【作者】 樊珊；

【导师】 陈俊英；

【作者基本信息】 西南交通大学 ， 生物医学工程， 2008， 硕士

【摘要】 本文采用非平衡磁控溅射沉积技术制备了具有金红石与锐钛矿混合结构的钛氧(Ti-O)薄膜,对其进行等离子体浸没H~+注入,并在此基础上,对Ti-O薄膜进行表面生物化改性。通过浓硫酸和双氧水活化Ti-O膜表面使其产生活性的羟基(-OH)基团,并通过硅烷偶联剂氨丙基三乙氧基硅烷(APTE)共价固定细胞外基质成分中的重要生物分子层粘连蛋白(Ln)和纤连蛋白(Fn)。另外,采用傅立叶变换红外光谱仪(FTIR)、X射线光电子能谱(XPS)、原子力显微镜(AFM)、扫描电子显微镜(SEM)、接触角测量等方法对每一步处理后的样品表面特征进行了测试与分析,最后通过体外人脐静脉血管内皮细胞(HUVEC)原代培养实验评价固定Ln和Fn前后Ti-O薄膜表面内皮细胞相容性。Ti-O薄膜以及注氢Ti-O薄膜经浓硫酸和双氧水活化后,FTIR结果显示表面可产生亲水性基团-OH;XPS结果显示-OH对应的O峰位增强;接触角减小,薄膜表面亲水性提高,并且注氢后产生的-OH基团更多。在活化的注氢Ti-O膜表面涂覆APTE后,FTIR及XPS结果表明APTE中的乙氧基水解后可以和Ti-O膜表面的羟基基团反应,使APTE化学键合到Ti-O膜表面;SEM及AFM结果显示表面更加平整,接触角增加,亲水性降低。在注氢Ti-O薄膜表面偶联固定Ln和Fn后,FTIR结果显示样品表面出现蛋白中的特征基团酰胺Ⅰ带、酰胺Ⅱ带基团,联合固定后峰值更强;免疫荧光染色结果显示Ln和Fn在表面出现,并且显示Ln和Fn联合固定成功。蛋白固定使样品表面粗糙度增大。体外人脐静脉内皮细胞培养实验结果表明,Ti-O薄膜本身具有一定的内皮细胞亲和性,在Ti-O薄膜表面固定Ln和Fn对内皮细胞的生长有良好的促进作用,在注氢Ti-O薄膜表面活化偶联后联合固定层粘连蛋白和纤连蛋白促内皮化效果非常显著,优于单种蛋白固定及在未注氢Ti-O薄膜表面固定蛋白的方法。以上的试验结果表明,通过对Ti-O薄膜表面注氢以及活化偶联,可以将Ln和Fn成功的共价固定在Ti-O薄膜表面。在Ti-O薄膜表面固定Ln和Fn对内皮细胞的生长有良好的促进作用。在注氢Ti-O薄膜表面活化偶联后联合固定层粘连蛋白和纤连蛋白促内皮化效果非常显著。更多还原

【Abstract】 In this work,titanium oxide(Ti-O)films were synthesized using Unbalance Magnetron Sputtering(UBMS).To obtain hydroxyl group,the films were treated by plasma immersion H~+ ion implantation.The films were then activated by acid solution.After that,aminopropyltriethoxysilane(APTE)was covalently interacted with the hydroxyl on the Ti-O film surface from activation.Then, laminin(Ln)and fibronectin(Fn)were covalently immobilized to the surface by the amidogen of APTE.Scanning electron microscopy(SEM),X-ray photoelectron spectroscopy(XPS),Fourier Transform Infrared spectroscopy (FTIR),Atomic Force Microscopy(AFM)and contact-angle measurement were used to investigate the characteristics of Ti-O films and modified Ti-O film surface.Human umbilical vein endothelial cells(HUVEC)were in vitro seeded to the modified and unmodified Ti-O films surface for evaluating the cell compatibility and endothelialization trend.The results of FTIR indicated that the functional group of hydroxyl was presented onto Ti-O film surface after being pretreated by plasma implantation and acid solution.The peak of O element corresponding to hydroxyl group increased in results of XPS,the contact angle of Ti-O film surface was lower.The results of FTIR and XPS indicated that APTE could be chemically reacted with the hydroxyl of Ti-O film surface.The results of SEM and AFM suggested that the surface roughness was lower after APTE coated and the contact angle of the film surface was increased.The result of FTIR indicated that albumen amideⅠ,and amideⅡadsorption peaks were presented after laminin and fibronectin immobilized onto Ti-O films. The peaks were stronger when laminin and fibronectin were immobilized together.The result of immunofluorescence indicated that laminin and fibronectin were immobilized to Ti-O film surface.The contact angle of Ti-O film surface showed no obvious change,and the surface roughness was increased.Cell culture investigation suggested that laminin and fibronectin could enhance HUVEC adhesion and proliferation,and improve the HUVE cell behavior.And the results of HUVECs cultured experiment suggested that Ti-O film which modified by plasma implantation and acid solution and immobilized with Ln and Fn is best.All these results indicated that laminin and fibronectin could be covalently immobilized onto Ti-O film surface by APTE.Laminin and fibronectin can promote the adhesion and growth of endothelial cells.So,it is a good surface endothelialization way to immobilized laminin onto and fibronectin Ti-O film surface.更多还原