Ionomycin causes activation of p38 and p42/44 mitogen-activated protein kinases in human neutrophils

Abstract

Many receptor-linked agents that prime or activate the NADPH oxidase in polymorphonuclear neutrophils (PMNs) elicit changes in cytosolic Ca2+concentration and activate mitogen-activated protein (MAP) kinases. To investigate the role of Ca2+in the activation of p38 and p42/44 MAP kinases, we examined the effects of the Ca2+-selective ionophore ionomycin on priming and activation of the PMN oxidase. Ionomycin caused a rapid rise in cytosolic Ca2+that was due to both a release of cytosolic Ca2+stores and Ca2+influx. Ionomycin also activated (2 μM) and primed (20–200 nM) the PMN oxidase. Dual phosphorylation of p38 MAP kinase and phosphorylation of its substrate activating transcription factor-2 were detected at ionomycin concentrations that prime or activate the PMN oxidase, while dual phosphorylation of p42/44 MAP kinase and phosphorylation of its substrate Elk-1 were elicited at 0.2–2 μM. SB-203580, a p38 MAP kinase antagonist, inhibited ionomycin-induced activation of the oxidase (68 ± 8%, P < 0.05) and tyrosine phosphorylation of 105- and 72-kDa proteins; conversely, PD-98059, an inhibitor of MAP/extracellular signal-related kinase 1, had no effect. Treatment of PMNs with thapsigargin resulted in priming of the oxidase and activation of p38 MAP kinase. Chelation of cytosolic but not extracellular Ca2+completely inhibited ionomycin activation of p38 MAP kinase, whereas chelation of extracellular Ca2+abrogated activation of p42/44 MAP kinase. These results demonstrate the importance of changes in cytosolic Ca2+for MAP kinase activation in PMNs.