THE REQUIREMENT FOR PHOSPHOLIPASE-A(2) FOR ACTIVATION OF THE ASSEMBLED NADPH OXIDASE IN HUMAN NEUTROPHILS

Phospholipase A(2) (PLA(2)) inhibitors suppressed simultaneously, in a dose-dependent manner, the activation of NADPH oxidase and the release of H-3-labelled arachidonic acid ([H-3]AA) stimulated by either phorbol 12-myristate 13-acetate (PMA) or opsonized zymosan (OZ) in human neutrophils. In spite of total inhibition of superoxide production in the presence of the PLA(2) inhibitors, 10 mu M bromophenacyl bromide (BPB) or 20 mu M quinacrine, a maximal phosphorylation of p47 and translocation of p47 and p67 to the neutrophil membranes induced by PMA or OZ was observed. Addition of 10 mu M free AA, which by itself did not stimulate superoxide generation, restored oxidase activity in neutrophils treated with PLA(2) inhibitors. These findings indicate that phosphorylation and translocation of the cytosolic factors to the membranes are not sufficient for generating superoxide; a functional PLA(2) is also needed to stimulate the oxidase activity. The inhibition of PLA(2) activity did not prevent the phosphorylation of p47, suggesting that the location of PLA(2) is downstream of and does not activate protein kinase C.