臺灣博碩士論文加值系統

摘要
組織工程（Tissue engineering）為結合生物、化學及工程三方面的原則，來發展具功能性的代替品，以便恢復、維持、或改善生物組織的功能。組織工程的基本架構主要是由細胞（Cell）、支架（Scaffold）及信號（Signal）所結合而成。
本篇實驗之主要目的便是以組織工程的方式來發展出一個新的創傷敷料。首先，在經過篩選數種真菌及生物醫材後，我們發現由匍枝根黴菌（Rhizopus stolonifer）細胞壁所萃取出的RHIZOCHITOSAN最適合作為此敷材的支架。經由電子顯微鏡的觀察後，發現此支架具有多性之特徵。另外，經過Elson-Morgan比色法亦證實，RHIZOCHITOSAN為一種含有幾丁聚醣（Chitosan）成份的基質材料，其含量約為13.33%。而在物理特性的觀察方面，此材料的吸水能力比同來源的RHIZOCHITIN好2.4倍且有一定的韌性。
再者，我們使用一Platelet-rich-Plasma（PRP）的相關產品，RegenplexTM，作為新敷材中的信號來源。 RegenplexTM是一個富含生長因子（Growth factor）的生物性材料，它主要的來源是把血液中的血小板加以濃縮再乾燥而製成。此材料不但可以長時間保留生長因子的活性，更可以在常溫下長期保存，並且在使用量很少情況之下對治療創傷便有不錯的效果。
根據上述兩者之特性所結合之複合材料，在經由電子顯微鏡觀察後已經發現RegenplexTM可以吸附在RHIZOCHITOSAN上面。而在測試生長因子的釋放活性實驗中，觀察到RegenplexTM並不會受到RHIZOCHITOSAN的影響，並且以可以長期釋放生長因子達一個月之久。另外在動物實驗中，我們觀察到RHIZOCHITOSAN與RegenplexTM的複合材，在前期對於老鼠背部的傷口癒合程度比起單獨使用或是對照組的效果來得更好。而在Zymography的實驗中，RHIZOCHITOSAN、RegenplexTM與兩者之複合材皆可以在傷口癒合前期降低MMP-9的表現，但對MMP-2來說只有複合材料會使其在癒合後期大量表現。再者，在組織切片中，我們也可以在第七天的時候就發現到實驗組的效果已比對照組好。
總而言之，在本研究中我們已經證實RHIZOCHITOSAN確實可以吸附RegenplexTM並可以穩定且長期地釋放生長因子。而此新的創傷敷材，也的確可以幫助老鼠傷口的前期癒合並抑制MMP-9及刺激MMP-2的表現。

RegenplexTM , a dry form of platelets and thrombin, produced growth factors when suspended in PBS. RegenplexTM is not only sustain to release growth factors for a long time, but also can stored in ambient environment. Nevertheless, because it’s cost too high to utilize on clinical directly. The main purpose of experiment is using the cell wall component isolated from Rhizopus stolonifer (RS-6) to combine with RegenplexTM and find a new wound dressing product. This new product provides a elastic mechanical support on which the adhered platelets were protected. In this way, it is possible for clinical-used and the growth factors releasing is not interfered. After screening a lot of biomaterials, we suggest that the chitosan-rich component of Rhizopus stolonifer was an ideal scafford for this purpose. We also find that this new product is a sponge-like porous material with a strong ability to absorb water. The ability of releasing growth factors and the enhancement of wound healing for this product is now under investigation.

目錄
第一章 緒論………………………………………………………………... 1
一、細胞（Cell）…………………………………………………… 1
二、支架（Scaffold）……………………………………………… 2
三、作為支架之生物性材料……………………………………….. 2
A、幾丁質及幾丁聚醣（Chitin & Chitosan）............................. 2
B、纖維蛋白原（Fibronectin）…………………………………. 6
C、膠原蛋白（Collagen）………………………………………. 6
四、信號（Signal）………………………………………………… 7
A、血小板（platelet）…………………………………………… 7
B、血小板豐厚血漿（Platelet-Rich-Plasma, PRP）…………… 9
C、Platelet-derived growth factor（PDGF）……………………. 10
D、Transforming growth factor-β（TGF-β）…………………… 10
五、皮膚傷口之生理機制………………………………………….. 11
A、傷口癒合過程之凝結期與發炎期…………………………… 12
B、傷口癒合過程之遷移及增生期與重塑期…………………… 12
六、Matrix metalloproteinase（MMPs）…………………………… 13
七、本實驗室所開發之生物性材料……………………………….. 14
第二章 研究動機…………………………………………………………... 16
一、RHIZOCHITOSAN.................................................................... 16
二、RegenplexTM 18
第三章 研究架構…………………………………………………………... 19
第四章 實驗材料與方法…………………………………………………... 20
一、實驗材料………………………………………………………. 20
二、實驗方法………………………………………………………. 22
第一部份 創傷敷料的製備流程……………………………........................... 22
第二部份 RHIZOCHITOSAN之基本化學特性測定...................................... 23
第三部份 RHIZOCHITOSAN之基本物理性質測定...................................... 25
第四部份 觀察RHIZOCHITOSAN與RegenplexTM 兩者間之關係……… 25
第五部份 動物實驗………………………………………………… 26
第五章 實驗結果…………………………………………………………... 30
一、創傷敷材的製備……………………………………………… 30
二、RHIZOCHITOSAN之化學性質探討...................................... 30
三、RHIZOCHITOSAN之物理特性探討……………………….. 31
四、RHIZOCHITOSAN與RegenplexTM 兩者間之關係………… 32
五、動物實驗………………………………………………………. 33
第六章 討論………………………………………………………………... 35
一、RHIZOCHITOSAN作為創傷敷材之可能性……………….. 35
二、RHIZOCHITOSAN作為組織工程中基質之可能性……….. 36
三、RHIZOCHITOSAN與RegenplexTM 複合敷材在動物實驗
中之表現……………………………………………………...
37
第七章 參考文獻…………………………………………………………... 38
附錄………………………………………………………………………… 43

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