Phagocytosis of IgG-coated Emulsion Droplets – PhagoDrop

Phagocytosis is a process that consists in the ability of a cell to internalize objects larger than 0.5 microns. Whereas unicellular organisms use phagocytosis to capture and eat preys, in multicellular organisms it represents a fundamental part of innate immunity, organ homeostasis and tissue remodeling. Innate immunity relies on a specialized subset of cells, the professional phagocytes, which patrol the organism, identify, ingest and eliminate pathogens. Among them, macrophages are versatile cells residing in tissues that are able to scavenge worn-out cells and participate to the activation of the adaptive immune response. Phagocytosis by macrophages is triggered by the binding of the target to specific receptors present at the surface of the phagocyte.
Several receptors have been identified so far, each involving different signaling pathways and ingestion mechanism. In the case of Fc\gamma receptors-mediated phagocytosis, antigens present at the surface of the target are bound by specific soluble immunoglobulins (IgGs). Fc regions of those IgGs are actively recognized by the Fc\gammareceptors (Fc\gamma Rs from the phagocyte surface, which form clusters and trigger the internalization. Engulfment then occurs by an actin-driven membrane extension and closure of a phagocytic cup around the foreign body to create a specific degradative compartment: the phagosome.
Most of our current quantitative knowledge on phagocytosis is based on the use of various model particles such as heat- or chemically inactivated bacteria or yeast and mostly polymer microparticles. The versatility and reproducibility of the latter has allowed monitoring the influence of parameters such as size and surface chemistry, shape and mechanical properties of the target on the mechanism of phagocytosis.
Antigens present on the surface of endogenous targets, such as erythrocytes, cancerous and apoptotic cells exhibit a lateral mobility that can’t be mimicked by adhesive proteins adsorbed on the solid surface of common polymeric targets. However, in the immunological context, phenomena as for example the formation of the immune synapse or the fabrication of artificial antigen presenting cells, require the ability of antigens or receptors to be laterally mobile at the interface of the target.
We confirmed that IgG-coated emulsions are synthetic particles allowing the lateral diffusion of the adhesive molecules bound to their surface that show a different behavior than solid particles in the phagocytic context, both in term of quantitative internalization and microscopic features.
In the project, we intend to first (WP1) study the influence of physico-chemical parameters of the particles (size, interfacial tension and viscosity) on the phagocytosis efficiency to rationalize the characterization of the engineered droplets; and second (WP2) to analyze the the macrophage response cascade occurring following the adhesion, the internalization and the phagosome maturation, by assaying the dynamic of the IgGs, their traffic, the respiratory burst and reactive oxygen species production, and finally the fate of the droplets on a long timescale.