Macrophage Activation & Substance P Receptor Expression

描述(申请人提供)：P物质是一种神经肽，可以影响周围组织和中枢神经系统的宿主反应。因此，P物质及其受体，即神经激肽-1受体，是神经系统和免疫系统之间最重要的联系之一。神经-免疫相互作用将是这项提议的重点，总体目标是明确定义神经激肽-1受体用于增强宿主对病原体的反应的机制。这些研究将利用体内和体外感染模型，重点放在巨噬细胞、树突状细胞和小胶质细胞，以及神经肽P物质如何增强宿主来源或病原体来源的信号。使用神经激肽-1受体缺陷小鼠将提供一个有价值的体内模型系统，以确定与同源野生型小鼠品系相比宿主反应的差异。细菌和病毒病原体将被用来确定P物质与神经激肽-1受体结合所介导的体内机制。这种机制将侧重于宿主反应的启动，以及抗原特异性反应的发展。研究还将侧重于P物质对巨噬细胞和树突状细胞上神经激肽-1受体介导的抗原处理/提呈和共刺激的影响，以此作为调节宿主反应的机制。体外培养的原代脑小胶质细胞将被用来研究它们对P物质的反应性。此外，小胶质细胞培养将被用来质疑是否可以确定神经激肽-1受体介导的机制，这可能有助于解释神经肽介导的中枢神经系统内的炎症反应。这些研究将使用的方法包括核糖核酸酶保护分析，T-PER匀浆中存在的分子的酶联免疫吸附分析，免疫荧光染色细胞的流式细胞术分析，以及三色染色的组织切片的共聚焦分析，以确定表达分子的细胞来源。基因缺陷和转基因小鼠也将被用作确定P物质介导的反应的模型。综上所述，这些研究将证明P物质是宿主对病原体免疫反应的组成部分。

DESCRIPTION (provided by applicant): Substance P is a neuropeptide that can affect host responses in peripheral tissues and in the central nervous system. As such, substance P and its receptor, the neurokinin-1 receptor, represent one of the most important links between the nervous and immune systems. Neuro-immunological interactions will be the focus of this proposal, with the overall goal of clearly defining the mechanisms used by neurokinin-1 receptors to augment the host response against pathogens. These investigations will make use of in vivo and in vitro models of infection, and focus on macrophages, dendritic cells, and microglia, and how the neuropeptide substance P can augment host-derived or pathogen-derived signals. The use of neurokinin-1 receptor-deficient mice will provide a valuable in vivo model system to define differences in the host responses when compared to congenic, wild-type mouse strains. Bacterial and viral pathogens will be used to define in vivo mechanisms mediated by substance P binding to neurokinin-1 receptors. Such mechanisms will focus on the initiation of host responses, as well as the development of antigen-specific responses. Studies will also focus on the effects of substance P on antigen processing/presentation and co-stimulation mediated by neurokinin-1 receptors on macrophages and dendritic cells as a mechanism for modulating the host response. In vitro cultures of primary brain microglia will be investigated for their responsiveness to substance P. In addition, microglia cultures will be used to question whether neurokinin-1 receptor-mediated mechanisms can be defined which might help to explain neuropeptide-mediated inflammatory responses within the central nervous system. The methods which will be used in these studies include ribonuclease protection assays, enzyme-linked immunosorbent assays of molecules present in T-PER homogenates, flow cytometric analyses of immunofluorescently stained cells, and confocal analyses of tissue sections which have been tri-color stained to identify the cellular source of expressed molecules. Genetically deficient and transgenic mice will also be used as models for defining substance P mediated responses. Taken together, these studies will demonstrate that substance P is an integral part of the host immune response against pathogens.