Mechanobiology of Phagocytosis

巨噬细胞是免疫细胞，是抵抗细菌、病毒、癌细胞和其他病原体的第一道防线。巨噬细胞将自身包裹在外来颗粒周围，吞噬并消化它。这个过程被称为吞噬作用，涉及化学和物理力量。虽然吞噬过程的生物化学研究已经很好，但本项目采取了不同的视角：它将巨噬细胞视为一台机器，将吞噬过程视为一个机械事件。这些力是什么？它们是如何协调的？这台机器是怎么工作的？不了解作用力，我们就缺乏对吞噬作用的完整科学描述，更广泛地说，缺乏对免疫反应本身的描述。该项目采用先进的显微技术和独特的力测量方法，实时研究巨噬细胞的三维结构，同时测量巨噬细胞产生的力。从这个项目产生的数据将提供一个更全面的模型，这个基本的生物过程，集成了生物化学，结构和生物物理力量。整个项目的另一个关键部分是将研究成果公诸于世，帮助人们了解实验室中最新的令人兴奋的科学成果。更广泛的影响项目包括将在当地博物馆开展的基于虚拟现实的展览，以及将在当地一所高中教授为期一周的生物物理学“迷你学期”。本项目的总体目标是利用先进的成像和力测量工具来揭示吞噬过程中力产生和机械传感的机制。吞噬作用是巨噬细胞、中性粒细胞和其他细胞吞噬大量外来病原体颗粒的过程，是免疫系统防御的第一道防线。为了吞噬目标粒子，一个或多个不同的肌动蛋白驱动的机械过程将目标吸入，包围并包围它。除了生化信号动力学外，吞噬还受到包括力和结构动力学在内的基本机械过程的驱动。最基本的问题没有答案：巨噬细胞产生什么力？是什么肌动蛋白动力学和相关的细胞形态动力学导致了它们的产生？外力和机械环境如何影响吞噬过程中的决策？直接将巨噬细胞靶力与肌动蛋白动力学联系起来的测量是建立吞噬过程模型的关键缺失部分。在这个项目中，一系列假设将被测试，这些假设是由类似的机制指导的，这些机制已经被提出用于其他细胞系统，如间充质运动中的“分子离合器”机制，它提供了力的产生和机械传感。在吞噬过程中巨噬细胞力产生的测量将与伴随的高质量的活细胞体积成像细胞形态和细胞骨架动力学一起进行。这将通过使用软珠作为目标来补充，以测量巨噬细胞对目标施加的局部压缩和剪切力。定量时空相关分析的体积肌动蛋白动力学和力的测量也将进行。这将提供丰富的详细数据，告知产生特定巨噬细胞力事件的局部动态肌动蛋白重塑的位置和时间。如果成功，该项目将产生一个全新的巨噬细胞动态图谱，绘制动态3D细胞形态和肌动蛋白动力学与局部力产生和机械传感的关系。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Macrophages are immune cells that act as one of the first lines of defense against bacteria, viruses, cancer cells and other pathogens. Macrophages wrap themselves around a foreign particle, engulfing and digesting it. This process, called phagocytosis, involves both chemistry and physical forces. Though the process of phagocytosis has been well studied biochemically, this project takes a different perspective: it treats the macrophage as a machine and the phagocytic process as a mechanical event. What are the forces? How do they coordinate? How does the machine work? Without understanding forces, we lack a complete scientific description of phagocytosis and, more broadly, the immune response itself. The project employs both advanced microscopic techniques as well as unique force measurement methods to investigate the three-dimensional structure of the macrophage in real time while simultaneously measuring the forces it generates. The data generated from this project will provide a more comprehensive model of this fundamental biological process that integrates the biochemistry, the structure, and the biophysical forces. Another critical component of the overall project is bringing the research to the public to help inform folks on the latest exciting science that is happening in the lab. Broader impact projects include a virtual-reality-based exhibit that will be developed at a local museum, and a week-long biophysics “mini-term” that will be taught at a local high school. The overarching goal of this project is to use advanced imaging and force measurement tools to reveal the mechanisms of force generation and mechanosensing in phagocytosis. Phagocytosis is the process through which macrophages, neutrophils and other cells engulf large foreign pathogen particles as a first line of immune system defense. To engulf a target particle, one or more distinct actin-driven mechanical processes draws in the target, surrounds and encloses it. Along with biochemical signaling dynamics, phagocytosis is also driven by fundamentally mechanical processes involving forces and structural dynamics. The most fundamental questions are unanswered: What forces does a macrophage produce? What actin dynamics and associated cell morphological dynamics give rise to them? How do external forces and mechanical environment affect decision making in phagocytosis? Measurements directly associating macrophage-target forces with actin dynamics are a crucial missing piece to developing models of the engulfment process. Within this project, a set of hypotheses will be tested that are guided by analogous mechanisms that have been proposed for other cell systems such as the “molecular clutch” mechanism in mesenchymal motility which provides force generation as well as mechanosensing. Measurements of macrophage force generation during engulfment will be performed along with accompanying high quality, live cell volumetric imaging of cell morphology and cytoskeletal dynamics. This will be complemented by use of soft beads as targets to measure local compressive and shear forces being imposed by the macrophage on the target. Quantitative spatio-temporal correlation analysis of volumetric actin dynamics and force measurements will also be performed. This will provide rich detailed data informing location and timing of the local dynamic actin remodeling that produce a specific macrophage force event. If successful, the project will produce a completely new dynamical atlas of the macrophage that maps the relationship of dynamic 3D cell morphology and actin dynamics with local force generation and mechanosensing.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.