委員-蘇益仁
口試委員-余俊強
指導教授-劉清泉
中文關鍵字 腸病毒71型  單核球 
英文關鍵字 enterovirus 71  PBMC 
學科別分類
中文摘要   腸病毒71 (Enterovirus 71, EV71)屬於微小RNA病毒科之腸病毒屬,在臨床上可以引起很多症狀,包括有嚴重的腦幹腦炎併發肺水腫。在1998年的時候,台灣爆發了腸病毒71的大流行。根據我們實驗室之前的結果發現,在肺水腫病人有較低的CD4+、CD8+及NK細胞數,顯示在嚴重的腸病毒感染之下,周邊血液單核球細胞可能受到各方面不同的影響而造成這樣的結果,可能是因病毒量(viral load)本身,或是由於宿主體內環境改變而造成細胞凋亡或重新分佈至其他器官。為研究腸病毒71感染後細胞免疫系統活化情況,於是我們建立了一個in vitro的感染模式,以不同的細胞株或人體的周邊血液單核球細胞,探討這些細胞在受到感染後,細胞被活化的情形。將採集的周邊血液單核球細胞或是臍帶血液單核球細胞利用梯度離心分離出來後,再以腸病毒71感染。在偵測方法方面,則是利用抗腸病毒71抗體偵測病毒抗原,或用溶菌斑試驗定量病毒效價。我們的結果顯示,周邊血液單核球細胞,包括T細胞、B細胞及巨噬細胞都可以被腸病毒71感染,而T細胞產生的病毒量又比B細胞及巨噬細胞多。在細胞激素的表現上,可能是因為感染使得細胞數目減少,因此測得的細胞激素在感染那一組反而比控制組來得少。除此之外,在感染的早期,細胞會表現FasL,同時伴隨著細胞凋亡的增加,顯示感染之後,細胞可能走向細胞凋亡的途徑。另外,我們也比較了不同階段病人其血清中細胞激素及趨化素的表現,只有在肺水腫病人其血清中細胞激素及趨化素明顯的比其他組來得高。我們證實了周邊血液淋巴球細胞都能被腸病毒71感染,而且細胞會有細胞凋亡的情況發生,重症病人體內的發炎反應也比輕症來的高,藉由以上結果,或許能對腸病毒71型感染的致病機轉有所了解。
英文摘要   Enterovirus 71 (EV71), the member of the Picornaviridae family, can cause serious brainstem encephalitis (BE) with pulmonary edema (PE). In Taiwan, a major outbreak of EV71 infection occurred in 1998. Our previous studies found that lymphopenia was significantly more frequent among PE patients who have lower circulating CD4+ T cells, CD8+ T cells, and natural killer (NK) cells. To elucidate the role of mononuclear cells in the pathogenesis of EV71 infection, cord blood mononuclear cells (CBMC) were isolated by Ficoll-Metriozate density gradient centrifugation, and then infected by EV71. The infected cells were analyzed by flow cytometry with monoclonal antibodies specific for EV71. The virus titer was quantitated by plaque assay. Supernatant were collected and the cytokine levels were detected by Cytometric Bead Assay (CBA). We found that all of the mononuclear cells including T cells, B cells, and monocytes can be infected by EV71. Among those infected cells, T cells generate high amounts of EV71 virions. The cytokine levels were decreased after infection as well as the total cell numbers, it may due to the decreased of cell counts. The expression of CD95L was increased in early infection, and the Annexin V-positive cells could be detected at the same time point as early as 12 hours post infection. In previous study, we detected the cytokines levels were higher in stage 3B patients. These results were similar with the chemokine profiles in stage 3B patients. Here, we demonstrate that mononuclear cells can be infected by EV71. And stage 3B patients showed more cytokine and chemokine levels. These finding maybe shed light on the understanding of the pathogenesis of EV71 infection.
論文目次 中文摘要
英文摘要
致謝
目錄
圖目錄
表目錄
壹、緒論
貳、材料與方法
一、材料
二、方法
三、統計方法
參、結果
一、腸病毒71型感染之病人臨床變化
A. 血球的變化
B. 細胞激素(cytokine)及趨化素(chemokine)的變化
C. 細胞表面抗原之分析
二、腸病毒71 in vitro感染模式的建立
A. 腸病毒71可以感染單核球細胞
B. 腸病毒71感染後影響細胞之功能
C. 細胞表面CD95、CD95L的變化
肆、討論
伍、結論
陸、參考文獻


附錄
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