Natural and inducible regulatory B cells are widely distributed in ovine lymphoid tissues

Regulatory B cells that produce IL-10 are now recognized as an important component of the immune system. We previously confirmed that IL-10 secreting CD21⁺ regulatory B cells (B_reg cells) were present in ovine jejunal Peyer's patches (JPP) and this IL-10 production suppressed IL-12 and IFN-γ secretion. It is not known, however, whether ovine B_reg cells are restricted to JPP or are present in other lymphoid tissues. Therefore, CD21⁺ B cells were purified from sheep JPP and from a variety of mucosal and systemic lymphoid tissues using magnetic cell sorting. Purified CD21⁺ B cells were stimulated with a TLR9-agonist, CpG oligodeoxynucleotide (CpG ODN), and the frequency of spontaneous and inducible (i) IL-10-secreting B cells was evaluated by ELISPOT. Spontaneous IL-10 secreting CD21⁺ B cells were present in mucosal (jejunal PP, parabronchial lymph nodes (LN), mesesnteric LN, and palatine tonsils) and systemic (spleen and blood) lymphoid tissues. Mucosal lymphoid tissues (parabronchial and mesenteric LNs and JPP) had the highest frequency of cells spontaneously secreting IL-10 while tonsils had the lowest. The frequency of B cells spontaneously secreting IL-10 was lowest in blood and spleen. There was large inter-animal variation in the frequency of CD21⁺ B cells spontaneously secreting IL-10 and no significant difference was detected following CpG ODN stimulation. When comparing within individual animals there was, however, a consistent increase in the frequency of CD21⁺ cells secreting IL-10 following CpG ODN stimulation versus stimulation with GpC control ODN. The presence of inducible (i)B_reg cells in ovine mucosal tissues supports previous evidence from mice indicating that B cells have the capacity to modulate inflammatory responses. The presence of iB_reg cells in ruminants may also provide a novel therapeutic target for both immunomodulatory drugs and vaccines designed to control antigen-specific mucosal inflammation.