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  • 學位論文

人類珍珠素蛋白多醣第五片段之結構分析及其對於背根神經結細胞神經纖維生長之影響

Structural Analysis and Biological Roles of Human Perlecan Domain V on Neurite Outgrowth in Dorsal Root Ganglion

指導教授 : 楊維中
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摘要


細胞外間質中的基底膜(Basement membrane)常扮演著維持細胞型態與細胞附著及生長支架(scaffold)的功用,主要由第四型膠原蛋白(type IV collagen)、層粘蛋白(Laminin)、硫酸肝素蛋白多醣(Heparan sulfate proteoglycan,HSPG) (例如:Perlecan及Agrin)、內肌動蛋白(Entaction)、Nidogen所組成。這些分子也被證實與多種細胞貼附有關。而本研究所要探討的是基底膜中的主成分之一-珍珠素(Perlecan),屬硫酸肝素蛋白多醣的一種,是一種蛋白多醣(Proteoglycan),由一條核蛋白(core protein)與2-3條硫酸肝素(Heparan sulfate,HS)黏多醣鏈(Glycosaminoglycan,GAG)所組成。胚胎分化時期若缺乏Perlecan時會造成腦部細胞的異位。腦部受到損傷時,會大量被星狀膠細胞所表現,且不只表現於血管壁週遭還表現於非血管的位置,推測Perlecan可促進神經纖維生長,並進一步促進修復機制的啟動。本研究以基因重組技術(recombinant DNA technology)合成類Perlecan domain V蛋白當做基質,用以培養背根神經結細胞(Dorsal root ganglion,DRG),再外加GAG中的硫化軟骨膠(Chondroitin sulfate)或硫酸肝素(Heparan sulfate),探討其對背根神經結細胞神經纖維生長能力的影響,以期能釐清Perlecan 在中樞神經系統受傷後可能扮演的角色。此外我們以旋光儀(Circular dichroism,CD)分析基因重組之人類Perlecan domain V的構型,藉以探討不同製備條件下,其構型及功能差異,以評估其作為未來神經組織修復之生物醫學材料之潛力。

並列摘要


The extracellular matrix (ECM) basement membranes (BM) can maintain the cell shape and act as a substrate for cell adhesion and as a scaffold for cell growth. The major components of BM include type IV collagen, laminin, heparan sulfate proteoglycan (HSPG) (for example: perlecan and agrin), entaction, and nidogen. Perlecan, the most abundant proteoglycan in BM is a kind of HSPG consisting of a core protein with 2-3 heparan sulfate glycosaminoglycan (GAG) chains. Perlecan null mice developed exencephaly. When the brain is damaged, astrocytes express increased amount of prlecan, not only present on blood vessel wall but also in other places such as hippocampus. In addition, perlecan can promote the neurite extension. In this study, we prepared bacteria expressed recombinant domain V of human perlecan and studied its effect on dorsal root ganglion (DRG) neuron neurite extension. In addition, the seconderary structure of the recombinant human Perlecan domain V by using different preparation protocol was analyzed by circular dichroism (CD). Finally, the potential of developing the recombinant perlecan domain V as a novel biomaterial for nerve tissue repair and regeneration is discussed.

參考文獻


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