Modulation of neutrophil complement receptor 3 expression by pneumococci

Complement receptor 3 (CR3; CD18/CD11b) plays an important role in the recognition and clearance of Streptococcus pneumoniae (pneumococci) by neutrophils. The purpose of the present study was to characterize the modulation of CR3 surface expression on neutrophils exposed to pneumococci and to assess its functional significance. CR3 was detected with fluorescent phytoerythrin-labelled anti-CR3 (CD11b) antibodies, quantified with a fluorescence cell counter (FACS) and localized by confocal fluorescence microscopy. Uptake of fluorescent FITC-labelled pneumococci was quantified by FACS. Whole blood from healthy volunteers was exposed at 37 °C to killed whole type III Streptococcus pneumoniae (KSP; 10⁸/ml) or to a positive control (Escherichia coli lipopolysaccharide) that enhanced CR3 surface expression on neutrophils to a comparable extent. Varying the concentration of KSP between 10⁵ and 10⁸ organisms/ml progressively augmented CR3 surface expression measured at 1 h, whereas the response declined at 10⁹/ml. The diminished response to 10⁹ KSP/ml proved to be time-dependent, with surface CR3 up-regulated maximally within 5 min, and down-regulated thereafter. Labelling of CR3 during exposure demonstrated accelerated receptor sequestration, and confocal fluorescence microscopy demonstrated internalized CR3. Cooling to 16 °C, to inhibit the up-regulation of CR3 surface expression, also inhibited the uptake of FITC-labelled KSP and morphological changes. Accelerated down-regulation of surface CR3 expression by exposure to 10⁹/ml unlabelled KSP diminished the uptake of labelled KSP added subsequently. In contrast, lipopolysaccharide-induced up-regulation of CR3 expression increased the uptake of labelled KSP. Together, these experiments reveal dynamic modulation of CR3 expression on the surface of neutrophils exposed to pneumococci and a functional correlate of this modulation. Thus neutrophil expression of CR3 changes dynamically in response to exposure of neutrophils to progressively higher concentrations of pneumococci, conditions that mimic early neutrophil recruitment to densely infected lung tissue in acute pneumococcal pneumonia.