Effect of tumor necrosis factor-alpha on the stimulus-coupled responses of neutrophils and their modulation by various inhibitors.

Preincubation of human peripheral blood polymorphonuclear leukocytes (HPPMN) with recombinant human tumor necrosis factor-alpha (rHuTNF-alpha) enhanced the formylmethionyl-leucylphenylalanine (FMLP)-induced superoxide (O2-.) generation in a concentration- and preincubation time-dependent manner. The enhancement was very high for the FMLP- or opsonized zymosan (OZ)-induced O2-. generation, but was low for arachidonic acid (AA)- and phorbol myristate acetate (PMA)-induced O.2- generation. The rHuTNF-alpha has no effect on the steady state of intracellular calcium ion concentration ([Ca2+]i) nor on the membrane potential of neutrophils. The rHuTNF-alpha-primed FMLP-induced O2-. generation was inhibited by nicotineamide (NA), pertussis toxin (PT), and by the tyrosine kinase (TK) inhibitor, genistein, but was enhanced by the protein kinase C (PKC) inhibitor, H-7 (1-(5-isoquinolinesulfonyl)-3-methyl-piperazine). The inhibitory actions of NA and PT were also observed in in vivo primed guinea pig peritoneal neutrophils (GPtPMN). However, FMLP-induced O2-. generation of GPtPMN was enhanced by genistein, but was inhibited by H-7. These data indicate that TNF-alpha does not induce changes in [Ca2+]i nor in membrane potential of HPPMN, and that TNF-alpha-primed FMLP-induced O.2- generation of HPPMN is coupled with ADP-ribosylation and activation of G-proteins, and that protein kinases, especially TK, seem to exert an important role in the priming action of TNF.