Activation of guinea-pig and bovine neutrophil NADPH oxidase by N,N'-dicyclohexylcarbodiimide

Dicyclohexylcarbodiimide (DCCD) is a potent stimulant of superoxide generation in guinea-pig peritoneal and bovine blood neutrophils. The dependence of DCCD-elicited respiratory burst on the composition of the medium was investigated. At 37 degrees C, the superoxide generation was short-lived and a rapid losses of enzymatic activity was observed; at 0 degree C, the activity could be preserved for hours. Superoxide generation by whole cells was accompanied by exocytic degranulation. Prolonged incubation with DCCD at 37 degrees C resulted also in a progressive loss of cellular integrity evidenced by the release of a fraction of lactate dehydrogenase. Km values of the particulate NADPH oxidase isolated from DCCD-triggered guinea-pig and bovine cells were 31.7 and 50.0 microM, respectively. Cells pre-equilibrated with the potential sensitive dye Di-S-C3-(5) exhibited changes in the transmembrane potential upon stimulation. Stimulation with DCCD resulted also in the release of membrane-associated calcium, indicated by quenching of the fluorescence of chlortetracycline-loaded neutrophils. Both effects were observed also in human neutrophils which did not generate superoxide upon exposure to DCCD. The mechanism of DCCD-induced responses is discussed.