Inhibition of superoxide generation and myeloperoxidase release by carvedilol after receptor and nonreceptor stimulation of human neutrophils

To compare three stimuli which activate human neutrophils with different signal transduction mechanisms, in order to better localize the effect of the beta-adrenoceptor antagonist carvedilol (CARV) on superoxide generation (O2*-) and myeloperoxidase release (MPO). The effect of CARV [0.1-100 micromol/l] on O2*- generation and MPO release from isolated human neutrophils was studied after specific receptor activator N-formyl-methionyl-leucyl-phenylalanine (fMLP) and nonreceptor phorbol-12-myristate-13-acetate (PMA) and calcium ionophor (A23187) stimuli. O2*- generation was measured as superoxide dismutase inhibitable reduction of cytochrome c and MPO release as the oxidation of o-dianisidine in the presence of hydrogen peroxide in a spectrophotometer Hewlet Packard 8452 A at respective 550 and 463 nm. CARV had no effect on O2*- generation and MPO release in nonstimulated cells. In the concentration 10 and 100 micromol/l, it significantly decreased fMLP and PMA stimulated O2*- generation and MPO release. Incubation of neutrophils with CARV [100 micromol/l] caused significant inhibition of O2*- generation and MPO release induced by A23187. Wortmannin, a specific inhibitor of 1-phosphatidylinositol-3-kinase, inhibited significantly only fMLP stimulated O2*- generation. CARV [100 micromol/l] with wortmannin [50 nmol/l] further decreased O2*- generation after the same stimulus. CARV decreased O2*- generation and MPO release from isolated human neutrophils both by membrane-operating stimulus - fMLP and membrane bypassing activators - PMA and A 23187. This fact, together with effect the of wortmannin, indicates that the inhibition may be attributed to the non-specific action of CARV and its interference with phospholipase D signaling pathway, which plays only a minor role in proteinkinase C stimulated O2*- generation.