Nanomolar arachidonic acid influences the respiratory burst in eosinophils and neutrophils induced by GTP‐binding protein

Abstract

To investigate a possible role of phospholipase A, (PLA,) in the respiratory burst in bovine eosinophilic and neutrophilic leukocytes dependent on GTP-binding protein (G-protein), we permeabilized these cells with Staphylococcus aureus a-toxin and induced NADPH oxidase activity with the non-hydrolysable GTP analogue GTP [S] or the aluminium tetrafluoro complex AlF,-. Under same experimental conditions, cells responded with different onset times. The onset time for eosinophils was 50-200 s, for neutrophils it was only a few seconds. GTP[S] stimulated in neutrophils only 5 % of the respiratory burst compared to eosinophils, whereas AlF,--induced comparable responses (neutrophils 120% of eosinophils). GDP inhibited these responses with an IC,, value of 2.4 mM. Arachidonic acid showed, with the exception of AIF,-stimulated neutrophils, on both stimuli and cell types an enhancing effect (150%) that reached its maximum at 0.1-1 pM. The PLA, inhibitor 4-bromophenacylbromide reduced the GTP[S]-and AlF,--induced response almost completely (10 pM) and the inhibition was not significantly different for eosinophils and neutrophils (IC50 1 -3 pM). If the respiratory burst was reduced with 4-bromophenacylbromide to 1 -4% of the original value, 10% of the basal NADPH oxidase activity could be restored by addition of only 20-100 nM arachidonic acid. In addition, the PLA, activator adriamycin enhanced the response in a dose-dependent manner and in the same order as arachidonic acid did. The results presented above suggest that the respiratory burst may be regulated by different low-molecular-mass and/or heterotrimeric G-proteins and an active role for arachidonic acid or its metabolites in the activation and the maintenance of the direct G-protein-stimulated respiratory burst in bovine eosinophils and neutrophils.When stimulated by particulate or soluble stimuli like opsonized zymosan, formyl-methionyl-leucyl-phenylalanine, leuktriene B,, platelet-activating factor or the anaphylatoxine C,a, phagocytes of the immune system generate superoxide (O;-) and other reduced oxygen species by using an NADPH oxidase, for reviews, see [l -31. of Fribourg, me du mussCe 5 , CH-1700 Fribourg, Switzerland cytosolic factors p47ph0" and p67ph"", the small G-protein p21r"'1'2 and an associated GDP dissociation inhibitor that have been identified as components of the activating mechanism at least in the cell-free system [4-61.The phosphorylation of several proteins has been reported. The phosphorylation of the p47phoX occurs in normal cells but has been reported to be absent in patients with autosomal recessive chronic granulomatous disease 171. There have been reports suggesting that phospholipase A, (PLA,) is involved in the activation of the respiratory burst oxidase. An increase in PLA, activity and a release of arachidonic acid has been shown to occur following the stimulation of NADPH oxidase by different stimuli in neutrophils as well as in macrophages [8-101. Quinacrine and 4-bromophenacylbromide, inhibitors of PLA,, have previously be...

Voltage‐activated proton current in eosinophils from human blood.

“…The threshold calcium concentration necessary to elicit detectable enhancement of voltage-gated proton conductance was found to lie between 041 and 1 UM, that is, within the range which has been detected during eosinophil activation (Kernen et al 1991). It should be noted that these observations differ from those made in snail neurones where elevation of intracellular calcium up to 10 /M was without effect on the H+ current (Byerly et al 1984 (Rossi, 1986;Henderson & Chappell, 1992;Aebischer, Pashe & Jorg, 1993). Eosinophils may also come into contact with high concentrations of arachidonic acid released by other cell types at sites of severe inflammation (Hammarstrom, Hamberg, Samulesson, Duell, Stawiski & Voorhees, 1975).…”

“…Activation of eosinophils by many receptor-directed agonists also results in the liberation of arachidonic acid with eosinophilic inflammation (Sanders, Zweier, Harrison, from its storage sites in membrane lipids and its subsequent oxidative metabolism by cyclo-oxygenase and lipoxygenase pathways. Mobilization of arachidonic acid within the eosinophil is an important component of the activation of the respiratory burst oxidase and degranulation (Rossi, 1986;Henderson & Chappell, 1992;Aebischer, Pashe & Jorg, 1993). Eosinophils may also come into contact with high concentrations of arachidonic acid released by other cell types at sites of severe inflammation (Hammarstrom, Hamberg, Samulesson, Duell, Stawiski & Voorhees, 1975).…”

Early signalling events implicated in leukotriene B4-induced activation of the NADPH oxidase in eosinophils: role of Ca2+, protein kinase C and phospholipases C and D

“…The intracellular sequence of events that precede, and ultimately promote, the assembly of an active oxidase complex is incompletely understood but appears to depend on the nature of the activating stimulus [11,21]. In neutrophils, persuasive evidence is available that soluble agonists utilize a pathway(s) that involves the participation of phospholipase A2 (PLA2), phospholipase C (PLC), PLD, protein kinase C (PKC), Ptdlns 3-kinase and one or more tyrosine kinases that are not necessarily mutually exclusive [22][23][24][25][26][27][28][29][30][31][32]. Mechanistically, perhaps the best studied stimulant of O°generation in the neutrophil is formylmethionylleucylphenylalanine (fMLP).…”

Effects of dietary arachidonic acid on human immune response

“…Results from a study with Syrian hamsters indicate that dietary AA was metabolized differently from the endogenously synthesized AA (11). Addition of AA or its metabolites to cultured immune cells altered their activity and the eicosanoids produced in vitro (12)(13)(14)(15)(16)(17). These studies suggest that AA and its metabolites may play an important role in modulating IR, and that the metabolism of AA may be altered depending on its source.…”