臺灣博碩士論文加值系統

中文摘要
尿調理素是孕婦尿中含量最豐富的蛋白質，然而，我們對於尿調理素的生理功能仍不清楚。多形核白血球是主要的吞噬細胞且在先天性免疫中扮演抵抗細菌感染的一個重要角色。在本論文中，我們想要研究尿調理素對人類和小鼠多形核白血球的吞噬能力和殺菌能力是否會有影響。此外，對於尿調理素活化人類多形核白血球的殺菌機轉，也加以研究。
我們用尿調理素25μg在體外刺激人類多形核白血球一個小時，發現它可以增強多形核白血球的吞噬能力，除此之外，我們也發現尿調理素可以活化人類多形核白血球，並增強它對革蘭氏陰性菌如創傷弧菌和沙門氏鼠傷寒桿菌的殺菌力，但卻無法增強對革蘭氏陽性菌如金黃色葡萄球菌和李斯特菌的殺菌能力，尿調理素並不會增強多形核白血球產生TNF-α的能力，但藉由 DHR123 的染色和流體細胞計數儀的分析，我們發現經過尿調理素的處理，可以促使多形核白血球對PMA 刺激產生更多的 respiratory burst，而且經尿調理素處理後可以增強人類的多形核白血球和 LPS 結合的能力。
除了研究人類的多形核白血球，我們也發現給予尿調理素的小鼠兩個小時後再給予細菌，可以增強對革蘭氏陰性菌如創傷弧菌和沙門氏鼠傷寒桿菌在體內的清除能力，但卻無法增強小鼠對革蘭氏陽性菌如金黃色葡萄球菌和李斯特菌在體內的清除能力，此外，我們發現給予尿調理素的小鼠，可以增加小鼠體內多形核白血球的數目，也可以增強多形核白血球的吞噬能力。從以上結果顯示，尿調理素可以增強人類和老鼠的多形核白血球的功能並幫助清除細菌，而且對於革蘭氏陰性菌的效果特別好。這些研究的結果，可以提供我們更了解尿調理素對多形核白血球的免疫刺激功能，並對開發一個新的免疫刺激劑和發展免疫療法提供一個研究方向。

英文摘要
Uromodulin (URO) is the most abundant protein in the urine of pregnant women. However, its physiological function is still unclear. Polymorphonuclear neutrophils (PMNs) are the major phagocytes playing important roles in the innate immunity against bacterial infection. In this study, the effects of URO on the functions of PMN, such as phagocytosis and bactericidal activities, were studied both in human and in the mice. In addition, the mechanism of URO enhancement of bactericidal activity of human PMN by URO was studied. The phagocytosis activities of human PMN were increased after pretreatment with URO (25μg) for 1h in vitro. Furthermore, the bacterial clearance rate against the gram-negative bacteria such as Vibrio vulnificus and Salmonella typhimurium but not gram-positive bacteria such as Staphylococcus aureus and Listeria monocytogenes were increased in URO-pretreated human PMNs. URO pretreatment did not induce TNF-α production and respiratory burst in PMNs as shown by dihydrorhodamaine 123 staining and flow cytometry. However, URO primed human PMN to produce more respiratory burst upon phorbol myristate acetate (PMA) challenge and enhance the binding of fluorescein isothiocyanate-lipopolysaccharide (FITC-LPS) to human PMNs. Not only human PMN, we also found that the bacterial clearance against gram-negative bacteria such as Vibrio vulnificus and Salmonella typhimurium but not gram-positive bacteria such as Staphylococcus aureus and Listeria monocytogenes were increased in mice treated with URO (200μg) 2h before challenge. The percentage and absolute number of PMN from URO-pretreated mice were increased. Furthermore, the phagocytosis activities of PMN from URO-pretreated mice were increased. Collectively, our results suggest that URO can enhance the function of PMN to clear bacterial infection in mice and human, especially against gram-negative bacteria. The results of this study provide us a better understanding of the immunostimulatory effects of URO on PMN and a potential target to develop new immunostimulating agent.

目錄
中文摘要---------------------------------------------------------------3
英文摘要---------------------------------------------------------------5
圖目錄------------------------------------------------------------------7
表目錄------------------------------------------------------------------8
誌謝---------------------------------------------------------------------9
縮寫--------------------------------------------------------------------10
實驗設計--------------------------------------------------------------12
緒論--------------------------------------------------------------------13
儀器與試劑-----------------------------------------------------------19
材料和方法-----------------------------------------------------------23
結果--------------------------------------------------------------------41
討論--------------------------------------------------------------------53
圖附錄-----------------------------------------------------------------65
表附錄-----------------------------------------------------------------77
參考文獻--------------------------------------------------------------78
作者簡介--------------------------------------------------------------86

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