Article

Effect of arachidonic acid reacylation on leukotriene biosynthesis in human neutrophils stimulated with granulocyte-macrophage colony-stimulating factor and formyl-methionyl-leucyl-phenylalanine.

Department of Pharmacology, University of Colorado at Denver and Health Sciences Center, 12801 E. 17th Avenue, Aurora, CO 80045, USA.
Journal of Biological Chemistry (impact factor: 4.77). 05/2006; 281(15):10134-42. DOI:10.1074/jbc.M510783200 pp.10134-42
Source: PubMed

ABSTRACT Priming of human neutrophils with granulocyte-macrophage colony-stimulating factor (GM-CSF) followed by treatment with formyl-methionyl-leucyl-phenylalanine (fMLP) stimulates cells in a physiologically relevant manner with modest 5-lipoxygenase activation and formation of leukotrienes. However, pretreatment of neutrophils with thimerosal, an organomercury thiosalicylic acid derivative, led to a dramatic increase (>50-fold) in the production of leukotriene B(4) and 5-hydroxyeicosatetraenoic acid, significantly higher than that observed after stimulation with calcium ionophore A23187. Little or no effect was observed with thimerosal alone or in combination with either GM-CSF or fMLP. Elevation of [Ca(2+)](i) induced by thimerosal in neutrophils stimulated with GM-CSF/fMLP was similar but more sustained compared with samples where thimerosal was absent. However, [Ca(2+)](i) was significantly lower compared with calcium ionophore-treated cells, suggesting that a sustained calcium rise was necessary but not sufficient to explain the effects of this compound on the GM-CSF/fMLP-stimulated neutrophil. Thimerosal was found to directly inhibit neutrophil lysophospholipid:acyl-CoA acyltransferase activity at the doses that stimulate leukotriene production, and analysis of lysates from neutrophil preparations stimulated in the presence of thimerosal showed a marked increase in free arachidonic acid, supporting the inhibition of the reincorporation of this fatty acid into the membrane phospholipids as a mechanism of action for this compound. The dramatic increase in production of leukotrienes by neutrophils when a physiological stimulus such as GM-CSF/fMLP is employed in the presence of thimerosal suggests a critical regulatory role of arachidonate reacylation that limits leukotriene biosynthesis in concert with 5-lipoxygenase and cytosolic phospholipase A(2)alpha activation.

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Keywords

5-hydroxyeicosatetraenoic acid
 
calcium ionophore-treated cells
 
critical regulatory role
 
cytosolic phospholipase A(2)alpha activation
 
dramatic increase
 
fatty acid
 
free arachidonic acid
 
GM-CSF/fMLP
 
GM-CSF/fMLP-stimulated neutrophil
 
granulocyte-macrophage colony-stimulating factor
 
human neutrophils
 
limits leukotriene biosynthesis
 
marked increase
 
membrane phospholipids
 
modest 5-lipoxygenase activation
 
neutrophil preparations
 
neutrophils
 
organomercury thiosalicylic acid derivative
 
physiological stimulus
 
physiologically relevant manner