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    題名: 經不同條件製備之多孔纖維支架對PC12細胞增生,分化及相關基因表現之影響
    作者: 楊織萍
    貢獻者: 醫學科學研究所
    日期: 2010
    上傳時間: 2010-10-20 10:46:48 (UTC+8)
    摘要: 神經系統受損後，很難自我再生，治療後，受損組織間常形成一個無血管供應及支持性基質的間隙，導致神經組織功能性的修復不易。本研究利用纖維組成的3D高孔洞網狀結構薄膜環境模擬細胞外間質，並利用類神經細胞：PC12細胞株為研究模式。研究主要以聚左乳酸(PLLA)與聚丁烯琥珀酸己二酸共聚物(PBSA) 製成一系列仿細胞外間質的絲狀薄膜，並以PC12細胞株為對象，將PC12細胞株培養在不同薄膜上，來探討PC12細胞株在不同薄膜上的貼附、增生情形。結果顯示細PLLA對於細胞貼附與增生均具較佳，細胞增生部分甚至比TCPS高於2.3倍，然而這都是以細胞現行表現探討所營造的環境是否利於細胞生長，無法知道細胞長遠的表現為何，因此本研究再利用基因表現的方式來探討細胞潛力。結果得知， 細PBSA有助於PC12細胞神經軸突生長，以細胞基因表現來看，由代表神經軸突錐生長的gene：GAP43及代表神經軸突生長的gene：MAP2可知，細胞培養在細PBSA於GAP43與MAP2表現上比TCPS分別高出19倍及1.7倍，其他披覆r-PGA、collagen等的薄膜效果亦相若。總結，本研究結果推論細PLLA及細PBSA較相似於細胞間質的結構，故在細胞貼附、增生及神經軸突生長較2D好，其提供促進神經細胞間的交互作用及神經細胞生長之良好環境，此對於神經再生組織工程中提供一個極佳的方向。


    After the neuron system injured, it is hard to regenerate. There will be a gap that has no blood vessel and matrix. Therefore, the regeneration of neuron system is hard to do. I fabricate the membranes of 3D structure to mimic the extra cellular matrix, and use the quasi-neuron cell: PC12 cell lines to be the cell model for my research. PBSA and PLLA of polymers are fabricated the fibrous membranes to mimic extra cellular matrix, and seeded PC12 cell line on the differential membranes. Then, confer PC12 cell proliferation and adhesion. The results exhibit that the fine fibrous PLLA membranes improve cell adhesion and proliferation rather than TCPS to 2.3 folds. But it is the external expression of cell, and we do not know the internal expression of cell. Therefore, keep the gene expression to research the internal expression of cell. The result exhibit that the fine fibrous PLLA membranes improve the growth of neurite outgrowth cone and neurite outgrowth, that show from the GAP43 and MAP2 gene expression rather than TCPS to 19 and 17 folds. And the results of membranes are coated collagen and r-PGA are similar of non-coating membranes. Conclusion, the structure of fine fibrous PLLA and PBSA membranes are similar of the extra cellular matrix, so they improve PC12 cell proliferation, adhesion and the growth of neurite outgrowth rather than 2D structure (ex: TCPS). The fibrous PLLA and PBSA membranes can provide a good environment to improve the interaction and growth of neuron cells that provide a good approach for the nerve regeneration of tissue engineering.
    關聯: 92頁
    描述: 目 錄

    誌謝 I
    摘要 III
    Abstract V
    縮寫表 VII
    目錄 VIII
    第一章 緒論 1
    1-1 前言 1
    1-2 研究動機 3
    1-3 研究目的 4
    第二章 文獻回顧 5
    2-1組織工程 5
    2-2電氣紡絲 6
    2-3周邊神經系統傷害與修復 7
    2-4生物支架與細胞外間質 9
    第三章 材料與方法 13
    3-1 材料與試劑 13
    3-2 儀器設備 14
    3-3 研究方法 16
    3-3-1 材料物化性質分析 17
    3-3-2 材料滅菌 19
    3-4 實驗流程圖 20
    3-5 實驗步驟 21
    3-5-1 薄膜製備 21
    3-5-2細胞培養與薄膜檢測 23
    3-5-3薄膜上細胞基因表現分析 26
    第四章 結果與討論 31
    4-1 纖維薄膜型態及其物理特性 32
    4-1-1 纖維薄膜之製備 33
    4-1-2 纖維薄膜之細胞毒性測試 35
    4-1-3 纖維薄膜之親疏水性測定 37
    4-2 纖維薄膜之細胞貼附試驗 38
    4-3 纖維薄膜之細胞增生試驗 41
    4-4 纖維薄膜對細胞活性檢測劑呈色劑吸收之
    preliminary study 43
    4-5 細胞電穿孔基因轉殖及細胞基因轉殖後於薄膜上生長之觀察 45
    4-6 PC12細胞株培養於纖維薄膜上之基因表現 46
    第五章 結論 56
    第六章 未來研究方向 58
    參考文獻 59
    附錄 實驗數據與圖 62


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    30. D.H. Reneker, I. Chun, Nanometre diameter fibres of polymer, produced by electrospinning. Nanotechnology, 1996. 7(3): p. 216-223.
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    顯示於類別:[醫學科學研究所] 博碩士論文

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