Involvement of cytosolic phospholipase A2 and secretory phospholipase A2 in arachidonic acid release from human neutrophils

The purpose of this study was to define the role of secretory phospholipase A(2) (sPLA(2)), calcium-independent PLA(2), and cytosolic PLA(2) (cPLA(2)) in arachidonic acid (AA) release from fMLP-stimulated human neutrophils, While fMLP induced the release of extracellular sPLA(2) activity and AA, 70% of sPLA(2) activity remained associated with the cell. Treatment with the cell-impermeable sPLA(2) inhibitors DTT or LY311-727, or the anti-sPLA(2) Ab 3F10 all inactivated extracellular sPLA(2) activity, but had minimal effect on neutrophil AA mass release. In contrast, coincubation of streptolysin-O toxin-permeabilized neutrophils with DTT, LY311-727, or 3F10 all decreased [H-3(8)]AA release from [H-3(8)]AA-labeled, fMLP-stimulated cells. Exposure to fMLP resulted in a decrease in the electrophoretic mobility of cPLA(2), a finding consistent with cPLA(2) phosphorylation,and stimulated the translocation of cPLA(2) from cytosolic to microsomal and nuclear compartments. The role of cPLA(2) was further evaluated with the cPLA(2) inhibitor methyl arachidonyl fluorophosphonate, which attenuated cPLA(2) activity in vitro and decreased fMLP-stimulated AA mass release by intact neutrophils, but had no effect on neutrophil sPLA(2) activity. Inhibition of calcium-independent PLA(2) with haloenol lactone suicide substrate had no effect on neutrophil cPLA(2) activity or AA mass release. These results indicate a role for cPLA(2) and an intracellular or cell-associated sPLA(2) in the release of AA from fMLP-stimulated human neutrophils, The Journal of Immunology, 2000, 164: 2084-2091.