ADHESION MOLECULE INDUCT ACTIVATION IN INFLAMMATORY INJURY

白细胞向缺血和再灌流心肌的迁移 再灌流后迅速恢复。在动物模型中，心肌组织 通过实验疗法显著减少伤害，限制了 中性粒细胞积聚和/或激活。建议进行的实验 这项应用将使用犬类和小鼠细胞的体外模型 确定中性粒细胞转变所需的黏附机制 从循环中的静止细胞到能够黏附的细胞- 对心肌细胞的依赖性损伤。这种转变在体内发生为 中性粒细胞通过血管壁迁移到心肌组织。这个 具体目标将解决这一过程中不同的实验阶段 转变定义，1)允许中性粒细胞 在切应力作用下的血管内皮细胞停顿 静脉范围，2)允许中性粒细胞 通过血管壁结构迁移并附着在心肌细胞上， 3)反应性中性粒细胞产生的粘附性决定因素 氧、细胞因子(IL-1β、TNFα和IL-6)和趋化因子(IL-8和IL-6) MCP-1)。将使用两种一般的实验策略--i) 分离已知的粘连机制(例如，通过使用重组粘连 分子)，以确定它是否能支持或影响白细胞 在这三个阶段中的作用，以及ii)确定相对 每种粘接机构对这些功能的贡献如下 白细胞与细胞因子刺激的内皮细胞相互作用， 细胞外表面和细胞因子刺激的心肌细胞。这个 以后的研究将使用单抗来封闭特定的粘附物 功能，或来自基因工程缺陷小鼠的细胞 特定的黏附分子。这些研究的重点将放在CD上 18整合素和刺激因子(IL-8和血小板激活因子) 上调该家族两个成员的功能(CD11b/CD18和 CD11a/CD18)、ICAM-1(CD54)在内皮和心肌细胞上的表达 选择素家族(CD62L和CD62P)和一种未知的黏附机制 这解释了中性粒细胞在24小时后的初次黏附 IL-1对内皮细胞的刺激作用。我们还将评估 允许犬单核细胞停留在内皮细胞上的粘连步骤 在流动条件下，影响上述生产的 细胞因子和趋化因子。这些研究将集中在两个粘连上 除了上面提到的分子，β1整合素，VLA4 (CD49d/CD29)和VCAM-1(CD106)。犬和犬的活体显微镜观察 小鼠模型也将被用于评估由此产生的一些概念 从体外获得的数据。这些研究将加强 了解以下关键的连续粘接步骤 中性粒细胞和单核细胞的迁移和激活 心肌再灌注损伤。

Emigration of leukocytes into ischemic and reperfused myocardium occurs rapidly following reperfusion. In animal models, myocardial tissue injury is reduced significantly by experimental therapies that limit the accumulation and/or activation of neutrophils. Experiments proposed in this application will use in vitro models with canine and murine cells to define adhesive mechanisms necessary for the transition of neutrophils from resting cells in the circulation to cells capable of adherence- dependent damage to cardiac myocytes. This transition occurs in vivo as neutrophils migrate through vessel walls into myocardial tissue. The Specific Aims will address experimentally distinct stages in this transition define, 1) the molecular mechanisms that allow neutrophils to stop on endothelium under conditions of flow with shear stresses in the venular range, 2) the adhesive mechanisms that allow neutrophils to migrate through vessel wall structures and attach to cardiac myocytes, and 3) the adhesive determinants for neutrophil production of reactive oxygen, cytokines (IL-1beta, TNFalpha, and IL-6) and chemokines (IL-8 and MCP-1). Two general experimental strategies will be used -- i) to isolate a known adhesive mechanism (e.g., by using recombinant adhesion molecules) in order to determine if it can support or influence leukocyte functions in these 3 stages, and ii) to determine the relative contributions of each adhesive mechanisms to these functions as leukocytes interact with cytokine-stimulated endothelial cells, extracellular surfaces, and cytokine-stimulated cardiac myocytes. The latter studies will use monoclonal antibodies to block specific adhesive functions, or cells from mice genetically engineered to be deficient in specific adhesion molecules. The focus of these studies will be on CD 18 integrins and stimuli (IL-8 and platelet activating factor) that upregulate the function of two members of this family (CD11b/CD18 and CD11a/CD18), ICAM-1 (CD54) on endothelium and myocytes, members of the selectin family (CD62L and CD62P), and an undefined adhesive mechanism that accounts for primary adhesion of neutrophils 24 hours after stimulation of endothelial cells with IL-1. We will also evaluate the adhesive steps that allow canine monocyte to stop on endothelial cells under conditions of flow, and that influence production of the above cytokines and chemokines. These studies will focus on two adhesion molecules in addition to those cited above, the beta1 integrin, VLA4 (CD49d/CD29), and VCAM-1 (CD106). Intravital microscopy in canine and murine models will also be used to assess some of the concepts resulting from the data obtained in vitro. These studies will enhance understanding of the critical sequential adhesive steps necessary for the emigration and activation of neutrophils and monocytes in the context of myocardial reperfusion injury.