臺灣博碩士論文加值系統

本研究目的在探討山藥 (Dioscorea alata L.) 黏多醣對於人類皮膚纖維母細胞促進傷口癒合之影響。實驗部分均為體外培養試驗，主要針對三個方向進行探討：1、山藥黏多醣對人類皮膚纖維母細胞的增生評估；2、山藥黏多醣對人類皮膚纖維母細胞遷移行為的影響；3、山藥黏多醣對人類皮膚纖維母細胞合成細胞外基質之能力評估；以期瞭解山藥黏多醣應用於皮膚傷口修復的影響。
研究發現山藥黏多醣對於人類皮膚纖維母細胞有增生的作用，於10％血清培養狀態下，發現在濃度1.56~25 μg/ ml時有較佳的促進生長效果；而藉由低血清培養模式 (1% FBS) 評估下，其增生的效果相對於控制組而言有更好的促進增生之能力，在濃度0.39~200μg/ ml時均有促進人類皮膚纖維母細胞生長的效果；且於高濃度山藥黏多醣處理下，效果較為顯著，於100μg/ ml時有最好的促進生長能力。
而山藥黏多醣對人類皮膚纖維母細胞有促進遷移能力的效果，利用顯微鏡觀察，於兩百倍隨機視野下發現遷移的細胞數隨加入山藥黏多醣的濃度有正相關性，在濃度6.25、50 μg/ ml時，與控制組有顯著差異。
在山藥黏多醣對皮膚纖維母細胞合成細胞外基質之能力評估方面，發現山藥黏多醣可以促進膠原蛋白及纖黏連蛋白 (fibronectin) 的產生；山藥黏多醣於濃度12.5μg/ ml時，相對於控制組而言，可以顯著促進膠原蛋白的產生；而在山藥黏多醣濃度大於12.5μg/ ml下，與控制組相比，達到顯著促進纖黏連蛋白生成的效果。

The purpose of this study is to explore the effects of yam mucopolysaccharide from Dioscorea alata L. on wound healing. The thesis includes three parts : 1. The effects of yam mucopolysaccharide on human skin fibroblasts proliferation in vitro. 2. The effects of yam mucopolysaccharide on human skin fibroblasts migration in vitro. 3. The effects of yam mucopolysaccharide on the extracellular matrices production by the fibroblasts in vitro. The results obtained might shed the light of the effects of yam mucopolysaccharide on skin wound healing.
The results show that yam mucopolysaccharide enhanced human skin fibroblasts proliferation. The higher the concentrations of yam mucopolysaccharide (ranged between 1.56~25 μg/ ml) in 10% serum medium, the better the results on the skin fibroblasts proliferation. The effects of yam mucopolysaccharide on fibroblasts proliferation are more significant for the skin fibroblasts cultured in 1% serum medium at concentrations range between 0.39~200μg/ ml. The enhancement is dose-dependent between 0.39~100μg/ ml on 1% serum medium. The optimal concentration is 100μg/ ml.
Yam mucopolysaccharide can also promote migration of human skin fibroblasts. The cells in 200 × times microscopic fields were counted. The beneficial effect of yam mucopolysaccharide on skin fibroblasts migration was dose-dependent. The effects of yam mucopolysaccharide on the fibroblasts migration is significantly different from the control when concentrations are 6.25 μg/ ml and 50 μg/ ml.
The results of the effects of yam mucopolysaccharide on the extracellular matrices excretion by the fibroblasts, showed that yam mucopolysaccharide facilitated fibroblasts to secrete either collagen or fibronectin.
At a concentration of 12.5μg/ ml of yam mucopolysaccharide, the collagen secretion by the skin fibroblasts is significantly higher than the control. So as the enhancement on the secretion of fibronectin at the slightly higher concentrations (over 12.5μg/ ml).

中文摘要................................................I
英文摘要………………………………………………………………Ⅱ
目錄……………………………………………………………………Ⅳ
壹、前言………………………………………………………………1
貳、文獻整理.………………………………………………….……3
一、 山藥簡介……………………………………………………...3
1. 概說……………………………………………………………...3
2. 栽培種類及品種………………………………………………….3
3. 成分特性及效能………………………………………………….6
二、 皮膚簡介—皮膚的構造……………………………………..11
三、 傷口的修復…….……….……………………………………12
1. 細胞的增生作用與細胞外間質的合成…………………………12
2. 受傷組織的修復與細胞的遷移行為………………………....14
2. 1. 膠原蛋白的來源與生合成……………………………...14
2. 2. 膠原蛋白對傷口組織之功能…………………………...16
2. 3. 纖黏連蛋白 (fibronectin) 的表現….……………….17
參、 材料與方法……………………………………………..……22
一、 實驗大綱……………………………………………….…22
二、 實驗流程………………………………………………………22
三、 實驗材料………………………………………………………24
1. 細胞株………………………………………………..….…24
2. 山藥材料之來源………………………………………...…24
3. 藥品………………………………………………………………24
4. 設備……………………………………………………………..26
肆、 實驗方法………………………………………………………28
1. 山藥黏多醣之製備….………………………………………….28
2. 一般成分分析………………………………………………....29
2. 1. 水分………………..…………………………………….29
2. 2. 灰份……………………………………………………….29
2. 3. 粗蛋白……………..…………………………………….30
2. 4. 全糖量……………..…………………………………….31
3. 皮膚纖維母細胞的培養.………………………………….....31
4. 山藥黏多醣對皮膚纖維母細胞生長能力之評估 (MTT assay)…….33
5. 山藥黏多醣對皮膚纖維母細胞遷移能力的評估….………….33
5. 1. Migration assay………………..……………........33
5. 2. Wound-healing assay…………………………........34
6. 山藥黏多醣對皮膚纖維母細胞合成胞外基質成份的影響……35
6. 1. 山藥黏多醣對皮膚纖維母細胞促進膠原蛋白合成之作用…..35
6. 2. 山藥黏多醣對皮膚纖維母細胞促進纖黏連蛋白合成之影響....35
7. 統計分析…………………………………………………………36
肆、 結果與討論……………………………………….…...……38
1. 山藥黏多醣的萃取率及基本成分之分析………………………38
1.1 萃取率………………………………………………………38
1.2. 基本成分分析 ……………………………………………38
2. 山藥黏多醣對皮膚纖維母細胞生長能力之影響………………39
2.1. 纖維母細胞……………………………………………….39
2.2. 山藥黏多醣對於皮膚纖維母細胞的生長能力評估…….39
2.2.1. 於10%正常血清狀態觀察山藥黏多醣對細胞增生的影響....40
2.2.2. 於1%低血清狀態下觀察山藥黏多醣對細胞增生的影響…..40
2.2.3. 於不同培養時間下山藥黏多醣對細胞增生的影響….41
3. 山藥黏多醣對皮膚纖維母細胞的遷移能力之評估……………42
3.1. Migration assay…………………………………………42
3.2. Wound-healing assay……………………………………43
4. 山藥黏多醣對皮膚纖維母細胞合成細胞外基質能力之評估…44
4.1. 山藥黏多醣對膠原蛋白 (Collagen) 合成之影響…….44
4.2. 山藥黏多醣對纖黏連蛋白 (Fibronectin) 合成之影響....45
伍、結論…………………………………………………………….47
陸、參考文獻…………………………….…………………………48
圖……………………………………….……………………………57
表……………………………………………………….……………73

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