Beschreibung:
<jats:title>Abstract</jats:title>
<jats:p>Macrophages are able to phagocytose vastly different targets, ranging from bacteria to apoptotic and cancer cells. Especially the difference in rigidity between such targets is striking, with bacteria being ~1000 fold stiffer than human cells. Phagocytosis can also be strongly affected by target rigidity and is seemingly less efficient for softer targets. However, mechanistic studies have largely focused on the uptake of stiff particles, and it is currently poorly understood how macrophages adapt the phagocytic mechanism for efficient uptake of soft targets. Here, we developed deformable hydrogel microparticles to study phagocytosis in unprecedented detail, using targets that more accurately mimic cellular physical properties. We used extrusion through Shirasu Porous glass (SPG) membranes to create emulsions with uniformly sized droplets containing acrylamide, bis(acrylamide) and acrylic acid. Subsequent polymerization yields monodisperse (CV &lt; 0.1) deformable microparticles with cell-like sizes (~10 μm) and rigidities (Young’s modulus 0.1 - 10 kPa). Furthermore, they could readily be conjugated with a variety of ligands and fluorescent dyes. We show that IgG-functionalized microparticles are taken up by J774.1 macrophage-like cells in a rigidity dependent manner. Soft particles (0.3 kPa) were strongly deformed during phagocytosis. We used confocal microscopy and subsequent image processing to resolve the three-dimensional shape of individual particles during engulfment with 50 nm precision. Then, we quantified the physical interaction between target and phagocyte using a novel strategy to infer cellular exerted forces (&gt;10 Pa) directly from the deformed particle shape. We observed highly localized force exertion by the phagocytes and 4 implied mechanically distinct steps in the phagocytic process. Initially, we observe outward-directed pushing forces from the phagocytic cup base. During subsequent pseudopod extension the majority of the deformation is localized in a ring that is initially irregular, but becomes uniform during cup closure. Surprisingly, strong localized punches at the cup base occur in these stages. After cup closure, we observe some of the strongest forces, seemingly pushing the engulfed target into the cell. Our novel approach gives us unprecedented detail on the mechanical interaction between phagocyte and target in phagocytosis. The presented method here is expected to find broad applications in the study of the immune system in vitro and in vivo.</jats:p>
<jats:p>Citation Format: Daan Vorselen, Yifan Wang, Matt Footer, Wei Cai, Julie Theriot. Quantifying the interaction between macrophages and deformable microparticles with cell-like mechanical properties [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr B194.</jats:p>