RU-521

Objective To explore the mechanism of the cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) axis in mediating neutrophil extracellular trap (NET) formation in gouty arthritis (GA). Methods Thirty-two C57BL/6J mice were randomly divided into four groups: Sham group, GA group, GA+RU.521 group and GA+RU.521+MSA-2 group, with 8 mice in each group. While the Sham group was injected with PBS buffer into the ankle joints, the other groups were injected with monosodium urate (MSU) crystals to induce GA. Neutrophils were isolated from peripheral blood samples obtained from healthy volunteers and then were divided into four groups: control group (Con), MSU group, MSU+RU.521 group and MSU+RU.521+MSA-2 group. The Con group was exposed to PBS buffer for 24 hours, while the other groups were exposed to 40 μg/mL MSU for 24 hours. The expression of citrulline histone H3 (CitH3) and myeloperoxidase (MPO) in ankle joints and neutrophils was analyzed using immunofluorescence staining. Western blot was used to analyze the protein expression of the NLRP3 pathway and the cGAS-STING signaling pathway in ankle joint tissues and neutrophils. The levels of tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β) and IL-6 in ankle joint tissues and neutrophil supernatants were determined by enzyme-linked immunosorbent assay (ELISA). Results Compared with the Sham group, the GA group exhibited significantly elevated levels of inflammatory mediators (IL-1β, TNF-α and IL-6), increased expressions of NLRP3, ASC and c-CASP1 and enhanced fluorescence intensity of MPO and CitH3 in the ankle joint tissues. However, treatment with RU.521 effectively reversed these changes. Compared with GA+RU.521 group, the GA+RU.521+MSA-2 group exhibited significantly elevated levels of IL-1β, TNF-α and IL-6, increased expressions of NLRP3, ASC and c-CASP1 and enhanced fluorescence intensity of MPO and CitH3 in the ankle joint tissues. Compared with the Con group, the MSU group exhibited significantly increased expressions of NLRP3, ASC, c-CASP1 and enhanced fluorescence intensity of MPO and CitH3 in neutrophils. The expressions of NLRP3, ASC and c-CASP1 and the fluorescence intensity of MPO and CitH3 in neutrophils in MSU+RU.521 group were lower than those in MSU group. Compared with the MSU+RU.521 group, the MSU+RU.521+MSA-2 group exhibited significantly increased expressions of NLRP3, ASC, c-CASP1, and STING and enhanced fluorescence intensity of MPO and CitH3 in neutrophils. Conclusion MSU crystal may promote the formation of NETs and induce inflammatory reaction by activating cGAS-STING signaling pathway. Therefore, targeting cGAS-STING signaling pathway may be a promising strategy for anti-GA therapy.