Regulation of Neutrophil Migration and Polarity

描述（由申请人提供）：中性粒细胞是人体中最丰富的白色血细胞，是人体抵御微生物感染的第一道细胞防线。虽然通常在血流中发现，但中性粒细胞迅速迁移到组织中的感染部位。这种迁移受到严格的调控;事实上，中性粒细胞的异常积聚会导致组织损伤。因此，了解控制中性粒细胞迁移的分子机制具有相当重要的临床意义。该应用程序的重点是了解小Rho GTdR Cdc 42在中性粒细胞极性迁移过程中的作用。我们对Cdc 42-null中性粒细胞的分析表明，Cdc 42是中性粒细胞极性的关键调节因子。在一个机制的水平上，我们已经确定了一个新的途径介导Cdc 42的功能，并涉及白细胞整合素CD 11b/CD 18。CD 11b/CD 18是一种与致命的人类免疫缺陷疾病白细胞粘附缺陷综合征相关的粘附分子。CD 11b是中性粒细胞向组织迁移的重要介质，其机制尚不清楚。我们发现CD 11b聚集在Cdc 42-null细胞中是有缺陷的，恢复CD 11b聚集拯救了极性-从而表明CD 11b参与了中性粒细胞极性。有趣的是，我们发现，CD 11b空中性粒细胞，类似于Cdc 42空细胞，在迁移过程中表现出缺陷的极性。这些数据表明Cdc 42通过CD 11b信号调节中性粒细胞极性的假设。我们建议在体外和体内确定Cdc 42-CD 11b轴在调节中性粒细胞迁移的分子机制和生物学后果。我们计划使用视频显微镜，免疫荧光成像和生物化学实验来检查（目的1）Cdc 42调节极化中性粒细胞中CD 11b聚集的分子机制;（目的2）确定中性粒细胞极性中Cdc 42-CD 11b轴下游激活的信号通路。(Aim 3）由于已知CD 11b在中性粒细胞迁移到肺中中起重要作用，如果Cdc 42-CD 11b轴在中性粒细胞极性中是关键的，那么Cdc 42的缺陷应导致体内肺部炎症过程中的等同缺陷。我们将使用CD 11b依赖性肺损伤模型对此进行测试。我们认为，初级Cdc 42空中性粒细胞的分析提供了一个难得的机会，研究Cdc 42在中性粒细胞极性的具体功能。如果我们的假设，Cdc 42调节中性粒细胞极性通过CD 11b信号被验证，这将显着改变的观点Cdc 42和CD 11b的功能，中性粒细胞迁移，从而在嗜中性粒细胞依赖性炎症。拟议的研究预计将导致了解并最终治疗与嗜中性粒细胞相关的疾病的策略。

DESCRIPTION (provided by applicant): Neutrophils are the most abundant white blood cell in humans, and form the body's first line of cellular defense against infecting microorganisms. While normally found in the blood stream, neutrophils migrate rapidly to sites of infection in tissue. This migration is tightly regulated; indeed, aberrant accumulation of neutrophils causes tissue injury. The understanding of the molecular mechanism(s) controlling neutrophil migration is thus of considerable clinical importance. This application focuses on understanding the role of the small Rho GTPase Cdc42 in neutrophil polarity during migration. Our analysis of Cdc42-null neutrophils shows that Cdc42 is a critical regulator of polarity in neutrophils. At a mechanistic level, we have identified a new pathway mediating Cdc42 functions and involving the leukocyte integrin CD11b/CD18. CD11b/CD18 is an adhesion molecule associated with a lethal human immunodeficiency disorder Leukocyte Adhesion Deficiency syndrome. CD11b is an important mediator of neutrophil migration into tissue, of ill-defined mechanism. We show CD11b clustering is defective in Cdc42-null cells, and restoring CD11b clustering rescues polarity - thereby suggesting the involvement of CD11b in neutrophil polarity. Interestingly, we show that CD11b-null neutrophils, similarly to Cdc42-null cells, exhibited defective polarity during migration. These data suggest the hypothesis that Cdc42 regulates neutrophil polarity through CD11b signaling. We propose to determine the molecular mechanism and biological consequences of the Cdc42-CD11b axis in modulating neutrophil migration both in vitro and in vivo. We plan to use video microcopy, immunofluorescence imaging and biochemistry experiments to examine (Aim1) the molecular mechanism by which Cdc42 regulates CD11b clustering in polarized neutrophils; (Aim 2) to determine the signaling pathway that is activated downstream of the Cdc42-CD11b axis in neutrophil polarity. (Aim 3) Since CD11b is known to play significant roles in neutrophil migration into lungs, if the Cdc42-CD11b axis is critical in neutrophil polarity, then deficiency of Cdc42 should result in equivalent defects in lung inflammatory processes in vivo. We will test this using a model of CD11b-dependent lung injury. We believe that analysis of primary Cdc42-null neutrophils offers a rare opportunity to study the specific function of Cdc42 in neutrophil polarity. If our hypothesis that Cdc42 regulates neutrophil polarity though CD11b signaling is validated, this will significantly change the view of Cdc42 and CD11b functions in neutrophil migration and thus in neutrophil-dependent inflammation. The proposed studies are expected to lead to strategies to understand and ultimately treat neutrophil-related disorders.