Stigmasterol is a phytosterol that presents pharmacologic properties. However, its anti-inflammatory mechanism and antinociceptive effect are not yet elucidated. Thus, the present study aimed to investigate the anti-inflammatory and antinociceptive activities of stigmasterol and its mechanism of action in mice. The antinociceptive activity was assessed by the acetic acid-induced writhing test, formalin test, and hot plate test. The anti-inflammatory activity was investigated by carrageenan-induced peritonitis and paw edema induced by arachidonic acid. The involvement of glucocorticoid receptors in the mechanism of stigmasterol anti-inflammatory action was investigated by molecular docking, also by pretreating mice with RU-486 (glucocorticoid receptor antagonist) in the acetic acid-induced writhing test. Mice motor coordination was evaluated by the rota-rod test and the locomotor activity by the open field test. The lowest effective dose of stigmasterol was standardized at 10 mg/kg (p.o.). It prevented abdominal writhes and paw licking, but it did not increase the latency time in the hot plate test, suggesting that stigmasterol does not show an antinociceptive effect in response to a thermal stimulus. Stigmasterol decreased leukocyte infiltration in peritonitis assay and reduced paw edema elicited by arachidonic acid. Molecular docking suggested that stigmasterol interacts with the glucocorticoid receptor. Also, RU-486 prevented the effect of stigmasterol in the acetic-acid abdominal writhing test, which might indicate the contribution of glucocorticoid receptors in the mechanism of stigmasterol action. Stigmasterol reduced the number of crossings but did not impair mice's motor coordination. Our results show that stigmasterol presents anti-inflammatory effects probably mediated by glucocorticoid receptors.

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