Activation of phospholipase A2 in human neutrophils by polyunsaturated fatty acids and its role in stimulation of superoxide production

Abstract

Although polyunsaturated fatty acids (PUFA) have been shown to stimulate neutrophil responses such as the oxygen-dependent respiratory burst (superoxide production), the mechanisms involved still remain undefined. Here we investigate the effect of PUFA on the phospholipase A2 (PLA2)-signal transduction process in human neutrophils. Exogenous eicosatetraenoic acid [arachidonic acid; C20:4(n-6)] or docosahexaenoic acid [C22:6(n-3)] promoted the release of [3H]C20:4(n-6) from prelabelled neutrophils in a time- and dose-dependent manner, which is indicative of PLA2 activation. The release of [3H]C20:4(n-6) from the cells by C20:4(n-6) and C22:6(n-3) was suppressed by PLA2 inhibitors. Other PUFA ¿eicosapentaenoic [C20:5(n-3)], octadecatrienoic [gamma-linolenic; C18:3(n-6)] and octadecadienoic [linoleic; C18:2(n-6)] acids¿ also had the ability to release [3H]C20:4(n-6); however, certain C20:4(n-6) derivatives [15-hydroperoxyeicosatetraenoic acid, 15-hydroxyeicosatetraenoic acid and C20:4(n-6) methyl ester] and saturated fatty acids [octadecanoic (stearic; C18:0) and eicosanoic (arachidic; C20:0) acids] had no significant effect. Treatment of the neutrophils with exogenous C22:6(n-3) caused the mass of endogenous unesterified C20:4(n-6) to increase. Incubation of the leucocytes with C20:4(n-6) or C22:6(n-3) evoked activation of the 85 kDa cytosolic PLA2 (cPLA2) and the 14 kDa secretory PLA2 (sPLA2), but not the cytosolic Ca2+-independent PLA2. In contrast, C20:0 did not activate any of the PLA2 isoforms. Activation of cPLA2 by PUFA was found to precede that of sPLA2. C22:6(n-3), C20:4(n-6) and other PUFA induced punctate localization of cPLA2 in the cells, which was not observed with saturated fatty acids. Pretreatment of the leucocytes with PLA2 inhibitors markedly decreased superoxide production induced by C20:4(n-6). These results show that PUFA activate PLA2 in neutrophils, which might have a mandatory role in biological responses.

Arachidonic acid triggers an oxidative burst in leukocytes

“…This action of AA has been shown by different investigators in various cell types, particularly phagocytes, endothelial cells, 3T3 fibroblasts, and cancer cell lines (11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23). Most published data indicate only a slight effect of AA on superoxide generation, probably because the superoxide assays normally used, such as the cytochrome c reduction system, can only determine extracellular superoxide.…”

Unique Effect of Arachidonic Acid on Human Neutrophil TNF Receptor Expression: Up-Regulation Involving Protein Kinase C, Extracellular Signal-Regulated Kinase, and Phospholipase A2

“…Indeed, we have demonstrated that exogenous AA caused the activation of the cPLA 2 and secretory PLA 2 (28). This was accompanied by the release of [ 3 H]AA from prelabeled cells, and this could be inhibited by AACOCF3.…”

“…We have previously demonstrated that AA stimulates the activities of PKC, ERK1/ERK2, and phospholipase A 2 (27,28). To understand the mechanisms by which AA caused the up-regulation of TNFR, the chemical inhibitors, GF109203X, PD098059, and AACOCF3, were used to target PKC, ERK1/ERK2, and the cPLA 2 , respectively.…”

Independent Functioning of Cytosolic Phospholipase A2 and Phospholipase D1 in Trp-Lys-Tyr-Met-Val-D-Met-Induced Superoxide Generation in Human Monocytes

“…Moreover, a number of previous publications have reported that stimulation of phagocytic cells with AA, the product of PLA 2 activity, leads to the activation of NADPH oxidase, which then results in superoxide generation (31,43). The involvement of the two phospholipases, cPLA 2 and PLD 1 , in superoxide generation after treatment of human monocytes with WKYMVm was, therefore, further investigated.…”