Platelet Aggregation In Response Research Articles

Assessment of Aspirin and Clopidogrel Resistance in Patients Undergoing Cardiovascular Surgery: A Single Center Cross-Sectional Study.

We aimed to investigate the antiplatelet drug resistance utilizing light transmission-lumiaggregometry (LT-LA) and platelet function analyzer-100 (PFA-100) in patients undergoing cardiovascular surgery. The study included 60 patients diagnosed with stable coronary artery disease and peripheral vascular diseases that required surgery. The participants were divided into three groups: patients receiving Aspirin (ASA) (n=21), patients receiving Clopidogrel (CLO) (n=19), and patients receiving dual therapy (ASA+CLO) (n=20). Aggregation and secretion tests in LT-LA and closure time in PFA-100 were used to measure antiplatelet drug resistance. Based on the ADP-induced aggregation test, 43% of patients were resistant to ASA, 22% to CLO, and 15% to dual therapy. Diabetes, hypertension, and hyperlipidemia were the most commonly identified comorbid disorders. In patients with comorbid risk factors, the median value of platelet aggregation response to ADP was significantly higher in the ASA group than in the CLO and dual therapy groups (p=0.0001). In patients receiving ASA monotherapy, the maximum amplitude of aggregation response to platelet agonists was equal to or more than 70% in 43% of patients for ADP and 28% for collagen in LT-LA. Elevated ADP (≥0.29 nmol) and collagen (≥0.41 nmol)-induced ATP release were found in 66% of patients utilizing ADP agonist and 80% of patients using collagen agonist undergoing ASA therapy in LT-LA. Closure times in PFA-100 were normal in 28% of patients using collagen-ADP cartridges and 62% of patients using collagen-epinephrine (CEPI) cartridges who received ASA. Recurrent thrombosis and bleeding were observed in 12 (20%) of patients with cardiovascular disease. Three of these individuals (25%) showed ASA resistance, which showed normal response to ADP-induced aggregation (≥70%) and secretion (≥0.29 nmol), as well as normal CEPI closure times. Our findings suggest that antiplatelet drug monitoring with LT-LA and PFA-100 may be useful in high-risk and complicated cardiovascular patients.

Potential Protein Signatures for Recurrence Prediction of Ischemic Stroke.

Acute ischemic stroke is a major cause of mortality and disability worldwide, with approximately 7.4% to 7.7% recurrence within the first 3 months. This study aimed to identify potential biomarkers for predicting stroke recurrence. We conducted a nested case-control study using a hospital-based cohort from the Third China National Stroke Registry selecting 214 age- and sex-matched patients with ischemic stroke with hypertension and no history of diabetes or heart disease. Using data-independent acquisition for discovery and multiple reaction monitoring for quantitative validation, we identified 26 differentially expressed proteins in large-artery atherosclerosis (Causative Classification of Ischemic Stroke [CCS]1), 16 in small-artery occlusion (CCS3), and 25 in undetermined causes (CCS5) among patients with recurrent stroke. In the CCS1 and CCS3 subgroups, differentially expressed proteins were associated with platelet aggregation, neuronal death/cerebroprotection, and immune response, whereas differentially expressed proteins in the CCS5 subgroup were linked to altered metabolic functions. Validated recurrence predictors included proteins associated with neutrophil activity and vascular inflammation (TAGLN2 [transgelin 2], ITGAM [integrin subunit α M]/TAGLN2 ratio, ITGAM/MYL9 [myosin light chain 9] ratio, TAGLN2/RSU1 [Ras suppressor protein 1] ratio) in the CCS3 subgroup and proteins associated with endothelial plasticity and blood-brain barrier integrity (ITGAM/MYL9 ratio and COL1A2 [collagen type I α 2 chain]/MYL9 ratio) in the CCS3 and CCS5 subgroups, respectively. These findings provide a foundation for developing a blood-based biomarker panel, using causative classifications, which may be used in routine clinical practice to predict stroke recurrence.

Impact of lipid emulsions in parenteral nutrition on platelets: a literature review.

Lipid emulsions are essential components of parenteral nutrition solutions that provide energy and essential fatty acids. The complexity of the formulations of lipid emulsions may lead to adverse outcomes such as platelet reactivity and changes in platelet aggregation and related coagulation. Platelets are responsible for haemostasis; they activate and demonstrate morphological changes upon extracellular factors to maintain blood fluidity and vascular integrity. Although parenteral nutrition lipid emulsions are generally found safe with regard to modulation of platelet activity, studies are still accumulating. Thus, this review aims to investigate platelet-related changes by parenteral nutrition lipid emulsions in human studies. Studies have pointed out patients at risk of bleeding and increased platelet aggregation responses due to the administration of lipid emulsions. Lipid emulsions may further benefit patients at high risk of thrombosis due to anti-thrombotic effects and should be cautiously used in patients with thrombocytopenia. The reported platelet-related changes might be associated with the fatty acid change in the plasma membranes of platelets following changes in platelet synthesis and plasma levels of eicosanoids. In conclusion, studies investigating platelets and parenteral nutrition should be supported to minimize the adverse effects and to benefit from the potential protective effects of parenteral nutrition lipid emulsions.

Association of platelet aggregation to amyloid and tau PET in the Framingham Heart Study

AbstractBackgroundPlatelets are key mediators of inflammation and vascular dysfunction. Prior work highlighted a role of platelet function in Ab and tau dynamics, and dementia risk. We hypothesized that platelet aggregation may associate with amyloid and tau brain levels in middled age people at risk of cardiovascular disease.MethodWe examined cross sectional associations of platelet aggregation with brain amyloid and tau PET levels (11C‐labeled Pittsburgh Compound‐B (PiB) and tau (18F‐Flortaucipir (FTP)) in middle aged cognitively normal participants at the Framingham Heart Study (FHS) Gen3 Cohort. Platelet aggregation was measured by light transmission aggregometry. The response to 0.98uM ADP stimulation was primarily evaluated followed by additional agonists and stimulation doses for sensitivity analysis. The associations were explored in multivariate regression models adjusted for age, age squared, sex, the time from blood to PET examination, and PET camera used.ResultThe final study sample included 356 participants (median [Q1,Q3] age, 55 [49, 61]; 51% women) of which 82 (23%) were on preventive platelet modifying treatments (PPT) (Table 1). We found significant interaction between PPT and platelet aggregation in their association with amyloid and tau PET levels, motivating a stratified analysis by the use or not of PPT. The platelet aggregation response was not associated with PiB or FTP retention in non‐PPT users (n = 274) (Table 2). In PPT users, the platelet aggregation response was positively associated with amyloid levels in the precuneus (β±SE: 0.36±0.14, P<0.05), and tau levels in the rhinal (β±SE: 0.49±0.15, P<0.01) and temporal regions (β±SE: 0.39±0.17, P<0.05).ConclusionOur results indicate that platelet aggregation is not associated with amyloid and tau PET levels in the general population. In people who are on PPT for primary prevention of CVD, platelet aggregation associates with amyloid and tau PET in Alzheimer’s disease (AD) relevant brain regions. This result reinforces the need for further examining the mediator role of platelet aggregation in the association of cardiovascular risk and AD.

Mice Lacking α-Actinin-1 in Megakaryocytes Display the Feature of Thrombocytopenia and Impair Platelet Functions

Cytoskeleton remodeling and mitochondrial bioenergetics play important roles in platelet biogenesis and functions. α-Actinin-1, a filament crosslinker, was widely expressed in nonmuscle cells, including megakaryocytes (MKs) and platelets. The cytoskeleton is essential for normal mitochondrial morphology, motility, and distribution. In recent years, the accumulated evidence suggests that inherited mutations in α-actinin-1 are implicated in congenital macrothrombocytopenia. However, the role and underlying mechanism of α-actinin-1 in platelet biogenesis and platelet functions remain poorly studied. In this study, we generated an MK-specific α-actinin-1 knockout mouse model (referred to herein as PF4- Actn1 -/-) to investigate the contribution of α-actinin-1 in platelet biogenesis and functions. Blood count tests showed that PF4- Actn1 -/- mice exhibited reduced platelet counts. The decreased platelet number in PF4- Actn1 -/- mice was not attributed to accelerated platelet clearance or impaired TPO generation in vivo. Platelet count changes observed in an anti-GPIbα antibody-induced thrombocytopenia model and splenectomy indicated that the diminished platelet counts in PF4- Actn1 -/- mice might be due to defects in platelet biogenesis. Compared to control mice, PF4- Actn1 -/- mice had normal numbers of MK progenitors, including in LSK, MPP, CMP, GMP, MEP, and PreMegE. H&E staining and flow cytometry revealed a decrease in the number of MKs in BM, but an increase in the number of MKs in the spleen. The absence of α-actinin-1 significantly increased the proportion of 2N-4N MKs and decreased the proportion of 8N-32N MKs. CFU-MK colony formation and MK migration in response to SDF-1 signaling was inhibited in PF4- Actn1 -/- mice. Furthermore, in vitro studies showed a reduced ratio of PPF-bearing MKs in fetal liver-derived PF4- Actn1 -/- MKs. Collectively, these data suggest that α-actinin-1 deficiency in mouse MKs could impair platelet biogenesis, resulting in thrombocytopenia in PF4- Actn1 -/- mice. Platelet adhesion and spreading on immobilized fibrinogen and collagen, thrombin-stimulated clot retraction, and platelet aggregation in response to various concentrations of ADP, thrombin, and collagen was significantly decreased in PF4- Actn1 -/- platelets. PF4- Actn1 -/- platelets also exhibited decreased integrin αIIbβ3 activation and reduced P-selectin exposure in response to various concentrations of ADP, ADP+EPI, thrombin, collagen, and convulxin. Notably, PF4- Actn1 -/- platelets showed inhibited calcium mobilization, reactive oxygen species (ROS) generation, and actin polymerization in response to collagen and thrombin. Furthermore, PF4- Actn1 -/- mice demonstrated significantly impaired hemostasis. No differences were observed in the PT, APTT, TT, and fibrinogen concentrations between PF4- Actn1 -/- mice and Actn1 f/f mice. However, tail, liver, and brain bleeding tests revealed that PF4- Actn1 -/- mice had significantly prolonged bleeding time and increased bleeding volume. Kaolin-activated whole thromboelastography (TEG) mapping assay indicated that there were no differences in the R-time, K-time, and α-angle values, but the maximum amplitude was decreased in PF4- Actn1 -/- mice. To explore the mechanism of α-actinin-1 in platelet biogenesis and platelet functions. Low- (2-4N) and high-ploidy (≥8N) Actn1 f/f and PF4- Actn1 -/- MKs were sorted for quantitative proteomics analysis. The results of 4D label-free quantitative proteomics revealed that protein expression was significantly decreased in high-ploidy MKs following α-actinin-1 knockdown. Functional enrichment analysis of differentially expressed proteins indicated that α-actinin-1 deletion reduced platelet activation and mitochondrial functions. PF4- Actn1 -/- platelets exhibited reduced mitochondrial membrane potential, mitochondrial ROS generation, mitochondrial respiration and glycolysis, suggesting alterations in mitochondrial function in platelets. In this study, we report that mice with α-actinin-1 deficiency in MKs recapitulate the features of thrombocytopenia and exhibit impaired platelet functions.

The bleeding diathesis in patients with hereditary haemorrhagic telangiectasia is not due to impaired platelet function

BackgroundPatients with the rare disease; Hereditary haemorrhagic telangiectasia (HHT) often bleed from telangiectatic lesions in mucosal surfaces. Studies suggest that impaired platelet function may also play a role in their bleeding tendency. The aim of the present study was to investigate whether HHT-patients with epistaxis have impaired platelet function.MethodWe conducted a case–control study based on a sample size calculation and included 22 HHT-patients (inclusion criteria: epistaxis severity score ≥ 4, no intake of medicine affecting platelet function the last 5 days, HHT-type 1 or 2, age ≥ 18 years) and 20 controls. We assessed the platelet function with standard haemostasis parameters, flow cytometry (platelet function and micro aggregation), rotational thromboelastometry and Platelet Function Analyzer 200.ResultsWe found no significant difference in mean platelet volume and immature platelet fraction and no difference in platelet activation as measured by exposure of CD62P, CD63P and PAC1 binding. Nor did we find a significant difference in platelet aggregation response in HHT-patients compared with the control group for all agonists (thrombin receptor activating peptide, adenosine diphosphate and collagen-related peptide). The PFA-200 analysis was without difference between the two groups and thromboelastometry showed no impairment of global haemostasis.ConclusionReduced platelet function is unlikely to contribute to the frequent and long bleeding episodes that HHT-patients suffer from. We propose that further studies should focus on whether patients with HHT have hypercoagulability.

Abstract 17453: In vitro Aspirin Responsiveness Can Predict Platelet Hyperreactivity and Cardiovascular Risk

Background: While antiplatelet therapy decreases cardiovascular events, insufficient platelet aggregation suppression by antiplatelet therapy is associated with an increased risk of cardiovascular events. In this study, we developed a diagnostic tool to establish an individual’s “aspirin (ASA) responsiveness” in vitro and investigated its association with other metrics of platelet activity and cardiovascular risk. Methods: In 115 subjects off all antiplatelet therapy, platelet aggregation in response to high-dose of arachidonic acid (AA; 1.5 mM) with and without incubation of platelet-rich plasma with ASA (3 mM) was measured by light transmission aggregometry (LTA). ASA responsiveness was defined as ([% aggregation to AA - % aggregation to AA + ASA] / % aggregation to AA) x 100. Next, we correlated ASA responsiveness with other metrics of platelet activity. Finally, we evaluated its clinical significance in a cohort of patients undergoing lower extremity revascularization (LER) followed longitudinally for cardiovascular events. Results: Among 115 participants (mean age 50.3 years old, 65% female, and 43.5% underrepresented minorities, off all antiplatelet therapy), mean % aggregation to high-dose AA was 89.5% ± 3.5. In vitro ASA responsiveness was variable, mean 73.72% ± 15 (range: 13.7% to 97.6%). ASA responsiveness was lower in older (P<0.05) and female participants (P<0.05) and was not associated with race/ethnicity, diabetes, or platelet count. Lower ASA responsiveness was associated with a significant increase in platelet aggregation to epinephrine (0.4 μM, P=0.0004), ADP (1.0 μM, P=8.89x10 -9 ), and collagen (0.2 μg/mL, P=1.16x10 -9 ). In an independent cohort of 67 patients undergoing LER, lower in vitro ASA responsiveness (e.g. a reduced platelet-inhibiting effect), was associated with a significantly increased risk of major adverse cardiac and limb events (1 st tertile-63.6%, 2 nd tertile-50.0% and 3 rd tertile-27.3%; P=0.016 for trend). Conclusions: Reduced ASA responsiveness is associated with increased platelet activity and a higher risk of cardiovascular events. We propose that this rapid, relatively simple assay may be used preclinically for ASA testing and to identify individuals at high cardiovascular risk.

Abstract 18621: Platelet Aggregation is Heightened in Those With Metabolic Syndrome and Less Effectively Mediated by Aspirin Therapy

Background: Patients with metabolic syndrome (MetS) are at a higher risk of developing cardiovascular disease (CVD); however, interventions to mitigate this risk are not fully known. Platelets play a significant role in atherothrombosis, and aspirin is an important, yet controversial therapy used for the primary prevention of CVD. In this study, we examined platelet phenotype in MetS and degree of inhibition from ex vivo aspirin administration. Methods: In a study examining the mechanisms of atherothrombosis in pro-inflammatory conditions, patients without clinical CVD were enrolled and underwent phenotyping. Patients with MetS (n = 34, age 53.3 ± 13 years, 59% male) were compared to patients with no MetS (n = 63, age 41.1 ± 16 years, 52% male). Platelet aggregation of freshly isolated platelets was measured via light transmission aggregometry (LTA) in resting and stimulated conditions, in response to agonists adenosine diphosphate (ADP), high and low dose of arachidonic acid (AA), and after ex vivo aspirin (ASA) administration. Results: MetS patients were older and similar in sex and race (Figure 1A). As expected, patients with MetS had higher individual components of MetS, yet similar LDL-C (Figure 1A). Platelet aggregation to low dose ADP (p=0.09) and AA (p=0.04) was significantly increased in MetS (Figure 1B). Platelet aggregation in response to AA and coincubation with ASA was significantly lower in the no MetS cohort (Figure 1C). The efficacy of aspirin therapy was significantly lower in MetS (Figure 1D). In response to high dose AA after ASA treatment, platelet aggregation increased as number of MetS parameters also increased (Figure 1E). However, no individual metabolic syndrome parameter appeared to be driving these findings (Figure 1E). Conclusion: Among MetS patients, platelet aggregation is heightened and less inhibited by aspirin than those with no MetS. Additional research is warranted to explore possible anti-platelet strategies in patients with MetS.

Abstract 15758: Platelet Aggregation is Elevated in Psoriasis and Associated With Cardiovascular Risk

Introduction: Psoriasis is associated with increased CV risk the mechanisms and prevention of which are not fully known. We evaluated the platelet phenotype (implicated in atherothrombosis) and in vitro response to aspirin in patients with psoriasis and its association with vascular stiffness and coronary atherosclerosis. Methods: Patients with psoriasis (n = 64, age 47.9 ± 15 years, 65% male), percent body surface area (BSA) of psoriasis 3%, were compared to controls (n = 40, age 40.8 ± 16 years, 45% male). Percent platelet aggregation in response to agonists adenosine diphosphate (ADP), collagen (Col), and arachidonic acid (AA - with and without aspirin) was measured in freshly isolated platelet-rich plasma via light transmission aggregometry (LTA). Vascular stiffness (via pulse wave velocity [PWV]) and CT coronary angiography were each performed as readouts of CV risk. Results: Compared to controls, psoriasis patients were older, of Caucasian ethnicity (Figure 1A), and overweight, but otherwise similar. Platelet aggregation after treatment with Col (p=0.01), ADP (p < 0.01), and AA (p = 0.05) were all higher in psoriasis vs. control (Figure 1B-D). Platelet aggregation after activation with high dose AA was similar between groups (Figure 1E) however co-incubation with high dose AA plus aspirin displayed incomplete platelet inhibition of psoriasis platelets vs. controls (Figure 1F). Platelet aggregation to low dose AA and high dose AA plus aspirin co-incubation were each higher in those with enhanced vascular stiffness (figure 1G/1H) and the presence of coronary plaque (Figure 1I/1J). Conclusion: Platelet aggregation is consistently elevated in psoriasis across a variety of agonists, incompletely inhibited by aspirin, and associated with biomarkers of CV risk such as vascular stiffness and the presence of coronary plaque. These findings highlight the need to evaluate various anti-platelet strategies to reduce atherothrombotic risk in patients with psoriasis.

Abstract 18385: Platelet Transcriptome in Type 2 Diabetes Provides Insight Into Peripheral Artery Disease and Cardiovascular Events

Background: Platelets are integral to atherothrombosis and peripheral artery disease (PAD), processes that are accelerated in diabetes. We evaluated the platelet transcriptome, generated a platelet RNA score, and investigated whether it could serve as a novel biomarker of PAD severity and major adverse cardiovascular events (MACE). Methods: We performed RNA sequencing on circulating platelet samples obtained from 133 patients enrolled in the platelet activity and cardiovascular events in PAD study. The transcriptome was (1) compared in PAD patients with Type 2 Diabetes (T2D) (n=73) vs without T2D (n=60), and (2) correlated with platelet aggregation responses to submaximal epinephrine (0.4uM). Using the overlap of transcripts identified by these approaches, we scored each sample based on a rank-based phenotypic metric (Singscore) to develop a T2D-specific platelet RNA score. This platelet score was compared in PAD patients with (vs without) critical limb ischemia (CLI), and PAD patients with (vs without) incident MACE (myocardial infarction, stroke, or death) after multivariable adjustment. Results: Overall, 384 transcripts were differentially expressed in PAD patients with vs without T2D (P<0.01; 195 upregulated and 189 downregulated). Gene set enrichment analysis noted upregulation of pathways related to cell chemotaxis, acute inflammatory response, and platelet alpha granules in T2D. Among these 384 genes, 102 significantly correlated with platelet aggregation (P<0.01) and were used to create a T2D platelet RNA score. To evaluate its clinical significance, this score was 3- times higher in PAD with (vs without) CLI (P=0.002). Further, after adjusting for age, sex, race, T2D, hypertension and coronary artery disease, the platelet score above (vs below) the median was associated with increased MACE (HR 2.8, 95% CI 1.4 to 5.6, P = 0.003). Conclusions: These findings demonstrate a unique platelet transcriptome in T2D that is associated with PAD severity and future risk of cardiovascular events. This work provides an important step toward the use of platelet RNA to unlock novel molecular mechanisms and inform future clinical outcomes.

PB0867 Alpha-2-Macroglobulin Precludes Platelet Aggregation in Response to the Brain-Derived Neurotrophic Factor (BDNF)

PB0867 Alpha-2-Macroglobulin Precludes Platelet Aggregation in Response to the Brain-Derived Neurotrophic Factor (BDNF)

No impact of polymorphism in the phosphodiesterase 5A gene in Cavalier King Charles Spaniels on pimobendan-induced inhibition of platelet aggregation response.

A variant in the canine phosphodiesterase (PDE) 5A gene (PDE5A:E90K) is associated with decreased concentrations of circulating cyclic guanosine monophosphate (cGMP) and response to PDE5 inhibitor treatment. Pimobendan is a PDE inhibitor recommended for medical treatment of certain stages of myxomatous mitral valve disease (MMVD) in dogs. PDE5A:E90K polymorphism attenuates the inhibitory effect of pimobendan on in vitro platelet aggregation and increases basal platelet aggregation in Cavalier King Charles Spaniels (CKCS). Selected clinical variables (MMVD severity, sex, age, hematocrit, platelet count in platelet-rich plasma [PRP], and echocardiographic left ventricular fractional shortening [LV FS]) will not show an association with results. Fifty-two privately owned CKCS with no or preclinical MMVD. Using blood samples, we prospectively assessed PDE5A genotype using Sanger sequencing and adenosine diphosphate-induced platelet aggregation response (area under the curve [AUC], maximal aggregation [MaxA], and velocity [Vel]) with and without pimobendan using light transmission aggregometry. Dogs also underwent echocardiography. Pimobendan inhibited platelet function as measured by AUC, MaxA, and Vel at a concentration of 10 μM (P < .0001) and Vel at 0.03 μM (P < .001). PDE5A:E90K polymorphism did not influence the inhibitory effect of pimobendan or basal platelet aggregation response. The PDE5A:E90K polymorphism did not influence in vitro basal platelet aggregation response or the inhibitory effect of pimobendan on platelet aggregation in CKCS. Dogs with the PDE5A:E90K polymorphism did not appear to have altered platelet function or response to pimobendan treatment.

Platelet Reactivity on ECMO: Role of VerifyNow

Viscoelastic assays help in identifying disorders of clotting factors, platelet function, and detection of excessive fibrinolysis. Rotational thromboelastometry (ROTEM) assesses quantitative and qualitative aspects of whole blood coagulation which allows for the determination of the mechanism of hemostatic disturbance. This helps clinicians in targeting the correct blood component therapy transfusion. Now, using verify we aim to study platelet reactivity, VA ECMO which will reflect the platelet aggregation response to an aggregating agent like Aspirin and thus know the etiology of bleeding on VA ECMO, to know the right component therapy to be transfused with a Point of Care test. For extracorporeal membrane oxygenation, although the conventional laboratory tests are used to monitor anticoagulation on whole blood, viscoelastic tests reflect coagulation status like no other standard laboratory tests. They show the status of color propagation, formation, and stabilization. It was reported that platelet function was impaired over time in all parameters with different activators.

Liposomal Resveratrol Alleviates Platelet Storage Lesion via Antioxidation and the Physical Buffering Effect.

Platelet transfusion is essential in the treatment of platelet-related diseases and the prevention of bleeding in patients with surgical procedures. Platelet transfusion efficacy and shelf life are limited mainly by the development of platelet storage lesion (PSL). Mitigating PSL is the key to prolonging the platelet shelf life and reducing wastage. Excess intracellular reactive oxygen species (ROS) are one of the main factors causing PSL. In this study, we explored a nanomedicine strategy to improve the quality and functions of platelets in storage. Resveratrol (Res), a natural plant product, is known for its antioxidative effect. However, medical applications of Res are limited due to its low water solubility and stability. Therefore, we used a resveratrol-loaded liposomal system (Res-Lipo) to better utilize the antioxidant effect of the drug. This study aimed to evaluate the effect of Res-Lipo on platelet oxidative stress and alleviation of PSL during the storage time. Res-Lipo scavenged intracellular ROS and inhibited platelet apoptosis and activation during storage. Res-Lipo not only maintained mitochondrial function but also improved platelet aggregation in response to adenosine 5'-diphosphate. These results revealed that Res-Lipo ameliorated PSL and prolonged the platelet survival time in vivo. The strategy provides a potential method for extending the platelet storage time and might be considered a potential and safe additive to alleviate PSL.

Alpha-2-macroglobulin prevents platelet aggregation induced by brain-derived neurotrophic factor

The brain-derived neurotrophic factor (BDNF) has been recently shown to have activating effects in isolated platelets. However, BDNF circulates in plasma and a mechanism to preclude constant activation of platelets appears necessary. Hence, we investigated the mechanism regulating BDNF bioavailability in blood.Protein-protein interactions were predicted by molecular docking and validated through immunoprecipitation. Platelet aggregation was assessed using light transmission aggregometry with washed platelets in response to classical agonists or BDNF, in the absence or presence of alpha-2-macroglobulin (α2M), and in platelet-rich plasma. BDNF signaling was assessed with phospho-blots.As little as 25% autologous plasma was sufficient to completely abolish platelet aggregation in response to BDNF. Docking predicted two forms of BDNF binding to native or activated α2M, in parallel and perpendicular arrangements, and the model suggested that the BDNF-α2M complex cannot bind to the high-affinity BDNF receptor, tropomyosin receptor kinase B (TrkB). Experimentally, native and activated α2M formed stable complexes with BDNF preventing BDNF-induced TrkB activation and signal transduction. Both native and activated α2M inhibited BDNF induced-platelet aggregation in a concentration-dependent manner with comparable half-maximal inhibitory concentrations (IC50≈ 125–150 nM).Our study implicates α2M as a physiological regulator of BDNF bioavailability, and as an inhibitor of BDNF-induced platelet activation in blood.

A correlation study between mean platelet volume and platelet aggregation study in acute coronary syndrome patients.

Platelets play an important role in cardiovascular disease mainly in the development of acute thrombotic events. Elevated platelet indices have been proposed as a risk factor for acute coronary syndrome (ACS). It remains uncertain whether increased platelet indices are the result or the cause of ACS. This study aimed to correlate mean platelet volume (MPV) and platelet aggregation response to know the functional status of platelets based on their size. A total of 50 patients with ST-segment elevation ACS (STE-ACS) or non-ST-segment elevation ACS (NSTE-ACS) were included and their MPV was measured and platelet aggregometry was performed. Patients were divided into two groups, patients with MPV ≤9.1 fl as group 1 and those with MPV >9.1 fl as group 2. The mean maximum platelet aggregation response (MMPAR) with ADP, Collagen, and Epinephrine, of both the groups, were compared. MMPAR to ADP, Collagen, and Epinephrine in group 1 was 74.47%, 66.13%, and 72.9%, respectively, and in group 2, 72.94%, 59.97%, and 72.43%, respectively. There was no statistically significant difference in the MMPAR to ADP, Collagen, and Epinephrine among the two groups. Increased MPV does not indicate the platelets are hyperreactive. An increase in MPV may be because of the increased release of immature platelets from bone marrow as there is increased consumption of platelets at the site of thrombus formation in ACS.

Image-based flow simulation of platelet aggregates under different shear rates.

Hemodynamics is crucial for the activation and aggregation of platelets in response to flow-induced shear. In this paper, a novel image-based computational model simulating blood flow through and around platelet aggregates is presented. The microstructure of aggregates was captured by two different modalities of microscopy images of in vitro whole blood perfusion experiments in microfluidic chambers coated with collagen. One set of images captured the geometry of the aggregate outline, while the other employed platelet labelling to infer the internal density. The platelet aggregates were modelled as a porous medium, the permeability of which was calculated with the Kozeny-Carman equation. The computational model was subsequently applied to study hemodynamics inside and around the platelet aggregates. The blood flow velocity, shear stress and kinetic force exerted on the aggregates were investigated and compared under 800 s-1, 1600 s-1 and 4000 s-1 wall shear rates. The advection-diffusion balance of agonist transport inside the platelet aggregates was also evaluated by local Péclet number. The findings show that the transport of agonists is not only affected by the shear rate but also significantly influenced by the microstructure of the aggregates. Moreover, large kinetic forces were found at the transition zone from shell to core of the aggregates, which could contribute to identifying the boundary between the shell and the core. The shear rate and the rate of elongation flow were investigated as well. The results imply that the emerging shapes of aggregates are highly correlated to the shear rate and the rate of elongation. The framework provides a way to incorporate the internal microstructure of the aggregates into the computational model and yields a better understanding of the hemodynamics and physiology of platelet aggregates, hence laying the foundation for predicting aggregation and deformation under different flow conditions.

Pharmacological Studies on the Ca2+ Influx Pathways in Platelet-Activating Factor (PAF)-Induced Mouse Urinary Bladder Smooth Muscle Contraction.

Platelet-activating factor (PAF) not only acts as a mediator of platelet aggregation, inflammation, and allergy responses but also as a constrictor of various smooth muscle (SM) tissues, including gastrointestinal, tracheal/bronchial, and pregnancy uterine SMs. Previously, we reported that PAF induces basal tension increase (BTI) and oscillatory contraction (OC) in mouse urinary bladder SM (UBSM). In this study, we examined the Ca2+ influx pathways involved in PAF-induced BTI and OC in the mouse UBSM. PAF (10-6 M) induced BTI and OC in mouse UBSM. However, the PAF-induced BTI and OC were completely suppressed by extracellular Ca2+ removal. PAF-induced BTI and OC frequencies were markedly suppressed by voltage-dependent Ca2+ channel (VDCC) inhibitors (verapamil (10-5 M), diltiazem (10-5 M), and nifedipine (10-7 M)). However, these VDCC inhibitors had a minor effect on the PAF-induced OC amplitude. The PAF-induced OC amplitude in the presence of verapamil (10-5 M) was strongly suppressed by SKF-96365 (3 × 10-5 M), an inhibitor of receptor-operated Ca2+ channel (ROCC) and store-operated Ca2+ channel (SOCC), but not by LOE-908 (3 × 10-5 M) (an inhibitor of ROCC). Overall, PAF-induced BTI and OC in mouse UBSM depend on Ca2+ influx and the main Ca2+ influx pathways in PAF-induced BTI and OC may be VDCC and SOCC. Of note, VDCC may be involved in PAF-induced BTI and OC frequency, and SOCC might be involved in PAF-induced OC amplitude.

Emergency management of patients with Glanzmann thrombasthenia: consensus recommendations from the French reference center for inherited platelet disorders

Glanzmann thrombasthenia (GT) is a genetic bleeding disorder characterised by severely reduced/absent platelet aggregation in response to multiple physiological agonists. The severity of bleeding in GT varies markedly, as does the emergency situations and complications encountered in patients. A number of emergency situations may occur in the context of GT, including spontaneous or provoked bleeding, such as surgery or childbirth. While general management principles apply in each of these settings, specific considerations are essential for the management of GT to avoid escalating minor bleeding events. These recommendations have been developed from a literature review and consensus from experts of the French Network for Inherited Platelet Disorders, the French Society of Emergency Medicine, representatives of patients’ associations, and Orphanet to aid decision making and optimise clinical care by non-GT expert health professionals who encounter emergency situations in patients with GT.

MAGT1 Deficiency Dysregulates Platelet Cation Homeostasis and Accelerates Arterial Thrombosis and Ischemic Stroke in Mice.

MAGT1 (magnesium transporter 1) is a subunit of the oligosaccharide protein complex with thiol-disulfide oxidoreductase activity, supporting the process of N-glycosylation. MAGT1 deficiency was detected in human patients with X-linked immunodeficiency with magnesium defect syndrome and congenital disorders of glycosylation, resulting in decreased cation responses in lymphocytes, thereby inhibiting the immune response against viral infections. Curative hematopoietic stem cell transplantation of patients with X-linked immunodeficiency with magnesium defect causes fatal bleeding and thrombotic complications. We studied the role of MAGT1 deficiency in platelet function in relation to arterial thrombosis and hemostasis using several in vitro experimental settings and in vivo models of arterial thrombosis and transient middle cerebral artery occlusion model of ischemic stroke. MAGT1-deficient mice (Magt1-/y) displayed accelerated occlusive arterial thrombus formation in vivo, a shortened bleeding time, and profound brain damage upon focal cerebral ischemia. These defects resulted in increased calcium influx and enhanced second wave mediator release, which further reinforced platelet reactivity and aggregation responses. Supplementation of MgCl2 or pharmacological blockade of TRPC6 (transient receptor potential cation channel, subfamily C, member 6) channels, but not inhibition of store-operated calcium entry, normalized the aggregation responses of Magt1-/y platelets to the control level. GP (glycoprotein) VI activation of Magt1-/y platelets resulted in hyperphosphorylation of Syk (spleen tyrosine kinase), LAT (linker for activation of T cells), and PLC (phospholipase C) γ2, whereas the inhibitory loop regulated by PKC (protein kinase C) was impaired. A hyperaggregation response to the GP VI agonist was confirmed in human platelets isolated from a MAGT1-deficient (X-linked immunodeficiency with magnesium defect) patient. Haploinsufficiency of TRPC6 in Magt1-/y mice could normalize GP VI signaling, platelet aggregation, and thrombus formation in vivo. These results suggest that MAGT1 and TRPC6 are functionally linked. Therefore, deficiency or impaired functionality of MAGT1 could be a potential risk factor for arterial thrombosis and stroke.