5-Lipoxygenase Products Modulate the Activity of the 85-kDa Phospholipase A2 in Human Neutrophils

Abstract

14C]arachidonoylphosphatidylcholine as substrate. A similar increase in cytosolic PLA 2 activity was induced by stimulation of neutrophils with leukotriene B 4 (LTB 4 ), 5-oxoeicosatetraenoic acid, or 5-hydroxyeicosatetraenoic acid (5-HETE). LTB 4 was the most potent of the agonists, showing maximal effect at 1 nM. Inhibition of 5-lipoxygenase with either eicosatetraynoic acid or zileuton prevented the AA-induced increase in PLA 2 activity but had no effect on the response induced by LTB 4 . Furthermore, pretreatment of neutrophils with a LTB 4 -receptor antagonist, LY 255283, blocked the AAand LTB 4 -induced activation of PLA 2 but did not influence the action of 5-HETE. Treatment of neutrophils with pancreatic PLA 2 also induced an increase in the activity of the cytosolic PLA 2 ; this response was inhibited by both eicosatetraynoic acid or LY 255283.The increases in PLA 2 activity in response to stimulation correlated with a shift in electrophoretic mobility of the 85-kDa PLA 2 , as determined by Western blot analysis, suggesting that phosphorylation of the 85-kDa PLA 2 likely underlies its increase in catalytic activity. Although stimulation of neutrophils with individual lipoxygenase metabolites did not induce significant mobilization of endogenous AA, they greatly enhanced the N-formylmethionyl-leucyl-phenylalanine-induced mobilization of AA as determined by mass spectrometry analysis. Our findings support a positive-feedback model in which stimulus-induced release of AA or exocytosis of secretory PLA 2 modulate the activity of the cytosolic 85-kDa PLA 2 by initiating the formation of LTB 4 . The nascent LTB 4 is then released to act on the LTB 4 receptor and thereby promote further activation of the 85-kDa PLA 2 . Since 5-HETE and LTB 4 are known to prime the synthesis of platelet-activating factor, the findings suggest that 85-kDa PLA 2 plays a role in platelet-activating factor synthesis.Arachidonic acid (AA) 1 is the precursor for prostaglandins and leukotrienes (collectively named eicosanoids), which together with platelet-activating factor (PAF) are important lipid mediators involved in allergic and inflammatory reactions. The initial step in the production of eicosanoids is hydrolysis of sn-2-arachidonate from membrane phospholipids generating free AA; phospholipase A 2 s (PLA 2 ) with different characteristics have been implicated in this hydrolysis (1-3). The 14-kDa PLA 2 s (type I and II) are secretory enzymes, and although they show no apparent preference for hydrolysis of AA-containing phospholipids (4, 5), they have been suggested to participate in the generation of eicosanoids after first being secreted by mobilizing AA from phospholipids on the outer leaflet of the plasma membrane (6 -12). The 85-kDa PLA 2 (13, 14) is an intracellular enzyme with clear preference for AA-containing phospholipids (5, 15, 16). Furthermore, the 85-kDa PLA 2 translocates to membranes in response to submicromolar concentrations of Ca 2ϩ (13,17,18) and is also regulated by phosphorylation, which results in an inc...

The Functions of Five Distinct Mammalian Phospholipase A2s in Regulating Arachidonic Acid Release

“…The simplest explanation for these events is that sPLA 2 s-IIA and -V activate intracellular cPLA 2 , which eventually releases AA in a selective manner. Indeed, this route is compatible with the recent studies using rat mesangial cells (77), human neutrophils (78), human astrocytoma (79), and mouse osteoblasts (19), in which sPLA 2 -IIA caused translational (19) or post-translational (77-79) activation of cPLA 2 , probably through the production of lipid mediators (19,77,78) or through the putative sPLA 2 receptor-dependent and catalysis-independent process (79). However, several lines of evidence argue against this speculation in our 293 cell system.…”

Functional Coupling between Secretory and Cytosolic Phospholipase A2 Modulates Tumor Necrosis Factor-α- and Interleukin-1β-induced NF-κB Activation

“…In addition, since the inhibitory effect of both snpPLA 2 and cPLA 2 inhibitors on [ 3 H]AA release is relieved by excess AA, we may suggest that snpPLA 2 and cPLA 2 upon cytokine stimulation act in a sequential manner, possibly in a positive feedback model. The AA metabolites LTB 4 , 5-oxoeicosatetraenoic acid, and 5-HETE has been found to activate cPLA 2 in neutrophils (63). We observed that the 5-LO inhibitor L-655,238 reduced cellular AA release by 85% in response to cytokines and that this effect was partially reversed by LTB 4 (Fig.…”

Atypical λ/ιPKC Conveys 5-Lipoxygenase/Leukotriene B4-mediated Cross-talk between Phospholipase A2s Regulating NF-κB Activation in Response to Tumor Necrosis Factor-α and Interleukin-1β

“…We have recently shown that both snpPLA 2 and cPLA 2 , through AA-derived 5-LO metabolites as LTB 4 acting through its G-protein-coupled receptor in an autocrine feedback mechanism, contribute to activation of the immunomodulatory transcription factor NF-B in response to TNF-␣ and IL-1␤ (29). Other authors (62,63) have also described that snpPLA 2 and LTB 4 (acting in an autocrine fashion) increase AA release, cPLA 2 phosphorylation, and cPLA 2 activation. However, the mechanisms explaining how snpPLA 2 /LTB 4 regulate cPLA 2 activation is not understood in detail.…”