17–19 However, several studies suggest that nTreg do not universally suppress all T helper cell subsets to the same extent. In newborns, human thymus-derived nTreg strongly suppress Th1 cells but not Th2 cells, and similar properties have been ascribed to nTreg in mice.20,21 Additionally, nTreg isolated from peripheral human blood have been shown to strongly suppress the production and secretion of interferon-γ (IFN-γ), IL-2

and IL-4, but not that of IL-10, in an allogenic model.22 Thus, diurnal changes in the Th1/Th2 balance could also be Quizartinib purchase regulated by the diurnal rhythm of nTreg-suppressive activity. We previously demonstrated that the suppression of CD4+ CD25− T-cell proliferation by nTreg followed a sleep-dependent rhythm.23 However, whether

this suppressive rhythm of nTreg affects the proliferation and cytokine secretion of Th1, Th2 and Th17 cells to the same extent is not yet clear. Furthermore, the signal-transduction mechanisms by which nTreg mediate their suppressive function in responder T cells (Tres) are largely unknown in humans. One possible mechanism of diurnal changes in the Th1/Th2/Th17 balance could be the hormonal priming of T cells and/or nTreg in vivo through Epigenetics inhibitor the diurnal secretion of hormones with known immunomodulatory effects, such as prolactin, growth hormone, cortisol, noradrenalin and melatonin.8,24–31 To address the vital question of whether nTreg or hormones regulate diurnal changes in the Th1/Th2/Th17 balance, and whether Th1, Ureohydrolase Th2 and Th17 cell activity follows a diurnal rhythm, we investigated the activity of the Th1/Th2/Th17 cells and their regulation by nTreg. We were able to demonstrate that nTreg suppressed IFN-γ, IL-2 and tumour necrosis factor-α (TNF-α), but not IL-4, IL-6, IL-10, or IL-17A. The suppression of IL-2 was reduced if nTreg-associated CD25 was inhibited. Highly purified nTreg secreted IL-6, IL-10 and IL-17, but not IL-2, IL-4, IFN-γ or TNF-α. Furthermore, we observed that secretion

of the cytokines IL-2, IFN-γ, TNF-α and IL-10 by naïve CD4+ T cells follows a diurnal rhythm. Multiple regression analysis, as well as subsequent in vitro experiments, suggested that serum levels of cortisol and prolactin contribute to the underlying mechanisms. Taken together, our findings imply that hormones and nTreg contribute to the diurnal secretion of cytokines from T helper cells. Cytokine secretion, and suppression of cytokine secretion by nTreg, was analyzed for Th1 (IFN-γ), Th2 (IL-4, IL-6) and Th17 (IL-17) cytokines, as well as for the cytokines IL-2, IL-10 and TNF-α. Furthermore, the proliferation of cytokine (IL-2, IL-4, IL-10, IL-17A, IFN-γ, TNF-α)-producing CD4+ CD25− Tres was investigated. For these analyses, T cells were isolated from blood samples taken from healthy male donors at 08:30 hr.