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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 黃義侑(Yi-You Huang) | |
dc.contributor.author | Jia-Ni Lee | en |
dc.contributor.author | 李佳霓 | zh_TW |
dc.date.accessioned | 2021-06-16T08:04:01Z | - |
dc.date.available | 2017-07-29 | |
dc.date.copyright | 2014-07-29 | |
dc.date.issued | 2014 | |
dc.date.submitted | 2014-07-02 | |
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/57969 | - |
dc.description.abstract | 肌腱組織由緻密的膠原纖維束所組成,主要功能為負責關節活動過程中之力量傳遞。目前臨床之腱傷可透過自體移植(autografts)、異體移植(allografts)、異種移植(xenografts)或人工肌腱置換治療。然而,提供移植處產生併發症、異體移植引發免疫排斥與人工肌腱疲乏等潛在問題,均促使仿生肌腱修復基材之發展。
過往文獻指出肌腱經去細胞處理後將可保留大量的胞外基質與相關生長因子,有鑑於此,我們將電紡所得之蠶絲蛋白纖維束浸泡於肌腱胞外基質液中,後續凍乾處理得結構與微環境近似天然組織之蠶絲/肌腱胞外基質(SF/t-ECM)複合支架。 將材料與肌腱細胞(tenocytes)共同培養七天後,SF/t-ECM複合支架細胞增生約60 %並開始分泌膠原蛋白;SEM及Confocal則可觀察到細胞於SF/t-ECM複合支架表層及深部細胞成紡錘狀貼附良好;另一方面,HE染色顯示SF/t-ECM複合支架於培養十六天後細胞已可浸潤到底層纖維,同時間點下SF支架則未觀察到此現象;動物實驗的部份,材料植入後十六天HE及Masson’s Trichrome染色均顯示SF/t-ECM複合支架相較SF之組別有更多的纖維母細胞(fibroblasts)浸潤及膠原纖維(collagen fibers)生成。 本研究設計之SF/t-ECM複合支架,解決了現階段肌腱組織工程上細胞無法深部浸潤支架增生並分泌胞外基質來提供機械強度之問題,結果顯示此近似天然組織之仿生基材未來應於肌腱修復上將相當具有潛力。 | zh_TW |
dc.description.abstract | Tendons, which play an important role in force transmission, are composed of densely packed collagen fibers. When tendons are injured, surgical treatments are used to repair the damaged tendon by autografts, allografts, xenografts, or prosthetic devices. However, the risks of donor-site morbidity, transplant rejection, and injury recurrence usually limit long-term function recovery. There is a need to engineer biomaterial scaffolds that mimic the native tendon to overcome these problems.
Previous studies have shown a lot of tendon extracellular matrix components as well as growth factors are still retained in decellularized tissues. Hence, we developed the silk fibroin/tendon-derived extracellular matrix (SF/t-ECM) scaffolds that are to be morphologically, biochemically, and histologically similar to the ECM of native tendon tissues. The results indicate cell proliferation on the SF/t-ECM scaffolds was significantly higher than that on the silk fibroin (SF) control groups after 7 days of cell culture. Histology demonstrated cells infiltrated through the full thickness of the SF/t-ECM scaffolds. Moreover, in rat tendon repair model, the SF/t-ECM scaffolds exhibited more fibroblasts proliferation and collagen fibers synthesis as compared with the SF control groups after 16 days of scaffold implanted. This investigation demonstrate that SF/t-ECM scaffolds have potential application in tendon repair. | en |
dc.description.provenance | Made available in DSpace on 2021-06-16T08:04:01Z (GMT). No. of bitstreams: 1 ntu-103-R01548010-1.pdf: 2928114 bytes, checksum: 203e715a14691cba5dd71c35e0f353b3 (MD5) Previous issue date: 2014 | en |
dc.description.tableofcontents | 誌謝 I
摘要 II Abstract III 目錄 V 圖目錄 IX 表目錄 XI 第一章 緒論 1 1.1 肌腱的結構與功能 1 1.1.1腱傷之治療 2 1.2 肌腱組織工程 2 1.2.1細胞 (Cell) 2 1.2.2支架 (Scaffold) 3 1.2.3生長因子 (Growth factor) 4 1.3 靜電紡絲法 5 1.3.1靜電紡絲法於肌腱組織工程上之應用 5 1.4 蠶絲 7 1.5 細胞外基質 9 1.6 肌腱組織工程之近年發展 11 第二章 研究動機與目的 13 第三章 材料與方法 15 3.1 實驗藥品 15 3.2 實驗儀器 17 3.3 萃取蠶絲蛋白 18 3.3.1準備溶液 18 3.3.2蠶繭脫膠 18 3.3.3萃取步驟 18 3.4 萃取豬肌腱組織之胞外基質 18 3.4.1準備溶液 18 3.4.2萃取步驟 19 3.5 靜電紡絲 19 3.5.1電紡機之架設 19 3.5.2製備電紡溶液 20 3.6 製備蠶絲蛋白支架 20 3.7 製備蠶絲/肌腱胞外基質支架 20 3.7.1準備溶液 20 3.7.2製備步驟 20 3.8 Collagen含量測試 21 3.8.1準備溶液 21 3.8.2實驗步驟 21 3.9 GAG含量測試 22 3.9.1準備溶液 22 3.9.2實驗步驟 22 3.10 肌腱細胞之初代培養 23 3.10.1準備溶液 23 3.10.2分離培養肌腱細胞 24 3.11 材料滅菌及細胞培養 24 3.11.1材料滅菌 24 3.11.2細胞培養 24 3.12 掃描式電子顯微鏡(SEM)觀察支架表層細胞型態 24 3.12.1準備溶液 24 3.12.2固定並乾燥細胞 25 3.13 共軛焦顯微鏡(Confocal Microscope)觀察支架內部細胞型態 25 3.13.1準備溶液 25 3.13.2實驗步驟 25 3.14 MTT細胞存活率分析 25 3.14.1準備溶液 25 3.14.2實驗步驟 26 3.15 動物實驗 26 3.15.1實驗動物 26 3.15.2準備溶液 26 3.15.3實驗步驟 26 3.16 HE Stain評估組織病理表現 27 3.16.1準備溶液 27 3.16.2石蠟切片 27 3.16.3染色步驟 28 3.17 Masson’s Trichrome Stain評估膠原蛋白分泌量 28 3.17.1準備溶液 28 3.17.2石蠟切片 29 3.17.3染色步驟 29 3.18 統計分析 30 第四章 研究結果與討論 31 4.1 蠶絲蛋白(SF)支架 31 4.1.1蠶絲纖維束之角度分佈 31 4.1.2蠶絲纖維束之孔隙度分析 32 4.2 蠶絲/肌腱胞外基質支架(SF/t-ECM) 33 4.2.1滅菌前後SEM觀察變化 33 4.2.2滅菌前後Collagen流失程度評估 34 4.2.3滅菌前後GAG流失程度評估 34 4.2.4培養液中Collagen釋放速率評估 35 4.3 分離培養肌腱細胞(Tenocyte) 36 4.3.1 SEM觀察表層細胞型態 37 4.3.2 Confocal觀察內部細胞型態 37 4.3.3 MTT細胞增生率分析 39 4.3.4 Collagen細胞分泌量分析 40 4.3.5 HE Stain細胞浸潤性評估 41 4.4 體內動物實驗 42 4.4.1 HE stain組織病理學分析 42 4.4.2 Masson’s trichrome膠原蛋白分泌評估 45 第五章 結論 46 參考文獻 47 圖目錄 圖1- 1 肌腱之膠原纖維組成 1 圖1- 2 間葉幹細胞可分化之中胚層細胞譜系 3 圖1- 3 間葉幹細胞之肌腱分化相關因子 4 圖1- 4 電紡之滾輪收集法 5 圖1- 5 濕性電紡絲法(wet electrospinning) 6 圖1- 6 氣隙電紡絲法(airgap electrospun) 7 圖1- 7 蠶絲蛋白之多元型態應用 9 圖1- 8 細胞外基質(extracellular matrix, ECM) 10 圖1- 9 經去細胞後之胞外基質應用於肌腱修復 11 圖1- 10 以編織法所得之蠶絲-膠原蛋白複合支架 12 圖2- 1 實驗架構流程 14 圖4- 1 蠶絲纖維束之角度分佈 32 圖4- 2 SEM×1000蠶絲纖維束移除PEO前後之變化 32 圖4- 3 SEM觀察SF/t-ECM支架滅菌前後之變化 33 圖4- 4 滅菌前後之Collagen含量變化 34 圖4- 5 滅菌前後之GAG含量變化 35 圖4- 6 培養液中Collagen釋放速率評估 36 圖4- 7 分離培養之肌腱細胞 36 圖4- 8 SEM觀察表層細胞型態 37 圖4- 9 Confocal觀察內部細胞型態 38 圖4- 10 MTT細胞增生率分析 39 圖4- 11 Collagen分泌量分析 40 圖4- 12 HE Stain細胞浸潤性評估 41 圖4- 13 於大鼠髕肌腱受傷處植入材料縫合 42 圖4- 14 植入八天後的組織染色圖 43 圖4- 15 植入十六天後的組織染色圖 44 圖4- 16 Masson’s trichrome 膠原蛋白分泌評估 45 表目錄 表1- 1蠶絲與其他生醫材料之機械強度比較 8 表3- 1動物實驗分組 26 | |
dc.language.iso | zh-TW | |
dc.title | 蠶絲蛋白/肌腱胞外基質支架於肌腱修復上之應用 | zh_TW |
dc.title | Application of Silk Fibroin/Tendon-Derived Extracellular Matrix Scaffold for Tendon Repair | en |
dc.type | Thesis | |
dc.date.schoolyear | 102-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 鐘次文(Tze-Wen Chung),黃意真(Yi-Cheng Huang),林森祥(Sen-Shang Lin) | |
dc.subject.keyword | 肌腱組織工程,氣隙電紡絲法,蠶絲蛋白,細胞外基質,肌腱修復, | zh_TW |
dc.subject.keyword | Tendon tissue engineering,Air-gap electrospun,Silk fibroin,Extracellular matrix,Tendon repair, | en |
dc.relation.page | 54 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2014-07-02 | |
dc.contributor.author-college | 工學院 | zh_TW |
dc.contributor.author-dept | 醫學工程學研究所 | zh_TW |
顯示於系所單位: | 醫學工程學研究所 |
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