摘要
摘要 在硬骨組織的細胞外間質中，其包含著有機物質及無機物質存在，而其有機物為膠原蛋白、葡萄胺聚醣等；而無機物的部分則為氫氧基磷灰石。在本研究中，吾人嘗試以生物仿效的方式將幾丁聚醣/動物明膠/三聚磷酸三元複合材料進行礦化而製成硬骨組織工程所需的材料。首先，先將幾丁聚醣/動物明膠膜材與三聚磷酸分子間的作用關係進行分析；由AFM中可以發現膜材在經過低pH值的三聚磷酸鹽溶液交聯後，膜材會形成奈米級的微相分離情形；而在紅外線光譜分析（FTIR）、X光繞射分析（XRD）、能量散佈分析儀（EDS）、動態黏彈性測試（DMA）、及機械測試中，皆顯示出三聚磷酸分子，隨著交聯溶液的pH值改變，其在膜材中扮演的角色亦會由單純的離子交聯劑，變為可以控制膜材中幾丁聚醣分子結晶型態變化的重要角色，且膜材的機械性質亦因此而受三聚磷酸分子的影響。 幾丁聚醣/動物明膠/三聚磷酸三元複合之奈米膜材，其仿生礦化(biomineralization)的研究上；吾人以兩種方式來進行，並輔以能量散佈分析儀（EDS）、掃瞄式電子顯微鏡（SEM）、X光繞射分析（XRD）來觀察；其結果發現先將膜材浸泡至氫氧化鈣水溶液中後，再接著浸泡模擬人體體液（SBF）的製程，可以提供膜材有足夠的成核因子，來形成膜材上氫氧基磷灰石的球狀結晶物。而之後再以穿透式電子顯微鏡（TEM）來觀察實際膜材中的氫氧基磷灰石的結晶型態變化，其顯示在此條件下的膜材，具有仿生礦化的能力，且隨著時間的增長，其膜材中的氫氧基磷灰石的結晶型態越完整且數目也越多。最後則將此膜材用冷凍造的方式製成多孔性的支架，再以同樣的條件下反應，其結果亦可以由SEM中觀察到，膜材的架構不變，而其孔洞周圍有球狀的氫氧基磷灰石結晶堆積形成。 最後，施以簡單的生物體外實驗（ROS細胞培養測試），其發現膜材對細胞並無毒性；且在經過鹼性磷酸酶（alkaline phosphatase；ALPase）的染色及活性測試中，更可以得知在經過仿生礦化後的膜材，其具有骨傳導（osteoconductivity）的性質，且隨著膜材中的氫氧基磷灰石的增加而明顯。綜合以上的研究，可以發現本研究所得之幾丁聚醣/物明膠/氧基磷灰石之仿生礦化材料，具有硬骨組織工程上的應用性。
並列摘要
Abstract The extracellular matrices (ECMs) of hard tissue are mainly composed of organic and inorganic such as collagen, glycosaminoglycans (GAGs) and hydroxyapatite (HA). This study proposed a method to produce size-controlled nanoapatites that are mineralized chitosan/gelatin/tripolyphosphate (TPP) nanocomposites for the use as biomimetic bone tissue-engineering materials. The chitosan/gelatin/TPP ternary composites demonstrated nano-sized phase separation as observed from the atomic force microscopy (AFM) study. FT-IR, X-ray diffraction (XRD), tensile strength and dynamic mechanical analysis (DMA) showed that TPP polyion played an important role in affecting the chemical structures, and in dominating the crystallization and mechanical properties of the chitosan/gelatin/TPP nanocomposites. The chitosan/gelatin/TPP nanocomposite films were used as biomimetic templates for nucleation and growth of hydroxyapatite crystals on the films from simulated body fluid (SBF). The spherical aggregates of hydroxyapatite on the film surface were identified with X-ray diffraction and energy-dispersive X-ray (EDS). The results suggested that chitosan/gelatin/TPP nanocomposites are promising materials for biomineralization of hydroxyapatite. Chitosan/gelatin/TPP ternary composite scaffolds were prepared by a freeze-drying technique, and could also form mineralized apatite on their pore wall surface. The biomimetic organic/inorganic nanocomposite scaffold is now under examination of its osteoconductivity. Finally, from a simple osteoblastic cell (ROS 17/2.8) in vitro evaluation, we found that the film is non-toxic to cell. Furthermore, through the ALP stain and activity tests of alkaline phosphatease, we could get that the film treated by biomineralization has a property of osteoconductivity. This property would be apparent with the increase of hydroxypatite in the film. As we studied above, we could find that the biomineralization materials of chitosan/gelatin/tripolyphosphate acquired in this study has its utilities in biomimetic bone tissue-engineering.
參考文獻
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