<b><i>Background:</i></b> Ouabain (OUA) is a cardiotonic glycoside originally extracted from African plants. It has also been described as an endogenous component in mammals, being released in stress situations mainly by the adrenal gland. OUA has been reported to be capable of inhibiting mitogen-induced lymphocyte proliferation and also affects B and T lymphocytes. <b><i>Objectives:</i></b> The aim of this work is to show the effects of OUA in peripheral T lymphocytes. <b><i>Methods:</i></b> In the in vivo experiments, mice were injected intraperitoneally for 3 consecutive days with RPMI medium (control group) or 0.56 mg/kg of OUA diluted in RPMI medium (OUA group). On the fourth day, spleen or mesenteric lymph nodes were removed. <b><i>Results:</i></b> OUA significantly reduced the number of CD4+ T lymphocytes in the spleen, especially regulatory T cells (Tregs). In vitro OUA did not inhibit the proliferation of CD4⁺T lymphocytes stimulated with anti-CD3 neither was able to induce the apoptosis of CD4⁺ nor Tregs. There was no increase in the number or percentage of T lymphocytes in the mesenteric lymph nodes, suggesting that there was no preferential accumulation of these cells in this organ. Secretion of IL-2 by activated T lymphocytes was decreased by the OUA, explaining at least in part the reduction of Tregs, since this cytokine is involved in the peripheral conversion and maintenance of Tregs. <b><i>Conclusion:</i></b> The impact of this reduction in autoimmune diseases, allergy and cancer as well as the potential use of OUA as a therapeutic approach in tumor treatment still needs more investigation.

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