電紡絲製備褐藻酸鈉/聚己內酯之奈米複合纖維進行原位轉染

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姓名

胡哲誠(Zhe-chen Hu) 查詢紙本館藏

畢業系所

化學工程與材料工程學系

論文名稱

電紡絲製備褐藻酸鈉/聚己內酯之奈米複合纖維進行原位轉染
(Electrospun Alginate/Polycaprolactone Composite Nanofibers for in-situ Transfection)

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至系統瀏覽論文 ( 永不開放)

摘要(中)

褐藻酸鈉所製備纖維支架，其負電特性可用來吸附帶正電的聚乙烯亞胺/基因納米粒子，藉由增加纖維的用量，可以增進轉染的效果，然而褐藻酸鈉對於細胞生物相容性不佳。因此我們導入了具生物相容性的聚己內酯來進行混合紡絲，SEM照片與纖維螢光染色中證實纖維的比例可以依照需求來調整，接觸角與紅外光譜儀之測試中也顯示複合纖維之性質會隨組成比例不同而改變。將不同組成比例之複合纖維來進行粒子吸附後進行轉染，發現聚己內酯纖維比例的提高可以增加材料的生物相容性，但是粒子的吸附量也隨之下降。為兼具轉染效果與生物適合性，我們利用EDTA來降解褐藻酸鈉纖維，結果發現在培養初期褐藻酸鈉纖維結構仍可維持以用於吸附粒子，在細胞攝入粒子後褐藻酸鈉纖維會逐漸被降解，只留下聚己內酯纖維，使細胞在攝入基因後得以存活於合適的環境。最後，為了要臨床應用需求，我們透過調整鈣離子的濃度來改變褐藻酸鈉纖維的交聯程度，並於結果發現，降低交聯程度會使纖維的結構較不穩定而會逐漸自行降解，亦可達到兼具轉染效果與生物適合性的效果。這些結果皆顯示利用可降解釋的複合支架來進行送藥並提升生物活性將有利於組織工程的應用。

摘要(英)

To regulate in situ gene delivery from biomaterial scaffolds, electrospun alginate nanofibers were applied to adsorb DNA/polyethylene (PEI) complex. The transfection efficiency increased with increasing deposited nanofibers. However, alginate was not favor for cell adhesion. Therefore, biocompatible poly (ε-poly olactone) (PCL) nanofibers was coelectrospun with alginate to increase biocompatibility. The scanning electron microscopy and fluorescent dye staining results suggested that the definite fiber ratios could be controlled. In addition, contact angle and FT-IR results also indicated that the properties of composite fibers can be regulated by the fiber ratios. The in situ transfection results demonstrated that the incorporated PCL fibers improved biocompatibility; however, the transfection efficiency was reduced. To preserve both gene transfer ability and biocompatibility, EDTA was applied to remove calcium ions for loosening alginate fiber structure. This treatment may initially maintain alginate fibers for nanoparticle adsorption, but these alginate fibers were gradually degraded in days to create a more appropriate environment for cell survival. For clinical application, we tried to regulate calcium concentration during fiber crosslinking to control the stability of alginate fibers. Though decreasing the levels of crosslinking, alginate fibers were degraded with time, which promoted both transfection efficiency and biocompatibility. These results supported biodegradable composite scaffolds should be potential for drug delivery with excellent bioactivity, which should be beneficial for tissue engineering applications.

關鍵字(中)

★ 電紡絲
★ 奈米粒子
★ 降解性材料

關鍵字(英)

★ Electrospinning
★ Nanoparticles
★ Degradable materials

論文目次

摘要 I
Abstract II
致謝 III
目錄 IV
圖目錄 VI
第一章緒論 1
1-1 背景 1
1-2 實驗目的 3
第二章文獻回顧 5
2-1 組織工程 5
2-2 電紡絲於組織工程 7
2-3 電紡絲簡介 10
2-4 褐藻酸鈉 13
2-4-1 褐藻酸鈉之來源 13
2-4-2 褐藻酸鈉之性質 13
2-4-3 電紡絲製備褐藻酸鈉於組織工程應用 14
2-5 聚己內酯 16
2-5-1 聚己內酯之來源 16
2-5-2 聚己內酯之性質 16
2-5-3 電紡絲製備聚己內酯於組織工程應用 17
2-6 基因治療 18
2-6-1 生長因子 18
2-6-2 基因載體 19
2-7 聚乙烯亞胺 21
2-7-1 聚乙烯亞胺之來源 21
2-7-2 聚乙烯亞胺之性質 21
2-8 電紡絲於基因治療之應用 23
2-9 帶電物質的作用 28
2-9-1 作用原理 28
2-9-2 帶電物質於組織工程中的運用 28
2-10 降解性材料 30
2-10-1 降解性材料之性質 30
2-10-2 降解性材料之應用 30
第三章材料與方法 35
3-1 實驗藥品 35
3-2 實驗儀器 37
3-3 實驗方法 39
3-3-1 電紡絲溶液之配製 39
3-3-1-1 儲備溶液配製 39
3-3-1-2 Alginate/PEO 紡絲液配製 39
3-3-1-3 Polycaprolactone/PEO 紡絲液配製 39
3-3-2 纖維製備 40
3-3-3 掃描式電子顯微鏡影像分析(SEM) 40
3-3-4 纖維收集量量測 40
3-3-5 混合纖維製備 41
3-3-6 材料表面分析 41
3-3-6-1 接觸角(contact angle)測定 41
3-3-6-2 FT-IR 測定 41
3-3-7 纖維之螢光染色測定 41
3-3-8 粒子轉染實驗 42
3-3-9 DNA吸附定量 42
3-3-10 EDTA降解實驗 43
3-3-11 氯化鈣降解實驗 44
3-3-12 MTT assay 45
3-3-13 LDH assay 45
第四章結果與討論 47
4-1 粒子於褐藻酸鈉纖維上的吸附與轉染 47
4-2 褐藻酸鈉/聚己內酯複合纖維的製備與性質 53
4-2-1 纖維之收集量 53
4-2-2 複合纖維之收集 57
4-2-3 纖維之螢光染色測定 59
4-2-4 材料表面性質測定 61
4-3 複合纖維的生物適合性 66
4-4 粒子於複合纖維上之轉染 71
4-5 複合纖維降解實驗 76
4-6 複合纖維自行降解實驗 82
第五章結論 90
第六章參考資料 92

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指導教授

胡威文(Wei-wen Hu)

審核日期

2013-7-26

推文