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 f irst non-mammalian cell to exhibit endogenous sulfidopeptide leukotrie ne (LT) synthesis. Non-nucleated mammalian platelets lack 5-LO, but co ntribute significantly to LTC(4) production by transcellular synthesis . However, nucleated bullfrog thrombocytes have not been examined for 5-LO activity. Endogenous leukotriene synthesis by bullfrog thrombocyt es 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 ana lysis demonstrated synthesis of LTB(4), LTB(4) isomers, 15(S)-monohydr oxyicosatetraenoic acid (HETE), 5(S),12(S)-diHETE, 5(S),15(S)-diHETE, lipoxin A(4) (LXA(4)) and LXB(4). Thrombocytes synthesized large quant ities of sulfidopeptide leukotrienes but no lipoxins. Sulfidopeptide l eukotriene and LTB(4) radioimmunoassay analysis and the radiological R P-HPLC profile of [H-3]AA metabolism further confirmed synthesis. Incu bations with [H-3]LTC(4) demonstrated slow and incomplete conversion t o [H-3]LTD(4). Thrombocyte leukotriene profile changed over time revea ling a significant shift from the LTC(4) synthase to LTA(4) hydrolase pathway, corresponding with release of large amounts of LTA(4). Thromb ocytes potentially play a pivotal role in inflammatory and cardiovascu lar responses. 5-LO activity in amphibian homologs to mammalian platel ets and erythrocytes compared with the lack of activity in the mammali an counterparts may correspond to the loss of the nucleus in the evolu tion of these cells.