Lipoxins formation by rat basophilic leukemia (RBL-1) cells.
Rat basophilic leukemia (RBL-1) cells (1.5 x 10(9) were incubated with (15S), 15-hydroperoxy-5,8,11-cis-13-trans-eicosatetraenoic acid (15-HPETE) (100 microM) and calcium ionophore A23187 (5 microM) for 30 min at 37 degrees C. The reaction products were extracted and separated on reverse-phase high performance liquid chromatography (RP-HPLC). The fraction of interest which showed the characteristic UV spectrum of lipoxins (LXs) (lambda max at 301 nm, shoulders at 289 nm and 317 nm) was further purified using a second RP-HPLC. This step produced four distinct peaks, three of which co-eluted with synthetic lipoxin A4 (LXA4), lipoxin B4 (LXB4), and authentic 6S-LXA4 standards, respectively. These fractions were analyzed by capillary gas chromatography and mass spectrometry (GC/MS) for structural identity. Based on these data (chromatographic profile, UV spectrum, mass spectra) and the reported criteria (Serhan/Samuelsson, 1984-88), these three fractions were identified with LXA4[(5S,6R,15S)-5,6,15-trihydroxy-7,9,13-trans-11-cis-eicosatetra enoic acid)], LXB4[(5S,14R,15S)-5,14,15-trihydroxy-6,10,12-trans-8-cis-eic osa-tetraenoic acid)] and 6S-LXA4[(5S,6S,15S)-5,6,15-trihydroxy-7,9,13-trans-11-cis-eicosate traenoic acid)]. It is concluded that RBL-1 cells have the capacity to generate LXs by metabolizing arachidonic acid (AA) derivatives (i.e., 15-HPETE) and that LXs produced by RBL-1 cells are indistinguishable from those derived from other cells.
Kim
S J
SJ
Membrane Research Institute, University City Science Center, Philadelphia, PA 19104.
D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D008401:Gas Chromatography-Mass Spectrometry; D006893:Hydroxyeicosatetraenoic Acids / Q000096:biosynthesis* / Q000302:isolation & purification; D015471:Leukemia, Basophilic, Acute / Q000378:metabolism; D015289:Leukotrienes / Q000494:pharmacology; D008054:Lipid Peroxides / Q000494:pharmacology; D044045:Lipoxins; D014407:Tumor Cells, Cultured
D006893:Hydroxyeicosatetraenoic Acids; D015289:Leukotrienes; D008054:Lipid Peroxides; D044045:Lipoxins; C040527:lipoxin A4; D000001:Calcimycin; C025086:15-hydroperoxy-5,8,11,13-eicosatetraenoic acid; C042651:lipoxin B4