Molecular and Force Dynamics: Leukocyte Adhesion Molecules

白细胞招募到急性炎症部位是一个精细的过程，由白细胞和内皮细胞粘附分子（CAMs）的膜表达和功能激活启动，包括选择素、整合素和igg超家族配体。在本R01的任期内，出现了一系列统一的主题，这些主题开始定义控制从白细胞滚动到坚定捕获和跨内皮迁移的分子和力动力学：1)选择素被赋予机械和生化特性，使它们能够同时作为粘附和信号转导受体；2)剪切应力和跨膜钙释放激活Ca2+ (CRAC)通道调节细胞内钙通量，同步整合素介导的阻滞和细胞迁移；3) β2-整合素的亲和力和亲和性提供了与跨内皮细胞迁移的内向外信号传导相关的看门人机制。中性粒细胞参与的这些规则导致了一种初步假设，即在急性炎症条件下，粘附和信号事件构成了一个系统，该系统通过设计促进快速和不可逆的转运，同时维持基底内皮屏障功能。在这一竞争更新中，我们将创新的血管模拟微流体通道与实时免疫荧光成像相结合，以实现以下具体目标：1)研究钙释放激活钙通量在中性粒细胞募集多步骤过程中的作用。2)确定中性粒细胞上的e -选择素配体如何作为触发和放大炎症内皮上整合素激活的趋化因子信号传导的机械传感器3)确定内皮屏障功能和中性粒细胞募集如何在皮肤伤口模型中从急性到慢性炎症的转变中耦合。我们的策略包括使用新分离的人类中性粒细胞和小鼠炎症模型，以确定炎症疾病预后和治疗的调节途径和分子靶点。

Recruitment of leukocytes to acute sites of inflammation is a finely orchestrated process initiated by membrane expression and functional activation of leukocyte and endothelial cell adhesion molecules (CAMs) including selectins, integrins, and Ig-super family ligands. A set of unifying themes have emerged over the tenure of this R01 which begin to define the molecular and force dynamics that govern the transition from leukocyte rolling to firm arrest and transendothelial migration: 1) Selectins are endowed with mechanical and biochemical properties which allow them to function as both adhesive and signal transduction receptors; 2) Shear stress and transmembrane calcium release activated Ca2+ (CRAC) channels regulate intracellular calcium flux which functions to synchronize integrin mediated arrest and cell migration; 3) β2-integrin affinity and avidity provide gatekeeper mechanisms associated with inside-out signaling of transendothelial migration. These rules of neutrophil engagement have lead to the primary hypothesis that under conditions of acute inflammation the adhesive and signaling events constitute a system that by design promotes rapid and irreversible transmigration while maintaining basal endothelial barrier function. In this competitive renewal we apply our innovative vascular mimetic microfluidic channels combined with real-time immunofluorescence imaging to pursue the following specific aims: 1) Examine the role of calcium release activated calcium flux in orchestration of the multistep process of neutrophil recruitment. 2) Determine how E-selectin ligands on neutrophils serve as mechano-transducers that trigger and amplify chemokine signaling of integrin actvation on inflamed endothelium 3) Determine how endothelial barrier function and neutrophil recruitment are coupled in the transition from acute to chronic inflammation studied in a cutaneous wound model. Our strategy entails the use of freshly isolated human neutrophils and murine models of inflammation with the overarching goal of identifying regulatory pathways and molecular targets for prognosis and treatment of inflammatory diseases.