Neutrophil Response to Chemoattractants

中性粒细胞在肺部炎症部位和其他部位的定位是通过一个多步骤的过程完成的，在许多情况下，由g蛋白偶联受体（gpcr）的配体触发。暴露于化学引诱剂配体可诱导血管中的初始隔离、长期滞留和迁移。它们在肺部炎症中的潜在重要性不仅体现在GPCR配体安装引起的显著中性粒细胞反应上，还体现在GPCR的靶向缺失对中性粒细胞积累的巨大影响上。尽管对gpcr的反应基本相似，但化学引诱剂诱导的功能反应却有显著差异。该提案的中心假设是，化学吸引剂激活的MAP激酶的光谱是由基于膜的信号平台组织的不同酪氨酸激酶的激活控制的。反过来，细胞的功能反应至少在一定程度上是由激活的MAP激酶阵列决定的。使用人类和小鼠中性粒细胞（后者允许使用基因靶向动物），PLB-985细胞（一种可分化为中性粒细胞样细胞的可转染细胞系）和中国仓鼠卵巢（CHO）细胞表达fMLP， PAF和LTB4受体，我们提出以4个特定目标为重点来验证这些假设。1)确定细胞质酪氨酸激酶在中性粒细胞从g蛋白偶联受体到MAP激酶的信号通路中的作用；2)确定响应化学引诱剂而组织信号复合物的平台和机制；3)确定中性粒细胞中连接酪氨酸激酶和p38的信号通路的组织；4)在体内检测g蛋白偶联受体激活p38在肺部炎症中的作用。这些研究将为GPC4的激活提供新的见解，并为炎症状态的干预提供新的靶点。

Localization of neutrophils at sites of inflammation in the lung and elsewhere is accomplished through a multi-step process triggered in many circumstances by ligands that act through G-protein-coupled receptors (GPCRs). Both initial sequestration in the vasculature, prolonged retention, and migration are induced by exposure to chemoattractant ligands. Their potential importance in pulmonary inflammation is shown not only by the striking neutrophilic response induced by installation of GPCR ligands, but also by the dramatic effects of the targeted deletion of GPCRs on neutrophil accumulation. Despite fundamental similarities in the response to GPCRs, the functional responses induced by chemoattractants differ strikingly. The central hypothesis of the proposal is that the spectrum of MAP kinases activated by chemoattractants is governed by the activation of distinct tyrosine kinases organized by membrane-based signaling platforms. In turn, the functional response of the cell is determined, at least in part, by the array of MAP kinases activated. Using human and murine neutrophils (the latter to permit use of gene-targeted animals), PLB-985 cells (a transfectable line that differentiates into a neutrophil-like cell) and chinese hamster ovary (CHO) cells expressing receptors for fMLP, PAF, and LTB4, we propose to test these hypotheses focusing on 4 specific aims. 1) Determine the role of cytoplasmic tyrosine kinases in signaling pathways that lead from G-protein coupled receptors to MAP kinases in the neutrophil; 2) Determine the platform and mechanisms by which signaling complexes are organized in response to chemoattractants; 3) Determine the organization of signaling pathways that connect tyrosine kinases and p38 in the neutrophil; and 4) Test the role of G-protein coupled receptor activation of p38 in lung inflammation in vivo. These studies will offer new insights into GPC4 activation in general, and offer new targets for intervention in inflammatory states.