臺灣博碩士論文加值系統

利用靜電紡絲技術將複合材料製備成細胞支架，可提供類細胞外間質的環境以及較大的表面積和孔洞率，並使支架具有多種功能，可以提升其在組織工程應用。本研究目的在提升聚鄰甲氧基苯胺 (POMA) 靜電紡絲之生物相容性，並探討其在組織工程之應用。本研究所合成 POMA 的重量平均分子量為 45880 Dalton，在電紡溶液中混合 20 分鐘，黏度快速上升至 71.9 cP；5% (w/v) 為電紡出奈米纖維之最佳濃度。將 5 % (w/v) POMA 與 17 % w/v 明膠以 70:30 體積比混合成功製備出 POMA/明膠 (PG) 電紡絲；以 5% (w/v) 檸檬酸作為摻雜劑可製備出具有導電率之電紡絲 (PGC)。POMA、PG 及 PGC 電紡絲的直徑分別為 356±58、306±55 及 269±40 nm。POMA 表面接觸角為 109±2o；而 PG 及 PGC 則不具接觸角，將其交聯後 PGE 及 PGCE 的接觸角提升為 28±3o 及 30±3o。PGE 及 PGCE 的交聯程度分別為 64.23±7.41 %、74.11±1.44 %；交聯後材料的最大熱失重溫度比交聯前高出 15-20 oC。循環伏安法分析發現 POMA、PGE 及 PGCE 電紡絲具有氧化還原能力；但只有添加摻雜劑的 PGC 及 PGCE 組別能夠量測到導電率，分別為 3.31±0.19 x 10-5、5.85±0.49 x 10-6 S/cm。相較於未摻雜的 PG，PGC 中 POMA 的苯環與醌環吸收峰由 316、623 nm 紅移至 424、760 nm，進一步驗證檸檬酸摻雜可使 POMA 由鹼式中間氧化態轉變成鹽式中間氧化態。POMA 不具降解性，而 PGE 及 PGCE 則具生物可降解性，纖維型態分別能維持 1、8 天。人類脂肪幹細胞 (hASCs) 及大鼠心肌原細胞 (H9c2) 貼附及生長於 PGE 及 PGCE 的情形均較 POMA 為佳。進一步施加電場刺激，可使 hASCs 細胞長度增加並且呈現兩極化生長，並提高 H9c2 細胞分化成肌小管的能力。本研究證實 POMA 混合明膠並摻雜檸檬酸的電紡絲可使其生物相容性大幅提升，未來可應用於組織工程。

Composite scaffold prepared by electrospinning technology can provide extracellular matrix-like environment, large surface area, high porosity, and multifunctionality, thus increases their application in tissue engineering. The purpose of this study was to enhance the biocompatibility of electrospun poly(o-methoxyaniline) (POMA) fibers and to explore its potential for tissue engineering application. Average molecular weight of POMA synthesized in this study was 45880 Dalton. Viscosity of the solution for electrospinning increased rapidly to 71.9 cP within 20-min mixing. The optimal concentration of POMA for electrospinning nanofibers was 5 % (w/v). POMA/gelatin (PG) electrospun fibers were successfully prepared using a mix of 5 % (w/v) POMA and 17 % (w/v) gelatin in a volume ratio of 70:30. Eletrospinning mats (PGC) with conductivity was made when doped with 5 % (w/v) citric acid. The average diameters of POMA, PG, and PGC nanofibers were 356±58, 306±55, and 269±40 nm, respectively. Contact angle of POMA was 109±2o, and those of PG and PGC were 0o. After crosslinking, contact angles of PGE and PGCE were 28±3o, and 30±3o, respectively. The degrees of crosslinking for PGE and PGCE were 64.23±7.41 % and 74.11±1.44 %, respectively. The maximal temperature of weight loss increased 15-20 oC after crosslinking. Cyclic voltammetric analysis found that POMA, PGE and PGCE had redox capability. Only doped PGC and PGCE had measured conductivity of 3.31±0.19 x 10-5 and 5.85±0.49 x 10-6 S/cm, respectively. Compared to undoped PG, the absorption wavelengths of 316 and 623 nm corresponding to benzene ring and the quinone ring of POMA were red-shifted to 424 and 760 nm in PGC, which further validated citric acid doping could change the state of POMA from emeraldine base to emeraldine salt. POMA nanofibers were non-biodegradable. PGE and PGCE nanofibers were biodegradable where fibrous morphology maintained for 1 and 8 days. Human adipose stem cells (hASCs) and rat cardiac myoblast (H9c2) showed improved attachment and proliferation on PGE and PGCE compared to POMA nanofibers. Furthermore, applied electric field could increase cell length as well as the cell polarization of hASCs, and enhance the differentiation ability of H9c2 cells into myotubes. In summary, citric acid doped nanofibers of POMA blended with gelatin were found to exhibit improved biocompatibility and could be applied in tissue engineering in the future.

摘要 I
Abstract II
致謝 IV
目錄 V
圖索引 VIII
表索引 X
縮寫表 XI
第一章 緒論 1
1.1. 前言 1
1.2. 理論基礎 2
1.2.1. 靜電紡絲 2
1.2.2. 導電高分子 4
1.2.3. 聚苯胺 5
1.2.3.1. 聚苯胺的合成 5
1.2.3.2. 聚苯胺氧化還原狀態 6
1.2.3.3. 聚苯胺的摻雜劑 7
1.2.4. 明膠 8
1.2.5. 電訊號在組織再生及細胞刺激之探討 9
1.3. 文獻回顧 11
1.3.1. 電紡絲支架於組織工程之應用 11
1.3.2. 靜電紡絲製備導電高分子纖維及應用 13
1.3.3. 導電高分子電紡絲支架於組織工程之應用 14
1.4. 研究動機與目的 15
第二章 材料與方法 17
2.1. 研究架構 17
2.2. 靜電紡絲奈米纖維製備 19
2.2.1. 試劑配置 19
2.2.1.1. 氯化鈣/鹽酸溶劑配製 19
2.2.1.2. 明膠溶液配製 19
2.2.2. 聚鄰甲氧基苯胺合成 19
2.2.3. 靜電紡絲 21
2.2.3.1. POMA 21
2.2.3.2. POMA-明膠電紡絲 (PG) 製備 21
2.2.3.3. POMA/明膠/檸檬酸共混電紡絲 (PGC) 製備 22
2.2.4. 電紡絲之 EDC 交聯 22
2.3. 高分子溶液黏度分析 23
2.4. 電紡絲特性分析 24
2.4.1. 掃描式電子顯微鏡 24
2.4.1.1. 型號 S-4100 FE-SEM 操作步驟 24
2.4.1.2. 型號 JSM-7600F FE-SEM 操作步驟 24
2.4.1.3. 電紡纖維直徑分析 25
2.4.2. 材料接觸角分析 25
2.4.3. 傅立葉轉換紅外線光譜分析 26
2.4.4. 紫外光-可見光光譜儀分析 27
2.4.5. 電化學分析 27
2.4.6. 四點探針導電率量測 28
2.4.7. 熱重分析法 29
2.4.8. 交聯度測試 29
2.4.9. 降解試驗 30
2.5. 體外電刺激系統 31
2.6. 細胞培養 32
2.6.1. 細胞培養相關試劑配製 32
2.6.1.1. 人類脂肪幹細胞 α-MEM 培養基 32
2.6.1.2. 大鼠心肌原細胞 DMEM 培養基 33
2.6.1.3. 磷酸鹽緩衝溶液 34
2.6.1.4. 胰蛋白酶-EDTA 34
2.6.1.5. Trypan blue 染劑 34
2.6.2. 細胞來源 35
2.6.2.1. 人類脂肪幹細胞 35
2.6.2.2. 大鼠心肌原細胞 35
2.6.3. 細胞培養技術 36
2.6.3.1. 細胞繼代培養 36
2.6.3.2. 細胞凍存 37
2.6.3.3. 細胞解凍 37
2.6.3.4. 細胞計數 38
2.7. 細胞生長於電紡絲纖維之型態觀察 38
2.7.1. 掃描式電子顯微鏡觀察 38
2.7.2. FDA/PI 螢光雙重染色觀察 39
2.7.3. Rhodamine-phalloidin/Hoechst 33258 雙重染色觀察 40
2.8. 細胞長度分析 41
2.9. 細胞增生試驗 41
2.10. 大鼠心肌原細胞肌小管分化誘導 42
2.10.1. 大鼠心肌原細胞肌小管分化液配製 42
2.10.2. 大鼠心肌原細胞肌小管誘導分化之步驟 43
2.10.3. 肌球蛋白重鏈螢光染色 43
2.10.4. 肌小管影像分析 43
2.11. 統計分析 44
第三章 結果 45
3.1. 電紡溶液黏度分析 45
3.2. POMA 電紡絲的製備及 SEM 型態觀察 47
3.2.1. POMA 電紡絲之 SEM 影像 47
3.2.2. 明膠電紡絲 SEM 影像 50
3.2.3. POMA/明膠電紡絲 52
3.2.4. POMA/明膠/檸檬酸電紡絲 55
3.3. 電紡絲之交聯與交聯後分析 59
3.4. 接觸角分析 61
3.5. 熱重分析 61
3.6. 傅立葉轉換紅外線光譜分析 66
3.7. 紫外光-可見光吸收光譜分析 70
3.8. 電化學及導電率分析 71
3.9. 電紡絲之體外降解評估 73
3.10. 電紡絲與 H9c2 心肌原細胞之生物相容性評估 75
3.10.1. 細胞型態評估 75
3.10.2. 細胞增生評估 79
3.10.3. 細胞骨架肌動蛋白結構評估 81
3.10.4. H9c2 肌小管分化評估 83
3.11. 電紡絲與人類脂肪間葉幹細胞之生物相容性評估 88
3.11.1. 細胞型態評估 88
3.11.2. 細胞增生評估 92
3.11.3. 細胞骨架肌動蛋白結構評估 94
第四章 討論 96
4.1. 高分子溶液黏度對電紡型態之影響 96
4.2. 電紡絲製備與纖維型態之探討 97
4.3. 檸檬酸摻雜與電紡絲導電率之探討 98
4.4. 電紡絲交聯度與降解速率探討 101
4.5. 電紡絲生物相容性探討 103
4.6. 電刺激及電紡絲誘導肌小管分化能力之探討 104
第五章 結論與未來展望 106
參考文獻 108
附錄表 118
附錄圖 122


圖 1-1. 靜電紡絲儀器架設簡圖 2
圖 1-2. 泰勒圓錐形成示意圖 3
圖 1-3. 聚苯胺結構通式 6
圖 1-4. 聚苯胺氧化還原反應 7
圖 1-5. 檸檬酸化學結構 8
圖 1-6. 檸檬酸與聚苯胺反應式 8
圖 2-1. 研究架構 18
圖 2-2. 鄰甲氧基苯胺結構式 20
圖 2-3. 聚鄰甲氧基苯胺結構式 20
圖 2-4. 接觸角測量示意圖 26
圖 2-5. 循環伏安法原理示意圖 28
圖 2-6. TNBS 與自由胺基之反應 30
圖 2-7. 體外電刺激系統裝置示意圖 32
圖 2-8. hASCs 放大 100 倍之細胞型態 35
圖 2-9. H9c2 細胞放大 100 倍之細胞型態 36
圖 2-10. Resazurin 轉變成 Resorufin 之化學反應 42
圖 3-1. POMA、Gelatin、PG、PGC 電紡溶液之黏度分析 46
圖 3-2. POMA 電紡絲之 SEM 影像 49
圖 3-3. 明膠電紡絲之 SEM 影像 51
圖 3-4. PG電紡絲 SEM 影像 53
圖 3-5. PG 在不同電壓或不同電紡距離下之纖維直徑變化 54
圖 3-6. PGC 在不同電紡參數下之 SEM 影像 56
圖 3-7. PGC 在不同溶液推進速率或不同電紡距離下之纖維直徑變化 58
圖 3-8. PG、PGC 電紡絲以 EDC 交聯後之 SEM 影像 59
圖 3-9. PGE 及 PGCE 的交聯程度 60
圖 3-10. TGA 分析電紡絲之熱重分析曲線 64
圖 3-11. EDC 交聯前後之熱重分析曲線比較 65
圖 3-12. POMA、PG PGC 之 FTIR 圖譜 67
圖 3-13. 電紡絲在 EDC 交聯前後之比較 68
圖 3-14. POMA、PG、PGC 之 UV-Vis 吸收光譜 70
圖 3-15. POMA、PGE 與 PGCE 之電化學活性比較 72
圖 3-16. POMA、PGE 及 PGCE 電紡絲 74
圖 3-17. H9c2 細胞在 POMA 電紡絲上生長之 SEM 影像 76
圖 3-18. H9c2 在 PGE 電紡絲生長之 SEM 影像 76
圖 3-19. H9c2 在 PGCE 電紡絲生長之 SEM 影像 77
圖 3-20. H9c2 細胞在 POMA、PGE 及 PGCE 電紡絲之 FDA/PI 螢光染色影像 78
圖 3-21. 電刺激對 H9c2 細胞在 POMA、PGE 及 PGCE 電紡絲之增生影響 80
圖 3-22. 電刺激 H9c2 細胞在 POMA、PGE 及 PGCE 電紡絲之細胞骨架分佈影響 82
圖 3-23. H9c2 細胞在 POMA、PGE 及 PGCE 電紡絲上分化之 MHC 螢光染色影像 85
圖 3-24. H9c2 細胞在 POMA、PGE 及 PGCE 電紡絲上分化成肌小管之融合指數分析 86
圖 3-25. H9c2 細胞在 POMA、PGE 及 PGCE 電紡絲上分化成肌小管之長度分析 87
圖 3-26. hASCs 在 POMA 電紡絲生長之 SEM 影像 89
圖 3-27. hASCs 在 PGE 電紡絲生長之 SEM 影像 89
圖 3-28. hASCs 在 PGCE 電紡絲生長之 SEM 影像 90
圖 3-29. hASCs 細胞在 POMA、PGE 及 PGCE 電紡絲之 FDA/PI 螢光染色影像 91
圖 3-30. 電刺激對 hASCs 在 POMA、PGE 及 PGCE 電紡絲細胞長度影響 91
圖 3-31. 電刺激對 hASCs 細胞在 POMA、PGE 及 PGCE 電紡絲之增生影響 93
圖 3-32. 電刺激 hASCs 在 POMA、PGE 及 PGCE 電紡絲之細胞骨架分佈影響 95
附圖 A. EDC 交聯明膠機制 122
附圖 B. 體外電刺激系統架構示意圖 123

表 2-1. 實驗材料組別縮寫 19
表 3-1. 電紡 POMA、PG、PGC 成纖維之最佳參數 48
表 3-2. 電紡 PG 調控參數 52
表 3-3. 電紡 PGC 調控參數及組別 55
表 3-4. POMA、PG、PGC、PGE 及 PGCE 電紡絲之纖維直徑 60
表 3-5. POMA、PG、PGC、PGE 及 PGCE 之 FTIR 特徵吸收峰值 69
附表 A. 儀器表 118
附表 B. 藥品資料表 120

[1]Ueda M, Tohnai I and Nakai H (2001) Tissue engineering research in oral implant surgery. Artificial Organs. 25(3): 164-171.
[2]Hutmacher D W (2000) Scaffolds in tissue engineering bone and cartilage. Biomaterials. 21(24): 2529-2543.
[3]Jun I, Jeong S and Shin H (2009) The stimulation of myoblast differentiation by electrically conductive sub-micron fibers. Biomaterials. 30(11): 2038-2047.
[4]Barnes C P, Sell S A, Boland E D, Simpson D G and Bowlin G L (2007) Nanofiber technology: Designing the next generation of tissue engineering scaffolds. Advanced Drug Delivery Reviews. 59(14): 1413-1433.
[5]Liang D, Hsiao B S and Chu B (2007) Functional electrospun nanofibrous scaffolds for biomedical applications. Advanced Drug Delivery Reviews. 59(14): 1392-1412.
[6]Liao S, Li B J, Ma Z W, Wei H, Chan C and Ramakrishna S (2006) Biomimetic electrospun nanofibers for tissue regeneration. Biomedical Materials. 1(3): R45-R53.
[7]Oren R, Sfez R, Korbakov N, Shabtai K, Cohen A, Erez H, Dormann A, Cohen H, Shappir J, Spira M E and Yitzchaik S (2004) Electrically conductive 2D-PAN-containing surfaces as a culturing substrate for neurons. Journal of Biomaterials Science-Polymer Edition. 15(11): 1355-1374.
[8]Gizdavic-Nikolaidis M, Ray S, Bennett J R, Easteal A J and Cooney R P (2010) Electrospun functionalized polyaniline copolymer-based nanofibers with potential application in tissue engineering. Macromolecular Bioscience. 10(12): 1424-1431.
[9]Cooley J F (1902) Apparatus for electrically dispersing fluids. US 692,631
[10]Morton W J (1902) Method of dispersing fluids. US 20120055673
[11]Formhals A (1934) Process and apparatus for preparing artificial threads. US 1,975,504
[12]Taylor G (1964) Disintegration of water drops in an electric field. Proceedings of the Royal Society of London A: Mathematical, Physical &; Engineering Sciences. 280(1382): 383-397.
[13]Badami A S, Kreke M R, Thompson M S, Riffle J S and Goldstein A S (2006) Effect of fiber diameter on spreading, proliferation, and differentiation of osteoblastic cells on electrospun poly(lactic acid) substrates. Biomaterials. 27(4): 596-606.
[14]Baji A, Mai Y W, Wong S C, Abtahi M and Chen P (2010) Electrospinning of polymer nanofibers: Effects on oriented morphology, structures and tensile properties. Composites Science and Technology. 70(5): 703-718.
[15]Geng X Y, Kwon O H and Jang J H (2005) Electrospinning of chitosan dissolved in concentrated acetic acid solution. Biomaterials. 26(27): 5427-5432.
[16]Homayoni H, Ravandi S A H and Valizadeh M (2009) Electrospinning of chitosan nanofibers: Processing optimization. Carbohydrate Polymers. 77(3): 656-661.
[17]Min B M, Lee S W, Lim J N, You Y, Lee T S, Kang P H and Park W H (2004) Chitin and chitosan nanofibers: electrospinning of chitin and deacetylation of chitin nanofibers. Polymer. 45(21): 7137-7142.
[18]Torres-Giner S, Ocio M J and Lagaron J M (2008) Development of active antimicrobial fiber based chitosan polysaccharide nanostructures using electrospinning. Engineering in Life Sciences. 8(3): 303-314.
[19]Bhattarai N, Edmondson D, Veiseh O, Matsen F A and Zhang M Q (2005) Electrospun chitosan-based nanofibers and their cellular compatibility. Biomaterials. 26(31): 6176-6184.
[20]Ohkawa K, Cha D I, Kim H, Nishida A and Yamamoto H (2004) Electrospinning of chitosan. Macromolecular Rapid Communications. 25(18): 1600-1605.
[21]林雅凡、余瑞琳、蔡蘊明、鄭淑芬 (2010) 導電塑膠-聚苯胺. 國立台灣大學普化教學組.
[22]Hideki Shirakawa, Edwin J. Louis, Alan G. MacDiarmid, Chiang C K and Heeger A J (1977) Synthesis of electrically conducting organic polymers: Halogen derivatives of polyacetylene, (CH)x. Journal of the Chemical Society, Chemical Communications. 4(16): 578-580.
[23]Feast (1986) Synthesis of conducting polymers. New York: Marcel Dekker.
[24]Hong S Y and Marnick D S (1992) Understanding the conformational stability and electronic structures of modified polymers based on polythiophene. Macromolecules. 25 (18): 4652-4657.
[25]Guimard N K, Gomez N and Schmidt C E (2007) Conducting polymers in biomedical engineering. Progress in Polymer Science. 32(8-9): 876-921.
[26]Wang B, Tang J and Wang F (1987) Electrochemical polymerization of aniline. Synthetic Metals. 18(1-3): 323-328.
[27]Gonçalves D, dos Santos J D S, Mattoso L H C, Karasz F E, Akcelrud L and Faria R M (1997) Poly (o-methoxy aniline): solubility, deprotonation-protonation process in solution and cast films. Synthetic Metals. 90(1): 5-11.
[28]Hall H K, Padias A B and Boone H W (2007) Condensation routes to polyaniline and its analogs. Journal of Polymer Science Part a-Polymer Chemistry. 45(21): 4751-4763.
[29]Albuquerque J E, Mattoso L H C, Balogh D T, Faria R M, Masters J G and MacDiarmid A G (2000) A simple method to estimate the oxidation state of polyanilines. Synthetic Metals. 113(1-2): 19-22.
[30]Hatchett D W, Josowicz M and Janata J (1999) Acid doping of polyaniline: Spectroscopic and electrochemical studies. The Journal of Physical Chemistry B. 103(50): 10992-10998.
[31]Mo Z L, Zhao Z L, Chen H, Niu G P and Shi H F (2009) Heterogeneous preparation of cellulose-polyaniline conductive composites with cellulose activated by acids and its electrical properties. Carbohydrate Polymers. 75(4): 660-664.
[32]Wang H J, Ji L W, Li D F and Wang J Y (2008) Characterization of nanostructure and cell compatibility of polyaniline films with different dopant acids. Journal of Physical Chemistry B. 112(9): 2671-2677.
[33]Ghadimi F, Safa K D, Massoumi B and Entezami A A (2002) Polyaniline doped with sulphosalicylic, salicylic and citric acid in solution and solid-state. Iranian Polymer Journal. 11(3): 159-166.
[34]Ballestar A, Yakuphanoglu F, Senkal B F, Munoz M and Farooq W A (2011) Electrical characterization with atomic force microscopy and low temperature transport properties of boric acid doped polyaniline with Fe3O4 nanoparticles composites. Optoelectronics and Advanced Materials-Rapid Communications. 5(1-2): 177-181.
[35]Shi R, Bi J L, Zhang Z Z, Zhu A C, Chen D F, Zhou X H, Zhang L Q and Tian W (2008) The effect of citric acid on the structural properties and cytotoxicity of the polyvinyl alcohol/starch films when molding at high temperature. Carbohydrate Polymers. 74(4): 763-770.
[36]Goodarzi A, Sahoo Y, Swihart M T and Prasad P N (2003) Aqueous ferrofluid of citric acid coated magnetite particles. MRS Proceedings. N6.6.1-N6.6.6.
[37]Edwards J V, Eggleston G, Yager D R, Cohen I K, Diegelmann R F and Bopp A F (2002) Design, preparation and assessment of citrate-linked monosaccharide cellulose conjugates with elastase-lowering activity. Carbohydrate Polymers. 50(3): 305-314.
[38]Jiang Q R, Reddy N and Yang Y Q (2010) Cytocompatible cross-linking of electrospun zein fibers for the development of water-stable tissue engineering scaffolds. Acta Biomaterialia. 6(10): 4042-4051.
[39]Li J X, He A H, Zheng J F and Han C C (2006) Gelatin and gelatin-hyaluronic acid nanofibrous membranes produced by electrospinning of their aqueous solutions. Biomacromolecules. 7(7): 2243-2247.
[40]Muyonga J H, Cole C G B and Duodu K G (2004) Extraction and physico-chemical characterisation of Nile perch (Lates niloticus) skin and bone gelatin. Food Hydrocolloids. 18(4): 581-592.
[41]Ghasemi-Mobarakeh L, Prabhakaran M P, Morshed M, Nasr-Esfahani M H and Ramakrishna S (2009) Electrical Stimulation of Nerve Cells Using Conductive Nanofibrous Scaffolds for Nerve Tissue Engineering. Tissue Engineering Part A. 15(11): 3605-3619.
[42]Shiyun M, Mahmoud R and Ze Z (2011) Electrical stimulation in tissue regeneration. Applied Biomedical Engineering.
[43]Chao P H G, Lu H H, Hung C T, Nicoll S B and Bulinski J C (2007) Effects of applied DC electric field on ligament fibroblast migration and wound healing. Connective Tissue Research. 48(4): 188-197.
[44]Balint R, Cassidy N J and Cartmell S H (2012) Electrical stimulation: A novel tool for tissue engineering. Tissue Engineering Part B Reviews.
[45]Sill T J and von Recum H A (2008) Electrospinning: applications in drug delivery and tissue engineering. Biomaterials. 29(13): 1989-2006.
[46]Li M Y, Mondrinos M J, Gandhi M R, Ko F K, Weiss A S and Lelkes P I (2005) Electrospun protein fibers as matrices for tissue engineering. Biomaterials. 26(30): 5999-6008.
[47]Pan H, Jiang H and Chen W (2006) Interaction of dermal fibroblasts with electrospun composite polymer scaffolds prepared from dextran and poly lactide-co-glycolide. Biomaterials. 27(17): 3209-3220.
[48]Li W J, Tuli R, Huang X, Laquerriere P and Tuan R S (2005) Multilineage differentiation of human mesenchymal stem cells in a three-dimensional nanofibrous scaffold. Biomaterials. 26(25): 5158-5166.
[49]Kang X H, Xie Y B, Powell H M, Lee L J, Belury M A, Lannutti J J and Kniss D A (2007) Adipogenesis of murine embryonic stem cells in a three-dimensional culture system using electrospun polymer scaffolds. Biomaterials. 28(3): 450-458.
[50]Rim N G, Kim S J, Shin Y M, Jun I, Lim D W, Park J H and Shin H (2012) Mussel-inspired surface modification of poly(l-lactide) electrospun fibers for modulation of osteogenic differentiation of human mesenchymal stem cells. Colloids and Surfaces B: Biointerfaces. 91(0): 189-197.
[51]Telemeco T A, Ayres C, Bowlin G L, Wnek G E, Boland E D, Cohen N, Baumgarten C M, Mathews J and Simpson D G (2005) Regulation of cellular infiltration into tissue engineering scaffolds composed of submicron diameter fibrils produced by electrospinning. Acta Biomaterialia. 1(4): 377-385.
[52]Stankus J J, Guan J J, Fujimoto K and Wagner W R (2006) Microintegrating smooth muscle cells into a biodegradable, elastomeric fiber matrix. Biomaterials. 27(5): 735-744.
[53]Ladd M R, Lee S J, Stitzel J D, Atala A and Yoo J J (2011) Co-electrospun dual scaffolding system with potential for muscle-tendon junction tissue engineering. Biomaterials. 32(6): 1549-1559.
[54]Pinto N J, Johnson A T, MacDiarmid A G, Mueller C H, Theofylaktos N, Robinson D C and Miranda F A (2003) Electrospun polyaniline/polyethylene oxide nanofiber field-effect transistor. Applied Physics Letters. 83(20): 4244-4246.
[55]Chronakis I S, Grapenson S and Jakob A (2006) Conductive polypyrrole nanofibers via electrospinning: Electrical and morphological properties. Polymer. 47(5): 1597-1603.
[56]Ju Y W, Park J H, Jung H R and Lee W J (2007) Electrochemical properties of polypyrrole/sulfonted SEBS composite nanofibers prepared by electrospinning. Electrochimica Acta. 52(14): 4841-4847.
[57]Li X F, Hao X F, Yu H B and Na H (2008) Fabrication of Polyacrylonitrile/polypyrrole (PAN/Ppy) composite nanofibres and nanospheres with core-shell structures by electrospinning. Materials Letters. 62(8-9): 1155-1158.
[58]Laforgue A and Robitaille L (2010) Production of conductive PEDOT nanofibers by the combination of electrospinning and vapor-phase polymerization. Macromolecules. 43(9): 4194-4200.
[59]Sundaray B, Choi A and Park Y W (2010) Highly conducting electrospun polyaniline-polyethylene oxide nanofibrous membranes filled with single-walled carbon nanotubes. Synthetic Metals. 160(9-10): 984-988.
[60]Li M Y, Guo Y, Wei Y, MacDiarmid A G and Lelkes P I (2006) Electrospinning polyaniline-contained gelatin nanofibers for tissue engineering applications. Biomaterials. 27(13): 2705-2715.
[61]Li W-T, Shie M-F, Dai C-F and Yeh J-M (2010) Electrospinning poly(o-methoxyaniline) nanofibers for tissue engineering applications. MEDICON 2010. 29 596-599.
[62]Prabhakaran M P, Ghasemi-Mobarakeh L, Jin G R and Ramakrishna S (2011) Electrospun conducting polymer nanofibers and electrical stimulation of nerve stem cells. Journal of Bioscience and Bioengineering. 112(5): 501-507.
[63]Macinnes Jr D and Funt B L (1988) Poly-o-methoxyaniline: A new soluble conducting polymer. Synthetic Metals. 25(3): 235-242.
[64]Shie M F, Li W T, Dai C F and Yeh J M (2009) In vitro Biocompatibility of Electrospinning Polyaniline Fibers. IFMBE Proceedings. 211-214.
[65]謝沐峰 (2009) 聚鄰甲氧基苯胺電紡絲於組織工程之應用. 中原大學生物醫學工程學系碩士學位論文.
[66]Liang H C, Chang W H, Liang H F, Lee M H and Sung H W (2004) Crosslinking structures of gelatin hydrogels crosslinked with genipin or a water-soluble carbodiimide. Journal of Applied Polymer Science. 91(6): 4017-4026.
[67]Shalel-Levanon S and Marmur A (2003) Validity and accuracy in evaluating surface tension of solids by additive approaches. Journal of Colloid and Interface Science. 262(2): 489-499.
[68]Tao L, Zhang G F, Zhou W B and Su Z G (2007) Determination of modification degree of succinylated gelatin by size exclusion chromatography coupled with multi angle laser light scatting. Chinese Journal of Analytical Chemistry. 35(1): 43-48.
[69]Sheu M T, Huang J C, Yeh G C and Ho H O (2001) Characterization of collagen gel solutions and collagen matrices for cell culture. Biomaterials. 22(13): 1713-1719.
[70]Zuk P A, Zhu M, Ashjian P, De Ugarte D A, Huang J I, Mizuno H, Alfonso Z C, Fraser J K, Benhaim P and Hedrick M H (2002) Human adipose tissue is a source of multipotent stem cells. Molecular Biology of the Cell. 13(12): 4279-4295.
[71]Fraser J K, Wulur I, Alfonso Z and Hedrick M H (2006) Fat tissue: an underappreciated source of stem cells for biotechnology. Trends in Biotechnology. 24(4): 150-154.
[72]Kimes B W and Brandt B L (1976) Properties of a clonal muscle cell line from rat heart. Experimental Cell Research. 98(2): 367-381.
[73]Hescheler J, Meyer R, Plant S, Krautwurst D, Rosenthal W and Schultz G (1991) Morphological, biochemical, and electrophysiological characterization of a clonal cell (H9c2) line from rat heart. Circulation Research. 69(6): 1476-1486.
[74]Chrzanowski T H, Crotty R D, Hubbard J G and Welch R P (1984) Applicability of the fluorescein diacetate method of detecting active bacteria in freshwater. Microbial Ecology. 10(2): 179-185.
[75]Moffat B D and Snell T W (1995) Rapid toxicity assessment using an in vivo enzyme test for brachionus plicatilis (Rotifera). Ecotoxicology and Environmental Safety. 30(1): 47-53.
[76]O'Brien J, Wilson I, Orton T and Pognan F (2000) Investigation of the Alamar Blue (resazurin) fluorescent dye for the assessment of mammalian cell cytotoxicity. European Journal of Biochemistry. 267(17): 5421-5426.
[77]Pagano M, Naviglio S, Spina A, Chiosi E, Castoria G, Romano M, Sorrentino A, Illiano F and Illiano G (2004) Differentiation of H9c2 cardiomyoblasts: The role of adenylate cyclase system. Journal of Cellular Physiology. 198(3): 408-416.
[78]Reneker D H and Yarin A L (2008) Electrospinning jets and polymer nanofibers. Polymer. 49(10): 2387-2425.
[79]Lee K H, Kim H Y, La Y M, Lee D R and Sung N H (2002) Influence of a mixing solvent with tetrahydrofuran and N,N-dimethylformamide on electrospun poly(vinyl chloride) nonwoven mats. Journal of Polymer Science Part B-Polymer Physics. 40(19): 2259-2268.
[80]Ji Y L, Wolfe P S, Rodriguez I A and Bowlin G L (2012) Preparation of chitin nanofibril/polycaprolactone nanocomposite from a nonaqueous medium suspension. Carbohydrate Polymers. 87(3): 2313-2319.
[81]Amarnath C A, Kim J, Kim K, Choi J and Sohn D (2008) Nanoflakes to nanorods and nanospheres transition of selenious acid doped polyaniline. Polymer. 49(2): 432-437.
[82]Ghasemi-Mobarakeh L, Prabhakaran M P, Morshed M, Nasr-Esfahani M-H and Ramakrishna S (2008) Electrospun poly(ε-caprolactone)/gelatin nanofibrous scaffolds for nerve tissue engineering. Biomaterials. 29(34): 4532-4539.
[83]Manandhar S, Vidhate S and D'Souza N (2009) Water soluble levan polysaccharide biopolymer electrospun fibers. Carbohydrate Polymers. 78(4): 794-798.
[84]Neo Y P, Ray S, Easteal A J, Nikolaidis M G and Quek S Y (2012) Influence of solution and processing parameters towards the fabrication of electrospun zein fibers with sub-micron diameter. Journal of Food Engineering. 109(4): 645-651.
[85]Ramakrishna S, Fujihara K, Teo W-E, Lim T-C and Ma Z (2005) An introduction to electrospinning and nanofibers. World Scientific.
[86]Lee J S, Choi K H, Do Ghim H, Kim S S, Chun D H, Kim H Y and Lyoo W S (2004) Role of molecular weight of atactic poly(vinyl alcohol) (PVA) in the structure and properties of PVA nanofabric prepared by electrospinning. Journal of Applied Polymer Science. 93(4): 1638-1646.
[87]Skotheim T A (1986) Handbook of conducting polymers. Marcel Dekker.
[88]Ayad M M and Zaki E A (2008) Doping of polyaniline films with organic sulfonic acids in aqueous media and the effect of water on these doped films. European Polymer Journal. 44(11): 3741-3747.
[89]Kobayashi T, Yoneyama H and Tamura H (1984) Electrochemical reactions concerned with electrochromism of polyaniline film-coated electrodes. Journal of Electroanalytical Chemistry and Interfacial Electrochemistry. 177(1–2): 281-291.
[90]Batich C D, Laitinen H A and Zhou H C (1990) Chromatic changes in polyaniline films. Journal of The Electrochemical Society. 137(3): 883-885.
[91]Zhang J J, Gu M M, Zheng T T and Zhu J J (2009) Synthesis of gelatin-stabilized gold nanoparticles and assembly of carboxylic single-walled carbon nanotubes/Au composites for cytosensing and drug uptake. Analytical Chemistry. 81(16): 6641-6648.
[92]Mukherjee P and Nandi A K (2010) Bimetallic Au-core-Ag-shell nanoparticles from interfacial redox process using poly(o-methoxyaniline). Journal of Colloid and Interface Science. 344(1): 30-36.
[93]Kim H S, Hobbs H L, Wang L, Rutten M J and Wamser C C (2009) Biocompatible composites of polyaniline nanofibers and collagen. Synthetic Metals. 159(13): 1313-1318.
[94]Dai C F, Weng C J, Yeh T C, Lai B C, Sung C Y, Wei Y, Chang K C and Yeh J M (2012) Preparation of electrospun electroactive POMA fiber mats. Polymer International. 61(2): 213-221.
[95]An J, Liu J, Zhou Y, Zhao H, Ma Y, Li M, Yu M and Li S (2012) A Polyaniline-grafted graphene hybrid with amide groups and its use in supercapacitors. The Journal of Physical Chemistry C. 3(P16): 19699-19708.
[96]Bhadra S, Khastgir D, Singha N K and Lee J H (2009) Progress in preparation, processing and applications of polyaniline. Progress in Polymer Science. 34(8): 783-810.
[97]Mattoso L H C, Faria R M, Bulhões L O S and MacDiarmid A G (1994) Synthesis, doping, and processing of high molecular weight poly(o-methoxyaniline). Journal of Polymer Science Part A: Polymer Chemistry. 32(11): 2147-2153.
[98]黃美鳳 (2005) 幾丁聚醣接枝半乳糖簇之材料性質及其肝靶向性研究. 中央大學.
[99]馬純媛 (1999) 以 Genipin 或 Carbodiimide 交聯生物組織材料的交聯結構與交聯性質探討. 中央大學.
[100]Zhang S, Huang Y Q, Yang X P, Mei F, Ma Q, Chen G Q, Ryu S and Deng X L (2009) Gelatin nanofibrous membrane fabricated by electrospinning of aqueous gelatin solution for guided tissue regeneration. Journal of Biomedical Materials Research Part A. 90A(3): 671-679.
[101]van Wachem P B, Beugeling T, Feijen J, Bantjes A, Detmers J P and van Aken W G (1985) Interaction of cultured human endothelial cells with polymeric surfaces of different wettabilities. Biomaterials. 6(6): 403-408.
[102]Gupta M C and Umare S S (1992) Studies on poly(o-methoxyaniline). Macromolecules. 25(1): 138-142.
[103]Saravana Sampath S and Babu B V (2006) Kinetic parameter estimation of gelatin waste by thermogravimetry. National Conference on Environmental Conservation. 573-579.
[104]LIN C L, KUO Y C and LIN T S (2005) Effects of dental implant length and bone quality on biomechanical responses in bone around implants: 3-D non-linear finite element analysis. Biomedical Engineering: Applications, Basis and Communications. 17(01): 44-49.
[105]Araujo P L B, Ferreira C R P C and Araujo F S (2011) Biodegradable conductive composites of poly(3-hydroxybutyrate) and polyaniline nanofibers: Preparation, characterization and radiolytic effects. Express Polymer Letters. 5(1): 12-22.
[106]Zareh E N, Moghadam P N, Azariyan E and Sharifian I (2011) Conductive and biodegradable polyaniline/starch blends and their composites with polystyrene. Iranian Polymer Journal. 20(4): 319-328.
[107]Alvarez-Barreto J F and Sikavitsas V I (2007) Improved mesenchymal stem cell seeding on RGD-modified poly(L-lactic acid) scaffolds using flow perfusion. Macromolecular Bioscience. 7(5): 579-588.
[108]Mohan N, Nair P D and Tabata Y (2009) A 3D biodegradable protein based matrix for cartilage tissue engineering and stem cell differentiation to cartilage. Journal of Materials Science-Materials in Medicine. 20(1): 49-60.
[109]Borriello A, Guarino V, Schiavo L, Alvarez-Perez M A and Ambrosio L (2011) Optimizing PANi doped electroactive substrates as patches for the regeneration of cardiac muscle. Journal of Materials Science-Materials in Medicine. 22(4): 1053-1062.
[110]Shi G X, Zhang Z and Rouabhia M (2008) The regulation of cell functions electrically using biodegradable polypyrrole-polylactide conductors. Biomaterials. 29(28): 3792-3798.
[111]Ghasemi-Mobarakeh L, Prabhakaran M P, Morshed M, Nasr-Esfahani M H, Baharvand H, Kiani S, Al-Deyab S S and Ramakrishna S (2011) Application of conductive polymers, scaffolds and electrical stimulation for nerve tissue engineering. Journal of Tissue Engineering and Regenerative Medicine. 5(4): e17-e35.
[112]Patel N and Poo M M (1982) Orientation of neurite growth by extracellular electric fields. Journal of Neuroscience. 2(4): 483-496.
[113]Kotwal A and Schmidt C E (2001) Electrical stimulation alters protein adsorption and nerve cell interactions with electrically conducting biomaterials. Biomaterials. 22(10): 1055-1064.
[114]Tandon N, Goh B, Marsano A, Chao P H, Montouri-Sorrentino C, Gimble J and Vunjak-Novakovic G (2009) Alignment and elongation of human adipose-derived stem cells in response to direct-current electrical stimulation. Conference of the IEEE Engineering in Medicine and Biology Society. 1(1): 6517-6521.
[115]Zhao M, Forrester J V and McCaig C D (1999) A small, physiological electric field orients cell division. Proceedings of the National Academy of Sciences of the United States of America. 96(9): 4942-4946.
[116]Sirivisoot S and Harrison B S (2011) Skeletal myotube formation enhanced by electrospun polyurethane carbon nanotube scaffolds. International Journal of Nanomedicine. 6(1): 2483-2497.
[117]Ku S H, Lee S H and Park C B (2012) Synergic effects of nanofiber alignment and electroactivity on myoblast differentiation. Biomaterials. 33(26): 6098-6104.
[118]Cooper A, Bhattarai N and Zhang M Q (2011) Fabrication and cellular compatibility of aligned chitosan-PCL fibers for nerve tissue regeneration. Carbohydrate Polymers. 85(1): 149-156.
[119]Jha B S, Colello R J, Bowman J R, Sell S A, Lee K D, Bigbee J W, Bowlin G L, Chow W N, Mathern B E and Simpson D G (2011) Two pole air gap electrospinning: Fabrication of highly aligned, three-dimensional scaffolds for nerve reconstruction. Acta Biomaterialia. 7(1): 203-215.
[120]Gupta D, Venugopal J, Prabhakaran M P, Dev V R G, Low S, Choon A T and Ramakrishna S (2009) Aligned and random nanofibrous substrate for the in vitro culture of Schwann cells for neural tissue engineering. Acta Biomaterialia. 5(7): 2560-2569.
[121]Ricotti L, Polini A, Genchi G G, Ciofani G, Iandolo D, Mattoli V, Menciassi A, Dario P and Pisignano D (2011) Nanostructured, highly aligned poly(hydroxy butyrate) electrospun fibers for differentiation of skeletal and cardiac muscle cells. IEEE Engineering in Medicine and Biology Society. 3597-3600.
[122]Charest J L, Garcia A J and King W P (2007) Myoblast alignment and differentiation on cell culture substrates with microscale topography and model chemistries. Biomaterials. 28(13): 2202-2210.