Objective: Recent studies suggest that crystals of monosodium urate (MSU), deposited in joints of patients with acute gouty arthritis, activate the NALP3 inflammasome. In the present study we have investigated whether production of pro-inflammatory cytokines by crystals was exacerbated during co-stimulation with TLR ligands.
Methods: Mononuclear cells of 22 healthy donors were stimulated by various concentrations of MSU crystals in the absence or presence of lipopolysaccharide (LPS), Pam3Cys and flagellin. Production of tumour necrosis factor-alpha (TNFα), interleukin (IL)-1β and IL-6, as well as the intracellular concentrations of pro-IL-1β were measured by ELISA. mRNA transcripts of TNFα and IL-1β were assessed by real-time PCR. Stimulation experiments were also performed with PBMCs of one patient carrying a NALP3 mutation.
Results: MSU induced a moderate release of IL-1β and IL-6, but not of TNFα. Urate crystals amplified IL-1*[beta] production stimulated by the TLR4 ligand LPS, while no synergy was apparent for IL-6 production. In addition, no synergy between urate crystals and Pam3Cys (TLR2-ligand) or flagellin (TLR5-ligand) was apparent. The synergy between urate crystals and LPS was directed at the level of the NALP3 inflammasome, as it was present only when active IL-1β was measured, but not at the level of IL-1 mRNA or pro-IL-1β. The synergy between LPS and MSU crystals ceased to exist in the presence of a caspase-1 inhibitor.
Conclusions: MSU crystals act in synergy with LPS for the induction of enhanced release of IL-1β. Increased cleavage of pro-IL-1β by urate-activated caspase-1 is proposed as the underlying mechanism.