臺灣博碩士論文加值系統

慢性氣喘(chronic asthma)是一種慢性呼吸道發炎疾病，患者在接觸到環境中的過敏原後出現呼吸道收縮、黏液分泌增加、免疫細胞大量浸潤，並發生呼吸困難、胸悶、咳嗽等症狀。泡沫狀巨噬細胞(foamy macrophage, foam cell)是巨噬細胞在吞噬修飾低密度脂蛋白後在細胞內形成脂肪滴的型態，在血管中泡沫狀巨噬細胞的堆積會導致血管壁增厚，增加心血管疾病發生的風險，但泡沫狀巨噬細胞在慢性氣喘發炎所扮演的角色仍未清楚。他汀類藥物(statin)是一種HMG-CoA還原酶抑製劑主要用於心血管疾病高風險患者，用來降低患者的膽固醇以降低心血管疾病病發的風險。許多研究顯示他汀類藥物可藉由抑制細胞訊息中間物生成以達到抑制發炎之效果。我們先前的研究顯示他汀類藥物可降低氣喘患者氣喘相關緊急就診或住院的風險，但是他汀類藥物是透過何種機制影響氣喘患者尚未明朗。因此本研究想了解他汀類藥物是否可藉由抑制泡沫狀巨噬細胞發炎以達到改善慢性氣喘的效果。
首先我們建立塵螨(Dermatophagoides pteronyssinus, Der p)誘導慢性氣喘的小鼠模式，並在此動物模式下給予他汀類藥物，評估肺部發炎及纖維化情形。實驗結果顯示慢性氣喘小鼠肺部有泡沫狀巨噬細胞堆積情形。在細胞實驗中，他汀類藥物可降低氧化低密度脂蛋白(oxidized low density lipoprotein, oxLDL)誘導泡沫狀巨噬細胞生成，且他汀類藥物可隨濃度梯度有效降低氧化低密度脂蛋白受體(SR-A1)表現量，但對於氧化低密度脂蛋白受體(CD36)、膽固醇排出蛋白(ABCG1)、細胞傳訊蛋白(STAT3, p38 MAPK)磷酸化的表現量則無明顯差異。在預防性刺激細胞實驗中，預先給予他汀類藥物具有降低TNF-α分泌的現象。然而，他汀類藥物治療的慢性氣喘小鼠組別的肺功能指數、氣管發炎現象、泡沫狀巨噬細胞堆積情形與慢性氣喘小鼠組別並沒有明顯改善。綜合以上結果，他汀類藥物可能通過下調SR-A1表現量來減少泡沫狀巨噬細胞的形成，未來我們可以透過將SR-A1表現抑制的相關實驗以證實我們的推論。在未來可以透過更多的實驗來了解泡沫狀巨噬細胞在慢性氣喘中扮演著怎樣的角色以及探討他汀類藥物是否可藉由改善泡沫狀巨噬細胞以達到改善慢性氣喘的效果以及可能的作用機制。

Chronic asthma is a chronic respiratory inflammation disease. Patients with chronic asthma may have airway hyperresponsiveness, mucus secretion, immune cells invasion in the lungs. Foamy macrophage is a type of macrophage that stored lipid droplets in cell after phagocytosis of modified low density lipoproteins (mLDL). In addition, foamy macrophages had high expression of inflammatory and fibrotic cytokines compared to normal macrophages. Previous studies indicated that foamy macrophages played the key role of atherosclerosis and pulmonary fibrosis. However, the role of foamy macrophages in chronic asthma is not yet clear. Several studies have suggested that statins, HMG-CoA reductase inhibitors, has multiple functions, such as inhibition of inflammation and regulation of cholesterol production. Our previously study showed that statins decreased the risk of asthma-related ED visits and/or hospitalizations in patients with asthma. The detail mechanisms of statins in chronic asthma are not yet clear. Therefore, we will investigate the role of foamy macrophages in chronic asthma and examine whether statins improve chronic asthma by decreasing formation of foamy macrophages. First, we established chronic asthma mouse model and found a lot of foamy macrophages were accumulated in the lungs of chronic asthma mice. Next, we examined whether atorvastatin decreased the number of foamy macrophages. The in vitro results showed that atorvastatin reduced the formation of foamy macrophages and the expression of oxLDL receptor (macrophage scavenger receptor 1, SR-A1). However, the expression of oxLDL receptor (cluster of differentiation 36, CD36), cholesterol excretion protein (ATP-binding cassette sub-family G member 1, ABCG1), and cellular signaling protein phosphorylation (STAT3, p38 MAPK) did not showed significant differences between atorvastatin-treated and control group. We further investigated the effects of atorvastatin in chronic asthma mouse model. However, atorvastatin-treated chronic asthma mice did not significantly improve lung function, inflammation, and the number of foamy macrophages in the lungs compared to chronic asthma mice. Several studies had shown that statins do work on asthma. As our study shows that statins decreased the formation of foamy macrophages might through down regulated SR-A1 expression. In this study, the relationship between chronic asthma, statins and foamy macrophages is still not clear yet. In the future, we should do further experiments to evaluate the relation between chronic asthma, statins and foamy macrophages.

中文摘要 I
英文論文 III
誌謝 XV
目錄 XVI
圖目錄 XVIII
縮寫檢索表 XIX
序論 1
一. 慢性氣喘簡介 1
二. 泡沫狀巨噬細胞簡介 2
三. 他汀類藥物簡介 3
四. 他汀類藥物與氣喘的關聯性 4
五. 研究方向 5
實驗材料與方法 6
一. 實驗材料 6
二. 實驗方法 9
1. 慢性氣喘小鼠模式的建立 9
2. 肺部呼吸系統功能測量 10
3. 血清分離、保存與測定 10
4. 肺部沖洗液的收集 10
5. 細胞計數與細胞鑑別性染色 11
6. 細胞激素的定量 11
7. 血清中IgE抗體分泌量測定 11
8. 血清中Derp特異性IgE抗體分泌量測定 12
9. 組織固定與染色 12
10. Masson’s trichrome染色 13
11. Oil Red O染色 13
12. Bone Marrow Derived Macrophage(BMDM)分化 14
13. 流式細胞術 14
14. 低密度脂蛋白純化與氧化 14
15. 蛋白質定量 15
16. 西方墨點法 16
結果 18
一. 使用BALB/c小鼠建立慢性氣喘動物模式 18
I. 慢性氣喘造成的肺功能降低 18
II. 慢性氣喘小鼠肺沖洗液中免疫細胞浸潤以及血清中IgE的表現 18
III. 小鼠肺部病理切片觀察 19
二. 測試阿托伐他汀在誘導單核球形成泡沫狀巨噬細胞的作用 19
I. 阿托伐他汀在泡沫狀巨噬細胞的作用 20
II. 阿托伐他汀在泡沫狀巨噬細胞可能的作用機制 20
III. 阿托伐他汀在泡沫狀巨噬細胞的抑制發炎作用 21
三. 阿托伐他汀在慢性氣喘小鼠的作用 22
I. 慢性氣喘造成的肺功能降低 22
II. 慢性氣喘小鼠肺沖洗液中免疫細胞浸潤以及血清中IgE的表現 22
III. 小鼠肺部病理切片觀察 23
IV. 慢性氣喘小鼠肺沖洗液中TARC與血清中TGF-β的表現 23
討論 25
圖表 28
參考文獻 40

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