R. van Oerle

Coagulation factors and the protein C system as determinants of thrombin generation in a normal population

Summary.  Background: Thrombin generation is a powerful tool to probe overall plasma coagulability.

Platelet‐ and erythrocyte‐derived microparticles trigger thrombin generation via factor XIIa

See also Shapiro S, Laffan M. Making contact with microparticles. This issue, pp 1352–4.

Preanalytic variables of thrombin generation: towards a standard procedure and validation of the method

Summary.  Background: Thrombin generation assays are sensitive methods for assessment of the overall clotting potential of plasma, but, despite their common use in thrombosis research, standardization of preanalytic conditions is lacking. In order to set up a standardized protocol, we analyzed different preanalytic variables and validated the calibrated automated thrombogram method.

Thrombin generation in patients with a first acute myocardial infarction

Summary.  Background: Despite improved treatment options, myocardial infarction is still an important cause of morbidity and mortality. One of the contributing mechanisms in the acute myocardial infarction (AMI) is plasma hypercoagulability. Methods: We investigated hypercoagulability in 135 (first) patients with AMI using thrombin generation (TG) testing. TG testing was performed in plasmas, drawn upon admission and before medication administration, and subsequently after 4 days, 3 and 6 months. Further, we evaluated determinants of thrombin generation using multiple regression analysis of major coagulation proteins and inhibitors. Admission TG results were also related to 1‐year outcome: cardiovascular death, recurrent myocardial infarction, a second coronary intervention [percutaneous coronary intervention (PCI) or coronary artery bypass grafting (CABG)] and ischemic stroke. Results: At day 0, the TG parameters peak height, endogenous thrombin potential (ETP) and lag time were increased compared with a reference population. Peak height and lag time stayed persistently increased in patients. The lowest half of the ETP values was statistically not significantly associated with an occurrence of endpoints. The lowest half of the ETP values combined with the upper half of the D‐dimer values were associated with endpoints; odds ratio 5.8 (1.1–30.7). Tissue factor pathway inhibitor (TFPI) seems to be an important determinant of TG in AMI and healthy persons. Conclusions: TG reflects acute hypercoagulability during AMI and partly also in the 6‐month period after the acute event. TG shows a trend of an inverse association with risk of recurrent ischemic cardiovascular complications. Unraveling mechanisms in TG might improve our understanding of the pathophysiology of AMI and direct future improvements in medical care.

Prothrombin complex concentrate reduces blood loss and enhances thrombin generation in a pig model with blunt liver injury under severe hypothermia

Although prothrombin complex concentrate (PCC) is increasingly used for the treatment of trauma-induced coagulopathy, few studies have investigated the impact and safety of PCC for this indication. The present study was performed to assess PCC for treatment of coagulopathy after blunt liver injury under severe hypothermia. Coagulopathy in 14 anaesthetised pigs was induced by haemodilution. Subsequently, standardised blunt liver injury was induced under severe hypothermia (32.8-33.2°C). Animals were randomised to receive either PCC (35 IU kg⁻¹) or saline (control). Coagulation was assessed over the following 2 hours by thromboelastometry and thrombin generation. Internal organs were examined to determine presence of emboli. The administration of PCC showed a significant reduction in blood loss (p=0.002 vs. controls) and a significant increase in the rate of survival (p=0.022 vs. controls). Plasma thrombin generation in the PCC group increased considerably above baseline levels, with significant increases in peak thrombin levels and endogenous thrombin potential versus controls throughout the follow-up period. In addition, PT decreased significantly in the PCC group versus the control group. However, only slight improvements in thromboelastometry variables were observed. Histology showed an equal degree of liver injury in both groups, and no thromboembolism. In severely hypothermic pigs, the application of PCC corrected trauma-induced coagulopathy and reduced blood loss. Thus, the infusion of PCC might be a reasonable approach to reduce the need for blood cell transfusion in trauma. Furthermore, the impact and safety of PCC application can be monitored through thrombin generation and thromboelastometry under hypothermia.

Factor XII activation is essential to sustain the procoagulant effects of particulate matter

See also Mutch NJ. Emerging roles for factor XII in vivo. This issue, pp 1355–8.

Active site inhibited factor VIIa attenuates myocardial ischemia/reperfusion injury in mice

Summary.  Background: Inhibition of specific coagulation pathways such as the factor VIIa‐tissue factor complex has been shown to attenuate ischemia/reperfusion (I/R) injury, but the cellular mechanisms have not been explored. Objectives: To determine the cellular mechanisms involved in the working mechanism of active site inhibited factor VIIa (ASIS) in the protection against myocardial I/R injury. Methods: We investigated the effects of a specific mouse recombinant in a mouse model of myocardial I/R injury. One hour of ischemia was followed by 2, 6 or 24 h of reperfusion. Mouse ASIS or placebo was administered before and after induction of reperfusion. Results: ASIS administration reduced myocardial I/R injury by more than 40% at three reperfusion times. Multiplex ligation dependent probe amplification (MLPA) analysis showed reduced mRNA expression in the ischemic myocardium of CD14, TLR‐4, interleukin‐1 (IL‐1) receptor‐associated kinase (IRAK) and IκBα upon ASIS administration, indicative of inhibition of toll‐like receptor‐4 (TLR‐4) and subsequent nuclear factor‐κB (NF‐κB) mediated cell signaling. Levels of nuclear activated NF‐κB and proteins influenced by the NF‐κB pathway including tissue factor (TF) and IL‐6 that were increased after I/R, were attenuated upon ASIS administration. After 6 and 24 h of reperfusion, neutrophil infiltration into the area of infarction was decreased upon ASIS administration. There was, however, no evidence of an effect of ASIS on apoptosis (Tunel staining and MLPA analysis). Conclusions: We conclude that the diminished amount of myocardial I/R injury after ASIS administration is primarily due to attenuated inflammation‐related lethal I/R injury, probably mediated through the NF‐κB mechanism.

Soluble tissue factor is a candidate marker for progression of microvascular disease in patients with Type 2 diabetes

Summary.  Objective: To determine the relationship between abnormalities in blood coagulation and prevalent or incident cardiovascular complications in Type 2 diabetes. Design and methods: Prospective cohort study of 128 patients with Type 2 diabetes in whom blood samples were collected at baseline and after 1 year of follow‐up. All cardiovascular complications at baseline and follow‐up were recorded. Forty‐three healthy, age‐matched subjects served as a control group. Results: Logistic analysis revealed an independent relationship between soluble tissue factor (TF) and microvascular disease [per pg mL−1 TF: Exp(B) = 1.008; CI(95%)1.002–1.014], or neurogenic disease [Exp(B) = 1.006; CI(95%)1.001–1.011]. The highest levels of soluble TF were observed in patients with microvascular and neurogenic disease (P < 0.001). Patients with Type 2 diabetes having a soluble TF concentration >300 pg mL−1 are at a 15‐fold higher risk for the presence of microvascular disease and at a 10‐fold higher risk for the presence of neurogenic disease compared with the patients with concentrations below 100 pg mL−1. Soluble TF was correlated with tissue type plasminogen activator, von Willebrand factor antigen, systolic blood pressure and age. Levels of F1′ + 2, D‐dimer, FVIII activity, t‐PA and vWFag were not different among patients with micro‐, macro‐ or neurogenic complications compared with patients without those complications. Forty‐eight new micro‐, macro‐ and/or neurogenic complications were diagnosed after 1 year follow‐up. With the exception of higher F1 + 2 levels after 1 year all other markers remained unchanged. A trend toward higher soluble TF levels was observed in patients with new microvascular events (P = 0.056). Conclusions: Soluble TF is associated with existing microvascular and neurogenic complications in patients with Type 2 diabetes and is a candidate marker for progression of microvascular disease.

Development of novel membranes for blood purification therapies based on copolymers of N-vinylpyrrolidone and n-butylmethacrylate

Developments in membrane based blood purification therapies often come with longer treatment times and therefore longer blood-material contact, which requires long-term membrane biocompatibility. In this study, we develop for the first time membranes for blood purification using the material SlipSkin™, which is a copolymer, made from N-vinylpyrrolidone (NVP) and butylmethacrylate (BMA). Specific attention is focused on understanding the mechanism of pore formation and the tailoring of the membrane mechanical and transport properties to obtain the optimal membrane for blood purification therapies. Polymer composition, solvent type and solvent evaporation time influence membrane morphology and membranes with sieving properties of cascade filters in plasma fractionation applications are developed. The new membranes have very good blood compatibility properties; in fact compared to benchmark flat membranes currently used in the clinic, they have lower platelet adhesion while all other properties (contact activation, thrombogenicity, leukocyte adhesion, hemolysis and complement activation) are also very good and comparable to the benchmarks.

ATP-gated P2X1 ion channels protect against endotoxemia by dampening neutrophil activation

Summary.  Background:  In sepsis, extracellular ATP, secreted by activated platelets and leukocytes, may contribute to the crosstalk between hemostasis and inflammation. Previously, we showed that, in addition to their role in platelet activation, ATP‐gated P2X1 ion channels are involved in promoting neutrophil chemotaxis.