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研究生:羅婉慈
研究生(外文):Wan-Tzu Lo
論文名稱:氧化幾丁聚醣/昆布胺酸水膠作為異體副甲狀腺移植免疫隔離之運用
論文名稱(外文):Oxidized Chitosan/Laminin Hydrogel for Allo-parathyroid Immunoisolation
指導教授:林峯輝
口試委員:張國基林建成郭士民楊凱強
口試日期:2013-06-21
學位類別:碩士
校院名稱:國立臺灣大學
系所名稱:醫學工程學研究所
學門:工程學門
學類:綜合工程學類
論文種類:學術論文
論文出版年:2013
畢業學年度:101
語文別:中文
論文頁數:47
中文關鍵詞:副甲狀腺副甲狀腺機能不足異體移植免疫隔離幾丁聚醣水膠微包覆
外文關鍵詞:parathyroid glandshypoparathyroidismallo-transplantationimmunoisolationchitosan-based hydrogelmicroencapsulation
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副甲狀腺機能不足症為目前難以完全治癒的內分泌疾病之一,在臨床上也並不建議在副甲狀腺異體移植後使用全身性免疫抑制的藥物,故使用具半通透性的膜來包覆副甲狀腺組織再進行異體移植,來達到植入組織在病人體內具有免疫隔離的效果。本研究使用氧化幾丁聚醣交聯昆布胺酸,將副甲狀腺組織塊外面包覆後滴入三聚磷酸鈉溶液中,利用離子吸附的方式在組織塊外圍成一層膜狀的水膠。在材料測試方面,氧化幾丁聚醣/昆布胺酸水膠在體外28天內可維持約80 %重量與型態,且具有良好的生物相容性,同時不具細胞毒性。以材料包覆組織後進行體外培養結果,組織外的膜狀水膠可使人類副甲狀腺素成功釋放至環境中,證實包覆材料並不會影響組織的分泌功能,並大致具有隨周圍環境的鈣離子濃度調整釋放量的趨勢,且在組織切片的觀察結果顯示包覆材料並不影響具分泌能力的主細胞的存活。在動物實驗方面雖有成功導致實驗動物具有副甲狀腺機能不足症,但在移植之後的組織對血鈣濃度的調節效果並不如預期,顯示本實驗模型還有待改進。

Permanent hypoparathyroidism is one of the endocrine disorders that are difficult to treat medically. Since hypoparathyroidism rarely is a life-threatening condition, systemic immunosuppression for recipients of allo-transplants are not justified. Coating the allo-transplanted tissue with a semipermeable membrane to protect the tissue from immune rejection is an alternative. In this study, oxidized chitosan cross-linked with laminin is prepared for microencapsulation of parathyroid tissue fragments. The parathyroid tissue particles with oxidized chitosan/laminin are dropped in sodium tripolyphophate solution. Tripolyphophate ion, the polyanion, would interact electrostatically with the cationic chitosan and become hydrogel to enclose the tissue particle. In the material analysis, oxidized chitosan/laminin hydrogel maintained about 80 % mass and it’s morphology in PBS until the 28th day. The material in vitro tests also showed that hydrogel had good biocompatibility and low cytotoxicity. The microencapsulation of parathyroid tissue in vitro study revealed that the hydrogel was freely permeable to the human parathyroid hormone and responded to different calcium concentration in the ex vivo test. The histological evaluation showed that the capsular hydrogel membrane were not affective to the chief cells which are the main secretory cell in the parathyroid glands. In animal study, hypocalcemia rats were induced successfully but the encapsulated tissue particles implanted were not functional in vivo. The result demonstrated that the animal model in this study must be improved.

口試委員審定書 i
誌謝 ii
中文摘要 iii
英文摘要 iv
目錄 v
圖目錄 ix
表目錄 xii
第一章 簡介 1
1.1 副甲狀腺(parathyroid) 1
1.2 副甲狀腺機能不足症(hypoparathyriodism) 3
1.3 副甲狀腺異體移植(parathyroid allotransplant) 4
1.4 免疫隔離(immunoisolation) 5
1.5 研究目的 8
第二章 文獻回顧 9
2.1 目前用於副甲狀腺異體移植免疫隔離的研究 9
2.2 幾丁聚醣水膠(chitosan-based hydrogel) 13
第三章 材料與實驗方法 15
3.1 實驗儀器與實驗藥品 15
3.1.1 實驗儀器 15
3.1.2 實驗藥品 15
3.2 氧化幾丁聚醣的製備方法與其特性 17
3.2.1 氧化幾丁聚醣的製備方法 17
3.2.2 特性官能基測定 17
3.2.3 核磁共振光譜儀分析 17
3.3 氧化幾丁聚醣/昆布胺酸水膠製備方法與其特性 18
3.3.1 氧化幾丁聚醣/昆布胺酸水膠製備方法 18
3.3.2 In vitro降解程度測定 18
3.3.3 切割分子量(molecular weight cut-off, MWCO)測定 18
3.4 氧化幾丁聚醣/昆布胺酸水膠細胞活性與細胞毒性測試 20
3.4.1 細胞存活/增生檢測 20
3.4.2 細胞毒性測試 21
3.5 氧化幾丁聚醣/昆布胺酸水膠包覆副甲狀腺組織 23
3.5.1 副甲狀腺組織in vitro培養與包覆 23
3.5.2 副甲狀腺組織in vitro分泌能力與ex vivo周流系統試驗 23
3.5.3 In vitro培養組織塊切片及染色 25
3.6 動物實驗 26
3.6.1 副甲狀腺機能不足動物模型建立 26
3.6.2 動物實驗分組 28
3.6.3 血液生化檢測 28
第四章 結果與討論 29
4.1 氧化幾丁聚醣性質分析 29
4.1.1 紅外線光譜儀分析 29
4.1.2 核磁共振光譜儀分析 31
4.2 氧化幾丁聚醣/昆布胺酸水膠特性分析 32
4.2.1 In vitro降解程度測定 32
4.2.2 切割分子量測定 34
4.3 氧化幾丁聚醣/昆布胺酸水膠細胞活性與細胞毒性測試 36
4.3.1 細胞存活/增生檢測 36
4.3.2 細胞毒性測試 37
4.4 副甲狀腺組織in vitro分泌能力與ex vivo周流系統試驗結果 38
4.4.1 副甲狀腺組織in vitro分泌能力 38
4.4.2 副甲狀腺組織ex vivo周流系統試驗結果 39
4.4.3 In vitro培養組織塊切片及染色 40
4.5 動物實驗 43
第五章 結論 45
參考資料 46


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