Abstract Background: Adenosine accumulates in inflamed tissue and can suppress proliferation and cytokine production in helper T cells (Th). Th cells from mice lacking the A2A Adenosine Receptor (A2AAR-/-) do not function properly in the CD45RB transfer model of colitis. Methods: To examine the role of A2AAR in regulating colitis, we tested colonic myeloperoxidase (MPO) activity after adoptive transfer of Th cell susbsets from wildtype or A2AAR-/- mice. Th cells from mesenteric lymph nodes (MLN) of colitic mice were treated with an A2AAR agonist to assess effects on cytokine production. To investigate the role of A2AAR on myeloid cells in the CD45RB model of colitis, we adoptively transferred wildtype Th cells into RAG1/A2AAR double knockout (DKO) mice. Results: Colons of mice that received A2AAR-/- CD45RBhi cells with wildtype or A2AAR-/- CD45RBlo Treg had significantly greater MPO activity than mice that received wildtype CD45RBhi and CD45RBlo cells. Cytokine production by MLN Th cells from colitic mice was suppressed by an A2AAR agonist. Adoptive transfer of CD45RBhi Th cells into either RAG1-/- or DKO mice induced colitis but co-transfer of CD45RBlo Tregs attenuated colitis only in RAG1-/- mice. Conclusions: These data indicate that adenosine signaling through A2AAR plays an important suppressive role in colitis and that this is mediated through effects on both T cells and myeloid cells. CK supported by a CCFA Fellowship.