溫度敏感型胺基酸水膠包覆胰腺β細胞在組織工程上之應用

Thermosensitive Polypeptide Hydrogel encapsulated Pancreatic β cell for Tissue Engineering Application

李俐瑩(Li-Ying Li)

指導教授 ： 朱一民

國立清華大學/工學院/化學工程學系/碩士(2020年)

摘要

本研究將甲基化聚乙二醇 (methoxy-poly(ethyleneglycol), mPEG) 之 -OH端改質成 -NH2，再共聚合上左旋丙胺酸 (ʟ-alanine)，形成一兩嵌段共聚物mPEG-polyalanine (mPEG-PA)，作為可應用於原位成膠之溫度敏感型胺基酸水膠。希望藉由材料之親疏水性質，提供良好之生長環境來促使細胞貼附，並藉由分析膠體性質及成膠機制，探討此膠體做為移植細胞的支架材料之潛力。
　　將此溫度敏感型胺基酸水膠以(_^1)H核磁共振光譜儀 ((_^1)H-NMR)、傅立葉轉換反射式紅外線光譜儀 (ATR-FTIR)、凝膠滲透層析儀 (GPC)、流變儀與掃描式電子顯微鏡 (SEM) 分析其各種性質及成膠溫度。接著探討材料對細胞毒性、水膠與細胞共培養之生物相容性，由MTT assay結果可以看到細胞存活率皆維持在90 % 以上；再來則進行胰腺 β cell功能性之體外測試，藉由酵素免疫分析法 (ELISA) 及即時定量聚合酶鏈式反應 (qRT-PCR) 確認β cell在包覆於水膠後，仍具有胰島素分泌的功能。
　　本研究最終期望開發一溫度敏感型胺基酸水膠，具有可注射、原位成膠以及良好之生物相容性等特性，成為具有優勢之組織工程的細胞支架，並使用此溫度敏感型胺基酸水膠mPEG-poly(alanine)作為胰島皮下移植的封裝材料，成為糖尿病治療之手段。

關鍵字

胺基酸水膠 ； 溫度敏感型 ； 原位成膠 ； 胰島素

並列摘要

In this study, we report the synthesis and characterization of a temperature-sensitive polypeptide hydrogel. The -OH end of methoxy-poly(ethylene glycol) (mPEG) was modified to -NH2. Then, ʟ -alanine was co-polymerized with mPEG at the end of N-terminal of mPEG to form a two-block copolymer mPEG-poly(alanine) (mPEG-PA). The hydrogel can be used for in-situ gelation. After that, we explores the potential of this hydrogel as a cell scaffold application by analyzing gelation properties.
　　On the basis of the result of (_^1)H nuclear magnetic resonance spectrometer ((_^1)H-NMR) , Fourier transform reflective infrared spectrometer (ATR-FTIR), gel permeation chromatography (GPC), sol-to-gel transition, rheometer and scanning electron microscope (SEM) studies, the molecular weight and gelation properties of the hydrogel are determined. Furthermore, MIN6 β-cells were encapsulated in the hydrogel to observe the cell viability. By MTT assay, the cell viability may be reached above 90%. In order to confirm the insulin secretion, pancreatic β cells in vitro were demonstrated by enzyme immunoassay (ELISA) and quantitative real time polymerase chain reaction (qRT-PCR).
　　Overall, we show a strategy to modify mPEG hydrogel with bioactive polypeptide and use the thermosensitive methoxy-poly(ethylene glycol)- poly(alanine) (mPEG-PA) hydrogel as cell-encapsulating materials for the subcutaneous transplantation of pancreatic β cells. The thermosensitive hydrogel will be looked forwarded to act as a superior cell scaffold for local treatment.

並列關鍵字

polypeptide hydrogel ； temperature sensitive ； in situ gelation ； insulin

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