SIZE OF IGG-OPSONIZED PARTICLES DETERMINES MACROPHAGE RESPONSE DURINGINTERNALIZATION

It is generally assumed that particles >1 mu m elicit a phagocytic res ponse. To determine whether this is the case, we examined the uptake a nd transport of IgG-opsonized polystyrene beads of defined size, rangi ng from 0.2 to 3 mu m, by mouse bone marrow-derived macrophages. The k inetics of opsonized bead internalization were comparable for each of the different beads examined. We used rhodamine phalloidin to examine particle-induced assembly of F-actin phagocytic cups by fluorescence m icroscopy. Phagocytic cup formation was size dependent in a nonlinear fashion. Less than 30% of 0.2- to 0.75-mu m particles and greater than 80% of 2- and 3-mu m particles were associated with F-actin. Cells tr eated with 0.25 mu m cytochalasin D showed decreased phagocytic cup fo rmation and a linear decrease in bead uptake as a function of particle surface area. In contrast, potassium depletion, which preferentially inhibits clathrin-mediated endocytosis, was more effective at inhibiti ng the uptake of smaller beads. Thus, with increasing particle size, I gG-opsonized particle uptake became less clathrin dependent and more a ctin dependent. The kinetics of ligand delivery to lysosomes was measu red using an immunoprecipitation assay based on the intermixing of int ernalized anti-dinitrophenol (DNP) IgG with DNP-derivitized beta-glucu ronidase (DNP-beta-glu) incorporated into lysosomes, Soluble mannosyla ted anti-DNP IgG: was delivered to lysosomes after an 8-min lag period . The kinetics of anti-DNP IgG-opsonized beads showed a size-dependent response, where beads sized 0.2, 0,5, and 0.75 mu m showed a lag peri od prior to delivery to lysosomes. In contrast, beads 1.0 mu m or larg er showed no lag in delivery to lysosomes. Since beads that had no lag in delivery to lysosomes also showed high levels of phagocytic cup fo rmation, this suggests that phagocytic cups may be important in the ra pid delivery of internalized particles to lysosomes. (C) 1998 Academic Press.