Effects of respiratory burst inhibitors on nitric oxide production by human neutrophils.
Human neutrophils (PMN) activated by N-formylmethionyl-leucyl-phenylalanine (fMLP) simultaneously release nitric oxide (.NO), superoxide anion (O2.-) 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 antagonists of intracellular signaling pathways and subsequently stimulated either with fMLP or with a phorbol ester (PMA). The G-protein inhibitor, pertussis toxin (1-10 micrograms/ml) decreased H2O2 yield without significantly changing .NO production in fMLP-stimulated neutrophils; no effects were observed in PMA-activated cells. The inhibition of tyrosine kinases by genistein (1-25 micrograms/ml) completely abolished H2O2 release by fMLP-activated neutrophils; conversely, .NO production increased about 1.5- and 3-fold with fMLP and PMA, respectively. Accordingly, orthovanadate, an inhibitor of phosphotyrosine phosphatase, markedly decreased .NO production and increased O2.- release. On the other hand, inhibition of protein kinase C with staurosporine and the use of burst antagonists like adenosine, cholera toxin or dibutyryl-cAMP diminished both H2O2 and .NO production. The results suggest that the activation of the tyrosine kinase pathway in stimulated human neutrophils controls positively O2.- and H2O2 generation and simultaneously maintains .NO production in low levels. In contrast, activation of protein kinase C is a positive modulator for O2.- and .NO production.
Carreras
M C
MC
University Hospital, School of Medicine, University of Buenos Aires, Argentina.
Riobó
N A
NA
Pargament
G A
GA
Boveris
A
A
Poderoso
J J
JJ
D000262:Adenylyl Cyclases / Q000378:metabolism; D004563:Electrochemistry; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000037:antagonists & inhibitors; D019833:Genistein; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D007529:Isoflavones / Q000494:pharmacology; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D019255:NADPH Oxidases / Q000378:metabolism; D009504:Neutrophils / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis*; D037342:Pertussis Toxin; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D016897:Respiratory Burst / Q000187:drug effects*; D015398:Signal Transduction; D019311:Staurosporine / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D004791:Enzyme Inhibitors; D007529:Isoflavones; D010566:Virulence Factors, Bordetella; D013481:Superoxides; D009569:Nitric Oxide; D009240:N-Formylmethionine Leucyl-Phenylalanine; D006861:Hydrogen Peroxide; D019833:Genistein; D019255:NADPH Oxidases; D037342:Pertussis Toxin; D011505:Protein-Tyrosine Kinases; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate