PASTEUR, Département de Chimie, École Normale Supérieure, PSL University, Sorbonne Université, CNRS, Paris, France; Institut Pierre-Gilles de Gennes pour la Microfluidique, Paris, France.
PASTEUR, Département de Chimie, École Normale Supérieure, PSL University, Sorbonne Université, CNRS, Paris, France; Institut Curie, PSL University, INSERM U932, Paris, France; Institut Pierre-Gilles de Gennes pour la Microfluidique, Paris, France.
Biophys J. 2019 Aug 6;117(3):408-419. doi: 10.1016/j.bpj.2019.06.021. Epub 2019 Jun 26.
Phagocytosis by macrophages represents a fundamental process essential for both immunity and tissue homeostasis. It consists in the uptake of pathogenic or cellular targets larger than 0.5 μm. For the biggest particles, the phagocytic process involves a massive reorganization of membrane and actin cytoskeleton as well as an important intracellular deformation all in a matter of minutes. The study of the role of the size of objects in their phagocytosis has led to contradictory results in the last decades. We designed a method using confocal microscopy, automated image analysis, and databases for fast quantitative analysis of phagocytosis assays. It yields comprehensive data on the cells and targets geometric and fluorescence intensity parameters, automatically discriminates internalized from external targets, and stores the relationship between a cell and the targets it has engulfed. We used two types of targets (solid polystyrene beads and liquid lipid droplets) to investigate the influence of size on the phagocytic uptake of macrophages. The method made it possible not only to perform phagocytic assays with functionalized droplets and beads of different sizes but to use polydisperse particles to further our understanding of the role of size in phagocytosis. The use of monodisperse and polydisperse objects shows that whereas smaller monodisperse objects are internalized in greater numbers, objects of different sizes presented simultaneously are internalized without preferred size. The total surface engulfed by the cell is thus the main factor limiting the uptake of particles, regardless of their nature or size. A meta-analysis of the literature reveals that this dependence in surface is consistently conserved throughout cell types, targets' nature, or activated receptors.
巨噬细胞的吞噬作用代表了免疫和组织稳态所必需的基本过程。它包括摄取大于 0.5μm 的病原体或细胞靶标。对于最大的颗粒，吞噬过程涉及膜和肌动蛋白细胞骨架的大规模重排以及重要的细胞内变形，所有这些都在几分钟内完成。在过去的几十年中，研究物体大小在吞噬作用中的作用导致了相互矛盾的结果。我们设计了一种使用共聚焦显微镜、自动图像分析和数据库的方法，用于快速定量分析吞噬作用测定。它提供了有关细胞和目标几何和荧光强度参数的综合数据，自动区分内化的和外部的目标，并存储细胞与其吞噬的目标之间的关系。我们使用两种类型的目标（固体聚苯乙烯珠和液体脂质液滴）来研究大小对巨噬细胞吞噬作用的影响。该方法不仅能够进行具有不同大小的功能化液滴和珠的吞噬作用测定，还能够使用多分散颗粒进一步了解大小在吞噬作用中的作用。使用单分散和多分散物体表明，虽然较小的单分散物体被内化的数量更多，但同时呈现的不同大小的物体被内化而没有优先大小。因此，细胞吞噬的总表面积是限制颗粒摄取的主要因素，而与颗粒的性质或大小无关。对文献的荟萃分析表明，这种对表面积的依赖性在细胞类型、靶标性质或激活受体中始终保持一致。
