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研究生: 郭廷雍
Kuo, Ting-Yung
論文名稱: 蘭嶼豬的傷口癒合模型及各年齡層的皮膚變化
Wound Healing Models and Age-Related Skin Changes in Lanyu Pigs (Sus scrofa)
指導教授: 黃玲惠
Huang, Lynn-Huei
學位類別: 博士
Doctor
系所名稱: 生物科學與科技學院 - 生物科技與產業科學系
Department of Biotechnology and Bioindustry Sciences
論文出版年: 2024
畢業學年度: 112
語文別: 英文
論文頁數: 249
中文關鍵詞: 蘭嶼豬傷口修復傷口陣列修復模型生長因子鹿茸
外文關鍵詞: Lanyu pig, Wound healing, Wound array model, Growth factor, Deer velvet antler
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  • 本研究旨在探討台灣特有品種蘭嶼豬(Sus scrofa)作為未來老年病學和美容保健品研究的動物模型,試驗涵蓋從胎兒階段到120月齡的豬隻。分析各年齡階段的豬隻皮膚分別為胎兒100天、出生後1天及1、2.5、5、7、9、12、18、24、32、42、48、60、70、90和120個月。使用組織病理學分析、免疫組織化學(IHC)和分子細胞學觀察各年齡階段皮膚組織隨年齡變化。組織病理學分析發現皮厚度: 從1月齡的1.63 ± 0.24 mm增加到32個月的6.71 ± 0.40 mm,隨後減少至120月齡。表皮層厚度從1月齡的25 ± 1 µm增加到18月齡的38 ± 1 µm,之後無顯著變化。真皮層總細胞數從1月齡到18月齡,其細胞數量減少一倍。基底膜皺褶數從1月齡的12個減少到120月齡的6個。免疫組織化學分析發現表皮幹細胞隨年齡增加而減少。膠原蛋白與膠原蛋白間隙隨年齡增加而增加,致使皮膚組織形態早期階段(1-18月齡)組織緻密,老年階段(32-120月齡)組織鬆散。膠原蛋白分布量在I型膠原蛋白(COL-1)嬰兒期顯現略有增加,隨年齡逐漸增加,另III型膠原蛋白(COL-3)在胎兒期最高,隨年齡增加逐漸減少。總膠原蛋白分布量於32月齡時最高,隨年齡減少。膠原蛋白束厚度隨豬隻年齡增加而增厚。彈性纖維蛋白型態結構在年輕豬隻為纖細,老年豬隻變短且變厚,試驗結果顯示在48月齡豬隻之皮膚型態及組織顯現重要的轉變期,為豬隻自青壯期進入老齡期的分界點,並與老齡的人類皮膚具高度相似性。與家豬(Landrace)的比較,蘭嶼豬其體型和體重較小且皮膚表面積增長率較家豬小。傷口收縮速度顯示前6週顯著收縮,之後減少,蘭嶼豬在傷口癒合觀察6週後,因生長而致皮膚表面積增長影響較小。傷口再上皮化顯示不同年齡之間無差異。組織學結果顯示蘭嶼豬與家豬具相似的癒合和重塑模式。傷口癒合研究的適用性以蘭嶼豬較適合,因其體型較小和穩定的傷口癒合特徵。應用皮膚傷口陣列評估傷口癒合期間血流信號的測量結果顯示傷口間距需大於4公分,而評估傷口癒合所需開創的最佳傷口尺寸應大於4 x 4 cm2。皮膚傷口癒合前後之分子和細胞學檢測分析結果顯示mRNA表現量與細胞數量變化於癒合期間顯著增加。
    隨後的一項研究評估了水鹿鹿茸 (DVA) 水萃取物在促進糖尿病蘭嶼豬傷口癒合方面的功效。結果顯示DVA 萃取物顯著改善傷口閉合併增加灌注,可以作為治療正常和受損傷口的經濟藥物製劑。蘭嶼豬在生理和形態學特徵上與人類皮膚相似,使其成為老年病學和美容保健品研究的理想動物模型。蘭嶼豬因其穩定的傷口癒合特徵和較小的體型,是開發治療傷口、評估敷料、細胞療法和藥物的重要動物模型。

    In this study, we stablished 5 research with Lanyu pigs in substitute human wound therapy study: (1) Age-related skin architecture transition time points were observed at 36~48-month-old pigs which physiological status at senior adult stage to aged adult at early stage. (2) Lanyu pig is a better model system for studying wound healing than domestic Landrance pig. (3) Golden standards for minimum wound size of 4 cm × 4 cm and distance by ≥4 cm between adjacent wounds for effectively differentiating interventions in considering 3R principles. (4) The mRNA expression level of GF and its receptors, Cytokine, Proteoglycan, Matrix metalloproteinases and inhibitors, COL-1, Bone morphogenetic protein-1 and HSP-47 were significantly higher but TIMP-3 were depressed at specific times after wounding. (5) A long-term hyperglycemia diabetes animal model of Lanyu pigs were successful induced and diabetic wound treated with deer velvet antler extract may accelerate the wound closure rate through angiogenesis and thus accelerate wound healing. Therefore, our ideal preclinical animal model is crucial for assessing wound healing and management for improving skin wound healing and to meet the growing demand for developing biomedical products in acute and chronic wound therapy.

    Chinese abstract (中文摘要) I Abstract IV Acknowledgement VIII Table of contents IX Contents of tables XI Contents of figures XII Abbreviation list XV Chapter 1. Research background 1 1-1 Skin structure and functions 1 1-2 Wound and wound healing 9 1-3 Wound care product 27 1-4 Purpose 28 1-5 Specific Aims 30 Chapter 2. Unveiling Skin Tales: A Comparative Morphometric Analysis of Age-Related Skin Changes Across the Lifespan of Lanyu pigs (Sus scrofa) 32 2-1 Introduction 32 2-2 Materials and methods 40 2-3 Result 49 2-4 Discussion 55 Chapter 3. Lanyu pigs (Sus scrofa): An Excellent Animal Model for Wound Healing Assessment 58 3-1 Introduction 58 3-2 Materials and methods 61 3-3 Result 64 3-4 Discussion 69 Chapter 4. Assessment of Skin Wound Healing via an Optimized Wound Array Model in Lanyu Pigs (Sus scrofa) 73 4-1 Introduction 73 4-2 Materials and methods 76 4-3 Result 86 4-4 Discussion 96 Chapter 5. Molecular and Cellular Mechanisms of Skin Wound Healing in the Lanyu Pigs (Sus scrofa) Model 100 5-1 Introduction 100 5-2 Materials and methods 103 5-3 Result 110 5-4 Discussion 121 Chapter 6. Effects of Taiwan Red Elk Deer velvet antler on Cutaneous Wound Healing in Alloxan-Induced Diabetic Lanyu Pigs (Sus scrofa) 125 6-1 Introduction 125 6-2 Materials and methods 126 6-3 Result 130 6-4 Discussion 133 Chapter 7. Conclusions 137 References 140 Tables 162 Figures 164 Publication list 230

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