Research ArticleIMMUNE REGULATION

Tfr cells lack IL-2Rα but express decoy IL-1R2 and IL-1Ra and suppress the IL-1–dependent activation of Tfh cells

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Science Immunology  08 Sep 2017:
Vol. 2, Issue 15, eaan0368
DOI: 10.1126/sciimmunol.aan0368
  • Fig. 1 Tfr and Tfh cell response to IL-2.

    (A to D) Proportions of Treg (A), Tfh (B), and Tfr (C) cells in CD4+ T cells and the Tfr/Tfh ratio (D) of untreated mice (filled circles), IL-2–treated mice (filled squares), immunized mice (empty circles), and immunized IL-2–treated mice (empty squares). n = 5; *P < 0.05, **P < 0.01, Mann-Whitney U test. Data are representative of three independent experiments.

  • Fig. 2 Tfr cells do not express CD25.

    (A) Flow cytometry contour plots (left) showing CXCR5hiPD-1hi Tfol (orange) and CXCR5int/loPD-1int/lo non-Tfol (blue) cells within CD4+ T cells and histograms showing their Bcl6 expression status (right). (B) Foxp3 expression of Tfol and non-Tfol cells defining Treg cells and Tfr cells (both Foxp3+). (C) Mean fluorescence intensity (MFI) of CD25 expression levels in Treg cells and Tfr cells (n = 5). Data are representative of three independent experiments (left). Representative histogram of the expression of CD25 in Tfr and Treg cells (right). (D and E) IL-4 (D) and IL-21 (E) production by Tfh cells from immunized mice, in the presence (n = 4) or absence (n = 4 to 5) of Tfr cells. Results are representative of two independent experiments. (F) Flow cytometry plot identifying different populations according to their PD-1 and CXCR5 expression within CD4+Foxp3+ T cells (left) and their expression of Bcl6 (middle) and CD25 (right). For (C) to (E), *P < 0.05, **P < 0.01, Mann-Whitney U test.

  • Fig. 3 Tfr cells’ transcriptomic profile distinguishes them from Treg and Tfh cells.

    (A) Heat map comparing the gene expression profiles of Treg (green), Tfr (blue), and Tfh (black) cells from two genetic backgrounds after immunization. Red, high gene expression; blue, low gene expression. (B) Dendrogram of Treg, Tfr, and Tfh cells representing the hierarchical cluster analysis performed with the pvclust R package.

  • Fig. 4 Gene and protein expression reveals a unique functional profile for Tfr cells compared with Treg and Tfh cells.

    (A) NFATc1 expression and its downstream regulated genes. (B and C) Variations in functions (B) and expression (C) of indicated molecules in Treg (white), Tfh (blue), and Tfr (pink) cells. (D and E) MFI of IL-1R2 (D) (n > 20) and OX40 (E) (n = 8) in Tfr, Treg, Teff, and Tfh cells. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, Mann-Whitney U test.

  • Fig. 5 IL-1β induces Tfh cell activation that is inhibited by Tfr cells.

    (A) Fold change in Tfh cell proportion among CD4+ cells from indicated conditions in comparison with OVA + Alum control condition (dotted line) (n = 6 to 7). Data shown are from one experiment and representative of two independent experiments. (B) Serum anti-OVA–specific IgG production by mice immunized with OVA, treated (n = 3) or not treated (n = 5) with IL-1β. (C) IL-4 production by B cells and Tfh cells from immunized mice, in the presence or absence of B cells or IL-1β. (D and E) IL-4 and IL-21 production by Tfh cells from immunized mice cocultured with B cells, with or without Tfr cells or Anakinra (D) and with or without IL-1β and Tfr cells (E). (C to E) n = 4 to 5; *P < 0.05, **P < 0.01, Mann-Whitney U test.

  • Fig. 6 Tfr cells respond better to immunization with self-antigens than with foreign antigens.

    (A to C) Tfh (A) and Tfr (B) cell proportions within Tfol cells and Tfr/Tfh ratio (C) after INS (n = 6) or OVA (n = 7) immunization. (D and E) GCB cell proportions within CD19+ B cells (D) and correlation between the percentage of Tfr and GCB cells from INS-immunized (filled circles) or OVA-immunized (empty circles) mice (E). (F and G) GCB cell proportions within CD19+ B cells (F) and correlation between the percentage of Tfr and GCB cells from INS-immunized mice receiving IL-1β (empty circles) or not (filled circles) (G). (A to C and F) **P < 0.01, Mann-Whitney U test. (E and G) Each symbol represents one mouse, and the Spearman rank correlation value is shown.

  • Fig. 7 Human Tfr cells do not express CD25.

    (A) Flow cytometry contour plots showing CD4+Foxp3 and CD4+Foxp3+ T cells (left); CXCR5hiPD-1hi Tfh cells (blue) and CXCR5int/loPD-1int/lo Teff cells (cyan) within CD4+Foxp3 T cells (middle); and CXCR5hiPD-1hi Tfr cells (red) and CXCR5int/loPD-1int/lo Treg cells (green) within CD4+Foxp3+ T cells (right). (B) Histograms showing CD25 expression level (left) and MFI (right) on gated subsets from (A). (C) Flow cytometry contour plot identifying different subsets according to PD-1 and CXCR5 expression within CD4+Foxp3+ T cells (left) and the corresponding expression of CD25 (right). (D) Histograms showing the IL-1R2 expression level of the subsets mentioned above (left) and their MFI (right). (B and D) n = 5; *P < 0.05, **P < 0.01, Mann-Whitney U test.

Supplementary Materials

  • immunology.sciencemag.org/cgi/content/full/2/15/eaan0368/DC1

    Materials and Methods

    Fig. S1. Representative flow cytometry gating for Tfol cells.

    Fig. S2. IL-2 increases the number of splenic Treg cells.

    Fig. S3. Expression of CD25 on Tfr and Tfh cells in three different genetic backgrounds.

    Fig. S4. Tfh and Tfr cells have similar expressions of Bcl6 and CD25.

    Fig. S5. Clustering of Treg, Tfr, and Tfh cells based on the expression of the entire 545-gene set from the NanoString mouse immunology panel.

    Fig. S6. Cytokines, CD40L, and OX40 expression of Tfr, Treg, and Tfh cells.

    Fig. S7. IL-1R2 and OX40 expression of Tfr, Treg, Teff, and Tfh cells.

    Fig. S8. IL-1β production in coculture of B and Tfh cells.

    Fig. S9. GCB representative gating after INS or OVA immunization.

    Table S1. Exact P values of the asterisk symbols shown in figures.

    Source data (Excel file)

  • Supplementary Materials

    Supplementary Material for:

    Tfr cells lack IL-2Rα but express decoy IL-1R2 and IL-1Ra and suppress the IL-1–dependent activation of Tfh cells

    Paul-Gydeon G. Ritvo, Guillame Churlaud, Valentin Quiniou, Laura Florez, Faustine Brimaud, Gwladys Fourcade, Encarnita Mariotti-Ferrandiz, David Klatzmann*

    *Corresponding authors. Email: david.klatzmann{at}upmc.fr

    Published 8 September 2017, Sci. Immunol. 2, eaan0368 (2017)
    DOI: 10.1126/sciimmunol.aan0368

    This PDF file includes:

    • Materials and Methods
    • Fig. S1. Representative flow cytometry gating for Tfol cells.
    • Fig. S2. IL-2 increases the number of splenic Treg cells.
    • Fig. S3. Expression of CD25 on Tfr and Tfh cells in three different genetic backgrounds.
    • Fig. S4. Tfh and Tfr cells have similar expressions of Bcl6 and CD25.
    • Fig. S5. Clustering of Treg, Tfr, and Tfh cells based on the expression of the entire 545-gene set from the NanoString mouse immunology panel.
    • Fig. S6. Cytokines, CD40L, and OX40 expression of Tfr, Treg, and Tfh cells.
    • Fig. S7. IL-1R2 and OX40 expression of Tfr, Treg, Teff, and Tfh cells.
    • Fig. S8. IL-1β production in coculture of B and Tfh cells.
    • Fig. S9. GCB representative gating after INS or OVA immunization.
    • Table S1. Exact P values of the asterisk symbols shown in figures.

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    Other Supplementary Material for this manuscript includes the following:

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