Cutting Edge: Suppression of GM-CSF Expression in Murine and Human T Cells by IL-27:suppression of GM-CSF expression in murine and human T cells by IL-27

GM-CSF is a potent pro-inflammatory cytokine that plays a pathogenic role in the CNS inflammatory disease, EAE. As IL-27 ameliorates EAE, we hypothesised that IL-27 suppresses GM-CSF expression by T cells. We found that IL-27 suppressed GM-CSF expression in CD4(+) and CD8(+) T cells in splenocyte and purified T cell cultures. IL-27 suppressed GM-CSF in Th1, but not Th17 cells. IL-27 also suppressed GM-CSF expression by human T cells in non-polarised and Th1 but not Th17 polarised PBMC cultures. In vivo, IL-27p28 deficiency resulted in increased GM-CSF expression by CNS infiltrating T cells during Toxoplasma gondii infection. While in vitro suppression of GM-CSF by IL-27 was independent of IL-2 suppression, IL-10 up-regulation or SOCS3 signalling, we observed that IL-27-driven suppression of GM-CSF was STAT1 dependent. Our findings demonstrate that IL-27 is a robust negative regulator of GM-CSF expression in T cells which likely inhibits T cell pathogenicity in CNS inflammation

IFNγ Signaling Endows DCs with the Capacity to Control Type I Inflammation during Parasitic Infection through Promoting T-bet+ Regulatory T Cells

IFNγ signaling drives dendritic cells (DCs) to promote type I T cell (Th1) immunity. Here, we show that activation of DCs by IFNγ is equally crucial for the differentiation of a population of T-bet+ regulatory T (Treg) cells specialized to inhibit Th1 immune responses. Conditional deletion of IFNγ receptor in DCs but not in Treg cells resulted in a severe defect in this specific Treg cell subset, leading to exacerbated immune pathology during parasitic infections. Mechanistically, IFNγ-unresponsive DCs failed to produce sufficient amount of IL-27, a cytokine required for optimal T-bet induction in Treg cells. Thus, IFNγ signalling endows DCs with the ability to efficiently control a specific type of T cell immunity through promoting a corresponding Treg cell population

IFNγ Signaling Endows DCs with the Capacity to Control Type I Inflammation during Parasitic Infection through Promoting T-bet+ Regulatory T Cells.

IFNγ signaling drives dendritic cells (DCs) to promote type I T cell (Th1) immunity. Here, we show that activation of DCs by IFNγ is equally crucial for the differentiation of a population of T-bet+ regulatory T (Treg) cells specialized to inhibit Th1 immune responses. Conditional deletion of IFNγ receptor in DCs but not in Treg cells resulted in a severe defect in this specific Treg cell subset, leading to exacerbated immune pathology during parasitic infections. Mechanistically, IFNγ-unresponsive DCs failed to produce sufficient amount of IL-27, a cytokine required for optimal T-bet induction in Treg cells. Thus, IFNγ signalling endows DCs with the ability to efficiently control a specific type of T cell immunity through promoting a corresponding Treg cell population

IFNγ signaling endows DCs with the capacity to control type I inflammation during parasitic infection through promoting T-bet+ regulatory T cells.

IFNγ signaling drives dendritic cells (DCs) to promote type I T cell (Th1) immunity. Here, we show that activation of DCs by IFNγ is equally crucial for the differentiation of a population of T-bet+ regulatory T (Treg) cells specialized to inhibit Th1 immune responses. Conditional deletion of IFNγ receptor in DCs but not in Treg cells resulted in a severe defect in this specific Treg cell subset, leading to exacerbated immune pathology during parasitic infections. Mechanistically, IFNγ-unresponsive DCs failed to produce sufficient amount of IL-27, a cytokine required for optimal T-bet induction in Treg cells. Thus, IFNγ signalling endows DCs with the ability to efficiently control a specific type of T cell immunity through promoting a corresponding Treg cell population

Interleukin-27 Priming of T Cells Controls IL-17 Production In trans via Induction of the Ligand PD-L1

Interleukin (IL)-27 is a key immunosuppressive cytokine that counters T helper 17 (Th17) cell-mediated pathology. To identify mechanisms by which IL-27 might exert its immunosuppressive effect, we analyzed genes in T cells rapidly induced by IL-27. We found that IL-27 priming of naïve T cells upregulated expression of programmed death ligand 1 (PD-L1) in a signal transducer and activator of transcription (STAT)1-dependent manner. When co-cultured with naïve CD4(+) T cells, IL-27-primed T cells inhibited the differentiation of Th17 cells in trans through a PD-1-PD-L1 interaction. In vivo, co-administration of naïve TCR transgenic T cells (2D2 T cells) with IL-27-primed T cells expressing PD-L1 inhibited the development of Th17 cells and protected from severe autoimmune encephalomyelitis. Thus, these data identify a suppressive activity of IL-27, by which CD4(+) T cells can restrict differentiation of Th17 cells in trans

Minimal role of Treg cell-intrinsic IFNγ signaling in promoting T-bet⁺CXCR3⁺ Treg cells.

<p>FACS analysis and frequencies of T-bet⁺ or CXCR3⁺ cells in Foxp3⁺CD4⁺ Treg cells in <b>(A)</b> the spleen (Spl) and <b>(B)</b> the lamina propria (LP) of small intestine. FACS analysis and frequencies of T-bet⁺ or IFNγ⁺ Foxp3^-CD4⁺ Teff cells in <b>(C)</b> Spl and <b>(D)</b> LP isolated from <i>Foxp3</i>^<i>cre</i><i>IFNγR2</i>^<i>fl/fl</i> or <i>Foxp3</i>^<i>cre</i><i>IFNγR2</i>^<i>+/+</i> mice. FACS plots shown are representative of three independent experiments.</p

IL-27 secreted from <i>T gondii</i> infected-DC promotes T-bet⁺ Th1-Treg cell differentiation through stimulating IL-27R on Treg cells.

<p><b>(A, B)</b> CD4⁺CXCR3^-GFP⁺ Treg cells isolated from naive Foxp3^GFP mice were cultured with DCs from <i>CD11c</i>^<i>cre</i><i>IFNγR2</i>^<i>fl/fl</i> or WT control mice day 6 post <i>T</i>. <i>gondii</i> infection. Isotype control or IL-27 neutralizing antibodies were added at the beginning of culture. <b>(C, D)</b> CD4⁺CXCR3^-CD25^hi Treg cells isolated from naive IL-27Rα-deficient or WT control mice were cultured with DCs from WT mice day 6 post <i>T</i>. <i>gondii</i> infection in the presence of antibodies as indicated. FACS plots and histograms represent two or three independent experiments (*p<0.05; **p<0.01; ***p<0.001). </p

DC-derived IL-27 is critical for maintaining normal T-bet⁺ Th1-Treg cell population in both physiological and <i>T</i>. <i>gondii</i> infection settings.

<p>FACS analysis and frequencies of T-bet⁺ cells within Foxp3⁺CD4⁺ T cell population from spleen and LP in <i>CD11c</i>^<i>cre</i><i>IL27p28</i>^<i>fl/fl</i> mice and WT littermate controls <b>(A)</b> at steady state or <b>(B)</b> 8 days after <i>T</i>. <i>gondii</i> infection. FACS plots are representative of three independent experiments (*p<0.05; **p<0.01; ***p<0.001).</p

Reduced T-bet⁺ Th1-Treg cells in <i>CD11c</i>^<i>cre</i><i>IFNγR2</i>^<i>fl/fl</i> mice resulted in unrestrained IFNγ-mediated Th1 inflammation during <i>T</i>. <i>gondii</i> infection.

<p><b>(A,B)</b> Histological assessment of ileum from infected <i>CD11c</i>^<i>cre</i><i>IFNγR2</i>^<i>fl/fl</i> and WT control mice (n = 12). <b>(C)</b> ELISA analysis of serum IFNγ levels and <b>(D)</b> PCR analysis of parasite burden in LP at days 8 after infection. <b>(E)</b> Frequencies of total Foxp3⁺ Treg cells from LP in <i>CD11c</i>^<i>cre</i><i>IFNγR2</i>^<i>fl/fl</i> and WT control mice at day 8 post <i>T</i>. <i>gondii</i> infection. FACS analysis and frequencies of T-bet⁺ cells in Foxp3⁺CD4⁺ Treg cells and IFNγ⁺ cells in Foxp3^-CD4⁺ Teff cells from LP in <i>CD11c</i>^<i>cre</i><i>IFNγR2</i>^<i>fl/fl</i><b>(F)</b> without or <b>(G)</b> with Treg cell collapse and their corresponding WT control mice at day 8 post <i>T</i>. <i>gondii</i> infection. FACS data are representative of three to four independent experiments. (*p<0.05; **p<0.01; ***p<0.001).</p

Dispensable role of IFNγR in DC maturation and function at steady state.

<p>FACS analysis of MHC class II and CD86 in <b>(A)</b> CD11c⁺ DCs with or without LPS stimulation for 24hr from <i>CD11c</i>^<i>cre</i><i>IFNγR2</i>^<i>fl/f</i> and WT control mice. <b>(B)</b> Proliferation of OTII T cells co-cultured with DCs isolated from indicated mice pulsed with different does of OVA protein was shown by CFSE dilution. <b>(C)</b> Gene expression volcano plot, with—log 10 of the p value on the y axis and log 2 fold change on the x axis, such that genes with higher expression in WT DCs are on the right and genes with higher expression in KO DCs are on the left. <b>(D)</b> Signals (log2 intensity) of individual probe sets of <i>Ifngr2</i> gene and their locations on corresponding exons. All data are representative of three independent experiments. (*p<0.05).</p