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Biomedical subjects

M Lagarde

Publications and source records attributed to M Lagarde.

At least 217 records · Page 12Linked to original sources
PubMed1986

Studies on polyenoic acid incorporation into human platelet lipid stores: interactions with linoleic and arachidonic acids.

Several polyunsaturated fatty acids (C18-C22 acids) have been compared in their uptake by human platelets and their acylation into glycerophospholipid subclasses. This was also studied in the presence of linoleic and/or arachidonic acids, the main fatty acids of plasma free fatty acid pool. Amongst C20 fatty acids, dihomogamma linolenic acid (20:3(n-6)), 5,8,11-icosatrienoic acid (20:3(n-9)) and arachidonic acid (20:4(n-6)) were better incorporated. The uptake of 5,8,11,14,17-icosapentaenoic acid (20:5(n-3)) was significantly lower and comparable to that of C22 fatty acids (7,10,13,16-docosatetraenoic acid (22:4(n-6)) and 4,7,10,13,16,19-docosahexaenoic acid (22:6(n-3)) and linoleic acid (18:2(n-6)). In this respect, linolenic acid (18:3(n-3)) appeared the poorest substrate. The bulk of each acid was acylated into glycerophospholipids although the presence of linoleic and/or arachidonic acids diverted a part towards neutral lipids. This was prominent for 18:3(n-3) and C22 fatty acids. The glycerophospholipid distribution of each acid differed substantially and was not affected by the presence of linoleic and or arachidonic acids, except for 18:3(n-3) and 22:6(n-3) that were strongly diverted towards phosphatidylethanolamine (PE) at the expense of phosphatidylcholine (PC). The main features were an efficient acylation of 20:3(n-9) into phosphatidylinositol (PI) followed by 20:3(n-6) and 20:4(n-6), then by 20:5(n-3) and 22:4(n-6), and finally 22:6(n-3) and C18 fatty acids. This was reciprocal to the acylation into PE and to a lesser extent into PC which remained the main storage species in all cases. We conclude that human platelets may exhibit a certain specificity for taking up polyunsaturated fatty acids both in terms of total uptake and glycerophospholipid subclass distribution. Also the presence of polyunsaturated fatty acids of normal plasma, like linoleic and arachidonic acids, may interact specifically with such an uptake and distribution.

Arachidonic Acids↗
PubMed1986

Effects of linoleic acid and gamma-linolenic acid intake on platelet functions in elderly people.

Sixteen old subjects were given daily dietary supplement of lg of linoleic acid and lg of gamma-linolenic acid (primerose oil) or 2g of linoleic acid (sunflower oil) for periods of two months. Haemostatic parameters, platelet aggregation, exogenous and endogenous arachidonic acid metabolism were investigated before and after the intake. Diets did not induce any significant change in haemostatic parameters (bleeding time, levels of anti-thrombin III, plasminogen and plasma beta-TG and PF4). Platelet rich plasma aggregation induced by collagen and arachidonic acid were significantly reduced after linoleic acid (18:2n-6) intake. In contrast, gamma-linolenic acid (18:3n-6) supplement did not alter aggregation. However, thromboxane B2 formation (under stimulation) and vitamin E level in platelets (but not in plasma) were decreased after 18:3n-6 as compared to 18:2n-6 intake. The mechanism of thromboxane B2 decrease is unclear. Nevertheless, we may speculate that beneficial effect of this decrease could be counterbalanced by the decreased platelet vitamin E. We conclude that intake of 18:2n-6 or 18:3n-6 does not affect much platelet functions in elderly people.

8,11,14-Eicosatrienoic Acid↗
PubMed1986

Arachidonic acid strongly stimulates prostaglandin I3 (PGI3) production from eicosapentaenoic acid in human endothelial cells.

Eicosapentaenoic acid (EPA) is a prominent polyunsaturated fatty acid in fish oil which inhibits blood platelet aggregation and thromboxane A2 formation but not prostacyclin-like material generation from vascular endothelium. In this study we investigated interaction between EPA and arachidonic acid (AA) during their oxygenation by cultured endothelial cells. As measured by gas chromatography-mass spectrometry (GC-MS), AA increased markedly prostaglandin I3 (PGI3) production from EPA while that of PGI2 from AA was decreased by EPA. However, increasing the ratio AA/EPA over one almost suppressed the inhibition of PGI2 formation by EPA, and the stimulation of PGI3 production by AA was even higher. The effect of AA on EPA conversion to minor prostaglandins like PGE3 and PGF3 alpha was similar then confirming the stimulating effect and suggesting it is occurring at the cyclooxygenase instead of the prostacyclin synthase level. Altogether these data indicate that, in certain nutritional states where the liberation of EPA from endothelial cells will be accompanied with that of endogenous AA, substantial amounts of PGI3 could contribute to the prostacyclin-like activity of the vessel wall in addition to PGI2.

Arachidonic Acid↗
PubMed1985

13-Hydroxyoctadecadienoic acid is the vessel wall chemorepellant factor, LOX.

We have previously reported that endothelial cells synthesize a cytosol-associated, lipoxygenase-derived metabolite, LOX, which acts as a chemorepellant and, in so doing, maintains the vessel wall thromboresistance. In this study we demonstrate that LOX is a 13-hydroxylinoleic acid (13-OH-18:2) derived from linoleic acid and identical to 13-hydroxy-9-cis,11-trans-octadecadienoic acid, as measured by both reverse phase high pressure liquid chromatography and gas chromatography/mass spectrometry. In addition, we demonstrate that 13-OH-18:2 is produced in significantly greater quantities by endothelial cells than by smooth muscle cels or by fibroblasts. Furthermore, we demonstrate that 13-OH-18:2 is produced in microgram amounts under basal conditions and is decreased by thrombin, calcium ionophore, and trypsin stimulation. And finally, we demonstrate that endothelial cells do not synthesize any significant amounts of lipoxygenase-derived arachidonic acid metabolites either under basal or stimulated conditions unless exogenous arachidonic acid is added. These observations indicate that the major lipoxygenase-derived, chemorepellant metabolite produced by the endothelial is 13-hydroxy-9-cis,11-trans-octadecadienoic acid.

Antithrombins↗
PubMed1985

Studies on platelet lipoxygenase specificity towards icosapolyenoic and docosapolyenoic acids.

Two docosapolyenoic acids (22:5(n-3) and 22:5(n-6)) were isolated from the liver of normal and 18:3(n-3)-deficient trout, respectively. They were prepared by combined thin-layer chromatography (TLC) and reversed-phase high performance liquid chromatography (HPLC). Their purity, checked by capillary gas liquid chromatography, was greater than 95%. Each fatty acid was oxygenated into monohydroxy derivatives by human platelets. The hydroxy compounds were purified by TLC and HPLC and then derivatized for gas chromatography-mass spectrometry analysis. Whereas 22:5(n-6) was only converted into 14-OH-22:5, three hydroxy derivatives (11, 13 and 14) were obtained from 22:5(n-3). However, 13-hydroxy was not formed in the presence of aspirin, indicating that platelet lipoxygenase catalyses the formation of both 11- and 14-hydroxy derivatives from 22:5(n-3), as described previously, from 22:6(n-3). Further studies showed that 22:4(n-6) and 20:5(n-3) were only converted into 14- and 12-hydroxy derivatives. We conclude then that, besides the well-known n-9 oxygenation, lipoxygenase of human platelets is able to catalyse an n-12 oxygenation on docosapolyenoic acids of the n-3 family.

Blood Platelets↗
PubMed1985

Differential effect of external calcium on the oxygenated metabolism of endogenous and exogenous arachidonic acid in platelets.

The oxygenation of arachidonic acid into thromboxane B2 (TXB2), 12-hydroxy-heptadecatrienoic (HHT) and 12-hydroxy-eicosatetraenoic (12-HETE) acids has been examined in human platelets in the absence or presence of 1mM calcium. From endogenous arachidonic acid, external calcium did not affect the formation of cyclo-oxygenase products (TXB2 and HHT) but enhanced that of 12-HETE when thrombin at high concentrations was the agonist. Dose-response curves performed with thrombin and collagen revealed that increased stimulation resulted in higher ratios of 12-HETE/HHT. On the other hand external calcium did not alter significantly the synthesis of either products from exogenous arachidonic acid and the total conversion of the substrate was unchanged. We conclude that extracellular calcium may facilitate the liberation of arachidonic acid from platelet phospholipids when induced by high thrombin concentrations. The excess of arachidonic acid liberated would then be diverted towards the lipoxygenase pathway.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1985

Incorporation of some eicosanoic acids into endothelial cells--effect on platelet inhibitory activity and prostacyclin production.

Primary cultures of endothelial cells from human umbilical veins were grown until confluency. Then, dihomogammalinolenic acid (DHLA or 20:3n-6) and eicosapentaenoic acid (EPA or 20:5n-3), precursors of monoenoic and trienoic prostanoids, respectively, as well as 5,8,11-eicosatrienoic acid (20:3n-9), and isomer of DHLA, were incorporated into endothelial lipids. DHLA-rich endothelial cells had a decreased capacity of prostacyclin production. By contrast EPA- or 20:3n-9-rich endothelial cells were comparable to controls in this respect. DHLA and EPA were efficiently acylated into cell phospholipids and triglycerides at the opposite of 20:3n-9. It is suggested that both DHLA and EPA could alter the liberation of endogenous arachidonic acid for prostacyclin synthesis but this might be counterbalanced in EPA-rich endothelial cells by PGI3 production. We conclude that DHLA enrichment of endothelial cell lipids may impair the possible beneficial effect of the acid upon platelet functions whereas that of EPA would not be modified.

8,11,14-Eicosatrienoic Acid↗
PubMed1985

Prostaglandin E2-like activity of 20:3n-9 platelet lipoxygenase end-product.

5,8,11-Icosatrienoic acid (20:3n-9), a fatty acid associated with platelet hyperactivity, was oxygenated by platelet lipoxygenase. The end-product of this pathway was purified by high-performance liquid chromatography (HPLC) and characterized as 12-hydroxy-5,8,10-icosatrienoic acid [12-OH-20:3(5,8,10)] by capillary gas-liquid mass spectrometry. When tested upon platelet aggregation, 12-OH-20:3(5,8,10) exhibited a biphasic effect. At low concentrations (below 5 X 10(-7) M) it potentiated aggregation but inhibited it at higher levels, a pattern similar to that obtained with prostaglandin E2. However, since the amounts of 12-OH-20:3(5,8,10) generated under thrombin stimulation are in the range of concentrations with potentiating effects, it seems that the 12-OH derivative is responsible for the hyperaggrebility of 20:3n-9-rich platelets.

8,11,14-Eicosatrienoic Acid↗
PubMed1985

Diacylglycerol lipase activity in human platelet intracellular and surface membranes. Some kinetic properties and fatty acid specificity.

Diacyl glycerol lipase activity has been examined of intracellular and surface membranes isolated from human blood platelets by free flow electrophoresis. Enzyme activity is present on both membranes but is activated at different substrate concentrations (Km 14 microM and 140 microM for intracellular and surface membrane, respectively). Both enzyme activities are stimulated by EGTA and GSH, and inhibited by added Ca2+. The specificity of the intracellular membrane enzyme has been investigated using a range of diacylglycerol substrates differing only in their '2' position fatty acid. Arachidonic acid is clearly the preferred '2' position moiety with activities towards eicosatrienoic, linoleic, oleic and palmitic acid-containing substrates, all substantially lower.

Blood Platelets↗
PubMed1985

In vitro incorporation and metabolism of some icosaenoic acids in platelets. Effect on arachidonic acid oxygenation.

Three icosaenoic acids (20:3(n-6), 20:5(n-3) and 20:3(n-9)) which may arise in platelet phospholipids under certain dietary conditions and which may affect platelet functions have been taken up by human platelets. Each acid was pre-coated onto delipidated albumin and then incubated with platelets isolated from their plasma. The distribution study of each acid in cellular lipids revealed that around 80% of the acid taken up was located in phospholipids, of which the bulk was in phosphatidylcholine. The percentage incorporation of each acid into the different glycerophospholipids was similar to their endogenous percentage profiles, therefore simulating the in vivo situation. The icosaenoic acids then incorporated were liberated from phospholipids when platelets were incubated with thrombin or calcium ionophore A23187 and subsequently oxygenated through the cyclooxygenase and/or lipoxygenase pathway. Whereas 20:3(n-6) was readily converted into cyclooxygenase products, 20:5(n-3) was more specifically converted into lipoxygenase products, and this latter conversion was comparable to that of 20:3(n-9) which is not a prostanoid precursor. Finally, only 20:3(n-6)- or 20:5(n-3)-rich platelets exhibited a reduced availability of endogenous arachidonic acid from phospholipids when induced by thrombin. It is concluded that inhibitory polyunsaturated fatty acids (20:3(n-6) and 20:5(n-3)) could act both by reducing prostaglandin H2/thromboxane A2 production from endogenous arachidonic acid and in generating platelet inhibitory substances (cyclooxygenase and/or lipoxygenase products of 20:3(n-6) and 20:5(n-3)). On the other hand, 20:3(n-9), a fatty acid which potentiates platelet aggregation through its lipoxygenase end product, could produce sufficient amounts of this compound to enhance the aggregation when platelets are triggered with inducers of phospholipase activity such as thrombin or calcium ionophore.

Arachidonic Acid↗
PubMed1985

Enhancement of eicosaenoic acid lipoxygenation in human platelets by 12-hydroperoxy derivative of arachidonic acid.

Human platelet lipoxygenase activity toward several eicosaenoic acids was measured in intact cells as well as in subcellular fractions (cytosol and membranes). In whole platelets, the lipoxygenation of eicosaenoic acids was enhanced greatly by high concentration of aspirin, which partially inhibit the peroxidase activity associated with the pathway. The lipoxygenation also was increased by arachidonic acid (AA) or its lipoxygenase product, 12-hydroxyperoxy-eicosatetraenoic acid (12-HPETE). Similarly, prostanoid precursors, dihomogammalinolenic (DHLA) and eicosapentaenoic (EPA) acids also were better converted by cyclooxygenase in the presence of AA or 12-HPETE. Among the eicosaenoic acids tested, EPA oxygenation was affected most. Using cytosol or membranes as the lipoxygenase source instead of whole cells led to completely different results. AA exerted a competitive inhibition upon the other eicosaenoic acid oxygenation except that of EPA, for which a dual effect of AA was observed. This makes questionable the use of platelet subfractions for investigating lipoxygenase activity. We conclude that platelet lipoxygenation of eicosaenoic acids appears peroxide-dependent, especially for apparent poor substrates like EPA. This might be relevant in respect to 12-HPETE, which is the main hydroperoxy derivative to be produced during platelet activation.

8,11,14-Eicosatrienoic Acid↗
PubMed1985

[Polyunsaturated fatty acids, atherosclerosis and thrombosis].

This article is a short update of the role played by dietary polyunsaturated fatty acids in atherosclerosis and thrombosis, with special reference to the interactions between the platelets and endothelium. The n-3 family of fatty acids, especially eicosapentaenoic (20:5n-3) and docosahexaenoic acids (22:6n-3), seem to be beneficial in the prevention of these diseases.

Animals↗
PubMed1984

Inhibition of platelet aggregation and thromboxane synthesis after intake of small amount of icosapentaenoic acid.

Elderly people ingested 150 mg/day of icosapentaenoic acid (20:5n-3) or a placebo for one month. Platelet aggregation, platelet arachidonate metabolism and the fatty acid composition of both plasma and platelet lipids were investigated before and after the intake. Platelet aggregation induced by collagen, epinephrine or low concentrations of ADP was significantly reduced after 20:5n-3 intake. Besides, the main oxygenated product formation from endogenous platelet arachidonate under thrombin stimulation was markedly decreased after the 20:5n-3 supplementation. Such a decrease was absent after placebo. Moreover, no modification in the fatty acid composition of both plasma lipids and platelet phosphatidylcholine could be observed. We conclude that intake of low amounts of 20:5n-3 by elderly people, is able to lower their platelet sensitivity to aggregating agents, probably by decreasing the endogenous formation of platelet thromboxane A2, although no modification in the fatty acid composition was detected.

Adenosine Diphosphate↗
PubMed1984

Subcellular localization and some properties of lipoxygenase activity in human blood platelets.

Lipoxygenase activity was measured in human platelet subcellular fractions. From a sonicated platelet preparation, a granule fraction, mixed membranes (surface and intracellular) and cytosol fractions were separated by differential centrifugation. With respect to activities in the sonicated preparation, the lipoxygenase was slightly enriched in both the cytosol and mixed-membrane fractions and consistently de-enriched in the granule fractions. Approx. 65% and 20% of the total cell enzyme activity were found in the cytosol and mixed membranes respectively, with only 8% present in the granule fraction. Additionally we measured the lipoxygenase activity in purified surface- and intracellular-membrane subfractions prepared from the mixed membranes by free-flow electrophoresis. There was a slight enrichment in activity in the intracellular membrane fraction compared with that in the mixed membranes, and a depletion of activity in the surface membranes. Characterization of the enzyme activity, i.e. time course, pH-dependence, Ca2+-dependence, Vmax. and Km for arachidonic acid, and the carbon-position specificity for this acid, failed to reveal any significant differences between the membrane-bound and soluble forms of the lipoxygenase. These findings suggest that in human platelets the same lipoxygenase is associated with the membranes as in the cytosol and that the membrane-bound activity predominates in intracellular membrane elements.

Arachidonate Lipoxygenases↗
PubMed1984

Relationship between cyclic AMP and thromboxane formation in platelet-endothelial cell interactions.

Human platelet aggregation was triggered in the presence of various numbers of cultured endothelial cells. Thromboxane B2 formation and platelet cyclic AMP were also measured. Using a low concentration of thrombin (0.025U/ml) as aggregating agent, the inhibition of platelet aggregation correlated with that of thromboxane formation and was directly related to both the number of endothelial cells and platelet cyclic AMP. In contrast, using arachidonic acid (10(-5)M) instead of thrombin, platelet aggregation could be abolished although thromboxane formation was not affected. These results suggest that platelet aggregation induced by low concentrations of thrombin might be dependent on prostaglandin endoperoxides/thromboxane A2 production which could be inhibited by cyclic AMP. The normal synthesis of thromboxane B2 from exogenous arachidonate indicates that cyclic AMP is only active upon the liberation of endogenous arachidonate from platelet phospholipids.

6-Ketoprostaglandin F1 alpha↗