Blockade of LTC4 synthesis caused by additive inhibition of gIV-PLA2 phosphorylation: Effect of salmeterol and PDE4 inhibition in human eosinophils.
BACKGROUND: Prior investigations have demonstrated that beta(2)-adrenoceptor stimulation is ineffective in inhibiting synthesis of eicosanoids in human eosinophils. This effect has been postulated to relate to density or structural differences in the beta(2)-adrenoceptor or its coupled G-protein. However, recent reports indicate that cAMP-specific PDE4 activity in eosinophils is 10-fold that of other inflammatory cells. We postulated that selective blockade of PDE4 in eosinophils would unmask the inhibitory effect of beta(2)-adrenoceptor stimulation and that this inhibition would result from decreased phosphor-ylation of cytosolic group IV-PLA(2) (gIV-PLA(2)). OBJECTIVE: To determine (a) whether PDE4 inhibition alone with rolipram blocked secretions of arachidonic acid (AA) and leukotriene C(4) (LTC(4)) caused by activation of eosinophils with formyl-met-leu-phe plus cytochalasin B (FMLP/B), (b) to determine if PDE4 inhibition plus beta(2)-adrenoceptor agonist act additively to augment endogenous cAMP concentration, and (c) to determine the mechanism by which additive inhibition of AA and LTC(4) synthesis is regulated by cAMP. METHODS: Human eosinophils were pretreated with buffer, salmeterol or rolipram (singly or combination) before FMLP/B activation. Release of AA and LTC(4), intracellular cAMP concentration, and phosphorylation and activation of gIV-PLA(2) were determined. RESULTS: Rolipram unmasked the inhibitory effect of beta(2)-adrenoceptor stimulation with salmeterol and significantly attenuated the stimulated release of AA and subsequent LTC(4). Inhibition corresponded to increased cAMP production caused by rolipram alone or rolipram plus salmeterol and blocked proportionately the phosphorylation and activation of gIV-PLA(2) in FMLP/B-activated eosinophils. CONCLUSIONS: Inhibition of PDE4 by rolipram unmasks beta(2)-adrenergic blockade of LTC(4) synthesis caused by FMLP/B.
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Rolipram salmeterol AA LTC(4) cAMP rolipram rolipram salmeterol PDE4 rolipram cAMP LTC(4) cAMP PDE4 PDE4 PDE4 rolipram arachidonic acid AA leukotriene C(4) LTC(4) AA formyl-met-leu-phe cytochalasin B PDE4 PDE4 LTC4 salmeterol gIV-PLA(2) beta(2)-adrenoceptor gIV-PLA(2) PDE4 LTC(4) beta(2)-adrenergic beta(2)-adrenoceptor G-protein cAMP-specific PDE4 PDE4 beta(2)-adrenoceptor beta(2)-adrenoceptor cytosolic group IV-PLA(2) gIV-PLA(2) PDE4 cAMP PDE4 beta(2)-adrenoceptor PDE4 gIV-PLA2 salmeterol rolipram AA LTC(4) cAMP