Pathways controlling the superoxide response during phagocyte differentiation: involvement of arachidonic acid and Ca²⁺in the response to bacterial endotoxin

Abstract

In contrast to the phorbol ester oxidative response, which only develops during dimethyl‐sulphoxide (DMSO)‐induced differentiation of the human leukemic myeloblast HL‐60 cell‐line, the endotoxin response was observed in undifferentiated and differentiated cells. The Ca2+ response to endotoxin, detected in both differentiated and undifferentiated HL‐60 cells, consisted of a transient 10–50 nM increase in intracellular Ca2+. A very slow, irreversible increase in intracellular Ca2+ was detected at high 1–100 μg/ml endotoxin concentrations, and this effect, and the inositol phosphate response, correlated with the surfactant activities of various endotoxins and Lipid A. Arachidonic acid and sodium arachidonate 1–50 μM stimulated a large 200–500 nM and transient Ca2+ response in undifferentiated HL‐60 cells, which was significantly greater than that elicited by 1–50 μM eicosapentaenoic acid, and was not observed at similar concentrations of arachidonic acid methyl ester or myristic acid. These concentrations (1–50 μM) of arachidonic acid were observed to have surfactant activities on the plasma membrane. At lower arachidonic acid concentrations a marked potentiation of both Ca2+ and oxidative responses to the chemotactic peptide fMet‐Leu‐Phe was detected. It is possible that the arachidonic acid released during phospholipase A2 activation of neutrophils may be involved in cellular cross‐talk and, at higher concentrations, in directly activating Ca2+ and superoxide production. It is also possible that previously reported effects of endotoxin at high concentrations are an vitro artefact of surfactant properties of endotoxin.

Arachidonic acid in the reticuloendothelial network

“…During this process, cytotoxic superoxide anions (0;) are produced by a wide variety of immunocompetent cells in response to appropriate stimulation. Arachidonic acid activates NADPH oxidase [7,26]. This arachidonic acid release may be obligatory in Fc y receptor stimulation [ 51, and subsequent G-protein activation [8].…”

“…This activation may in turn depend on the activation of leucocyte G-proteins by arachidonic acid. In addition, arachidonic acid exerts direct effects on phagocyte H + and Ca2+ ion flux [6,7,19,21] and effects have been characterized at single channel level. Thus free arachidonic acid, with a limited range owing to enzymes of esterification and metabolism, activates phagocyte NADPH oxidase over short distances [5,11,20] (Figure 1).…”

“…In some pathways, such as Fc y-mediated superoxide generation [S] and in tumour necrosis factor (TNF)-induced cytotoxicity [lo], arachidonic acid appears to be an obligatory mediator. In other pathways such as lipopolysaccharide priming, the release of arachidonic acid may facilitate the responses to other stimuli [7,11,15,16,20].…”