Eicosanoids and lung macrophage antimicrobial mechanisms

描述(申请人提供)：肺部感染对全球健康的影响比任何其他类别的疾病都要大，迫切需要更好地了解肺的抗菌素防御。肺泡巨噬细胞(Am)是肺远端固有的天然免疫防御者，其免疫功能与其他吞噬细胞有很大不同。在感染过程中阐述的物质中，来自花生四烯酸(二十烷基类)的脂质介质，包括白三烯(LTS)B4(LTB4)和D4(LTD4)以及前列腺素E2(PGE2)，已成为天然免疫功能的重要调节因子。二十烷类化合物通过连接细胞表面特定的G蛋白偶联受体并启动信号事件来发挥作用。我们已经证明，LTBB4(通过B LT受体1[BLT1])和LTD4(通过半胱氨酰LT受体1[CysLT1])都促进AM的吞噬，而PGE2(通过E前列腺素受体2和4[EP2和EP4])抑制AM吞噬和杀死免疫球蛋白调理的微生物的能力。然而，这些二十烷类化合物的细胞内作用机制还不完全清楚。虽然这两个LT类都促进AM的先天免疫功能，但它们通过不同的信号通路发挥作用。同样，EP2和EP4连接后AM天然免疫功能的抑制是通过不同的途径介导的。随着针对特定二十烷类受体的药理药物的开发进展迅速，了解这种靶向治疗对肺部抗菌防御的影响是很重要的。本项目旨在了解这些二十烷基类化合物影响由调理免疫球蛋白受体Fc3受体(FCR)触发的AM信号通路中的关键事件的机制。为了进行比较，将研究与通过甘露糖受体(MR)摄取白色念珠菌相关的事件。我们将关注的中心信号事件是：1)磷脂酰肌醇3-激酶(PI3K)/10号染色体上缺失的磷酸酶和张力蛋白同源物(PTEN)的激活；2)蛋白激酶C(PKC)的异构体；以及3)小GTP酶。假设BLT1与CysLT1和EP2与EP4的连接对这些吞噬细胞受体触发的信号成分的不同影响反映了这些受体在脂筏膜微区的不同定位，以及它们与两种不同的环磷酸腺苷效应因子-蛋白激酶A(PKA)和环磷酸腺苷激活的交换蛋白(EPAC-1)的不同偶联。利用原代大鼠AM，我们将针对特定的目标确定：1)PKA和EPAC-1对FCR和MR诱导的信号事件的调节；2)FCR、MR及其下游信号成分在脂筏上的定位以及RAFT在吞噬和杀伤中的作用；3)PKA和EPAC-1以及脂筏定位在解释BLT1和CysLT1结扎对FCR和MR信号的差异调制中的作用；4)PKA和EPAC-1以及脂筏定位在解释EP2和EP4连接对FCR和MR信号的差异调制中的作用。这些拟议的研究将为AM的抗菌功能提供基本的见解，并为理解和调节肺内的天然免疫提供一个临床相关的框架。与公共卫生相关：肺炎对全球健康的影响比任何其他类别的疾病都要大，因此必须了解肺部如何保护自己免受感染。包括白三烯和前列腺素在内的类二十烷基脂类物质对肺内关键免疫细胞--肺泡巨噬细胞的抗菌防御功能具有强烈的相反作用。这项拟议的研究旨在了解二十烷类化合物对肺泡巨噬细胞的这些作用背后的细胞内事件，从而为调节肺防御机制提供新的临床相关见解。

DESCRIPTION (provided by applicant): Infections of the lung have a greater impact on global health than any other category of disease, and there is an urgent need to better understand antimicrobial defense of the lung. Alveolar macrophages (AMs) are the resident innate immune defenders of the distal lung, and the immunologic repertoire of these cells often differs substantially from that of other phagocytic cells. Among the substances elaborated during infection, lipid mediators derived from arachidonate (eicosanoids), including leukotrienes (LTs) B4 (LTB4) and D4 (LTD4) as well as prostaglandin E2 (PGE2), have emerged as important modulators of innate immune function. Eicosanoids act by ligating specific G protein-coupled receptors on the cell surface and initiating signaling events. We have shown that both LTBB4 (via B LT receptor 1 [BLT1]) and LTD4 (via cysteinyl LT receptor 1 [cysLT1]) promote, while PGE2 (via E prostanoid receptors 2 and 4 [EP2 and EP4]) inhibits, AM capacity for phagocytosis and killing of IgG-opsonized microbes. However, the intracellular mechanisms by which these eicosanoids act are incompletely understood. Although both LT classes promote innate immune functions in AMs, they act via distinct signaling pathways. Likewise, suppression of AM innate immune functions following ligation of EP2 and EP4 is mediated by distinct pathways. As development of pharmacologic agents targeting specific eicosanoid receptors is proceeding rapidly, it is important that the effects of such targeted therapies on antimicrobial defenses of the lung are understood. This project seeks to understand the mechanisms by which these eicosanoids influence key events in the AM signaling pathway triggered by the receptor for opsonic IgG - the Fc3 receptor (FcR). For comparison, events associated with ingestion of the yeast Candida albicans via the mannose receptor (MR) will be studied. The central signaling events we will focus on are activation of: 1) phosphoinositide 3-kinase (PI3K)/phosphatase and tensin homolog deleted on chromosome 10 (PTEN); 2) isoforms of protein kinase C (PKC); and 3) small GTPases. The hypothesis is that divergent effects on these phagocytic receptor-triggered signaling components by ligation of BLT1 vs. cysLT1 and EP2 vs. EP4 reflect the differential localization of these receptors to lipid raft membrane microdomains and their differential coupling to two distinct cyclic AMP effectors - protein kinase A (PKA) and exchange protein activated by cyclic AMP (Epac-1). Employing primary rat AMs, we will address specific aims to determine: 1) regulation by PKA and Epac-1 of FcR- and MR-induced signaling events; 2) localization to lipid rafts of FcR, MR, and their downstream signaling components and the role of rafts in phagocytosis and killing; 3) the roles of PKA vs. Epac-1 and of lipid raft localization in explaining divergent modulation of FcR and MR signaling by BLT1 vs. cysLT1 ligation; 4) the roles of PKA vs Epac-1 and of lipid raft localization in explaining divergent modulation of FcR and MR signaling by EP2 vs. EP4 ligation. These proposed studies will provide fundamental insights into AM antimicrobial function, and a clinically relevant framework for understanding and modulating innate immunity in the lung. PUBLIC HEALTH RELEVANCE: Pneumonia has more of an impact on global health than any other category of disease, making it imperative to understand how the lung defends itself from infection. Lipid mediators termed eicosanoids and including leukotrienes and prostaglandins exert potent and opposing effects on the antimicrobial defense functions of the key immune cell in the lung, the alveolar macrophage. The proposed research aims to understand the intracellular events that underlie these eicosanoid effects on alveolar macrophages, thereby providing new clinically relevant insights into regulation of lung defense mechanisms.