Beschreibung:
Abstract The alternative pathway (AP) of complement is an essential immune effector mechanism. Properdin (P), the only positive regulator of the AP, is required for stabilization of AP enzymatic convertases. P also binds to certain surfaces, in vitro, activating complement by recruiting a de novo C3bBb convertase. P is present in serum as dimers (P2), trimers (P3) and tetramers (P4) in a 1:2:1 ratio, with tetramers the most active, followed by trimers and dimers. Activated neutrophils release properdin from secondary granules into the cellular microenvironment. However, the distribution of neutrophil-secreted P oligomers (which would correlate with P function), and whether this distribution modulates local inflammatory processes, are unknown. We developed a novel ELISA-based assay that detects functional differences between P oligomers, based on the immobilization of P and assessment of its ability to generate C3 convertases leading to C3b deposition. Pure P2, P3, and P4 were functionally distinct in this assay and serum-derived P behaved similar to pooled P2-P4, as expected. Surprisingly, P from PMA-activated neutrophil supernatants had significantly lower activity than serum P (p<0.0001), and varied between healthy donors. To understand whether serum is required for enhancing P function (i.e. by allowing P polymerization), neutrophil supernatants were pre-incubated with P-depleted serum, followed by measurement of P function by ELISA. No increase in neutrophil-derived P function was observed. Additional studies are underway to understand the mechanisms involved in the lower function of neutrophil-derived P, with the aim of contributing to understanding molecular mechanisms of AP regulation in inflammatory microenvironments.