Antithrombin Activity Research Articles

Zn2+ Differentially Influences the Neutralisation of Heparins by HRG, Fibrinogen, and Fibronectin.

For coagulation to be initiated, anticoagulant glycosaminoglycans (GAGs) such as heparins need to be neutralised to allow fibrin clot formation. Platelet activation triggers the release of several proteins that bind GAGs, including histidine-rich glycoprotein (HRG), fibrinogen, and fibronectin. Zn2+ ions are also released and have been shown to enhance the binding of HRG to heparins of a high molecular weight (HMWH) but not to those of low molecular weight (LMWH). The effect of Zn2+ on fibrinogen and fibronectin binding to GAGs is unknown. Here, chromogenic assays were used to measure the anti-factor Xa and anti-thrombin activities of heparins of different molecular weights and to assess the effects of HRG, fibrinogen, fibronectin, and Zn2+. Surface plasmon resonance was also used to examine the influence of Zn2+ on the binding of fibrinogen to heparins of different molecular weights. Zn2+ had no effect on the neutralisation of anti-factor Xa (FXa) or anti-thrombin activities of heparin by fibronectin, whereas it enhanced the neutralisation of unfractionated heparin (UFH) and HMWH by both fibrinogen and HRG. Zn2+ also increased neutralisation of the anti-FXa activity of LMWH by fibrinogen but not HRG. SPR showed that Zn2+ increased fibrinogen binding to both UFH and LMWH in a concentration-dependent manner. The presented results reveal that an increase in Zn2+ concentration has differential effects upon anticoagulant GAG neutralisation by HRG and fibrinogen, with implications for modulating anti-coagulant activity in plasma.

The state of hemostasis system in patients after mild coronavirus infection

Introduction. One of the complications of the new coronavirus infection (COVID-19) is the development of venous thromboembolic complications. In this regard, changes in the hemostasis system that persist in the process of recovery in patients who have had a mild form of the disease are of interest.Aim. To evaluate plasma hemostasis in patients after mild coronavirus infection Materials and methods. 39 patients after mild coronavirus infection were examined. The following parameters were assessed: APTT, PT, fibrinogen, factor VIII, von Willebrand factor, D-dimer, proteins C and S, and thrombin generation. Results. In patients who underwent COVID-19, in comparison with healthy individuals, an increase in the plasma concentration of fibrinogen, von Willebrand factor antigen, and D-dimer was detected. Furthermore, increased activity of antithrombin III, decreased sensitivity to thrombomodulin, and an increase in the coagulation index were detected. Conclusion. Patients who have undergone COVID-19 are characterized by a procoagulant orientation of changes in plasma hemostasis that persist up to 3 months after the disease. It is necessary to take into account the state of the hemostasis system even in patients who have undergone COVID-19 in mild form.

The antithrombin activity recovery after substitution therapy is associated with improved 28-day mortality in patients with sepsis-associated disseminated intravascular coagulation

BackgroundDisseminated intravascular coagulation (DIC) is a common and critical complication in sepsis. Antithrombin activity, which is considered a biomarker for disease severity, was measured in septic DIC treated with antithrombin concentrates in this study.MethodsWe conducted a retrospective analysis of post-marketing survey data that included 1,800 patients with sepsis-associated DIC and antithrombin activity of 70% or less who were treated with antithrombin concentrates. The changes in sequential organ failure assessment (SOFA) score, DIC score, and antithrombin activity were sequentially assessed. Logistic regression analysis and receiver operating characteristic (ROC) curve analysis were performed to evaluate the performance of antithrombin activity to assess 28-day survival. Furthermore, the relationship between post-treatment antithrombin activity and survival was examined by Logistic regression analysis.ResultsSex, baseline SOFA score, baseline antithrombin activities, and the presence of pneumonia and soft tissue infection were significantly associated with 28-day mortality. The area under the curve for mortality was 0.639 for post-treatment antithrombin activity, and higher than those of baseline- and delta antithrombin activities. Logistic regression analysis revealed that higher post-treatment antithrombin activity was associated with better 28-day survival. When post-treatment antithrombin activity was more than 80%, the estimated survival was 88.2%. Whereas, the survival was 74.4% when the antithrombin activity was 80% or less (P < 0.0001). However, the relationship between post-treatment antithrombin activity and 28-day survival was considerably different between patients who recovered from DIC by Day 6 compared to those who did not. Similarly, the estimated 28-day survival, based on antithrombin activity, varied among patients with high and low SOFA scores, and the calculation needs to be adjusted based on the severity of the condition.ConclusionsPost-treatment antithrombin activity measurement was helpful in estimating the 28-day survival in patients with sepsis-associated DIC. However, patient outcomes vary considerably depending on factors that include baseline SOFA score, age, and baseline antithrombin activity. These variables play a substantial role in determining patient prognosis and should be considered when evaluating and interpreting the results.

Hemostasis Biomarkers and Long-Term Risk of Recurrent Cardiovascular Events and Bleeding Complications in Young Patients with an Ischemic Stroke

Background: Patients with ischemic stroke at a young age (18-50 years) have a persistent increased risk of recurrent ischemic cardiovascular events, despite antithrombotic treatment. Additionally, antithrombotic agents result in a higher risk of bleeding complications. Balancing hemostasis to prevent thrombotic events and minimizing the risk of hemorrhagic events is challenging. Previous research observed a hypercoagulable state and a significant degree of inflammation years after an acute ischemic stroke in young adults. It is known that an increased clotting tendency is a risk factor for a first ischemic stroke, especially in young adults. However, the risk profile of a recurrent event in these individuals can be different. To this end, our study aims to investigate the role of plasma pro- and anticoagulation biomarkers, platelet activation markers and thrombin generation parameters in the risk of recurrent ischemic arterial events and bleeding complications after a first ischemic stroke or transient ischemic attack (TIA) at a young age. Method: Patients from a single-center cohort study (the FUTURE cohort) with a first-ever TIA or ischemic stroke event between 1980 and 2010 were included. Blood was collected in 2010 and patients were followed on recurrent ischemic events or bleeding complications between 2010 and 2023. The following coagulation parameters were measured: blood cell count, Von Willebrand factor, active Von Willebrand factor, factor VIII, prothrombin, fragment 1.2, d-dimer, fibrinogen, fibrin degradation products, antithrombin, alpha-2-macroglobulin, serum CXCL-4 and CXCL-7. In addition, thrombin generation (TG) was determined using 1pM and 5pM tissue factor, and thrombomodulin in platelet-poor plasma to assess the functional capacity of plasma to form thrombin. The primary outcome was a composite of recurrent ischemic stroke, TIA, acute coronary syndrome or peripheral artery disease. The secondary outcome was the occurrence of a bleeding complication using the Bleeding Academic Research Consortium (BARC) criteria. We applied Cox proportional hazard models to estimate hazard ratios (HR) with 95% confidence intervals (CI). Results : Among 378 patients (median age 42.1 years, 56% women), 74 experienced a recurrent ischemic event during a mean follow-up of 6.5 years. An increased platelet and leukocyte count, along with elevated levels of fragment 1.2, fibrinogen and D-dimer were associated with a higher risk of ischemic events (table 1). Patients with low plasma antithrombin activity had a higher risk of recurrent ischemic events, especially in women (HR 0.67 [0.47-0.96]). After adjustment for age, sex and traditional risk factors, the association between leukocytes numbers, fragment 1.2 and fibrinogen levels and antithrombin activity and the risk of recurrent ischemic events remained: HR 1.14 [1.05-1.25], 1.07 [1.01-1.14], 0.80 [0.65-0.98], and 1.09 [1.00-1.17], respectively. In patients without vitamin K antagonist treatment, the association between antithrombin activity (HR 0.78 [0.62-0.98]) and leukocytes numbers (HR 1.15 [1.05-1.25]) with recurrent ischemic events were stronger. Thrombin generation parameters were not related to recurrence. Bleeding complications were observed in 16% of patients, mainly among women and predominantly attributable to gynecological bleedings. Hemoglobin, platelets, CXCL-4 and -7, prothrombin, fragment 1.2 and alpha-2-macroglobulin were associated with a bleeding complication (table 2). The association for platelets (HR 0.93 [0.89-0.97]), CXCL-4 (HR 0.90 [0.82-0.99]) and alpha-2-macroglobulin (HR 1.15 [1.03-1.29]) remained after adjustment. Conclusion: Lower antithrombin activity and, higher fragment 1.2, fibrinogen levels, and leukocytes counts were linked to an increased risk of recurrent ischemic events. Bleeding was associated with platelets, CXCL4, and alpha-2-macroglobulin. Overall, especially platelets appear to be significantly involved in the risk of both recurrent arterial thrombosis and bleeding, whereas the observed association with risk of recurrent ischemic events implies a more dominant role of the coagulation system.

Antithrombin activity and central venous catheter-associated thrombosis in critically ill children at high risk of bleeding

BackgroundNormalization of antithrombin activity may prevent catheter-associated thrombosis in critically ill children at high risk of bleeding. ObjectivesTo characterize the temporal pattern of antithrombin activity, assess its association with catheter-associated thrombosis and clinically relevant bleeding, and evaluate its relationship with thrombin generation in these children. MethodsIn this prospective cohort study, critically ill children <18 years old at high risk of bleeding with central venous catheter were eligible. Antithrombin activity and thrombin generation were measured from platelet-poor plasma and after in vitro antithrombin supplementation. Systematic surveillance ultrasound was performed to diagnose thrombosis. Children were followed for bleeding. ResultsWe enrolled 8 infants (median age: 0.2 years, IQR: 0.2, 0.3 years) and 72 older children (median age: 14.3 years, IQR: 9.1, 16.1 years). Mean antithrombin on the day of catheter insertion was 64 IU/dL (SD: 32 IU/dL) in infants and 83 IU/dL (SD: 35 IU/dL) in older children. Antithrombin normalized by the day of catheter removal.Thrombosis developed in 27 children, while 31 children bled. Thrombosis (regression coefficient: 0.008, 95% CI: -0.01, 0.03) and bleeding (regression coefficient: -0.0007, 95% CI: -0.02, 0.02) were not associated with antithrombin. Antithrombin was not correlated with in vivo change in endogenous thrombin potential (correlation coefficient: -0.07, 95% CI: -0.21, 0.08). In vitro supplementation reduced endogenous thrombin potential (correlation coefficient: -0.78; 95% CI: -0.95, -0.23). ConclusionThese findings may not support normalization of antithrombin activity to prevent catheter-associated thrombosis in critically ill children at high risk of bleeding.

Comparison of antithrombin activity assays in detection of patients with heparin binding site antithrombin deficiency: systematic review and meta-analysis

Antithrombin (AT) deficiency increases the risk for venous thromboembolism, therefore, a highly sensitive assay to identify this condition is crucial. The aim of this paper was to perform a meta-analysis comparing AT activities measured by different AT activity assays in patients with heparin binding site AT deficiency. In addition, the diagnostic sensitivity of selected assays was compared depending on the available data. An extensive literature search was performed considering results with publication date up to July 10, 2021. Seven relevant English-language observational studies, comparing AT activity measured by different AT activity assays in Caucasian Europeans with either the AT Budapest III or AT Padua I mutation were included in meta-analyses. There was no significant difference in AT activity between Labexpert and Innovance in patients with AT Budapest III (P = 0.567) and AT Padua I (P = 0.265), while AT activity determined by HemosIL was significantly higher compared to Innovance for both mutations (AT Budapest III: P < 0.001; AT Padua I: P < 0.001). These results are in line with the results of comparison of diagnostic sensitivity. In patients with AT Budapest III, the AT activity was also higher when measured with Berichrom compared to Innovance (P = 0.002), however, the results of comparison of diagnostic sensitivity across studies were variable. No significant difference (P = 0.117) in AT activity as well as diagnostic sensitivity was observed between Sta-Stachrom and Innovance. The results of our study suggest that Innovance, Labexpert and Sta-Stachrom are the most sensitive activity assays for detection of AT Budapest III and AT Padua I, whereas HemosIL showed considerably lower sensitivity for these two variants. As revealed in our study, the diagnostic sensitivity of AT activity assays to type II heparin binding site AT deficiency is different, and in some assays mutation dependent.

PB0576 Correlation between High Affinity Material for Antithrombin and Anticoagulant Activities of Heparin

PB0576 Correlation between High Affinity Material for Antithrombin and Anticoagulant Activities of Heparin

Immunity and hemostasis system indicators in age groups of laboratory rats during the use of cyclosporine A

Dysregulation of the T-cell link increases the susceptibility to cardiovascular disease with age. Therefore, we modeled immunosuppression and experimentally compared the state of the hemostasis system in young and old animals. White outbred male rats were used for the experiment. Cyclosporine in Sandimmun-Neoral (Novartis, Switzerland) was administered intramuscularly to experimental rats. Complement system proteins C3, C4, immunoglobulins (Ig) A, M, G and interleukin-6 were examined by enzyme immunoassay. Phenotyping of lymphocytes was performed by indirect immunofluorescence using monoclonal antibodies (subpopulations CD3+, CD19+, CD4+, CD8+, CD16+). The data obtained indicate that inhibition of the T-lymphocyte link (CD3+) does not necessarily lead to a compensated increase in the number of B-lymphocytes (CD19+), but stimulates their function: immunoglobulins are elevated (M, G). Under conditions of a reduced reaction from the cellular level, components of the complement system (C3, C4) are activated. Despite a slight decrease in phagocytic function, there was an increase in the number of natural killers (CD16+), active consumption of T-helpers (CD4+), a decrease in T-suppressors (CD8+), and an increase in the level of interleukin-6 in the plasma of experimental rats. Increased activity of antithrombin in old animals prevents procoagulant shifts against the background of excessive growth of "areas" with tissue factor. We found desquamated endotheliocytes, that opens the subendothelial layer and its level was higher in the experimental groups. Thus, we see an activation of the humoral link of the immune system. Experimental excessive concentration of the calcineurin blocker exposes the first pathogenetic links from the hemostasis system. An increase in the coagulation time of the initial stages of coagulation is combined with an acceleration of the final one. Despite the growth of fibrinogen in young animals, the mismatch in the regulation of the hemostasis system manifested itself mainly in old specimens of laboratory rats.

COVID-19-associated coagulopathy and acute kidney injury in critically ill patients.

The incidence of thrombotic events and acute kidney injury is high in critically ill patients with COVID-19. We aimed to evaluate and compare the coagulation profiles of patients with COVID-19 developing acute kidney injury versus those who did not, during their intensive care unit stay. Conventional coagulation and platelet function tests, fibrinolysis, endogenous inhibitors of coagulation tests, and rotational thromboelastometry were conducted on days 0, 1, 3, 7, and 14 following intensive care unit admission. Out of 30 patients included, 13 (43.4%) met the criteria for acute kidney injury. Comparing both groups, patients with acute kidney injury were older: 73 (60-84) versus 54 (47-64) years, p=0.027, and had a lower baseline glomerular filtration rate: 70 (51-81) versus 93 (83-106) mL/min/1.73m2, p=0.004. On day 1, D-dimer and fibrinogen levels were elevated but similar between groups: 1780 (1319-5517) versus 1794 (726-2324) ng/mL, p=0.145 and 608 (550-700) versus 642 (469-722) g/dL, p=0.95, respectively. Rotational thromboelastometry data were also similar between groups. However, antithrombin activity and protein C levels were lower in patients who developed acute kidney injury: 82 (75-92) versus 98 (90-116), p=0.028 and 70 (52-82) versus 88 (78-101) µ/mL, p=0.038, respectively. Mean protein C levels were lower in the group with acute kidney injury across multiple time points during their stay in the intensive care unit. Critically ill patients experiencing acute kidney injury exhibited lower endogenous anticoagulant levels. Further studies are needed to understand the role of natural anticoagulants in the pathophysiology of acute kidney injury within this population.

A prospective evaluation of the prevalence of thromboemboli and associated hemostatic dysfunction in dogs with carcinoma or sarcoma.

Knowledge of the prevalence of thromboemboli and the associated hemostatic status in dogs with carcinoma or sarcoma is unknown and might allow earlier intervention. Estimate prevalence of thromboemboli and their association with hemostatic changes in dogs with carcinomas or sarcomas; estimate predictive values of hemostatic variables for thromboembolic disease in tumor-bearing dogs. Thirty-two dogs with sarcoma, 30 with carcinoma, 20 healthy age-controlled dogs. Prospective cross-sectional study. A hemostasis panel (platelet concentration, thromboelastography, fibrinogen and D-dimer concentration, factor X, VII and antithrombin activity) was performed in all dogs. Tumor-bearing dogs underwent complete post mortem and histopathological evaluation. Comparisons between healthy dogs and tumor-bearing dogs with and without intracavitary hemorrhage; and tumor-bearing dogs with and without microthrombi were analyzed. Thromboembolic disease was identified in 32/62 (52%, 95% CI: 39%-65%) tumor-bearing dogs. Microthrombi were identified in 31/62 (50%, 95% CI: 37%-63%) dogs, 21/31 (68%, 95% CI: 49%-83%) had exclusively intra-tumoral microthrombi, 10/31 (32%, 95% CI: 17%-51%) had distant microthrombi. Macrothrombi were identified in 3 tumor-bearing dogs. Hemostatic changes potentially consistent with overt and non-overt disseminated intravascular coagulation were identified in some tumor-bearing dogs. D-dimer concentrations were significantly higher (P = .02) and platelet concentration significantly lower (P = .03) in tumor-bearing dogs with microthrombi compared to tumor-bearing dogs without microthrombi. D-dimer concentration above 500 ng/mL was 80% sensitive and 41% specific for the prediction of microthrombi presence. The high microthrombi prevalence and concomitant hemostatic dysfunction in dogs with carcinomas or sarcomas has not previously been reported, though the clinical importance is unknown. Increased D-dimer concentration might increase suspicion of microthrombi.

High prevalence of thrombophilic risk factors in patients with central retinal artery occlusion

IntroductionCentral retinal artery occlusion (CRAO) is a common cause of blindness and visual morbidity. In the majority of cases, it is related to thrombotic embolism. Nevertheless, the role of inherited or acquired thrombophilic risk factors in CRAO pathogenesis has not been comprehensively studied.MethodsIn 126 CRAO patients (66 [52.4%] men, median age 55 [range: 18–80] years) and 107 matched controls (56 [52.3%] men, median age 53 [range: 34–78] years) we evaluated classical atherosclerotic risk factors, including serum lipid profile and glucose level, analyzed intima-media complex thickness (IMT) of external carotid arteries, and performed transthoracic echocardiography. Furthermore, we established the prevalence of inherited and acquired thrombophilic risk factors, such as factor V Leiden (FVL) and prothrombin 20210 G/A genetic variants, plasma activity of factor (F) VIII, protein C and antithrombin activity, and free protein S levels. We also assessed the presence of antiphospholipid antibodies (APLA) and evaluated blood homocysteine in all enrolled subjects. Additionally, we estimated the occurrence of Val34Leu polymorphism of the A subunit of coagulation factor XIII (FXIII-A) in both groups as a potential thrombosis-protecting factor.ResultsAmong traditional atherosclerotic risk components, obesity/overweight and hypercholesterolemia were the most common in the CRAO group and occurred in 103 (81.7%) and 85 (67.5%) patients, respectively. CRAO patients also had elevated IMT and altered echocardiographic parameters, indicating diastolic cardiac dysfunction. In thrombophilia investigations, at least one laboratory risk factor occurred in 72.2% (n = 91) of CRAO patients, with APLA as the most frequent, detected in 38.1% (n = 48) of them (almost seven times more frequent than in controls, p < 0.001). Deficiencies in protein C activity and free protein S levels were also common in the CRAO group, reported in 17.5% (n = 22) and 19.8% (n = 25) of patients, respectively. Interestingly, among two analyzed prothrombotic genetic variants, only the FVL was related to CRAO, with the allelic frequency 2.4 times more prevalent than in controls (p = 0.044). Finally, the CRAO group was characterized by hyperhomocysteinemia, almost twice as common as in controls (p = 0.026). Antithrombin deficiency, elevated FVIII, and FXIII-A Val34Leu polymorphism were not associated with CRAO.ConclusionsOur findings suggest that thrombophilia plays a vital role in the pathogenesis of CRAO. Thus, proper laboratory screening should be considered in the primary and secondary prevention of those episodes, with implementing appropriate therapy as needed.

Comparative Studies of Complex Products Based on Heparins of Animal and Plant Origin

Complex preparations based on heparins of animal and vegetable origin with the inclusion of glutamic acid in a weight ratio of 1 : 4 were obtained. High molecular weight (HMН) and low molecular weight (LMWH) commercial preparations of heparins were used as heparins of animal origin, heparinoid was used as heparin of plant origin, isolated from the roots of the peony Akademik Sadovnichy (heparinoid). Their influence on the state of plasma hemostasis under in vitro conditions was studied in a comparative aspect. It has been established that all complex compounds have anticoagulant activity, which, according to the results of our studies, is realized by a different mechanism of their action on blood coagulation – on the internal, external, or general pathways of blood coagulation. HMН complexes with glutamic acid had an antithrombin effect, affecting the internal and general coagulation pathways, while LMWH complexes with glutamic acid, exerting anti-Xa activity, affected the internal pathway of coagulation activation, and the combination of heparinoid with glutamic acid inhibited factors not only internal, but also external ways of blood coagulation. In addition, the latter compound exhibited weak antithrombin activity. It was also shown that all heparin complexes prevented fibrin polymerization, increasing the fibrin depolymerization activity of plasma, and in the heparinoid compound, this activity was higher by 10–17% compared to other commercial agents. Thus, the best anticoagulant in terms of efficacy and safety is the heparinoid complex with glutamic acid.

In-vitro Evaluation of Antithrombin and Thrombolytic Activity of Leaves Extract of Lantana camara Linn

Thrombosis is possible outcomes of excessive coagulation or inhibition of anticoagulant processes. The pathophysiology of thrombosis is primarily caused by irregularities of the vascular wall, changes in blood flow, and changes in blood composition. Now a day’s, various natural antithrombotic or thrombolytic medications are used for the treatment. This study evaluated the pharmacognostical, antithrombin and thrombolytic activity of leaves extract of plant Lantana camara. The extractive value of leaves extract of plant L. camara was higher (20.58%) in methylene chloride as solvent. Total ash of plant L. camara was about 9.00% w/w. In comparison with heparin L. camara extract shown 50.10, 60.23, 62.32, 68.48 and 71.32% antithrombin activity at 10, 20, 40, 60, 80 and 100 mg/mL concentration, respectively. When different extract concentrations were added in the tubes containing blood clots, L. camara showed a good thrombolytic activity. We conclude that the leaves extract of plant L. camara shows significant antithrombin and thrombolytic activity.

Coagulation abnormalities in a prospective cohort of 50 patients with PMM2-congenital disorder of glycosylation.

Given the lack of reliable data on the prevalence of bleeding abnormalities and thrombotic episodes in PMM2-CDG patients, and whether coagulation abnormalities change over time, we prospectively collected and reviewed natural history data. Patients with PMM2-CDG often have abnormal coagulation studies due to glycosylation abnormalities but the frequency of complications resulting from these has not been prospectively studied. We studied fifty individuals enrolled in the Frontiers in Congenital Disorders of Glycosylation Consortium (FCDGC) natural history study with molecularly confirmed diagnosis of PMM2-CDG. We collected data on prothrombin time (PT), international normalized ratio (INR), activated partial thromboplastin time (aPTT), platelets, factor IX activity (FIX), factor XI activity (FXI), protein C activity (PC), protein S activity (PS) and antithrombin activity (AT). Prothrombotic and antithrombotic factor activities were frequently abnormal in PMM2-CDG patients, including AT, PC, PT, INR, and FXI. AT deficiency was the most common abnormality in 83.3% of patients. AT activity was below 50% in 62.5% of all patients (normal range 80-130%). Interestingly, 16% of the cohort experienced symptoms of spontaneous bleeding and 10% had thrombosis. Stroke-like episodes (SLE) were reported in 18% of patients in our cohort. Based on the linear growth models, on average, patients did not show significant change in AT (n=48; t(23.8)=1.75, p=0.09), FIX (n=36; t(61)=1.60, p=0.12), FXI (n=39; t(22.8)=1.88, p=0.07), PS (n=25; t(28.8)=1.08, p=0.29), PC (n=38; t(68)=1.61, p=0.11), INR (n=44; t(184)=-1.06, p=0.29), or PT (n=43; t(192)=-0.69, p=0.49) over time. AT activity positively correlated with FIX activity. PS activity was significantly lower in males. Based on our natural history data and previous literature, we conclude that caution should be exercised when the AT levels are lower than 65%, as most thrombotic events occur in patients with AT below this level. All five, male PMM2-CDG patients in our cohort who developed thrombosis had abnormal AT levels, ranging between 19% and 63%. Thrombosis was associated with infection in all cases. We did not find significant change in AT levels over time. Several PMM2-CDG patients had an increased bleeding tendency. More long-term follow-up is necessary on coagulation abnormalities and the associated clinical symptoms to provide guidelines for therapy, patient management, and appropriate counseling. Most PMM2-CDG patients display chronic coagulation abnormalities without significant improvement, associated with a frequency of 16% clinical bleeding abnormalities, and 10% thrombotic episodes in patients with severe antithrombin deficiency.

Curcumol, a major terpenoid from Curcumae Rhizoma, attenuates human uterine leiomyoma cell development via the p38MAPK/NF-κB pathway.

Uterine fibroids (UFs) are the most common benign tumors in women of reproductive age. Curcumae Rhizoma, the main essential oil component of which is curcumol, is widely used for the treatment of phymatosis in China due to its antitumor, anti-inflammatory, antithrombin, anti-tissue fibrosis and anti-oxygen pharmacological activities, but its potential for the treatment of UFs has not been evaluated. This study aimed to investigate the effects and mechanisms of curcumol intervention in human uterine leiomyoma cells (UMCs). Putative targets of curcumol intervention in UFs were identified using network pharmacology strategies. Molecular docking was performed to assess the binding affinity of curcumol to core targets. A concentration gradient of curcumol (0, 50, 100, 200, 300, 400 and 500μM) or RU-486 (mifepristone, 0, 10, 20, 40, 50, and 100μM) was applied to UMCs, and cell viability was detected by the CCK-8 assay. Cell apoptosis and cell cycle were examined by flow cytometry, and cell migration was assessed by a wound-healing assay. Additionally, the mRNA and protein expression levels of critical pathway components were evaluated by RT‒PCR and western blotting. Finally, the actions of curcumol on different tumor cell lines were summarized. Network pharmacology predicted 62 genes with roles in the treatment of UFs with curcumol, and MAPK14 (p38MAPK) displayed a higher interaction degree. GO enrichment and KEGG analyses revealed that the core genes were abundantly enriched in the MAPK signaling pathway. The molecular binding of curcumol to core targets was relatively stable. In UMCs, 200, 300 and 400μM curcumol treatment for 24h decreased cell viability compared with that in the control group, and the greatest effect was detected at 48h and maintained until 72h. Curcumol arrested cells in the G0/G1 phase and subsequently suppressed mitosis, promoted early apoptosis and reduced the degree of wound healing in a concentration-dependent manner in UMCs. Furthermore, 200μM curcumol decreased the mRNA and protein expression of p38MAPK, the mRNA expression of NF-κB, and the protein expression of Ki-67 and increased the mRNA and protein expression of Caspase 9. Curcumol (300 and 400μM) decreased the mRNA and protein expression of p38MAPK, NF-κB, and Ki-67 and increased the protein expression of Caspase 9 in UMCs. Curcumol was demonstrated to treat tumor cell lines, including breast cancer, ovarian cancer, lung cancer, gastric cancer, liver cancer and nasopharyngeal carcinoma, but its effects on benign tumors have not yet been reported. Curcumol suppresses cell proliferation and cell migration while arresting the cell cycle in the G0/G1 phase and inducing cell apoptosis in UMCs via a mechanism related to p38MAPK/NF-κB pathway regulation. Curcumol may be a potential therapeutic and preventive agent in the treatment of benign tumors such as UFs.

Antithrombin Deficiency Is Associated with Prothrombotic Plasma Fibrin Clot Phenotype.

Deficiency of antithrombin increases risk of venous thromboembolism. We hypothesized that antithrombin deficiency affects fibrin clot structure and function. We evaluated 148 patients (age: 38 [32-50] years; 70% women) with genetically confirmed antithrombin deficiency and 50 healthy controls. Fibrin clot permeability (Ks) and clot lysis time (CLT) along with thrombin generation capacity were assessed before and after antithrombin activity normalization in vitro. Antithrombin-deficient patients had lower antithrombin activity (-39%) and antigen levels (-23%) compared with controls (both p < 0.01). Prothrombin fragment 1 + 2 levels were 26.5% higher in patients with antithrombin deficiency than in controls along with 94% increased endogenous thrombin potential (ETP) and 108% higher peak thrombin (all p < 0.01). Antithrombin deficiency was associated with 18% reduced Ks and 35% prolonged CLT (both p < 0.001). Patients with type I (n = 65; 43.9%) compared with type II antithrombin deficiency (n = 83; 56.1%) had 22.5% lower antithrombin activity (p < 0.001) and despite similar fibrinogen levels, 8.4% reduced Ks, 18% prolonged CLT, and 30% higher ETP (all p < 0.01). Reduced Ks was associated with lower antithrombin antigen level (β = - 6.1, 95% confidence interval [CI]: -1.7 to -10.5), while prolonged CLT was associated with lower antithrombin antigen (β = - 69.6, 95% CI: -9.6 to -129.7), activity (β = - 2.4, 95% CI: -0.3 to -4.5), higher PAI-1 (β = 12.1, 95% CI: 7.7-16.5), and thrombin-activatable fibrinolysis inhibitor levels (β = 3.8, 95% CI: 1.9-5.7). Addition of exogenous antithrombin reduced ETP (-42%) and peak thrombin (-21%), and improved Ks (+8%) and CLT (-12%; all p < 0.01). Our study suggests that enhanced thrombin generation and prothrombotic plasma fibrin clot phenotype can contribute to increased risk of thrombosis in patients with antithrombin deficiency.

Investigation of real-world heparin resistance and anticoagulation management prior to cardiopulmonary bypass: report from a nationwide survey by the Japanese Association for Thoracic Surgery heparin resistance working group

ObjectiveHeparin resistance is often encountered during cardiopulmonary bypass. Heparin dose and activated clotting time target values for the initiation of cardiopulmonary bypass are not yet universally standardized; further no consensus exists on the management of heparin resistance. This study aimed to investigate the current real-world practice on heparin management and anticoagulant treatment for heparin resistance in Japan.MethodsA questionnaire survey was conducted at medical institutions nationwide with which The Japanese Society of Extra-Corporeal Technology in Medicine members are affiliated, targeting surgical cases with cardiopulmonary bypass performed from January 2019 through December 2019.ResultsAmong 69% (230/332) of the participating institutions, the criterion for heparin resistance was defined as “the target activated clotting time value not reached even with an additional dose of heparin administration”. Cases of heparin resistance were reported in 89.8% (202/225) of the responded institutions. Of note, 75% (106/141) of the responded institutions reported heparin resistance associated with antithrombin activity ≥ 80%. Antithrombin concentrate was used in 38.4% (238/619 responses) or third dose of heparin in 37.8% (234/619 responses) for advanced heparin resistance treatment. Antithrombin concentrate was found to be effective in resolving heparin resistance in patients having normal, as well as lower antithrombin activity.ConclusionHeparin resistance has occurred in many cardiovascular centers, even among patients with normal antithrombin activities. Interestingly, the administration of antithrombin concentrate resolved heparin resistance, regardless of the baseline antithrombin activity value.

Renal transplant and hemostasis: early postoperative changes in recipients and donors

BackgroundThe benefit of administering pharmacologic thromboprophylaxis following renal transplantation remains uncertain. ObjectivesTo compare hemostatic parameters before and after renal transplant surgery in both recipients and their donors at predetermined time points. MethodsBlood samples were collected at baseline (T1), immediately after surgery (T2), and at 24 hours after surgery (T3) in both recipients and donors and at 72 (T4) and 120 hours (T5) from recipients only. Assays included in vitro thrombin generation, factor VIII (FVIIIc) activity, von Willebrand factor (VWF) antigen, D-dimer, antithrombin activity, prothrombin fragment 1 + 2 (F1 + 2), thrombin-antithrombin complexes, and plasminogen activator inhibitor-1 (PAI-1) antigen. ResultsFifty-two patients (28 recipients and 24 donors) were enrolled. Both donors and recipients had increased FVIIIc, VWF, F1 + 2, D-dimer, and PAI immediately after surgery but reduced antithrombin. Mixed-model analysis showed that the magnitude of change over time (between T1 and T3) for FVIIIc (mean estimated difference [MED], 72; 95% CI, 41-102; P < .0001), VWF (MED, 89; 95% CI, 35-142; P = .001), F1 + 2 (MED, 283; 95% CI, 144-422; P < .0001), thrombin-antithrombin complexes (MED, 3.5; 95% CI, 1.9-5.1; P < .0001), D-dimer (MED, 2.2; 95% CI, 1.0-3.3; P < .0001), PAI-1 (MED, 9.2; 95% CI, 3.4-14.9; P = .002), and time to peak thrombin generation (MED, 1.5; 95% CI, 0.35-2.7; P = .01) was more significant in recipients than in donors. ConclusionPersistence of a hypercoagulable state was more prominent in recipients after 24 hours despite recovery in renal function and initiation of thromboprophylaxis.

Common Coagulopathies Associated With COVID-19 Patients.

The coronavirus disease 2019 (COVID-19) outbreak, which first appeared in the Chinese province of Hubei city of Wuhan, has been spreading internationally since December 2019. The World Health Organization (WHO) declared the coronavirus illness from 2019 to be a pandemic on March 11, 2020. Patients hospitalised with severe coronavirus or comorbid conditions (like cardiovascular disease and obesity) are linked to a worse prognosis. The rise in D-dimer and its relationship to prognosis are the most often documented aberrations in coagulation/fibrinolysis in COVID-19. However, the D-dimer assessment's utility is not limitless. Since the coagulation/fibrinolytic state might occasionally change over a short period of time, routine exams are also advantageous in understanding the relevance of the inquiry. Both thrombotic and hemorrhagic diseases should be taken into consideration, despite the fact that the pathophysiology of disseminated intravascular coagulation (DIC) linked with coronavirus disease 19 differs significantly from that of septic disseminated intravascular coagulation. Coagulation as well as fibrinolysis indicators are used to make the diagnosis of COVID-19 thrombosis, which encompasses both macro- and micro-thrombosis. Compared to bacterial-sepsis-associated coagulopathy/DIC, COVID-19 has a lower prevalence of prolonged prothrombin time, activated partial thromboplastin time, and decreased antithrombin activity. However, the causes of coagulopathy remain poorly understood. Hypoxia, endothelial injury, dysregulated immunological responses mediated by inflammatory cytokines, and lymphocyte cell death are thought to be implicated. While blood loss tends to be rare, it is uncertain if COVID-19 suffers from thrombosis or whether the current recommendations for regular venous thromboembolic dose are appropriate. It is important to decide on the COVID-19 therapy phases. Antiviral therapy, cytokine storm therapy, and thrombosis therapy are the steps. Future advancements are predicted, such as a therapy that combines heparin and nafamostat.

Galectin-3 as a Marker for Increased Thrombogenicity in COVID-19.

Galectin-3 is a beta-galactoside-binding lectin involved in inflammation and lung fibrosis and postulated to enhance thrombosis. In COVID-19, it is considered to be a prognostic marker of severity. The aim of this study was to evaluate whether galectin-3 is associated with thrombogenicity in COVID-19. Patients with moderate-to-severe COVID-19 (COVpos; n = 55) and patients with acute respiratory diseases, but without COVID-19 (COVneg; n = 35), were included in the study. We measured the amount of galectin-3, as well as other platelet and coagulation markers, and correlated galectin-3 levels with these markers of thrombogenicity and with the SOFA Score values. We found that galectin-3 levels, as well as von Willebrand Factor (vWF), antithrombin and tissue plasminogen activator levels, were higher in the COVpos than they were in the COVneg cohort. Galectin-3 correlated positively with vWF, antithrombin and D-dimer in the COVpos cohort, but not in the COVneg cohort. Moreover, galactin-3 correlated also with clinical disease severity, as measured by the SOFA Score. In patients with acute respiratory diseases, galectin-3 can be considered as a marker not only for disease severity, but also for increased hypercoagulability. Whether galectin-3 might be a useful therapeutic target in COVID-19 needs to be assessed in future studies.