High expression of IL-21 and/or IL-21R has been described in T cell-mediated inflammatory diseases characterized by defects of counterregulatory mechanisms. CD4+ CD25+ regulatory T cells (Treg) are a T cell subset involved in the control of the immune responses. A diminished ability of these cells to inhibit T cell activation has been documented in immune-inflammatory diseases, raising the possibility that inflammatory stimuli can block the regulatory properties of Treg. We therefore examined whether IL-21 controls CD4+ CD25+ T cell function. We demonstrate in this study that IL-21 markedly enhances the proliferation of human CD4+ CD25− T cells and counteracts the suppressive activities of CD4+ CD25+ T cells on CD4+ CD25− T cells without affecting the percentage of Foxp3+ cells or survival of Treg. Additionally, CD4+ CD25+ T cells induced in the presence of IL-21 maintain the ability to suppress alloresponses. Notably, IL-21 enhances the growth of CD8+ CD25− T cells but does not revert the CD4+ CD25+ T cell-mediated suppression of this cell type, indicating that IL-21 makes CD4+ T cells resistant to suppression rather than inhibiting CD4+ CD25+ T cell activity. Finally, we show that IL-2, IL-7, and IL-15, but not IL-21, reverse the anergic phenotype of CD4+ CD25+ T cells. Data indicate that IL-21 renders human CD4+ CD25− T cells resistant to Treg-mediated suppression and suggest a novel mechanism by which IL-21 could augment T cell-activated responses in human immune-inflammatory diseases.