Novel pathways controlling macrophage inflammation and resolution in atherosclerosis

摘要 尽管有有效的策略来降低胆固醇、动脉粥样硬化和相关的心血管疾病 仍然是全球主要的健康负担。近年来的研究突出了这种令人兴奋的治疗潜力 减少炎症的干预措施，以及确定新的和选择性靶点的必要性。我们的龙 学期目标是了解脂类代谢和炎症是如何相互作用的，以及这些相互作用是如何 在治疗上很有用。当前研究提案的目标是确定转录的作用 核因子红系相关因子-1(Nrf1；又称NFe2L1)在连接这两个过程中起重要作用。 中心假设是巨噬细胞中的Nrf1是预防慢性炎症和 维持胆固醇稳态，增强Nrf1的这些作用可能有益于 动脉硬化。这是基于令人信服的新的初步数据显示，Nrf1在巨噬细胞中被激活 对炎性刺激作出反应，并对脂类代谢途径基因的表达是必需的 对于炎症状态的正确解决至关重要。先前的结果还表明，Nrf1的缺失 极大地损害了细胞胆固醇稳态。这项拟议研究的基本原理是 巨噬细胞Nrf1的功能和机制可能为研究巨噬细胞的调控提供重要的线索 在一般情况下，炎症和消退，并揭示了治疗动脉粥样硬化的新的治疗靶点。 中心假说将通过追求三个具体目标来检验：1)检查Nrf1在 2)阐明Nrf1在巨噬细胞中作用的分子机制；3)探讨Nrf1在巨噬细胞中的作用。 巨噬细胞-Nrf1在体内动脉粥样硬化中的重要性。在第一个目标下，组织特异性可诱导缺失 在申请人的实验室中建立和测试的模型将用于对功能进行全面的表征 Nrf1在炎症输出、脂质代谢和炎症消退中的作用。这一目标还将探索 胆固醇对Nrf1功能的影响及其在调节胆固醇动态平衡中的相互作用 巨噬细胞。在第二个目的中，免疫沉淀容易获得的标记蛋白与DNA偶联 测序和MS/MS分析将确定Nrf1的蛋白质相互作用和翻译后修饰 潜在地调解了它的功能。第三个目标将测试Nrf1缺乏和过度表达对 用已建立的西方饮食喂养带有巨噬细胞的LDLR缺陷小鼠的体内动脉粥样硬化 Nrf1的特异性缺失，或者说腺相关病毒(AAV)介导的巨噬细胞特异性过表达Nrf1。 这种方法是创新的，因为它试图通过使用以下方法克服支持解决调解人交付的问题 巨噬细胞内自然存在的系统。这项拟议的研究意义重大，因为目前 在动脉粥样硬化的背景下，激活内源性炎症分解的方法有限，并且 这项研究可能为了解分辨率在动脉粥样硬化中的作用提供重要的见解，并揭示一种新的 治疗靶点。

Summary In-spite of effective strategies for lowering cholesterol, atherosclerosis and associated cardiovascular diseases remain a major global health burden. Work in recent years has highlighted the exciting therapeutic potential for interventions that reduce inflammation, as well as the need for identifying novel and selective targets. Our long term goal is to understand how lipid metabolism and inflammation interact, and how these interactions can be useful therapeutically. The objective of the current research proposal is to determine the role of the transcription factor Nuclear factor erythroid 2 related factor-1 (Nrf1; also known as Nfe2L1) in linking these two processes. The central hypothesis is that Nrf1 in the macrophage is a critical factor for preventing chronic inflammation and maintaining cholesterol homeostasis, and that enhancing these actions of Nrf1 may be beneficial in atherosclerosis. This is based on compelling new preliminary data showing that Nrf1 is activated in macrophages in response to inflammatory stimuli and is necessary for the expression of lipid metabolism pathway genes that are crucial for proper resolution of the inflammatory state. Previous results also showed that deletion of Nrf1 drastically impairs cellular cholesterol homeostasis.The rationale for the proposed research is that understanding the function and mechanism of macrophage Nrf1 may provide important insight into regulation of macrophage inflammation and resolution in general and reveal a novel therapeutic target for the treatment of atherosclerosis. The central hypothesis will be tested by pursuing three specific aims: 1) Examine the function of Nrf1 in macrophages; 2) Elucidate the molecular mechanism of Nrf1 action in macrophages; and 3) Explore the importance of macrophage-Nrf1 in atherosclerosis in-vivo. Under the first aim, tissue-specific inducible deletion models established and tested in the applicant's lab will be used for a thorough characterization of the function of Nrf1 in inflammatory output, lipid metabolism and resolution of inflammation. This aim will also explore the impact of cholesterol on Nrf1 function, and the reciprocal role of Nrf1 in regulating cholesterol homeostasis in macrophages. In the second aim, immunoprecipitation of readily available tagged protein coupled to DNA sequencing and MS/MS analysis will identify, protein interactors and post translational modifications of Nrf1 potentially mediating its function. The third aim will test the impact of Nrf1 deficiency and overexpression on atherosclerosis in-vivo using already established Western-diet fed Ldlr-deficient mice with either macrophage specific deletion of Nrf1, or adeno-associated-virus (AAV) mediated macrophage specific overexpression of Nrf1. The approach is innovative because it attempts to overcome issues of pro-resolving mediator delivery by using naturally existing systems within macrophages. The proposed research is significant, because current approaches to activate endogenous resolution of inflammation in the context of atherosclerosis are limited, and this study might provide important insight into the role of the resolution in atherosclerosis and reveal a novel therapeutic target.