Separation of agonist-stimulated arachidonate mobilization from subsequent leukotriene B4 synthesis in human neutrophils: different effects of oleoylacetylglycerol and phorbol myristate acetate as priming agents.
Preincubation of human neutrophils with phorbol esters or soluble diglycerides enhances subsequent f-Met-Leu-Phe (fMLP)-stimulated arachidonate mobilization and leukotriene B4 (LTB4) synthesis. We have recently reported that 1,3-dioctanoylglycerol (1,3-diC8) is equipotent with 1,2-sn-dioctanoylglycerol (1,2-diC8) as priming agent, thus suggesting that the priming effects of diacylglycerols are protein kinase C (PKC) independent (Rosenthal et al., 1993, Biochim. Biophys. Acta 1177:79-86). In order to further investigate this question, the present study has directly compared the effects of oleoylacetylglycerol (OAG) and the PKC activator, phorbol 12-myristate 13-acetate (PMA), on agonist-stimulated lipid metabolism. The results indicate that both OAG and PMA dose dependently enhance f-Met-Leu-Phe (fMLP)-stimulated release of [3H]arachidonate. Optimal concentrations of OAG (5 microns) and PMA (10 nM) are equipotent in increasing fMLP-stimulated arachidonate mobilization as quantitated either with total radioactivity or by mass measurements of free arachidonate. By contrast OAG is sixfold more effective than PMA in enhancing synthesis of 5-lipoxygenase (5-LO) metabolites by mass and two to threefold more effective than PMA in enhancing synthesis of [3H]eicosanoids. Furthermore, OAG, but not PMA, enhances fMLP-stimulated synthesis of platelet-activating factor. By contrast, PMA directly stimulates [3H]arachidonate mobilization, while OAG (20 microM) does not; despite these differences, the combined effects of PMA + OAG on subsequent agonist-stimulated arachidonate release are not greater than those of PMA alone. In cells challenged with subthreshold concentrations (< 0.1 microM) of the calcium ionophore A23187, both OAG and PMA stimulate [3H]arachidonate release but not [3H]LTB4 synthesis. These findings suggest that OAG does not directly activate 5-LO, but instead couples arachidonate mobilization to leukotriene synthesis in a PKC-independent manner.
Rosenthal
M D
MD
Department of Biochemistry, Eastern Virginia Medical School, Norfolk 23501.
Franson
R C
RC
D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase; D016718:Arachidonic Acid / Q000378:metabolism*; D001692:Biological Transport; D000001:Calcimycin / Q000494:pharmacology; D004075:Diglycerides / Q000494:pharmacology*; D015777:Eicosanoids / Q000096:biosynthesis; D005227:Fatty Acids / Q000378:metabolism; D006801:Humans; D006893:Hydroxyeicosatetraenoic Acids / Q000096:biosynthesis; D007211:Indoles / Q000494:pharmacology; D020024:Leukotriene Antagonists; D007975:Leukotriene B4 / Q000096:biosynthesis*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D009504:Neutrophils / Q000378:metabolism*; D010741:Phospholipases A / Q000096:biosynthesis; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D002722:Chlorobenzenes
D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase; D016718:Arachidonic Acid; D000001:Calcimycin; D004075:Diglycerides; D015777:Eicosanoids; D005227:Fatty Acids; D006893:Hydroxyeicosatetraenoic Acids; D007211:Indoles; D020024:Leukotriene Antagonists; D007975:Leukotriene B4; D009240:N-Formylmethionine Leucyl-Phenylalanine; D010741:Phospholipases A; D013755:Tetradecanoylphorbol Acetate; C060893:MK-886; D002722:Chlorobenzenes; C022022:5-hydroxy-6,8,11,14-eicosatetraenoic acid; C038237:1-oleoyl-2-acetylglycerol