Platelet Lipid Research Articles

Signs of platelet activation, but not lipid peroxidation, in fetal blood associated with functional and structural umbilicoplacental lesions in pregnancies complicated by impaired glucose metabolism

Objectives: To study the influence of platelet activation and lipid peroxidation in fetal blood on umbilical vascular prostanoid synthesis and placental morphology in diabetic pregnancy.Methods:The concentrations of thromboxane A2 (TxA2) and malondialdehyde (MDA) were determined in umbilical cord plasma in 21 women with diabetes mellitus/impaired glucose tolerance (DM/IGT)and ten healthy women. Segments from the umbilical artery and vein were incubated and prostacyclin (PGI2) and TxA2 metabolites were determined. Prostanoid synthesis was stimulatedwith calcium ionophore at a second incubation. Histological examination was carried out in samples from the umbilical cord, membranes and placental parenchyma. Non-parametric statistical analysis was used,with a two-tailed p < 0.05 considered statistically significant.Results: Cord plasma TxA2, but not MDA, was higher among DM/IGT women (p = 0.07). There were indicationsthat cord plasma TxA2, but not MDA, was positively correlated with vascular prostanoid synthesis and synthesis capacity. In the umbilical vein, both the basal and stimulated PGI2 productionand the stimulated TxA2 production were lower in the DM/IGT group. Ischemic placental lesions were associated with a high TxA2 and a low MDA concentration in cord plasma.Conclusions:Even in less severe forms of impaired glucose metabolism, disturbances in platelet activation significantly affect both biochemical and morphological vessel wall and tissue functions in the umbilicoplacentalunit. This could indicate an abnormal programming of fetal cell functions and designate cases at increased risk of developing cellular and organ damage.

Anti-platelet effects of different phenolic compounds from Yucca schidigera Roezl. Bark

Resveratrol (3,4',5-trihydroxystilbene) has been reported to have a variety of anti-inflammatory, anti-carcinogenic, anti-fungal and anti-platelet effects. It occurs naturally in different medicinal plants. Recently, resveratrol and other related phenolic compounds including trans -3,3',5,5'-tetrahydroxy-4'-methoxystilbene and yuccaols A and C were isolated from the bark of Yucca schidigera . The aim of the present study was to evaluate in vitro the effects of these compounds on platelet aggregation induced by thrombin and ADP. Pretreatment of platelets with resveratrol or other tested phenolics (1-25 w g/ml) slightly reduced platelet aggregation stimulated by 5 w M ADP ( P < 0.05) or 10 w M ADP ( P < 0.005). The comparison of the inhibitory effects of tested compound in thrombin-induced platelet aggregation revealed that phenolic showed even stronger antiplatelet actions than resveratrol. These compounds also had an inhibitory effect on the thrombin-induced enzymatic platelet lipid peroxidation determined as the level of thiobarbituric acid reactive substances.

Infrared spectroscopic study of diabetic platelets

In the present study, we use infrared difference spectroscopy to correlate spectroscopic changes with biochemical and physical profiles in diabetic platelets. The IR spectroscopic analysis confirms the increased glycosylation of diabetic platelets compared to that of control platelets. The protein composition and/or structure in diabetic platelets also changed. Furthermore, the overall content of membrane lipids in diabetic platelets increased compared to that in control platelets as did the lipid peroxidation while their membrane fluidity decreased. To investigate the chronic hyperglycemia effect on platelet membranes, we also incubated the platelets with a high concentration of glucose. Key changes observed in the glucose-incubated platelets were increased glycosylation of platelet membranes, increased lipid content and peroxidation, however, the membrane fluidity was not altered. Our results provide direct information on the overall changes in diabetic platelets, and suggest that IR spectroscopy may have a place in the characterization of platelets in diabetes.

Platelet lipid(s) bound to albumin increases endothelial electrical resistance: mimicked by LPA

The objectives were to determine whether the permeability-decreasing activity of platelet-conditioned medium (PCM) is associated with a lipid bound to albumin and whether lysophosphatidic acid (LPA) is present in the PCM. A decrease in permeability was assessed by an increase in electrical resistance across endothelial cell monolayers derived from bovine pulmonary arteries and microvessels. The Sephacryl S-200 fraction of PCM that contained albumin, the albumin immunoprecipitate from the PCM, and the methanol extract from the albumin immunoprecipitate all increased endothelial electrical resistance. Increased electrical resistance induced by PCM was not abolished by boiling and was mimicked by 1-oleoyl-LPA and 1-palmitoyl-LPA. Analysis of a methanol-chloroform extract of one sample of PCM by electrospray mass spectrometry revealed many fatty acids, ceramide, diacylglycerol, phosphatidic acid, and palmitoyl-LPA, but analysis of a second sample of PCM and the methanol extract of its albumin immunoprecipitate revealed no LPA, only lipids. These findings indicate that a bioactive lipid(s), possibly LPA, released from platelets and subsequently bound to albumin forms an active complex that decreases endothelial permeability.

Oxidative stress of platelets and thrombocytopenia in patients with vivax malaria

Oxidative stress and antioxidative capacity of platelets and the relationship with thrombocytopenia were determined in patients with vivax malaria and compared with those of healthy subjects. Whole blood thrombocyte count, platelet superoxide dismutase and glutathione peroxidase activities of patients with vivax malaria were lower and platelet lipid peroxidation levels were higher in patients than those of healthy subjects. There was an important negative correlation between whole blood thrombocyte count and platelet lipid peroxidation level. The antioxidative mechanisms of thrombocytes were insufficient in malaria patients and caused oxidative stress. The oxidative damage of thrombocytes might be important in the ethiopathogenesis of thrombocytopenia occurring in malaria.

Studies on pig blood platelet responses to peroxynitrite action

The action of peroxynitrite (ONOO - ) on pig blood platelet function was studied. Incubation of washed blood platelets with ONOO - (0.062-2 mM) caused strong, dose- and time-dependent inhibitory effects on adhesion of resting and thrombin-stimulated platelets to collagen and on platelet aggregation induced by thrombin. The platelet responses were measured 10 s after addition of peroxynitrite to platelets ('zero time') and after 60 min incubation of platelets with ONOO - . Exposure of blood platelets to peroxynitrite (60 min) brought about platelet lipid peroxidation assessed by the thiobarbituric acid method but did not cause any changes in blood platelet proteins as determined by SDS-polyacrylamide gel electrophoresis.

Lack of effect of fish oil supplementation on coagulation and transcapillary escape rate of albumin in insulin-dependent diabetic patients with diabetic nephropathy

Objective: We studied the effect of a diet supplementation with fish oil in insulin-dependent diabetic patients with nephropathy in order to evaluate whether abnormal transcapillary escape rate of albumin and procoagulant activity in these patients could be modified. Methods: A double-blind, randomized, controlled study was carried out at a tertiary referral centre. The subjects were 29 insulindependent diabetic patients with nephropathy. One year of fish oil supplementation (4.6 g n-3 fatty acids/day) was compared with placebo (olive oil). The main outcome measures were N-3 fatty acid proportions of platelet lipids, transcapillary escape rate of albumin, prothrombin fragment 1 + 2, thrombin-antithrombin complexes, markers of fibrinolysis, fibrinogen, factor VII antigen and activity, thrombomodulin, von Willebrand factor, platelet factor 4 and beta-thromboglobulin. These were measured every 6 months. Results: Neither transcapillary escape rate of albumin (7.4 (median) (5.0-9.8) (range) % vs. 7.0 (4.6-10.6) %) nor prothrombin fragment 1 + 2 (0.97 (0.72-2.40) nmol/L vs. 1.01 (0.59-3.11) nmol/L) changed after 12 months of fish oil supplementation. Conclusion: Increased transcapillary escape rate of albumin and activity could not be modified during diet supplementation with fish oil in insulin-dependent diabetic patients with nephropathy.

1H-NMR lipid profiles of human blood platelets; links with coronary artery disease

Blood platelets are closely involved in the early development of atherosclerosis and in the events that lead to thrombosis, both of which are dominating factors in coronary artery disease (CAD). The aim of the present study was to evaluate the platelet lipid profiles of patients suffering from CAD and explore the possibility of a link between platelet lipids and CAD, using high-resolution high-field proton nuclear magnetic resonance spectroscopy as the analytical tool. The total platelet lipid profiles of healthy volunteers were compared with those of patients presenting with chest pain requiring coronary angiography. Two lipid groups changed significantly: cholesterol increased by 16.5% and total diacylglycerophospholipids decreased by 15.7%. There was also a significant decrease of the ethanolamine-containing phospholipids, by 4.7%; the extent of unsaturation of the fatty acid chains, by 0.2, and increase of the linoleate content of the fatty acid chains, by 1.9%. Our results suggest that platelet lipid abnormalities occur in patients with CAD and these changes may predate the development of overt atherosclerosis.

Vitamin C suppresses the cisplatin toxicity on blood platelets.

The effects of vitamin C on the oxidative stress in blood platelets induced by cisplatin were studied. In the presence of vitamin C we measured in blood platelets the production of thiobarbituric acid reactive substances (TBARS), the generation of superoxide radicals (O2*-), other reactive oxygen species (H2O2, singlet oxygen and organic radicals) and catalase activity. Vitamin C at a low concentration (0.1 mM), like cisplatin (20 microM), induced blood platelet oxidative stress: an increase of TBARS, chemiluminescence and generation of superoxide radicals. After treatment of blood platelets with vitamin C at a high concentration (3 mM), chemiluminescence (p>0.05), the levels of O2*- (p<0.01) and TBARS (p<0.002) were reduced. We have shown that vitamin C at a high concentration (3 mM) had a protective effect against oxidative stress in platelets caused by cisplatin (20 microM). It diminished platelet lipid peroxidation and reactive oxygen species generation induced by cisplatin. In the presence of vitamin C, the catalase activity was suppressed.

Platelet aggregation and lipid peroxidation in essential hypertension

Platelet aggregation and lipid peroxidation in essential hypertension

The effect of dietary trans α-linolenic acid on plasma lipids and platelet fatty acid composition: the TransLinE study

To collect (i) baseline data and (ii) execute a large multicentre study examining the effect of trans alpha-linolenic acid on its incorporation into plasma lipids and on risk factors for coronary heart disease. Male volunteers were recruited and the habitual diet assessed by a 4-d weighed record. Fatty acid composition of plasma and platelet lipids were determined by gas chromatography at baseline. After a 6 week run-in period on a trans 'free' diet, male volunteers were randomised to consume 0.6 % of energy trans alpha-linolenic acid or to continue with a diet 'low' in trans alpha-linolenic acid for 6 weeks. Three European university research departments supported by the research and development departments of the food industry. Male volunteers (88) recruited by local advertisement. Replacement of 30 % of the fat of the habitual diet by margarine, oil and foods. Rapeseed oil was deodorised especially to produce the trans 'free' and 'high' trans foods for this study. The incorporation and conversion of trans alpha-linolenic acid into plasma lipids and platelets was assessed by gas chromatography and dietary compliance was verified by 4-d weighed record. Less trans alpha-linolenic acid isomers are incorporated into human plasma lipids in French volunteers than in Dutch or Scottish volunteers consuming their habitual diets. Trans 'free' alpha-linolenic acid-rich oil can be produced by careful deodorization during refining. The 'high' trans diet provided 1410+/-42 mg/d trans isomers of alpha-linolenic acid, whilst the 'low' trans group consumed 60+/-75 mg/d. The change in plasma lipid and platelet fatty acid composition documented that trans linolenic isomers are incorporated and converted to a trans isomer of eicosapentaenoic acid. Only the 15-trans alpha-linolenic acid is incorporated into plasma cholesteryl esters. The group consuming low trans diet had a slightly higher intake of fat, especially saturated and monounsaturated fat. Trans 'free' rapeseed oil, rich in alpha-linolenic acid, can be produced by careful deodorization. Dietary records show good compliance. Dietary trans isomers of alpha-linolenic acid are incorporated in plasma lipids and converted to long-chain polyunsaturated fatty acids. Their effects on risk factors for coronary heart disease and their metabolism will be reported elsewhere. European Commission (FAIR 95-0594 grant). European Journal of Clinical Nutrition (2000) 54, 104-113

Effects of catechins on human blood platelet aggregation and lipid peroxidation

Effects of catechins on human blood platelet aggregation and lipid peroxidation

Comparison between in vitro lipid peroxidation in fresh sheep platelets and peroxidative processes during sheep platelet ageing under storage at 4°C

Incubation of sheep platelet crude membranes with xanthine oxidase (XO)/hypoxanthine/Fe 2+-ADP revealed: (i) a fast peroxidative response – with a maximal linear rate of 14 nmol malondialdehyde (MDA) equivalents/mg protein, as evidenced by the thiobarbituric acid test – and a decrease in the polyunsaturated fatty acid (PUFA) content of the platelet crude membranes; (ii) a decrease in the lipid fluidity in the deep lipid core of the membranes but not at the membrane surface; (iii) a dramatic inhibitory effect on glucose 6-phosphatase (Glc-6-Pase) but not on acetylcholinesterase activity. Platelets were also aged by storage at 4°C in their own plasma or in Seto additive solution. In these media, platelet aggregates were visible and the effects on platelet phospholipids, PUFA, lipid extract fluorescence, crude membrane fluidity and membrane-bound enzyme activities were assessed for comparison with those observed in in vitro lipid peroxidation. The sensitivity of membranes from stored platelets to lipid peroxidation was also assessed. Storage of platelets in plasma for 5 days was associated with different changes in their crude membranes such as decreases in arachidonic acid contents, the decrease not being avoided by the presence of phospholipase A 2 inhibitors, increases in MDA equivalents, conjugated dienes and lipid extract fluorescence, decreases in the amounts of MDA equivalents formed by platelet crude membranes treated with the oxidizing agents, changes in membrane fluidity and inhibition of Glc-6-Pase. All these alterations were less pronounced or even abolished after platelet storage in Seto. These findings suggest that platelet lipid peroxidation due to XO/hypoxanthine/Fe 2+-ADP and platelet membrane alterations observed after platelet ageing under storage at 4°C share common features. Also, as regards the prevention of peroxidative processes, Seto solution permits better storage of sheep platelets than plasma.

Effect of Antioxidant Vitamin E as a Protective Factor in Experimental Atherosclerosis in Rhesus Monkeys

Atherosclerosis has been known for many years, yet its etiology remains unknown. Hypercholesterolemia is a major risk factor for atherosclerosis. The mechanism by which it triggers endothelial injury is not known. Since the role of the antioxidant vitamin E on experimental atherosclerosis is inconsistent, the present study was undertaken to evaluate platelet lipid peroxidation and the role of vitamin E (α-tocopherol) as protective factor in atherosclerosis in rhesus monkeys. A significant decrease in serum cholesterol and serum triglyceride levels was found in the group of animals which were reverted to stock diet along with vitamin E injections after 9 months of atherogenic diet feeding. Decreases in malonyldialdehyde levels and antioxidant enzyme activities were less significant in animals continued on an atherogenic diet feeding along with vitamin E as compared with animals fed a stock diet with vitamin E supplementation. The overall observations in this study suggest that antioxidant status and lipid peroxidation could be partly restored with vitamin E supplementation in experimental atherosclerosis. Damage to endothelial cells destroys their antithrombotic status and leads to fatal thrombosis. α-Tocopherol offers the best hope, but the question is how much of it should be administered for the prevention of atherosclerosis.

Age-related changes in platelet microviscosity and aggregation in rats.

1. To study possible changes in platelet microviscosity in aged animals, 1,6-diphenyl-1,3,5-hexatriene (DPH) was used as a nonpolar probe embedded in thrice-washed platelets from young, adult and aged rats. With the known values of maximum limiting anisotropy and the structural parameter of DPH and by estimating the steady state of fluorescence anisotropy and the average fluorescence of lifetime, we applied the Perrin equation to calculate the microviscosity. 2. We measured platelet aggregation, platelet lipids and platelet polyunsaturated fatty acids to determine any causal relationship between these parameters. Platelet aggregation, the platelet molar ratio of cholesterol to phospholipid ([C]/[PL]) and platelet microviscosity increased with age (P < 0.05) and were correlated with one another (P < 0.05). 3. Age-dependent increases in the steady state of fluorescence anisotropy, order parameters and the short component of fluorescence lifetime of the fluorophore were expressed as functions of variables, such as microviscosity or the [C]/[PL] ratio. 4. Platelet concentrations of arachidonic and eicosapentaenoic acids increased with age, but were not associated with aggregation. Age-related changes in microviscosity and the [C]/[PL] ratio seemed to be determinants affecting biophysical properties of platelet aggregation.

Changes in blood platelets exposed to UV-B radiation.

The effects of UV-B radiation (312 nm) on the pig-blood platelet secretory process (platelet activation) and platelet lipid peroxidation have been studied. The responses of platelets to UV-B radiation are compared with the response of these cells to thrombin, which is a strong platelet agonist. The obtained results show that exposure of blood platelets to UV-B radiation (1.2 mW/cm2, 0.072-8.64 J/cm2) causes dose-dependent platelet lipid peroxidation (measured as thiobarbituric acid reactive substances, TBARS) and release of adenine nucleotides and proteins from irradiated platelets. The dose-dependent release of platelet compounds from irradiated platelet does not correlate with the activity of platelet lactic dehydrogenase (marker of cell lysis) in the extracellular medium. It seems that UV-B radiation can partly activate platelets by stimulating the platelet secretory process and metabolism of arachidonate.

Serum-induced platelet procoagulant activity: An assay for the characterization of prothrombotic disorders

Platelets contribute to hemostasis by forming a platelet plug and by providing a procoagulant surface for the assembly and activation of the coagulation factors. The contribution of platelets to prothrombotic disorders has been difficult to analyze. Recently an assay was reported that measured the procoagulant activity of test platelets by making the platelet lipid surface the limiting factor in the production of thrombin. In this report we describe a novel technique, based on this assay, that we used to study patient serum factors that activate control platelets and in turn initiate measurable procoagulant activity. Using this assay we investigated a group of patients with prothrombotic disorders. The patient test serum was incubated with normal platelets in the presence of activated factor Xa. The resultant thrombin was measured in a chromogenic assay. The rate-limiting step was the presence of any potential platelet-activating factors, such as antibodies in the heat-treated test serum, that would allow the Xa to bind to the platelet phospholipid surface. Serum samples from patients with heparin-induced thrombocytopenia (HIT) and the anti-phospholipid antibody syndrome enhanced platelet procoagulant activity, while samples from patients with idiopathic thrombocytopenic purpura and disseminated intravascular coagulation (DIC) did not. HIT serum samples also induced platelet activation, as measured by platelet microparticle shedding, carbon 14-labeled serotonin release, and platelet aggregation. The measurement of serum-induced platelet procoagulant activity provides a method for the investigation of circulating platelet agonists in prothrombotic disorders.

Effect of He-Ne laser radiation on lipids in platelets

Low-intensity He-Ne laser radiation induces metabolic rearrangements in platelet lipids and activates some lipid-dependent systems of cell regulation as well as the synthesis of lipid-like second messengers. Modification of the lipid phase and physicochemical state of biological membranes are the molecular basis of biochemical adaptation of the platelets to laser radiation.

Persistent Impairment of Platelet Aggregation following Cessation of a Short-course Dietary Supplementation of Moderate Amounts of N-3 Fatty Acid Ethyl Esters

The duration of the effect of a short-course (1-mo twice-daily) supplementation of moderate amounts (2.28 g) of n-3 fatty acid ethyl esters (FA) on platelet lipid composition and aggregation was compared with that of olive oil (3 g/d) supplementation in 14 healthy volunteers. The FA preparation employed contained eicosapentaenoic acid (EPA) and docosahexaenoic acids (DHA) in a ratio of 1:1.4. A marked rise (p <0.05) in the plasma and platelet content of EPA and DHA, and minimal changes in the content of arachidonic acid (AA) were documented at withdrawal of the n-3 FA supplementation. EPA/AA and DHA/AA ratios in platelet phospholipids showed that the FA accumulation persisted 8-12 wks after stopping the supplementation (p <0.05). The aggregation of platelets in response to collagen or ADP, and thromboxane B2 (TXB2) formation were impaired at withdrawal. The impaired aggregation lasted 8-12 weeks (p always <0.05), whereas TXB2 formation returned to basal values 4 weeks after stopping the n-3 supplementation. No correlation was found between impaired aggregation and TXB2 formation. In contrast, the impaired sensitivity to ADP (p = 0.036) and, to a lesser extent, to collagen (p = 0.068) were related to changes in the intracellular pH (pHi) of the Na+/H+ reverse transport. No changes in platelet composition or function were observed either during or following olive oil supplementation. These results document a long-lasting impairment of platelet sensitivity to ADP and collagen; changes in the pHi values of the Na+/H+ reverse transport, and a simultaneous persistent accumulation of EPA and DHA in platelet phospholipids, after stopping a short-course dietary supplementation of moderate amounts of n-3 fatty acid ethyl esters.

Association of age-related decrease in platelet membrane fluidity with platelet lipid peroxide

The influence of age on platelet lipid peroxide (LPO), platelet membrane fluidity and the composition of fatty acid was investigated in female Wistar rats widely ranging in age from 14 to 720 days old. LPO levels were significantly higher (p < 0.05) in the platelets of upper age groups than in those of lower age groups, showing a significantly positive correlation with age (r = 0.84, p < 0.0001). Membrane fluidity, assessed by 1,6-diphenyl-1,3,5-hexatriene (DPH) fluorescence polarization, was significantly reduced with age. The composition of fatty acid demonstrated an age-related elevation (p < 0.05) in the unsaturation index. The rises in the LPO levels revealed a significantly positive correlation with DPH-polarization (r = 0.73, p < 0.0001). Thus our results suggested that the age-related deterioration of platelet membrane fluidity, despite a significant elevation in the unsaturation index, was due to the age-related higher basal levels of LPO in platelets.