Figure 5. P₄s increase PGA formation via C5a-mediated enhancement of CD11b expression.

Lepirudin anticoagulated whole blood was incubated with TRAP alone (10μM) or in combination with P₄s (5μg/ml) and anti-complement reagents (anti-properdin - 100μg/ml; PMX53 – 16.7μM; Comp - 50μM). PGA formation (A), C5a in supernatants (B), and CD11b expression (C) were determined as described in Materials and Methods. Results shown in A and C are representative of 1 of 5 independent experiments carried out with different human volunteer blood samples, and in 2 of these 5 experiments C5a was also determined (1 representative experiment shown in B). Two additional C5a ELISAs were carried out using supernatants harvested from experiments carried out under identical conditions, but that did not include duplicate samples for flow cytometry analysis. In the 5 independent experiments, effects of P₄s on PGA formation varied between 2.5-14-fold increases over TRAP alone, depending on the donor (variation not shown). Results are shown as mean and SD of duplicate observations. The data was analyzed by one-way ANOVA with Dunnett’s multiple comparison test against TRAP+P₄. p<0.001 compared to TRAP+P₄ unless indicated as n.s.