Adenosine is produced during inflammation and modulates different functional activities in macrophages. In murine bone marrow-derived macrophages, adenosine inhibits M-CSF-dependent proliferation with an IC 50 of 45 μM. Only specific agonists that can activate A 2B adenosine receptors such as 5′-N-ethylcarboxamidoadenosine, but not those active on A 1 (N 6-(R)-phenylisopropyladenosine), A 2A ([p-(2-carbonylethyl) phenylethylamino]-5′-N-ethylcarboxamidoadenosine), or A 3 (N 6-(3-iodobenzyl) adenosine-5′-N-methyluronamide) receptors, induce the generation of cAMP and modulate macrophage proliferation. This suggests that adenosine regulates macrophage proliferation by interacting with the A 2B receptor and subsequently inducing the production of cAMP. In fact, both 8-Br-cAMP (IC 50 85 μM) and forskolin (IC 50 7 μM) inhibit macrophage proliferation. Moreover, the inhibition of adenylyl cyclase and protein kinase A blocks the inhibitory effect of adenosine and its analogues on macrophage proliferation. Adenosine causes an arrest of macrophages at the G 1 phase of the cell cycle without altering the activation of the extracellular-regulated protein kinase pathway. The treatment of macrophages with adenosine induces the expression of p27 kip-1, a G 1 cyclin-dependent kinase inhibitor, in a protein kinase A-dependent way. Moreover, the involvement of p27 kip-1 in the adenosine inhibition of macrophage proliferation was confirmed using macrophages from mice with a disrupted p27 kip-1 gene. These results demonstrate that adenosine inhibits macrophage proliferation through a mechanism that involves binding to A 2B adenosine receptor, the generation of cAMP, and the induction of p27 kip-1 expression.