EFFECTS OF RESPIRATORY BURST INHIBITORS ON NITRIC-OXIDE PRODUCTION BYHUMAN NEUTROPHILS

Human neutrophils (PMN) activated by N-formyl-methionyl-leucyl-phenyla lanine (fMLP) simultaneously release nitric oxide (.NO), superoxide an ion (O-2(.-)) and its dismutation product, hydrogen peroxide (H2O2) TO assess whether .NO production shares common steps with the activation of the NADPH oxidase, PMN were treated with inhibitors and antagonist s of intracellular signaling pathways and subsequently stimulated eith er with FMLP or with a phorbol ester (PMA). The G-protein inhibitor, p ertussis toxin (1-10 mu g/ml) decreased H2O2 yield without significant ly changing .NO production in fMLP-stimulated neutrophils; no effects were observed in PMA-activated cells. The inhibition of tyrosine kinas es by genistein (1-25 mu g/ml) completely abolished H2O2 release by fM LP-activated neutrophils; conversely, .NO production increased about 1 .5- and 3-fold with fMLP and PMA, respectively. Accordingly, orthovana date, an inhibitor of phosphotyrosine phosphatase, markedly decreased .NO production and increased O-2(.-) release. On the other hand, inhib ition of protein kinase C with staurosporine and the use of burst anta gonists like adenosine, cholera toxin or dibutyryl-cAMP diminished bot h H2O2 and .NO production. The results suggest that the activation of the tyrosine kinase pathway in stimulated human neutrophils controls p ositively O-2(.-) and H2O2 generation and simultaneously maintains NO production in low levels. In contrast, activation of protein kinase C is a positive modulator for O-2(.-) and .NO production.