COBRE: OK MED RES FOUND: P1: MEMBRANE TETHER FORMATION DURING LEUKOCYTE ROLLING

该子项目是利用NIH/NCRR资助的中心赠款提供的资源的许多研究子项目之一。子项目和研究者（PI）可能从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。所列机构为中心机构，不一定为研究者机构。在炎症和血栓形成过程中，白细胞在活化的内皮细胞和血小板上滚动，允许趋化因子和其他介质的区域采样，这导致白细胞的整合素依赖性停滞和迁移到下层组织中。滚动需要选择素-配体键的快速形成和快速解离，所述选择素-配体键经受由壁剪切应力施加的张力。滚动elocities是非常稳定的，在很宽的范围内的剪切应力，并被认为需要补充选择素配体键的分子特性的细胞功能。这些细胞特征的性质知之甚少。我们观察到当中性粒细胞在P-选择素、E-选择素和L-选择素上滚动时，长的膜栓系快速形成。我们假设，这些系绳发挥了关键作用，稳定滚动速度，促进多个粘合剂接触，并通过减少对个人选择素配体键的力。我们建议研究膜系链的性质和功能，具体目标如下：目标1。我们将通过比较中性粒细胞在固定的P-，E-或L-选择素或分子定义的L-选择素配体上滚动时膜系链的数量，长度和寿命来确定不同的分子相互作用是否影响膜系链的形成或性质。我们还将确定在非选择素依赖性相互作用过程中是否形成系链：表达整合素α 4131的单核白细胞在血管细胞粘附分子-1（VCAM-1）上的滚动。目标2.我们将通过比较中性粒细胞、HL-60细胞、转染的CHO细胞和配体偶联的K562细胞的系链性质来确定不同的细胞特征是否影响膜系链的形成或性质，每种细胞在固定的选择素上滚动时表达共同的选择素配体。细胞的粘弹性将通过细胞分裂剂、固定剂和胆固醇螯合剂来调节。目标3。我们将确定流体动力学是否会影响膜系的形成或性质，通过检查增加的壁面剪切应力，红细胞和改变流动模式的影响。这些研究将提供深入了解特定的细胞和分子特性如何合作，以优化白细胞滚动在广泛的血液动力学条件。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Leukocyte rolling on activated endothelial cells and platelets, during inflammation and thrombosis, allows regional sampling of chemokines and other mediators, which leads to integrin-dependent arrest and emigration of leukocytes into the underlying tissue. Rolling requires the rapid formation and rapid dissociation of selectin-ligand bonds that are subjected to tensile forces applied by wall shear stress. Rolling elocities are remarkably stable over a wide range of shear stresses and are thought to require cellular features that complement the molecular properties of selectin-ligand bonds. The nature of these cellular features is poorly understood. We have visualized rapid formation of long membrane tethers as neutrophils roll on Pselectin, E-selectin, and L-selectin. We hypothesize that these tethers play a key role in stabilizing rolling velocities by facilitating multiple adhesive contacts and by reducing force on individual selectin-ligand bonds. We propose to study the properties and functions of membrane tethers in the following specific aims: Aim 1. We will determine whether distinct molecular interactions affect the formation or properties of membrane tethers by comparing the numbers, lengths, and lifetimes of membrane tethers as neutrophils roll on immobilized P-, E-, or L-selectin or on molecularly defined L-selectin ligands. We will also determine whether tethers form during a non-selectin-dependent interaction: rolling of mononuclear leukocytes expressing integrin a4131 on vascular cell adhesion molecule-1 (VCAM-1). Aim 2. We will determine whether distinct cellular features affect the formation or properties of membrane tethers by comparing tether properties of neutrophils, HL-60 cells, transfected CHO cells, and ligand-coupled K562 cells, each expressing a common selectin ligand, as they roll on an immobilized selectin. The viscoelastic properties of the cells will be modulated by cytoskeletal-disrupting agents, fixation, and cholesterol-chelating agents. Aim 3. We will determine whether fluid dynamics affect the formation or properties of membrane tethers by examining the effects of increased wall shear stress, erythrocytes, and altering flow patterns. These studies will provide insights into how specific cellular and molecular properties cooperate to optimize leukocyte rolling over a wide range of hemodynamic conditions.