Activation of phospholipase A(2) in human neutrophils by polyunsaturated fatty acids and its role in stimulation of superoxide production

Although polyunsaturated fatty acids (PUFA) have been shown to stimulate ne utrophil responses such as the oxygen-dependent respiratory burst (superoxi de production), the mechanisms involved still remain undefined. Here we inv estigate the effect of PUFA on the phospholipase A(2) (PLA(2))-signal trans duction process in human neutrophils. Exogenous eicosatetraenoic acid [arac hidonic acid; C-20:4(n-6)] or docosahexaenoic acid [C-22:6(n-3)] promoted t he release of [H-3](C22:6)(n-6) from prelabelled neutrophils in a time- and dose-dependent manner, which is indicative of PLA(2) activation. The relea se of [H-3](C20:4)(n-6) from the cells by C-20:4(n-6) and C-22:6(n-3) was s uppressed by PLA(2) inhibitors. Other PUFA {eicosapentaenoic [C-20:5(n-3)], octadecatrienoic [gamma-linolenic; C-18:3(n-6)] and octadecadienoic [linol eic; C-18:2(n-6)] acids} also had the ability to release [H-3]C-20:4(n-6), however, certain C-20:4(n-6) derivatives [15-hydroperoxyeicosatetraenoic ac id, 15-hydroxyeicosatetraenoic acid and C-20:4(n-6) methyl ester] and satur ated fatty acids [octadecanoic (stearic; C-18:0) and eicosanoic (arachidic; C-20:0) acids] had no significant effect. Treatment of the neutrophils wit h exogenous C-22:6(n-3) caused the mass of endogenous unesterified C-20:4(n -6) to increase. Incubation of the leucocytes with C-20:4(n-6) or C-22:6(n- 3) evoked activation of the 85 kDa cytosolic PLA(2) (cPLA(2)) and the 14 kD a secretory PLA(2) (sPLA(2)), but not the cytosolic Ca2+-independent PLA(2) . In contrast, C-20:0 did not activate any of the PLA(2) isoforms. Activati on of cPLA(2) by PUFA was found to precede that of sPLA(2). C-22:6(n-3), C- 20:4(n-6) and other PUFA induced punctate localization of cPLA(2) in the ce lls, which was not observed with saturated fatty acids. Pretreatment of the leucocytes with PLA(2) inhibitors markedly decreased superoxide production induced by C-20:4(n-6). These results show that PUFA activate PLA(2) in ne utrophils, which might have a mandatory role in biological responses.