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系統識別號 U0026-0812200912091256
論文名稱(中文) 動情素及山藥對於ApoE基因剔除小鼠動脈粥狀硬化進程與活性氧分子生成之影響
論文名稱(英文) Effects of Estrogen and Dioscorea on Atherosclerotic Progression and Superoxide Production in Apolipoprotien E Deficient (ApoE-/-) Mice
校院名稱 成功大學
系所名稱(中) 生理學研究所
系所名稱(英) Department of Physiology
學年度 94
學期 2
出版年 95
研究生(中文) 吳佳靜
研究生(英文) Jia-Jing Wu
學號 s3693401
學位類別 碩士
語文別 英文
論文頁數 58頁
口試委員 口試委員-江美治
召集委員-陳洵瑛
指導教授-陳麗玉
中文關鍵字 動脈粥狀硬化  山藥  動情素  apoE基因剔除小鼠 
英文關鍵字 Estrogen  Dioscorea  Atherosclerosis  apoE-/- mice 
學科別分類
中文摘要   過多的活性氧分子 (ROS) 會造成氧化壓力並且促使動脈粥狀硬化的病程進展。目前已知動情素 (Estrogen) 對於心血管系統具保護作用,其中的一種機制是經由抑制動脈粥狀硬化的發展達到其保護效果。山藥(Dioscorea)是東方國家的傳統食物,亦同時被認為藥用植物之一,其成分中含有植物性雌激素。本篇論文主題是使用ApoE基因剔除小鼠做為動物模式研究動情素及山藥是否能透過抗氧化機制影響動脈粥狀硬化的進程。ApoE- 基因剔除雌鼠經去卵巢手術後,分為實驗組及對照組,分別以灌食方式每日給予動情素或高低劑量 [10mg/體重(g), 2 mg/體重(g)]的山藥水煎萃取物TA-C。進行16週灌食之後,我們將小鼠麻醉後犧牲並取其主動脈進行Oil-red O染色,用以評估動脈粥狀硬化生成的情況;並收集血漿測量動脈中各類膽固醇含量。此外,我們利用化學螢光染色的方式偵測動脈組織中超氧化物 (Superoxide)的生成情形。研究結果顯示,動情素能減少動脈粥狀硬化以及超氧化物的生成,而TA-C對於血管組織中的動脈粥狀硬化及超氧化物的生成並無影響。TA-C對於ApoE基因剔除小鼠血漿中總膽固醇、三酸甘油酯(Triglyceride)、低密度脂蛋白(LDL)含量亦無顯著性影響。我們測試NADPH氧化酵素活性於U937這株人類單核球細胞株之變化,以探討動情素與皂苷(diosgenin)之抗氧化效果。實驗結果顯示,即使於10-7 M濃度的動情素與皂苷處理之下,U937細胞之NADPH氧化酵素活性仍未受到顯著影響。就以上結果而論,動情素可能藉由減少ApoE基因剔除小鼠動脈中活性氧分子的生成達到減低動脈粥狀硬化之效果,然而山藥水煎粹取物TA-C所含成分可能不足以對動脈粥狀硬化之發展造成影響。


英文摘要   Excessive production of reactive oxygen species causes oxidative stress that may leads to the development and progression of atherosclerosis. Previous evidences have shown that estrogen has protective effects against atherosclerotic progression. Dioscorea is a traditional food that has also been used as a medical herb. It contains phytoestrogen. In this study, we have studied whether estrogen and Dioscorea had anti-atherosclerotic action and antioxidant effect in ApoE-/- mice. Adult ovariectomized ApoE-/- mice received the daily oral gavage of 17β-estradiol (E2), Dioscorea water extract TA-C at low dose (2 mg/g B.W./day) or high dose (10 mg/g B.W./day). After 16 weeks, mice were sacrificed and their aortae were removed for oil-red O staining to evaluate atherosclerotic lesion area. The production of superoxide in aortic tissues was evaluated by using the DHE chemiluminescent staining. Our results showed estrogen reduced atherosclerotic progression and the production of superoxide in the vessel wall. However, TA-C had no effect. The lipid profile showed that TA-C treatment did not affect the serum levels of total cholesterol, triglyceride, and LDL in ApoE-/- mice. We also compared the antioxidant effect of estrogen and diosgenin (a pure compound isolated from Dioscorea) in U937 monocytic cell. We used NAD(P)H oxidase activity assay to evaluate the antioxidant effect of E2 and diosgein. The results showed that even at the concentration of 10-7 M, E2 and diosgenin did not reduce PMA-activated NAD(P)H oxidase activity. In conclusion, estrogen may exert its anti-atherosclerotic effects partially through reducing the superoxide production in the vessel wall in ApoE-/- mice. Dioscorea extract TA-C may not contain enough substances for the prevention of atherosclerotic progression.


論文目次 中文摘要.......................................1

ABSTRACT.......................................3

誌謝...........................................4

TABLE OF CONTENTS..............................5

LIST OF FIGURES................................7

LIST OF TABLE..................................8

INTRODUCTION...................................9

 PATHOGENESIS OF ATHEROSCLEROSIS:.............9
  Morphological features of atherosclerosis..9
  Mechanism of Atherogenesis................10

 OXIDATIVE STRESS IN ATHEROSCLEROSIS.........12

 NAD(P)H OXIDASE.............................13

 ESTROGEN AND ATHEROSCLEROSIS................15

 PHYTOESTROGEN AND DIOSCOREA.................17

 APOE-/- MOUSE MODEL OF ATHEROSCLEROSIS......18

 THE PURPOSE OF THIS STUDY...................20



MATERIAL AND METHODS..........................21

 ANIMAL AND TISSUE PREPARATION...............21

 PREPARATION OF DIOSCOREA TA-C EXTRACT.......22

 QUANTIFICATION OF ATHEROSCLEROSIS...........22

 DIHYDROETHIDIUM CHEMILUMINESCENCE FOR

 DETECTING SUPEROXIDE PRODUCTION.............23

 IMMUNOFLUORESCENT STAINING..................23

 CELL CULTURE................................24

 NAD(P)H OXIDASE ACTIVITY ASSAY..............24

  Treatment Protocols.......................24

  Preparation of cell membrane fraction.....26

  Measurement of NAD(P)H oxidase activity...26



RESULT........................................27

 PART I:.....................................27

  Effects of 17β-estradiol and Dioscorea extract TA-C on atherosclerotic progression in the aorta of ApoE-/- mice

 PART II .....................................28

  Effects of 17β-estradiol and Dioscorea extract TA-C on superoxide production in the vessel wall of aorta

 PART III....................................29

  Effects 17β-estradiol and diosgenin on NAD(P)H oxidase activity in U937 monocytic cells

DISCUSSION....................................31

REFERENCES....................................45

APPENDIX......................................50

自述..........................................58
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