Phagocytosis by macrophages: III. Effects of heat-labile opsonin and poly(L-lysine)

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Abstract
To study the mechanism of action of fresh serum (heat-labile opsonin) and poly(L-lysine) (a non-physiological opsonin) on particle uptake by phagocytes, a kinetic analysis of attachment and ingestion of lipopolysaccharide-coated particles during phagocytosis by mouse peritoneal macrophages was undertaken. The relative attachment rate was obtained from the initial overall rate of phagocytosis. The ingestion rate constant was obtained from the time-course of ingestion of the particles attached to the cell surface into the cytoplasm, using spin-labelled paraffin particles and ascorbate, a membrane-impermeable reducing reagent. Although the lipopolysaccharide-coated particles readily attached to the cell surface, they were ingested to only a small extent. Treatment of the particles with poly(L-lysine) and fresh serum influenced both attachment and ingestion. Poly(L-lysine) greatly increased the attachment rate, whereas fresh serum decreased it. The attachment rate was found to be closely correlated with the difference in surface charge density of the particles caused by such treatments, increasing with the decrease in negative charge density. Attachment of particles treated with fresh serum was strongly dependent on incubation temperature, and was inhibited by treatment of the cells with cytochalasin B or trypsin, whereas attachment of poly(L-lysine)-treated particles was only slightly, if at all, dependent on incubation temperature or treatment. Divalent cations in the incubation medium stimulated attachment of fresh-serum-treated particles. Treatment of the particles with both poly(L-lysine) and fresh serum made it possible for the particles to be ingested into the cytoplasm. The ingestion rate constants (k) for the two types of particles were almost identical (k = 1’2 × 10−1/min at 37°C), and they showed a similar temperature dependence. They were independent of the divalent cations in the incubation medium. Treatment of the cells with cytochalasin B inhibited ingestion. These results suggest that ingestion of the poly(L-lysine)-treated particles results from electrostatic non-specific interaction between the particles and the cell surface, whereas ingestion of the fresh-serum-treated particles results from specific interaction via ‘ligand-specific receptor)’ on the cell surface. Ingestion proceeded independently of such differences in the interaction, though attachment was strongly dependent on the electrostatic environment of the particle surface.

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