利用蛋白質分析法比較氣喘病人的CD4+T淋巴球在穩定控制期與不穩定期間的差異; Proteomic Analysis of CD4+ T-lymphocytes in Patients with Asthma between Controlled and Uncontrolled Level

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Title:	利用蛋白質分析法比較氣喘病人的CD4+T淋巴球在穩定控制期與不穩定期間的差異;Proteomic Analysis of CD4+ T-lymphocytes in Patients with Asthma between Controlled and Uncontrolled Level
Authors:	柯仰謦;Yang-Ching Ko
Contributors:	中國醫藥大學：醫學研究所碩士班
Keywords:	氣喘;T-淋;巴球;蛋白質分析法;asthma;T-lymphocyte;proteomic analysis
Date:	2009-06-11
Issue Date:	2009-08-13 14:50:34 (UTC+8)
Abstract:	背景：氣喘屬慢性發炎疾病，多數研究焦點放在引發發炎的機制。在氣喘病因學中，T-淋巴球衍生的細胞激素被廣為討論。在後基因領域中，蛋白質分析技術快速發展且成為基因領域裡重要的輔助工具。我們利用蛋白質分析法中兩個重要的技術：二維式膠片電泳分析法與蛋白質身份鑑定質譜分析，來比較氣喘病人從不穩定期到穩定控制期時 T-淋巴球蛋白質間的差異。材料與方法：對連續 6 位到院表現為不穩定氣喘患者抽血並萃取到 CD4+ T-淋巴球，患者在三個月內經藥物控制為穩定性氣喘，再次抽血取得 CD4+ T-淋巴球。接著利用二維式膠片電泳分析法比較不穩定期氣喘與穩定控制期氣喘時 T-淋巴球蛋白質間的差異。對於在膠片上有明顯差異的小點，我們再取出並利用蛋白質身份鑑定質譜儀分析與比對資料庫來確定為何種蛋白質。結果：氣喘患者體內的 CD4+ T-淋巴球有超過一百個以上的小點顯現在二維式膠片上。這些小點經由考馬斯亮藍染色液染色後，兩組患者的小點分佈呈現類似情況，但是其中有 13 個小點卻出現有意義的差異表現。若以從不穩定控制組到穩定控制組來看，其中有 6 種蛋白質減少而 7 種蛋白質增加。結論：經由蛋白質分析法的應用，確實讓我們發現在氣喘患者體內 CD4+ T-淋巴球在穩定控制期與不穩定控制期間的差異，我們需要更多數量與不同範疇的研究來確認何者為氣喘發作的標記或作為治療不穩定控制的方法。 Background As a chronic inflammatory disease, much of the research related to asthma has focused on proinflammatory mechanisms. T-lymphocyte (T-LC)-derived cytokines have been implicated in asthmatic pathogenesis. Proteomic technology has rapidly developed in the postgenomic era, and it is now widely accepted as a complementary technology to genetic profiling. We investigated the changes of proteins in T-LC of asthmatic patients from the uncontrolled to controlled level by using standard proteome technology: two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), liquid chromatography/mass spectrometry (LC/MS), and a database search. Methods The proteins of CD4+ T-LC were isolated from the whole blood of six asthmatic patients with uncontrolled to controlled levels over three months. 2D-PAGE was performed and coomassie blue stained protein spots were comparatively analyzed between the uncontrolled and controlled groups using an image analyzer. Some differentially expressed spots were identified by LC-MS/MS and database search. Results More than 100 spots were identified in the 2D-PAGE gels from the CD4+ T-LC of the asthmatic patients. The general distribution pattern of the spots in the Coomassie blue-stained gel was similar in both groups, and 13 proteins showed different intensity, suggesting differential expression. Six protein spots in the CD4+ T-LC of the uncontrolled asthmatic patients were increased and 7 spots were decreased compared to those of the controlled subjects. Conclusions The proteomic examination of the CD4+ T-LC revealed some differentially expressed proteins in the uncontrolled and controlled asthmatic patients. The possibility of using the differentially expressed proteins as important biomarkers and therapeutic targets in uncontrolled asthmatic patients warrants further study.
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