Advanced Imaging Core

摘要 高级成像核心C对于准确定义流程的空间和时间组织至关重要 控制巨噬细胞的可塑性。通过流式细胞术测定的表面标记主要用于 鉴定和分类不同的巨噬细胞亚群。然而，每个项目都明确指出，巨噬细胞 从根本上说，多样性是特定时空细胞内信号和协调的结果 转录节目由利基环境决定。这些过程控制巨噬细胞的可塑性。 炎症性肺损伤及其消退过程中。所有项目都将解决细胞间和旁分泌如何 信号与细胞内的信号相互作用，改变巨噬细胞的命运，以响应肺损伤和分解。因此， 核心C的功能是提供体外和体外所需的先进试剂和成像方法 进行活体研究，并能够对每个项目中的假设进行测试。我们将提供工具来解决 一个基本的问题，即给定的信号通路或转录程序对活跃的人是必不可少的 巨噬细胞分化、特化和功能的调节。要获得机械性的信号洞察力 每个项目的成功都需要核心C提供常规和超分辨率的方法 水平以询问活巨噬细胞中的信号通路。时空代谢、转录和 三磷酸腺苷、β-连环蛋白、CREB、STAT家族激活巨噬细胞后信号转导谱的变化 转录因子和其他关键信号将通过多色超分辨率显微镜和超分辨率显微镜进行研究。 分辨率活动成像。这些对巨噬细胞的分析将允许对时间进行细粒度评估- 巨噬细胞变种所使用的程序的序列和空间组织。虽然在生命中 成像是询问肺损伤和恢复结果、活体呼吸运动的最直接方法 动物是通过活体肺成像获得有意义结果的主要障碍。因此，一个主要的核心 C功能还将提供先进的双光子活体内肺成像，使用一种新的方法来可靠地 识别活体小鼠肺中的髓系细胞(在血管和组织中)并跟踪单个细胞的运动，使每个细胞 项目直接检查他们感兴趣的通路在肺损伤和修复阶段的功能影响。 通过提供对巨噬细胞分化、迁移、 和解决免疫挑战，核心C对该计划的整体成功至关重要。

Abstract Advanced Imaging Core C is essential to accurately define spatial and temporal organization of processes governing macrophage plasticity. Surface markers determined by flow cytometry have been primarily used to identify and sort diverse macrophage subpopulations. However, each Project makes clear that macrophage diversity is fundamentally a consequence of specific spatiotemporal intracellular signaling and coordinated transcriptional programming dictated by the niche environment. These processes control macrophage plasticity during both inflammatory lung injury and its resolution. All Projects will address how inter-cellular and paracrine signals interact with intracellular cues to alter macrophage fate in response to lung injury and resolution. Thus, the function of Core C is to provide advanced reagents and imaging methodologies required for both in vitro and in vivo studies and enable testing of hypotheses in each Project. We will provide the tools to address the fundamental question that a given signaling pathway or transcriptional program is essential for the active regulation of macrophage differentiation, specialization, and function. To gain the mechanistic signaling insights needed for the success of each Project, Core C will provide methods at both conventional and super-resolution levels to interrogate the signaling pathways in live macrophages. Spatiotemporal metabolic, transcriptional, and signaling landscape changes in response to macrophage activation by ATP, β-catenin, CREB, STAT family transcription factors and other key signals will be studied by multi-color super-resolution microscopy and super- resolution activity imaging. These analyses of macrophages will permit a fine-grained assessment of the time- sequence and spatial organization of the programs employed by macrophage variants. Although intravital imaging is the most direct way to interrogate lung injury and recovery outcomes, respiratory motion in the live animal represents a major obstacle for obtaining meaningful results by in vivo lung imaging. Thus, a major Core C function will also be to provide advanced two-photon intravital lung imaging using a novel approach to reliably identify myeloid cells in living mouse lung (in vessels and tissue) and track single cell motion, allowing each Project to directly examine the functional effects of their pathway of interest during lung injury and repair phases. By providing a seamless visual examination of signaling activity in macrophages as they differentiate, migrate, and resolve immune challenges, Core C is essential for the Program’s overall success.