Combination of inorganic nitrate and vitamin C prevents collagen-induced arthritis in rats by inhibiting pyroptosis

Rheumatoid arthritis (RA) is a chronic autoimmune inflammatory disease characterized by synovial inflammation, cartilage damage, and bone loss. Although effective treatments are currently lacking, early interventions hold promise for alleviating RA symptoms. Inorganic nitrates and vitamin C (VC) are essential bioactive substances abundant in fruits and vegetables. Notably, nitrates and VC exhibit synergistic effects in a series of physiological and pathological conditions. In this study, we aim to examine the combination of nitrate and VC for preventing RA in a collagen-induced arthritis (CIA) rat model. Nitrate partly reduced foot swelling and arthritis scores and was more effective when combined with VC. Histopathological and radiological analyses revealed that nitrate + VC treatment alleviated synovial hyperplasia and bone loss. Additionally, nitrate + VC lowered the levels of TNF-α and IL-1β in serum as well as synovial tissue, decreased the expression of NF-κB and reduced the number of macrophages in synovial tissue. Compared to the CIA group, nitrate + VC decreased the levels of NLRP3 and GSDMD in macrophages, thus inhibiting pyroptosis. According to <i>in vitro</i> experiments, nitrate inhibited the activation of the NLRP3/caspase-1/GSDMD pathway in macrophages by conversion into nitrite. VC reduced the expression and phosphorylation of NF-κB in macrophages and thus reduced the expression levels of TNF-α and IL-1β. Therefore, nitrate/nitrite and VC may exert synergistic effects by blocking the interaction between NF-κB and NLRP3, further alleviating the inflammation and pyroptosis of macrophages, which provides a new strategy for RA prevention.