ACTIVATION OF NF-KAPPA-B BY THE RESPIRATORY BURST OF MACROPHAGES

H2O2 and other reduced oxygen species have been proposed as activators of the transcription factor, NF kappa B. Stimulated macrophages produ ce superoxide and H2O2 (the respiratory burst). We tested the hypothes is that production of these species could serve as part of the NF kapp a B activation pathway in rat alveolar macrophages and the J774A.1 mou se monocyte/macrophage cell line. Phorbol myristate acetate (PMA) and ADP, which stimulate the respiratory burst, caused NF kappa B activati on in both cells. Catalase abolished NF kappa B activation, while supe roxide dismutase produced little inhibition. Thus, H2O2 was the princi pal agent of respiratory burst-associated NF kappa B activation. Aboli tion of NF kappa B activation by catalase also suggested that intermed iate signaling pathways, such as protein kinase C activation or intrac ellular free calcium elevation must not be involved. Exogenous H2O2 ad ded as a bolus greater than or equal to 50 mu M (greater than or equal to 50 nmol/10(6) macrophages) also activated NF kappa B in macrophage s. Nevertheless, the maximum endogenous production of H2O2 by stimulat ed alveolar macrophages during a 30-min incubation was less than or eq ual to 1.3 nmol H2O2/10(6) cells for PMA stimulation and less than or equal to 0.2 nmol H2O2/10(6) cells for ADP stimulation. Thus, relative ly little endogenous H2O2 generation was required to produce NF kappa B activation compared to the required amount of exogenous H2O2. As H2O 2 rapidly diffuses and is consumed, these results suggest that the sit e of action for endogenously generated H2O2 is probably close to its o rigin, the plasma membrane.