Thrombocytes are the predominant source of endogenous sulfidopeptide leukotrienes in the American bullfrog (Rana catesbeiana).
Nucleated bullfrog erythrocytes have 5-lipoxygenase (LO) and are the first non-mammalian cell to exhibit endogenous sulfidopeptide leukotriene (LT) synthesis. Non-nucleated mammalian platelets lack 5-LO, but contribute significantly to LTC4 production by transcellular synthesis. However, nucleated bullfrog thrombocytes have not been examined for 5-LO activity. Endogenous leukotriene synthesis by bullfrog thrombocytes and mixed leukocytes was analyzed by reverse-phase high-performance liquid chromatography (RP-HPLC). Calcium ionophore activated (A23187) leukocytes demonstrated 5-LO, 12-LO, and 15-LO activity. Spectral analysis demonstrated synthesis of LTB4, LTB4 isomers, 15(S)-monohydroxyicosatetraenoic acid (HETE), 5(S),12(S)-diHETE, 5(S),15(S)-di-HETE, lipoxin A4 (LXA4) and LXB4. Thrombocytes synthesized large quantities of sulfidopeptide leukotrienes but no lipoxins. Sulfidopeptide leukotriene and LTB4 radioimmunoassay analysis and the radiological RP-HPLC profile of [3H]AA metabolism further confirmed synthesis. Incubations with [3H]LTC4 demonstrated slow and incomplete conversion to [3H]LTD4. Thrombocyte leukotriene profile changed over time revealing a significant shift from the LTC4 synthase to LTA4 hydrolase pathway, corresponding with release of large amounts of LTA4. Thrombocytes potentially play a pivotal role in inflammatory and cardiovascular responses. 5-LO activity in amphibian homologs to mammalian platelets and erythrocytes compared with the lack of activity in the mammalian counterparts may correspond to the loss of the nucleus in the evolution of these cells.
Gronert
K
K
Department of Biology, New Mexico State University, Las Cruces 88003, USA.
Virk
S M
SM
Herman
C A
CA
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D001792:Blood Platelets / Q000378:metabolism*; D000001:Calcimycin / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D006893:Hydroxyeicosatetraenoic Acids / Q000096:biosynthesis / Q000097:blood; D007962:Leukocytes / Q000201:enzymology / Q000378:metabolism; D015289:Leukotrienes / Q000096:biosynthesis* / Q000097:blood / Q000737:chemistry; D008084:Lipoxygenase / Q000097:blood*; D016859:Lipoxygenase Inhibitors / Q000494:pharmacology; D009637:Masoprocol / Q000494:pharmacology; D011863:Radioimmunoassay; D011892:Rana catesbeiana / Q000097:blood*; D013057:Spectrum Analysis; D005723:gamma-Glutamyltransferase / Q000097:blood
D006893:Hydroxyeicosatetraenoic Acids; D015289:Leukotrienes; D016859:Lipoxygenase Inhibitors; D016718:Arachidonic Acid; D000001:Calcimycin; D009637:Masoprocol; D008084:Lipoxygenase; D005723:gamma-Glutamyltransferase