Administration of Low-Dose Endotoxin to Healthy Humans Increases C5a Binding to Circulating Neutrophils

We examined the binding of C5a to neutrophils of healthy human volunteers given endotoxin. Male volunteers aged 18-30 years were enrolled. After they fasted overnight, subjects had intravenous catheters placed in the forearm. In 5 volunteers, 3 ng/kg US standard reference endotoxin (lot EC-5, Escherichia coli O113; Bureau of Biologics, US Food and Drug Administration, Bethesda, MD) was given as an intravenous bolus injection; 4 injected with saline under identical conditions served as controls. At the time of injection and 3, 6, and 24 h afterward, heparinized (20 units/mL final concentration) blood was obtained. Neutrophils were isolated by ficoll-hypaque centrifugation followed by dextran sedimentation as previously described. Volunteers who received endotoxin developed neutrophilia that was first evident 2 h after injection. Compared with baseline, the absolute neutrophil count was increased twofold at 3 h and threefold at 6 h after injection. There was no increase in absolute neutrophil count at 24 h. Before the endotoxin injection and 3, 6, and 24 h afterward, the mean numbers of bands constituted 0, 5%, 6%, and 1% of the absolute neutrophil counts, respectively. Compared with baseline, neutrophils from volunteers given endotoxin bound 31% more super(125)I-labeled C5a 3 h after endotoxin injection. In contrast, in neutrophils from volunteers given saline, there were significant decreases in C5a binding. Compared with baseline, neutrophils from saline-injected subjects bound 46% less super(125)I-labeled C5a at 3 h and 51% less at 6 h.