生醫材料的仿生性改質為目前的主要研究之一，因此奈米級粗糙度及仿生性的塗層可說是目前研究的重點，本論文主要探討三個部分，第一個部分，探討鈦六鋁四釩合金奈米級粗糙度的差異對於表面特性及細胞的反應，以機械研磨及濕式拋光方式對鈦合金表面進行奈米級粗糙度的製備，粗糙度的Ra value 介於2.75~ 30.34 nm，並且使用人類類骨細細胞及纖維細胞評估生醫材料的生物相容性。實驗的方法包含材料表面性質測試，如奈米級表面粗糙度量測、表面型態、濕潤性及表面化學組成分析，生物相容性測試包含細胞增生、細胞型態，以及免疫螢光染色觀察細胞的骨架及貼附情形。第二個部分，利用精確及敏銳的細胞刮取儀評估細胞的黏附力表現，此方法可正確的定量細胞與基材之間的黏附強度，並與表面特性做討論以了解影響黏附力的因素與關係。第三個部分，藉由奈米級表面及仿生性的塗層促進材料表面的親和性，分別比較四種不同前處理、奈米級粗糙度表面、鈣磷塗層，膠原蛋白塗層及鈣磷/膠原蛋白複合塗層的細胞表現，實驗的方法包含仿生塗層的製備、表面塗層型態觀察、成分分析、細胞增生及型態觀察。
研究結果顯示，奈米級粗糙度會造成表面性質改變，包含表面形態、親疏水性、兩性氫氧基的含量百分比，以及表面蛋白質的吸附，也因此進而影響到纖維母細胞和骨母細胞的生長行為，包含細胞型態，細胞黏附力，細胞增殖，及細胞黏附蛋白的表現。而在仿生塗層的製備上，利用簡單且有效率的方式成功製備了均勻的塗層，也確實證實了仿生性塗層能促進細胞的親和性表現。實驗結果歸納出下列結論：
(1) 鈦六鋁四釩表面親水性及表面蛋白質的吸附會隨著奈米級表面粗糙度增加而增加。
(2) 細胞黏附力會隨著表面親水性與奈米級粗糙度增加而增加。
(3) 在相同的時間及表面前處理下，類骨細胞的細胞黏附強度比纖維細胞大。
(4) 類骨細胞及纖維細胞其細胞黏附力會隨著培養時間增加而增加。
(5) 在奈米尺度下不管是類骨細胞及纖維細胞，較粗糙的表面細胞型態有較多的偽足產生，並且黏附蛋白F-actin及vinculin的定量結果也較平滑的表面多。
(6) 在奈米尺度下不管是類骨細胞及纖維細胞，越平滑的表面越能促進細胞增生的表現。
(7) 無CaP前處理的組別，塗佈膠原蛋白於鈦合金表面無法形成均勻塗層
(8) 只有CaP塗層的組別，細胞增生的表現最差。
(9) 鈣磷/膠原蛋白複合塗層確實能促進細胞增生及較佳的細胞貼附表現。
本研究證實了不同的奈米粗糙度等級會造成材料表面特性改變，並且也進一步影響纖維母細胞與骨母細胞的反應，另一方面仿生性collagen/CaP 塗層確實能影響細胞的行為，促進細胞生長，因此鈦六鋁四釩選擇適當的仿生性表面改質處理程序，可使鈦合金更具細胞親和性。此外，應用細胞刮取儀方法來量測細胞與生醫材料的黏附力，可提供評估生醫材料親和性的另一選擇參考。

The biomimetic approach for the fabrication of function biomaterial is expected to promote cell response in recent years. Biomimetic modification of biomaterial becomes one of the current main researches, thus nano roughness and biomimetic coatings also become research priorities of current studies. Three aspects are discussed in this research. The first one aims at investigating the effect of roughness changes in nanometric scale on the surface properties and cell response. Proceeding preparation of nanometric roughness on titanium surface through mechanical grinding and wet polishing methods. Two cell types, human osteoblastlike cells and NIH-3T3 fibroblast cells, were used to evaluate the compatibility of biomaterial. Experiment includes tests of material surface properties, such as nanometric roughness, surface topography, wettability and surface chemical composition analysis; biocompatibility testing includes cell proliferation, cell morphology, cell skeleton and cell attachment. Secondly, we employ a cyto-detacher that quantitatively measures the cell adhesion strength between cells with substrates, and it’s an accurate and sensitive method. At the same time, we discuss about the surface characteristics to understand the factors and relationships that affect adhesion forces. Thirdly, to improve surface biocompatibility by nanomertric surface and biomimetic coating, and respectively compare four different pretreatments such as nanometric roughness surface, CaP coating, Collagen coating and Collagen/CaP coating. The experimental methods include preparation for biomimetic coating, observation to surface coating morphology, chemical composition analysis, cell proliferation and morphological observation.
The result shows that nanometric roughness may change surface properties such as surface topography, wettability, amount percentage of amphoteric hydroxyl and protein adsorption. Thus, it does further affect the growth behavior of fibroblast and osteoblast which includes the performance of cell morphology, cell adhesion, cell proliferation, and cell adhesive protein. In preparation for biomimetic coating, it successfully prepared continuous and homogenous coatings through a simple and efficient method, which confirmed the performance that biomimetic coating does promote the cytocompatibility. The followings are the summaries of the results:
(1) Both wettability and protein adsorption of Ti6Al4V are increasing with surface nanometric roughness.
(2) Cell adhesion force is increasing with nanometric roughness and wettability.
(3) In a same time and same surface pretreatment, cell adhesion force of osteoblast-like cell is greater than fibroblasts.
(4) Cell adhesion force of osteoblast-like cell and fibroblast are increasing with culturing time.
(5) In the nanometer scale, osteoblast-like cell and fibroblast have more lamellipodia and filopodia, and quantitative result of adhering to protein F-actin and vinculin are more compared with the smoother groups on the type of rough surface cell.
(6) In the nanometer scale, the smoother surface has the better cell proliferation for osteoblast-like cell and fibroblasts.
(7) Groups without CaP pre-treatment do not form uniform collagen coatings on titanium surface.
(8) Only groups with CaP coating have the worst cell proliferation.
(9) Collagen / CaP coating promotes the cell proliferation and cell attachment.
This study demonstrates that different levels of nanometric roughness change surface properties and also further affect responses of fibroblast and osteoblast. On the other hand, biomimetic Collagen/CaP coatings actually affect cell behavior and promote growth of cells. Thus, Ti6Al4V is selected the appropriate treatment of biomimetic surface modification so that makes titanium alloy more cytocompatibility. In addition, measuring cell adhesion force and biomaterials by application of cytodetachment provides an alternative reference to the affinity assessment of biomaterial.

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