Towards a Molecular Signature of Neutrophil Priming

中性粒细胞启动的分子特征

• 批准号: 7708311
• 负责人: ARTHUR Robert SALOMON
• 金额: $ 20.66万
• 依托单位: BROWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-17 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7708311
• 关键词: Address; Affect; Applications Grants; Arts; Biochemical; Bone Marrow; Cell Line; Cells; Cessation of life; Collaborations; Complex; Data; Detection; Development; Disease; Dose; Environmental Risk Factor; Eukaryotic Cell; Event; Experimental Designs; Exposure to; Functional disorder; Future; Global Change; Goals; Host Defense; Human; Imagery; Infection; Inflammation; Inflammatory; Inflammatory Response; Injury; Laboratories; Lead; Life; Lipopolysaccharides; Mass Spectrum Analysis; Mediator of activation protein; Methods; Modeling; Molecular; Molecular Profiling; Noise; Organ failure; Pathway interactions; Patients; Peptides; Phosphorylation; Phosphorylation Site; Platelet Activating Factor; Protein Array; Proteins; Proteome; Publishing; Reactive Oxygen Species; Reagent; Receptor Activation; Relative (related person); Reperfusion Injury; Reporting; Rest; Role; Sampling; Sepsis; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Signaling Molecule; Site; System; Systems Analysis; T-Lymphocyte; Techniques; Technology; Testing; Therapeutic; Time; Tissues; Trauma; Tyrosine Phosphorylation; Tyrosine Phosphorylation Site; Work; cytokine; design; in vivo; injury and repair; mast cell; methionyl-leucyl-phenylalanine; microbial; network models; neutrophil; novel diagnostics; public health relevance; response

DESCRIPTION (provided by applicant): Neutrophil priming is an integral aspect of a well-regulated inflammatory response and is the first step in controlling microbial infection and repair of injury. In this model, an initial injury "primes" neutrophils such that a secondary insult, which would otherwise be tempered in an unprimed cell, now results in an exaggerated inflammatory response. The excessive and unregulated release of cytokines and toxic oxygen radicals causes damage to adjacent tissues leading to tissue dysfunction, organ failure and death. The precise molecular pathways initiated upon priming of human neutrophils are poorly understood. The present proposal focuses on the application of quantitative phosphoproteomic technology to understand the unique proteome-wide tyrosine phosphorylation events associated with human neutrophil priming by exposure to low dose N-formyl-met-leu-phe, lipopolysaccharide or platelet activating factor. Using currently available quantitative phosphoproteomic technology and expertise with primary human neutrophils in our laboratories, we will sample primed neutrophils at various time points and quantify relative changes in wide-scale tyrosine phosphorylation from total cell lysates of cellular peptides. Information to be derived from this project include: global identification of tyrosine phosphorylation events that occur over time after neutrophil priming; visualization of the network of signal transduction pathways that are activated in the primed neutrophils; comparison of activation events that take place over time from neutrophils primed with different proinflammatory mediators. This type of quantitative analysis of global tyrosine phosphorylation sites during a time course of neutrophil priming along with traditional biochemical analysis will permit the construction of a signaling pathway network model for neutrophil priming. Together these findings will test the hypothesis that the primed state of the neutrophil includes signaling events that are common among proinflammatory mediators and can be presented as a molecular signature of this fundamental cellular response. Given that responding neutrophils must interpret a number of signals at a site of inflammation, the results from these studies may assist in the development of novel diagnostic or therapeutic reagents aimed at limiting the auto-inflammatory tissue damage patients suffer as a result of sepsis and trauma. PUBLIC HEALTH RELEVANCE: Neutrophil priming is an integral aspect of a well-regulated inflammatory response and is the first step in controlling microbial infection and repair of injury. The precise molecular pathways initiated upon priming of human neutrophils are poorly understood. In this proposal we introduce modern methods in quantitative mass spectrometry to facilitate the characterization of the cellular signaling pathways initiated upon neutrophil priming by a variety of agents by providing a global view of the phosphorylation state of normal and primed cells.

描述（由申请人提供）：中性粒细胞启动是良好调节的炎症反应的一个组成部分，并且是控制微生物感染和损伤修复的第一步。在这个模型中，最初的损伤会“引发”中性粒细胞，从而导致二次损伤，否则在未引发的细胞中会受到缓和，现在会导致过度的炎症反应。细胞因子和有毒氧自由基的过度和不受调节的释放会对邻近组织造成损害，导致组织功能障碍、器官衰竭和死亡。人们对人类中性粒细胞启动时启动的精确分子途径知之甚少。本提案重点关注定量磷酸蛋白质组学技术的应用，以了解与人中性粒细胞暴露于低剂量 N-甲酰基-met-leu-phe、脂多糖或血小板激活因子相关的独特的蛋白质组酪氨酸磷酸化事件。利用我们实验室目前可用的定量磷酸化蛋白质组学技术和原代人类中性粒细胞的专业知识，我们将在不同时间点对引发的中性粒细胞进行采样，并量化细胞肽总细胞裂解物中大规模酪氨酸磷酸化的相对变化。从该项目中获得的信息包括：中性粒细胞启动后随时间推移发生的酪氨酸磷酸化事件的全局识别；启动中性粒细胞中激活的信号转导通路网络的可视化；比较用不同促炎介质引发的中性粒细胞随时间发生的激活事件。这种对中性粒细胞启动过程中全局酪氨酸磷酸化位点的定量分析以及传统的生化分析将允许构建中性粒细胞启动的信号通路网络模型。这些发现将共同检验这样的假设：中性粒细胞的启动状态包括促炎介质中常见的信号事件，并且可以作为这种基本细胞反应的分子特征呈现。鉴于反应性中性粒细胞必须解释炎症部位的许多信号，这些研究的结果可能有助于开发新型诊断或治疗试剂，旨在限制患者因脓毒症和创伤而遭受的自身炎症组织损伤。公共卫生相关性：中性粒细胞启动是良好调节的炎症反应的一个组成部分，也是控制微生物感染和损伤修复的第一步。人们对人类中性粒细胞启动时启动的精确分子途径知之甚少。在本提案中，我们引入了定量质谱分析中的现代方法，通过提供正常细胞和引发细胞的磷酸化状态的全局视图，以促进对各种药物引发的中性粒细胞引发的细胞信号传导途径的表征。