Regulation of 5-lipoxygenase activity by the glutathione status in human polymorphonuclear leukocytes

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Abstract
The influence of the glutathione status of human polymorphonuclear leukocytes (PMN) on 5-lipoxygenase activity was studied by treating cells with increasing concentrations of 1-chloro-2,4-dinitrobenzene (Dnp-Cl) or azodicarboxylic acid bis(dimethylamide) (Diamide). Subsequent incubation with arachidonate resulted in an up to tenfold-stimulated formation of 5-hydroxyeicosatetraenoic acid, leukotriene B4, leukotriene B4 isomers and ω-hydroxyleukotriene B4. Higher concentrations of the GSH reagents were inhibitory. At maximal stimulation by Dnp-Cl, 5-hydroperoxyeicostetraenoic acid started to be built up at the expense of 5-HETE at glutathione levels which were diminished by about 50% compared to resting cells. No increase in cytosolic Ca2+ could be measured under these conditions by the fura-2 method. In PMN homogenates Dnp-Cl and Diamide were without effect and even caused inhibition when 5-lipoxygenase was stimulated by Ca2+ and ATP. 15-Lipoxygenase was either unchanged in the case of Diamide, or even increased after pretreatment with Dnp-Cl. The results allow us to conclude that 5-lipoxygenase activity in intact PMN is regulated not only by Ca2+ but in a complex manner also by the glutathione redox status. Conditions of oxidative stress increase the activity which may reflect the in vivo situation under phagocytosis and oxidative burst.