Integration of Leukotriene and Prostaglandin Receptor Signaling in Mast Cell Activation and Pulmonary Inflammation during Asthma

项目摘要 肥大细胞(Mast cell，MC)是哮喘的效应细胞，其激活可导致半胱氨酸基的分泌 白三烯(Cys-LT)和前列腺素(PGs)。MCS不仅分泌这些调解人，而且他们还 拥有它们的感受器，来感知它们的信号。Cys-LT是强有力的支气管收缩药，强大的 血管渗漏的诱导剂，呼吸道高反应性的增强剂，在 哮喘和其他炎症性疾病。Cys-Lts主要通过两种G蛋白偶联受体发挥作用 (GPCR)、CysLT1R和CysLT2R。另一种GPCRGPR17被LTD4激活。CysLT2R和 Gpr17负性调节CysLT1R功能。LTE4是Cys-Lts中含量最丰富、最稳定的一种，是一种 CysLT1R和CysLT2R的弱、部分激动剂。然而，LTE4在体内诱导了独特的反应 这不能用LTC4或LTD4概括。在MC中，LTE4比LTD4更强大，并传递信号 通过过氧化物酶体增殖物激活受体-γ和P2Y12R。最近，GPR99被 被标识为另一个优先使用LTE4的CysLTR。了解所有这些Cys-LT受体是如何 (CysLTR)相互作用以响应多个配体是至关重要的，特别是考虑到它们的 在呼吸道生理学中的作用。此外，Cys-Lts与PGE2协同增强钙离子通量，c-LTS与PGE2协同作用。 Fos、COX-2、PGD2和巨噬细胞炎性蛋白-1β(MIP-1β；CCl))的产生。 有趣的是，只有联合使用CysLT1R拮抗剂才能阻断LTD4-PGE2的协同作用 (MK571/Singulair)和EP3拮抗剂(L-798)，提示CysLT1R需要结合 用于治疗哮喘炎症的拮抗剂和EP3R拮抗剂。LTD4和PGE2的协同作用 增强DER致敏小鼠的肺部炎症(免疫细胞、杯状细胞的募集 化生，炎性转录上调)。与LTD4类似，LTE4与PGE2有协同作用，但它 不同于LTD4的是涉及PPARγ的信号。基于这些观察，我们假设Cys- LTS诱导Cys-LT反应性受体之间的复杂相互作用深刻影响 通过将抗炎PGE2信号转换为促炎信号，上调下游信号 MC中COX-2和PGD2的产生影响Th2炎症和哮喘。我们将检验这一假设 在以下具体目标中：1)确定已知(CysLT1R和CysLT2R)之间的相互作用 和可能的(GPR99、P2Y12R和PPARɣ)CysLTRs对Cys-LTS的反应，影响MC功能，2) 揭示Cys-LT-PGE2协同诱导PGD2产生和MC的机制 3)确定Cys-LT-PGE2相互作用和MC的生理意义。 体内肺部炎症。我们将分析其病理、生理和免疫学特征。 观察MCs的免疫应答，评价MCs的贡献。这些研究将带来大量的 对哮喘和过敏性疾病的病原学和治疗意义，以及为 未来调节炎症的治疗分子的开发和翻译。

Project Summary Mast cells (MCs) are effector cells in asthma and their activation causes secretion of cysteinyl leukotrienes (cys-LTs) and prostaglandins (PGs). MCs not only secrete these mediators, but they also possess receptors for them, to perceive their signals. Cys-LTs are potent bronchoconstrictors, powerful inducers of vascular leakage, potentiators of airway hyper-responsiveness and play an important role in asthma and other inflammatory disorders. Cys-LTs mainly act through two G-protein-coupled receptors (GPCR), CysLT1R and CysLT2R. Another GPCR, GPR17 is activated by LTD4. Both CysLT2R and GPR17 negatively regulate CysLT1R function. LTE4, the most abundant and stable of the cys-LTs, is a weak, partial agonist for the CysLT1R and CysLT2R. However, LTE4 induces unique responses in vivo that cannot be recapitulated by LTC4 or LTD4. In MC, LTE4 is more potent than LTD4 and relays signals through peroxisome proliferator activating receptor (PPAR)-γ and P2Y12R. Recently, GPR99 was identified as another CysLTR with a preference for LTE4. Understanding how all these cys-LT receptors (CysLTR) interact with each other in response to multiple ligands is critical, especially considering their role in airway physiology. Further, cys-LTs together with PGE2 synergistically potentiate calcium flux, c- Fos, COX-2, PGD2 and Macrophage Inflammatory Protein-1β (MIP-1β; CCL4)) generation in MCs. Interestingly, LTD4-PGE2 synergism is blocked only by combined treatment of CysLT1R antagonist (MK571/ singulair) and EP3 antagonist (L-798), suggesting the need for a combination of CysLT1R antagonists and EP3R antagonists to treat inflammation in asthma. LTD4+PGE2 synergism also potentiates pulmonary inflammation in der f sensitized mice (recruitment of immune cells, goblet cell metaplasia, up-regulation of inflammatory transcripts). Similar to LTD4, LTE4 synergizes with PGE2 but it differs from LTD4 via signals involving PPARγ. Based on these observations, we hypothesize that cys- LTs induce complex interactions between cys-LT-responsive receptors to profoundly influence the downstream signaling by switching anti-inflammatory PGE2 signaling to pro-inflammatory, upregulating COX-2 and PGD2 production in MC impacting Th2 inflammation and asthma. We will test this hypothesis in the following specific aims: 1) To determine the interplay between the known (CysLT1R and CysLT2R) and putative (GPR99, P2Y12R and PPARɣ) CysLTRs in response to cys-LTs, influencing MC function, 2) To uncover the mechanism by which cys-LT-PGE2 synergism induces PGD2 production and MC activation and 3) To determine the physiological significance of cys-LT+PGE2 interactions and MC in pulmonary inflammation in vivo. We will analyze pathologic, physiologic, and immunologic signatures of the immune response and evaluate the contribution of MCs. These studies will carry substantial pathogenic and therapeutic implications for asthma and allergic diseases as well as provide the basis for development and translation of future therapeutic molecules that regulate inflammation.