scholarly journals Leukotriene B4 synthesis and metabolism by neutrophils and granule-free cytoplasts

1986 ◽  
Vol 233 (2) ◽  
pp. 583-588 ◽  
Author(s):  
K A Haines ◽  
K N Giedd ◽  
G Weissmann
Keyword(s):  
Arachidonic Acid ◽  
Plasma Membrane ◽  
Leukotriene B4 ◽  
Ionophore A23187 ◽  
Tetraenoic Acid ◽  
Exogenous Arachidonic Acid ◽  
Neutrophil Degranulation ◽  
Arachidonate Metabolites

Leukotriene B4 [LTB4, (5S,12R)-hydroxyeicosa-6,14-cis-8,10-trans-tetraenoic acid], a potent mediator of inflammation, is released from neutrophils by agonists that provoke degranulation of the cell. To examine whether degranulation is a necessary requirement for synthesis and metabolism of LTB4 (or of other arachidonate metabolites), we prepared neutrophil-derived cytoplasts (neutroplasts), organelle-depleted vesicles of cytoplasm surrounded by the plasma membrane. In the presence of extracellular Ca2+ with or without exogenous arachidonic acid (150 microM), neutroplasts were exposed to the Ca2+ ionophore A23187 (10 microM) and the resultant lipoxygenation products of arachidonate were determined. Neutrophils metabolize arachidonic acid to 5-HETE greater than 15-HETE greater than LTB4 greater than all-trans-LTB4 isomers. Neutroplast products of arachidonate lipoxygenation were 15-HETE greater than 5-HETE greater than LTB4 greater than all-trans-LTB4 isomers. Neutroplasts, like neutrophils, were capable of converting LTB4 into its 20-hydroxy and 20-carboxy metabolites. Finally, neutroplasts could utilize intrinsic arachidonate, since the neutroplasts synthesized LTB4 (30 pmol/mg of protein) in the absence of added arachidonic acid. The data demonstrate that neutrophil degranulation is not required for synthesis or metabolism of LTB4 by neutrophils.

scholarly journals Na+/H+ exchange modulates the production of leukotriene B4 by human neutrophils

1989 ◽  
Vol 257 (3) ◽  
pp. 751-758 ◽  
Author(s):  
M Osaki ◽  
H Sumimoto ◽  
K Takeshige ◽  
E J Cragoe ◽  
Y Hori ◽  
...  
Keyword(s):  
Arachidonic Acid ◽  
Leukotriene B4 ◽  
Human Neutrophils ◽  
Ionophore A23187 ◽  
Cytoplasmic Ph ◽  
Chemotactic Peptide ◽  
Trans Isomers ◽  
Lipoxygenase Pathway ◽  
Exogenous Arachidonic Acid ◽  
Specific Inhibitors

Human neutrophils produce various compounds of the 5-lipoxygenase pathway, including (5S)-hydroxyeicosatetraenoic acid, leukotriene B4, its 6-trans isomers and omega-oxidation metabolites of LTB4, when the cells are stimulated with the Ca2+ ionophore A23187. The elevation in the extracellular pH (pHo) facilitated the cytoplasmic alkalinization induced by the ionophore as determined fluorometrically using 2',7'-bis(carboxyethyl)carboxyfluorescein and enhanced the production of all the 5-lipoxygenase metabolites. The production decreased when the alkalinization was blocked by the decrease in the pHo, the removal of the extracellular Na+ or the addition of specific inhibitors of the Na+/H+ exchange, such as 5-(NN-hexamethylene)amiloride, 5-(N-methyl-N-isobutyl)amiloride and 5-(N-ethyl-N-isopropyl)amiloride. The alkalinization of the cytoplasm with methylamine completely restored the production suppressed by the removal of Na+ from the medium. These findings suggest that the change in the cytoplasmic pH (pHi) mediated by the Na+/H+ exchange regulates the production of the lipoxygenase metabolites. The site of the metabolism controlled by the pHi change seemed to be the 5-lipoxygenase, because the production of all the metabolites decreased in parallel and the release of [3H]arachidonic acid from the neutrophils in response to the ionophore was not affected by the pHi change. Furthermore, the production of the 5-lipoxygenase metabolites stimulated by A23187 with or without exogenous arachidonic acid showed a similar pHo-dependence and the production induced by N-formylmethionyl-leucylphenylalanine (chemotactic peptide) with exogenous arachidonic acid also decreased when the cytoplasmic alkalinization was inhibited.

Human alveolar macrophage arachidonic acid metabolism

1988 ◽  
Vol 254 (6) ◽  
pp. C809-C815 ◽  
Author(s):  
G. P. Brown ◽  
M. M. Monick ◽  
G. W. Hunninghake
Keyword(s):  
Arachidonic Acid ◽  
Alveolar Macrophages ◽  
Biological Effects ◽  
Leukotriene B4 ◽  
Arachidonic Acid Metabolism ◽  
Ionophore A23187 ◽  
Lipoxygenase Pathway ◽  
Cyclooxygenase Inhibition ◽  
Human Alveolar Macrophage ◽  
Lipoxygenase Products

Metabolites of arachidonic acid are potent modulators of many biological events, and their release from macrophages appears to play an important role in immune and inflammatory processes. In addition, metabolites of the cyclooxygenase or lipoxygenase pathway exhibit distinct biological effects. We used a method to determine if human alveolar macrophages (HAM) could be selectively activated to release products of cyclooxygenase or lipoxygenase pathway of arachidonic acid. HAM obtained by bronchoalveolar lavage from individuals were [3H]arachidonic acid labeled and then stimulated with lipopolysaccharide (LPS) or Ca ionophore A23187. Essentially no arachidonate metabolites were released by unstimulated cells. LPS caused dose- and time-dependent release of arachidonate and only cyclooxygenase products; no lipoxygenase products were detected, even in presence of cyclooxygenase inhibition. Metabolites released in response to LPS included thromboxane B2, prostaglandins D2, F2a, E2, and hydroxyheptadecatrienoic acid. A23187 caused a rapid release of arachidonate and 5-lipoxygenase products, leukotriene B4 and 5-hydroxyeicosatetraenoic acid; no cyclooxygenase inhibition. This demonstrates that HAM are specifically activated to release metabolites derived from cyclooxygenase or lipoxygenase pathway of arachidonic acid. Additionally, shunting down an alternate pathway is not induced by use of inhibitors of either pathway. This suggests alveolar macrophages may enhance or suppress various inflammatory or immune processes in lung, in part, by selective release of various derivatives of arachidonic acid.

scholarly journals Leukotriene generation by eosinophils.

1982 ◽  
Vol 155 (2) ◽  
pp. 390-402 ◽  
Author(s):  
A Jörg ◽  
W R Henderson ◽  
R C Murphy ◽  
S J Klebanoff
Keyword(s):  
Arachidonic Acid ◽  
Calcium Ionophore ◽  
Leukotriene B4 ◽  
Arachidonic Acid Metabolism ◽  
Calcium Ionophore A23187 ◽  
Eicosatetraenoic Acid ◽  
Gas Chromatography Mass Spectrometry ◽  
Ionophore A23187 ◽  
Lipoxygenase Pathway ◽  
Leukotriene D4

Horse eosinophils purified to greater than 98% generated slow reacting substance (SRS) when incubated with the calcium ionophore A23187. On a per cell basis, eosinophils generated four to five times the SRS produced by similarly treated horse neutrophils. Eosinophil SRS production was inhibited by 5,8,11,14-eicosatetraynoic acid and augmented by indomethacin and arachidonic acid, suggesting that it was a product(s) of the lipoxygenase pathway of arachidonic acid metabolism. Compounds with SRS activity were purified by high-pressure liquid chromatography (HPLC) and identified by ultraviolet spectra, spectral shift on treatment with lipoxygenase, incorporation of [14C]arachidonic acid, gas chromatography-mass spectrometry, and comparison of retention times on HPLC to authentic standards. The eosinophil products characterized were 5-(S), 12-(R)-dihydroxy-6-cis-8, 10-trans-14-cis-eicosatetraenoic acid (leukotriene B4) and its 5-(S), 12-(R)-6-trans and 5-(S), 12-(S)-6-trans isomers, 5-(S)-hydroxy-6-(R)-S-glutathionyl-7,9-trans-11, 14-cis-eicosatetraenoic acid (leukotriene C4) and its 11-trans isomer, and 5-(S)-hydroxy-6-(R)-S-cysteinylglycine-7,9-trans-11,14-cis-eicosatetraenoic acid (leukotriene D4).

scholarly journals The suppression of 5-lipoxygenation of arachidonic acid in human polymorphonuclear leucocytes by the 15-lipoxygenase product (15S)-hydroxy-(5Z,8Z,11Z,13E)-eicosatetraenoic acid: structure-activity relationship and mechanism of action

1996 ◽  
Vol 314 (3) ◽  
pp. 911-916 ◽  
Author(s):  
Karen PETRICH ◽  
Peter LUDWIG ◽  
Hartmut KÜHN ◽  
Tankred SCHEWE
Keyword(s):  
Fatty Acids ◽  
Arachidonic Acid ◽  
Leukotriene B4 ◽  
Eicosatetraenoic Acid ◽  
Ionophore A23187 ◽  
Polymorphonuclear Leucocytes ◽  
Lipoxygenase Inhibitor ◽  
Acid Structure ◽  
Selection Of

(15S)-Hydroxy-(5Z,8Z,11Z,13E)-eicosatetraenoic acid (15-HETE) suppresses in ionophore-A23187-stimulated human polymorphonuclear leucocytes (PMN) the conversion of exogenous arachidonic acid into leukotriene B4 (LTB4) and (5S)-hydroxy-(6E,8Z,11Z,14Z)-eicosatetraenoic acid (5-HETE). However, contrary to earlier suggestions, 15-HETE is not a genuine 5-lipoxygenase inhibitor under these conditions, but rather suppresses the 5-lipoxygenation of arachidonic acid by switching-over of substrate utilization, as judged from a sizeable formation of labelled (5S,15S)-dihydroxy-(6E,8Z,11Z,13E)-eicosatetraenoic acid (5,15-diHETE) from 15-[1-14C]HETE. Identical results were obtained with human recombinant 5-lipoxygenase. In PMN the formation of 5,15-diHETE is strongly stimulated by either hydroperoxypolyenoic fatty acids or arachidonic acid, suggesting a crucial role of the hydroperoxide tone of the cell. A comparison of a selection of hydroxypolyenoic fatty acids with respect to their capability of suppressing 5-lipoxygenation of arachidonic acid revealed that 15-monohydroxyeicosanoids throughout exhibit the highest inhibitory potencies, whereas the other HETEs, 5,15-diHETE as well as octadecanoids, are modest or poor inhibitors. The R and S enantiomers of 15-HETE do not differ from each other, excluding a receptor-like binding of the 15-hydroxy group.

Polarity of arachidonic acid metabolism by bovine aortic endothelial cell monolayers

1987 ◽  
Vol 253 (5) ◽  
pp. H1177-H1183
Author(s):  
D. M. Shasby ◽  
L. L. Stoll ◽  
A. A. Spector
Keyword(s):  
Arachidonic Acid ◽  
Arachidonic Acid Metabolism ◽  
Biologically Active ◽  
Bovine Aortic Endothelial Cell ◽  
Ionophore A23187 ◽  
Vascular Events ◽  
Aortic Endothelial Cells ◽  
Cell Monolayers ◽  
Arachidonate Metabolites ◽  
Aortic Endothelial

Monolayers of bovine aortic endothelial cells cultured on micropore filters were used to determine the polarity of endothelial uptake, release, and transfer of arachidonic acid and some of its metabolites. Uptake and spontaneous release of arachidonic acid were more rapid at the luminal than at the interstitial surface. Transfer of arachidonic acid was more rapid from the luminal to the interstitial compartment than from the interstitial to the luminal compartment. After stimulation with the ionophore A23187, monolayers released arachidonate metabolites, including prostacyclin, to both the luminal and the interstitial compartments. The ability of the endothelium to rapidly take up and release arachidonic acid from the luminal surface and the ability to release biologically active eicosanoids to both the lumen and interstitium could be important for endothelial modulation of vascular events.

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