Background Rheumatoid arthritis (RA) is a debilitating, chronic autoimmune polyarthritis, characterized by synovial hypertrophy and inflammatory cell infiltration. Although the pathogenesis of RA is not understood clearly, altered oxidative stress is considered to play a critical role during the development and perpetuation of the disease. Although the pathogenesis of RA is not understood clearly, altered oxidative stress is considered to play a critical role during the development and perpetuation of the disease. Over the course of evolution, defensive mechanisms against oxidative stress have been developed. One major mechanism in the cellular defense against oxidative stress is activation of the nuclear-related factor 2 (Nrf2), a key transcription factor that contributes to the maintenance of cellular redox homeostasis. One of the major stress-responsive players regulated by Nrf2 is the antioxidant enzyme heme oxygenase-1 (HO-1). Nrf2 signaling has received recent attention as a participant in the pathogenesis of RA. Leflunomide is an oral drug that inhibits de novo pyrimidine synthesis by inhibiting dihydroorotate dehydrogenase. Leflunomide exerts immunoregulatory properties and is useful in RA treatment. Despite the clinical usefulness of the drug, the underlying mechanisms of the drug remain elusive.
Objectives We investigated the antiarthritis effects and mechanisms of action of the active metabolite of leflunomide, A77 1726, in interleukin-1 receptor antagonist-knockout mice (IL-1Ra-KO mice).
Methods To assess the therapeutic effect of A77 1726 on symptom severity in the IL-1Ra-KO mouse model, 14–15-week-old male IL-1Ra-KO mice were treated with 10 mg/kg or 30 mg/kg A77 1726. On day 42 after the first A77 1726 injection, the spleen was removed from each mouse in the different groups. Histologic analysis of the joints was performed using Safranin O, toluidine blue, and immunohistochemical staining. Sandwich ELISA kits were used to measure the amounts of IL-1β, TNF-α and malondialdehyde in sera obtained from the mice. The epxression of HO-1 in splenocytes was analyzed by immunostaining and western blotting.
Results A77 1726 reduced the arthritis severity and cartilage destruction. The joints isolated from A77 1726-treated mice showed decreased expression of inducible nitric oxide synthase, nitrotyrosine, TNF-α, and IL-1β. Interestingly, HO-1 expression was significantly higher in splenic CD4+ T cells isolated from A77 1726-treated mice compared with those from vehicle-treated mice. A77 1726 treatment in Jurkat cells significantly increased nuclear HO-1 activity. Proteome profiler array revealed that A77 1726 slightly activated the phosphorylation of Akt in Jurkat cells. The ability of A77 1726 to induce HO-1 was diminished when Akt activity was blocked.
Conclusions We suggest that the inhibitory effects of A77 1726 on joint inflammation and oxidative stress occur through Akt-dependent HO-1 induction in CD4+ T cells.
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Disclosure of Interest None declared