钛氧薄膜表面固定PEG及硒代胱胺构建催化活性层的研究

发布时间：2018-08-04 09:52

【摘要】：血管支架等血液接触材料抗凝抗增生性能不足严重制约了其临床应用。NO是心血管系统的信号因子,可抑制血小板的粘附及平滑肌细胞的增生,其在生物材料改性研究领域的应用受到广泛的关注。本论文在氧化钛表面通过PEG偶联固定NO的催化分子硒代胱胺,从而构建催化活性层。并对催化活性层进行了系列表征和抗凝、抗增生等生物学评价。采用非平衡磁控溅射在单晶硅材料表面沉积氧化钛薄膜,通过在材料表面沉积聚多巴胺的方式引入有机官能团,双端修饰的聚乙二醇可作为偶联剂将硒代胱胺固定于材料表面。论文通过FTIR、XPS跟踪检测了各改性阶段样品的表面微特征变化,证实各分子在样品表面成功固定。称重实验比较了两种工艺制备的聚多巴胺样品,显示以摇床震荡沉积5次得到的聚多巴胺的量最大；氨基定量实验显示以1.5mmol/l浓度接枝的PEG样品表面的氨基量最大。对改性样品进行体外生物学评价,GPx酶活力检测得到最终改性样品的酶活力为30.43U/cm2。ELISA检测证实改性样品能够明显抑制纤维蛋白原的粘附及变性。体外血小板粘附样品通过荧光染色、SEM等手段进行表征。结果显示,无供体组中,经聚乙二醇修饰的样品表面的血小板数量明显少于对比样；供体组中,胶原蛋白严重激活了对比样表面的血小板,而由于NO的作用,Se样品表面的血小板粘附、聚集、激活状态都得到了显著抑制,cGMP检测证实此效应是通过NO-cGMP信号通路实现的。体外SMC实验中,无供体组显示经修饰的样品均可在一定程度上抑制SMC的粘附；在供体存在条件下,SMC在Se样品表面的粘附受到强烈抑制,胞内cGMP的含量也明显升高。综上所述,本文通过PEG偶联固定硒代胱胺,成功在材料表面构建了催化活性层,改性后表面可催化内源性NO供体释放NO,进而升高细胞内cGMP含量,发挥其生物效应。在PEG、NO双重机制的作用下,材料表面抗凝及抗增生性能得到明显改善,为血液接触材料的表面改性提供了一种新途径。
[Abstract]:The lack of anticoagulant and anti-proliferative properties of blood contact materials such as vascular stents seriously restricts its clinical application. No is a signal factor of cardiovascular system, which can inhibit platelet adhesion and smooth muscle cell proliferation. Its application in the field of biomaterial modification has received extensive attention. In this paper, the catalytic active layer was constructed by coupling the no catalyst molecule selenocysteine on the surface of titanium oxide by PEG. A series of characterization of catalytic active layer and biological evaluation of anticoagulant and anti-proliferation were carried out. Titanium oxide thin films were deposited on the surface of monocrystalline silicon by unbalanced magnetron sputtering. Organic functional groups were introduced by deposition of dopamine on the surface of the materials. The double end modified polyethylene glycol could be used as a coupling agent to immobilize selenocysteamine on the surface of the materials. In this paper, FTIR XPS was used to trace and detect the changes of surface microfeatures of the samples at different modification stages, and it was proved that the molecules were successfully immobilized on the surface of the samples. The results showed that the maximum amount of polydopamine was obtained by shaking deposition 5 times, and the amino content on the surface of PEG grafted with 1.5mmol/l concentration was the largest. The enzyme activity of the modified sample was determined by biological evaluation in vitro. The result showed that the modified sample could inhibit the adhesion and denaturation of fibrinogen by 30.43U/cm2.ELISA. Platelet adhesion in vitro was characterized by fluorescence staining and SEM. The results showed that in the non-donor group, the number of platelet on the surface of the sample modified by polyethylene glycol was significantly less than that on the control group, while in the donor group, the platelet on the surface of the contrast sample was seriously activated by collagen. Because of the effect of no on platelet adhesion, aggregation and activation of se samples, the inhibition of NO-cGMP signaling pathway was demonstrated. In vitro SMC assay, no donor group showed that the modified samples could inhibit the adhesion of SMC to a certain extent, and the adhesion of SMC to the surface of se samples was strongly inhibited in the presence of donor, and the content of intracellular cGMP was also significantly increased. To sum up, the catalytic active layer was successfully constructed on the surface of the material by PEG coupling immobilization of selenocysteine. The modified surface could catalyze the release of no from endogenous no donors, and then increase the content of cGMP in the cells and exert its biological effect. The anticoagulant and anti-proliferative properties of the materials were improved obviously under the dual mechanism of PEGN and no, which provided a new way for the surface modification of blood contact materials.
【学位授予单位】：西南交通大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R318.08

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