Neutrophil Development During Inflammation and Atherosclerosis

项目总结 中性粒细胞对组织中的炎症信号做出敏锐的反应，并在心血管疾病中发挥作用。 (CAD)。中性粒细胞被胆固醇和氧化低密度脂蛋白激活后，产生细胞因子， 脱颗粒释放髓过氧化物酶、天青素和弹性蛋白酶，产生中性粒细胞胞外陷阱 (蚊帐)，并触发裂解细胞死亡。中性粒细胞是由干细胞前体细胞在骨髓中产生的 一旦成熟，就会进入血液和组织来对抗炎症。我们最近发现了一个早期的 中性粒细胞前体干细胞(NEP)存在于人和小鼠的骨髓中。在这里，我们 目前的新数据表明，NEP存在于小鼠和人的外周血中 CAD，提示NEP调节骨髓外的粒细胞生成和炎症。在这 项目中，我们将专注于通过激活Nlrp3炎症小体来实现炎症，因为我们发现Nlrp3和 111b在NEP中高表达。炎症小体传感器Nlrp3的激活触发了一种强大的 炎症反应，包括IL-1β的产生。在项目1中，我们假设激活 NEP内的NLRP3炎症小体触发粒细胞生成和中性粒细胞异质性 促进动脉粥样硬化的进展。我们将通过研究炎症体来检验我们的假设 心血管疾病(CAD)小鼠和人类的激活。在特定的目标1中，我们将测试 人类冠心病患者中性粒细胞亚群的表型和功能变化 与健康的受试者相比。这些受试者正在接受医学上必要的冠状动脉 血管造影术作为正在进行的弗吉尼亚州冠状动脉评估(CAVA)研究的一部分，及其范围 动脉粥样硬化是通过临床测量的。我们将分析NEP、中性粒细胞异质性，以及 这些受试者血液中中性粒细胞Nlrp3炎症小体，并将这些发现与临床参数联系起来。 在特定的目标2中，我们将使用动脉粥样硬化小鼠模型来测试Nlrp3的内在作用 中性粒细胞中的炎性小体影响粒细胞生成和动脉粥样硬化。我们将使用混合 用Nlrp3-/-、Gsdmd-/-和Gsdme-/-小鼠建立嵌合小鼠模型以评估炎症小体途径 在NEP中调节粒细胞的生成。新的中性粒细胞特异性和巨噬细胞特异性Nlrp3基因敲除小鼠 杂交到动脉粥样硬化易感的载脂蛋白E缺陷(apoE-/-)小鼠将直接用于 比较中性粒细胞和巨噬细胞Nlrp3炎性小体的作用对中性粒细胞异质性的影响 和动脉粥样硬化。我们将使用一种新的小鼠模型来评估NEP和ARMARE中Nlrp3的作用 中性粒细胞对动脉粥样硬化的影响。我们将定义Nlrp3炎症体是如何在中性粒细胞前体细胞中激活的 影响粒细胞生成和动脉粥样硬化，进而可能导致新的治疗方法 靶向中性粒细胞中的Nlrp3成分治疗动脉粥样硬化和其他炎症性疾病。

PROJECT SUMMARY Neutrophils acutely respond to inflammatory signals in tissues and play a role in cardiovascular disease (CAD). After activation by cholesterol and oxidized low density lipoproteins, neutrophils produce cytokines, degranulate to release myeloperoxidase, azurocidin, and elastase, generate neutrophil extracellular traps (NETs), and trigger lytic cell death. Neutrophils are produced in the bone marrow by stem cell progenitors and, once mature, egress to blood and tissues to fight inflammation. We recently discovered an early committed neutrophil progenitor stem cell (NeP) present in human and mouse bone marrow. Here, we present new data demonstrating the presence of NeP in the peripheral blood of mice and humans with CAD, suggesting that NeP modulate granulopoiesis and inflammation outside of the bone marrow. In this project, we will focus on inflammation via activation of the Nlrp3 inflammasome, as we find that Nlrp3 and ll1b are highly expressed in NeP. Activation of the inflammasome sensor Nlrp3 triggers a robust inflammatory response, including IL-1β production. In Project 1, we hypothesize that activation of the Nlrp3 inflammasome within NeP triggers granulopoiesis and neutrophil heterogeneity which promotes atherosclerosis progression. We will test our hypothesis by studying inflammasome activation in both mice and humans with cardiovascular disease (CAD). In Specific Aim 1 we will test how neutrophil subsets are phenotypically and functionally changed in human CAD patients compared to healthy subjects. These subjects are undergoing medically necessary coronary angiography as part of the ongoing Coronary Assessment in Virginia (CAVA) study, and their extent of atherosclerosis is measured clinically. We will analyze NeP, neutrophil heterogeneity, and the role of the neutrophilic Nlrp3 inflammasome in blood of these subjects and link these findings to clinical parameters. In Specific Aim 2 we will use atherosclerotic mouse models to test how intrinsic action of the Nlrp3 inflammasome in neutrophils impacts granulopoiesis and atherosclerosis. We will use mixed chimeric mouse models with Nlrp3-/-, Gsdmd-/-, and Gsdme-/- mice to assess how inflammasome pathways in NeP regulate granulopoiesis. New neutrophil-specific and macrophage-specific Nlrp3 knock-out mice crossed to atherosclerosis-susceptible, apolipoprotein E-deficient (apoE-/-) mice will be used to directly compare how action of neutrophil vs macrophage Nlrp3 inflammasomes impact neutrophil heterogeneity and atherosclerosis. We will use a new mouse model to assess Nlrp3 action in NeP versus mature neutrophils on atherosclerosis. We will define how Nlrp3 inflammasome activation in neutrophil progenitors impacts granulopoiesis and atherosclerosis, which in turn could lead to new therapeutic approaches targeting Nlrp3 components in neutrophils for atherosclerosis and other inflammatory diseases.