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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 林峰輝 | |
| dc.contributor.author | Chunching Li | en |
| dc.contributor.author | 李淳敬 | zh_TW |
| dc.date.accessioned | 2021-06-16T06:52:41Z | - |
| dc.date.available | 2024-12-31 | |
| dc.date.copyright | 2014-08-14 | |
| dc.date.issued | 2014 | |
| dc.date.submitted | 2014-07-22 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/57584 | - |
| dc.description.abstract | 相較於硬骨於體內之自然修復能力,自體修復之軟骨無法回復受損前的組 織功能。小範圍的軟骨缺損若無法即早修復,其將導致嚴重之關節炎疾病。然目 前臨床上之治療方式,皆無法根本性的修補軟骨缺損。
時至今日,軟骨組織工程之研究,仍未有適用於生物反應器之最佳組合。 明膠/玻尿酸/軟骨素三共聚物已於過去的研究展示其能維持軟骨細胞之型態。本 研究中,將使用間質性幹細胞做為軟骨組織工程之細胞來源,播種間質性幹細胞 於三共聚物支架,並結合誘導幹細胞軟骨分化之小分子藥物 Kartogenin ,作為 分化訊號。 於生物反應器培養 21 日後,本組合成功產生俱軟骨特性之構物。其軟骨相 關基因包含: Aggrecan 、︑ SOX9 以及 Col2a1 皆有顯著性之上昇,此外聚集之軟 骨性團塊可於掃描式電子顯微鏡下檢視,而其組織切片亦能發現類似軟骨陷窩之 結構。於免疫染色中,亦可檢視到軟骨蛋白聚醣 (aggrecan) 以及第二型膠原蛋白 (type II collagen) 表現。 相較於傳統上使用蛋白質誘發分化,使用小分子藥物於培養基消耗量大之 生物反應器為一較俱臨床可行性之選項。本研究以明膠/玻尿酸/軟骨素、 KGN 以及間質性幹細胞之組合提供了小分子藥物調控幹細胞之一範例。 | zh_TW |
| dc.description.abstract | Contrary to the capability of bone repair, the cartilage regeneration in body couldn’t recover the cartilage lesions to proper function, and the failure of small defect repairing could lead to severe osteoarthritis disease. However, current clinical treatment options are not satisfying to fully recover the lesions.
Till now, there is no optimized combination of scaffold and signal for cartilage tissue engineering in bioreactor system. Previously we have demonstrated tri- copolymer composed of gelatin, hyaluronic acid and chondroitin-6-sulfate can retain the phenotype of chondrocyte. In this study we demonstrated the combination of mesenchymal stem cells (MSCs) with tri-copolymer and Kartogenin (KGN) , a small molecule drug which promotes MSCs toward chondrogenetic differentiation. The combination successfully induced the chondrogenesis. After 21 days culture in homemade perfusion bioreactor system, the construct shows chondrogenetic characteristic in both genetic and morphology aspect. Aggrecan, Sox-9 and Col2a1 are all up-regulated, and the condensed cell mass are found in the porous scaffold structure. The lacunae-like structure were also found by Hematoxylin/Eosin staining. Since culturing in the bioreactor with cytokine or growth factor is impractical due to its need for great amount of medium, culturing with small molecule has become much more feasible. We expect that this research provides an example for small molecule regulated practice and thus provide a new combination for cartilage tissue engineering. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-16T06:52:41Z (GMT). No. of bitstreams: 1 ntu-103-R01548007-1.pdf: 22062717 bytes, checksum: 6dc87f8821b4c674fca0af8037eda876 (MD5) Previous issue date: 2014 | en |
| dc.description.tableofcontents | 摘 要 ........I
Abstract........II 目 錄 ............III 圖目錄..........VII 表目錄...........IX 縮寫表...........X 第一章 簡介 1-1 軟骨:結構、組成與功能.......1 1-1-1 關節軟骨:結構與組成........................................1 1-1-2 關節軟骨:代謝 .................................4 1-2 軟骨損傷與機轉:以分離性骨軟骨炎與骨關節炎為例...................................5 1-3 軟骨損傷之治療...............................8 1-3-1 注射式治療...................................................................8 1-3-2 鑽孔 .............................8 1-3-3 自體骨軟骨移植 .............................................................9 1-3-4 自體軟骨細胞移植 ........................................9 1-4 關節軟骨之組織工程方法...............................................10 1-4-1 支架 .......................................................................10 1-4-2 細胞 ...........................................................................11 1-4-3 訊號 ....................................................................................13 1-4-4 生物反應器...............................................13 1-5 研究目的.................................................15 第二章 理論基礎 2-1 關節軟骨組織工程支架:明膠/透明質酸/軟骨素共聚物..................16 2-1-1 由軟骨細胞外基質衍生之立體海綿狀支架 .................................16 2-1-2 明膠/透明質酸/軟骨素三共聚物.................................................17 2-2 關節軟骨組織工程:細胞-間質性幹細胞 .......................19 2-2-1 間質性幹細胞用於軟骨組織工程...........................................19 2-2-2 軟骨分化.........................................................................21 2-3 關節軟骨組織工程:訊號 - Kartogenin ..........................23 2-4 封閉式生物反應器............................................26 第三章 材料與方法 3-1 實驗架構................................................................27 3-2 實驗儀器.......................................................................28 3-3 實驗藥品.....................................................................................29 3-4 實驗方法...........................................................30 3-4-1 明膠/透明質酸/軟骨素三共聚物之製備 ..................................30 3-4-2 明膠/透明質酸/軟骨素三共聚物材料分析 .....................................31 3-4-2-1 以掃描式電子顯微鏡觀測材料微結構 ..................31 3-4-2-2 壓汞儀孔隙度分析......................................31 3-4-2-3 交聯度分析 ........................................31 3-4-2-4 傅氏紅外線吸收光譜...............................32 3-4-3 間質幹細胞..............................................33 3-4-3-1 間質幹細胞的分離與培養 ......................................33 3-4-3-2 細胞標記之測定..........................................................34 3-4-3-3 KGN分化效果檢測..............................34 3-4-4 細胞播種於明膠/透明質酸/軟骨素三共聚物及培養 ..................35 3-4-5 細胞支架複合體分析....................................35 3-4-5-1 細胞存活染色 ................................................35 3-4-5-2 硫化醣胺多醣定量分析....................................36 3-4-5-3 基因表現檢視 - 即時聚合酶鏈式反應................................37 3-4-5-4 組織學結構研究..................................................40 3-5 統計方法..............................................................41 第四章 結果 4-1 明膠/透明質酸/軟骨素三共聚物材料分析..................42 4-1-1 明膠/透明質酸/軟骨素三共聚物之微結構 ........................42 4-1-2 三共聚物交聯度量測....................................................45 4-1-3 三共聚物於交聯後之官能基分析......................45 4-2 幹細胞族群確立與 KGN 分化效果檢測 ............................47 4-2-1 幹細胞標記分析 ....................................47 4-2-2 軟骨形成 (Chondrogenesis) 檢視..................................47 4-2-2-1 於平面培養下幹細胞聚集檢視............................................47 4-2-2-2 於平面培養下之基因表現 ...................................49 4-3 幹細胞於明膠/透明質酸/軟骨素三共聚物支架之分化...................52 4-3-1 於三共聚物內培養下之細胞聚集.............................52 4-3-2 於三共聚物中培養之基因表現 ...................................52 4-4 三共聚物與幹細胞共構物於生物反應器之分化........................55 4-4-1 基因檢視:軟骨及肥大軟骨細胞相關基因......................55 4-4-2 以掃描式電子顯微鏡檢視細胞於三共聚內之型態...................59 4-4-3 三共聚物與細胞共構物之組織切片檢視..................................61 4-4-4 以免疫染色檢視軟骨特徵蛋白表現.......................................63 4-4-4 硫化醣胺多醣於細胞團塊之表現.............................................65 第五章 討論 5-1 軟骨缺損初期之修復及挑戰...................................................67 5-2 間質性幹細胞之軟骨形成作用與三共聚物材料.........................69 5-2-1 間質性幹細胞之軟骨形成作用 .................................................69 5-2-2 Kartogenin 作為軟骨分化訊號..................................69 5-2-3 軟骨形成作用與三共聚物材料 ...............................................70 5-3 三共聚物與細胞共構物之長期培養.........................71 5-3-1 間質性幹細胞之軟骨肥大特性 .......................................71 5-3-2 Kartogenin 與軟骨肥大............................................71 5-3-3 於生物反應器培養之結果.....................................73 5-3-3-1 基因表現層面 ..................................................73 5-3-3-2 組織學檢視 .......................................................................73 5-3-3-3 GAGs 之分泌 ....................................................................74 5-4 本研究之未來展望............................................75 5-4-1 動物模型之驗證 ........................................................................75 5-4-2 小規模軟骨損傷之修復.........................................................75 第六章 結論..........................................................77 參考文獻 ............................................................78 | |
| dc.language.iso | zh-TW | |
| dc.title | 明膠/玻尿酸/軟骨素三共聚物於間質性幹細胞之軟骨組織工程應用 | zh_TW |
| dc.title | Application of Gelatin/Hyaluronic Acid/ Chondroitin-6-Sulfate Tri-copolymer on MSCs Based Cartilage Tissue Engineering | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 102-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.coadvisor | 孫瑞昇 | |
| dc.contributor.oralexamcommittee | 楊禎明,徐善慧,張國基 | |
| dc.subject.keyword | 軟骨組織工,幹細胞,明膠/玻尿酸/軟骨素三共聚物, | zh_TW |
| dc.subject.keyword | Cartilage tissue engineering,mesenchymal stem cells,gelatin/hyaluronic acid/ chondroitin-6-sulfate tri-copolyemer, | en |
| dc.relation.page | 88 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2014-07-22 | |
| 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 | |
文件中的檔案：
| 檔案 | 大小 | 格式 | |
|---|---|---|---|
| ntu-103-1.pdf 目前未授權公開取用 | 21.55 MB | Adobe PDF |
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