Abstract
Herein, we demonstrate that nitric oxide is a potent (> 20% release) and highly selective inducer of [3H]arachidonic acid mobilization in the macrophage-like cell line RAW 264.7. Treatment of RAW 264.7 cells with (E)-6-(bromomethylene)-3-(1-naphthalenyl)-2H-tetrahydropyran-2-one resulted in the inhibition of the large majority (86%) of nitric oxide-induced [3H]arachidonic acid release into the medium (IC50 < 0.5 microM) and the concomitant inhibition of in vitro measurable calcium-independent phospholipase A2 activity (92% inhibition) without demonstrable effects on calcium-dependent phospholipase A2 activity. Since nitric oxide is a potent stimulator of glycolysis (and therefore glycolytically derived ATP) and since cytosolic calcium-independent phospholipase A2 exists as a catalytic complex comprised of ATP-modulated phosphofructokinase-like regulatory polypeptides and a catalytic subunit, we examined the role of glucose in facilitating nitric oxide-mediated arachidonic acid release. Nitric oxide-induced release of [3H]arachidonic acid possessed an obligatory requirement for glucose, was highly correlated with the concentration of glucose in the medium, and was dependent on the metabolism of glucose. Thus, [3H]arachidonic acid release is coupled to cellular glucose metabolism through alterations in the activity of calcium-independent phospholipase A2. Collectively, these results identify a unifying metabolic paradigm in which the generation of lipid second messengers is coordinately linked to the signalstimulated acceleration of glycolytic flux, thereby facilitating integrated metabolic responses to cellular stimuli.
Highlights
From the Division of Bioorganic Chemistry and Molecular Pharmacology, Departments of Internal Medicine, Chemistry, and Molecular Biology & Pharmacology, Washington University School of Medicine, St
Since nitric oxide is a potent stimulator of glycolysis and since cytosolic calcium-independent phospholipase ~ exists as a catalytic complex comprised of ATP-modulated phosphofructokinase-like regulatory polypeptides and a catalytic subunit, we examined the role of glucose in facilitating nitric oxidemediated arachidonic acid release
Since nitric oxide is a potent stimulator of macrophage glucose uptake and glycolytic flux (7, 17), we hypothesized that some of the effects of nitric oxide on macrophage phospholipid catabolism and physiologic function could result from the calciumindependent phospholipase ~-mediated mobilization of arachidonic acid in response to the nitric oxide-induced accelerated production of glycolytically derived ATP
Summary
Macrophage activation induced by a variety of different ligands (e.g. lipopolysaccharide, zymosan, etc.) results in increased cellular glycolytic flux, accelerated phospholipid catabolism, and stimulation of many macrophage physiologic functions (e.g. phagocytosis and chemotaxis) (7, 8) Macrophages contain both calcium-dependent (9, 10) and calciumindependent phospholipase A2 (11) activities that are catalyzed by separate and distinct classes of polypeptides Since nitric oxide is a potent stimulator of macrophage glucose uptake and glycolytic flux (7, 17), we hypothesized that some of the effects of nitric oxide on macrophage phospholipid catabolism and physiologic function could result from the calciumindependent phospholipase ~-mediated mobilization of arachidonic acid in response to the nitric oxide-induced accelerated production of glycolytically derived ATP. We report that: 1) nitric oxide is a potent activator of arachidonic acid mobilization in macrophages; 2) the highly selective mobilization of arachidonic acid by nitric oxide is predominantly mediated by calcium-independent phospholipase A2 ; and 3) nitric oxide-mediated arachidonic acid mobilization by calcium-independent phospholipase ~ possesses an obligatory dependence on glucose uptake and metabolism
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