513 IL-17 Role in the Regulatory Function of B Cells

BACKGROUND: B lymphocytes are known to be important cytokine sources in inflammation and play a pathogenic role by producing autoantibodies in a number of chronic immunological diseases. However, B cell depletion therapy induced an exacerbation of symptoms in some patients with autoimmune disorders, revealing that B cells play a critical anti-inflammatory role mediated by IL-10 release. We therefore investigated the human B cell regulatory subset producing IL-10 in response to stimulation. METHODS: Highly purified B cells were obtained from tonsils by using a multiple-step separation procedure which included rosette depletion, adherence depletion, CD3+ cell magnetic-activated depletion and CD19+ magnetic-activated positive cell selection. CD20+ purity was verified by flow cytometry. The CD19+CD20+ B cells were stimulated with CpG oligonucleotide, IL-4, IFN-gamma, anti-CD40, IL-17A and IL-17F, either alone or in combination. The expression of both IL-6 and IL-10 mRNA was analyzed by quantitative RT-PCR and by ELISA. B regulatory cell subsets expressing IL-10 and the markers CD5 and CD1d were quantified by FACS analysis. B cell proliferation was determined by 3H thymidine incorporation or CFSE labeling. RESULTS: Expression of IL-10 mRNA and protein in purified B cells from tonsils was weakly stimulated by anti-CD40 antibody, CpG oligonucleotide or with IL-17. When B cells were simultaneously stimulated with IL-17, anti-CD40 antibody and CpG oligonucleotide, the mRNA and protein expression of IL-10 was strongly increased (n = 3; P ≤ 0.001). B cells proliferation was also significantly increased. In contrast, stimulation with IL-4 alone or in combination with anti-CD40 antibody, decreased the expression of IL-10 (n = 3; P ≤ 0.001). CONCLUSIONS: TLR9 receptor stimulation synergizes with CD40 and IL-17 receptors stimulation in the induced proliferation and potent release of IL-10 cytokine while decreasing IL-6 production in B cells. These novel findings provide evidence that B lymphocytes might be an important source of the anti-inflammatory IL-10 cytokine, and provide novel evidence that stimulation of B lymphocytes with IL-17 cytokine could be an important regulatory mechanism in immune responses.