探討生物界面之蛋白質吸附機制及其在建構功能性生醫材料之應用

Sustained and Balanced Biological Responses of Biomaterial Surfaces by Adsorption of Functional Proteins

黃媺晴(Mei-Ching Huang)

指導教授 ： 陳賢燁

國立臺灣大學/工學院/化學工程學研究所/碩士(2015年)

https://doi.org/10.6342/NTU.2015.10617

摘要

由於大部分的生物反應都發生在表面，因此表面在生物學及醫學上扮演一個很重要的角色，在生物材料表面設計上，因為蛋白質吸附會影響材料表面的性質，以致於很多研究都專注於如何透過表面改質來阻止蛋白質的吸附。在本論文中，相反地，我們利用吸附來固定蛋白質使表面具有生物功能性，在應用蛋白質吸附於生物材料表面設計時，瞭解蛋白質吸附行為是相當有幫助的，因此，在表面親疏水性對吸附行為影響的部分，利用石英晶體微天平來檢測纖連蛋白於金表面及Parylene-C表面的吸附行為，在蛋白質大小對吸附行為影響的部分，以石英晶體微天平、表面電漿共振及原子力顯微鏡來檢驗纖連蛋白(440kDa)及骨型態發生蛋白(26kDa)於Parylene-C表面的吸附行為。在吸附有骨型態發生蛋白或纖連蛋白的Parylene-C表面，利用石英晶體微天平進行表面特性的驗證，包含了蛋白質的存在及抗蛋白質吸附性質之驗證。在骨型態發生蛋白及纖連蛋白的生物功能展現方面，進行了骨誘導分化測試，並也進行了生物相容性實驗及細胞增生測試，此外，我們更進一步應用高濃度血小板血漿吸附於Parylene-C表面，並檢驗高濃度血小板血漿於骨誘導分化上的影響力。實驗的結果顯現，纖連蛋白對於疏水的表面較親水的表面有較高的親和力，而較大的纖連蛋白吸附於表面上相較於較小的骨型態發生蛋白有較大的吸附質量，但較小的骨型態發生蛋白相較於較大的纖連蛋白有較多的分子數，再者，在飽和吸附的情形下，較大的纖連蛋白相較於較小的骨型態發生蛋白有較多的蛋白質吸附層數，而表面上吸附有纖連蛋白或骨型態發生蛋白展現抗蛋白質吸附的特性及良好的生物相容性，在生物功能表現部分，纖連蛋白及骨型態發生蛋白吸附於Parylene-C表面後皆能表現出骨誘導分化及細胞增生的能力，高濃度血小板血漿吸附於Parylene-C表面後也在骨誘導分化上展現出不錯的成效。

應用蛋白質吸附於生物材料設計上，是相當有發展性的，我們也期望這個方法能在不久的將來被廣泛應用於生物材料設計上！

關鍵字

蛋白質吸附 ； 功能性蛋白質 ； 生物材料 ； 骨誘導 ； 聚對二甲苯

並列摘要

Surfaces play an important role in biology and medicine by reason of most biological reactions occur at surfaces. In biomaterial surface design, efforts have focused on modifying surface to prevent protein adsorption since surface properties of a material are directly related to protein adsorption. In this thesis, by contrast, we utilize an adsorption process for the immobilization of functional proteins to install biological functions on surfaces. It is helpful to understand protein adsorption behavior on surfaces for applying adsorption process in biomaterial surface design. Thus, the effect of adsorbent surface energy (water wettability) on adsorption behavior was studied by using a quartz crystal microbalance (QCM) to examine fibronectin adsorption on Au surface and parylene-C surface. In addition, to investigate how protein size influence adsorption behavior, fibronectin (440kDa) and bone morphogenic protein-2 (26kDa) were used to adsorb on parylene-C, and quartz crystal microbalance (QCM), surface plasmon resonance (SPR) and atomic force microscope (AFM) were employed. In order to characterize surface properties of parylene-C with adsorbed bone morphogenic protein-2 or fibronectin, a quartz crystal microbalance (QCM) was used to inspect the existence of adsorbed proteins and protein-resistant property. To assess the demonstration of biological functions of adsorbed BMP-2 and fibronectin, an osteoinductivity experiments was performed, and cytocompatibility experiment and cell proliferation experiment were conducted as well. Besides, we further employed platelet-rich plasma to adsorb to parylene-C surface, and examined the effectiveness of platelet-rich plasma in osteoinduction. The results reveal that fibronectin showed higher affinities to hydrophobic surfaces than hydrophilic surfaces, and that a greater mass of fibronectin adsorbed to a particular fixed adsorbent surface than BMP-2, whereas more molecules of BMP-2 adsorbed than fibronectin. Also, fibronectin occupied more layers in adsorbed state than BMP-2 at surface saturation. Surfaces with adsorbed fibronectin or BMP-2 exhibited protein-resistant property and good biocompatibility. In the demonstration of biological function, both fibronectin and BMP-2, which were adsorbed on parylene-C surfaces, showed abilities in osteoinduction and cell proliferation. Platelet-rich plasma adsorbed on parylene-C surfaces performed well in osteoinduction, too.

The application of protein adsorption in biomaterial surface design is promising. We envision that this technique can be widely used in biomaterial surface design in the near future.

並列關鍵字

protein adsorption ； functional protein ； biomaterial ； osteoinduction ； parylene

參考文獻

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