Influence of polysialic acid on leukocyte migration

项目摘要 细胞表面蛋白质和脂质糖基化模式的变化越来越被认为是一个重要的因素。 是控制细胞活动的重要因素。人们对如何监管这些变化越来越感兴趣 影响健康和疾病的生理过程。本提案的目标是了解一个特定的 免疫系统细胞表面的碳水化合物修饰影响这些细胞的功能 在炎症和感染的状态下。作为细胞天然免疫和适应性免疫的关键成员， 在免疫应答中，单核细胞被募集到肺部感染部位，在那里它们分化成巨噬细胞。 和树突状细胞在通过淋巴系统迁移之前。这种有针对性的迁移和编程 单核细胞和单核细胞衍生细胞的成熟由特异性的、高度组织化的细胞-细胞和 受体-配体相互作用，其中许多通过细胞表面受体的糖基化调节， 粘附分子聚唾液酸（polySia）是至少三种这样的蛋白质的独特聚糖修饰， 神经细胞粘附分子（NCAM/CD 56）、神经纤毛蛋白-2（NRP-2）和E-选择素配体-1（ESL-1）， 在单核细胞成熟的不同阶段表达。我们将检验这一假设，即调控表达的 在单核细胞分化为巨噬细胞和树突状细胞时， 是聚唾液酸化的，有助于引导细胞归巢和肺疾病期间精心策划的免疫反应。 病毒和细菌病原体感染。缺乏这种酶表达的小鼠， 在白细胞中合成polySia（ST 8 SiaIV-/-），载体蛋白（NCAM/CD 56-/-和NRP-2-/-）将 用于体外和体内研究。我们的具体目标是：i）确定polySia在体内对 单核细胞、巨噬细胞、DC和嗜中性粒细胞在细菌感染小鼠模型中的靶向免疫应答中的作用 和病毒性肺炎; ii）定义通过该机制， 人和鼠单核细胞、巨噬细胞、DC和嗜中性粒细胞控制粘附和迁移 肺微血管单层;和iii）定义和分析多聚Sia和多聚Sia的调节表达， 在原代单核细胞分化为DC和巨噬细胞期间，多聚唾液酸化蛋白质的表达。我们期望 鉴定polySia促进或干扰的特定细胞-细胞和受体-配体相互作用， 单核细胞/巨噬细胞/树突状细胞成熟和迁移过程的不同阶段，以及 中性粒细胞募集我们的研究结果将提供一个蓝图，以工程师的表达水平， 多聚Sia和/或载体蛋白在骨髓细胞中，以优化迁移到和从位点 感染/炎症，以改善整体免疫反应。

PROJECT SUMMARY Changes in the glycosylation pattern of cell surface proteins and lipids are increasingly being recognized as an important factor in controlling cellular activity. There is growing interest in how these changes can be regulated to affect physiologic processes in health and disease. The goal of this proposal is to understand how a specific carbohydrate modification on the surface of cells of the immune system influences the function of these cells during states of inflammation and infection. As a key member of cellular innate and adaptive immune responses, monocytes are recruited to pulmonary sites of infection where they differentiate into macrophages and dendritic cells before migrating through the lymphatic system. This targeted migration and programmed maturation of monocytes and monocyte-derived cells is guided by specific, highly-organized cell-cell and receptor-ligand interactions, many of which are modulated by glycosylation of cell surface receptors and adhesion molecules. Polysialic acid (polySia) is a unique glycan modification of at least three such proteins, neural cell adhesion molecule (NCAM/CD56), neuropilin-2 (NRP-2), and E-selectin ligand-1 (ESL-1), that are expressed at different stages of monocyte maturation. We will test the hypothesis that regulated expression of polySia on monocytes as they differentiate into macrophages and dendritic cells, and on neutrophils that also are polysialylated, helps direct cell homing and a well-orchestrated immune response during pulmonary infection with viral and bacterial pathogens. Mice that are deficient in expression of the enzyme that synthesizes polySia in leukocytes (ST8 SiaIV-/-) and of the carrier proteins (NCAM/CD56-/- and NRP-2-/-) will be used for in vitro and in vivo studies. Our Specific Aims will i) establish the impact in vivo of polySia on monocytes, macrophages, DC and neutrophils in the targeted immune response in a murine model of bacterial and viral pneumonia; ii) define the mechanism(s) through which polysialylated proteins on the surface of human and murine monocytes, macrophages, DC and neutrophils control adhesion to and migration across pulmonary microvascular monolayers; and iii) define and analyze the regulated expression of polySia and of polysialylated proteins during differentiation of primary monocytes into DC and macrophages. We expect to identify specific cell-cell and receptor-ligand interactions that polySia promotes or interferes with during different stages of the monocyte/macrophage/dendritic cell maturation and migratory processes, as well during neutrophil recruitment. The results from our studies will provide a blueprint to engineer levels of expression of polySia and/or carrier proteins in myeloid cells in order to optimize migration to and from sites of infection/inflammation to improve the overall immune response.