Role In Hemostasis Research Articles

Platelets’ Functionality in Hemostasis, Inflammation, and Immune Response to Infections

Platelets play a central role in hemostasis and are increasingly recognized for their critical functions in immune responses and inflammation. This review explores the mechanisms by which platelets contribute to these processes, from their involvement in clot formation and wound healing to their emerging role in modulating immune cell activity during infections. The interaction between platelets and the inflammatory cascade creates a complex inflammation-hemostasis loop, with implications for both acute and chronic disease states. Platelets aggregate at the site of tissue damage, and adhere to the white blood cells, with subsequent release of cytokines and chemokines which are required for targeting lymphocytes, monocytes, and neutrophils to promote inflammation. Similarly, platelets engulf microbes and affect the prognosis of bacterial and viral infections. Recent advances in understanding platelet function during bacterial and viral infections are discussed, with a focus on the therapeutic potential of targeting platelet-mediated pathways in various pathological conditions.

Small but mighty: Platelets as multifunctional architects of tumor metastasis and immune regulation

AbstractPlatelets play an irreplaceable role in hemostasis and wound healing. However, beyond these classical roles, as the smallest anucleate cells in the blood stream, they are crucial for immune response which have inflammatory functions through specialized receptors and different signaling pathways, influencing both innate and adaptive immune response. Furthermore, many research have proved that platelets significantly contribute to tumor metastasis and are associated with poor prognoses in cancer patients through its coagulability and supporting an immunosuppressive tumor microenvironment. When tumor cells detach from the primary tumor mass and enter the bloodstream, they rapidly initiate the direct activation and adhesion of platelets, forming a protective microenvironment. This environment shields circulating tumor cells (CTCs) from the mechanical shear forces of blood flow and immune surveillance. Here we delve into the interaction between platelets and immunomodulation and explore the multifaceted roles and underlying mechanisms by which platelets influence tumor cell metastasis and tumor growth. Furthermore, we also discussed the diagnostic role of platelets in cancer occurrence and progression, as well as the feasibility and prospects of targeting platelets for antitumor immunotherapy. This review provides a multidimensional perspective and reference for platelet‐related cancer treatment strategies and diagnosis.

A Novel P-III Metalloproteinase from Bothrops barnetti Venom Degrades Extracellular Matrix Proteins, Inhibits Platelet Aggregation, and Disrupts Endothelial Cell Adhesion via α5β1 Integrin Receptors to Arginine-Glycine-Aspartic Acid (RGD)-Containing Molecules.

Viperid snake venoms are notably abundant in metalloproteinases (proteins) (SVMPs), which are primarily responsible for inducing hemorrhage and disrupting the hemostatic process and tissue integrity in envenomed victims. In this study, barnettlysin-III (Bar-III), a hemorrhagic P-III SVMP, was purified from the venom of the Peruvian snake Bothrops barnetti. Bar-III has a molecular mass of approximately 50 kDa and is a glycosylation-dependent functional metalloproteinase. Some biochemical properties of Bar-III, including the full amino acid sequence deduced from its cDNA, are reported. Its enzymatic activity is increased by Ca2+ ions and inhibited by an excess of Zn2+. Synthetic metalloproteinase inhibitors and EDTA also inhibit its proteolytic action. Bar-III degrades several plasma and ECM proteins, including fibrin(ogen), fibronectin, laminin, and nidogen. Platelets play a key role in hemostasis and thrombosis and in other biological process, such as inflammation and immunity, and platelet activation is driven by the platelet signaling receptors, glycoprotein (GP)Ib-IX-V, which binds vWF, and GPVI, which binds collagen. Moreover, Bar-III inhibits vWF- and convulxin-induced platelet aggregation in human washed platelets by cleaving the recombinant A1 domain of vWF and GPVI into a soluble ectodomain fraction of ~55 kDa (sGPVI). Bar-III does not reduce the viability of cultured endothelial cells; however, it interferes with the adhesion of these cells to fibronectin, vitronectin, and RGD peptides, as well as their migration profile. Bar-III binds specifically to the surface of these cells, and part of this interaction involves α5β1 integrin receptors. These results contribute to a better comprehension of the pathophysiology of snakebite accidents/incidents and could be used as a tool to explore novel and safer anti-venom therapeutics.

Lipopolysaccharide from Proteus mirabilis Slows Platelet Plug Formation in Human Whole Blood

Platelets are well known for their role in hemostasis. Additionally, platelets play a crucial role in immune and inflammatory responses. Toll-like receptors (TLRs) can mediate bacterial interactions during infection, triggering platelets to initiate an inflammatory response. TLR-4 receptors enable direct interactions between platelets and the bacterial lipopolysaccharide (LPS) endotoxin. The aim of this study was to assess platelet plug formation in response to LPS from Proteus mirabilis. Human whole blood was treated with varying concentrations of LPS over a range of incubation times. Then, platelet plug formation time was measured, under high shear conditions using the platelet function analyzer PFA-100, as aperture closure time (CT). The addition of either 2 or 10 µg/mL of LPS to 80% whole blood significantly prolonged the CTs even in the absence of preincubation (p = 0.028 or p = 0.049, respectively). With added preincubation of LPS with whole blood, the measured CTs were further prolonged. If the preincubation time was set to 35 min, then even the addition of 0.2 µg/mL of LPS resulted in significant CT prolongation (p < 0.001). Taken together, the platelet plug formation in the presence of collagen/ADP is significantly prolonged by the presence of LPS in a concentration and preincubation time-dependent manner. Exposure to P. mirabilis LPS reduces the platelet aggregation response in human whole blood.

Identification and Characterization of Candidate Platelet Proteins as Potential Biomarkers for Alcoholism Patients

ABSTRACT Chronic alcoholism is a significant health issue associated with severe liver complications, including alcohol-related liver disease (ARLD), progresses from fatty liver to cirrhosis and hepatocellular carcinoma. ARLD affects liver function and hemostasis, leading to increased mortality. This study intended to investigate the platelet proteome and detect platelet cell apoptosis via Annexin V in ARLD patients in order to identify potential biomarkers for disease progression. Twenty-three patients with ARLD were recruited, and their platelet proteins were analyzed via mass spectrometry. Approximately 441 proteins were identified, with 73 candidate biomarkers selected on the basis of clinical importance. Key proteins involved in coagulation, inflammation, and cellular interactions are highlighted. Gene ontology and STRING analyses revealed functional clusters and pathways associated with these proteins. This study identified critical platelet proteins, such as von Willebrand factor, vitronectin, and serpin family members, which play essential roles in hemostasis and liver disease pathogenesis. These findings suggest that changes in platelet proteomes could serve as noninvasive biomarkers for diagnosing, monitoring, and treating ARLD.

A-144 Liquid chromatography multiple reaction monitoring-based assay for quantitative assessment of fibrinogen levels in clinical samples

Abstract Background Fibrinogen plays a critical role in hemostasis and is essential to clot formation. Changes in the quantity or activity of fibrinogen can lead to disorders affecting coagulation. Thus, it is essential to precisely quantify fibrinogen levels to differentiate between hyperfibrinogenemia in hypercoagulable disorders and dysfibrinogenemia, hypo/afibrinogenemia, and irregular bleeding. Current diagnostic tests that use antibodies to detect fibrinogen or activity-based measures are susceptible to interference from heparin contamination, hemolysis, and hyperbilirubinemia. Here, we evaluated the use of targeted proteomics for developing a quantitative assay for fibrinogen in plasma samples. Methods Four peptides each for alpha and gamma chains of fibrinogen were synthesized using FMOC chemistry on a Liberty Blue (CEM Corp) peptide synthesizer. A targeted multiple reaction monitoring (MRM) assay was developed and optimized using Dionex Ultimate 3000 HPLC connected to a TSQ Altis triple quadrupole mass spectrometer (Thermo Scientific). Evaluations were conducted using a range of standards, such as pure protein, WHO plasma and commercially available plasma with known fibrinogen amounts for the accurate quantitation of fibrinogen. Calibration curves were generated using different dilutions in various matrices. The repeatability and precision of the assay were evaluated across five different days. The plasma samples were processed and digested in a 96-well plate and subjected to C18 clean-up using a Bravo automated liquid handler (Agilent). All samples were spiked-in with corresponding heavy-labeled peptides prior to digestion with trypsin. Multiple reaction monitoring assays were performed on tryptic digests of plasma samples. Results Calibration curves using commercial plasma for all peptides showed linearity from 20 mg/dl to 1,308 mg/dl. This covers the entire clinical range including normal (200-350 mg/dl), deficient states (<100 mg/dl) and excess production (>350 mg/dl) that can be used for evaluation of bleeding disorders including consumption coagulopathy and hyperfibrinogenemia. Based on the overall performance in terms of linearity, limit of detection/quantitation, carryover and coefficient of variation (CV) across five different days, two peptides per chain were finally selected for developing the quantitative assay. The intra-assay variability was <5% and intra-day CV was ∼20%. This assay is currently being tested on a variety of clinical samples representing a range of fibrinogen levels. Conclusions Our study focused on assessing whether multiplexed MRM assay could be implemented to measure fibrinogen levels in plasma. Our data indicate the feasibility of MRM-based assays for determining fibrinogen levels as an alternative to existing non-mass spectrometry-based activity or antigen assays.

The role of factor XIII in patient blood management.

Factor XIII (FXIII), a plasma transglutaminase, is a coagulation factor that plays a crucial role in blood clotting and patient blood management. The studies have demonstrated that FXIII targets a wide range of additional substrates that have an important role in hemostasis, especially in posttraumatic patients, patients undergoing surgery or obstetrics, being involved in wound healing and tissue repair. Morover, FXIII deficiency has also been described and an extensive research has shown that FXIII deficiency is a rare coagulopathy that leads to longer bleeding time, perioperative and postoperative complications and slower wound healing. Present article aims to overview the diverse functions of FXIII and to highlight its role in patient blood management.

Beyond basic characterization and omics: Immunomodulatory roles of platelet-derived extracellular vesicles unveiled by functional testing.

Renowned for their role in haemostasis and thrombosis, platelets are also increasingly recognized for their contribution in innate immunity, immunothrombosis and inflammatory diseases. Platelets express a wide range of receptors, which allows them to reach a variety of activation endpoints and grants them immunomodulatory functions. Activated platelets release extracellular vesicles (PEVs), whose formation and molecular cargo has been shown to depend on receptor-mediated activation and environmental cues. This study compared the immunomodulatory profiles of PEVs generated via activation of platelets by different receptors, glycoprotein VI, C-type lectin-like receptor 2 and combining all thrombin-collagen receptors. Functional assays in vivo in zebrafish and in vitro in human macrophages highlighted distinct homing and secretory responses triggered by the PEVs. In contrast, omics analyses of protein and miRNA cargo combined with physicochemical particle characterization found only subtle differences between the activated PEV types, which were insufficient to predict their different immunomodulatory functions. In contrast, constitutively released PEVs, formed in the absence of an exogenous activator, displayed a distinct immunomodulatory profile from the receptor-induced PEVs. Our findings underscore that PEVs are tunable through receptor-mediated activation. To truly comprehend their role(s) in mediating platelet functions among immune cells, conducting functional assays is imperative.

How Protein Depletion Balances Thrombosis and Bleeding Risk in the Context of Platelet's Activatory and Negative Signaling.

Platelets are small cell fragments that play a crucial role in hemostasis, requiring fast response times and fine signaling pathway regulation. For this regulation, platelets require a balance between two pathway types: the activatory and negative signaling pathways. Activatory signaling mediators are positive responses that enhance stimuli initiated by a receptor in the platelet membrane. Negative signaling regulates and controls the responses downstream of the same receptors to roll back or even avoid spontaneous thrombotic events. Several blood-related pathologies can be observed when these processes are unregulated, such as massive bleeding in activatory signaling inhibition or thrombotic events for negative signaling inhibition. The study of each protein and metabolite in isolation does not help to understand the role of the protein or how it can be contrasted; however, understanding the balance between active and negative signaling could help develop effective therapies to prevent thrombotic events and bleeding disorders.

A highly sensitive and specific fluorescent probe for thrombin detection and high-throughput screening of thrombin inhibitors in complex matrices

Thrombin plays a critical role in hemostasis and hemolysis, and is a significant biomarker for blood-related diseases. Detection and inhibitors screening of thrombin are essential in medical research. In this study, we developed a fluorescent sensor based on the interaction between quantum dots (QDs) and fibrinogen (Fib) for thrombin detection and its inhibitors screening. Upon the presence of thrombin, the fibrinogen of soluble QDs-Fib were converted into insoluble fibrin precipitate, causing a change of fluorescence intensity in the supernatant. Under optimized conditions, our method exhibited an excellent linearity (R2 ≥0.99) over the range of 2∼100 U/L with a limit of detection (LOD) as low as 0.29 U/L. Moreover, we employed this method to screen for thrombin inhibitors using dabigatran as an exemplary direct thrombin inhibitor (DTI), even at concentrations as low as 1 nM. Finally, the established method was successfully used to screen thrombin inhibitors in 23 different extracts from Eupolyphaga sinensis walker. The method provided not only a sensitive, specific and high throughput assay for the detection of thrombin activity in biological samples, but also a reliable strategy for the screening of thrombin inhibitors in complex matrices.

Genetic engineering of transfusable platelets with mRNA-lipid nanoparticles is compatible with blood banking practices

AbstractPlatelets contribute to a variety of physiological processes, including inflammation, sepsis, and cancer. However, because of their primary role in hemostasis, platelet transfusions are largely restricted to managing thrombocytopenia and bleeding. One way to expand the utility of platelet transfusions would be to genetically engineer donor platelets with new or enhanced functions. We have previously shown that lipid nanoparticles containing mRNA (mRNA-LNP) can be used to genetically modify authentic platelets in a nonclinical crystalloid solution. Currently, platelets collected for transfusion are stored in plasma or in plasma supplemented with platelet additive solution (PAS) at supraphysiological concentrations at room temperature, or at 4°C if intended for use in acute hemorrhage. Here, we describe a new plasma-optimized mRNA-LNP for transfecting platelets directly in plasma and plasma supplemented with PAS that is scalable to physiological and supraphysiological platelet concentrations. Transfecting platelets in clinical solutions with mRNA-LNP does not affect aspects of in vitro physiology, and transfected platelets are storable. The compatibility of this transfection system with current clinical practices could enable future mRNA-LNP–based platelet products and cell therapies.

Clinical, Laboratory, and Molecular Aspects of Factor V Deficiency.

Factor V (FV) is a glycoprotein that plays a pivotal role in hemostasis, being involved in coagulant and anticoagulant pathways. Congenital FV deficiency is a rare bleeding disorder with an incidence of 1 per million live births, considering the most severe homozygous form. FV deficiency is diagnosed using routine coagulation tests and FV activity assays. Several mutations, including missense, nonsense, and frameshift, have been detected in the F5 gene. Clinical symptoms are variable, ranging from mild ecchymoses and mucosal bleeding to life-threatening intracranial hemorrhage. The mainstay of treatment includes fresh-frozen plasma, preferentially virus-inactivated. In this narrative review, we provide an update of the main laboratory, molecular, clinical, and therapeutic features of inherited FV deficiency.

Beyond Blood Clotting: The Many Roles of Platelet-Derived Extracellular Vesicles.

Platelet-derived extracellular vesicles (pEVs) are emerging as pivotal players in numerous physiological and pathological processes, extending beyond their traditional roles in hemostasis and thrombosis. As one of the most abundant vesicle types in human blood, pEVs transport a diverse array of bioactive molecules, including growth factors, cytokines, and clotting factors, facilitating crucial intercellular communication, immune regulation, and tissue healing. The unique ability of pEVs to traverse tissue barriers and their biocompatibility position them as promising candidates for targeted drug delivery and regenerative medicine applications. Recent studies have underscored their involvement in cancer progression, viral infections, wound healing, osteoarthritis, sepsis, cardiovascular diseases, rheumatoid arthritis, and atherothrombosis. For instance, pEVs promote tumor progression and metastasis, enhance tissue repair, and contribute to thrombo-inflammation in diseases such as COVID-19. Despite their potential, challenges remain, including the need for standardized isolation techniques and a comprehensive understanding of their mechanisms of action. Current research efforts are focused on leveraging pEVs for innovative anti-cancer treatments, advanced drug delivery systems, regenerative therapies, and as biomarkers for disease diagnosis and monitoring. This review highlights the necessity of overcoming technical hurdles, refining isolation methods, and establishing standardized protocols to fully unlock the therapeutic potential of pEVs. By understanding the diverse functions and applications of pEVs, we can advance their use in clinical settings, ultimately revolutionizing treatment strategies across various medical fields and improving patient outcomes.

Soluble platelet selectin and platelets in COVID-19: a multifaceted connection.

The COVID-19 pandemic has brought to light the intricate relationship between platelets, soluble platelet selectin (sP-selectin), and disease pathogenesis. Platelets, traditionally recognized for their role in hemostasis, have emerged as key contributors to the immunothrombotic complications observed in COVID-19 patients. Concurrently, elevated levels of sP-selectin, indicative of platelet activation and endothelial injury, have been consistently identified in COVID-19 patients and have shown associations with disease severity and adverse outcomes. This multifaceted connection underscores the pivotal role of platelets and sP-selectin in orchestrating thromboinflammation, vascular dysfunction, and disease progression in COVID-19. Platelet activation triggers the release of inflammatory mediators and promotes platelet-leukocyte interactions, amplifying the systemic inflammatory response and exacerbating endothelial injury. Additionally, platelet-derived factors contribute to microvascular thrombosis, further exacerbating tissue damage and organ dysfunction in severe COVID-19. Elevated sP-selectin levels serve as biomarkers for disease severity and prognostication, aiding in risk stratification and early identification of patients at higher risk of adverse outcomes. Therapeutic strategies targeting platelet dysfunction and sP-selectin-mediated pathways hold promise in mitigating thromboinflammation and improving outcomes in COVID-19 patients. Antiplatelet agents, platelet inhibitors, and anti-inflammatory therapies represent potential interventions to attenuate platelet activation, inhibit platelet-leukocyte interactions, and alleviate endothelial dysfunction. A comprehensive understanding of the multifaceted connection between platelets, sP-selectin, and COVID-19 pathogenesis offers opportunities for tailored therapeutic approaches aimed at mitigating thromboinflammation and improving patient outcomes in this complex and challenging clinical setting.

Knowledge, attitudes, and practices of orthopedic operating room personnel regarding the use of pneumatic tourniquets.

Pneumatic tourniquets (PTs) play a crucial role in hemostasis during orthopedic surgery. This study aimed to investigate the current knowledge, attitudes, and practices (KAP) of orthopedic operating room personnel concerning the use of PTs. This cross-sectional study was conducted from Jul. 2023 to Aug. 2023. An online questionnaire was used to collect demographic information and KAP score data of the orthopedic operating room personnel from Hangzhou Regional Hospitals. A total of 336 participants included orthopedic physicians (37.8%), orthopedic nurses (31.5%), anesthesiologists (8.9%), operating room nurses (19.9%) and medical students (1.8%). The median knowledge score was 28.5 (24, 32), with a maximum score of 38; the median attitude score was 31 (28, 35), of a maximum of 35; the median practice score was 41 (36, 44), of a maximum of 45. Correlation analysis showed links between knowledge and attitude (r = 0.388, p<0.001), knowledge and practice (r = 0.401, p<0.001), and attitude and practice (r = 0.485, p<0.001). Multivariate logistic regression analysis confirmed that female gender (OR = 0.294, 95% CI: 0.167-0.520; p<0.001), working in a specialized hospital (OR = 0.420, 95% CI: 0.219-0.803; p = 0.009), and occupation as a surgical anesthesiologist (OR = 3.358, 95% CI: 1.466-7.694; p = 0.004) were associated with better knowledge scores. A higher educational degree (OR = 0.237, 95% CI: 0.093-0.608; p = 0.003) was associated with better practice scores. No previous training was associated with lower knowledge (OR = 0.312, 95% CI: 0.187-0.520; p<0.001) and practice (OR = 0.325, 95% CI: 0.203-0.521; p<0.001) scores. Orthopedic personnel in this study had acceptable knowledge, attitude, and practice concerning the use of PTs; additional training and guidance might enhance proficiency.

An RNA aptamer exploits exosite-dependent allostery to achieve specific inhibition of coagulation factor IXa

Hemostasis relies on a reaction network of serine proteases and their cofactors to form a blood clot. Coagulation factor IXa (protease) plays an essential role in hemostasis as evident from the bleeding disease associated with its absence. RNA aptamers specifically targeting individual coagulation factors have potential as anticoagulants and as probes of the relationship between structure and function. Here, we report X-ray structures of human factor IXa without a ligand bound to the active site either in the apo-form or in complex with an inhibitory aptamer specific for factor IXa. The aptamer binds to an exosite in the catalytic domain and allosterically distorts the active site. Our studies reveal a conformational ensemble of IXa states, wherein large movements of Trp215 near the active site drive functional transitions between the closed (aptamer-bound), latent (apo), and open (substrate-bound) states. The latent state of the apo-enzyme may bear on the uniquely poor catalytic activity of IXa compared to other coagulation proteases. The exosite, to which the aptamer binds, has been implicated in binding VIIIa and heparin, both of which regulate IXa function. Our findings reveal the importance of exosite-driven allosteric modulation of IXa function and new strategies to rebalance hemostasis for therapeutic gain.

P2Y12 Receptor Inhibitor for Antiaggregant Therapies: From Molecular Pathway to Clinical Application.

Platelets play a significant role in hemostasis, forming plugs at sites of vascular injury to limit blood loss. However, if platelet activation is not controlled, it can lead to thrombotic events, such as myocardial infarction and stroke. To prevent this, antiplatelet agents are used in clinical settings to limit platelet activation in patients at risk of arterial thrombotic events. However, their use can be associated with a significant risk of bleeding. An enhanced comprehension of platelet signaling mechanisms should facilitate the identification of safer targets for antiplatelet therapy. Over the past decade, our comprehension of the breadth and intricacy of signaling pathways that orchestrate platelet activation has expanded exponentially. Several recent studies have provided further insight into the regulation of platelet signaling events and identified novel targets against which to develop novel antiplatelet agents. Antiplatelet drugs are essential in managing atherothrombotic vascular disease. The current antiplatelet therapy in clinical practice is limited in terms of safety and efficacy. Novel compounds have been developed in response to patient variability and resistance to aspirin and/or clopidogrel. Recent studies based on randomized controlled trials and systematic reviews have definitively demonstrated the role of antiplatelet therapy in reducing the risk of cardiovascular events. Antiplatelet therapy is the recommended course of action for patients with established atherosclerosis. These studies compared monotherapy with a P2Y12 inhibitor versus aspirin for secondary prevention. However, in patients undergoing percutaneous coronary intervention, it is still unclear whether the efficacy of P2Y12 inhibitor monotherapy after a short course of dual antiplatelet therapy depends on the type of P2Y12 inhibitor. This paper focuses on the advanced-stage evaluation of several promising antiplatelet drugs.

Heat stress–induced platelet dysfunction is associated with loss of fibrinogen and is improved by fibrinogen supplementation

IntroductionHeatstroke is a critical heat-related condition characterized by coagulopathy and multiple organ dysfunction. One of the most severe complications of heatstroke is disseminated intravascular coagulation. This condition manifests as excessive clot formation and bleeding that are primarily due to platelet depletion and dysfunction. Fibrinogen plays a crucial role in hemostasis because it links integrin αIIbβ3 on adjacent platelets, thereby promoting the platelet activation and aggregation necessary for clot formation. However, reduced fibrinogen levels may impair the formation of the initial platelet plug and increase the risk of bleeding. The current study explored the effect of fibrinogen on platelet dysfunction in a heatstroke model. Materials and methodsMale Wistar rats were subjected to heat stress, and subsequent changes in hemodynamic, biochemical, and coagulation parameters were analyzed. Platelet viability, aggregation, adhesion, spreading and fibrin clot retraction were assessed. ResultsThe rats with heatstroke exhibited a variety of clinical symptoms, including hypotension, tachycardia, multiple organ dysfunction, and coagulopathy. Platelet viability in the heatstroke group was comparable to that in the healthy control group. However, the heatstroke group exhibited significant reductions in plasma fibrinogen levels and platelet aggregation, adhesion, spreading, and fibrin clot retraction. Notably, fibrinogen supplementation markedly augmented the aggregation responses of platelets in the heatstroke group. The impairment of platelet adhesion, spreading, and fibrin clot retraction in the rats with heatstroke was partially ameliorated by fibrinogen supplementation. ConclusionsAn early use of fibrinogen replacement may serve as a therapeutic intervention to alleviate platelet hyporeactivity and prevent the complications in patients with heatstroke.

Implications of von Willebrand Factor in Inflammatory Bowel Diseases: Beyond Bleeding and Thrombosis.

Inflammatory bowel disease (IBD) displays an increased venous and arterial thrombotic risk despite the common occurrence of intestinal bleeding. While some of the mechanisms leading to these thrombotic complications have been studied, other specific changes in the hemostasis profile of IBD patients have been less explored. One such example relates to von Willebrand factor (VWF) whose plasma levels have been reported to be modulated in IBD. Von Willebrand factor is a plasma glycoprotein crucial for hemostatic functions via roles both in platelet function and coagulation. High plasma VWF is a known risk factor for venous thromboembolism. In addition to its canonical roles in hemostasis, VWF is known to be directly or indirectly involved in other vascular processes such as maintenance of endothelial barrier integrity or proliferation of vascular smooth muscle cells. The purpose of this review is to recapitulate and update the existing data about VWF biology in IBD and to highlight its role both in the existing procoagulant phenotype and in vascular alterations that may occur in IBD.

Lysophosphatidylcholine induces oxidative stress and calcium-mediated cell death in human blood platelets.

Platelets are essential component of circulation that plays a major role in hemostasis and thrombosis. During activation and its demise, platelets release platelet-derived microvesicles, with lysophosphatidylcholine (LPC) being a prominent component in their lipid composition. LPC, an oxidized low-density lipoprotein, is involved in cellular metabolism, but its higher level is implicated in pathologies like atherosclerosis, diabetes, and inflammatory disorders. Despite this, its impact on platelet function remains relatively unexplored. To address this, we studied LPC's effects on washed human platelets. A multimode plate reader was employed to measure reactive oxygen species and intracellular calcium using H2DCF-DA and Fluo-4-AM, respectively. Flow cytometry was utilized to measure phosphatidylserine expression, mitochondrial membrane potential (ΔΨm), and mitochondrial permeability transition pore (mPTP) formation using FITC-Annexin V, JC-1, and CoCl2/calcein-AM, respectively. Additionally, platelet morphology and its ultrastructure were observed via phase contrast and electron microscopy. Sonoclot and light transmission aggregometry were employed to examine fibrin formation and platelet aggregation, respectively. The findings demonstrate that LPC induced oxidative stress and increased intracellular calcium in platelets, resulting in increased phosphatidylserine expression and reduced ΔΨm. LPC triggered caspase-independent platelet death and mPTP opening via cytosolic and mitochondrial calcium, along with microvesiculation and reduced platelet counts. LPC increased the platelet's size, adopting a balloon-shaped morphology, causing membrane fragmentation and releasing its cellular contents, while inducing a pro-coagulant phenotype with increased fibrin formation and reduced integrin αIIbβ3 activation. Conclusively, this study reveals LPC-induced oxidative stress and calcium-mediated platelet death, necrotic in nature with pro-coagulant properties, potentially impacting inflammation and repair mechanisms during vascular injury.