【作者】 雷丽娟；

【导师】 杨苹；

【作者基本信息】 西南交通大学 ， 材料科学与工程， 2011， 硕士

【摘要】 本文采用非平衡磁控溅射沉积技术在硅基底上制备了锐钛矿晶形为主的钛氧薄膜(Ti-O)。利用3-氨丙基瞵酸(APPA)对Ti-O进行活化,并使用傅里叶变换红外图谱分析(FTIR),接触角,X射线光电子能谱(XPS),扫描电子显微镜(SEM)和石英晶体微天平(QCM)对组装膦酸前后的Ti-O薄膜进行检测。此外,通过4-叠氮基苯胺对肝素进行修饰,利用紫外可见分光光度计和红外对改性后的肝素进行定性分析。在此基础上,利用365 nm波长的紫外灯在组装膦酸后的Ti-O薄膜(APPA/Ti-O)上固定肝素。利用甲苯胺蓝法对固定后的肝素进行定量,并进行部分凝血活酶时间(APTT)的测定,从而优化出实验所用肝素溶液的浓度。借助掩模板在APPA/Ti-O表面制备四种尺寸的肝素条纹微图形。使用光镜和台阶仪对甲苯胺蓝染色后的肝素图形样品进行形貌分析,最后研究图形对血小板和对内皮细胞行为的影响。红外、XPS、接触角、SEM和QCM测定结果显示：膦酸化后的Ti-O表面,出现微弱的APPA的亚甲基伸缩振动峰、N和P元素含量升高、亲水性显著增加、平整度增加、吸附质量增加,可以判定瞵酸APPA组装在Ti-O薄膜表面。经紫外可见分光光度计测定出改性后肝素溶液中出现苯环最大吸收峰,且红外检测结果表明改性后肝素分子中出现-N3和Ar-H伸缩振动峰,表明叠氮基苯胺已成功接枝到肝素分子中。甲苯胺蓝定量结果表明,在40mg/ml肝素浓度条件下,光固定于表面的肝素密度为3.8±0.7μg/cm2;该表面的APTT相对于血浆有所延长。通过甲苯胺蓝对肝素图形染色后发现利用光化学方法能够精确制备出肝素图形,经台阶仪检测图形轮廓可得出图形高度大约为30 nm。血小板和内皮细胞粘附实验表明,组装膦酸的Ti-O表面能够促进内皮细胞的生长但也促进血小板的粘附。肝素固定表面能够降低血小板的粘附和激活,但同时也不利于内皮细胞的增殖。肝素图形由上述两种表面按比例构成,其对血小板和内皮细胞的作用也发生了变化。肝素图形样品能够减少血小板的粘附,其效果与肝素平板样品相当。肝素图形样品还能促进内皮细胞粘附、铺展和增殖。由于图形上两种界面的综合作用,图形尺寸对细胞的生长也产生不同的影响。与细胞尺寸相近的的图形能够使细胞拉长并产生引导作用。而小尺寸的图形更有利于内皮细胞的增殖。细胞粘附力实验表明图形样品可促进细胞actin沿流体方向排布,提高细胞与基底的粘附作用。本研究为同时抑制血小板粘附和促进内皮细胞生长提供了可能,是血液接触材料表面改性的一种新的方法。更多还原

【Abstract】 The anatase titanium oxide films (Ti-O) were synthesized on silicon substrates by Unbalance Magnetron Sputtering. The Ti-O was modified by 3-aminopropylphosphonic acid (APPA), and tested by Fourier transform infrared spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), water contact angle, scanning electron microscopy (SEM) and Quartz crystal microbalance (QCM). In addition, heparin was modified by 4-azidoaniline to obtain photoreactive heparin, which confirmed by ultraviolet absorbance spectra and FTIR. The heparin was immobilized on APPA modified Ti-O (APPA/Ti-O) using a UV lamp with a wavelength of 365 nm. The amount of heparin immobilized on the surface was determined using the toluidine blue assay. And activated partial thromboplastin time (APTT) was tested in order to optimize the concentration of photoactive heparin solution. The patterns of heparin with four different sizes of stripes were fabricated by photo-immobilization of photoreactive heparin on APPA/Ti-O in presence of photomask. The surface morphology of the patterned samples dyed by toluidine blue was examined using optical microscopy and surface profiler. The influences of patterns on the platelets and endothelial cells behaviour were investigated by in vitro tests.After treatment with APPA, two weak peaks of methylene of APPA could be found in FTIR spectrum of Ti-O. It was observed from XPS spectra that surface content of N and P increased. The surface became more hydrophilic, and the particle size of surface diminished. The quality increased after APPA assembly which was tested by QCM. The above results revealed that APPA had assembled on Ti-O surface.After heparin was modified, a maximum absorption assigned to azidophenyl group was observed in the ultraviolet spectrum, and stretching vibrations of C-N3 and Ar-H were found in the FTIR spectrum. The results above confirmed that the azide group had been introduced into heparin molecule.The photoactive heparin with a concentration of 40 mg/ml was used to make heparin patterns, and the non-patterned heparin sample (HEP40) was used as control. Upon UV irradiation, the heparin was immobilized on APPA/Ti-O surface which confirmed by FTIR results. The results of toluidine blue assay showed that the amount of heparin immobilized on HEP40 was 3.8±0.7μg/cm2. The APTT of HEP40 was prolonged compared to plasma. Optical images of patterned heparin showed the patterns were clear and could reproduce the mask pattern precisely. The height of pattern was about 30 nm confirmed by surface profiler.The results of platelets and endothelial cells adhesion showed that APPA modified Ti-O surface could enhance both endothelial cell growth and platelets adhesion. Nonpatterned heparin surface could reduce adhesion and activation of platelets but inhibit the proliferation of endothelial cell as well. Heparin pattern was fabricated in proportion with above two surfaces (APPA and heparin). The patterned heparin samples could reduce the platelet adhesion which represented the similar effect as HEP40. At the same time, the patterned heparin promoted cell adhesion, spreading and proliferation. Due to the compositive effects of two surfaces, the size of pattern could make different effects on the growth of endothelial cells. The micro stripes with a similar size as cell could induce the cells to elongate and arrange along the stripe direction. But the patterns with smaller size were benefit for proliferation of endothelial cell. The experiment of cell adhesive force indicated that the pattern could induce endothelial cells arranged as the direction of flow and improve the adhesive force between the cells and substrate. It may suggest a new modification method for blood-contacting device, providing the functions of both inhibiting thrombus formation and accelerating endothelialization simultaneously.更多还原