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Summary: The interaction of the human FcγRIIA with immune complexes (ICs) promotes neutrophil activation and thus must be tightly controlled to avoid damage to healthy tissue. Here, we demonstrate that a fungal-derived soluble β-1,3/1,6-glucan binds to the glycosphingolipid long-chain lactosylceramide (LacCer) to reduce FcγRIIA-mediated recruitment to immobilized ICs under flow, a process requiring high-affinity FcγRIIA-immunoglobulin G (IgG) interactions. The inhibition requires Lyn phosphorylation of SHP-1 phosphatase and the FcγRIIA immunotyrosine-activating motif. β-glucan reduces the effective 2D affinity of FcγRIIA for IgG via Lyn and SHP-1 and, in vivo, inhibits FcγRIIA-mediated neutrophil recruitment to intravascular IgG deposited in the kidney glomeruli in a glycosphingolipid- and Lyn-dependent manner. In contrast, β-glucan did not affect FcγR functions that bypass FcγR affinity for IgG. In summary, we have identified a pathway for modulating the 2D affinity of FcγRIIA for ligand that relies on LacCer-Lyn-SHP-1-mediated inhibitory signaling triggered by β-glucan, a previously described activator of innate immunity.

Properdin (P) is a positive regulatory protein that stabilizes the C3 convertase and C5 convertase of the complement alternative pathway (AP). Several studies have suggested that properdin can bind directly to the surface of certain pathogens regardless of the presence of C3bBb. Saprophytic Leptospira are susceptible to complement-mediated killing, but the interaction of properdin with Leptospira spp. has not been evaluated so far. In this work, we demonstrate that properdin present in normal human serum, purified properdin, as well as properdin oligomers P2, P3, and P4, interact with Leptospira. Properdin can bind directly to the bacterial surface even in the absence of C3b. In line with our previous findings, AP activation was shown to be important for killing non-pathogenic L. biflexa, and properdin plays a key role in this process since this microorganism survives in P-depleted human serum and the addition of purified properdin to P-depleted human serum decreases the number of viable leptospires. A panel of pathogenic L.interrogans recombinant proteins was used to identify putative properdin targets. Lsa30, an outer membrane protein from L. interrogans, binds to unfractionated properdin and to a lesser extent to P2-P4 properdin oligomers. In conclusion, properdin plays an important role in limiting bacterial proliferation of non-pathogenic Leptospira species. Once bound to the leptospiral surface, this positive complement regulatory protein of the AP contributes to the formation of the C3 convertase on the leptospire surface even in the absence of prior addition of C3b.

Deposited immune complexes (IC) promote neutrophil recruitment, but the fine tuning of this process is still unclear. Here the authors show that the cis interaction of the IC receptor, FcγRIIA and CD18 integrin, Mac-1, on the neutrophil surface modulates neutrophil adhesion, with FcγRIIA sialylation specifically implicated in this interaction.

Neutrophils are critical in preventing the transition of acute peritoneal infection to sepsis. Here the authors show in three mouse models of peritonitis that neutrophils enter the abdominal cavity via high endothelial venules of the greater omentum, and characterize adhesion molecules involved.

Platelet/granulocyte aggregates (PGAs) increase thromboinflammation in the vasculature, and PGA formation is tightly controlled by the complement alternative pathway (AP) negative regulator, Factor H (FH). Mutations in FH are associated with the prothrombotic disease atypical hemolytic uremic syndrome (aHUS), yet it is unknown whether increased PGA formation contributes to the thrombosis seen in patients with aHUS. Here, flow cytometry assays were used to evaluate the effects of aHUS-related mutations on FH regulation of PGA formation and characterize the mechanism. Utilizing recombinant fragments of FH spanning the entire length of the protein, we mapped the regions of FH most critical for limiting AP activity on the surface of isolated human platelets and neutrophils, as well as the regions most critical for regulating PGA formation in human whole blood stimulated with thrombin receptor-activating peptide (TRAP). FH domains 19–20 were the most critical for limiting AP activity on platelets, neutrophils, and at the platelet/granulocyte interface. The role of FH in PGA formation was attributed to its ability to regulate AP-mediated C5a generation. AHUS-related mutations in domains 19–20 caused differential effects on control of PGA formation and AP activity on platelets and neutrophils. Our data indicate FH C-terminal domains are key for regulating PGA formation, thus increased FH protection may have a beneficial impact on diseases characterized by increased PGA formation, such as cardiovascular disease. Additionally, aHUS-related mutations in domains 19–20 have varying effects on control of TRAP-mediated PGA formation, suggesting that some, but not all, aHUS-related mutations may cause increased PGA formation that contributes to excessive thrombosis in patients with aHUS.