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系統識別號 U0026-2807201515484200
論文名稱(中文) 研究凝血酶調節素之類凝集素結構域在皮膚傷口癒合中所扮演的角色
論文名稱(英文) Study on the role of the lectin-like domain of thrombomodulin in cutaneous wound healing
校院名稱 成功大學
系所名稱(中) 醫學檢驗生物技術學系
系所名稱(英) Department of Medical Laboratory Science and Biotechnology
學年度 103
學期 2
出版年 104
研究生(中文) 吳懿軒
研究生(英文) Yi-Xuan Wu
學號 T36021138
學位類別 碩士
語文別 英文
論文頁數 81頁
口試委員 指導教授-吳華林
口試委員-施桂月
口試委員-江美治
口試委員-林淑華
中文關鍵字 凝血酶調節素  傷口癒合  巨噬細胞 
英文關鍵字 Thrombomodulin  Wound healing  Macrophages 
學科別分類
中文摘要 凝血酶調節素(TM) 是一種穿膜醣蛋白,主要表現在血管內皮細胞及其他角質細胞及巨噬細胞等。過去的研究證明在皮膚傷口癒合的過程中,角質細胞會大量表現凝血酶調節素,過去研究指出凝血酶調節素之胺基端類凝集素結構域 (第一結構域) 具有抗發炎的功能。皮膚傷口癒合主要由三種過程所組成,包含發炎期、組織增生期及組織修飾期。巨噬細胞在組織修復期間會大量浸潤的細胞,主要功能有二,分別為清除外來物質及細胞殘渣,防止組織感染;其次是分泌發炎前驅物質及數種成長因子來促使細胞增生。高遷移率族蛋白1 (HMGB1) 是屬於細胞核中與DNA進行結合之蛋白,一旦受到發炎刺激,HMGB1被視為一種發炎前驅物質從細胞核中釋放到細胞外,執行下游的反應。在本篇研究,我們證明全身性缺乏凝血酶調節素之類凝集素結構域的小鼠(TMLeD/LeD 小鼠)其皮膚傷口癒合速度明顯延遲。此外,TMLeD/LeD 小鼠的巨噬細胞其白血球介素-6, 白血球介素-1β及單核球趨化激素-1表現量上升。在巨噬細胞內高遷移率族蛋白之釋放在腫瘤壞死因子-α 刺激之下也有上升的趨勢。而且,在TMLeD/LeD小鼠的傷口處,巨噬細胞與嗜中性球的浸潤量以及白血球介素-6與單核球趨化激素-1的表現量亦有較多的現象。最後,TMLeD/LeD 小鼠與鏈脲佐菌素所誘發之糖尿病小鼠在傷口給予rTMD1重組蛋白具有促進傷口癒合的效果。此外,給予重組蛋白rTMD1可以有效減少糖尿病小鼠傷口處促發炎分子白血球介素-6與白血球介素-1beta的產生及巨噬細胞與嗜中性球的浸潤。綜合以上結果,我們發現凝血酶調節素之類凝集素結構域在皮膚傷口癒合過程中具有調控發炎的功能,而rTMD1重組蛋白具有治療皮膚傷口之潛力。
英文摘要 Thrombomodulin (TM) is a type I transmembrane glycoprotein that is predominantly expressed on the surface of vascular endothelial cells and some other cells such as keratinocytes and macrophages. Previous studies have demonstrated that TM is highly expressed in keratinocytes during wound healing. Previous reports have indicated that the N-terminal lectin-like domain of TM (TM domain 1, TMD1) plays a role in inhibition of inflammatory reaction. Cutaneous wound healing consists of three stages, including inflammatory phase, tissue proliferation phase, and tissue remodeling phase. Macrophages represent one of the most abundant inflammatory cell types during all stages of tissue repair. Macrophages not only cleanse foreign bacteria and particles but also secrete pro-inflammatory mediators and numerous growth factors, contributing to tissue growth. High mobility group box protein 1 (HMGB1) is a nuclear DNA binding protein, which can be secreted as a pro-inflammatory cytokine from nucleus to cytosol or out of various cells stimulated with pro-inflammatory cytokines. In this study, we demonstrated that the wound healing rate was slower in mice lacking the lectin-like domain of TM (TMLeD/LeD mice) than in wild-type mice in cutaneous wound healing assay in vivo. Furthermore, peritoneal macrophages from TMLeD/LeD mice had higher gene expression levels of interleukin-6 (IL-6) and monocyte chemoattractant protein-1 (MCP-1). Secretion of HMGB1 in cytosol and conditioned medium was higher in macrophages from TMLeD/LeD mice after stimulation with tumor necrosis factor-α (TNF-α). In addition, macrophage and neutrophil infiltration were increased at the wound sites of TMLeD/LeD mice on day 3 after wounding, and so did the protein expression of IL-6, interleukin-1 beta (IL-1β), and MCP-1. Finally, local administration of recombinant TMD1 (rTMD1) to the wound site significantly improved the wound healing in TMLeD/LeD mice and streptozotocin (STZ)-induced diabetic mice. In addition, rTMD1 inhibited pro-inflammatory cytokine IL-6 and IL-1β production and decreased neutrophil and macrophage infiltration at the wound site in diabetic mice. In summary, we found that TMD1 functions as a key regulator of inflammatory reaction in cutaneous wound healing and has therapeutic potential for the treatment for cutaneous wounds.
論文目次 Contents
中文摘要 1
Abstract 2
Acknowledgement 3
Contents 4
List of Figures 7
Abbreviation 8
Instruments 11
Reagents and Chemicals 13
Introduction 16
Objective of this study 21
Material and Methods 22
Results 48
Discussion 54
Reference 58
Figures 64
Appendixes 78
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