The role of adenosine A2A and A2B receptors in the regulation of TNF-α production by human monocytes

Adenosine is an endogenous nucleoside that regulates many physiological processes through the activation of its four receptors: A1, A2A, A2B and A3. Previous studies have identified the involvement of A2 receptors in the inhibitory activity of adenosine analogues on tumor necrosis factor-α (TNF-α) production by lipopolysaccharide (LPS) activated monocytes, but the relative contributions of A2A versus A2B receptors have not been determined in human primary monocytes. Nor has the role of A1 and A3 been clearly identified in the system. The lack of such information impacts on the selection of adenosine receptor agonists for disease intervention. Using LPS-stimulated human primary monocytes, we found that the adenosine receptor agonist, 5′-N-ethylcarboxamidoadenosine (NECA) or the A2A receptor agonist, 4-[2-[[6-amino-9-(N-ethyl-b-d-ribofuranuronamidosyl)-9H-purin-2-yl]amino]ethyl]benzenepropanoic acid hydrochloride (CGS21680) produced a concentration-dependent inhibition of TNF-α production, with IC50s of 58.4 nM (32.7-104.5 nM, 95% confidence interval) and 49.2 nM (22.7-105.9 nM, 95% confidence interval), respectively. The selective A2A receptor blocker, 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-ylaminso]ethyl)phenol (ZM241385, 30 nM), antagonized the effects of NECA and CGS21680 (pKB estimates were 8.7 ± 0.1 and 8.9 ± 0.1, respectively), while the selective A2B antagonist, N-(4-cyano-phenyl)-2-[4-(2,6-dioxo-1,3-dipropyl-2,3,4,5,6,7-hexahydro-1H-purin-8-yl)-phenoxy]-acetamide (MRS1754, 100 nM), failed to antagonize the effects of either agonist. Furthermore, neither the A1 receptor agonist, 2-chloro-N6-cyclopentyladenosine (CCPA) nor the A3 receptor agonist, 1-[2-chloro-6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-b-d-ribofuranuronamide (2-Cl-IB-MECA) showed significant inhibitory activity at concentrations that effectively bind to their respective receptors. We conclude that A2A receptor activation is predominantly responsible for the inhibitory effects of adenosine receptor agonists on TNF-α production from LPS-stimulated monocytes.