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研究生:洪慈璟
研究生(外文):Thu-Ching Hung
論文名稱:檳榔素對於人類臍靜脈內皮細胞氧化傷害之探討
論文名稱(外文):Effect of Arecoline on Oxidative stress in Human Umbilical Vein Endothelial Cells
指導教授:張基隆張基隆引用關係
指導教授(外文):Kee-Lung Chang
學位類別:博士
校院名稱:高雄醫學大學
系所名稱:醫學研究所
學門:醫藥衛生學門
學類:醫學學類
論文種類:學術論文
論文出版年:2011
畢業學年度:99
語文別:中文
論文頁數:147
中文關鍵詞:檳榔素血管內皮細胞血紅素氧化酶-1
外文關鍵詞:ArecolineHuman Umbilical Vein Endothelial CellsHO-1
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檳榔素(Arecoline)為檳榔子中最主要的生物鹼成份,長期以來被認為是嚼食檳榔者導致口腔癌之重要化學致病因子。已有研究指出,嚼食檳榔可能提高心血管疾病風險,但詳細的分子作用機制並不清楚。根據衛生署統計,心臟和腦血管疾病高居國人十大死因之第二、三名。造成心血管疾病致死的主要原因由動脈粥狀硬化所引起,動脈粥狀硬化是一種緩慢且複雜的過程,引起血管內皮細胞功能異常。因此,本研究探討檳榔素對於人類臍靜脈內皮細胞之影響,包括細胞毒性、細胞週期分析、氧化傷害及細胞黏著分子的表現等。
本研究發現人類血管內皮細胞經檳榔素刺激後,細胞週期會停滯在G2/M期,並抑制DNA合成。檳榔素會促進人類臍靜脈內皮細胞產生大量ROS,並促進細胞黏著分子intercellular cell adhesion molecule-1 (ICAM-1)及vascular cell adhesion molecule-1 (VCAM-1)大量表現。由於細胞內ROS的產生,造成細胞內抗氧化酵素及成分含量減少:包括superoxide dismutase (SOD)、catalase以及glutathione (GSH)。在加入檳榔素處理的細胞中再加入抗氧化劑GSH、N-acetyl-L-cysteine (NAC)可以減少ROS的產生與ICAM-1及VCAM-1的表現,而加入buthionine sulfoximine (BSO) 則會有加重的情形。也發現檳榔素會促使氧化壓力蛋白HO-1大量表現,利用HO-1 抑制劑zinc protoporphyrin-Ⅸ (ZnPP) 則會增加細胞的氧化傷害及ICAM-1與VCAM-1 的表現增加。實驗中分別利用ERK、JNK、p38 MAPK以及tyrosine kinase pathway的抑制劑加以作用,發現檳榔素主要是藉由JNK、p38 MAPK以及tyrosine kinase pathway調控細胞內HO-1的表現,而非ERK pathway。並利用細胞轉殖的方式,使人類臍靜脈內皮細胞大量表現HO-1蛋白,實驗結果得知,HO-1的大量表現可抑制檳榔素對於血管內皮細胞的毒性作用。
經由上述研究可了解檳榔素作用人類血管內皮細胞後,影響抗氧化分子的表現及相關訊息傳遞路徑,可作為預防心血管疾病及營養保健上的臨床營養防治學理依據和應用評估。


Arecoline, the most abundant areca alkaloid, has been reported to stimulate reactive oxygen species (ROS) production in several cells. Overpeoduction of ROS has been implicated to atherogenesis. Hemeoxygenase-1 (HO-1) has cytoprotective activities in vascular tissues. This study investigated the effect of arecoline on the pathogenesis of atherosclerosis and explored the role of HO-1 in this process. Human Umbilical Vein Endothelial Cells (HUVECs) were treated with arecoline, followed by analysis of ROS, adhesion molecules, and HO-1 expressions, meanwhile, the potential signaling pathways were determined. After arecoline treatment, ROS production was stimulated, adhesion molecules ICAM and VCAM expressed coincidently. Glutathione pretreatment could totally inhibit arecoline-stimulated ROS production and VCAM expression but not ICAM expression. Arecoline simultaneously induced HO-1 expression and which was partially relative to ROS stimulation. Inhibition of c-jun N-terminal kinase (JNK) by SP600125, p38 by SB 203580 or tyrosine kinase by genistein reduced arecoline-induced HO-1 expression. In contrast, inactivation of ERK (extracellular signal-related MAP kinase) by PD98059, had no effect. Transfection of GFP/HO-1 gene to HUVEC with 5-folds HO-1 activities could efficiently but not completely attenuate the decrease in cell viability by arecoline. This study demonstrates in HUVEC arecoline stimulated ROS production associated with ICAM and VCAM expressions. Under this condition, HO-1 expression was up-regulated through ROS, tyrosine kinase and MAPK (JNK and p38) signaling pathways.

目 錄

壹、中文摘要----------------------------------------------- 1
貳、英文摘要----------------------------------------------- 3
參、前言----------------------------------------------------- 4
血管內皮細胞----------------------------------------- 4
動脈粥狀硬化----------------------------------------- 9
檳榔---------------------------------------------------- 19
自由基------------------------------------------------- 23
血紅素氧化酶---------------------------------------- 26
肆、研究動機與目的------------------------------------- 31
伍、實驗材料---------------------------------------------- 32
陸、實驗方法---------------------------------------------- 39
柒、實驗結果---------------------------------------------- 54
捌、討論---------------------------------------------------- 63
玖、參考文獻---------------------------------------------- 73
拾、圖表--------------------------------------------------- 102
拾壹、附錄------------------------------------------------ 135


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