Emese Ugor1, Lilla Prenek1, Ramóna Pap1, Gergely Berta2, Dávid Ernszt3, József Najbauer1, Péter Németh1, Ferenc Boldizsár1, Tímea Berki4. 1. Department of Immunology and Biotechnology, Clinical Center, University of Pécs, Pécs H-7624, Hungary. 2. Department of Medical Biology, University of Pécs Medical School, Pécs H-7624, Hungary. 3. Department of Pharmaceutical Biotechnology, School of Pharmacy, University of Pécs, Pécs H-7624, Hungary. 4. Department of Immunology and Biotechnology, Clinical Center, University of Pécs, Pécs H-7624, Hungary. Electronic address: berki.timea@pte.hu.
Abstract
OBJECTIVE: Despite the fact that glucocorticoids (GC) are important therapeutic tools, their effects on regulatory T cells (Treg) are not well defined. The aim of our work was to investigate how GCs influence in vivo the thymic (tTreg) and peripheral Treg (pTreg) differentiation, survival and cytokine production. METHODS: Tregs were detected with flow cytometry in lymphatic organs of 4-6 weeks old BALB/c mice after repeated (2-4days), high-dose in vivo GC treatment using CD4/CD25 cell surface and Foxp3/IL-10/TGFβ/glucocorticoid receptor (GR) intracellular staining. Cytokine, Foxp3, and GR mRNA levels of sorted CD4+CD25high T cells were analyzed using RT-PCR. Foxp3 and GR localization in Treg cells was investigated with confocal microscopy. RESULTS: GC treatment of mice resulted in increased relative tTreg frequency in the thymus, which was due to decreased total thymocyte numbers with unchanged absolute tTreg cell count. In contrast the relative pTreg cell ratio in secondary lymphatic organs decreased or showed no changes after GC treatment, while the absolute number of pTregs decreased. Elevated intracellular IL-10+ and TGFβ+ tTreg and pTreg ratios were measured in GC-treated animals, accompanied with elevated Foxp3 mRNA expression. In addition, GC treatment caused increased TGFβ and IL-35 mRNA expression in CD4+CD25high+ splenic and elevated IL-10 mRNA level in thymic tTregs. GR expression of thymic tTreg cells was lower than in pTregs. GC treatment caused an opposite change in GR levels, elevating GR in tTregs but decreasing it in pTregs. We observed a nuclear localization of GR in both tTregs and pTregs, which showed high colocalization (∼60%) with Foxp3 transcription factor. These data suggest an interaction of these two transcription factors with further increase due to GC treatment in splenic pTregs. CONCLUSION: Our data show selective survival of tTregs and elevated production of immunosuppressive cytokines by Treg cells after GC treatment, which may contribute to the immunosuppressive effects of GCs.
OBJECTIVE: Despite the fact that glucocorticoids (GC) are important therapeutic tools, their effects on regulatory T cells (Treg) are not well defined. The aim of our work was to investigate how GCs influence in vivo the thymic (tTreg) and peripheral Treg (pTreg) differentiation, survival and cytokine production. METHODS: Tregs were detected with flow cytometry in lymphatic organs of 4-6 weeks old BALB/c mice after repeated (2-4days), high-dose in vivo GC treatment using CD4/CD25 cell surface and Foxp3/IL-10/TGFβ/glucocorticoid receptor (GR) intracellular staining. Cytokine, Foxp3, and GR mRNA levels of sorted CD4+CD25high T cells were analyzed using RT-PCR. Foxp3 and GR localization in Treg cells was investigated with confocal microscopy. RESULTS: GC treatment of mice resulted in increased relative tTreg frequency in the thymus, which was due to decreased total thymocyte numbers with unchanged absolute tTreg cell count. In contrast the relative pTreg cell ratio in secondary lymphatic organs decreased or showed no changes after GC treatment, while the absolute number of pTregs decreased. Elevated intracellular IL-10+ and TGFβ+ tTreg and pTreg ratios were measured in GC-treated animals, accompanied with elevated Foxp3 mRNA expression. In addition, GC treatment caused increased TGFβ and IL-35 mRNA expression in CD4+CD25high+ splenic and elevated IL-10 mRNA level in thymic tTregs. GR expression of thymic tTreg cells was lower than in pTregs. GC treatment caused an opposite change in GR levels, elevating GR in tTregs but decreasing it in pTregs. We observed a nuclear localization of GR in both tTregs and pTregs, which showed high colocalization (∼60%) with Foxp3 transcription factor. These data suggest an interaction of these two transcription factors with further increase due to GC treatment in splenic pTregs. CONCLUSION: Our data show selective survival of tTregs and elevated production of immunosuppressive cytokines by Treg cells after GC treatment, which may contribute to the immunosuppressive effects of GCs.
Authors: Lourdes Rocamora-Reverte; Selma Tuzlak; Laura von Raffay; Marcel Tisch; Heidi Fiegl; Mathias Drach; Holger M Reichardt; Andreas Villunger; Denise Tischner; G Jan Wiegers Journal: Front Immunol Date: 2019-03-18 Impact factor: 7.561