Search Results (45)

Background: Given the pivotal role of diet in cardiovascular diseases (CVDs), there is a growing demand for new sources of bioactive phytochemicals that can contribute to CVD prevention and treatment. Previous research has unveiled the cardioprotective properties of several parts of sea buckthorn (Hippophae rhamnoides L.). For example, various fractions isolated from raw and roasted sea buckthorn seeds showed antioxidant properties in vitro. In addition, the serotonin-rich fraction obtained from roasted seed extract had the strongest antioxidant activity. However, it was unclear which chemical constituents contribute to the anti-platelet potential of sea buckthorn seeds. Methods: The anti-platelet activity of two fractions (fraction b and fraction c) from raw sea buckthorn seed extract, two fractions (fraction d and fraction g) from roasted sea buckthorn seed extract, and two chemical compounds—isorhamnetin 3-O-β-glucoside-7-O-α-rhamnoside (a major component of fraction b), and serotonin (5-HT, 5-hydroxytryptamine), present in fraction c was estimated in several in vitro assays. Results: Isorhamnetin 3-O-β-glucoside-7-O-α-rhamnoside significantly inhibited platelet activation. It lowered the exposition of the active form of GPIIb/IIIa on the surface of 20 μM ADP-stimulated platelets by about 26%. It also inhibited the exposition of P-selectin on the surface of 10 and 20 μM ADP-stimulated platelets. In addition, isorhamnetin 3-O-β-glucoside-7-O-α-rhamnoside (at 50 µg/mL) significantly prolonged the time of thrombus formation. The results also indicate that fractions d and g (from roasted seeds) are more effective anti-adhesive factors than fractions from raw sea buckthorn seeds. Conclusions: It can be suggested that sea buckthorn seeds can serve as a new source of anti-platelet compounds (especially derivatives of isorhamnetin) beneficial in CVD prevention and treatment; however, in vivo research is needed to clarify their mechanism of action, physiologically relevant concentrations, and therapeutic potential. Full article

Apolipoprotein C3 (apoC3) is a key regulator of triglyceride metabolism and has emerged as a potential therapeutic target for reducing the risk of cardiovascular disease. However, its broader physiological functions are not fully understood. This study investigates the role of apoC3 in platelet function and thrombus formation. Interestingly, human apoC3 was found to rapidly inhibit platelet activation over the tested concentration range of 0.1–10 µg/mL, with significant effects observed at low concentrations and brief pre-incubation times (from 1 min). At a concentration of 10 µg/mL, apoC3 suppressed platelet activation by approximately 70% in response to ADP and by approximately 40% in response to collagen stimulation. Depleting apoC3 from human serum enhanced platelet aggregation by more than 25 % (1.28 ± 0.19 vs. vehicle), indicating an endogenous regulatory function of apoC3. Mechanistically, apoC3 binding to platelets reduced both GPIIb/IIIa activation and P-selectin expression by around 20%. ApoC3 binding to platelets increased when platelets were activated by ADP and was partially mediated by GPIIb/IIIa, implicating this integrin as a functionally relevant receptor. Taken together, these findings reveal a novel link between apoC3 and platelet biology with potential implications for thrombotic risk and vascular homeostasis. Full article

Background/Objectives: Platelet counts can be affected by storage conditions, potentially leading to pseudothrombocytopenia. The present study aimed to investigate temperature-dependent changes in platelet counts and morphology in whole blood samples anticoagulated with heparin or EDTA. We also examined the molecular mechanism of cold-induced aggregation via integrin GPIIb/IIIa–fibrinogen interaction using established bioinformatics technologies (docking simulation). Methods: Peripheral blood was collected from healthy volunteers (n = 6) and treated with either heparin or EDTA. The samples were stored at 4 °C, room temperature, or incubated at 37 °C. Platelet counts were measured using an automated hematology analyzer. The morphology of various blood cells in smears was assessed using the May-Grünwald Giemsa staining method. Docking simulations using an available software (HADDOCK 2.4) were performed to evaluate integrin–fibrinogen binding at different temperatures. Results: In automated blood cell counting, platelet counts in heparinized blood were significantly decreased under low-temperature conditions (4 °C), but this decrease was restored to levels comparable to those at room temperature upon warming to 37 °C (p < 0.05). No significant changes were observed in EDTA-treated samples. Microscopical findings showed platelet aggregation only in heparinized samples at 4 °C, with normal morphology restored upon warming (37 °C). Docking simulations estimated stronger integrin GPIIb/IIIa–fibrinogen binding at 4 °C than at 37 °C (p = 0.0286), suggesting temperature-dependent enhancement of molecular interactions. Conclusions: These findings indicate that heparin can induce reversible platelet aggregation at low temperatures in whole blood samples, leading to pseudothrombocytopenia. This phenomenon may be mediated by increased integrin GPIIb/IIIa–fibrinogen binding. Full article

In healthy conditions and cardiovascular diseases, the most abundant microparticles (MPs) in the bloodstream are those of platelet origin, but the direct effect of these microparticles on endothelial activation is poorly understood. The objective of this paper is to measure endothelial cell activation, as evaluated by the expression of the adhesion molecules E-selectin, VCAM-1, ICAM-1, and PECAM-1 in endothelial cell line HMEC-1 when stimulated with MPs produced by platelets stimulated in vitro with thrombin (TH), adenosine diphosphate (ADP), calcium ionophore (ICa), N-acetylglucosamine (NAcGlc), and without any stimulus. Platelets from healthy individuals induced the formation of MPs with different agonists. The results from the determination of the phenotype of the MPs showed that the expression of GPIIb/IIIa was significant, with median fold changes of TH = 2.2, ADP = 5.2, Ica = 7.0, and NAcGlc = 10.0. However, in HMEC-1 cells, the expression of adhesion molecules stimulated with MPs had a median change slightly higher for E-Sel expression (ranging from 1.4 to 4.2) and ICAM-1 expression (range 2.2 to 3.0), especially VCAM-1 expression (ranging from 15 to 18.8), all of which were significant. For PECAM-1, only stimulation with ICa (1.5) was significant, demonstrating that MPs elicit stimulus-dependent responses in endothelial cells. Platelet-derived MPs may have a potential role in modulating inflammation and other endothelial functions. Full article

Platelet satellitism (PS) is an in vitro phenomenon of platelets adhering around white blood cells, especially in blood samples anticoagulated with K₃EDTA. This, in some cases, can lead to spurious thrombocytopenia, without platelet dysfunction or bleeding events. Diagnosis is made by peripheral blood smear examination. The potential mechanism for PS remains largely unknown; however, it possibly involves the formation of IgG antibodies against the platelet glycoprotein receptor IIb/IIIa (GPIIb/IIIa). PS has been observed in various medical conditions, including infectious, autoimmune, and lymphoproliferative disorders, without an obvious causative relationship. Here, we describe a case of PS in a patient who presented with infection in the setting of underlying Immune Thombocytopenic Purpura and Multiple Sclerosis. Full article

Inherited platelet disorders (IPDs) are rare bleeding disorders characterized by impaired platelet function and/or reduced blood platelet count. Their diagnosis typically relies on complex laboratory methods, including flow cytometry, aggregometry, and molecular genetic analysis. In recent years, immunofluorescence microscopy has been established as an alternative diagnostic method for IPDs. Background/Objectives: This study aims to validate a quantitative approach enhancing reproducibility through automated image analysis for diagnosing IPDs using immunofluorescence microscopy, with Bernard–Soulier Syndrome (BSS) and Glanzmann thrombasthenia (GT) as model IPDs. Methods: Native blood smears from patients with suspected BSS or GT were stained using a standardized immunofluorescence protocol targeting platelet surface glycoproteins, granules, and cytoskeletal components. The slides were analyzed using an automated fluorescence microscope, and a rule-based subpopulation analysis was implemented to quantify fluorescence signals. The results were compared to those of a healthy control group, as well as data from flow cytometry and molecular genetic testing. Results: The automated analysis successfully differentiated BSS and GT patients from healthy controls based on distinct fluorescence signal patterns. In BSS samples, CD42b (GPIbα) expression was absent or severely reduced, while GT samples showed a deficiency of CD41/CD61 (GPIIb/IIIa). The platelet size distribution confirmed macrothrombocytopenia in BSS patients. Flow cytometry and molecular genetic testing corroborated these findings, supporting the diagnostic reliability of the automated immunofluorescence microscopy approach. Conclusions: This proof-of-principle study demonstrates that automated quantitative immunofluorescence microscopy is a viable alternative for diagnosing IPDs, offering a standardized, objective, and efficient method, particularly in settings where flow cytometry is not feasible. Full article

The close interplay between thrombotic and immunologic processes plays an important physiological role in the immune defence after tissue injury and has the aim to reduce damage and to prevent the spread of invading pathogens. However, the uncontrolled or exaggerated activation of these processes can lead to pathological thromboinflammation. Thromboinflammation has been shown to worsen the outcome of cardiovascular, autoinflammatory, or even infectious diseases. Imaging of thromboinflammation is difficult because many clinically relevant imaging techniques can only visualize either inflammatory or thrombotic processes. One interesting option for the noninvasive imaging of thromboinflammation is multispectral ¹⁹F magnetic resonance imaging (MRI). Due to the large chemical shift range of the ¹⁹F atoms, it is possible to simultaneously visualize immune cells as well as thrombus components with specific ¹⁹F tracer that have individual spectral ¹⁹F signatures. Of note, the ¹⁹F signal can be easily quantified and a merging of the ¹⁹F datasets with the anatomical ¹H MRI images enables precise anatomical localization. In this review, we briefly summarize the background of ¹⁹F MRI for inflammation imaging, active targeting approaches to visualize thrombi and specific immune cells, introduce studies about multispectral ¹⁹F MRI, and summarize one study that imaged thromboinflammation by multispectral ¹⁹F MRI. Full article

Background: There are no data regarding the outcomes of patients with stent thrombosis (ST) being treated with drug-coated balloon (DCB) angioplasty. Our aim was to compare the outcomes of patients with ST treated with DCB vs. a drug eluting stent (DES). Methods: In this registry analysis, we identified all patients treated for ST in our institution from June 2011 until November 2019. We excluded patients who died in the cath lab, patients with uncrossable lesions, and patients treated with thrombectomy only. Patient outcomes were obtained from Hospital Episodes Statistics from NHS England. The primary endpoint of this study was the composite of cardiovascular mortality, acute coronary syndrome, or target lesion revascularisation. The data were analysed with Cox regression and Kaplan–Meier estimator plots. Results: A total of 173 patients were identified; 92 treated with DCB-only, 36 with balloon angioplasty (BA), 26 with DES, and 19 with a combination of DES and DCB. We compared the outcomes of 92 patients with DCB versus 20 patients with DES, all of which had presented with late or very late ST. There was no difference between DCB and DES in terms of the primary endpoint (p = 0.06). Multivariate analysis identified diabetes (adverse) and the use of GPIIbIIIa inhibitor (favourable) as the only independent predictors of the primary endpoint. Implantation of a DES was independently associated with worse cardiovascular mortality. Conclusions: This is the first study assessing the outcomes of patients with ST treated with DCB only. It has demonstrated that DCBs are an attractive therapeutic option with a tendency towards favourable outcomes when compared to DESs. Full article

Schisandra chinensis Turcz. (Baill.) is a dioecious vine belonging to the Schisandraceae family. Its berries show beneficial activities, including cardioprotective, antioxidant, and anti-inflammatory. We examined the chemical content of S. chinensis berry extract and its antiplatelet potential in vitro. The antiplatelet activity assays included measurements of thrombus formation in full blood (with Total Thrombus-formation Analysis System) and platelet activation and adhesion. We also assessed the extract’s effect on coagulation times in human plasma and its cytotoxicity toward blood platelets based on extracellular lactate dehydrogenase activity. The most important constituents of the extract were dibenzocyclooctadiene lignans; schisandrin was the dominant compound. S. chinensis berry extract at the concentration of 50 μg/mL inhibited thrombus formation by approximately 15%. The adhesion of unstimulated and thrombin-activated blood platelets to collagen was inhibited by all used concentrations of the extract (0.5–50 μg/mL), while the adhesion of adenosine diphosphate (ADP)-activated platelets to fibrinogen was inhibited only by the concentrations of 10 and 50 μg/mL. The extract also inhibited the exposition of the active form of GPIIb/IIIa on the surface of platelets stimulated with 10 μM ADP (at 0.5–50 μg/mL) and 20 μM ADP (at 50 μg/mL). The exposition of P-selectin was inhibited only by the extract at the concentrations of 5–50 μg/mL in platelets stimulated with 10 μg/mL collagen. Moreover, the extract was not cytotoxic toward blood platelets. This indicates that S. chinensis berries hold promise as new antiplatelet agents, but more studies are needed to determine their mechanisms of action and in vivo efficiency. Full article

Background: Platelets have been shown to promote ovarian cancer; however, the mechanism is poorly understood. Previously, we demonstrated that platelets reduce the size and increase the density of multi-cellular ovarian cancer spheroids in cell cultures. The objectives of this study were to determine if platelet inhibitors could counteract these effects, and to explore the mechanisms involved. Methods: FDA-approved platelet inhibitors were screened for their abilities to alter platelet effects on ovarian cancer spheroids. Mass spectrometry was used to identify proteins significantly altered in cancer cells upon exposure to platelets. The effects of platelets and/or liver x receptor agonists or antagonists on LXR activity were measured using ES-2 ovarian cancer cells transduced with an LXR-reporter vector. Results: Eptifibatide, a GPIIB-IIIA integrin inhibitor, and dipyridamole, an adenosine reuptake inhibitor, reduced and enhanced platelet effects on ovarian cancer spheroids, respectively. Proteomic studies identified the LXR/RXR and integrin pathways as mediators of platelet effects on ovarian cancer, and downstream effectors of eptifibatide. Conclusions: Integrin pathways and their downstream LXR/RXR effectors are implicated in how platelets alter ovarian cancer spheroid morphology. These results support studying eptifibatide and LXR/RXR agonists as candidate drugs for repurposing as therapeutic strategies to counteract platelet promotion of ovarian cancer. Full article

The levels of platelet-derived extracellular vesicles (pEVs) have been reported as elevated in acute ischemic stroke (IS). However, the results of studies remain equivocal. This prospective, case-control study included 168 patients with IS, 63 matched disease controls (DC), and 21 healthy controls (HC). Total pEVs concentration, the concentration of phosphatidylserine-positive pEVs (PS⁺pEVs), the percentage of PS⁺pEVs (%PS⁺pEVs) and the concentration of pEVs with expression of CD62P⁺, CD40L⁺, CD31⁺, and active form of GPIIb/IIIa receptor (PAC-1⁺) were assessed on days 1, 3, 10, and 90 with the Apogee A50-Micro flow cytometer. The concentrations of pEVs, PS⁺pEVs, and %PS⁺pEVs were significantly higher after IS vs. HC (p < 0.001). PS⁺pEVs were higher after stroke vs. controls (p < 0.01). The concentrations of pEVs with expression of studied molecules were higher on D1 and D3 after stroke vs. controls. The concentration of pEVs after platelet stimulation with ADP was significantly diminished on D3. IS most notably affects the phenotype of pEVs with a limited effect on the number of pEVs. Ischemic stroke moderately disturbs platelet microvesiculation, most notably in the acute phase, affecting the phenotype of pEVs, with a limited impact on the number of pEVs. Full article

Glanzmann’s thrombasthenia (GT) is a rare autosomal recessive disorder of platelet function. The frequent occurrence of alloimmunization due to repeated platelet transfusions is the major complication of the disease. Achieving hemostasis in these patients with anti-GPIIb-IIIa antibodies during surgical procedures is a significant challenge due to the high risk of bleeding. Recombinant activated factor VII (rFVIIa) is an effective agent for achieving hemostasis in alloimmunized Glanzmann’s thrombasthenia patients. The key clinical question was to determine whether abdominal aortic aneurysm surgery can be safely performed with rFVIIa in Glanzmann’s thrombasthenia patients with anti-GPIIb/IIIa antibodies and whether long-term antiplatelet therapy is suitable for these patients. The patient underwent endovascular aneurysm repair with intensive rFVIIa administration, experiencing neither bleeding nor thrombosis. Data regarding the surgical management of Glanzmann’s thrombasthenia patients with anti-GPIIb-IIIa antibodies and the use of antithrombotics in this high-risk population are still very limited. Sharing clinical experience can be valuable for hematologists managing similar cases. Full article

Sea buckthorn (Hippophae rhamnoides L.) is a tree or shrub with small, orange berries. Sea buckthorn seeds have shown many properties beneficial to human health, including antioxidant, anti-hypertensive, anti-hyperlipidemic, and retinoprotective activities. Seeds, as a component of food, are often exposed to high temperatures, which can increase or decrease their biological activity. In our previous study, we showed that both raw and roasted sea buckthorn seeds had significant antioxidant activity, which was measured in human plasma in vitro. In this paper, we evaluated the effect of extracts from raw and roasted sea buckthorn seeds on several parameters of hemostasis in vitro, including thrombus formation in full blood (measured by the Total Thrombus formation Analysis System—T-TAS), blood platelet activation (based on the exposition of P-selectin, the active form of GPIIb/IIIa on their surface and platelet-derived microparticles formation), aggregation (measured with impedance aggregometry), adhesion to fibrinogen and collagen, arachidonic acid metabolism in washed platelets stimulated by thrombin, and COX-1 activity. We also measured the levels of free 8-isoprostane in plasma and the total non-enzymatic antioxidant status of plasma. The extract from roasted seeds (50 µg/mL) significantly prolonged the time of occlusion measured by T-TAS—the AUC₁₀ (area under the curve) value was decreased by approximately 18%. Both extracts decreased the exposition of the active form of GPIIb/IIIa on the surface of platelets activated with 10 μM ADP (by 38.4–62.2%) and 20 μM ADP (by 39.7–51.3%). Moreover, the extract from raw seeds decreased the exposition of P-selectin on the surface of platelets stimulated with 20 μM ADP (by 31.2–34.9%). The adhesion of thrombin-stimulated platelets to fibrinogen and collagen was inhibited only by the extract from roasted sea buckthorn seeds (by 20–30%). Moreover, the extract from raw seeds inhibited the level of TBARS (thiobarbituric acid-reactive substances, an indicator of enzymatic peroxidation of arachidonic acid) in washed platelets stimulated with thrombin; the activity of COX-1 was inhibited by both extracts, although the effect of the extract from raw seeds was stronger. These results indicate that sea buckthorn seeds have anti-platelet activity that is not decreased by thermal processing, but more research is needed to determine which exact chemical compounds and mechanisms are responsible for this phenomenon. Full article

Glanzmann Thrombasthenia (GT) is an inherited platelet disorder caused by defects in platelet integrin α_IIbβ₃ (GPIIb/IIIa), which is a platelet receptor essential for the binding of fibrinogen. This can lead to severe bleeding, especially after trauma or perioperatively, and to microcytic anemia because of chronic blood loss. We report on a 40-year-old female patient with extensive bleeding complications and platelet antibody formation who presented in Homburg and Freiburg for extensive platelet function analyses and molecular genetic analyses. According to platelet aggregometry, the patient had previously been diagnosed with Glanzmann Thrombasthenia (GT). In addition, an MRI scan had been performed due to an unsteady gait and had revealed bilateral para-ophthalmic aneurysms of both internal carotid arteries (ICAs). Assuming a 5% rupture risk per 5 years for each aneurysm, the patient was offered and accepted endovascular treatment. Next-generation sequencing (NGS) panel analysis identified a previously undescribed homozygous one-base-pair deletion in ITGA2B, which leads to a loss of function of the α_IIb-subunit of the receptor. This case illustrates the difficulties that can arise regarding the treatment of patients with rare platelet bleeding disorders, and supports the importance of continuous medical care by a specialized hemophilia center for these patients. Full article

Vaccine-induced thrombotic thrombocytopenia (VITT) is a rare but severe complication following COVID-19 vaccination, marked by thrombocytopenia and thrombosis. Analogous to heparin-induced thrombocytopenia (HIT), VITT shares similarities in anti-platelet factor 4 (PF4) IgG-mediated platelet activation via the FcγRIIa. To investigate the involvement of platelet-antibodies in VITT, we analyzed the presence of platelet-antibodies directed against glycoproteins (GP)IIb/IIIa, GPV and GPIb/IX in the serum of 232 clinically suspected VITT patients determined based on (suspicion of) occurrence of thrombocytopenia and/or thrombosis in relation to COVID-19 vaccination. We found that 19% of clinically suspected VITT patients tested positive for anti-platelet GPs: 39%, 32% and 86% patients tested positive for GPIIb/IIIa, GPV and GPIb/IX, respectively. No HIT-like VITT patients (with thrombocytopenia and thrombosis) tested positive for platelet-antibodies. Therefore, it seems unlikely that platelet-antibodies play a role in HIT-like anti-PF4-mediated VITT. Platelet-antibodies were predominantly associated with the occurrence of thrombocytopenia. We found no association between the type of vaccination (adenoviral vector vaccine versus mRNA vaccine) or different vaccines (ChAdOx1 nCoV-19, Ad26.COV2.S, mRNA-1273, BTN162b2) and the development of platelet-antibodies. It is essential to conduct more research on the pathophysiology of VITT, to improve diagnostic approaches and identify preventive and therapeutic strategies. Full article