Thrombosis and haemostasis最新文献

A DOAC concentration threshold above which an impact on surgical hemostasis starts to occur is unknown. Thrombin generation assays (TGAs) provide a measure of the coagulation phenotype. This study aimed to determine whether preoperative TGA parameters are associated with postoperative bleeding, and whether this is partly due to residual DOAC levels. We conducted a nested case-control study using samples from apixaban/rivaroxaban-treated patients with atrial fibrillation from the PAUSE perioperative study. Cases were participants with postoperative major or clinically relevant non-major bleeding; controls were participants without bleeding. TGA parameters were measured using Calibrated Automated Thrombography (5 pM tissue factor). Generalized linear mixed models and causal mediation analyses were used to evaluate the relationship between DOAC levels, TGA parameters, and bleeding. Forty-eight cases were matched to 474 controls. Residual DOAC levels were higher in cases than controls (p = 0.03) and TGA parameters were correlated with residual DOAC levels (p < 0.05). A longer lag time (LT; OR = 1.319 per minute [95%CI 1.077-1.617]) and time-to-peak (TTP; OR = 1.154 per minute [95%CI 1.028-1.296]) were associated with an increased odds of bleeding; higher peak (OR = 0.994 per nM [95%CI 0.989-0.998]) and higher mean velocity rate index (mVRI; OR = 0.986 per nM/min [95%CI 0.976-0.996]) were associated with a lower odds of bleeding. The effect of apixaban/rivaroxaban levels on bleeding was mediated by altered TGA parameters (LT, TTP, peak, mVRI). TGA parameters are associated with postoperative bleeding and the impact of DOAC levels on bleeding is mediated by effects on thrombin generation.

Background:  The Hemophilia Functional Ability Scoring Tool (Hemo-FAST), consisting of a patient-reported outcome (PRO) part and a clinician-reported outcome (ClinRO) part, was developed as a rapid and effective tool to assess functional mobility in clinical practice. This study (NCT04731701) aimed to validate the psychometric properties of Hemo-FAST for assessment of joint health in people with haemophilia (PwH).

Methods:  PwH A or B aged ≥18 years completed questionnaires including the PRO part of Hemo-FAST and the short-form 36 health survey (SF-36) during one study visit. Clinicians completed the Haemophilia Joint Health Score (HJHS) and the ClinRO part of Hemo-FAST at the same visit. Validation was performed using reliability, construct validity, and structure validity assessments.

Results:  The study enrolled 180 PwH A or B from 14 centres across France. Estimated time to complete the PRO part was mean (standard deviation) 4.6 (5.4) minutes. PRO items showed good test-retest reliability (intraclass correlation coefficient value ≥0.70). Inter-rater values were >0.70 for 7/9 ClinRO items, indicating good reliability. All items (15 PRO; 9 ClinRO) had high internal consistency (Cronbach's coefficient alpha: 0.97). Hemo-FAST demonstrated convergent construct validity with HJHS and the SF-36 physical component and discriminant construct validity with the SF-36 mental health component. Hemo-FAST scores distinguished between subgroups of people with expected differences in joint health status, including by haemophilia severity (p < 0.0001).

Conclusion:  This study successfully validated Hemo-FAST as a rapid and reliable tool for the functional assessment of joint health in adults with haemophilia, both in clinical practice and clinical research settings.

Background:  We previously identified a factor (F)VIII molecular defect associated with an R2159C mutation in the C1 domain (named "FVIII-Ise") together with undetectable FVIII antigen (FVIII:Ag) levels measured by two-site sandwich ELISA using an anti-C2 domain alloantibody (alloAb). The patient had clinically mild hemophilia A, and his reduced FVIII:C correlated with FVIII:Ag measured by ELISA using monoclonal antibodies (mAbs) with A2 and A2/B domain epitopes, suggesting that the R2159C mutation modified C2 domain antigenicity.

Aim:  To investigate functional and structural characteristics of the FVIII-R2159C mutant.

Methods and results:  ELISAs using a previous anti-C2 domain alloAb confirmed that the antigen level of recombinant FVIII-R2159C mutant prepared in BHK cells was 56% lower relative to wild-type (WT), consistent with our earlier reports. This anti-C2 domain alloAb competitively inhibited FVIII and anti-C1 domain mAb binding, indicating the involvement of specificity for C1 and C2 epitopes. The K _m for FVIII-R2159C with FIXa or FX in the tenase complex was similar to that of FVIII-WT. Thrombin- and FXa-catalyzed cleavage reactions of FVIII-R2159C were similar to those of WT. The K _d for FVIII-R2159C binding to phospholipids was moderately greater than for FVIII-WT, however, while there were no significant differences in von Willebrand factor binding. In silico molecular dynamic simulation analyses revealed subtle differences between FVIII-WT and FVIII-R2159C.

Conclusion:  The FVIII-R2159C mutation was not different from FVIII-WT in interactions with FIXa, FX, and thrombin, but reduced binding potential to phospholipids and to an anti-C1/C2 domain alloAb was evident apparently due to subtle changes in conformational structure.

Background:  To evaluate residual fibrinolysis resistance activity (FRA) in plasma, a detergent-modified plasma clot lysis assay time (dPCLT) was established in which α2-antiplasmin (A2AP) and plasminogen activator inhibitor type 1 (PAI-1) are inactivated without impacting protease activity. We applied this novel assay to severely injured trauma patients' plasma.

Material and methods:  Tissue-type plasminogen activator (tPA)-induced plasma clot lysis assays were conducted after detergents- (dPCLT) or vehicle- (sPCLT) treatment, and time to 50% clot lysis was measured ("transition midpoint", T _m). Residual FRA was then calculated as ([sPCLT T _m] - [dPCLT T _m]/[sPCLT T _m]) x100% = Δ T_m PCLT (%). Assay results were compared to rapid thromboelastography (TEG) LY30, tPA TEG LY30, and plasma fibrinolysis biomarkers in polytrauma patients' plasma (N=43).

Results:  Δ T_m PCLT(%) in normal plasma (N=5) was 63.0 ± 8.3 whereas in A2AP-depleted plasma was -19.1 ± 1.3%, Plasmin-antiplasmin (PAP) complex increased after complete lysis of sPCLT, whereas that in dPCLT was negligible in normal plasma. In trauma plasma, significant correlations between Δ T_m PCLT and active PAI-1 (r = 0.85, p<0.0001), PAP complex (r = -0.85, p<0.0001), free A2AP (r = 0.66, p<0.0001), total A2AP levels (r = 0.52, p=0.001) and tPA TEG LY30 (r = -0.85, p<0.0001) were found. dPCLT in hyperfibrinolysis patients diagnosed by tPA TEG was significantly shorter than those with low fibrinolysis [10.2 ± 6.4 minutes versus 20.2 ± 2.1 minutes, p=0.0006].

Conclusion:  Hyperfibrinolysis after trauma is significantly related to exhaustion of FRA, and our novel assay appears to quickly assess this state and may be a useful clinical diagnostic after additional validation.

Key points: · We established a new clot lysis assay to measure residual fibrinolysis resistance activity after inactivating PAI-1 and A2AP by detergents without impacting protease function.. · This novel clot lysis assay unmasked the mechanism of hyperfibrinolysis after trauma as exhaustion of fibrinolysis resistance activity, and appeared useful in quickly identifying these patients..

Background:  Neutrophil extracellular traps can contribute to thrombosis via stabilization of fibrin network, which is normally conducted by plasma transglutaminase, Factor XIII-A as part of coagulation cascade. The possible presence and activity of FXIII-A in neutrophils or during NETosis are unknown. Here, we investigated potential presence of FXIII-A in neutrophils and participation in NET-fibrin(ogen) interaction in vitro. METHODS:  Data mining of human and mouse F13A1/F13a1 mRNA expression in whole-body scRNA sequence atlases was conducted. F13a1 mRNA and protein expression was assessed in isolated mouse bone marrow neutrophils. NETosis was induced using 12-phorbol 13-myristate acetate (PMA), and the transglutaminase activity was assessed with 5-(biotinamido)pentylamine incorporation to plasma fibronectin and a fluorescence-fibrin(ogen)-based activity assay using ATTO488-Cadaverine. Externalization of FXIII-A and its interaction with neutrophil extracellular trap (NET) markers, namely, decondensed DNA, CitH3, and MPO, were examined with immunofluorescence microscopy. NET-fibrin(ogen) interaction was investigated with and without serum and/or transglutaminase inhibitor, NC9. Effect of soluble fibrinogen and fibrin(ogen) network on NETosis was also assessed.

Results:  Data mining of RNAseq atlases showed F13A1/F13a1 expression in adipose tissue, blood, and bone marrow neutrophils. mRNA expression and protein production were confirmed in isolated neutrophils where expression was comparable to that of macrophages and monocytes. FXIII-A was externalized and active as a transglutaminase and colocalized with NET markers during NETosis. FXIII-A transglutaminase activity promoted NET-fibrin(ogen) interaction and entrapment of neutrophils within fibrin(ogen) matrix. Soluble fibrinogen or fibrin(ogen) network did not induce NETosis.

Conclusion:  This study identifies neutrophils as a source of FXIII-A and suggests its role in stabilizing NET-fibrin(ogen) matrix structures.

The McMaster Immune Thrombocytopenia (ITP) Summit, held on October 27, 2023, was an educational seminar from leading experts in immune thrombocytopenia and related disorders geared toward hematologists, internists, immunologists, and clinical and translational scientists. The focus of the Summit was to review the mechanisms, diagnosis, and treatment of primary versus secondary ITP. Specific objectives were to describe the unique features of secondary ITP, and to review its mechanisms in the context of autoimmune disease and infection. The key messages in this Summit were: (1) ITP is a heterogeneous disease, and genetic and immunologic insights may help classify patient subtypes; (2) exploring the autoimmune mechanisms and their association with hypogammaglobulinemia in patients with secondary ITP could improve our understanding of ITP and its subtypes; (3) investigating the mechanisms of ITP in the context of infections caused by viruses such as CMV, HIV, dengue, and hepatitis C, or bacteria such as H. pylori, or vaccinations could provide insight into the causes of ITP. A better understanding of secondary ITP could help elucidate the pathogenesis of ITP.

Background:  Portal vein system-specific risk factors contributing to portal vein thrombosis in cirrhosis are poorly investigated.

Aim:  This study aimed to quantify contact system and intrinsic pathway activation in the peripheral compared to portal venous blood in patients with decompensated cirrhosis.

Methods:  Adult patients with cirrhosis undergoing transjugular intrahepatic portosystemic shunt underwent simultaneous blood sampling from a peripheral vein and the portal vein. Complexes of serine proteases with their respective inhibitors were measured by ELISA to quantify contact system (PKa:C1-INH [plasma kallikrein:C1-esterase inhibitor] and FXIIa:C1-INH) and intrinsic pathway activation (FXIa:C1-INH, FXIa:α1at [α-1 antitrypsin], FXIa:AT [antithrombin], and FIXa:AT).

Results:  Twenty patients with cirrhosis (mean age 55 ± 7 years, M = 58%, Child-Pugh A/B/C 6/11/3) and 25 healthy controls (mean age 45 ± 12 years, M = 60%) were enrolled. The etiology of cirrhosis was primarily alcohol abuse, followed by chronic viral infection. Log-transformed peripheral levels of all the complexes were significantly higher in patients compared with controls. While levels of PKa:C1-INH, FXIIa:C1-INH, FXIa:C1-INH and FXIa:α1at were similar in peripheral and portal venous blood in cirrhotic patients, FXIa:AT and FIXa:AT levels were significantly higher in portal blood (p = 0.013 and 0.011, respectively). FXIa:C1-INH significantly correlated with both contact system complexes (FXIIa:C1-INH and PKa:C1-INH) and with FIX:AT.

Conclusion:  Markers of contact system and intrinsic pathway activation in the systemic circulation were significantly higher in cirrhosis versus controls. Complexes of FXIa and FIXa with AT were significantly higher in the portal than in peripheral plasma in cirrhosis, possibly indicating a unique heparin-like effect in portal venous blood.

Background:  Mendelian mutations in the Prothrombin gene (F2) and the factor V Leiden gene (F5) genes are established risk factors for venous thromboembolism (VTE). Walking pace is associated with the risk of coronary artery diseases, but no study has investigated its association with VTE. This study aimed to investigate the association and causality between walking pace and VTE, compare its population risk with established Mendelian mutations, and determine if blood biomarkers mediate its effect.

Methods:  We followed up 445,261 UK Biobank participants free of VTE at baseline. Self-reported walking pace was collected via touchscreen questionnaire at baseline. The carrier status of two Mendelian mutations in F2 and F5 genes was determined by the genotypes of rs1799963 (G20210A, c.*97 G > A) and rs6025 (p.R534Q), respectively. Cox proportional hazard model was used to estimate the effect of walking pace on incident VTE. We conducted a bidirectional Mendelian randomization (MR) analysis, by using 70 single-nucleotide polymorphisms (SNPs) from a walking pace genome-wide association studies (GWAS) and 93 SNPs from a VTE GWAS as instrumental variables. We used both individual-level data and GWAS summary statistics for mediation analysis.

Results:  Over a median follow-up period of 12.8 years, 11,155 incident VTE cases were identified. The 10-year incidence rates for brisk and slow walking pace were 1.32% (confidence interval [CI]: 1.27-1.37%) and 3.90% (CI: 3.71-4.09%), respectively. For noncarriers, F2 and F5 carriers, the 10-year incidence rates were 1.70% (CI: 1.66-1.73%), 2.94% (CI: 2.66-3.22%), and 3.62% (CI: 3.39-3.84%), respectively. The overall risk of VTE for F5 mutation carriers with a brisk walking pace (2.65%) was smaller than that for noncarriers with a slow walking pace (3.66%). For F5 mutation carriers, brisk pace (but not steady pace) reduces the risk of VTE (p interaction < 0.05). MR analyses displayed a causal relationship (inverse variance weighted: p = 3.21 × 10^-5) from walking pace to VTE incidence. Mediation analysis showed that serum albumin (ALB) and cystatin C (CYS) levels partially mediated the effect of brisk walking pace on the risk of VTE incidence, with mediation proportions of 8.7 to 11.7%, respectively.

Conclusion:  On the population scale, the protective effect of brisk walking pace offsets the risk of VTE caused by Mendelian mutations. We provided preliminary evidence that a brisk walking pace causally reduces the risk of VTE. Serum ALB and CYS partially mediate this effect.

Background:  The benefits and risks of extending anticoagulant treatment beyond the first 3 to 6 months in patients with venous thromboembolism (VTE) in clinical practice are not well understood.

Methods:  ETNA-VTE Europe is a prospective, noninterventional, post-authorization study in unselected patients with VTE treated with edoxaban in eight European countries for up to 18 months. Recurrent VTE, major bleeding, and all-cause death were the primary study outcomes.

Results:  The median age of the 2,644 patients was 65 years; 46.6% were female, and 22.8% had a history of VTE. The median treatment duration was 50.6 weeks (interquartile range: 23.4-77.7). VTE recurrence occurred in 100 patients (3.8% at an annual rate of 2.7%/year); 37 patients (1.4%) were on edoxaban at the time of the event, with a corresponding annualized rate of 1.6%/year. Major bleeding was experienced by 37 patients (1.4%) during edoxaban treatment, corresponding to an annualized rate of 1.5%/year. Overall, 95 patients died (3.6%; annualized rate 2.6%/year), with the majority for reasons other than VTE- and cardiovascular (CV)-related causes. Out of 15 deaths (1.9%; annualized rate 2.1%/year) that occurred during edoxaban treatment, 1 was related to VTE and 11 related to CV (annualized rate 0.0%/year and 0.5%/year).

Conclusions:  ETNA-VTE Europe provides evidence for the real-world effectiveness of edoxaban treatment (up to 18 months) based on a low rate of VTE recurrence, all-cause death, and major bleeding, and is aligned with the results of the randomized clinical trial reassuring the use of edoxaban in the treatment of VTE in routine clinical practice.