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研究生:李愛薇
研究生(外文):Ai-Wei Lee
論文名稱:探討熊果酸抑制動脈粥狀硬化與促進缺血後血管新生之相關性研究
論文名稱(外文):Ursolic acid inhibits atherogenesis and promotes post-ischemia angiogenesis
指導教授:馮琮涵馮琮涵引用關係
學位類別:博士
校院名稱:臺北醫學大學
系所名稱:醫學科學研究所
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:2013
畢業學年度:101
語文別:中文
論文頁數:111
中文關鍵詞:熊果酸動脈粥狀硬化
外文關鍵詞:ursolic acidatherosclerosis
相關次數:
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  • 收藏至我的研究室書目清單書目收藏:1
熊果酸(Ursolic acid;UA)是存在於天然植物中的一種五環三萜類植物皂苷(triterpenoid compound),普遍存在於人們的飲食以及中草藥,具有廣泛性的生物意義,包含抗發炎以及抗癌症等作用;然而,直到現在,熊果酸與血管新生以及抗動脈粥狀硬化的相關研究仍然十分罕見;因此,我們分別以人類冠狀動脈內皮細胞(human coronary artery endothelial cells;HCAECs)進行細胞實驗,以及C57BL/B6老鼠進行動物實驗,來探討熊果酸與血管疾病相關議題;並且探討可能的相關機制。本實驗證明,熊果酸在下肢缺血的C57BL/B6老鼠中,可以增加側枝血流的恢復,以及促進血管新生而增加微血管密度;在細胞實驗中,熊果酸可以增加人類血管內皮細胞的管柱生成、以及移行作用,同時發現內皮細胞透過一氧化氮相關機制,進而增加異體移植發炎因子ㄧ(allograft inflammatory factor-1;AIF-1)的表現;另外,已知HMGㄧ(high mobility group protein 1;HMGB1)與血管發炎有十分的相關性,本實驗證明HMGB1透過活化人類血管內皮細胞,或是高膽固醇C57BL/B6老鼠的氧化型低密度脂蛋白受體-1 (lectin-like oxidized low-density lipoprotein receptor -1;LOX-1),進而增加氧化低密度脂蛋白的吞噬,熊果酸可以透過抑制環氧化酶-2(cyclooxygenase-2;COX-2)與一氧化氮相關機制,進而減少氧化型低密度脂蛋白的吸收。這些結果證實熊果酸對於高膽固醇飲食所引發的動脈粥狀硬化,以及血管新生具有治療的效果,並對相關的疾病提供新的研究機制。

Ursolic acid (UA), a triterpenoid compound found in plants, is used in the human diet and in medicinal herbs and possesses a wide range of biological benefits including antioxidative, anti-inflammatory, and anticarcinogenic effects. However, until now, there are rare evidences to promote the benefit of UA on neovascularization and anti-atherogenesis. Therefore, we performed the studies using human coronary artery endothelial cells (HCAECs) in vitro and C57BL/B6 mice in vivo to identify the effects of UA in these issues. Furthermore, we explored the probably underlying mechanisms in this study. The results demonstrated that UA enhances collateral blood flow recovery and capillary density through induction of neovascularization in hind limb ischemia C57BL/B6 mice. In in vitro data showed that UA increases tube formation and migration capacities in human endothelial cells, and exposing HCAECs to UA increased allograft inflammatory factor-1(AIF-1) expression through a nitric oxide-related mechanism. Additionally, high-mobility group box 1 (HMGB1) is known to mediate vascular inflammation. We have demonstrated that HMGB1 enhances oxidized low density lipoprotein (oxLDL) uptake through induction of lectin-like oxidized low-density lipoprotein receptor 1 (LOX-1) in HCAECs and C57BL/B6 mice in this study. Exposing HMGB1-stimulated HCAECs and hypercholesterolemic C57BL/B6 mice to UA decreased the LOX-1-mediated absorption of oxLDL through a cyclooxygenase (COX)-2-related NO signaling pathway. These findings suggest that UA may be a potential therapeutic agent in the hypercholesterolemia-induced atherosclerosis and induction of neovascularization. It provides a novel mechanistic insight into the potential effects of UA on ischemic vascular diseases and atherosclerosis.

目錄
頁次
縮寫表‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥ i
目錄‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥iii
中文摘要‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥ vi
英文摘要‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥ vii
第一章、緒論
第一節 引言‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥2
第二節 熊果酸
一 探討熊果酸的抗發炎機制‥‥‥‥‥‥‥‥‥‥‥‥2
二 熊果酸對血管功能之影響‥‥‥‥‥‥‥‥‥‥‥‥4
第三節 High mobility group protein B1(HMGB1)
一 HMGB1在發炎反應中所扮演的角色‥‥‥‥‥ ‥‥7
二 HMGB1對血管內皮細胞功能的影響以及與心血管
疾病的關係‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥9
第四節 氧化型低密度脂蛋白受體-1
一 氧化型低密度脂蛋白受體-1在血管內皮細胞發炎反
應的角色以及動脈粥狀硬化的關係‥‥‥‥‥ ‥‥11
第五節 同種異體移植發炎因子一
一 同種異體移植發炎因子一與發炎反應的機制‥‥‥14
二 同種異體移植發炎因子一在血管內皮細胞的角色‥16
第六節 研究動機與目的‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥17
第二章、研究方法與材料
第一節 體外實驗
一 試劑‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥19
二 人類之HMGB1蛋白製備‥‥‥‥‥‥‥‥‥‥‥19
三 細胞培養‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥19
四 細胞存活率分析‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥20
五 內皮細胞管柱形成實驗‥‥‥‥‥‥‥‥‥‥‥‥21
六 傷口癒合試驗‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥21
七 纖維肌動蛋白染色與免疫螢光染色法‥‥‥‥‥‥21
八 細胞蛋白質萃取‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥22
九 西方點墨分析法‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥22
十 血管內皮細胞吞噬被DiI標示的氧化低密度脂蛋白 23
十一 細胞酵素免疫分析法‥‥ ‥‥‥‥‥‥‥‥‥‥‥24
十二 測量前列腺素E2 (prostaglandin E2;PGE2)的含量 ‥25
第二節 動物實驗
一 下肢缺血實驗‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥25
二 注射藥物引發動脈粥狀硬化的實驗‥‥‥‥‥‥‥26
三 血液生化值的測量‥‥‥‥‥‥‥‥‥‥‥‥‥‥27
四 形態學分析與免疫組織染色‥‥‥‥‥‥‥‥‥‥28
第三節 統計分析‥‥‥‥‥‥‥‥‥‥‥ ‥‥‥‥‥‥ 29
第三章、結果
第一部分 藉由熊果酸與血管內皮細胞的研究探討動脈粥狀
硬化的形成
一 熊果酸對實驗動物的毒性分析‥‥‥‥‥‥‥‥‥31
二 人類重組蛋白HMGB1對血管內皮細胞的作用‥‥ 32
三 熊果酸處理血管內皮細胞,可以抑制由HMGB1引
起細胞表面LOX-1的表現,以及增加血管內皮細胞
吞噬DiIoxLDL‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥32
四 熊果酸藉由抑制COX-2蛋白質的表現,而減少HMGB1
刺激血管內皮細胞所引起的LOX-1表現‥‥‥‥‥33
五 以HMGB1刺激血管內皮細胞,熊果酸藉由增加eNOS
的活性進而減少LOX-1的表現‥‥‥‥‥‥‥‥‥34
六 實驗動物注射重組蛋白HMGB1,會使血漿中HMGB1
以及Lactate的濃度增加‥‥‥‥‥‥‥‥‥‥‥‥35
七 實驗動物給予高膽固醇飲食並且注射HMGB1,會
明顯增加動脈粥狀硬化斑塊的形成以及LOX-1的
表現‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥36
第二部分 藉由熊果酸與血管內皮細胞的研究探討血管新生
一 熊果酸藉由增加老鼠微血管密度進而提高血流回復
的能力‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥37
二 熊果酸促進血管內皮細胞增生、管柱形成以及移行
‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥38
三 熊果酸透過活化內皮細胞的eNOS進而增加AIF-1
的表現‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥40
四 熊果酸可以增加動物的微血管密度以及eNOS與AIF-1
的表現‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥41
第四章、討論與結論
一 熊果酸與血管新生‥‥‥‥‥‥‥‥‥‥‥‥‥‥43
二 同種異體移植發炎因子一與血管新生‥‥‥‥‥‥44
三 HMGB1與心臟血管疾病‥‥‥‥‥‥‥‥‥ ‥‥46
四 氧化型低密度脂蛋白受體-1的表現與調控‥‥‥‥48
五 熊果酸會調控環氧化酵素-2引起的相關發炎反應‥49
六 熊果酸與脂質代謝‥‥‥‥‥‥‥‥‥‥‥‥‥‥50
七 結論‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥51
第五章、參考文獻‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥53
第六章、附圖‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥65
第七章、附表‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥ 77
第八章、論文相關著作‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥80


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