DYNAMIC STUDIES OF COAGULATION AND THROMBOSIS

描述：(改编自申请人的摘要)。这是一场竞争激烈的 续签ROI研究项目拨款。拟议研究的目的是 应用最先进的流变学方法，荧光光学切片 显微镜及细胞和分子生物学技术在研究中的应用 动脉粥样硬化和血栓形成。 概述了四个具体目标： 1)了解观察到的基因表达模式的差异 血管内皮细胞(ECS)和平滑肌细胞(SMC) 流体剪应力和周期性机械应变的生理水平。 2)解释剪切预适应对基因表达谱的影响 响应于随后机械环境或细胞因子的变化 刺激。 3)了解不同内皮细胞之间的潜在差异 它们对机械力的反应。 4)解释了流致剪应力的早期信号转导。 通过细胞骨架连接和/或扩散的第二个细胞核 信使。 PI假设应力和应变对应力和应变的影响完全不同 基因表达与细胞表型调控。重要的蛋白质在 血栓形成和动脉粥样硬化(凝血酶受体1[激活的蛋白酶 受体-1或PAR-1]，血管内皮生长因子，前列腺素H合成酶，ICAM，VCAM，GP1b[a血小板 可在内皮细胞上表达的受体]和von Willebrand因子) 已经成为研究的目标。PI之前已经显示出戏剧性的效果 动脉切应力对TPA和ET合成和分泌的影响。在……里面 此外，将使用新的DNA微阵列技术，大大提高 每次实验获得的信息量。这种方法可能会导致 对以协调方式调节的一组基因的理解 机械力，从而导致重要的新假说的血管 生物学。 一个主要的问题是机械信号是如何被传递到 细胞内的变化。光学切片荧光显微镜与数字化 图像处理将被用来研究压力对钙离子通量的影响和 胞内pH。该设备能够获得两个多细胞平均值 单细胞的荧光变化。这种能力也已经发展到 用电子显微镜观察单细胞内离子的三维分布 光学切片和图像分析与重建。原子核的变化 响应电池机械载荷的几何形状也将使用 这项技术。最后一个要考察的问题是压力的相对作用。 (或应变)大小与应力(或应变)变化率的关系。自.以来 动脉血流固有的搏动性，PI‘s在体内的应用 体外实验结果将需要这些知识。

DESCRIPTION: (Adapted from the applicant's abstract). This is a competitive renewal of an ROI research project grant. The aim of the proposed research is to apply state-of-the-art rheological methods, fluorescent optical sectioning microscopy and cell and molecular biology techniques to the study of atherosclerosis and thrombosis. Four specific aims are outlined: 1) To understand the observed differences in gene expression patterns in endothelial cells (Ecs) and smooth muscle cells (SMCs) produced by physiological levels of fluid shear stress and cyclical mechanical strain. 2) To interpret the effect of shear preconditioning on gene expression profiles in response to subsequent change in mechanical environment or cytokine stimulation. 3) To understand the potential differences between various endothelial cell types in their response to mechanical forces. 4) To interpret early signal transduction of flow induced shear stress to the cellular nucleus via either cytoskeletal connections and/or diffusible second messengers. The PI hypothesizes that both stress and strain have quite different effects on gene expression and regulation of cell phenotype. Proteins important in thrombosis and atherosclerosis (thrombin receptor 1 [protease activated receptor - 1 or PAR - 1], VEGF, PGH Synthases, ICAM, VCAM, GP1b [a platelet receptor that can be expressed on endothelial cells] and von Willebrand factor) have been targeted for study. The PI has shown previously dramatic effects of arterial shear stress on TPA and endothelin syntheses and secretion. In addition, new DNA micro array technology will be used to greatly increase the amount of information obtained per experiment. This approach could lead to an understanding of groups of genes that are regulated in a coordinated manner by mechanical forces and thus result in important new hypotheses in vascular biology. A major question is how are mechanical signals transduced to produce intracellular changes. Optical sectioning fluorescence microscopy and digital image processing will be used to study stress effects on calcium fluxes and intracellular pH. The equipment is able to obtain both multicellular averages and single cell changes in fluorescence. The ability has also been developed to examine the three-dimensional distribution of ions in a single-cell using optical sectioning and image analysis and reconstruction. Changes in nuclear geometry in response to cell mechanical loading will also be examined using this technology. A final question to be examined is the relative role of stress (or strain) magnitude versus the rate of change of stress (or strain). Since arterial blood flow is inherently pulsatile, in vivo application of the PI's in vitro results will require this knowledge.