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Memory B cells IgM only Marginal zone B cells without plasmablasts

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Presentation on theme: "Memory B cells IgM only Marginal zone B cells without plasmablasts"— Presentation transcript:

1 Memory B cells IgM only Marginal zone B cells without plasmablasts CD19+ IgM Plasmablasts Memory B cells IgD only Switched memory CD27 CD27 IgD Naive B cells Naive B cells CD27- IgD- CD38 IgD Transitional CD24 Mature naive CD38 Figure e-1 Gating strategy of B cell subset characterization: a representative example from an HC. B cell subsets were gated in CD19 positive peripheral mononuclear cells. Plasmablasts were defined as CD19+ CD38hi CD27hi. Among the remaining B cells, CD27 and IgD expression distinguished memory B cells (CD27+), naive B cells (IgD+ CD27-) and exhausted B cells (IgD- CD27-). Among CD27+ B cells, the expression of IgM and IgD distinguished marginal zone B cells (CD27+ IgMhi IgDlo), switched memory B cells (CD27+ IgM- IgD-), memory IgM only (CD27+ IgM+ IgD-) and IgD only B cells (CD27+ IgM- IgD+). Among CD27- B cells, transitional B cells were characterized by high expression of both CD38 and CD24, whereas mature naive B cells expressed intermediate level of CD24 and CD38.

2 IL-10+ : 6.1% IL-10 Isotype control IL-6 CD19 IL-6+ : 61.1% Figure e-2 Representative illustration of IL-10 and IL-6 intracellular expression in CD19+ B cells after 5h (A) and 48 hours (B) stimulation.

3 A B Figure e-3 Lymphocyte count and distribution in peripheral blood of healthy controls and MS patients at baseline. Total lymphocyte counts in the different groups (A). Comparison of percentages of total T cells, CD4 and CD8 T cells, NK and total B cells in the different groups (B). HC: healthy controls, RIS: radiologically isolated syndrome, CIS: clinically isolated syndrome, RRMS: relapsing remitting multiple sclerosis patients.

4 A wo evolution w evolution ≤ 6 months > 6 months wo evolution w evolution ≤ 6 months > 6 months wo evolution w evolution ≤ 6 months > 6 months wo evolution w evolution ≤ 6 months > 6 months wo evolution w evolution ≤ 6 months > 6 months wo evolution w evolution ≤ 6 months > 6 months B Transitional Mature naive Marginal zone Switched memory wo evolution w evolution ≤ 6 months > 6 months wo evolution w evolution ≤ 6 months > 6 months wo evolution w evolution ≤ 6 months > 6 months wo evolution w evolution ≤ 6 months > 6 months IgD only IgM only CD27- IgD- Plasmablasts RIS/CIS wo evolution w evolution ≤ 6 months > 6 months wo evolution w evolution ≤ 6 months > 6 months wo evolution w evolution ≤ 6 months > 6 months wo evolution w evolution ≤ 6 months > 6 months Figure e-4 Figure e-4: Lymphocyte population and B cell subset distribution in evolving and non evolving RIS/CIS patients In RIS/CIS patients, comparison of total lymphocyte counts, percentages of total T cells, CD4 and CD8 T cells, NK and total B cells (A) and B cell subset percentages (B) between evolving and non-evolving patients. RIS: radiologically isolated syndrome, CIS: clinically isolated syndrome, w: with, wo: without.

5 A B Figure e-5 RR MS wo evolution w evolution ≤ 6 months wo evolution

6 Figure e-5 Lymphocyte population and B cell subset distribution in evolving and non-evolving in RRMS patients In RRMS patients, comparison of total lymphocyte counts, percentages of total T cells, CD4 and CD8 T cells, NK and total B cells (A) and B cell subset percentages (B) between evolving and non-evolving patients. RRMS: relapsing remitting multiple sclerosis patients, w: with, wo: without.

7 A B C D E F < 6 mois : NS IL-6/IL-10 B cell ratio IL-6+ B cells
RIS/CIS wo evolution w evolution > 6 months IL-6+ B cells RIS/CIS wo evolution w evolution > 6 months IL-10+ B cells RIS/CIS wo evolution w evolution > 6 months D E F IL-6/IL-10 B cell ratio RRMS w evolution ≤ 6 months wo evolution IL-6+ B cells RRMS wo evolution w evolution ≤ 6 months IL-10+ B cells RRMS wo evolution w evolution ≤ 6 months < 6 mois : NS Figure e-6 Functional properties of B cells according to the disease evolutivity Percentages of IL-10 producing B cells (A), IL-6 producing B cells (B), and comparison of IL-6/IL-10 producing B cell ratio (C), between evolving and non-evolving RRMS patients at 6 months. RRMS: relapsing remitting multiple sclerosis patients, w: with, wo: without.

8 A B C IL-6/IL-10 B cell ratio wo evolution w evolution ≤ 6 months IL-6+ B cells wo evolution w evolution ≤ 6 months IL-10+ B cells wo evolution w evolution ≤ 6 months Figure e-7 Functional properties of B cells according to the disease evolution in RIS/CIS patients after short polyclonal stimulation A, B, C: Comparison of IL-6/IL-10 producing B cell ratio (A), percentages of IL-6 producing B cells (B) and IL-10 producing B cells after 5 hours stimulation (C) between RIS/CIS patients who evolved and who did not at 6 months, w: with, wo: without.

9 Short stimulation Long stimulation
B Long stimulation IL-10+ B cells IL-10+ B cells % % Does IL-6 production by IL-10 producing B cells influence, perturb, unsettle their regulatory properties ? Switched memory Switched memory Transitional Mature naive Transitional Marginal zone Mature naive Marginal zone Figure e-8 Cytokine production capacities of peripheral blood B cell subsets after short (5h) or long (48h) polyclonal stimulation Percentages of IL-10 producing B cells among transitional, mature naïve, marginal zone and switched memory B cell subsets from healthy controls after 5h (A) and 48h (B) stimulation. RIS: radiologically isolated syndrome, CIS: clinically isolated syndrome.

10 p > 0.05 Does IL-6 production by IL-10 producing B cells influence, perturb, unsettle their regulatory properties ? Figure e-9 IL-10 B cell production induced by long stimulation of healthy control PBMC is independent of type I IFN. Percentage of IL-10 producing B cells after stimulation for 48H with type I IFN blocking antibodies (Ab): anti-IFN R2, anti-IFN-α, anti-IFN-β 10µg/ml each, or with corresponding isotype controls. IFN: interferon, Ab: antibodies.

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