Formation of a 5-oxo metabolite of 5,8,11,14,17-eicosapentaenoic acid and its effects on human neutrophils and eosinophils.

Open AccessPublished:December 01, 1995DOI:
      This paper is only available as a PDF. To read, Please Download here.
      We recently showed that human neutrophils convert arachidonic acid to its 5-oxo metabolite, 5-oxo-6,8,11,14-eicosatetraenoic acid (5-oxo-ETE). 5-Oxo-ETE, which is synthesized by oxidation of 5-hydroxy-6,8,11,14-eicosatetraenoic acid (5-HETE) by a highly specific microsomal dehydrogenase, is a potent stimulator of human neutrophils and eosinophils. The objective of the current investigation was to determine whether neutrophils can convert 5,8,11,14,17-eicosapentaenoic acid (EPA) to its 5-oxo metabolite, 5-oxo-6,8,11,14,17-eicosapentaenoic acid (5-oxo-EPE) and, if so, to compare the biological activities of 5-oxo-EPE and 5-oxo-ETE. The two major eicosanoids formed by neutrophils incubated with EPA in the presence of A23187 were 5-hydroxy-6,8,11,14,17-eicosapentaenoic acid (5-HEPE) and 5-oxo-EPE. Smaller amounts of LTB5 and 20-hydroxy-LTB5 were also formed. Phorbol myristate acetate stimulated the formation of 5-oxo-EPE from both EPA and 5-HEPE. 5-HEPE and 5-HETE were equally good substrates for 5-hydroxyeicosanoid dehydrogenase (Km, ca. 0.85 microM; Vmax, ca. 1.4 pmol/min per microgram protein). 5-Oxo-EPE mobilized calcium in neutrophils with an EC50 of 36 nM, about 10 times higher than that of 5-oxo-ETE. 5-Oxo-EPE was also about one-tenth as active as 5-oxo-ETE in stimulating the migration of both human neutrophils and human eosinophils. It is concluded that 5-oxo-EPE is readily formed from EPA via 5-HEPE. However, it is only about one-tenth as potent as 5-oxo-ETE in stimulating human neutrophils and eosinophils. These results support the contention that EPA can alleviate certain inflammatory diseases by reducing the contribution of arachidonate-derived eicosanoids.


        • Borgeat P.
        • Samuelsson B.
        Metabolism of arachidonic acid in polymorphonuclear leukocytes.
        Structural analysis of novel hydroxylated compounds. J Biol.Chem. 1979; 254: 7865-7869
        • Murphy R.C.
        • Pickett W.C.
        • Culp B.R.
        • Lands W.E.
        Tetraene and pentaene leukotrienes: selective production from murine mastocytoma cells after dietarymanipulation.
        Prostaglandins. 1981; 22: 613-622
        • Lee T.H.
        • Mencia-Huerta J.M.
        • Shih C.
        • Corey E.J.
        • Lewis R.A.
        • Austen K.F.
        Effects of exogenous arachidonic, eicosapentaenoic,and docosahexaenoic acids on the generation of 5-lipoxygenase pathway products by ionophore-activated human neutrophils.
        J Clin. Invest. 1984; 74: 1922-1933
        • Powell W.S.
        Properties of leukotriene B4 20-hydroxylase from polymorphonuclear leukocytes.
        J. Biol.Chem. 1984; 259: 3082-3089
        • Shak S.
        • Goldstein I.M.
        Leukotriene B4 omega-hydroxylase in human polymorphonuclear leuko-cytes.
        Partial purification and identification as a cyto-chrome P-450.J. Clin. Invest. 1985; 76: 1218-1228
        • Hansson G.
        • Lindgren J.A.
        • Dahlen S.E.
        • Hedqvist P.
        • Samuelsson B.
        Identification and biological activity of novel omega-oxidized metabolites of leukotriene B4 from human leukocytes.
        FEBS Lett. 1981; 130: 107-112
        • Wainwright S.L.
        • Powell W.S.
        Mechanism for the formation of dihydro metabolites of 12-hydroxyei-cosanoids.
        Conversion ofleukotriene B4 and 12-hydroxy- 5,8, 10,14-eicosatetraenoic acid to 12-oxo intermediates.J.Biol. Chem. 1991; 266: 20899-20906
        • Powell W.S.
        • Gravelle F.
        Metabolism ofleukotriene B4 to dihydro and dihydro-oxo products by porcine leukocytes.
        J. Bioi. Chem. 1989; 264: 5364-5369
        • Ford-Hutchinson A.W.
        • Rackham A.
        • Zamboni R.
        • Rokach J.
        • Roy S.
        Comparative biological activities of synthetic leukotriene B4 and its omega-oxidation products.
        Prostaglandins. 1983; 25: 29-37
        • Jubiz W.
        • Radmark O.
        • Malmsten C.
        • Hansson G.
        • Lindgren J.Å.
        • Palmblad J.
        • Uden A.M.
        • Samuelsson B.
        A novel leukotriene produced by stimulation of leukocytes with formylmethionylleucylphenylalanine.
        J. Biol. Chem. 1982; 257: 6106-6110
        • Soberman R.J.
        • Okita R.T.
        • Fitzsimmons B.
        • Rokach J.
        • Spur B.
        • Austen K.F.
        Stereochemical requirements for substrate specificity of LTB4 20-hydroxylase.
        J.Biol. Chem. 1987; 262: 12421-12427
        • Wainwright S.
        • Falck J.
        • Yadagiri P.
        • Powell W.
        Metabolism of 12(S)-hydroxy-5,8, 10,14-eicosatetraenoic acid and other hydroxylated fatty acids by the reductase pathway in porcine polymorphonuclear leukocytes.
        Biochemistry. 1990; 29: 10126-10135
        • O'Flaherty J.
        • Wykle R.
        • Redman J.
        • Samuel M.
        • Thomas M.
        Metabolism of 5-hydroxyicosatetraenoate by human neutrophils: production of a novel omega-oxidized derivative.
        J. Immunol. 1986; 137: 3277-3283
        • Maas R.L.
        • Turk J.
        • Oates J.A.
        • Brash A.R.
        Formation of a novel dihydroxy acid from arachidonic acid by lipoxygenase-catalyzed double oxygenation in rat mononuclear cells and human leukocytes.
        J. Bioi. Chem. 1982; 257: 7056-7067
        • Borgeat P.
        • Picard S.
        • Vallerand P.
        • Sirois P.
        Transformation of arachidonic acid in leukocytes.
        Isolation and structural analysis of a novel dihydroxy derivative. Prostaglandins Med. 1981; 6: 557-570
        • Powell W.S.
        • Gravelle F.
        • Gravel S.
        Metabolism of 5(S)-hydroxy-6,8, 11,14-eicosatetraenoic acid and other 5(S)-hydroxyeicosanoids by a specific dehydrogenase in human polymorphonuclear leukocytes.
        J. Biol. Chem. 1992; 267: 19233-19241
        • Powell W.S.
        • Chung D.
        • Gravel S.
        5-0xo- 6,8,11,14-eicosatetraenoic acid is a potent stimulator of human eosinophil migration.
        J.Immunol. 1995; 154: 4123-4132
        • Powell W.S.
        • Gravelle F.
        • Gravel S.
        Phorbolmyristate acetate stimulates the formation of 5-oxo- 6,8,11,14-eicosatetraenoic acid by human neutrophils by activating NADPH oxidase.
        J. Biol. Chem. 1994; 269: 25373-25380
        • Powell W.S.
        • Gravel S.
        • MacLeod R.J.
        • Mills E.
        • Hashefi M.
        Stimulation of human neutrophils by 5-oxo-6,8, 11,14-eicosatetraenoic acid by a mechanism independent of the leukotriene B4 receptor.
        J. Biol. Chem. 1993; 268: 9280-9286
        • O'Flaherty J.T.
        • Cordes J.
        • Redman J.
        • Thomas M.J.
        5-0xo-eicosatetraenoate, a potent human neutrophil stimulus.
        Biochern. Biophys. Res. Commun. 1993; 192: 129-134
        • O'Flaherty J.T.
        • Cordes J.F.
        • Lee S.L.
        • Samuel M.
        • Thomas M.J.
        Chemical and biological characterization of oxo-eicosatetraenoic acids.
        Biochim. Biophys.Acta. 1994; 1201: 505-515
        • Schwenk U.
        • Schröder J.M.
        5-0xo-eicosanoids are potent eosinophil chemotactic factors: functional characterizationand structural requirements.
        J. Biol. Chem. 1995; 270: 15029-15036
        • Simopoulos A.P.
        Omega-3 fatty acids in health and disease and in growth and development.
        Am.J. Clin. Nutr. 1991; 54: 438-463
        • Powell W.S.
        • Zhang Y.
        • Gravel S.
        Effects of phorbol myristate acetate on the synthesis of 5-oxo-6,8, 11,14-eicosatetraenoic acid by human polymorphonuclear leukocytes.
        Biochemistry. 1994; 33: 3927-3933
        • Borgeat P.
        • Picard S.
        19-Hydroxypro-staglandin B2 as an internal standard for on-line extraction-high-performance liquid chromatography analysis of lipoxygenase products.
        Anal. Biochem. 1988; 171: 283-289
        • Boyum A.
        Isolation of mononuclear cells and granulocytes from human blood.
        Isolation of monuclear cells by one centrifugation, and of granulocytes by combining centrifugation and sedimentation at 1 g. Scand.J. Clin. Lab. Invest. 21, Suppl. 1968; 97: 77-89
        • Hansel T.T.
        • De Vries I.J.
        • Iff T.
        • Rihs S.
        • Wandzilak M.
        • Betz S.
        • Blaser K.
        • Walker C.
        An improved immunomagnetic procedure for the isolation of highly purified human blood eosinophils.
        J. Immunol. Methods. 1991; 145: 105-110
        • Powell W.S.
        Precolumn extraction and reversed-phase high-pressure liquid chromatography of pro-staglandins and leukotrienes.
        Anal. Biochem. 1987; 164: 117-131
        • Gelfand E.W.
        • Cheung R.K.
        • Grinstein S.
        Mitogen-induced changes in Ca2+ permeability are not mediated by voltage-gated K+ channels.
        J. Biol. Chem. 1986; 261: 11520-11523
        • Grynkiewicz G.
        • Poenie M.
        • Tsien R.Y.
        A new generation of Ca2+ indicators with greatly improved fluorescence properties.
        J. Biol. Chem. 1985; 260: 3440-3450
        • Falk W.
        • Goodwin Jr., R.H.
        • Leonard E.J.
        A 48-well micro chemotaxis assembly for rapid and accurate measurement ofleukocyte migration.
        J. Immunol. Methods. 1980; 33: 239-247
        • Sehmi R.
        • Wardlaw A.J.
        • Cromwell O.
        • Kurihara K.
        • Waltmann P.
        • Kay A.B.
        Interleukin-5 selectively enhances the chemotactic response of eosinophils obtained from normal but not eosinophilic subjects.
        Blood. 1992; 79: 2952-2959
        • Kay A.B.
        Studies on eosinophil leucocyte migration.
        II. Factors specifically chemotactic for eosinophils and neutrophils generated from guinea-pig serum by antigen-antibody complexes. Clin. Exp. Immunol. 1970; 7: 723-737
        • Lin L.L.
        • Lin A.Y.
        • Knopf J.L.
        Cytosolic phospholipase A2 is coupled to hormonally regulated release of arachidonic acid.
        Proc. Natl. Acad. Sci. USA. 1992; 89: 6147-6151
        • Needleman P.
        • Whitaker M.O.
        • Wyche A.
        • Watters K.
        • Sprecher H.
        • Raz A.
        Manipulation of platelet aggregation by prostaglandins and their fatty acid precursors: pharmacological basis for a therapeutic approach.
        Prostaglandins. 1980; 19: 165-181
        • Powell W.S.
        • Gravelle F.
        Metabolism of eicos-apentaenoic acid by aorta: formation of a novel 13-hydroxylated prostaglandin.
        Biochim. Biophys. Acta. 1985; 835: 201-211
        • Yerram N.R.
        • Moore S.A.
        • Spector A.A.
        Eicosapentaenoic acid metabolism in brain microvessel endothelium: effect on prostaglandin formation.
        J Lipid Res. 1989; 30: 1747-1757
        • Thien F.C.
        • Hallsworth M.P.
        • Soh C.
        • Lee T.H.
        Effects of exogenous eicosapentaenoic acid on generation of Jeukotriene C4 and leukotriene C5 by calcium ionophore-activated human eosinophils in vitro.
        J. Immunol. 1993; 150: 3546-3552
        • Nathaniel D.J.
        • Evans J.F.
        • Leblanc Y.
        • Leveille C.
        • Fitzsimmons B.J.
        • Ford-Hutchinson A.W.
        Leukotriene A5 is a substrate and an inhibitor of rat and humanneutrophil LTA4 hydrolase.
        Biochem. Biophys. Res. Commun. 1985; 131: 827-835
        • Goldman D.W.
        • Pickett W.C.
        • Goetz) E.J.
        Human neutrophil chemotactic and degranulating activities of leukotriene B5 (LTB5) derived from eicosapentaenoic acid.
        Biochem. Biophys. Res. Commun. 1983; 117: 282-288
        • Lee T.H.
        • Mencia-Huerta J.M.
        • Shih C.
        • Corey E.J.
        • Lewis R.A.
        • Austen K.F.
        Characterization and biologic properties of 5,12-dihydroxy derivatives of eicosapentaenoic acid, including Jeukotriene B5 and the double lipoxygenase product.
        J. Biol. Chem. 1984; 259: 2383-2389
        • Seya A.
        • Terano T.
        • Tamura Y.
        • Yoshida S.
        Comparative effect of leukotriene B4 and leukotriene B5 on calcium mobilizationin human neutrophils.
        Prostaglandins Leukot. Essent. Fatty Acids. 1988; 34: 47-50
        • Leitch A.G.
        • Lee T.H.
        • Ringel E.W.
        • Prickett J.D.
        • Robinson D.R.
        • Pyne S.G.
        • Corey E.J.
        • Drazen J.M.
        • Austen K.F.
        • Lewis R.A.
        Immunologically induced generation of tetraene and pentaene leukotrienes in the peritoneal cavities of menhaden-fed rats.
        J. Immunol. 1984; 132: 2559-2565
        • Dahlen S.E.
        • Hedqvist P.
        • Hammarstrom S.
        Contractile activities of several cysteine-containing leukotrienes in the guinea-pig lung strip.
        Eur.J. Pharmacal. 1982; 86: 207-215