臺灣博碩士論文加值系統

異位性皮膚炎 (Atopic dermatitis；AD)是一種容易復發的慢性炎症皮膚疾病，由於體內免疫系統的過度反應，伴隨血清免疫球蛋白E (IgE)的上升，造成皮膚出現潮紅、濕疹、皮膚損傷及劇烈搔癢。金盞花 (Calendula officinalis Linn)是原產於 地中海地區的一年生草本植。金盞花從中古世紀開始就已經被普遍認識的菊科(Asteraceae family)植物，早期被製成軟膏用於傷口癒合的治療、抗敗血症和抗炎症等效果之應用。本實驗的目的為評估金盞花萃取物對於誘導異位性皮膚炎大鼠的之療效用，同時針對20%、30%和40%三種不同濃度之萃取物進行治療效果的比較。實驗對象為30隻雄性、六週齡的Sprague-Dawley品系大鼠，使用2, 4 - 二硝基氯苯 (2, 4-dinitirochlorobenzene；DNCB)進行異位性皮膚炎之誘導，每週兩次，將誘導試劑塗抹於頸背部區域，達誘導異位性皮膚炎的狀態，持續誘導至實驗結束，共六週。確認誘導成功後，治療組分別給予濃度40%、30%和20%的金盞花萃取物軟膏，塗抹部位與誘導部位相同，每週五次，連續治療四週，並於實驗結束後進行大鼠犧牲。本實驗的評估項目包含：皮膚炎臨床狀態評分、皮膚鏡觀察、皮膚組織切片H & E (hematoxylin and eosin)染色下表皮厚度及炎症細胞浸潤情形、皮膚組織切片TB (toluidine blue)染色下肥大細胞浸潤情形。結果顯示，皮膚炎臨床狀態評分數值方面，於治療中期並無明顯改善，但在治療期結束時，三組治療組與陽性對照組皆有極顯著性差異 (P < 0.01)；治療濃度30%和40%的組別在皮膚厚度測量上，與陽性組分別有極顯著性差異 (P < 0.01)和顯著性差異 (P < 0.05)，治療濃度20%的組別則不具有顯著差異 (P > 0.05)；僅治療濃度30%組別於炎症細胞浸潤數量上與陽性組有顯著性差異 (P < 0.05)；肥大細胞浸潤數量方面，治療濃度20%及30%兩組，分別與陽性組具有顯著性差異 (P < 0.05)和極顯著性差異 (P < 0.01)，其餘濃度40%組別則無顯著性差異 (P > 0.05)；血清總IgE抗體力價方面，治療濃度30%組和治療濃度40%組與陽性控制組都具有顯著性差異 (P < 0.05)，對於降低血清總IgE抗體力價有較佳的效果，且以治療濃度40%組為最佳。實驗過程中亦無產生明顯異常或副作用。本實驗結果支持金盞花萃取物能夠減緩異位性皮膚炎病程的效果。

Atopic dermatitis (AD) is a recurrent and chronic inflammation of skin disease. This disease is presented with immune system hyper-reactivity that progresses the high immunoglobulin IgE level and expressed erythematous lesion, eczema, skin damage and intense itching clinically. Calendula officinalis Linn is usually grown as an annual plant, it’s a native of the Mediterranean area. Since the Antiquity, medicinal properties have been attributed to the flower commonly known as C. officinalis, from the Asteraceae family. It was initially made by unguent and ointment to utilize for wound healing, antiseptic and anti-inflammatory. The aims of this study were to assess the efficacy of the extracts of C. officinalis on 2, 4-dinitrochlorobenzene (DNCB)-induced atopic dermatitis in rats. Thirty male 6-weeks-old Sprague-Dawley rats were randomly divided into 6 groups. The rats received DNCB topically application on the back for 4 weeks to induce AD-like lesions, and then treated with 40%, 30% and 20% concentration ethanol extracts ointment of C. officinalis for each treatment groups five times in one week for four weeks. After rats were sacrificed, the dermatitis scores and histological characteristics, including epidermal hyperplasia, infiltrating mast cells and inflammatory cells were analyzed. The dermatitis scores in all treatment groups were extreme significant lower than the control group (P < 0.01). The thickness of epidermis in the 30% and 20% groups were extreme significant lower than the control group (P < 0.01). The number of infiltrating inflammatory cells in the 30% group was significant lower than the control group (P < 0.05). The number of infiltrating mast cells in the 40% and 30% groups were extreme significant lower than the control group (P < 0.01). The level of IgE in 40% and 30% groups were significant lower than the control group (P < 0.05). The results indicated that C. officinalis could alleviate the histological change, decrease inflammatory effects and regulate immune responses of atopic dermatitis in rats. In conclusion, C. officinalis could be as an alternative drug for atopic dermatitis.

誌謝 i
摘要 ii
Abstract iii
目次 v
表目次 viiii
圖目次 ixx
第一章　緒言 1
第二章　文獻探討 2
第一節　皮膚解剖構造 2
一、表皮 2
二、基底膜帶 4
三、真皮 4
四、上皮附屬組織 4
第二節　異位性皮膚炎 5
一、異位性皮膚炎之簡介 5
二、異位性皮膚炎之病因與致病機轉 6
三、臨床診斷犬貓異位性皮膚炎的方法 7
四、臨床上治療異位性皮膚炎的方法 9
五、誘導異位性皮膚炎之實驗動物模型 10
第三節　金盞花 14
一、金盞花所含主要植物化學物質 14
二、金盞花的應用領域和使用方式 14
三、藥用植物的萃取方法 16
第三章　材料與方法 19
第一節　實驗動物 19
第二節　實驗材料 19
一、藥物與試劑 19
二、儀器與設備 20
三、其他 21
第三節　實驗設計 22
一、實驗分組 22
二、建立異位性皮膚炎的動物模型 23
三、萃取物製備與應用 24
四、評估異位性皮膚炎改善的效力 25
五、數據分析 27
第四章　結果 29
第一節　臨床症狀觀察 29
一、環境適應期 29
二、異位性皮膚炎誘導期 29
三、金盞花萃取物治療期 29
第二節　體重變化紀錄 29
第三節　皮膚狀態肉眼觀察與皮膚炎狀態肉眼觀察評分 33
第四節　皮膚切片H & E染色後皮膚厚度觀察 44
第五節　皮膚組織切片H & E染色後炎症細胞浸潤情形 48
第六節　皮膚組織切片TB染色後肥大細胞浸潤情形 51
第七節　血清總IgE抗體力價檢測 55
第八節　血清總IgG抗體力價檢測 57
第五章　討論 61
第一節　誘導異位性皮膚炎效果評估 61
第二節　實驗中動物的體重變化 61
一、環境適應期 61
二、異位性皮膚炎誘導期 61
三、金盞花萃取物治療期 61
第三節　治療效果評估 62
一、肉眼下皮膚炎狀態評分 62
二、皮膚組織切片H & E染色後皮膚厚度觀察 62
三、皮膚組織切片H & E染色後炎症細胞浸潤之觀察 63
四、皮膚組織切片TB染色後肥大細胞浸潤之觀察 63
五、血清總IgE抗體力價檢測 64
六、血清總IgG抗體力價檢測 65
第四節　比較三種不同濃度的金盞花萃取物在治療上的差異 66
第五節　實驗中可能影響治療結果的因素 67
一、實驗動物品系的差異 67
二、萃取方式的不同與給予方式的差異 67
三、金盞花原產地與季節的不同 68
第六章　結論 69
參考文獻 70

表目次
表 1、犬異位性皮膚炎臨床診斷標準 8
表 2、各組大鼠於實驗期間之體重數值 30
表 3、各組大鼠於實驗期間之皮膚炎臨床狀態評分 34
表 4、各組大鼠於實驗期間之皮膚表皮層厚度 45
表 5、各組大鼠於實驗治療期結束後之皮膚組織切片-炎症細胞浸潤數量 48
表 6、各組大鼠於實驗治療期結束後之皮膚組織切片-肥大細胞浸潤數量 51
表 7、各組大鼠於實驗各階段之血清總IgE抗體力價 55
表 8、各組大鼠於實驗各階段之血清總IgG抗體力價 58

圖目次
圖1、皮膚組織的分層與結構圖 3
圖2、異位性皮膚炎的致病機轉的免疫途徑 7
圖3、實驗設計流程圖 22
圖4、各組大鼠於實驗期間之體重數值 29
圖5、各組大鼠於實驗誘導期結束後之體重數值 32
圖6、各組大鼠於實驗治療期結束後之體重數值 33
圖7、各組大鼠於實驗環境適應期結束後之皮膚炎臨床狀態評分 35
圖8、各組大鼠於實驗誘導期結束後之皮膚炎臨床狀態評分 36
圖9、各組大鼠於實驗治療期結束後之皮膚炎臨床狀態評分 37
圖10、各組大鼠於環境適應期之肉眼 38
圖11、各組大鼠於環境適應期之皮膚鏡觀察 (20X) 39
圖12、各組大鼠於誘導期結束後之肉眼病變 40
圖13、各組大鼠於誘導期結束後之皮膚鏡觀察 (20X) 41
圖14、各組大鼠於治療期結束後之肉眼病變 42
圖15、各組大鼠於治療期結束後之皮膚鏡觀察 (20X) 43
圖16、各組大鼠於治療期結束後之皮膚表皮層厚度 45
圖17、各組大鼠於實驗治療期結束後之組織切片-皮膚厚度之觀察 [H & E染色] (100X) 46
圖18、各組大鼠於實驗治療期結束後之組織切片-皮膚厚度之觀察 [H & E染色] (200X) 47
圖19、各組大鼠於實驗治療期結束後之炎症細胞浸潤數量 49
圖20、各組大鼠於實驗治療期結束後之組織切片-炎症細胞浸潤之觀察 [H & E染色] (200X) 50
圖21、各組大鼠於實驗治療期結束後之肥大細胞浸潤數量 52
圖22、各組大鼠於實驗治療期結束後之組織切片-肥大細胞浸潤之觀察 [TB染色] (200X) 53
圖23、各組大鼠於實驗治療期結束後之組織切片-肥大細胞浸潤之觀察 [TB染色] (400X) 54
圖24、各組大鼠實驗各階段之血清總IgE抗體力價 56
圖25、各組大鼠實驗各階段之血清總IgG抗體力價 59

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