- 學位論文
軟骨細胞培養於雙醛澱粉交聯明膠凝膠體外評估之研究
In vitro Evaluation of Chondrocyte on Dialdehyde-Starch-Crosslinked Gelatin Hydrogel
摘要
本研究之目的乃利用生物相容性良好的明膠（Gelatin）天然高分子材料作為膠體支架的主要材料，由於明膠的機械性質不理想，因此會利用交聯劑來提升其機械強度，但有鑑於目前常用的交聯劑各有其優缺點，如：戊二醛（glutaraldehyde）、1-ethyl-3-(3-dimethylaminopropyl) carbodiimide（EDC）、綠梔子素（genipin），雖然戊二醛對支架的機械強度提升顯著，但對細胞有很強的細胞毒性；而像EDC交聯後，最後會以尿素的形式脫離，對細胞不會造成細胞毒性，但支架的機械強度提升不顯著；綠梔子素對支架的機械強度提升顯著，且也不會造成很強的細胞毒性，但其產物會產生暗黑色，所以目前使用的交聯劑都有其缺點；因此本研究利用NaIO4氧化開環後的雙醛澱粉（Dialdehyde stach，DAS）來當天然材料交聯劑與明膠交聯，以提升水膠支架的機械強度和降低水膠支架細胞毒性，並在體外培養出軟骨組織，以達修復受損關節軟骨之目的。 由實驗結果可知，水膠支架可於室溫下成膠並有適當提升其機械強度和熱穩定性，並於細胞共混後，幾乎沒有細胞毒性(cytotoxicity)並不會影響細胞在二維和三維結構的水膠支架中的增生(proliferation)，且培養於三維結構的水膠支架中能促進軟骨細胞分泌細胞外基質(extracellular matrix，ECM)，因此可證實水膠的三維結構對於軟骨細胞是個理想的生長環境。相信未來應用於受損軟骨的修復有很大的潛力。
並列摘要
The purpose of research is to use gelatin which has biological compatibility to be hydrogel material .Gelatin is not ideal as the mechanical property, therefore,we promote its mechanical property using the crosslinking agent. In the present commonly used cross-linking agents, for instance: glutaraldehyde , 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide(EDC) , genipin etc.Although glutaraldehyde has ideal mechanical strength, but has very strong cytotoxicity; Using EDC to cross-link,and be separated finally from by the urea form, is non-cytotoxicity, but mechanical strength of hydrgel will not be remarkable; Genipin will promote to mechanical strength of hydrgel, and have weak cytotoxicity, but its product can have the dark color, therefore cross-linking agents have its shortcoming at present.In this reseach use sodium periodate to oxidized starch to form Dialdehyde stach(DAS), sodium periodate is a highly selective oxidizing agent, which cleaves the C-2 – C-3 linkage of anhydroglucose units with the formation of dialdehyde groups. The highly reactive dialdehyde groups in starch can be used as cross-linking agents.DAS was crosslinked with gelatin in order to improve the mechanical properties of gelatin/DAS copolymer, and culture the cartilage tissue in vitro, in order to reach the purpose to repair damaged articular cartilage. From the results, DAS/gelatin hydrogel can form gel at room temperature.The gel strength and thermostability of DAS/gelatin hydrogel was increased when the oxidation percentage of DAS to cross-link gelatin increased.DAS/gelatin hydrogel is non-cytotoxicity and will not influence the cell proliferation.Three-dimensional hydrogel culture model is benefit to chondrocyte to increased the secretion of the extracellular matrix.The results can confirm that the hydrogel three-dimensional structure is the ideal environment for chondrocyte. It has potential to repair local defect of articular cartilage in the future.