Neutrophil Microscopy and Quantitative Imaging Core B

总结 中性粒细胞是抵抗细菌和真菌的第一道防线，并诱导炎症。的研究 因此，介导嗜中性粒细胞活化的分子机制具有根本的重要性。想象这些 机制需要独特的、最先进的技术和方法。为了实现这一点，设计了内核B 协调、设计、开发和执行新的技术方法，以了解中性粒细胞 炎症的生物学核心B将与所有三个项目合作，并将提供相关的成像和显微镜检查 在小鼠和人样品中的测定。核心利用现有的尖端技术来实现新的 实验方法，并帮助阐明研究计划中提出的复杂的细胞生物学过程 项目1（赫德里克博士）、项目2（Catz博士）和项目3（霍夫曼博士）。为此，我们将在 技术核心总监（Kiosses博士）的专门工作，他将分配特定的时间和精力来支持 PPG的目标。核心B优化了中性粒细胞成像、图像定量和数据的工作流程 共享所有项目。它提供了严格的质量控制，并确保多个性能的一致性。 应用于冠心病中性粒细胞和前体细胞研究的关键技术。 核心B将实施独特的活细胞方法来研究中性粒细胞囊泡运输、细胞骨架 重塑和细胞死亡途径。核心B在技术上和概念上都是创新的， 专为生物学家设计的新方法，包括超分辨率显微镜和定量分析， 中性粒细胞动力学以阐明中性粒细胞功能并阐明其在炎症中的作用。在关系中 在项目1中，核心B将识别NETosis期间动脉粥样硬化斑块中的中性粒细胞及其前体细胞 以及在高脂饮食中牙菌斑发展的各个阶段的炎性小体活化条件下的焦亡， 使用CAD小鼠的ApoE-/- Western饮食模型的离体和体内。与项目2，核心 B将利用单分子超分辨率显微镜和全内反射荧光显微镜 研究调节中性粒细胞前体中囊泡运输和肌动蛋白重塑的分子机制。 核心B还将利用相关光电子显微镜（CLEM）来表征新发现的 嗜酸性粒细胞（PSG）。与项目3合作，核心B将检查三维 当细胞经历凋亡或坏死时，线粒体网络中的定位和空间变化 来自小鼠模型的中性粒细胞及其前体细胞和来自患有 CAD.总而言之，Core B的独特设计充分利用了拉霍亚的宝贵技术 尽可能最大限度地提高与PPG相关的专门研究和生产力。

SUMMARY Neutrophils are the first line of defense against bacteria and fungi and induce inflammation. The study of the molecular mechanisms mediating neutrophil activation is therefore of fundamental importance. Visualizing these mechanisms requires unique, state-of-the-art technologies and approaches. To achieve this, Core B is designed to coordinate, design, develop and execute new technical approaches applied to the understanding of neutrophil biology in inflammation. Core B will work with all three Projects and will provide relevant imaging and microscopy assays in mice and human samples. The core utilizes available cutting-edge technology to implement new experimental approaches and to help elucidate the intricate cell biology processes proposed in research plans of Projects 1 (Dr. Hedrick), Project 2 (Dr. Catz) and Project 3 (Dr. Hoffman). To this end, we will have the dedicated effort of a Technology Core director (Dr. Kiosses) who will allocate specific time and efforts to support the goals of the PPG. Core B optimizes the workflows of neutrophil imaging, image quantification and data sharing for all projects. It provides stringent quality controls and ensures uniformity of performance of multiple key technologies applied to the study of neutrophils and precursors in the setting of coronary artery disease. Core B will implement unique live-cell approaches to study neutrophil vesicular trafficking, cytoskeleton remodeling and cell death pathways. Core B is technically and conceptually innovative because it utilizes, neutrophil-dedicated, novel approaches that include super-resolution microscopy and quantitative analyses of neutrophil dynamics to elucidate neutrophil functions and to elucidate their role in inflammation. In relationship with Project 1, Core B will identify neutrophils and their precursors in atherosclerotic plaques during NETosis and pyroptosis in conditions of inflammasome activation at various stages of plaque development in high-fat diet, both ex-vivo and in vivo using the ApoE-/- Western diet model of CAD mice. In association with Project 2, Core B will utilize single-molecule super-resolution microscopy and Total Internal Reflection Fluorescence Microscopy to study molecular mechanisms regulating vesicular trafficking and actin remodeling in neutrophil precursors. Core B will also utilize Correlated-Light Electron Microscopy (CLEM) to characterize the newly identified precursor-specific granules (PSGs). In collaboration with Project 3, Core B will examine three-dimensional localization and spatial changes in the mitochondrial network as the cells undergo apoptotic or necroptotic cell death in both neutrophils and their precursors from mouse models and human neutrophils from patients with CAD. In conclusion, Core B is uniquely designed to use the valuable technology available in La Jolla to the fullest extent possible to maximize neutrophil-dedicate research and productivity associated with the PPG.