Exposure to inflammatory cytokines selectively limits GM-CSF production by induced T regulatory cells.
Bottom Line: Understanding processes that can limit this potentially deleterious effect of Treg cells in a therapeutic setting is therefore important.We show that iTreg cells can produce significant amounts of three proinflammatory cytokines (IFN-γ, GM-CSF and TNF-α) upon secondary TCR stimulation.Furthermore, we show that IL-6 and IL-27 individually, or IL-2 and TGF-β in combination, can mediate the selective loss of GM-CSF production by iTreg cells.
Affiliation: MRC Centre for Inflammation Research, University of Edinburgh, Edinburgh, UK.Show MeSH
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Mentions: iTreg cells were generated by 5 day culture of naive (Foxp3-GFP− CD4+) T cells with anti-CD3, anti-CD28, TGF-β, and IL-2. Highly pure iTreg cells were isolated by FACS-sorting based on Foxp3-GFP expression, (Fig. 1A) and were then restimulated with plate-bound anti-CD3 and anti-CD28 for 72 h. We used cytokine bead arrays to screen for the presence of inflammatory cytokines in supernatants from these secondary cultures. The majority of cytokines tested (IL-2, IL-4, IL-5, IL-6, IL-10, IL-13, IL-17, IL-21, and IL-22) were undetectable (data not shown). However, IFN-γ, TNF-α, and GM-CSF were detectable. Intracellular cytokine staining of restimulated iTreg cells demonstrated the loss of Foxp3 expression, as previously described 16, and confirmed the presence of cells staining positive for IFN-γ, GM-CSF, and TNF-α (Fig.1B–E). The majority of cells were TNF-α+ (Fig. 1D). Production of IFN-γ and GM-CSF was more restricted, and approximately 25% of cells were double positive for IFN-γ and GM-CSF (Fig. 1E). While IFN-γ and GM-CSF staining was seen chiefly in cells that had lost Foxp3 expression, TNF-α was also seen in a clear population that had retained Foxp3.
Affiliation: MRC Centre for Inflammation Research, University of Edinburgh, Edinburgh, UK.