Hemodynamic regulation of perivalvular endothelial gene expression prevents deep venous thrombosis.
Welsh John D,Hoofnagle Mark H,Bamezai Sharika,Oxendine Michael,Lim Lillian,Hall Joshua D,Yang Jisheng,Schultz Susan,Engel James Douglas,Kume Tsutomu,Oliver Guillermo,Jimenez Juan M,Kahn Mark L
The Journal of clinical investigation
Deep venous thrombosis (DVT) and secondary pulmonary embolism cause approximately 100,000 deaths per year in the United States. Physical immobility is the most significant risk factor for DVT, but a molecular and cellular basis for this link has not been defined. We found that the endothelial cells surrounding the venous valve, where DVTs originate, express high levels of FOXC2 and PROX1, transcription factors known to be activated by oscillatory shear stress. The perivalvular venous endothelial cells exhibited a powerful antithrombotic phenotype characterized by low levels of the prothrombotic proteins vWF, P-selectin, and ICAM1 and high levels of the antithrombotic proteins thrombomodulin (THBD), endothelial protein C receptor (EPCR), and tissue factor pathway inhibitor (TFPI). The perivalvular antithrombotic phenotype was lost following genetic deletion of FOXC2 or femoral artery ligation to reduce venous flow in mice, and at the site of origin of human DVT associated with fatal pulmonary embolism. Oscillatory blood flow was detected at perivalvular sites in human veins following muscular activity, but not in the immobile state or after activation of an intermittent compression device designed to prevent DVT. These findings support a mechanism of DVT pathogenesis in which loss of muscular activity results in loss of oscillatory shear-dependent transcriptional and antithrombotic phenotypes in perivalvular venous endothelial cells, and suggest that prevention of DVT and pulmonary embolism may be improved by mechanical devices specifically designed to restore perivalvular oscillatory flow.
10.1172/JCI124791
PET/MR Imaging of Malondialdehyde-Acetaldehyde Epitopes With a Human Antibody Detects Clinically Relevant Atherothrombosis.
Senders Max L,Que Xuchu,Cho Young Seok,Yeang Calvin,Groenen Hannah,Fay Francois,Calcagno Claudia,Meerwaldt Anu E,Green Simone,Miu Phuong,Lobatto Mark E,Reiner Thomas,Fayad Zahi A,Witztum Joseph L,Mulder Willem J M,Pérez-Medina Carlos,Tsimikas Sotirios
Journal of the American College of Cardiology
BACKGROUND:Oxidation-specific epitopes (OSEs) are proinflammatory, and elevated levels in plasma predict cardiovascular events. OBJECTIVES:The purpose of this study was to develop novel positron emission tomography (PET) probes to noninvasively image OSE-rich lesions. METHODS:An antigen-binding fragment (Fab) antibody library was constructed from human fetal cord blood. After multiple rounds of screening against malondialdehyde-acetaldehyde (MAA) epitopes, the Fab LA25 containing minimal nontemplated insertions in the CDR3 region was identified and characterized. In mice, pharmacokinetics, biodistribution, and plaque specificity studies were performed with Zirconium-89 (Zr)-labeled LA25. In rabbits, Zr-LA25 was used in combination with an integrated clinical PET/magnetic resonance (MR) system. F-fluorodeoxyglucose PET and dynamic contrast-enhanced MR imaging were used to evaluate vessel wall inflammation and plaque neovascularization, respectively. Extensive ex vivo validation was carried out through a combination of gamma counting, near infrared fluorescence, autoradiography, immunohistochemistry, and immunofluorescence. RESULTS:LA25 bound specifically to MAA epitopes in advanced and ruptured human atherosclerotic plaques with accompanying thrombi and in debris from distal protection devices. PET/MR imaging 24 h after injection of Zr-LA25 showed increased uptake in the abdominal aorta of atherosclerotic rabbits compared with nonatherosclerotic control rabbits, confirmed by ex vivo gamma counting and autoradiography. F-fluorodeoxyglucose PET, dynamic contrast-enhanced MR imaging, and near-infrared fluorescence signals were also significantly higher in atherosclerotic rabbit aortas compared with control aortas. Enhanced liver uptake was also noted in atherosclerotic animals, confirmed by the presence of MAA epitopes by immunostaining. CONCLUSIONS:Zr-LA25 is a novel PET radiotracer that may allow noninvasive phenotyping of high-risk OSE-rich lesions.
10.1016/j.jacc.2017.11.036
Next-generation sequencing in the diagnosis of non-cirrhotic splanchnic vein thrombosis.
Magaz Marta,Alvarez-Larrán Alberto,Colomer Dolors,López-Guerra Mónica,García-Criado M Ángeles,Mezzano Gabriel,Belmonte Ernest,Olivas Pol,Soy Guillem,Cervantes Francisco,Darnell Anna,Ferrusquía-Acosta José,Baiges Anna,Turon Fanny,Hernández-Gea Virginia,García-Pagán Juan Carlos
Journal of hepatology
BACKGROUND & AIMS:Myeloproliferative neoplasms (MPNs) are the most frequent cause of non-tumoural non-cirrhotic splanchnic vein thrombosis (NC-SVT). Diagnosis of MPN is based on blood cell count alterations, bone marrow histology, and detection of specific gene mutations. Next-generation sequencing (NGS) allows the simultaneous evaluation of multiple genes implicated in myeloid clonal pathology. The aim of this study was to evaluate the potential role of NGS in elucidating the aetiology of NC-SVT. METHODS:DNA samples from 80 patients (75 with idiopathic or exclusively local factor [Idiop/loc-NC-SVT] and 5 with MPN and NC-SVT [SVT-MPN] negative for Janus kinase 2 gene [JAK2] [V617F and exon 12], calreticulin gene [CALR], and thrombopoietin gene [MPL] mutations by classic techniques) were analysed by NGS. Mutations involved in myeloid disorders different from JAK2, CALR, and MPL genes were categorised as high-molecular-risk (HMR) variants or variants of unknown significance. RESULTS:In 2/5 triple-negative SVT-MPN cases (40%), a mutation in exon 12 of JAK2 was identified. JAK2-exon 12 mutation was also identified in 1/75 patients with Idiop/loc-NC-SVT. Moreover, 28/74 (37.8%) of the remaining Idiop/loc-NC-SVT had at least 1 HMR variant. Sixty-two patients with Idiop/loc-NC-SVT were not receiving long-term anticoagulation and 5 of them (8.1%) had recurrent NC-SVT. This cumulative incidence was significantly higher in patients with HMR variants than in those without. CONCLUSIONS:NGS identified JAK2-exon12 mutations not previously detected by conventional techniques. In addition, NGS detected HMR variants in approximately one-third of patients with Idiop/loc-NC-SVT. These patients seem to have a higher risk of splanchnic rethrombosis. NGS might be a useful diagnostic tool in NC-SVT. LAY SUMMARY:Next-generation sequencing (NGS) performs massive sequencing of DNA allowing the simultaneous evaluation of multiple genes even at very low mutational levels. Application of this technique in a cohort of patients with non-cirrhotic non-tumoral portal vein thrombosis (NC-SVT) and a negative study for thrombophilic disorders was able to identify patients with a mutation in exon 12 not previously detected by conventional techniques. Moreover, NGS detected High Molecular Risk (HMR)-variants (Mutations involved in myeloid disorders different from JAK2, CALR and MPL genes) in approximately one third of patients. These patients appear to be at increased risk of rethrombosis. All these findings supports NGS as a potential useful tool in the management of NC-SVT.
10.1016/j.jhep.2020.06.045
Left Ventricular Thrombus After Acute Myocardial Infarction: Screening, Prevention, and Treatment.
McCarthy Cian P,Vaduganathan Muthiah,McCarthy Killian J,Januzzi James L,Bhatt Deepak L,McEvoy John W
JAMA cardiology
Importance:Left ventricular (LV) thrombus is a complication of acute myocardial infarction (MI) and is associated with systemic thromboembolism. With randomized clinical trials investigating the optimal antithrombotic regimen in patients with MI who require concomitant chronic anticoagulation and with the emergence of the direct-acting oral anticoagulants, treatment options for post-MI LV thrombus have become more complicated. Herein, we review the epidemiology, pathogenesis, diagnosis, prevention, and treatment of LV thrombus after acute MI. Observations:Contemporary epidemiologic data suggest the incidence of LV thrombus, detected using optimal imaging modalities, may be as high as 15% in patients with ST-segment elevation MI and up to 25% in patients with anterior MI. While a standard transthoracic echocardiogram is commonly used for screening, it is limited by low sensitivity for LV thrombus detection, necessitating the addition of contrast (unless contraindicated) and/or use of cardiac magnetic resonance imaging when pretest probability is high. To our knowledge, there are no existing randomized clinical trials evaluating the safety and efficacy of anticoagulation in the prevention or treatment of LV thrombus after MI, and clinicians must rely on available epidemiologic and trial-generated data from related entities to guide treatment. Randomized clinical trials have confirmed that triple therapy increases bleeding rates compared with less potent antithrombotic regimens after MI, and observational data suggest that triple therapy regimens may not prevent LV thrombus formation. On the other hand, if an LV thrombus is detected, anticoagulation is essential to prevent systemic thromboembolism. We offer 1 approach to treatment, grounded in the best available data. Conclusions and Relevance:Uncertainties remain regarding the optimal screening pathway, frequency of follow-up imaging, candidate selection for thromboprophylaxis, and treatment strategies for post-MI LV thrombus. Ongoing studies from related therapeutic areas of varying antithrombotic regimens will continue to inform the optimal approach to treatment; however, more dedicated study of this clinical conundrum is also needed.
10.1001/jamacardio.2018.1086
PCSK9 (Proprotein Convertase Subtilisin/Kexin 9) Enhances Platelet Activation, Thrombosis, and Myocardial Infarct Expansion by Binding to Platelet CD36.
Qi Zhiyong,Hu Liang,Zhang Jianjun,Yang Wenlong,Liu Xin,Jia Daile,Yao Zhifeng,Chang Lin,Pan Guanxing,Zhong Haoxuan,Luo Xinping,Yao Kang,Sun Aijun,Qian Juying,Ding Zhongren,Ge Junbo
Circulation
BACKGROUND:PCSK9 (proprotein convertase subtilisin/kexin 9), mainly secreted by the liver and released into the blood, elevates plasma low-density lipoprotein cholesterol by degrading low-density lipoprotein receptor. Pleiotropic effects of PCSK9 beyond lipid metabolism have been shown. However, the direct effects of PCSK9 on platelet activation and thrombosis, and the underlying mechanisms, as well, still remain unclear. METHODS:We detected the direct effects of PCSK9 on agonist-induced platelet aggregation, dense granule ATP release, integrin αIIbβ3 activation, α-granule release, spreading, and clot retraction. These studies were complemented by in vivo analysis of FeCl-injured mouse mesenteric arteriole thrombosis. We also investigated the underlying mechanisms. Using the myocardial infarction (MI) model, we explored the effects of PCSK9 on microvascular obstruction and infarct expansion post-MI. RESULTS:PCSK9 directly enhances agonist-induced platelet aggregation, dense granule ATP release, integrin αIIbβ3 activation, P-selectin release from α-granules, spreading, and clot retraction. In line, PCSK9 enhances in vivo thrombosis in a FeCl-injured mesenteric arteriole thrombosis mouse model, whereas PCSK9 inhibitor evolocumab ameliorates its enhancing effects. Mechanism studies revealed that PCSK9 binds to platelet CD36 and thus activates Src kinase and MAPK (mitogen-activated protein kinase)-extracellular signal-regulated kinase 5 and c-Jun N-terminal kinase, increases the generation of reactive oxygen species, and activates the p38MAPK/cytosolic phospholipase A2/cyclooxygenase-1/thromboxane A signaling pathways downstream of CD36 to enhance platelet activation, as well. Using CD36 knockout mice, we showed that the enhancing effects of PCSK9 on platelet activation are CD36 dependent. It is important to note that aspirin consistently abolishes the enhancing effects of PCSK9 on platelet activation and in vivo thrombosis. Last, we showed that PCSK9 activating platelet CD36 aggravates microvascular obstruction and promotes MI expansion post-MI. CONCLUSIONS:PCSK9 in plasma directly enhances platelet activation and in vivo thrombosis, and MI expansion post-MI, as well, by binding to platelet CD36 and thus activating the downstream signaling pathways. PCSK9 inhibitors or aspirin abolish the enhancing effects of PCSK9, supporting the use of aspirin in patients with high plasma PCSK9 levels in addition to PCSK9 inhibitors to prevent thrombotic complications.
10.1161/CIRCULATIONAHA.120.046290
Venous Thromboembolism: Advances in Diagnosis and Treatment.
Tritschler Tobias,Kraaijpoel Noémie,Le Gal Grégoire,Wells Philip S
JAMA
IMPORTANCE:Venous thromboembolism (VTE), comprising deep vein thrombosis (DVT) and pulmonary embolism (PE), is a common and potentially fatal disease. OBJECTIVE:To summarize the advances in diagnosis and treatment of VTE of the past 5 years. EVIDENCE REVIEW:A systematic search was conducted in EMBASE Classic, EMBASE, Ovid MEDLINE, and other nonindexed citations using broad terms for diagnosis and treatment of VTE to find systematic reviews and meta-analyses, randomized trials, and prospective cohort studies published between January 1, 2013, and July 31, 2018. The 10th edition of the American College of Chest Physicians Antithrombotic Therapy Guidelines was screened to identify additional studies. Screening of titles, abstracts, and, subsequently, full-text articles was performed in duplicate, as well as data extraction and risk-of-bias assessment of the included articles. FINDINGS:Thirty-two articles were included in this review. The application of an age-adjusted D-dimer threshold in patients with suspected PE has increased the number of patients in whom imaging can be withheld. The Pulmonary Embolism Rule-Out Criteria safely exclude PE when the pretest probability is low. The introduction of direct oral anticoagulants has allowed for a simplified treatment of VTE with a lower risk of bleeding regardless of etiology or extent of the VTE (except for massive PE) and has made extended secondary prevention more acceptable. Thrombolysis is best reserved for patients with massive PE or those with DVT and threatened limb loss. Insertion of inferior vena cava filters should be avoided unless anticoagulation is absolutely contraindicated in patients with recent acute VTE. Graduated compression stockings are no longer recommended to treat DVT but may be used when acute or chronic symptoms are present. Anticoagulation may no longer be indicated for patients with isolated distal DVT at low risk of recurrence. CONCLUSIONS AND RELEVANCE:Over the past 5 years, substantial progress has been made in VTE management, allowing for diagnostic and therapeutic strategies tailored to individual patient characteristics, preferences, and values.
10.1001/jama.2018.14346
Venous thromboembolism.
Khan Faizan,Tritschler Tobias,Kahn Susan R,Rodger Marc A
Lancet (London, England)
Venous thromboembolism, comprising both deep vein thrombosis and pulmonary embolism, is a chronic illness that affects nearly 10 million people every year worldwide. Strong provoking risk factors for venous thromboembolism include major surgery and active cancer, but most events are unprovoked. Diagnosis requires a sequential work-up that combines assessment of clinical pretest probability for venous thromboembolism using a clinical score (eg, Wells score), D-dimer testing, and imaging. Venous thromboembolism can be considered excluded in patients with both a non-high clinical pretest probability and normal D-dimer concentrations. When required, ultrasonography should be done for a suspected deep vein thrombosis and CT or ventilation-perfusion scintigraphy for a suspected pulmonary embolism. Direct oral anticoagulants (DOACs) are the first-line treatment for almost all patients with venous thromboembolism (including those with cancer). After completing 3-6 months of initial treatment, anticoagulation can be discontinued in patients with venous thromboembolism provoked by a major transient risk factor. Patients whose long-term risk of recurrent venous thromboembolism outweighs the long-term risk of major bleeding, such as those with active cancer or men with unprovoked venous thromboembolism, should receive indefinite anticoagulant treatment. Pharmacological venous thromboembolism prophylaxis is generally warranted in patients undergoing major orthopaedic or cancer surgery. Ongoing research is focused on improving diagnostic strategies for suspected deep vein thrombosis, comparing different DOACs, developing safer anticoagulants, and further individualising approaches for the prevention and management of venous thromboembolism.
10.1016/S0140-6736(20)32658-1
Innovative Molecular Testing Strategies for Adjunctive Investigations in Hemostasis and Thrombosis.
Seminars in thrombosis and hemostasis
Clinicians and scientists in the fields of hemostasis and thrombosis have been among those first to integrate new molecular strategies for the purpose of enhancing disease diagnosis and treatment. The molecular diagnosis and introduction of gene therapy approaches for hemophilia are obvious examples of this tendency. In this review, the authors summarize information concerning three molecular technologies that have reached various stages of translational potential for their incorporation into the clinical management of disorders of hemostasis. Chromatin conformation assays are now being used to capture structural knowledge of long-range genomic interactions that can alter patterns of gene expression and contribute to quantitative trait pathogenesis. Liquid biopsies in various forms are providing opportunities for early cancer detection, and in the context of tumor-educated platelets, as described here, can also characterize tumor type and the extent of tumor progression. This technology is already being trialed in patients with unprovoked venous thrombosis to assess the potential for occult malignancies. Lastly, advances in single cell transcriptome analysis, provide opportunities to definitively determine molecular events in rare cells, such as antigen-specific regulatory T cells, within the context of heterogeneous cell populations.
10.1055/s-0039-1692977
Basic science research opportunities in thrombosis and hemostasis: Communication from the SSC of the ISTH.
Journal of thrombosis and haemostasis : JTH
Bleeding and thrombosis are major clinical problems with high morbidity and mortality. Treatment modalities for these diseases have improved in recent years, but there are many clinical questions remaining and a need to advance diagnosis, management, and therapeutic options. Basic research plays a fundamental role in understanding normal and disease processes, yet this sector has observed a steady decline in funding prospects thereby hindering support for studies of mechanisms of disease and therapeutic development opportunities. With the financial constraints faced by basic scientists, the ISTH organized a basic science task force (BSTF), comprising Scientific and Standardization Committee subcommittee chairs and co-chairs, to identify research opportunities for basic science in hemostasis and thrombosis. The goal of the BSTF was to develop a set of recommended priorities to build support in the thrombosis and hemostasis community and to inform ISTH basic science programs and policy making. The BSTF identified three principal opportunity areas that were of significant overarching relevance: mechanisms causing bleeding, innate immunity and thrombosis, and venous thrombosis. Within these, five fundamental research areas were highlighted: blood rheology, platelet biogenesis, cellular contributions to thrombosis and hemostasis, structure-function protein analyses, and visualization of hemostasis. This position paper discusses the importance and relevance of these opportunities and research areas, and the rationale for their inclusion. These findings have implications for the future of fundamental research in thrombosis and hemostasis to make transformative scientific discoveries and tackle key clinical questions. This will permit better understanding, prevention, diagnosis, and treatment of hemostatic and thrombotic conditions.
10.1111/jth.15718
Snake Venoms in Diagnostic Hemostasis and Thrombosis.
Moore Gary William
Seminars in thrombosis and hemostasis
Snake venoms have evolved primarily to immobilize and kill prey, and consequently, they contain some of the most potent natural toxins. Part of that armory is a range of hemotoxic components that affect every area of hemostasis, which we have harnessed to great effect in the study and diagnosis of hemostatic disorders. The most widely used are those that affect coagulation, such as thrombin-like enzymes unaffected by heparin and direct thrombin inhibitors, which can help confirm or dispute their presence in plasma. The liquid gold of coagulation activators is Russell's viper venom, since it contains activators of factor X and factor V. It is used in a range of clotting-based assays, such as assessment of factor X and factor V deficiencies, protein C and protein S deficiencies, activated protein C resistance, and probably the most important test for lupus anticoagulants, the dilute Russell's viper venom time. Activators of prothrombin, such as oscutarin C from Coastal Taipan venom and ecarin from saw-scaled viper venom, are employed in prothrombin activity assays and lupus anticoagulant detection, and ecarin has a valuable role in quantitative assays of direct thrombin inhibitors. Snake venoms affecting primary hemostasis include botrocetin from the jararaca, which can be used to assay von Willebrand factor activity, and convulxin from the cascavel, which can be used to detect deficiency of the platelet collagen receptor, glycoprotein VI. This article takes the reader to every area of the diagnostic hemostasis laboratory to appreciate the myriad applications of snake venoms available in diagnostic practice.
10.1055/s-0041-1732465
Use of Thromboelastography in Clinical Practice.
Burton Andrew G,Jandrey Karl E
The Veterinary clinics of North America. Small animal practice
Viscoelastic testing, such as thromboelastography or thromboelastometry, is performed on whole-blood samples, which include both soluble plasma factors as well as blood cells and platelets bearing tissue factor and phospholipid. This methodology allows identification of fibrinolysis and can provide analysis of platelet function. Viscoelastic testing has become increasingly accessible and popular in emergency and critical care settings in recent years and can provide important information for the diagnosis and management of patients with hemostatic disorders. This article discusses the principles and interpretation of viscoelastic testing, application to small animal emergency and critical care medicine, and potential advantages and disadvantages.
10.1016/j.cvsm.2020.08.001
Disseminated intravascular coagulation and its immune mechanisms.
Blood
Disseminated intravascular coagulation (DIC) is a syndrome triggered by infectious and noninfectious pathologies characterized by excessive generation of thrombin within the vasculature and widespread proteolytic conversion of fibrinogen. Despite diverse clinical manifestations ranging from thrombo-occlusive damage to bleeding diathesis, DIC etiology commonly involves excessive activation of blood coagulation and overlapping dysregulation of anticoagulants and fibrinolysis. Initiation of blood coagulation follows intravascular expression of tissue factor or activation of the contact pathway in response to pathogen-associated or host-derived, damage-associated molecular patterns. The process is further amplified through inflammatory and immunothrombotic mechanisms. Consumption of anticoagulants and disruption of endothelial homeostasis lower the regulatory control and disseminate microvascular thrombosis. Clinical DIC development in patients is associated with worsening morbidities and increased mortality, regardless of the underlying pathology; therefore, timely recognition of DIC is critical for reducing the pathologic burden. Due to the diversity of triggers and pathogenic mechanisms leading to DIC, diagnosis is based on algorithms that quantify hemostatic imbalance, thrombocytopenia, and fibrinogen conversion. Because current diagnosis primarily assesses overt consumptive coagulopathies, there is a critical need for better recognition of nonovert DIC and/or pre-DIC states. Therapeutic strategies for patients with DIC involve resolution of the eliciting triggers and supportive care for the hemostatic imbalance. Despite medical care, mortality in patients with DIC remains high, and new strategies, tailored to the underlying pathologic mechanisms, are needed.
10.1182/blood.2020007208
Communication from the Scientific and Standardization Committee of the International Society on Thrombosis and Haemostasis on sepsis-induced coagulopathy in the management of sepsis.
Journal of thrombosis and haemostasis : JTH
Disseminated intravascular coagulation (DIC) is a life-threatening complication in sepsis and other critical conditions. The International Society on Thrombosis and Haemostasis (ISTH) released the diagnostic criteria for overt DIC in 2001. Since then, ISTH overt DIC has been used as the global standard criterion for a decompensated stage of DIC. Because detecting an earlier stage of DIC would be useful for therapeutic considerations, the scientific standardization committees of the ISTH introduced the sepsis-induced coagulopathy (SIC) scoring system in 2019. The SIC scoring system is specifically designed to detect the compensated phase of DIC in sepsis, which can lead to overt DIC with disease progression. Studies examining the performance of the SIC scoring system have reported its usefulness over the past 5 years. The reported incidence of SIC was approximately 60% in patients with sepsis, which was twice as much as that of overt DIC. Almost all patients with overt DIC were diagnosed with SIC earlier. The reported mortality of SIC was ≥30% and, thus, can be used for patient selection for anticoagulant therapy. Despite the limited data, some continue to suggest the potential efficacy of anticoagulant therapy in patients with SIC. Although heparin, antithrombin, and thrombomodulin are the candidates for anticoagulation, none of them have proven to be effective with robust evidence, and future trials are warranted.
10.1016/j.jtha.2022.10.022