Anti-CD200R2, anti-IL-9, anti-IL-35, or anti-TGF-β abolishes increased graft survival and Treg induction induced in cromolyn-treated CD200R1KO.CD200tg mice.

Rejection is associated with early degranulation (≥80%) of graft-infiltrating CD200R1 receptor-positive mast cells (MCs). Survival is increased, and MC degranulation is decreased, in CD200 mice but not in CD200R1KO mice. CD200 engagement of CD200R2 (not present on MCs) alters dendritic cell differentiation and enhances induction of Foxp3 regulatory T cells (Tregs). We investigated whether attenuation of MC degranulation by sodium cromoglycate allowed CD200 to increase survival in CD200R1KO mice. C57BL/6 control, CD200R1KO, CD200, or CD200R1KO.CD200 mice received BALB/c grafts with or without treatment with cromoglycate. Survival was monitored daily from day 10, with mixed lymphocyte culture responses measured on day 14 or 21 and graft immunohistology performed on day 14. Decreased MC degranulation and increased graft Foxp3 Treg infiltration/survival occurred in CD200 mice and in CD200-treated control mice or CD200R1KO.CD200 mice receiving cromoglycate. Neutralizing anti-CD200 or anti-CD200R1/R2 monoclonal antibody caused graft rejection, as did anti-interleukin (IL)-9, anti-IL-35, or anti-transforming growth factor-β antibodies, with the latter also decreasing graft-infiltrating Tregs. These data imply a coordinated effect of MCs and Tregs on increased graft survival induced by CD200, with a critical role for IL-9, IL-35, and transforming growth factor-β in the development/function of Tregs.