Apparent involvement of the A(2A) subtype adenosine receptor in the anti-inflammatory interactions of CGS 21680, cyclopentyladenosine, and IB-MECA with human neutrophils

This study was undertaken to identify the adenosine receptor (AR) subtypes which downregulate the proinflammatory activities of human neutrophils, as well as the involvement of adenosine 3',5'-cyclic monophosphate (cAMP) and its relationship to cellular handling of Ca2+ in mediating these effects. N eutrophils were treated with varying concentrations (0.01-1 mu M) of AR ago nists operative at A(1) (N-6-cyclopencyladenosine, CPA), A(2A) (2(4-[(2-car boxyethyl)phenyl]ethylamino)-5'-N-ethylcarboxamidoadenosine, CGS 21680), an d A(3) (N-6-(3-iodobenzyl-5)-N-methylcarbamoyladenosine, IB-MECA) receptors , after which they were activated with the chemoattractant, N-formyl-L-meth ionyl-L-leucyl-L-phenylalanine (FMLP, 1 mu M). Intracellular cAMP, superoxi de, and elastase were assayed using radioimmunoassay, lucigenin-enhanced ch emiluminescence (LECL), and colorimetric procedures, respectively, while ch anges in the concentrations of cytosolic Ca2+ were monitored by fura-2-base d spectrofluorimetry. CGS 21680, at all concentrations tested, inhibited su peroxide production in a dose-related manner, while CPA and IB-MECA were ef fective only at the highest concentrations tested (0.5-1 mu M). The release of elastase from activated neutrophils was also inhibited by all three AR agonists, but was more sensitive to CGS 21680 and IB-MECA than was superoxi de production. The inhibitory effects of all 3 agonists on superoxide produ ction and elastase release were associated with accelerated clearance of Ca 2+ from the cytosol of activated neutrophils, and were effectively neutrali zed by pretreatment of the cells with the highly selective A(2A)R antagonis t, ZM 241385 (4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin -5yl amino]ethyl)phenol). Increased cAMP was detected in neutrophils treate d with CGS 21680 and IB-MECA (1 mu M) These data support the involvement of the A(2A)R subtype in the suppression of superoxide production and degranu lation by activated human neutrophils, probably by cAMP-mediated alteration s in Ca2+ handling. BIOCHEM PHARMACOL 60;7:993-999, 2000. (C) 2000 Elsevier Science Inc.