Defined particle ligands trigger specific defense mechanisms of macrophages.

Phagocytosis is a receptor-mediated process for sequestration and inactivation of infectious microbes. It can be triggered by microbial surface compounds or particle-attached host proteins. We monitored the effector functions of murine bone marrow-derived macrophages (BMMs) in response to polystyrene-streptavidin beads coated with the defined ligands IgG1, ß-glucan, mannan, complement factors C1q or iC3b, or fibronectin (FN). Cell-autonomous effector mechanisms (uptake, phagosome maturation, cytokine responses and killing activity) were differentially triggered. All particle-ligand complexes stimulated the release of nitric oxide, but only beads coated with IgG, complement factors or FN caused production of superoxide. Beads coated with C1q, iC3b or FN strongly stimulated the secretion of pro-inflammatory TNF-α, IL-6, and IL-1ß and also of anti-inflammatory IL-10. Escherichia coli coated with C1q, iC3b or FN was killed much less efficiently than with any of the other ligands, depending on the presence of IL-10 activity. This indicated an important role of IL-10 as regulator of cell-autonomous immune functions of macrophages. Our data show that defined ligands on microbial surfaces are interesting candidates to activate innate defense mechanisms selectively and specifically.

Lipid microdomain-dependent macropinocytosis determines compartmentation of Afipia felis.

Phagocytic compartments are specialized endocytic organelles and usually mature along the degradative pathway into phagolysosomes. The rare human pathogen Afipia felis localizes to a compartment that is different from canonical phagocytic compartments. Here, we present evidence that internalization of Afipia by macrophages and unusual phagosome development are considerably decreased by attachment of cholera toxin B subunit to macrophage ganglioside GM1 or by extraction or oxidation of plasma membrane cholesterol. Amiloride (an inhibitor of Na(+)/H(+) exchanger and macropinocytosis) strongly inhibited uptake of A. felis at a late step, i.e. the closure of macropinocytic structures rather than the production of membrane ruffles. Ultrastructural evidence showed that A. felis was taken up by macrophages via macropinocytosis. In contrast, A. felis opsonized with a monoclonal IgG antibody was ingested by a zipper-like mechanism, resulting in normal phagosome maturation. Hence, while the preferred path of A. felis uptake is dependent on the integrity of lipid microdomains and on macropinocytosis, and while this uptake leads to an unusual phagosome and to intracellular survival of A. felis, those bacteria that enter using Fcgamma receptors are delivered to a late endocytic compartment.