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Background: Chronic kidney disease patients show changes in the endothelial surface layer (ESL). Whether hemodialysis (HD) itself or low molecular weight heparins (LMWH) induce ESL alterations is unknown. Methods: We studied the ESL in 20 HD patients with Sidestream Dark Field Imaging [measuring perfused boundary region (PBR)] and measurement of ESL constituents in plasma during HD in 2 studies. LMWH was administered at the start of HD in study A, and 120 min after the start of HD in study B. Mean platelet volume (MPV) and platelet large cell ratio (P-LCR) were also measured. Results: Syndecan-1 increased significantly 30 min after LMWH administration. sP-Selectin increased 120 min after HD start, and MPV and P-LCR decreased significantly during HD. No significant changes of PBR, sE-Selectin, sICAM-1, or sVCAM-1 were perceived. Conclusions: HD caused a significant increase in Syndecan-1 without a change in PBR. The administration of LMWH appeared to precede the rise in Syndecan-1. © 2014 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background: The use of PCC for the treatment of trauma-induced coagulopathy potentially increase the risk of thromboembolism and disseminated intravascular coagulation, which is addressed to an imbalance of both pro- and anticoagulants. As PCCs differ in composition, we used an in vitro dilutional approach to assess the overall thrombin generation of five different PCCs through various laboratory assays. Methods: The vitamin K-dependent coagulation factors, heparin, and antithrombin were assessed in five commercially available PCCs. The procoagulant potential of the PCCs was assessed in plasma and whole blood from 4 healthy donors by means of classical coagulation assays, thrombin generation assay and thromboelastometry. In order to reflect coagulopathy, whole blood was diluted to 80, 60, 40, and 20% with Ringer’s lactate solution. Results: The five different PCCs were characterised by comparable levels of factors II, VII, IX and X (all around 20–30 IU/mL), whereas the heparin (0 to 17.6 IU/mL) and antithrombin (0.06 to 1.29 IU/mL) levels were remarkably different between manufactures. In vitro dilution of blood induced a prolongation of the PT and aPTT, and attenuation of thrombin generation and ExTem induced thromboelastometry. Overall, non- or low-heparin containing PCCs restored the in vitro dilutional coagulopathy, whereas PCCs containing heparin have an anticoagulant effect. The thrombin generation assay showed to be the most sensitive method for assessment of PCC effects. Conclusions: This study shows that most available PCCs are not balanced regarding their pro- and anticoagulants. The effect of measured differences in thrombin generation among different PCCs requires further investigations to elaborate the clinical meaning of this finding in the treatment of trauma induced coagulopathy. [ABSTRACT FROM AUTHOR]

Background: Variations in the blood coagulation activity, determined genetically or by medication, may alter atherosclerotic plaque progression, by influencing pleiotropic effects of coagulation proteases. Published experimental studies have yielded contradictory findings on the role of hypercoagulability in atherogenesis. We therefore sought to address this matter by extensively investigating the in vivo significance of genetic alterations and pharmacologic inhibition of thrombin formation for the onset and progression of atherosclerosis, and plaque phenotype determination. Methodology/principal findings: We generated transgenic atherosclerosis-prone mice with diminished coagulant or hypercoagulable phenotype and employed two distinct models of atherosclerosis. Gene-targeted 50% reduction in prothrombin \((FII^{−/WT}:ApoE^{−/−})\) was remarkably effective in limiting disease compared to control \(ApoE^{−/−}\) mice, associated with significant qualitative benefits, including diminished leukocyte infiltration, altered collagen and vascular smooth muscle cell content. Genetically-imposed hypercoagulability in \(TM^{Pro/Pro}:ApoE^{−/−}\) mice resulted in severe atherosclerosis, plaque vulnerability and spontaneous atherothrombosis. Hypercoagulability was associated with a pronounced neutrophilia, neutrophil hyper-reactivity, markedly increased oxidative stress, neutrophil intraplaque infiltration and apoptosis. Administration of either the synthetic specific thrombin inhibitor Dabigatran etexilate, or recombinant activated protein C (APC), counteracted the pro-inflammatory and pro-atherogenic phenotype of pro-thrombotic \(TM^{Pro/Pro}:ApoE^{−/−}\) mice. Conclusions/significance: We provide new evidence highlighting the importance of neutrophils in the coagulation-inflammation interplay during atherogenesis. Our findings reveal that thrombin-mediated proteolysis is an unexpectedly powerful determinant of atherosclerosis in multiple distinct settings. These studies suggest that selective anticoagulants employed to prevent thrombotic events may also be remarkably effective in clinically impeding the onset and progression of cardiovascular disease.

Background: COVID-19 associated coagulopathy (CAC) is associated with an increase in thromboembolic events. Current guidelines recommend prophylactic heparins in the management of CAC. However, the efficacy of this strategy in the intensive care population remains uncertain., Objective: We aimed to measure thrombin generation (TG) to assess CAC in intensive care unit (ICU) patients receiving thromboprophylaxis with low molecular weight heparin (LMWH) or unfractionated heparin (UFH). In addition, we performed statistical modeling to link TG parameters to patient characteristics and clinical parameters. Lastly, we studied the potency of different anticoagulants as an alternative to LMWH treatment in ex vivo COVID-19 plasma., Patients/methods: We included 33 patients with confirmed COVID-19 admitted at the ICU. TG was measured at least twice over the course of 6 weeks after admission. Thrombin generation parameters peak height and endogenous thrombin potential (ETP) were compared to healthy controls. Results were subsequently correlated with a patient characteristics and laboratory measurements. In vitro spiking in TG with rivaroxaban, dabigatran, argatroban and orgaran was performed and compared to LMWH., Results: Anti-Xa levels of all patients remained within the therapeutic range throughout follow-up. At baseline, the mean (SE) endogenous thrombin potential (ETP) was 1,727 (170) nM min and 1,620 (460) nM min for ellagic acid (EA) and tissue factor (TF), respectively. In line with this we found a mean (SE) peak height of 353 (45) nM and 264 (96) nM for EA and TF. Although fluctuating across the weeks of follow-up, TG parameters remained elevated despite thromboprophylaxis. In vitro comparison of LMWHs and direct thrombin inhibitors (e.g., agratroban, dabigatran) revealed a higher efficacy in reducing coagulation potential for direct thrombin inhibition in both ellagic acid (EA) and tissue factor (TF) triggered TG., Conclusion: In a sub-group of mechanically ventilated, critically ill COVID-19 patients, despite apparent adequate anti-coagulation doses evaluated by anti-Xa levels, thrombin generation potential remained high during ICU admission independent of age, sex, body mass index, APACHE II score, cardiovascular disease, and smoking status. These observations could, only partially, be explained by (anti)coagulation and thrombosis, inflammation, and multi-organ failure. Our in vitro data suggested that direct thrombin inhibition compared with LMWH might offer an alternate, more effective anticoagulant strategy in COVID-19., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 van de Berg, Mulder, Alnima, Nagy, van Oerle, Beckers, Hackeng, Hulshof, Sels, Henskens, van der Horst, ten Cate, Spronk, van Bussel and MaastrICCht Collaborators.)

Thrombin generation may be a potential tool to improve risk stratification for cardiovascular diseases. This study aims to explore the relation between thrombin generation and cardiovascular risk factors, cardiovascular diseases, and total mortality. For this study, N=5000 subjects from the population-based Gutenberg Health Study were analysed in a highly standardized setting. Thrombin generation was assessed by the Calibrated Automated Thrombogram method at 1 and 5 pM tissue factors trigger in platelet poor plasma. Lag time, endogenous thrombin potential, and peak height were derived from the thrombin generation curve. Sex-specific multivariable linear regression analysis adjusted for age, cardiovascular risk factors, cardiovascular diseases and therapy, was used to assess clinical determinants of thrombin generation. Cox regression models adjusted for age, sex, cardiovascular risk factors and vitamin K antagonists investigated the association between thrombin generation parameters and total mortality. Lag time was positively associated with obesity and dyslipidaemia for both sexes (p<0.0001). Obesity was also positively associated with endogenous thrombin potential in both sexes (p<0.0001) and peak height in males (1 pM tissue factor, p=0.0048) and females (p<0.0001). Cox regression models showed an increased mortality in individuals with lag time (1 pM tissue factor, hazard ratio=1.46, [95% CI: 1.07; 2.00], p=0.018) and endogenous thrombin potential (5 pM tissue factor, hazard ratio = 1.50, [1.06; 2.13], p=0.023) above the 95th percentile of the reference group, independent of the cardiovascular risk profile. This large-scale study demonstrates traditional cardiovascular risk factors, particularly obesity, as relevant determinants of thrombin generation. Lag time and endogenous thrombin potential were found as potentially relevant predictors of increased total mortality, which deserves further investigation.

The process of thrombin generation involves numerous plasma proteases and cofactors. Interaction with the vessel wall, in particular endothelial cells (ECs), influences this process but data on this interaction is limited. We evaluated thrombin generation on EA.hy926, human coronary arterial ECs (HCAECs) and patient-derived human venous ECs (HVECs) by means of a modified calibrated automated thrombogram (CAT) method and especially looked into contribution of the intrinsic and extrinsic pathways. Thrombin generation was measured in presence of confluent ECs with normal pooled and factor XII-deficient (FXII-deficient) platelet-poor plasma, with/without active site inhibited factor VIIa (ASIS) to block the extrinsic pathway and corn trypsin inhibitor for blocking contact activation (intrinsic pathway). Fetal bovine serum (FBS) was removed from culture conditions as FXIIa from the serum retained on ECs apparently, thereby inducing strong contact activation. In serum-free conditions, EA.hy926 and patient-derived HVECs induced thrombin generation mainly via the contact activation pathway with minor influence of ASIS on peak height and very low thrombin generation curves in FXII-deficient plasma. HVECs derived from coronary arterial bypass graft (CABG) patients showed increased thrombin generation compared to control patients, which could be ascribed to increased contact activation. Contribution of the extrinsic pathway on patient-derived ECs was limited. We conclude that the CAT method in combination with serum-free cultured ECs offers a valuable high-throughput method to evaluate endothelial influences on thrombin generation, which appears to involve predominantly contact activation on ECs. Contact activation-mediated thrombin generation was increased on ECs from CABG patients compared to controls.