C5a Regulates IL-1ß Production and Leukocyte Recruitment in a Murine Model of Monosodium Urate Crystal-Induced Peritonitis.

Gouty arthritis results from the generation of monosodium urate (MSU) crystals within joints. These MSU crystals elicit acute inflammation characterized by massive infiltration of neutrophils and monocytes that are mobilized by the pro-inflammatory cytokine IL-1ß. MSU crystals also activate the complement system, which regulates the inflammatory response; however, it is unclear whether or how MSU-mediated complement activation is linked to IL-1ß release in vivo, and the various roles that might be played by individual components of the complement cascade. Here we show that exposure to MSU crystals in vivo triggers the complement cascade, leading to the generation of the biologically active complement proteins C3a and C5a. C5a, but not C3a, potentiated IL-1ß and IL-1α release from LPS-primed MSU-exposed peritoneal macrophages and human monocytic cells in vitro; while in vivo MSU-induced C5a mediated murine neutrophil recruitment as well as IL-1ß production at the site of inflammation. These effects were significantly ameliorated by treatment of mice with a C5a receptor antagonist. Mechanistic studies revealed that C5a most likely increased NLRP3 inflammasome activation via production of reactive oxygen species (ROS), and not through increased transcription of inflammasome components. Therefore we conclude that C5a generated upon MSU-induced complement activation increases neutrophil recruitment in vivo by promoting IL-1 production via the generation of ROS, which activate the NLRP3 inflammasome. Identification of the C5a receptor as a key determinant of IL-1-mediated recruitment of inflammatory cells provides a novel potential target for therapeutic intervention to mitigate gouty arthritis.

The Syk-NFAT-IL-2 Pathway in Dendritic Cells Is Required for Optimal Sterile Immunity Elicited by Alum Adjuvants.

Despite a long history and extensive usage of insoluble aluminum salts (alum) as vaccine adjuvants, the molecular mechanisms underpinning Ag-specific immunity upon vaccination remain unclear. Dendritic cells (DCs) are crucial initiators of immune responses, but little is known about the molecular pathways used by DCs to sense alum and, in turn, activate T and B cells. In this article, we show that alum adjuvanticity requires IL-2 specifically released by DCs, even when T cell secretion of IL-2 is intact. We demonstrate that alum, as well as other sterile particulates, such as uric acid crystals, induces DCs to produce IL-2 following initiation of actin-mediated phagocytosis that leads to Src and Syk kinase activation, Ca2+ mobilization, and calcineurin-dependent activation of NFAT, the master transcription factor regulating IL-2 expression. Using chimeric mice, we show that DC-derived IL-2 is required for maximal Ag-specific proliferation of CD4+ T cells and optimal humoral responses following alum-adjuvanted immunization. These data identify DC-derived IL-2 as a key mediator of alum adjuvanticity in vivo and the Src-Syk pathway as a potential leverage point in the rational design of novel adjuvants.

A novel human anti-interleukin-1ß neutralizing monoclonal antibody showing in vivo efficacy.

The pro-inflammatory cytokine interleukin (IL)-1ß is a clinical target in many conditions involving dysregulation of the immune system; therapeutics that block IL-1ß have been approved to treat diseases such as rheumatoid arthritis (RA), neonatal onset multisystem inflammatory diseases, cryopyrin-associated periodic syndromes, active systemic juvenile idiopathic arthritis. Here, we report the generation and engineering of a new fully human antibody that binds tightly to IL-1ß with a neutralization potency more than 10 times higher than that of the marketed antibody canakinumab. After affinity maturation, the derived antibody shows a>30-fold increased affinity to human IL-1ß compared with its parent antibody. This anti-human IL-1ß IgG also cross-reacts with mouse and monkey IL-1ß, hence facilitating preclinical development. In a number of mouse models, this antibody efficiently reduced or abolished signs of disease associated with IL-1ß pathology. Due to its high affinity for the cytokine and its potency both in vitro and in vivo, we propose that this novel fully human anti-IL-1ß monoclonal antibody is a promising therapeutic candidate and a potential alternative to the current therapeutic arsenal.

The NLRP3 inflammasome affects DNA damage responses after oxidative and genotoxic stress in dendritic cells.

The NOD-like receptor (NLR) family pyrin domain-containing 3 (NLRP3) inflammasome is a cytoplasmic protein complex that mediates inflammatory responses to a broad array of danger signals. The inflammasome drives caspase-1 activation and promotes secretion of the pro-inflammatory cytokines IL-1ß and IL-18, and might also participate in other cellular processes. Here, we tried to identify new pathways regulated by the NLRP3 inflammasome in murine dendritic cells (DCs) in response to monosodium urate (MSU) crystals. Using a transcriptomic approach, we found that DCs from Nlrp3(-/-) mice responded to MSU with differential expression of genes involved in the DNA damage response and apoptosis. Upon exposure to MSU or other ROS-mobilizing stimuli (rotenone and γ-radiation), DNA fragmentation was markedly ameliorated in Nlrp3(-/-) and casp-1(-/-) DCs compared with WT DCs. Moreover, Nlrp3(-/-) DCs experienced significantly less oxidative DNA damage mediated by ROS. A significant decrease of the expression of several genes involved in double-strand and base-excision DNA repair was observed in WT DCs. Basal DNA repair capacity in WT DCs resulted in activation and stabilization of p53 in vitro and in vivo, which resulted in increased cell death compared with that in Nlrp3(-/-) DCs. These data provide the first evidence for the involvement of the NLRP3 inflammasome in DNA damage responses induced by cellular stress.