Genetic and metabolic regulation of macrophage activation at steady state

项目摘要 炎症对于组织修复和有害病原体和分子的清除是必不可少的，但如果被激活 不合时宜的是，先天免疫系统会引发自体炎症。巨噬细胞扮演着关键角色 并对人类健康产生了巨大的影响，既有积极的，也有消极的，但仍存在许多问题 关于这些细胞的问题仍未得到答复。它们是异质的，并采用一种激活状态的光谱，两者 作为独特的组织驻留巨噬细胞，并作为单核细胞在循环中。这些激活状态的驱动因素 仍然难以捉摸。代谢起着一定的作用，巨噬细胞在 前炎性激活，包括从氧化磷酸化转换为糖酵解。然而，无论是 新陈代谢或其他细胞内过程本身就可以决定激活状态，目前尚不清楚，这对 了解与不适当的炎症有关的一系列人类疾病。目前建议的 研究计划解决了这个问题，并极大地扩展了我们之前的工作。 我们正在利用斑马鱼提供的独特优势进行精巧的遗传操作，高产量 筛选和体内成像以剖析细胞内过程和细胞内过程之间的复杂关系 巨噬细胞激活。在Nod样受体nlrc3l中发现了一种失活突变，它导致 不适当的巨噬细胞激活和小胶质细胞的丢失导致我们在斑马鱼中创建了一种新的基因筛查 产生了几个新的巨噬细胞突变，我们将进行研究。这些新的突变导致了不适当的 巨噬细胞激活，破坏小胶质细胞发育，或两者兼而有之。我们正在调查nlrc3l和 相关基因以及我们从筛选中发现的新基因通过改变 代谢和其他细胞内过程。我们正在多个层面采取综合方法--使用 差异转录组学、蛋白质组学和代谢组学，体内单细胞和整体的高功率 成像、基于CRISPR的基因工程、蛋白质-蛋白质相互作用和跨物种 实验--来解决这些问题。此外，我们之前的结果提出了关键的新问题 关于如何调节激活状态以确保组织巨噬细胞的正常发育，以及 存在控制多个器官的巨噬细胞激活的共同机制。我们的研究计划将 使用一组不同的组织巨噬细胞(小胶质细胞、肠道细胞)扩展以解决体内这些关键区域 巨噬细胞和库普弗细胞)。这些研究旨在提供对巨噬细胞的必要见解。 疾病相关背景下的差异性、可塑性和潜力，并解决看似不同的问题 功能(发育和激活调节)编码在分子程序管理中 巨噬细胞。总而言之，我们的研究将带来利用 巨噬细胞用于疾病预防和治疗的特性。

Project Summary Inflammation is essential for tissue repair and removal of harmful pathogens and molecules, but if activated inappropriately, the innate immune system triggers autoinflammatory conditions. Macrophages play a key role in inflammation and have tremendous impact on human health, both positive and negative, yet many questions about these cells remain unanswered. They are heterogeneous and adopt a spectrum of activation states, both as distinct tissue-resident macrophages and in circulation as monocytes. The drivers of these activation states remain elusive. Metabolism plays a role, with macrophages undergoing metabolic changes during proinflammatory activation, including switching from oxidative phosphorylation to glycolysis. However, whether metabolism or other intracellular processes alone can dictate activation states remains unclear and is critical to understanding a host of human diseases associated with inappropriate inflammation. The current proposed research program addresses this question and significantly expands our previous work. We are leveraging the unique advantages zebrafish provide for exquisite genetic manipulations, high throughput screening, and in vivo imaging to dissect the complex relationship between intracellular processes and macrophage activation. The discovery of an inactivating mutation in a NOD-like receptor, nlrc3l, that causes inappropriate macrophage activation and loss of microglia led us to create a new genetic screen in zebrafish that yielded several novel macrophage mutations that we will investigate. These new mutations cause inappropriate macrophage activation, disrupt microglia development or both. We are investigating whether nlrc3l and associated genes as well as new genes we identify from the screen regulate macrophage activation by altering metabolic and other intracellular processes. We are taking an integrated approach at multiple levels—using differential transcriptomics, proteomics, and metabolomics, high power in vivo single cellular and whole-body imaging, CRISPR based genetic engineering, protein-protein interactions, and cross-species experimentations—to address these questions. Furthermore, our previous results raise critical new questions regarding how activation states are regulated to ensure normal tissue macrophage development, and whether common mechanisms exist to control macrophage activation across multiple organs. Our research program will expand to address these key areas in vivo using a diverse set of tissue macrophages (microglia, intestinal macrophages, and Kupffer cells). These studies aim to provide necessary insights into macrophage differentiation, plasticity, and potential in a disease-relevant context, and address how seemingly disparate functions (development and activation regulation) are encoded in the molecular program governing macrophages. In summary, our research will lead to new knowledge and technologies needed to harness the properties of macrophages for disease prevention and treatment.