Suppression of monocyte 85-kDa phospholipase A2 by antisense and effects on endotoxin-induced prostaglandin biosynthesis.
Studies were conducted to characterize a human monocyte model where the role of the 85-kDa phospholipase A2 (PLA2) in prostanoid formation could be evaluated. The presence of an immunologically related 85-kDa PLA2 and type II 14-kDa PLA2 was demonstrated in human monocytes and their roles examined in lipopolysaccharide (LPS)-induced monocyte prostaglandin E2 (PGE2) formation. Exposure of human monocytes to LPS over 18 h resulted in the up-regulation of the mitogen-inducible cyclooxygenase-2 and was accompanied by production and release of prostaglandin E2 but not leukotriene C4. This coincided with a 2-fold increase in the 85-kDa PLA2 protein and activity levels. In contrast, there was no effect on the type II 14-kDa-like PLA2 activity measured in the 100,000 x g particulate fraction nor did LPS induce the release of type II 14-kDa PLA2 into the medium. Treatment with cycloheximide over 18 h resulted in a time-dependent decrease in cytosolic 85-kDa PLA2 protein and activity (half-life = 4 h), but there was no change in the particulate type II 14-kDa-like PLA2 activity. Monocytes were therefore exposed to an 85-kDa PLA2 initiation site-directed antisense oligonucleotide which specifically decreased the cytosolic 85-kDa PLA2 protein levels and activity in a concentration-dependent manner. This had no effect on the cyclooxygenase-2 (protein mass or the ability to convert arachidonic acid to PGE2) or the particulate fraction sn-2 acylhydrolytic activity but was associated with a decrease in LPS-induced PGE2 production. Taken together, these data support a role for the cytosolic 85-kDa PLA2 in LPS-induced monocyte PGE2 formation.
Roshak
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Department of Inflammation and Respiratory Pharmacology, SmithKline Beecham Pharmaceuticals, King of Prussia, Pennsylvania 19406.
Sathe
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Marshall
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D016718:Arachidonic Acid / Q000378:metabolism; D001483:Base Sequence; D015232:Dinoprostone / Q000096:biosynthesis*; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008297:Male; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009000:Monocytes / Q000187:drug effects / Q000201:enzymology*; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D010741:Phospholipases A / Q000502:physiology*; D054467:Phospholipases A2; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis
D008070:Lipopolysaccharides; D016376:Oligonucleotides, Antisense; D016718:Arachidonic Acid; D011451:Prostaglandin-Endoperoxide Synthases; D010741:Phospholipases A; D054467:Phospholipases A2; D015232:Dinoprostone