Adenosine 3 ',5 '-cyclic monophosphate (cAMP)-dependent inhibition of IL-5from human T lymphocytes is not mediated by the cAMP-dependent protein kinase A(1)

IL-5 is implicated in the pathogenesis of asthma and is predominantly relea sed from T lymphocytes of the Th2 phenotype. In anti-CD3 plus anti-CD28-sti mulated PBMC, albuterol, isoproterenol, rolipram, PGE(2), forskolin, choler a toxin, and the cAMP analog, 8-bromoadenosine cAMP (8-Br-cAMP) all inhibit ed the release of IL-5 and lymphocyte proliferation. Although all of the ab ove compounds share the ability to increase intracellular cAMP levels and a ctivate protein kinase (PK) A, the PRA inhibitor H-89 failed to ablate the inhibition of IL-5 production mediated by 8-Br-cAMP, rolipram, forskolin, o r PGE(2). Similarly, H-89 had no effect on the cAMP-mediated inhibition of lymphocyte proliferation. Significantly, these observations occurred at a c oncentration of H-89 (3 muM) that inhibited both PKA activity and CREB phos phorylation in intact cells. Additional studies showed that the PKA inhibit ors H-8, 8-(4-chlorophenylthio) adenosine-3 ' ,5 ' -cyclic monophosphorothi oate Rp isomer, and a myristolated PKA inhibitor peptide also failed to blo ck the 8-Br-cAMP-mediated inhibition of IL-5 release from PBMC. Likewise, a role for PKG was considered unlikely because both activators and inhibitor s of this enzyme had no effect on IL-5 release. Western blotting identified Rap I, a downstream target of the cAMP-binding proteins, exchange protein directly activated by cAMP/cAMP-guanine nucleotide exchange factors 1 and 2 , in PBMC. However, Rap1 activation assays revealed that this pathway is al so unlikely to be involved in the cAMP-mediated inhibition of IL-5. Taken t ogether, these results indicate that cAMP-elevating agents inhibit IL-5 rel ease from PBMC by a novel cAMP-dependent mechanism that does not involve th e activation of PKA.