Immobilization of human thrombomodulin onto polymeric thinfilms for biomedical material

将人血栓调节蛋白固定在用于生物医学材料的聚合物薄膜上

基本信息

批准号：
16300156
负责人：
AKASHI Mitsuru
金额：
$ 9.22万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2006
项目状态：
已结题

项目摘要

Human thrombomodulin (hTM) is an endothelial cell-associated protein with potent natural anticoagulant activity by converting thrombin from a procoagulant protease to an anticoagulant. We previously reported the immobilization of hTM onto polymeric film surface by covalent bond formation with the surface of the substrates. We discovered that hTM-immobilized materials had excellent properties for inhibiting both the coagulation and platelet aggregation of human blood. However, the activity of bioactive proteins is decreased through the covalent immobilization process, and chemical immobilization also has the risk of leaving remnant chemical contaminants such as condensation reagents and chemical linkers. In this study, we focused on the physical adsorption of hTM onto a polymeric biomaterial surface because physical adsorption can prevent denaturation and leave no remnant chemical reagents.The adsorption of hTM onto polysulfone (PSF) films was analyzed quantitatively by quartz crystal microbalance (QCM) analysis. The adsorption constant and the maximum adsorption amount, calculated by the assumption of a Langmuir-type adsorption, showed that hTM adsorbed with a relatively weak interaction onto the PSF film. The physically adsorbed hTM showed high co-enzymatic activity for the activation of protein C, as well as anticoagulant activity. Furthermore, the surface wettability of the PSF film was easily controllable by the physical adsorption of hydrophobic and hydrophilic bioactive proteins. The anticoagulant activity of hTM adsorbed on the surface of dialyzer was also confirmed by ex vivo animal studies. The various biocompatible nanofilms were fabricated by layer-by-layer assembly, which can be applied for hTM adsorption.We have successfully developed the blood-compatible biomedical materials by physical adsorption of hTM onto polymeric thinfilms, clarified the adsorption mechanism of hTM, and obtained significant knowledge about blood-compatible biomedical materials.

人血栓形成蛋白（HTM）是一种内皮细胞相关的蛋白质，具有有效的天然抗凝剂活性，通过将凝血酶从促凝剂蛋白酶转化为抗凝剂。我们先前通过与底物表面的共价键形成将HTM固定在聚合物膜表面上。我们发现，HTM-Immobilized材料具有极好的特性，可抑制人体血液的凝结和血小板聚集。然而，通过共价固定过程降低了生物活性蛋白的活性，化学固定化还具有离开残留化学污染物（如冷凝试剂和化学接头）的风险。在这项研究中，我们专注于HTM在聚合物生物材料表面上的物理吸附，因为物理吸附可以防止变性，并且没有保留的化学试剂。石英晶体微生物（QCM）对HTM的吸附在多硫酮（PSF）膜上的吸附进行了定量分析。通过假设Langmuir型吸附计算的吸附常数和最大吸附量表明，HTM在PSF膜上以相对较弱的相互作用吸附。物理吸附的HTM显示出高的共酶活性，可激活蛋白C以及抗凝剂活性。此外，通过疏水和亲水生物活性蛋白的物理吸附，PSF膜的表面润湿性很容易控制。离体动物研究也证实了HTM吸附在透析表面上的HTM的抗凝活性。各种生物相容性的纳米膜是通过一层组件制造的，可以用于HTM吸附。我们通过将HTM的物理吸附到聚合物薄膜上，成功地开发了与血液兼容的生物医学材料，并阐明了HTM的吸附机制，并获得了材料的大量知识。