Molecular and Force Dynamics: Leukocyte Adhesion Molecules

描述（由申请人提供）：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，这些主题提供了分子尺度上的见解，了解PMNs如何将力作为从滚动过渡到坚定停滞和跨内皮迁移的空间机械线索：1)选择素具有机械和生化特性，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) 22-整合素的亲和力和结合ICAM-1的结合机制通过调节跨内皮迁移的外向内信号提供了一个看门人机制。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)确定中性粒细胞形状极化LFA-1外向内信号传导的分子和机械机制。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。

DESCRIPTION (provided by applicant): Recruitment of leukocytes to sites of acute 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 provide molecular scale insight into how PMNs integrate force as a spatio-mechanical cue in the transition from 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) 22-integrin affinity and bond mechanics in binding ICAM-1 provide a gatekeeper mechanism via regulation of outside-in signaling of transendothelial migration. 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 activation on inflamed endothelium 3) define the molecular and mechanical mechanisms underlying LFA-1 outside-in signaling of neutrophil shape polarization. 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.