Complement alternative pathway acts as a positive feedback amplification of neutrophil activation

Abstract

Complement alternative pathway plays an important, but not clearly understood, role in neutrophil-mediated diseases. We here show that neutrophils themselves activate complement when stimulated by cytokines or coagulation-derived factors. In whole blood, tumor necrosis factor/ formyl-methionyl-leucyl-phenylalanine or phorbol myristate acetate resulted in C3 fragments binding on neutrophils and monocytes, but not on T cells. Neutrophils, stimulated by tumor necrosis factor, triggered the alternative pathway on their surface in normal and C2-depleted, but not in factor B-depleted serum and on incubation with purified C3, factors B and D. This occurred independently of neutrophil proteases, oxidants, or apoptosis. Neutrophil-secreted properdin was detected on the cell surface and could focus "in situ" the alternative pathway activation. Importantly, complement, in turn, led to further activation of neutrophils, with enhanced CD11b expression and oxidative burst. Complement-induced neutrophil activation involved mostly C5a and possibly C5b-9 complexes, detected on tumor necrosis factor-and serum-acti- IntroductionNeutrophils and the complement alternative pathway (AP) are major effectors of cell-mediated and humoral innate immunity. The analysis of complement knockout mice, in experimental inflammatory diseases, shed new light on the participation of complement AP in neutrophil-mediated diseases, such as rheumatoid arthritis, 1 membranoproliferative glomerulonephritis, 2 ischemia-reperfusion injury, 3 and, more recently, antineutrophil cytoplasmic autoantibody (ANCA)-associated small-vessel vasculitis. 4 In an experimental model of ANCA-associated small-vessel vasculitis, the unexpected observation that factor B-deficient mice were protected from the disease, whereas C4-deficient mice were not, 4 revealed the alternative complement pathway as a new partner of this neutrophilmediated disease. It was thus reasonable to propose that neutrophils could participate in the activation of complement AP.If the activation of neutrophils by complement fragments, such as C3a or C5a, is well known, data on complement triggering by neutrophils are scarce. Neutrophil proteases and oxidants have been reported to activate complement in cell-free systems, 5-7 and supernatants of ANCA-activated neutrophils were shown to release unknown complement activating factors. 4 We investigated the ability of the neutrophil surface to activate complement and deposit active complement fragments on their plasma membrane.Complement activation on blood cells is kept under control by fluid phase regulators (ie, plasma factor H and C4b-binding protein) and by cell membrane regulators (ie, decay accelerating factor DAF [CD55] and membrane cofactor protein MCP [CD46]). These regulators prevent the formation and accelerate the decay of C3bBb and C4b2a alternative and classic C3-convertases and act as cofactors for factor I-mediated C3b and C4b proteolysis. Neutrophils also express CR1 (CD35), which is the only cofactor allowing factor I to fully degrade ...

Properdin binding to complement activating surfaces depends on initial C3b deposition

“…Thus, MPO could be the molecule that links properdin binding to the surface of activated neutrophils, as has been reported previously (49). By studying properdin binding to immobilized MPO, the authors proposed that MPO promotes complement activation by directly binding properdin (49).…”

“…Thus, MPO could be the molecule that links properdin binding to the surface of activated neutrophils, as has been reported previously (49). By studying properdin binding to immobilized MPO, the authors proposed that MPO promotes complement activation by directly binding properdin (49). We question those findings, however, because they were based on a detection system using HRP-labeled antibodies in which we found it impossible to distinguish substrate turnover caused by peroxidase activity by the antibody-linked HRP and that caused by the immobilized MPO.…”

Feedback Amplification of Neutrophil Function

“…Such conditioned supernatants were then able to prime neutrophils for respiratory burst in a C5a-receptor-dependent manner, indicating existence of a feedback amplification loop [68]. Neutrophil activation by various stimuli also triggered the alternative pathway of complement activation which in turn resulted in neutrophil activation primarily through C5a generation [69].…”

Production of complement components by cells of the immune system

“…Stimulated PMNs were reported to secrete C3 with an intact thioester and it was speculated that the PMN-derived proteases, which are also released locally, can subsequently activate this newly secreted C3 [35,36]. Neutrophils not only store C3 but also FP, the only complement regulator that enhances complement activation, which can be secreted upon stimulation [30,35,37]. Besides the release of C3 and FP, FB was also demonstrated to be secreted upon activation of neutrophils [38].…”