Estimated Intake of Potassium, Phosphorus and Zinc with the Daily Diet Negatively Correlates with ADP-Dependent Whole Blood Platelet Aggregation in Older Subjects.

The aggregation of blood platelets is the pivotal step that leads to thrombosis. The risk of thrombotic events increases with age. Available data suggest that minerals taken with diet can affect the course of thrombosis. However, little is known about the relationship between platelet aggregability and mineral intake with diet among elderly people. Thus, we evaluated the associations between the reactivities of platelets to arachidonic acid, collagen or ADP and the estimated quantities of minerals consumed as a part of the daily diet in 246 subjects aged 60-65 years (124 men and 122 women). The found simple (not-adjusted) Spearman's rank negative correlations are as follows: 1. arachidonate-dependent aggregation and the amounts of potassium, zinc, magnesium, phosphorus, iron, copper and manganese; 2. collagen-dependent aggregation and the amounts of potassium, phosphorus, iron and zinc; and 3. ADP-dependent aggregation and the amounts of potassium, phosphorus and zinc. The negative associations between ADP-dependent platelet reactivity and the amount of potassium, phosphorus and zinc and between collagen-dependent aggregability and the amount of phosphorus were also noted after adjusting for a bunch of cardiovascular risk factors. Overall, in older subjects, the intake of minerals with diet is negatively related to blood platelet reactivity, especially in response to ADP. Diet fortification with some minerals may possibly reduce the thrombotic risk among elderly patients.

Estradiol as the Trigger of Sirtuin-1-Dependent Cell Signaling with a Potential Utility in Anti-Aging Therapies.

Aging entails the inevitable loss of the structural and functional integrity of cells and tissues during the lifetime. It is a highly hormone-dependent process; although, the exact mechanism of hormone involvement, including sex hormones, is unclear. The marked suppression of estradiol synthesis during menopause suggests that the hormone may be crucial in maintaining cell lifespan and viability in women. Recent studies also indicate that the same may be true for men. Similar anti-aging features are attributed to sirtuin 1 (SIRT1), which may possibly be linked at the molecular level with estradiol. This finding may be valuable for understanding the aging process, its regulation, and possible prevention against unhealthy aging. The following article summarizes the initial studies published in this field with a focus on age-associated diseases, like cancer, cardiovascular disease and atherogenic metabolic shift, osteoarthritis, osteoporosis, and muscle damage, as well as neurodegenerative and neuropsychiatric diseases.

Contribution of Physical Activity to the Oxidative and Antioxidant Potential in 60-65-Year-Old Seniors.

Both acute exercise and regular physical activity (PA) are directly related to the redox system. However, at present, there are data suggesting both positive and negative relationships between the PA and oxidation. In addition, there is a limited number of publications differentiating the relationships between PA and numerous markers of plasma and platelets targets for the oxidative stress. In this study, in a population of 300 participants from central Poland (covering the age range between 60 and 65 years), PA was assessed as regards energy expenditure (PA-EE) and health-related behaviors (PA-HRB). Total antioxidant potential (TAS), total oxidative stress (TOS) and several other markers of an oxidative stress, monitored in platelet and plasma lipids and proteins, were then determined. The association of PA with oxidative stress was determined taking into the account basic confounders, such as age, sex and the set of the relevant cardiometabolic factors. In simple correlations, platelet lipid peroxides, free thiol and amino groups of platelet proteins, as well as the generation of superoxide anion radical, were inversely related with PA-EE. In multivariate analyses, apart from other cardiometabolic factors, a significant positive impact of PA-HRB was revealed for TOS (inverse relationship), while in the case of PA-EE, the effect was found to be positive (inverse association) for lipid peroxides and superoxide anion but negative (lower concentration) for free thiol and free amino groups in platelets proteins. Therefore, the impact of PA may be different on oxidative stress markers in platelets as compared to plasma proteins and also dissimilar on platelet lipids and proteins. These associations are more visible for platelets than plasma markers. For lipid oxidation, PA seems to have protective effect. In the case of platelets proteins, PA tends to act as pro-oxidative factor.

Carbosilane ruthenium metallodendrimer as alternative anti-cancer drug carrier in triple negative breast cancer mouse model: A preliminary study.

The carbosilane metallodendrimer G1-[[NCPh(o-N)Ru(η6- p-cymene)Cl]Cl]4 (CRD13), based on an arene Ru(II) complex coordinated to imino-pyridine surface groups, has been conjugated with anti-cancer drugs. Ruthenium in the positively-charged dendrimer structure allows this nanoparticle to be considered as an anticancer drug carrier, made more efficient because ruthenium has anticancer properties. The ability of CRD13 to form complexes with Doxorubicin (DOX), 5-Fluorouracil (5-Fu), and Methotrexate (MTX) has been evaluated using zeta potential measurement, transmission electron microscopy (TEM) and computer simulation. The results show that it forms stable nanocomplexes with all those drugs, enhancing their effectiveness against MDA-MB-231 cancer cells. In vivo tests indicate that the CRD13/DOX system caused a decrease of tumor weight in mice with triple negative breast cancer. However, the tumors were most visibly reduced when naked dendrimers were injected.

Relationships between Plasma Concentrations of Testosterone and Dihydrotestosterone and Geriatric Depression Scale Scores in Men and Women Aged 60-65 Years-A Multivariate Approach with the Use of Quade's Test.

The potential role of testosterone and dihydrotestosterone in the pathogenesis of depression in older subjects is poorly recognized and understood. The current study examines the symptoms of depression in males and females at the age of 60-65 using a short version (15 questions) of the Geriatric Depression Scale (GDS) questionnaire. Blood plasma levels of androgens were estimated by LC/MS/MS. Total GDS score calculated for males were not found to be significantly associated with plasma levels of testosterone or dihydrotestosterone. Older men with higher plasma testosteronemia were more likely to report being in good spirits most of the time, but more willing to stay at home than undertake outside activities. The men with higher plasma levels of dihydrotestosterone also perceived themselves as being in good spirits most of the time. Older men with higher testosterone were more likely to report having more problems with their memory than others. No significant associations were found between plasma levels of androgens and GDS scores in older women; however, some tendencies suggest that testosterone and dihydrotestosterone may act as antidepressants in older women.

The amino acid content in the daily diet of seniors negatively correlates with the degree of platelet aggregation in a sex- and agonist-specific manner.

Aging is a significant risk factor for the development of thrombotic diseases, dependent on blood platelet reactivity. However, the risk of thrombosis also appears to be significantly modulated by dietary nutrient content. The aim of the current study was to assess the relationship between the amount of amino acids present in the daily diet (not supplemented) and the reactivity of blood platelets to arachidonate, collagen and ADP in 246 women and men aged 60-65 years. Platelet reactivity was tested using whole blood impedance aggregometry. Amino acid intake was assessed with a 24-hour Recall Questionnaire and calculated with Dieta 5.0 software. Older subjects receiving higher amounts of all essential amino acids with their daily diet exhibit significantly lower platelet responsiveness to AA-, COL- and ADP in a sex-specific manner: dietary amino acid content was more closely associated with AA- and, to some extent, ADP-induced platelet reactivity in women, and with COL-induced platelet aggregability in men. Therefore, dietary amino acid content may be a novel factor responsible for attenuating platelet reactivity in a sex- and agonist-specific manner.

The Association of Oxidative and Antioxidant Potential with Cardiometabolic Risk Profile in the Group of 60- to 65-Year-Old Seniors from Central Poland.

Pathogenesis of cardiovascular diseases is caused by, inter alia, oxidative stress. On the other hand, cardiovascular risk factors may cause redox imbalance. The pathological pathways between those components are to be determined. In the group comprised of 300 sex-matched subjects, we evaluated a number of cardiovascular risk factors: blood pressure, body mass, lipids, glucose, homocysteine, uric acid, von Willebrand factor (vWF), VCAM-1 and ICAM-1. The presence of cardiovascular diseases and drugs for their treatment were examined. Secondly, we assessed total antioxidative status (TAS), total oxidative status (TOS) and other markers of oxidative stress. TAS was inversely related to LDL cholesterol. TOS was positively associated with BMI and female sex, but negatively associated with the use of angiotensin II receptor antagonists. Plasma lipid peroxides concentration was positively related to ICAM-1 and presence of stroke, whereas platelet lipid peroxides were positively associated with vWF. Platelets proteins thiol groups were in a positive relationship with vWF, but in a negative relationship with uric acid and diagnosed lipid disorders. Both free thiol and amino groups were positively associated with plasma glucose. Platelets free amino groups were related to platelets count. Superoxide generation by blood platelets (both with and without homocysteine) was positively connected to glucose level. Among women, oxidative markers appear to be more related to glucose level, whereas among men they are related to body mass indices. TAS, TOS and oxidative markers are largely related to modifiable cardiovascular risk factors such as body mass, and intake of drugs such as angiotensin II receptor blockers. Plasma and platelet oxidation markers appear to be especially associated with glucose concentration. The presented analyses unanimously indicate strong connections between cardiovascular risk factors and redox potential and specify how cardiometabolic interventions may counter-balance oxidative stress.

Protein disulfide isomerase-A1 regulates intraplatelet reactive oxygen species-thromboxane A2 -dependent pathway in human platelets.

BACKGROUND: Platelet-derived protein disulfide isomerase 1 (PDIA1) regulates thrombus formation, but its role in the regulation of platelet function is not fully understood. AIMS: The aim of this study was to characterize the role of PDIA1 in human platelets. METHODS: Proteomic analysis of PDI isoforms in platelets was performed using liquid chromatography tandem mass spectometry, and the expression of PDIs on platelets in response to collagen, TRAP-14, or ADP was measured with flow cytometry. The effects of bepristat, a selective PDIA1 inhibitor, on platelet aggregation, expression of platelet surface activation markers, thromboxane A2 (TxA2 ), and reactive oxygen species (ROS) generation were evaluated by optical aggregometry, flow cytometry, ELISA, and dihydrodichlorofluorescein diacetate-based fluorescent assay, respectively. RESULTS: PDIA1 was less abundant compared with PDIA3 in resting platelets and platelets stimulated with TRAP-14, collagen, or ADP. Collagen, but not ADP, induced a significant increase in PDIA1 expression. Bepristat potently inhibited the aggregation of washed platelets induced by collagen or convulxin, but only weakly inhibited platelet aggregation induced by TRAP-14 or thrombin, and had the negligible effect on platelet aggregation induced by arachidonic acid. Inhibition of PDIA1 by bepristat resulted in the reduction of TxA2 and ROS production in collagen- or thrombin-stimulated platelets. Furthermore, bepristat reduced the activation of αIIbß3 integrin and expression of P-selectin. CONCLUSIONS: PDIA1 acts as an intraplatelet regulator of the ROS-TxA2 pathway in collagen-GP VI receptor-mediated platelet activation that is a mechanistically distinct pathway from extracellular regulation of αIIbß3 integrin by PDIA3.

Plasma Concentration of Cortisol Negatively Associates with Platelet Reactivity in Older Subjects.

The interaction of platelets with steroid hormones is poorly investigated. Age is one of the factors that increase the risk of pathological platelet reactivity and thrombosis. The aim of this study was to assess whether there were associations between platelet reactivity and plasma cortisol levels in volunteers aged 60-65 years. For this purpose, impedance aggregometry in whole blood measured after arachidonic acid, collagen, or ADP stimulation was used to estimate platelet reactivity and mass spectrometry was used to measure peripheral plasma cortisol concentration. Statistically significant negative correlations were observed between cortisol concentration and platelet reactivity in response to arachidonic acid and ADP, but not to collagen. The presented results suggest for the very first time that cortisol is a new endogenous modulator of platelet reactivity in the elderly population.

Melatonin as a Reducer of Neuro- and Vasculotoxic Oxidative Stress Induced by Homocysteine.

The antioxidant properties of melatonin can be successfully used to reduce the effects of oxidative stress caused by homocysteine. The beneficial actions of melatonin are mainly due to its ability to inhibit the generation of the hydroxyl radical during the oxidation of homocysteine. Melatonin protects endothelial cells, neurons, and glia against the action of oxygen radicals generated by homocysteine and prevents the structural changes in cells that lead to impaired contractility of blood vessels and neuronal degeneration. It can be, therefore, assumed that the results obtained in experiments performed mainly in the in vitro models and occasionally in animal models may clear the way to clinical applications of melatonin in patients with hyperhomocysteinemia, who exhibit a higher risk of developing neurodegenerative diseases (e.g., Parkinson's disease or Alzheimer's disease) and cardiovascular diseases of atherothrombotic etiology. However, the results that have been obtained so far are scarce and have seldom been performed on advanced in vivo models. All findings predominately originate from the use of in vitro models and the scarcity of clinical evidence is huge. Thus, this mini-review should be considered as a summary of the outcomes of the initial research in the field concerning the use of melatonin as a possibly efficient attenuator of oxidative stress induced by homocysteine.

Synthesis and evaluation of adenosine derivatives as A1, A2A, A2B and A3 adenosine receptor ligands containing boron clusters as phenyl isosteres and selective A3 agonists.

A series of adenosine and 2'-deoxyadenosine pairs modified with a 1,12-dicarba-closo-dodecaborane cluster or alternatively with a phenyl group at the same position was synthesized, and their affinity was determined at A1, A2A, A2B and A3 adenosine receptors (ARs). While AR affinity differences were noted, a general tendency to preferentially bind A3 AR over other ARs was observed for most tested ligands. In particular, 5'-ethylcarbamoyl-N6-(3-phenylpropyl)adenosine (18), N6-(3-phenylpropyl)-2-chloroadenosine (24) and N6-(3-phenylpropyl)adenosine (40) showed nanomolar A3 affinity (Ki 4.5, 6.4 and 7.5 nM, respectively). Among the boron cluster-containing compounds, the highest A3 affinity (Ki 206 nM) was for adenosine derivative 41 modified at C2. In the matched molecular pairs, analogs bearing boron clusters were found to show lower binding affinity for adenosine receptors than the corresponding phenyl analogs. Nevertheless, interestingly, several boron cluster modified adenosine ligands showed significantly higher A3 receptor selectivity than the corresponding phenyl analogs: 7vs. 8, 15vs. 16, 17vs. 18.

Adenosine Receptor Agonist HE-NECA Enhances Antithrombotic Activities of Cangrelor and Prasugrel in vivo by Decreasing of Fibrinogen Density in Thrombus.

Blood platelets' adenosine receptors (AR) are considered to be a new target for the anti-platelet therapy. This idea is based on in vitro studies which show that signaling mediated by these receptors leads to a decreased platelet response to activating stimuli. In vivo evidence for the antithrombotic activity of AR agonists published to date were limited, however, to the usage of relatively high doses given in bolus. The present study was aimed at verifying if these substances used in lower doses in combination with inhibitors of P2Y12 could serve as components of dual anti-platelet therapy. We have found that a selective A2A agonist 2-hexynyl-5'-N-ethylcarboxamidoadenosine (HE-NECA) improved the anti-thrombotic properties of either cangrelor or prasugrel in the model of ferric chloride-induced experimental thrombosis in mice. Importantly, HE-NECA was effective not only when applied in bolus as other AR agonists in the up-to-date published studies, but also when given chronically. In vitro thrombus formation under flow conditions revealed that HE-NECA enhanced the ability of P2Y12 inhibitors to decrease fibrinogen content in thrombi, possibly resulting in their lower stability. Adenosine receptor agonists possess a certain hypotensive effect and an ability to increase the blood-brain barrier permeability. Therefore, the effects of anti-thrombotic doses of HE-NECA on blood pressure and the blood-brain barrier permeability in mice were tested. HE-NECA applied in bolus caused a significant hypotension in mice, but the effect was much lower when the substance was given in doses corresponding to that obtained by chronic administration. At the same time, no significant effect of HE-NECA was observed on the blood-brain barrier. We conclude that chronic administration of the A2A agonist can be considered a potential component of a dual antithrombotic therapy. However, due to the hypotensive effect of the substances, dosage and administration must be elaborated to minimize the side-effects. The total number of animals used in the experiments was 146.

Blood Platelets as an Important but Underrated Circulating Source of TGFß.

When treating diseases related primarily to tissue remodeling and fibrosis, it is desirable to regulate TGFß concentration and modulate its biological effects. The highest cellular concentrations of TGFß are found in platelets, with about 40% of all TGFß found in peripheral blood plasma being secreted by them. Therefore, an understanding of the mechanisms of TGFß secretion from platelets may be of key importance for medicine. Unfortunately, despite the finding that platelets are an important regulator of TGFß levels, little research has been carried out into the development of platelet-directed therapies that might modulate the TGFß-dependent processes. Nevertheless, there are some very encouraging reports suggesting that platelet TGFß may be specifically involved in cardiovascular diseases, liver fibrosis, tumour metastasis, cerebral malaria and in the regulation of inflammatory cell functions. The purpose of this review is to briefly summarize these few, extremely encouraging reports to indicate the state of current knowledge in this topic. It also attempts to better characterize the influence of TGFß on platelet activation and reactivity, and its shaping of the roles of blood platelets in haemostasis and thrombosis.

Time-dependent interactions of blood platelets and cancer cells, accompanied by extramedullary hematopoiesis, lead to increased platelet activation and reactivity in a mouse orthotopic model of breast cancer - implications for pulmonary and liver metastasis.

Aging has become a significant risk factor for several diseases, including breast cancer.Platelet activation and platelet-cancer cell aggregate fractions were found to increase with tumor progression in a mouse model of breast cancer. At advanced stages of tumor development, platelets from mice with breast cancer were hyperreactive to low agonist concentrations and hyporeactive to high ones. Platelet activation and reactivity were strongly associated with breast cancer metastasis in the lungs and extramedullary hematopoiesis in the liver. A greater fraction of platelet aggregates was observed in 4T1-injected mice at the advanced stages of breast cancer. In vitro, platelet activation was elevated after incubation with 4T1 cells, and thrombin-stimulated platelets formed aggregates with 4T1 cells. Neither GPIbα, nor GPIIb/IIIa blocking antibodies, were able to affect platelet-cancer cell aggregation in vitro.The primed circulating platelets became more sensitive to subthreshold stimuli at advanced stages of tumor development, and the formation of platelet-cancer cell aggregates increased with cancer progression. Our findings demonstrate that the age-associated progression of breast cancer cells is connected with increased platelet functioning, and that it can be manifested by the increased number of metastases and extramedullary hematopoiesis in a time-dependent-manner.

Functional inhibition of F11 receptor (F11R/junctional adhesion molecule-A/JAM-A) activity by a F11R-derived peptide in breast cancer and its microenvironment.

PURPOSE: To examine the involvement of the F11R/JAM-A protein in breast cancer metastasis, we utilized the F11R/JAM-A antagonistic peptide 4D (P4D) in experiments of transendothelial migration (TEM) of breast cancer cells. METHODS: Experiments were conducted in the mouse 4T1 breast cancer model utilizing the human mammary epithelial cell and endothelial cell lines. The levels of soluble F11R/JAM-A (sJAM-A) in the murine plasmas were measured by ELISA. Levels of F11R/JAM-A mRNA and protein in cell lines were assessed by qRT-PCR and Western blot, respectively. Cell surface expression of F11R/JAM-A was demonstrated by flow cytometry. Functional tests included the TEM of breast cancer cells and adhesion of breast cancer cells to the endothelium. The endothelial permeability was studied by fluorescent tracer assay and by the Real-Time Cell Analysis (RTCA). RESULTS: The tumor inducers Tß4 and TGF-ß1 reduced the levels of sJAM-A in murine plasma, and reduced the F11R/JAM-A protein levels in the human microvascular endothelial cell line HMEC-1. The adhesion and TEM measured between breast cancer cells and inflamed or Tß4-treated endothelium were inhibited by P4D. The presence of P4D did not destabilize the pre-existing tight junctions in the endothelial monolayer. The barrier-protecting effect of P4D was stronger than that of forskolin, when a booster dose of P4D was applied to the inflamed endothelium. CONCLUSIONS: F11R/JAM-A protein can be considered as a novel target in the treatment of breast cancer metastasis. In vivo and clinical studies are needed to further investigate the effectiveness of F11R/JAM-A-derived peptide as a possible anti-metastatic drug.

What is the most important determinant of cardiometabolic risk in 60-65-year-old subjects: physical activity-related behaviours, overall energy expenditure or occupational status? A cross-sectional study in three populations with different employment status in Poland.

OBJECTIVES: The aim of the study was to determine whether cardiovascular risk factors may differ according to occupational status and whether physical activity related to total energy expenditure (PA-EE) and related to health-related behaviours (PA-HRB) is associated with common cardiovascular risk factors or metabolic syndrome in pre-elderly subjects. METHODS: Three hundred subjects aged 60-65 were recruited and divided into three equal groups of white-collar, blue-collar workers and unemployed subjects; 50% were women. The subjects were tested for major cardiovascular risk factors such as smoking, anthropometric indices, blood pressure, lipid levels, glucose, uric acid and homocysteine. PA-EE and PA-HRB were assessed with PA questionnaires. RESULTS: Blue-collar workers displayed higher anthropometric indices, blood pressure and higher PA-EE in comparison with other two groups. PA-HRB had a positive impact on body mass indices, lipids, glucose, uric acid and the prevalence of metabolic syndrome, with no such relationship observed for PA-EE. CONCLUSIONS: The greatest cardiovascular risk was observed in the blue-collar workers group. Only PA-HRB had a positive association with cardiometabolic risk profile. No relationship was observed for PA-EE. Thus, promoting everyday life and leisure time PA behaviours is crucial for preventing cardiometabolic risk in pre-elderly subjects, even in blue-collar workers with high work-related EE.

The Mystery behind the Pineal Gland: Melatonin Affects the Metabolism of Cholesterol.

Melatonin may be considered a cardioprotective agent. Since atherogenesis is partly associated with the metabolism of lipoproteins, it seems plausible that melatonin affects cardiovascular risk by modulating the metabolism of cholesterol and its subfractions. Moreover, cholesterol-driven atherogenesis can be hypothetically reduced by melatonin, mainly due to the minimalization of harmful reactions triggered in the cardiovascular system by the reactive oxygen species-induced toxic derivatives of cholesterol. In this review, we attempted to summarize the available data on the hypolipemizing effects of melatonin, with some emphasis on the molecular mechanisms underlying these reactions. We aimed to attract readers' attention to the numerous gaps of knowledge present in the reviewed field and the essential irrelevance between the findings originating from different sources: clinical observations and in vitro mechanistic and molecular studies, as well as preclinical experiments involving animal models. Overall, such inconsistencies make it currently impossible to give a reliable opinion on the action of melatonin on the metabolism of lipoproteins.

Platelet and Red Blood Cell Counts, as well as the Concentrations of Uric Acid, but Not Homocysteinaemia or Oxidative Stress, Contribute Mostly to Platelet Reactivity in Older Adults.

PURPOSE: The goal of this study was to estimate the hierarchical contribution of the most commonly recognized cardiovascular risk factors associated with atherogenesis to activation and reactivity of blood platelets in a group of men and women at ages 60-65. METHODS: Socioeconomic and anthropometric data were taken from questionnaires. Blood morphology and biochemistry were measured with standard diagnostic methods. Plasma serum homocysteine was measured by immunochemical method. Plasma concentrations of VCAM, ICAM, total antioxidant status, and total oxidant status were estimated with commercial ELISA kits. Markers of oxidative stress of plasma and platelet proteins (concentrations of protein free thiol and amino groups) and lipids (concentrations of lipid peroxides) and generation of superoxide anion by platelets were measured with colorimetric methods. Platelet reactivity was estimated by impedance aggregometry with arachidonate, collagen, and ADP as agonists. Expression of selectin-P and GPIIb/IIIa on blood platelets was tested by flow cytometry. RESULTS: Platelet aggregation associated significantly negatively with HGB and age and significantly positively with PLT, MPV, PCT, PDW, and P-LCR. When platelet reactivity ("cumulative platelet reactivity_aggregation") was analyzed in a cumulated manner, the negative association with serum concentration of uric acid (R s = -0.169, p = 0.003) was confirmed. Multivariate analysis revealed that amongst blood morphological parameters, platelet count, plateletcrit, and number of large platelets and uric acid are the most predictive variables for platelet reactivity. CONCLUSIONS: The most significant contributors to platelet reactivity in older subjects are platelet morphology, plasma uricaemia, and erythrocyte morphology.

Testosterone and dihydrotestosterone reduce platelet activation and reactivity in older men and women.

The cardiovascular effects of testosterone and dihydrotestosterone are generally attributed to their modulatory action on lipid and glucose metabolism. However, no ex vivo studies suggest that circulating androgen levels influence the activation and reactivity of blood platelets - one of the main components of the haemostasis system directly involved in atherosclerosis. The levels of testosterone, dihydrotestosterone and oestradiol in plasma from men and women aged from 60 to 65 years were measured by LC-MS; the aim was to identify any potential relationships between sex steroid levels and the markers of platelet activation (surface membrane expression of GPII/IIIa complex and P-selectin) and platelet reactivity in response to arachidonate, collagen or ADP, monitored with whole blood aggregometry and flow cytometry. The results of the ex vivo part of the study indicate that the concentrations of testosterone and its reduced form, dihydrotestosterone are significantly negatively associated with platelet activation and reactivity. These observations were confirmed in an in vitro model: testosterone and dihydrotestosterone significantly inhibited platelet aggregation triggered by arachidonate or collagen. Our findings indicate that testosterone and dihydrotestosterone are significant haemostatic steroids with inhibitory action on blood platelets in older people.

Inhibition of glutamate receptors reduces the homocysteine-induced whole blood platelet aggregation but does not affect superoxide anion generation or platelet membrane fluidization.

Homocysteine (Hcy) is an excitotoxic amino acid. It is potentially possible to prevent Hcy-induced toxicity, including haemostatic impairments, by antagonizing glutaminergic receptors. Using impedance aggregometry with arachidonate and collagen as platelet agonists, we tested whether the blockade of platelet NMDA (N-methyl-D-aspartate), AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) and kainate receptors with their inhibitors: MK-801 (dizocilpine hydrogen maleate, [5R,10S]-[+]-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine), CNQX (7-nitro-2,3-dioxo-1,4-dihydroquinoxaline-6-carbonitrile) and UBP-302 (2-{[3-[(2S)-2-amino-2-carboxyethyl]-2,6-dioxo-3,6-dihydropyrimidin 1(2H)-yl]methyl}benzoic acid) may hamper Hcy-dependent platelet aggregation. All the tested compounds significantly inhibited Hcy-augmented aggregation of blood platelets stimulated either with arachidonate or collagen. Hcy stimulated the generation of superoxide anion in whole blood samples in a concentration-dependent manner; however, this process appeared as independent on ionotropic glutamate receptors, as well as on NADPH oxidase and protein kinase C, and was not apparently associated with the extent of either arachidonate- or collagen-dependent platelet aggregation. Moreover, Hcy acted as a significant fluidizer of surface (more hydrophilic) and inner (more hydrophobic) regions of platelet membrane lipid bilayer, when used at the concentration range from 10 to 50 µmol/l. However, this effect was independent on the Hcy action through glutamate ionotropic receptors, since there was no effects of MK-801, CNQX or UBP-302 on Hcy-mediated membrane fluidization. In conclusion, Hcy-induced changes in whole blood platelet aggregation are mediated through the ionotopic excitotoxic receptors, although the detailed mechanisms underlying such interactions remain to be elucidated.