臺灣博碩士論文加值系統

導電高分子如聚苯胺、聚吡咯不易被加工成纖維，而同軸靜電紡絲可以製
備出具有殼核結構的纖維，可將不易被電紡的殼層材料與易被電紡的核層材料
組合在一起。本研究利用靜電紡絲技術成功製備聚鄰甲氧基苯胺
(Poly-(o-methoxyaniline), POMA) 為殼層，聚己內酯為核層的同軸電紡絲 PA / PC，並摻雜樟腦磺酸 (Camphorsulfonic acid, CPSA) 製備成 C-PA / PC 電紡絲。最後評估大鼠骨髓間葉幹細胞 (Mesenchymal stem cell, MSCs) 在此電紡絲上生長及神經分化能力。穿透式電子顯微鏡 (Transmission electron microscopy, TEM) 影像中可觀測到 PA / PC 與 C-PA / PC 電紡絲具有雙層結構的特性，其纖維直徑分別為 194±37 nm (核：177±6 nm；殼：14±3 nm) 及 199±31 nm (核：172±8 nm；殼：16±2 nm)。傅立葉轉換紅外線光譜分析 (Fourier transform infrared spectroscopy, FTIR) 與熱重分析儀 (Thermogravity analysis, TGA) 分析顯示同軸電紡絲含有 POMA 及 PCL 的特徵峰質與熱裂解溫度值 (Decomposition temperature, Td)。接觸角試驗顯示摻雜 CPSA 可增加電紡絲表面的親水性。循環伏安法 (Cyclic voltammeter, CV) 證實 PA / PC 及 C-PA / PC 電紡絲均保有 POMA 的電活性。四點探針分析發現摻雜 CPSA 的 C-PA / PC 電紡絲的導電率較 PA / PC 電紡絲高。掃描式電子顯微鏡 (Scanning electron microscopy, SEM) 觀察發現 C-PA / PC 及 PA / PC 電紡絲不具降解能力，長期浸泡在培養基中會使纖維膨脹。在巨觀與微觀的力學分析結果發現 PA / PC 電紡絲的延展性較 C-PA / PC 電紡絲良好。MTS 分析、螢光染色及 SEM 影像中均顯示此纖維對於 MSCs 具有良好的生物相容性，且可作為神經細胞分化的支架，早期神經分化標記 βⅢ-tubulin 表現以 C-PA / PC 電紡絲較佳，晚期神經分化標記 MAP-2 表現則以 PA / PC 電紡絲較佳。

Conductive polymers such as polyaniline and polypyrrole are known for their difficulty for fiber processing. With coaxial electrospinning, core-shell fibers may comprise of an electrospinnable core material and a non-electrospinnable shell material. Here, core–shell structured poly(ε-caprolactone) (PCL)-poly(o-methoxyaniline) (POMA) (PA/PC) nanofibers were successfully prepared by coaxial electrospinning technique. PA/PC fibers were doped with camphorsulfonic acid (CPSA) to form C-PA/PC nanofibers. Cell proliferation and neural differentiation of mesenchymal stem cells (MSCs) from rat bone marrow were studied on PA/PC and C-PA/PC nanofibers. Transmission Electron Microscopy (TEM) images showed that the diameters of PA/PC and C-PA/PC nanofibers were 194±37nm (core: 177±6 nm; shell: 14±3 nm) and 199±31 nm (core: 172±8 nm; shell: 16±2 nm), respectively. Fourier Transform Infrared (FTIR) spectroscopy and Thermogravity Analysis (TGA) showed that coaxial electrospun nanofibers had characteristic wave numbers and Decomposition temperature (Td) of POMA and PCL. Contact angle analysis revealed that CPSA doping enhanced surface hydrophilicity of PA/PC nanofibers. Cyclic voltammetry (CV) analysis demonstrated electroactivity of PA/PC and C-PA/PC nanofibers. Four point probe analysis found that C-PA/PC nanofibers had higher conductivity than PA/PC nanofibers. Scanning Electron Microscopy (SEM) images showed that both nanofibers were not biodegradable. The fibers had swollen appearance after long-term immersion in culture media. Mechanical analysis at macroscopic and microscopic level found that PA/PC nanofibers had better ductility than C-PA/PC nanofibers. MTS, fluorescence staining and SEM analyses exhibited improved attachment and proliferation of MSCs on both types of nanofibers. It is feasible to use core–shell nanofibers as a scaffold for neural differentiation of MSCs as demonstrated by immunostaining with neuronal cell markers. C-PA/PC nanofibers had higher expression on early neural differentiation marker-β III tubulin, whereas PA/PC nanofibers had higher expression on late neural differentiation marker-MAP-II.

摘要
Abstract
致謝
目錄
圖目錄
表目錄
縮寫表
第一章 緒論
1.1 前言
1.2 理論基礎
4.4 間葉幹細胞於電紡絲上之神經分化誘導
4.4.1 神經分化誘導測試
4.4.2 神經分化標記免疫螢光染色分析
第五章 討論
5.1 同軸靜電紡絲參數與纖維形態探討
5.2 靜電紡絲特性探討
5.3 大鼠骨髓間葉幹細胞於電紡絲貼附、增生與活性探討
5.4 大鼠骨髓間葉幹細胞於電紡絲神經分化之探討
第六章 結論與未來展望
參考文獻
附錄
A. 儀器表
B. 藥品表
附錄圖


圖目錄
圖 1-1 泰勒錐成形示意圖
圖 1-2 同軸裝置示意圖
圖 1-3 複合式泰勒錐成形示意圖
圖 1-4 聚苯胺結構通式
圖 1-5 聚苯胺氧化還原反應
圖 1-6 聚己內酯結構式
圖 1-7 維生素A酸結構式
圖 1-8 2-巰基乙醇結構式
圖 1-9 本論文研究架構圖
圖 3-1 聚鄰甲氧基苯胺結構式
圖 3-2鄰甲氧基苯胺單體結構式
圖 3-3 二氧化矽結構式
圖 3-4 同軸針組裝結構
圖 3-5 循環伏安法示意圖
圖 3-6 四點探針裝置與原理示意圖
圖 3-7 AFM裝置與原理示意圖
圖 3-8 骨髓MSCs分離步驟示意圖
圖 3-9 骨髓MSCs形態
圖 3-10 細胞生長曲線圖
圖 4-1 5 wt% POMA 及 6 wt% PCL 電紡絲 SEM影像圖
圖 4-2 SNF-R2 電紡絲 SEM 影像
圖 4-3 PA / PC與C-PA / PC 電紡絲 SEM影像圖
圖4-4 PA / PC、C-PA / PC TEM影像圖
圖4-5 POMA、PCL、PA / PC、C-PA / PC 電紡絲接觸角影像
圖4-6 POMA、PCL、CPSA、PA / PC與C-PA / PC FTIR圖譜
圖 4-7 POMA、PCL、CPSA、PA / PC 與 C-PA / PC 的 TGA 分析曲線
圖 4-8 POMA、C-PA、PA / PC 與 C-PA / PC 的循環伏安法分析曲線
圖 4-9 原子力顯微鏡之力學分析定量圖
圖 4-10 PCL 及 PA / PC電紡絲之應力-應變
圖 4-11 PA / PC、C-PA / PC 及 SNF-R2 電紡絲的膨潤降解 SEM 影像
圖 4-12 間葉幹細胞培養於 PA / PC、C-PA / PC 及 SNF-R2 電紡絲之增生情形
圖 4-13 間葉幹細胞於 PA / PC、C-PA / PC 及 SNF-R2 電紡絲之 SEM 影像
圖 4-14 間葉幹細胞於 PA / PC、C-PA / PC 及 SNF-R2 電紡絲之 FDA / PI MSCs 培養於電紡絲之 FDA/PI 螢光染色影像
圖 4-15 間葉幹細胞於 PA / PC、C-PA / PC 及 SNF-R2 電紡絲之細胞骨架螢光染色影像
圖 4-16 間葉幹細胞培養於 PA / PC、C-PA / PC 及 SNF-R2 電紡絲之 βⅢ-tubulin 免疫螢光染色影像
圖 4-17間葉幹細胞培養於 PA / PC、C-PA / PC 及 SNF-R2 電紡絲之MAP-2 免疫螢光染色影像
圖 4-18 間葉幹細胞培養於 PA / PC、C-PA / PC 及 SNF-R2 電紡絲之
附圖 A-1 不同神經誘導分化試劑濃度對MSCs的形態影響
附圖 A-2 MSCs於神經分化誘導分化試劑的形態變化
附圖 A-3 MSCs神經誘導分化第14天之MAP-2與βⅢ-Tubluin免疫螢光染色圖


表目錄
表 3-1 電紡絲組別縮寫表
表 3-2 Rhodamine-phalloidin / Hoechst 33258 螢光染劑配製表
表 3-3 神經誘導分化因子配方表
表 3-4 神經分化免疫螢光染色抗體種類與稀釋倍率
表 4-1 PA / PC、C-PA / PC 及 SNF-R2 電紡最佳參數表
表 4-2 POMA、PCL、CPSA、PA / PC 及 C-PA / PC 的 FTIR特徵峰值
表 4-3 PA / PC 與 C-PA / PC 電紡絲組成
表 4-4 POMA 摻雜前後、PA/PC 及 C-PA/PC 電紡絲導電率比較表
表 4-5 PA / PC、C-PA / PC 及 SNF-R2電紡絲膨潤降解纖維直徑變化表
表 5-1 5 wt% POMA單獨利用外針以不同推進速率電紡結果
表 5-2 6 wt% PCL 單獨利用內針以不同推進速率、不同溶液比例電紡結果
表 5-3 外針 5 wt% POMA、內針 6 wt% PCL 固定流速，不同溶劑比例電紡結果
表 5-4 間葉幹細胞培養於 PA / PC、C-PA / PC 及 SNF-R2 電紡絲之βⅢ-Tubulin 及 MAP-2 表現量化表

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