臺灣博碩士論文加值系統

慢性傷口指未依照預期的時間癒合通常伴隨著慢性疾病，癒合時間長達四週以上，例如糖尿病足部潰瘍、壓瘡等。傳統的傷口敷料主要功能為覆蓋於傷口表面保護傷口，卻沒有主動癒合的功效，在治療傷口的效果有限且未能滿足現今需求，開發具有主動癒合的先進敷料是刻不容緩。本篇研究將開發具有生物活性之金屬離子濺鍍於高分子材料聚丙烯（Polypropylene, PP）以促進傷口癒合能力的敷料，透過一系列的物化試驗、細胞實驗、動物實驗與微生物試驗，完成傷口敷料上市前的製程參數與功效的確認。研究顯示鎂具生物可吸收降解，其離子更有促細胞增生、分化、遷移與膠原蛋白生成等功效，近幾年已被廣泛應用於生醫材料的開發上，但目前市場上卻沒有使用於傷口敷料的相關應用，所以衍生出將鎂離子與高分子材料結合成為新一代的新型複合傷口敷料。
此研究中實驗組別分成對照組(無濺鍍金屬離子)、含鎂離子敷料與含銀鎂離子敷料，在材料的物化試驗中證明金屬離子成功濺鍍於材料表面且能夠有效釋放，且其疏水特性不易沾黏傷口組織;在細胞實驗方面進行細胞遷移與細胞毒性試驗，發現含鎂離子和含銀鎂離子敷料能夠加快小鼠纖維母細胞(NIH-3T3)爬行速度與對照組相比達顯著性的差異，且對於人類臍靜脈內皮細胞(HUVECs)無毒性;在動物實驗分成兩組大鼠傷口模型（每組6隻），分別為大鼠切割傷口模型 (急性傷口)與糖尿病鼠傷口模型(慢性傷口)，在實驗期間每兩天更換一次敷料並拍照紀錄傷口大小，於第九天將大鼠犧牲取下傷口組織進行病理切片染色分析，包含蘇木精-伊紅染色(hematoxylin and eosin stain, H＆E stain)觀察組織結構其上皮生長情形，與免疫組織化學染色(immunohistochemistry, IHC)觀察新生角質細胞數量(Ki67 marker)與血管內皮再生面積(CD31 marker)。在急性傷口實驗結果發現含鎂離子敷料其上皮化程度、新生角質細胞數量和血管內皮細胞面積，與對照組相比皆達顯著性差異有較好的癒合效果;而慢性傷口是透過餵食4週60 %高脂飼料(High fat diets, HFD)並注射35 mg/kg低劑量的鏈脲佐菌(Streptozotocin, STZ)誘發成糖尿病鼠，實驗結果發現同時具有兩種金屬離子的含銀鎂離子敷料，其在上皮化程度和血管內皮細胞面積與對照組相比達顯著性差異，對於糖尿病鼠傷口有較好的癒合效果，於抑菌試驗中對大腸桿菌的抑菌率達到40%。從所述的結果表明，含有鎂離子的新型複合傷口敷料具有治療傷口的潛力。

Chronic wounds do not heal as expected, with healing times longer than four weeks, often accompanied by chronic diseases such as diabetic foot ulcers and pressure ulcers. The traditional wound dressing function is to cover the wound surface to protect the wound, but it does not have the effect of active healing. The effect of treating wounds is limited and cannot meet their current needs. It is urgent to develop advanced dressings with active healing. This research develops a dressing with bioactive metal ion sputtered on polypropylene (PP) to promote wound healing. Through a series of physicochemical tests, in vitro, in vivo and microbiological tests, parameters and confirmation effects are found. Magnesium used in this study is bioabsorbable and degradable, and its ions are found could promote cell proliferation, differentiation, migration, and collagen deposition. So far, magnesium ion coated wound dressings have been investigated. So we developed novel composite wound dressings combining magnesium ions on polymer dressing materials.
In this study, the experimental group was divided into a control group (without sputtered metal ions), a dressing containing magnesium ions, and a dressing containing silver-magnesium ions. In the physiochemical test of the material, it was proved that metal ions were successfully sputtered on the surface of the material and could be effectively released, and its hydrophobic property was not easy to stick to the wound tissue; in vitro, cell migration and cytotoxicity tests were performed and it was found that magnesium ions and silver-magnesium ions dressings can promote NIH-3T3 migration and have significant differences compared to the control group, and have no toxicity to HUVECs; in vivo, the rat 1-cm open wound model was divided into two groups (6 rats in each group), a normal rat wound model (acute wound) and a diabetic rat wound model (chronic wound). Dressings were replaced every two days during the experiment and photos were taken to record the wound size. On day 9, the rats were sacrificed and the wound tissue was removed for pathological analysis. Hematoxylin and eosin stain (H＆E stain) was used to observe the epithelial growth of the tissue structure, and immunohistochemistry (IHC) was used to observe the number of keratinocytes (Ki67 marker) and vascular endothelial regeneration area (CD31 marker).
In acute wounds, it was found that the dressing containing magnesium ions, whose percentage of re-epithelialization, the number of keratinocytes, and the area of ​​vascular endothelial cells, had significant differences compared to the control group and had better healing effects; chronic wounds were induced into diabetic rats by feeding 60% High fat diets (HFD) for 4 weeks and injecting Streptozotocin (STZ) at a low dose of 35 mg/kg. It was found that the silver-magnesium ions dressing with two kinds of metal ions, the percentage of re-epithelialization and the area of ​​vascular endothelial cells, had significant differences compared with the control group, and had better healing effects on chronic wounds. The bacteriostatic activity of the dressing in E. coli was 40%.
In summary, novel composite wound dressings containing magnesium ions have the potential to promote wound healing.

中文摘要 I
Abstract III
誌謝 V
Table of Contents VII
List of Tables X
List of Figures XI
Chapter 1: Introduction 1
1.1 Wound dressing 1
1.2 The structure of human skin 2
1.2.1 Epidermis layer 3
1.2.2 Dermis layer 5
1.2.3 Hypodermis layer 5
1.3 Stage of skin wound healing 6
1.3.1 Hemostasis phase 6
1.3.2 Inflammatory phase 6
1.3.3 Proliferation phase 7
1.3.4 Remodeling phase 7
1.3.5 Wound type 9
1.4The wound healing of active ions background-Mg2 + and Ag + 11
1.5 Motivation and aims of the research 12
Chapter 2: Material and Methods 13
2.1 Flow chart of experiment 13
2.2 Experimental materials 14
2.3 Experimental equipment 15
2.4 Physicochemical properties 16
2.4.1 Immersion test 16
2.4.2 Micrographs 16
2.4.3 Surface wettability 16
2.5 In vitro tests 17
2.5.1 Cell culture 17
2.5.2 Cytotoxicity assay 17
2.5.3 Cell migration 19
2.6 In vivo tests 20
2.6.1 Acute wound model 20
2.6.2 Chronic wound model 21
2.7 Antimicrobial assay 30
Chapter 3: Results and discussion 32
3.1 Physicochemical properties 32
3.1.1 Immersion test 32
3.1.2 Micrographs 35
3.1.3 Surface wettability 37
3.2 In vitro tests 38
3.2.1 Cytotoxicity assay 38
3.2.2 Cell migration 43
3.3 In vivo tests 45
3.3.1 Pre-test for dose-dependent parameters 45
3.3.2 Acute wound model 46
3.3.3 Chronic wound model 57
3.4 Antimicrobial 70
Chapter 4: Conclusion 72
Chapter 5: Future work 74
References 75

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