scholarly journals Synthesis of leukotriene C and other arachidonic acid metabolites by mouse pulmonary macrophages.

1982 ◽  
Vol 155 (3) ◽  
pp. 720-733 ◽  
Author(s):  
C A Rouzer ◽  
W A Scott ◽  
A L Hamill ◽  
Z A Cohn
Keyword(s):  
Arachidonic Acid ◽  
Prostaglandin E2 ◽  
Specific Activity ◽  
Thromboxane A2 ◽  
Fatty Acid Analysis ◽  
Cell Protein ◽  
Experimental Conditions ◽  
Pulmonary Macrophages ◽  
Hydroxyeicosatetraenoic Acids

Mouse resident pulmonary macrophages were subdivided into alveolar (PAM) and interstitial (PTM) populations on the basis of accessibility to pulmonary lavage, and zymosan-induced arachidonic acid (20:4) metabolism was examined in both populations labeled with [3H]20:4. Maximal phagocytic doses of unopsonized zymosan induced the specific release of 11% of phospholipid 20:4 by PTM and 4.6% by PAM. Direct fatty acid analysis of [3H]20:4-labeled PTM cultured in the presence or absence of zymosan indicated that the specific activity of the [3H]20:4 in cell phospholipid provided an accurate measure of 20:4 released by the cells, and could therefore be used to quantitate the synthesis of 20:4 metabolites by PTM in vitro. The single major 20:4 metabolite of PTM was the slow-reacting substance leukotriene C, which was synthesized in quantities of 3-4 pmol/microgram cell protein (280-370 pmol/10(6) cells), and comprised 20-25% of the released 20:4. PTM also synthesized prostaglandin E2, prostacyclin, thromboxane A2, and hydroxyeicosatetraenoic acids. In contrast, PAM produced leukotrienes D and E in addition to leukotriene C, prostaglandin E2, thromboxane A2, and hydroxyeicosatetraenoic acids. Prostacyclin formation by PAM was not observed. These studies define a set of experimental conditions for the study of 20:4 metabolism by pulmonary macrophages, and demonstrate that these cells are rich sources of LTC as well as other 20:4 oxygenated products.

In Vitro Incorporation and Metabolism of Icosapentaenoic and Docosahexaenoic Acids in Human Platelets - Effect on Aggregation

1986 ◽  
Vol 56 (01) ◽  
pp. 057-062 ◽  
Author(s):  
Martine Croset ◽  
M Lagarde
Keyword(s):  
Arachidonic Acid ◽  
Fatty Acid ◽  
Platelet Aggregation ◽  
Fatty Acid Distribution ◽  
Thromboxane A2 ◽  
Human Platelets ◽  
Aggregation Response ◽  
Docosahexaenoic Acids ◽  
Membrane Level

SummaryWashed human platelets were pre-loaded with icosapentaenoic acid (EPA), docosahexaenoic acid (DHA) or EPA + DHA and tested for their aggregation response in comparison with control platelets. In fatty acid-rich platelets, an inhibition of the aggregation could be observed when induced by thrombin, collagen or U-46619. The strongest inhibition was observed with DHA-rich platelets and it was reduced when DHA was incorporated in the presence of EPA.Study of fatty acid distribution in cell lipids after loading showed that around 90% of EPA or DHA taken up was acylated into phospholipids and a very small amount (less than 2%) remained in their free and hydroxylated forms. DHA was more efficiently acylated into phosphatidylethanolamine (PE) than into phosphatidylinositol (PI) in contrast to what observed with EPA, and both acids were preferentially incorporated into phosphatidylcholine (PC). EPA inhibited total incorporation of DHA and increased its relative acylation into PE at the expense of PC. In contrast, DHA did not affect the acylation of EPA. Upon stimulation with, thrombin, EPA was liberated from phospholipids and oxygenated (as judged by the formation of its monohydroxy derivative) whereas DHA was much less metabolized, although consistently transferred into PE.It is concluded that EPA and DHA might affect platelet aggregation via different mechanisms when pre-loaded in phospholipids. Whereas EPA is known to alter thromboxane A2 metabolism from endogenous arachidonic acid, by competing with it, DHA might act directly at the membrane level for inhibiting aggregation.

scholarly journals Dynamics of leukotriene C production by macrophages.

1980 ◽  
Vol 152 (5) ◽  
pp. 1236-1247 ◽  
Author(s):  
C A Rouzer ◽  
W A Scott ◽  
A L Hamill ◽  
Z A Cohn
Keyword(s):  
Time Course ◽  
Culture Medium ◽  
Peritoneal Macrophages ◽  
Experimental Conditions ◽  
Silicic Acid Chromatography ◽  
Pg Release ◽  
Antigen Antibody ◽  
Mouse Peritoneal Macrophages ◽  
Hydroxyeicosatetraenoic Acids

A method for the radiochemical assay of LTC production by mouse peritoneal macrophages in vitro is presented. The method involves labeling macrophages in culture with [5,6,8,9,11,12,14,15-3H]20:4 followed by stimulation of arachidonic acid (20:4) release under the experimental conditions desired. Radiolabeled leukotriene C (LTC) is recovered from the culture medium by extraction and silicic acid chromatography in 40% yield with full retention of biological activity. Because this LTC is radiochemically pure, the quantity of LTC release may be estimated from the amount of radioactivity in the sample. Use of the radioassay to study parameters affecting LTC synthesis by macrophages indicated that the time course of LTC synthesis and its relationship to the dose of a phagocytic stimulus (zymosan) were very similar to those of prostaglandin (PG) release. LTC release was also similar to that of PG in that lower levels of both metabolites were produced by Corynebacterium parvum-elicited macrophages than by resident cells. Finally, LTC release was stimulated in response to a challenge with antigen-antibody complexes, but lower maximal levels were attained than those with zymosan. The data presented here are consistent with the hypothesis that challenge of macrophages with a phagocytic stimulus leads to the release of 20:4 by an inducible phospholipase. Cyclooxygenase and lipoxygenase then compete for the released 20:4, leading to the production of PG, hydroxyeicosatetraenoic acids, and LTC.

scholarly journals Endothelial-secreted arachidonic acid metabolites modulate polymorphonuclear leukocyte chemotaxis and diapedesis in vitro

Blood ◽  
1988 ◽  
Vol 71 (3) ◽  
pp. 771-779 ◽  
Author(s):  
J Doukas ◽  
HB Hechtman ◽  
D Shepro
Keyword(s):  
Arachidonic Acid ◽  
Polymorphonuclear Leukocyte ◽  
Thromboxane A2 ◽  
In Vitro Assays ◽  
General Effect ◽  
Random Motility ◽  
Acid Metabolites ◽  
Leukocyte Chemotaxis ◽  
High Doses

Abstract The influence of endothelial cells (ECs) on polymorphonuclear leukocyte (PMN) motility was examined using in vitro assays of PMN diapedesis and chemotaxis. ECs are seen to release arachidonic acid (20:4) metabolites that directly increase or decrease PMN movement, with their general effect being enhanced motility. This effect can be increased or decreased by treating ECs with stimulators or inhibitors of 20:4 metabolism, respectively. The metabolites include thromboxane B2, which increases PMN random motility, chemotaxis, and diapedesis in a dose- responsive manner and which acts as a chemoattractant; 6-keto-PGF1 alpha, which increases chemotaxis and diapedesis at high doses but decreases these responses at low doses; and a lipoxygenase-derived metabolite, suggested to be 5-hydroxyeicosatetraenoic acid, which increases chemotaxis and diapedesis. Thromboxane A2 and prostacyclin mimetics also affect chemotaxis in qualitatively similar manners as TxB2 and 6-keto-PGF1 alpha, respectively, but display greater potency. EC release of these metabolites is also seen to be substratum modulated, with an increased production by cells cultured on extracellular matrices. These results suggest that ECs are capable of modulating PMN motility and suggest a role for ECs in the control of PMN diapedesis.

scholarly journals Activation of macrophages for enhanced release of superoxide anion and greater killing of Candida albicans by injection of muramyl dipeptide.

1980 ◽  
Vol 152 (6) ◽  
pp. 1659-1669 ◽  
Author(s):  
N P Cummings ◽  
M J Pabst ◽  
R B Johnston
Keyword(s):  
Candida Albicans ◽  
Superoxide Anion ◽  
Specific Activity ◽  
Muramyl Dipeptide ◽  
Peritoneal Macrophages ◽  
Cell Protein ◽  
Bacillus Calmette Guerin ◽  
Myristate Acetate ◽  
Recipient Animal

The adjuvant muramyl dipeptide (MDP) has been shown to affect a number of macrophage functions in vitro. We studied the effect of subcutaneous injection of MDP into mice. Cultured peritoneal macrophages from treated mice displayed increased spreading, total cell protein, and specific activity of beta-glucosaminidase a constituent of macrophage lysosomes, and of lactate dehydrogenase. Generation of superoxide anion (O2-) by MDP-treated macrophages stimulated by contact with phorbol myristate acetate was enhanced by over fivefold to levels achieved by macrophages from bacillus Calmette-Guérin-infected mice. The enhancement in stimulated O2- release was noted by 1 h after injection of MDP, peaked by 3 h, and remained high for at least 48 h. Priming for enhancement of O2- release by MDP was similar in athymic nude mice and in normal littermates, suggesting that mature T lymphocytes are not involved in this MDP effect. Priming for enhanced stimulated O2- release, and morphologic and enzymic changes, were not achieved by injection of the D-D stereoisomer of MDP. Phagocytosis of Candida albicans was only slightly greater by macrophages from mice give MDP, but MDP-stimulated cells killed two times more C. albicans in vitro than did cells from untreated animals. When MDP was given 18 h before, simultaneously with, or 24 h after lethal infectious challenge with C. albicans, treated mice were protected compared with controls. These results suggest that injection of MDP effectively and rapidly activates macrophages in the recipient animal. This agent should serve as an important probe of macrophage physiology and, perhaps ultimately, as a means of enhancing host defense in humans.

scholarly journals Phospholipase A activity in the skin. Modulators of arachidonic acid release from phosphatidylcholine

1979 ◽  
Vol 184 (2) ◽  
pp. 283-290 ◽  
Author(s):  
V A Ziboh ◽  
J T Lord
Keyword(s):  
Arachidonic Acid ◽  
Prostaglandin E2 ◽  
Specific Activity ◽  
Feedback Regulation ◽  
Phospholipase A ◽  
Particulate Fraction ◽  
Prostaglandin F2 Alpha ◽  
Low Concentrations ◽  
Freeze Dried ◽  
Hydrolysis Of

The distribution of the hydrolysis of 1-acyl-2-[1-14C]arachidonoyl-sn-glycero-3-phosphocholine and the simultaneous biosynthesis of prostaglandins by subcellular fractions from human and rat skin membrane preparations were determined. The phospholipase A2 activity was distributed among the subcellular particulate preparations with the highest specific activity in the 105000g particulate fraction. The activity was optimal at pH 7.5 in the presence of 1.0 mM-CaCl2 and was inhibited by EDTA. The hydrolysis of phosphatidylcholine by the skin 105000g particulate fraction was inhibited by cortisol and triamcinolone acetonide and it was stimulated by histamine, bradykinin, retinoic acid and cholera enterotoxin (freeze-dried Vibrio cholerae). Furthermore hydrolysis of phosphatidylcholine by the skin phospholipase A was also enhanced by low concentrations of prostaglandin E2 and prostaglandin F2 alpha. These last results suggest that the amplication of the hydrolysis of phosphatidylcholine by prostaglandin E2 and prostaglandin F2 alpha, with the consequent release of arachidonic acid (the substrate of prostaglandin synthesis) is likely a positive-feedback regulation of the arachidonic acid-prostaglandin cascade.

scholarly journals Endothelial-secreted arachidonic acid metabolites modulate polymorphonuclear leukocyte chemotaxis and diapedesis in vitro

Blood ◽  
1988 ◽  
Vol 71 (3) ◽  
pp. 771-779
Author(s):  
J Doukas ◽  
HB Hechtman ◽  
D Shepro
Keyword(s):  
Arachidonic Acid ◽  
Polymorphonuclear Leukocyte ◽  
Thromboxane A2 ◽  
In Vitro Assays ◽  
General Effect ◽  
Random Motility ◽  
Acid Metabolites ◽  
Leukocyte Chemotaxis ◽  
High Doses

The influence of endothelial cells (ECs) on polymorphonuclear leukocyte (PMN) motility was examined using in vitro assays of PMN diapedesis and chemotaxis. ECs are seen to release arachidonic acid (20:4) metabolites that directly increase or decrease PMN movement, with their general effect being enhanced motility. This effect can be increased or decreased by treating ECs with stimulators or inhibitors of 20:4 metabolism, respectively. The metabolites include thromboxane B2, which increases PMN random motility, chemotaxis, and diapedesis in a dose- responsive manner and which acts as a chemoattractant; 6-keto-PGF1 alpha, which increases chemotaxis and diapedesis at high doses but decreases these responses at low doses; and a lipoxygenase-derived metabolite, suggested to be 5-hydroxyeicosatetraenoic acid, which increases chemotaxis and diapedesis. Thromboxane A2 and prostacyclin mimetics also affect chemotaxis in qualitatively similar manners as TxB2 and 6-keto-PGF1 alpha, respectively, but display greater potency. EC release of these metabolites is also seen to be substratum modulated, with an increased production by cells cultured on extracellular matrices. These results suggest that ECs are capable of modulating PMN motility and suggest a role for ECs in the control of PMN diapedesis.

Antivasoconstrictor and Antiaggregatory Activities of Picotamide Unrelated to Thromboxane A2 Antagonism

1997 ◽  
Vol 78 (05) ◽  
pp. 1385-1391 ◽  
Author(s):  
Roberta Vezza ◽  
Domenico Spina ◽  
Ronald J Tallarida ◽  
Malevika Nathan ◽  
Clive P Page ◽  
...  
Keyword(s):  
Receptor Antagonist ◽  
Shape Change ◽  
Thromboxane A2 ◽  
Inhibitory Effects ◽  
Experimental Conditions ◽  
Vasoactive Substances ◽  
Vasoconstrictor Effect ◽  
Synthase Inhibitor

SummaryPicotamide is a dual thromboxane (Tx) A2 receptor antagonist/Tx synthase inhibitor although some observations suggest an anti-vasoconstrictor effect independent of TxA2 inhibition/antagonism. The aim of our study was to assess whether picotamide antagonises vascular contractions induced by different vasoactive substances in vitro. Picotamide inhibited competitively the contraction of rabbit aortic rings induced by the TxA2 mimetic U46619 (pA2 = 3.59) but also the contractions induced by phenylephrine (pA2 = 3.93) and serotonin (5-HT) (pA2 = 5.81) although in a not competitive way. Picotamide did not inhibit potassium-induced contractions, thus excluding aspecific effects on vascular smooth muscle. Picotamide inhibited 5-HT-induced platelet aggregation in vitro with an IC50 (212 μM) similar to that found when other aggregating stimuli are used, but it did not affect shape change (IC50> 1 mM) suggesting that the effects of picotamide can not be ascribed to 5-HT2-receptor antagonism; in the same experimental conditions neither a Tx-receptor antagonist (BM13.177) nor a dual Tx-receptor antagonist/synthase inhibitor (ridogrel) affected 5-HT-induced platelet responses.Our studies demonstrate that picotamide exerts antivasoconstrictor and platelet inhibitory effects unrelated to TxA2 antagonism. This activity may contribute to the anti-thrombotic/anti-ischaemic effects of the drug in vivo.

Trace-Element Contamination in Biopsies of the Liver

1973 ◽  
Vol 19 (5) ◽  
pp. 472-475 ◽  
Author(s):  
J Versieck ◽  
A Speecke ◽  
J Hoste ◽  
F Barbier
Keyword(s):  
Trace Elements ◽  
Trace Element ◽  
Nuclear Reactor ◽  
Specific Activity ◽  
Trace Element Analysis ◽  
High Specific Activity ◽  
Experimental Conditions ◽  
Element Analysis ◽  
Liver Biopsies

Abstract Contamination by trace elements during collection of liver biopsies has been investigated qualitatively and quantitatively. Menghini needles for percutaneous liver biopsy and disposable surgical blades were irradiated in a nuclear reactor until their constituents had a high specific activity. Sampling of liver biopsies was simulated in vitro with use of the irradiated instruments. Under these experimental conditions, radioactivity found in the samples corresponds to contaminations as may be expected in clinical practice. The results demonstrate that needle biopsies are heavily contaminated by numerous trace elements and unsuitable for trace-element analysis. Liver biopsies taken during laparotomy, with a steel surgical blade, are markedly less contaminated but still not acceptable for determinations of some trace elements, especially chromium and nickel.

Effects of atrial natriuretic peptide on renal medullary prostaglandin E2 production

1989 ◽  
Vol 257 (3) ◽  
pp. F336-F340
Author(s):  
R. J. Bolterman ◽  
M. D. Bentley ◽  
S. M. Sandberg ◽  
M. J. Fiksen-Olsen ◽  
J. C. Romero
Keyword(s):  
Arachidonic Acid ◽  
Atrial Natriuretic Peptide ◽  
Prostaglandin E2 ◽  
Natriuretic Peptide ◽  
Inhibitory Effect ◽  
Pge2 Production ◽  
In Vitro Effects ◽  
Different Doses ◽  
Atrial Natriuretic

Like arachidonic acid (AA) and bradykinin (BK), the intrarenal administration of atrial natriuretic peptide (ANP) has been shown to increase the urinary excretion of prostaglandin E2 (PGE2). In the present study, the direct in vitro effects of ANP on PGE2 production were compared with those of AA and BK. Canine renal inner medullary slices were preincubated for 30 min and washed in aerated Krebs-Ringer buffer (37 degrees C). During the final incubation period, with the use of varied concentrations of AA, BK, or ANP in Krebs-Ringer buffer, samples were obtained at 0 and 30 min to be used for radioimmunoassay of PGE2. Although the rate of PGE2 production was significantly increased 11-fold with AA and threefold with BK, it was unaffected by four different doses of ANP (10(-5) to 10(-11) M). Furthermore, the production of PGE2 during basal and stimulated (BK or AA) conditions was significantly blocked by indomethacin but not by ANP. These results indicate that ANP had no direct stimulatory or inhibitory effect on the medullary production of PGE2.

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