Regulation of phospholipase A2 activation and arachidonic acid metabolism in an interleukin-3-dependent macrophage-like cell line.
An interleukin 3 (IL-3)-dependent macrophage-like cell line, 11-1-B3, was newly established from CBA/J mouse bone marrow cell cultures. Assay of eicosanoids in the culture supernatants of the intact and [3H]arachidonic acid (AA)-prelabeled cells showed that, after stimulation with the Ca2+ ionophore A23187, the 11-1-B3 cells synthesized and released relatively large amounts of prostaglandin E2 (PGE2) and leukotriene B4 (LTB4) but not LTC4. In addition, 11-1-B3 cells showed Ca(2+)-dependent and alkaline pH-optimal phospholipase A2 (PLA2) activity that preferentially hydrolyzed cleavage of sn-2-arachidonyl- but not sn-2-oleoylphosphatidylcholine. The cellular enzyme was distributed with 90% of the activity in the cytosol and 10% in the membrane fraction. Treatment of cells with A23187 for 5-10 min resulted in five- to sevenfold increases in the membrane-associated PLA2 but activity in the cytosol was unchanged. This increase in membrane-associated enzyme activity was transient, returning to the pretreatment distribution after 30 min. In sharp contrast, phorbol myristate acetate (PMA) stimulation failed to induce either eicosanoid release or PLA2 activation, although PMA induced translocation of protein kinase C (PKC) to the membrane fraction within 10 min. The data suggest that increases in cellular Ca2+ directly activate membrane-associated PLA2 and consequently initiate AA metabolism; PKC activation by PMA requires additional steps to activate PLA2, a mechanism that is apparently deficient in the IL-3-dependent M phi-like cells.
Shibata
Y
Y
Fujisaki Cell Center, Hayashibara Biochemical Laboratories Inc., Okayama, Japan.
McCaffrey
P G
PG
Minowada
J
J
Volkman
A
A
Oghiso
Y
Y
D000818:Animals; D001095:Arachidonic Acids / Q000378:metabolism*; D001692:Biological Transport / Q000502:physiology; D001853:Bone Marrow / Q000378:metabolism / Q000502:physiology; D001854:Bone Marrow Cells; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000502:physiology; D002458:Cell Fractionation; D002478:Cells, Cultured; D015232:Dinoprostone / Q000378:metabolism; D015777:Eicosanoids / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D006863:Hydrogen-Ion Concentration; D007377:Interleukin-3 / Q000502:physiology*; D007975:Leukotriene B4 / Q000378:metabolism; D008264:Macrophages / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D051379:Mice; D008808:Mice, Inbred CBA; D010741:Phospholipases A / Q000378:metabolism* / Q000502:physiology; D054467:Phospholipases A2; D011493:Protein Kinase C / Q000493:pharmacokinetics; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014316:Tritium
D001095:Arachidonic Acids; D015777:Eicosanoids; D007377:Interleukin-3; D014316:Tritium; D007975:Leukotriene B4; D000001:Calcimycin; D011493:Protein Kinase C; D010741:Phospholipases A; D054467:Phospholipases A2; D015232:Dinoprostone; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium