Figure 2.

Complement component 5a (C5a) generated by the alternative pathway of complement during coagulation is responsible for priming of polymorphonuclear leucocytes (PMN) by products of coagulation. (a) Screen of potential PMN priming agents that could be responsible for the observed PMN priming by products of coagulation. PMN were preincubated with the specified agent(s), followed by addition of luminol + horseradish peroxidase (HRP) and then treatment with 100 nM N‐formylmethionine‐leucyl‐phenylalanine (fMLP). Total reactive oxygen species (ROS) production over 10 min was measured, with results normalized to an internal positive priming control [20 ng/ml tumour necrosis factor (TNF)‐α] and compared with fMLP alone (i.e. no priming agent, ‘none’) for reference. (b–f) PMN priming by products of coagulation [50% (v/v)] measured by total ROS generation in response to fMLP in the presence or absence of C5aR blocking antibody [versus immunoglobulin (Ig)G control] or the C5aR antagonist W‐54011 (versus vehicle control). Results normalized to an internal positive priming control (20 ng/ml TNF‐α). (g–i) Enzyme‐linked immunosorbent assay (ELISA) for complement fragments C5a, Bb, and C4d from n = 4 healthy volunteers. Results reported as mean ± standard error of the mean (s.e.m.). Significance was set at P ≤ 0·05. *P ≤ 0·05; **P ≤ 0·01; ***P ≤ 0·001; ****P ≤ 0·0001. FP = fibrinopeptides; PF1+2 = prothrombin fragment 1+2; AP = activation peptide.