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論文中文名稱:以幾丁聚醣為基質探討電磁場對成骨組織之影響 [以論文名稱查詢館藏系統]
論文英文名稱:Utilized Chitosan as Matrix Discuss the Effect of Electromagnetic Fields on Bone Tissue Engineering [以論文名稱查詢館藏系統]
院校名稱:臺北科技大學
學院名稱:工程學院
系所名稱:化學工程研究所
出版年度:97
中文姓名:呂科賢
英文姓名:Ko-Hsien Lu
研究生學號:95738044
學位類別:碩士
語文別:中文
口試日期:2008-07-31
論文頁數:70
指導教授中文名:林忻怡
口試委員中文名:蔡偉博;胡勇誌
中文關鍵詞:電磁場組織工程幾丁聚醣骨母細胞
英文關鍵詞:electromagnetic fieldstissue engineeringchitosanosteoblast
論文中文摘要:電磁場(electromagnetic field, EMF)用在骨頭治療等已經有很好的成果,而用於臨床治療骨質疏鬆、骨不癒合也有很好的療效。然而超過1公分以上的骨缺陷以電磁場治療無法有很好的效果。目前,許多研究以組織工程的方式,將具有生物相容性多孔的支架植入骨缺陷處,用以促進細胞生長及減少骨缺陷修復的時間。
本研究將探討骨母細胞生長在幾丁聚醣支架及幾丁聚醣薄膜上經過電磁場刺激的變化。將實驗分為控制組及實驗組,實驗組部份每天連續以2小時的電磁場(18-30 Gauss, 75Hz, impulse width 1.3 ms, induced electrical amplitude 3.5±1mv)刺激,連續21天,在第1、2、3週測定其細胞活性(MTT assay)、鹼性磷酸酶(ALP)、蛋白質(Total protein)、鈣質含量(Calcium)、DNA含量;並且以螢光染色(Fluorescent stain)、電子式顯微鏡(SEM)觀察骨母細胞生長形態,並以鈣質染色(Calcium stain)觀察鈣質的產生。而材料方面,實驗組部分同樣以電磁場每天2小時,連續21天,並對其壓縮、降解及接觸角作測試。
從實驗結果發現,經過電磁場刺激後,細胞活性、DNA、鈣質都有明顯的增加,而鹼性磷酸酶在第3週經過電磁場刺激的實驗組則是下降。從電子式顯微鏡及螢光染色可以發現細胞在幾丁聚醣支架上及薄膜上生長的情形(呈現多角形及紡錘體)。而在材料上,經過電磁場刺激後,其壓縮、降解及接觸角跟控制組比較並無太大的差別,説明電磁場並不會影響幾丁聚醣在物理上的性質。
由結果得知,電磁場可以促進骨母細胞培養在幾丁聚醣上的增殖及分化情況。為了更近一步模擬人體的骨組織的環境,因此未來仍需在不同細胞或動物實驗,探討電磁場與幾丁聚醣的影響。
論文英文摘要:Electromagnetic Field (EMF) is shown to be able to accelerate bone healing and improve on osteoporosis. However the healing effect of EMF on large size bone fracture(>1cm) is not significant. Tissue engineering uses biocompatible porous scaffolds to fill up large defects, promote cell in-growth and can help reduce bone fracture healing time.
In this study we will explore effects of EMF on cells growing on tissue engineering scaffolds. We cultivated osteoblast (7F2) on chitosan scaffold and chitosan film, and subjected the cells to EMF (Intensity 18-30 Gauss, 75Hz, impulse width 1.3 ms, induced electrical amplitude 3.5mv) for 2 hours each day for three weeks. On week 1、week 2 and week 3, tested cell viability(MTT assay), alkaline phosphatase activity(ALP), total protein, calcium content, DNA content, cell morphology (SEM、fluorescent stain) and calcium stain. For chitosan we subjected material to EMF for 2 hours for three weeks and examined its compression strength, degradation rate and contact angle.
We observed increased cell viability, proliferation, calcium deposition and DNA content in cells exposed to EMF; ALP activity decreased by week 3 in cells exposed to EMF. From SEM and fluorescent stain pictures, we observed osteoblast typical and cell morphology on chitosan scaffold(Polygonal and Spindle-like shape) after EMF exposure. For chitosan, we found there is no difference in compression strength, degradation rate and contact angle after EMF exposure.
In conclusion, our results show EMF promote osteoblast proliferation and differentiation on chitosan scaffold. For mimic the environment of human bone tissue, it is necessary to do in vivo tests in the future for investigating the effect of EMF on tissue engineering.
論文目次:中文摘要 i
英文摘要 iii
誌謝 v
目錄 vi
表目錄 ix
圖目錄 x
第一章 緒論 1
1.1 幾丁聚醣的介紹 1
1.2.1 幾丁聚醣的結構 1
1.2.2 幾丁聚醣性質 3
1.2.3 幾丁聚醣在生醫的應用 4
1.2 電磁場 7
1.2.1 電磁場簡介 7
1.2.2 電磁場的生醫應用 10
1.2.3 電磁場的作用機制 12
1.3 骨組織 13
1.3.1 骨結構 13
1.3.2 骨母細胞 15
1.3.3 骨骼重塑 16
1.4 研究目的 18
第二章 實驗方法、藥品與儀器 19
2.1 實驗藥品 19
2.1.1 骨母細胞來源 19
2.1.2 細胞培養用藥品 19
2.1.3 實驗藥品 21
2.2 實驗儀器 24
2.3 實驗方法 25
2.3.1 電磁場產生器 25
2.3.2 幾丁聚醣薄膜及支架的製備 26
2.3.3 幾丁聚醣物理性質測試 28
2.3.3.1 壓縮測試(Compression) 28
2.3.3.2 接觸角(Contact angle) 28
2.3.3.3 降解測試(Degradation) 29
2.3.4 骨母細胞在幾丁聚醣上活性測試 30
2.3.4.1 細胞接種 30
2.3.4.2 鹼性磷酸酶活性(ALP) 31
2.3.4.3 蛋白質(Total protein) 31
2.3.4.4 細胞活性(MTT assay) 32
2.3.4.5 DNA含量 32
2.3.4.6 鈣質含量(Calcium) 32
2.3.4.7 骨母細胞培養在幾丁聚醣支架上3週壓縮測試 33
2.3.4.8 電子式顯微鏡(SEM) 33
2.3.4.9 螢光染色(Fluorescent stain) 33
2.3.4.10 鈣質染色(Calcium stain) 34
2.3.4.11 統計分析 34
第三章 結果與討論 35
3.1 材料測試結果 35
3.1.1 幾丁聚醣外觀 35
3.1.2 以SEM觀察幾丁聚醣支架 36
3.1.3 壓縮測試(Compression) 37
3.1.4 接觸角測試(Contact angle) 38
3.1.5 降解測試(Degradation) 39
3.2 骨母細胞在幾丁聚醣上活性測試結果 40
3.2.1 細胞活性(MTT assay) 40
3.2.2 DNA含量 41
3.2.3 以SEM觀察骨母細胞培養在幾丁聚醣支架上情形 42
3.2.4 螢光染色(Fluorescent stain) 46
3.3.5 鹼性磷酸酶活性(ALP) 50
3.3.6 鈣質染色(Calcium stain) 51
3.3.7 鈣質含量(Calcium content) 55
3.3.8 骨母細胞培養在幾丁聚醣支架上3週壓縮測試 56
第五章 討論 57
第六章 結論 60
參考文獻 61
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論文全文使用權限:同意授權於2010-08-21起公開