PLATELET ACTIVATION AND POLYMER SURFACE THROMBOGENICITY

与任何血液接触的人工表面，无论是长期或 短期应用，生物反应有其基础， 在材料表面发生的分子和细胞相互作用。 这是材料是否被设计为生物学上 “惰性”和脱落的血液蛋白质和/或细胞，或者被开发为 具有生物学“活性”并提供蛋白质附着的底物， 细胞生长 我们的研究一直是，并将继续是， 阐明了参与粘附的基本细胞机制， 随后粘附的血小板和细胞与基质的相互作用。 这项工作还旨在提供有关“先驱” 蛋白质和细胞/血小板在静态和流动条件下相互作用， 随后通过蛋白质重排，选择性地 蛋白质缺失或添加，细胞扩散，配体受体相互作用， 和蛋白质或细胞分离。 对这些主要事件的理解 相对于材料（聚合物）结构和相对于次级事件 例如血栓形成、栓塞、细胞附着和生长， 建议研究的主要目标。 为此，相关光和 电子显微镜技术与各种 免疫标记和配体标记方法以及生物化学， 生理和药理学评估。 这种方法允许 在纳米水平上显示聚合物的体相和表面结构 的分辨率，并允许材料结构直接与 蛋白质附着和取向，包括蛋白质-蛋白质相互作用。 这反过来又与随后的血小板粘附、扩散相关， 活化，以及血小板-血小板相互作用，导致或不导致， 血栓形成和栓塞。 特别是几类相分离的结构 聚氨酯，其具有不同的软链段/硬链段比率， 类是确定立体高电压（1兆电子伏）TEM和 低电压（1-5 kV）高分辨率SEM。 常规ESCA和接触 还将对这些聚合物进行角度研究。 吸附， 各种血清蛋白的构象和表位表达， 聚合物，将通过蛋白质的直接成像来观察， 胶体金免疫标记或直接配体标记方法。 空间分辨率在2-5 nm的数量级上（远低于纳米颗粒的尺寸）。 单个聚合物结构域和蛋白质的大小）。 血小板 附着，铺展，Ca++动员，分泌，细胞骨架 重组、受体可用性和运动性将被评估 相对于聚合物的超微结构、表面特征和蛋白质， 吸附/取向 这是要做的裸以及蛋白质 暴露的聚合物和在静态和流动条件下。 血小板-血小板 也要研究相互作用，如表面改性， 蛋白质重排，或选择性蛋白质添加或蛋白质去除， 血小板 这些研究将加深对血液材料的认识 在分子和细胞水平上的相互作用，并将有助于设计 “生物相容性”材料的惰性，细胞“脱落” 表面或设计用于细胞粘附和生长的表面。 另外 这些使用人类蛋白质/细胞的体外研究有助于比较 使用其他物种的体内和体外研究。

With any blood contacting artificial surface, whether for long-term or short-term application, the biological response has its basis in the molecular and cellular interactions occurring at the material surface. This is the case whether the material is designed to be biologically "inert" and shed blood proteins and/or cells or is developed to be biologically "active" and provide a substrate for protein attachment and cell growth. Our studies have been, and continue to be, directed toward elucidating the basic cellular mechanisms involved in adhesion and the subsequent interactions of adherent platelets and cells with the substrate. The work is also designed to provide information on how the "pioneer" proteins and cells/platelets interact in static and in flow conditions to subsequently modify the surface through protein rearrangement, selective protein deletion or addition, cell spreading, ligand receptor interactions, and protein or cell detachment. An understanding of these primary events relative to material (polymer) structure and relative to secondary events such as thrombus formation, embolization, cell attachment and growth is a key goal of the proposed studies. To this end correlative light and electron microscopic techniques are used in conjunction with various immunolabeling and ligand labeling procedures and with biochemical, physiological and pharmacological assessments. This approach permits visualization of bulk and surface polymer structure at the nanometer level of resolution and allows material structure to be directly correlated with protein attachment and orientation including protein-protein interactions. This in turn can be correlated to subsequent platelet adhesion, spreading, activation, and platelet-platelet interactions leading, or not leading, to thrombus formation and embolization. Specifically the structure of several classes of phase separated polyurethanes with differing soft segment/hard segment ratios within each class are to be determined with stereo high voltage (1 MeV) TEM and with low voltage (1-5 kV) high resolution SEM. Conventional ESCA and contact angle studies on these polymers are also to be performed. Adsorption, conformation, and epitope expression of various serum proteins, to these polymers, is to be observed by direct imaging of proteins and with colloidal gold immunolabeling or direct ligand labeling procedures. Spatial resolution is on the order of 2-5 nm (well below the size of individual polymer domains and the size of the proteins). Platelet attachment, spreading, Ca++ mobilization, secretion, cytoskeletal reorganization, receptor availability and motility are to be evaluated relative to polymer ultrastructure, surface characteristics, and protein adsorption/orientation. This is to be done on bare as well as protein exposed polymer and in static and flow conditions. Platelet-platelet interactions are also be to be studied as are surface modifications such as protein rearrangement, or selective protein addition or protein removal by platelets. These studies will enhance the understanding of blood-materials interactions at the molecular and cellular level and will aid in the design of "biocompatible" materials either in the sense of inert, cell "shedding" surfaces or surfaces designed for cell adhesion and growth. Additionally these in vitro studies with human proteins/cells facilitate comparisons with in vivo, and in vitro studies using other species.