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研究生： 王文甫
Wen-fu Wang
論文名稱： 設計與製作圓錐平板型生物反應器
Designed and Validated Cone and Plate Bioreactor
指導教授： 鍾志昂
Chih-Ang Chung
口試委員:
學位類別： 碩士
Master
系所名稱： 工學院 - 機械工程學系
Department of Mechanical Engineering
畢業學年度： 96
語文別： 中文
論文頁數： 89
中文關鍵詞： 生物反應器 、圓錐平板 、幹細胞 、組織工程 、剪應力
外文關鍵詞： Bioreactor, Tissue engineering, Cone and plate, stem cell, Shear stress
相關次數： 點閱：11  下載：0 
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    • 醫學的進步和發展往往和科技的進步有著不可分割的關係，近代發展的新領域-組織工程，就是希望可以於體外培養的方式，置換我們體內失去功能或是受損的器官及組織，其中生物反應器在組織工程中更是不可或缺的裝置，除了提供細胞基本生存所需的環境之外，還可模擬細胞生長時的環境，或是提供不同的機械刺激調控細胞的基因表現或是分化路徑。
    本研究是以圓錐平板黏度計的原理為範本，並依據許多學者數值計算的結果為基礎，設計且製作出，可於體外培養細胞並可針對所需定量提供剪應力刺激的生物反應器，最後再利用此生物反應器，並以老鼠骨髓幹細胞為對象分別提供0、0.8、1.2、1.6 Pa的剪應力刺激4小時，以探討老鼠骨髓幹細胞受剪應力後細胞形狀及增生速率的改變。結果發現，施加不同的剪應力刺激老鼠骨髓幹細胞，細胞的形狀和細胞增生速率都不隨著剪應力增加而和靜態有所不同。
    系統經過測試，證明此生物反應器為一穩定且可提供寬廣剪應力範圍之裝置，未來更可以利用此系統提供不同的刺激條件，以提供組織工程領域相關研究發展。

    The progress and development of medical science has an inseparable relation with technology. The new frontier in latest development - tissue engineering, is a technology using cell-scaffold constructs developed in vitro to replace injury or function losing organs and tissues. Bioreactors are an indispensable device for the development of tissue engineering. They provide not only basic living environments for cells, but also mock up the environments for functional tissues to form. Besides, bioreactors may also be used to apply various different mechanical stimuli to control cells’ gene expression or differential pathway.
    In this study, a bioreactor which can be used to culture cells in vitro and provide quantitative shear stress has been designed. The model of the design is based on the regulation of the cone and plate viscometer, and the stress values are computed using the experiential formula proposed in previous literature. The in-house bioreactor was used to provide four hours of shear stress in a day in 0, 0.8, 1.2, 1.6 Pa respectively to stimulate the rat bone marrow stem cells. Results show when different shear stresses were applied, the cell shape and proliferation rate were not changing obviously comparing to the case of no shear imposed.
    This study demonstrates that the bioreactor constructed in house can be a stable device providing broaden shear stresses. With this devices, further tests on the cell responses to the shear stimulation can be conducted for tissue engineering applications.

    中文摘要 英文摘要 誌謝 目錄.....................................................Ⅰ 表目錄...................................................Ⅳ 圖目錄...................................................Ⅴ 第一章 緒論............................................1 1.1 前言............................................1 1.1.1 組織工程........................................2 1.1.2 生物反應器......................................4 1.2 研究背景........................................5 1.2.1 機械刺激........................................5 1.2.2 剪應力生物反應器................................8 1.3 研究動機.............................................10 第二章 生物反應器設計與製作........................... 11 2.1 設計概念....................................... 11 2.2 剪應力產生原理及定常轉速剪應力計算............. 12 2.2.1 剪應力產生原理..................................13 2.2.2 定常轉速剪應力計算..............................14 2.3 培養室之設計....................................14 2.3.1 圓錐平板尺寸設計................................14 2.3.2 培養液性質量測..................................16 2.3.3 系統剪應力性質表現..............................16 2.4 培養環境系統設計................................17 2.4.1溫度控制...........................................17 2.4.2二氧化碳濃度控制...................................19 2.5 馬達驅動控制....................................20 2.6 機構設計........................................21 2.6.1 圓錐平板間距調整..................................22 2.6.2 支撐及密封機構...................................23 2.6.2.1 密封機構.......................................23 2.6.2.2 支撐機構.......................................25 第三章 實驗方法.........................................42 3.1 細胞來源.............................................42 3.2 玻片消毒與細胞種植...................................42 3.3 材料對細胞增生影響測試...............................43 3.4 系統操作步驟.........................................44 3.5 系統培養效能測試.....................................46 3.6 實驗條件及步驟.......................................48 3.7 馬達轉速.............................................50 3.8實驗結果..............................................50 3.8.1 細胞數量測定...................................51 3.8.2 細胞型態觀察...................................52 第四章 結果與討論.......................................62 4.1 0 Pa培養............................................62 4.2 0.8 Pa刺激..........................................63 4.3 1.2 Pa刺激..........................................64 4.4 1.6 Pa刺激..........................................65 第五章 結論.............................................81 參考文獻.................................................84 表目錄 表3.1實驗試藥及儀器......................................54 圖目錄 圖2.1a全系統示意圖.......................................26 圖2.1b全系統實體圖.......................................26 圖2.2圓錐平板原理示意圖..................................27 圖2.3a圓錐平板示意圖.....................................28 圖2.3b圓錐平板實體圖.....................................28 圖2.3c圓錐尺寸圖.........................................29 圖2.3d平板培養室尺寸圖...................................30 圖2.2e平板培養室外蓋尺寸圖...............................31 圖2.4培養液於37℃剪應力與剪應變率關係圖..................32 圖2.5系統剪應力性能圖....................................33 圖2.6a溫度控制系統原理示意圖.............................34 圖2.6b溫度控制系統實體圖.................................34 圖2.7a二氧化碳濃度控制示意圖.............................35 圖2.7b二氧化碳濃度控制實體圖.............................35 圖2.8a馬達機構簡圖.......................................36 圖2.8b馬達機構實體圖.....................................36 圖2.9圓錐平板間距調整示意圖..............................37 圖2.10密封套筒示意圖.....................................38 圖2.11馬達撐機構示意圖...................................39 圖2.12升降平台支撐架示意圖...............................40 圖2.12平板支撐架示意圖...................................41 圖3.1材料對細胞增生影響測試結果..........................56 圖3.2系統操作前準備工作..................................57 圖3.3系統操作步驟........................................58 圖3.4系統培養效能結果....................................59 圖3.5實驗流程圖..........................................60 圖3.6a 玻片記號示意圖....................................61 圖3.6b記號後之玻片圖.....................................61 圖4.1 0 Pa培養結果......................................67 圖4.2 0 Pa培養前細胞形狀................................68 圖4.3a 0 Pa培養控制組細胞形狀...........................69 圖4.3b 0 Pa培養系統內細胞形狀...........................69 圖4.4 0.8 Pa 培養結果...................................70 圖4.5 0.8 Pa 培養前細胞形狀.............................71 圖4.6a控制組細胞形狀.....................................72 圖4.6b 0.8 Pa刺激之細胞形狀..............................72 圖4.7 1.2 Pa培養結果....................................73 圖4.8 1.2 Pa 培養前細胞形狀.............................74 圖4.9a控制組細胞形狀.....................................75 圖4.9b 1.2 Pa刺激之細胞形狀.............................75 圖4.10 1.6 Pa培養結果...................................76 圖4.11 1.6 Pa 培養前細胞形狀............................77 圖4.12a控制組細胞形狀....................................78 圖4.12b 1.6 Pa刺激之細胞形狀............................78 圖4.13a控制組細胞形狀....................................79 圖4.13b 1.6 Pa刺激結束立即取出之細胞形狀................79 圖4.14刺激後立刻取出之細胞增生速率結果...................80

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