篩選體內表現基因載體的建立與心內膜炎免疫反應之特徵

Construction of a plasmid for screening in vivo gene expression and characteristic of immune response in infective endocarditis

吳淑惠(Shu-Hui Wu)

指導教授 ： 賈景山

國立臺灣大學/醫學院/口腔生物科學研究所/碩士(2005年)

https://doi.org/10.6342/NTU.2005.01882

摘要

轉糖鏈球菌(Streptococcus mutans)屬於草綠色鏈球菌，存在於人體口腔中並可引起齲齒。當轉糖鏈球菌進入血流中形成菌血症時，便有機會導致心臟瓣膜缺損的病人產生感染性心內膜炎（infective endocarditis, IE）。本實驗的第一部分中，為了找尋轉糖鏈球菌的新的毒性因子，我們建立一個適用於in vivo expression technology的表現載體，利用pDLDlacZ帶有兩個不具啟動子的報導基因：km-lac，而去篩選體內表現的基因。
本實驗第二部分是利用先前已建立好之實驗性心內膜炎之動物模型，探討S. mutans所引起心內膜炎的免疫反應。同時收集臨床人類心內膜炎檢體，觀察人類心內膜炎的慢性發炎的免疫反應特徵，並與實驗性心內膜炎相比較。實驗發現，造成感染的轉糖鏈球菌會存在於vegetation當中、纖維化的區域形成一小菌落，而避免受到白血球的吞噬作用。利用免疫染色發現，感染24小時後的巨噬細胞與纖維母細胞皆有NF-κB的活化。而在人類的組織切片中則可發現巨噬細胞、內皮細胞與纖維母細胞皆有細胞激素：IL-6、TNF-α與IL-1β的大量表現，並且伴隨著NF-κB的活化。因此我們推測纖維母細胞中的NF-κB活化在導致感染性心內膜炎持續性發炎中扮演一個重要的角色。

關鍵字

轉糖鏈球菌

並列摘要

Streptococcus mutans, a member of the viridans streptococci, can cause dental caries in the oral cavity and infective endocarditis (IE) when entering into blood circulation. Major processes in the pathogenesis of IE are the formation of vegetations at the damaged surface on valves and inflammation of the endocardium. In the first part of this study, we developed a novel plasmid, pDLDlacZ for in vivo expression technology (IVET) in order to identify novel virulent factors of S. mutans. A Streptococcus-E. coli shuttle vector which carries dual (Km-lac) promoterless reporter genes has been constructed.
In the secondary part of the study, we used an experimental endocarditis rat model established previously in our laboratory to investigate the immune response induced by S. mutans. Human biopsies of infective endocarditis at chronic inflammatory stage was also analyzed and compared with the immune response identified on experimental endocarditis. Nests of S. mutans, with positive staining for secretory proteins, was walled-off in the fibrosis region of VG to avoid phagocytosis. Immunohistochemistry staining of the tissue sections showed that NF-κB was activated in both macrophage and fibroblast 24 hours after infection. In human biopsies, IL-6, TNF-α, and IL-1β secretion was found in macrophages, fibroblasts, and endothelium in addition to activated NF-κB. Therefore, NF-κB activation in the fibroblasts in endocardium might play an impartment role in the persistence of inflammation in IE

並列關鍵字

S. mutans ； IVET ； NF-kappa B

參考文獻

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