15-Lipoxygenase Metabolism of 2-Arachidonylglycerol

GENERATION OF A PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR α AGONIST*

Abstract

The recent demonstrations that cyclooxygenase-2 and leukocyte-type 12-lipoxygenase (LOX) efficiently oxygenate 2-arachidonylglycerol (2-AG) prompted an investigation into related oxygenases capable of metabolizing this endogenous cannabinoid receptor ligand. We evaluated the ability of six LOXs to catalyze the hydroperoxidation of 2-AG. Soybean 15-LOX, rabbit reticulocyte 15-LOX, human 15-LOX-1, and human 15-LOX-2 oxygenate 2-AG, providing 15(S)-hydroperoxyeicosatetraenoic acid glyceryl ester. In contrast, potato and human 5-LOXs do not efficiently metabolize this endocannabinoid. Among a series of structurally related arachidonyl esters, arachidonylglycerols serve as the preferred substrates for 15-LOXs. Steady-state kinetic analysis demonstrates that both 15-LOX-1 and 15-LOX-2 oxygenate 2-AG comparably or preferably to arachidonic acid. Furthermore, 2-AG treatment of COS-7 cells transiently transfected with human 15-LOX expression vectors or normal human epidermal keratinocytes results in the production and extracellular release of 15-hydroxyeicosatetraenoic acid glyceryl ester (15-HETE-G), establishing that lipoxygenase metabolism of 2-AG occurs in an eukaryotic cellular environment. Investigations into the potential biological actions of 15-HETE-G indicate that this lipid, in contrast to its free-acid counterpart, acts as a peroxisome proliferator-activated receptor α agonist. The results demonstrate that 15-LOXs are capable of acting on 2-AG to provide 15-HETE-G and elucidate a potential role for endocannabinoid oxygenation in the generation of peroxisome proliferator-activated receptor α agonists.

  • Abbreviations:
    AG
    arachidonylglycerol
    COX
    cyclooxygenase
    AEA
    anandamide
    LOX
    lipoxygenase
    HETE
    hydroxyeicosatetraenoic acid
    HpETE
    hydroperoxyeicosatetraenoic acid
    G
    glyceryl ester
    MS
    mass spectrometry
    LC
    liquid chromatography
    RP-HPLC
    reversed phase high performance liquid chromatography
    PPAR
    peroxisome proliferator-activated receptor
    AA
    arachidonic acid
    NHEK
    normal human epidermal keratinocyte
    HBSS
    Hepes-buffered saline solution
    • Received February 1, 2002.
    • Revision received April 14, 2002.
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    This Article

    1. The Journal of Biological Chemistry 277, 23278-23286.
    1. All Versions of this Article:
      1. M201084200v1
      2. 277/26/23278 (most recent)

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