- We have recently demonstrated that angiotensin II stimulates NADPH-oxidase and phospholipase A2 activities in phagocytic cells and the essential requirement of cytosolic phospholipase A2 for NADPH-oxidase activity in phagocytic cells. Since the activation of cytosolic phospholipase A2 is attributed to serine-505 phosphorylation mediated by MAP-kinases, the role of p38 MAP-kinases and ERK1/2 in NADPH-oxidase and phospholipase A2 activation was studied in neutrophils and monocytes from normotensive volunteers. Neutrophils and monocytes were separated from whole blood. NADPH-oxidase activity was measured by reduction of cytochrome C. Phospholipase A2 activity was measured by the release of arachidonic acid from prelabled cells. Cells were incubated with MEK inhibitors UO126 (0.1μM–5μM), or with p38 MAP-kinases inhibitor SB 202190 (0.5μM–10μM) for 30 min in 37°C prior to stimulation with angiotensin II (10−6M). The activation of ERK1/2 and p38 MAP-kinase was determined by western blot analysis with anti-active antibodies. Angiotensin II stimulated a time-dependent activation of both type of MAP-kinases ERK2 and p38. ERK activation was immediate, and was detected 30 sec after stimulation, peaked at 3 min and decreased thereafter. Phosphorylation of p38 MAP kinase was detected in unstimulated cells slightly increased at 3 min and stayed elevated for, at least, 15 min. Activation of NADPH-oxidase in angiotensin II-stimulated neutrophils and monocytes was inhibited in a dose dependent manner by both the p38 MAP-kinases inhibitor, SB 202190, and the MEK inhibitor, UO126. The p38 MAP kinase inhibitor, SB 202190, completely inhibited phospholipase A2 activity stimulated by angiotensin II in correlation with its effect on NADPH oxidase activity. In contrast the MEK inhibitor, UO126, did not affect phospholipase A2 activity stimulated by angiotensin II. Our results demonstrated that both types of MAP-kinases, ERK and p38 mediate the activation of NADPH-oxidase by angiotensin II in phagocytic cells. While p38 MAP-kinase mediate the phospholipase A2 activity, ERK does not participate in phospholipase A2 activation and its role in NADPH-oxidase activation remains to be elucidated.