Background
Most tissue factor (TF) activity assays are based on measurement of factor X (FX) activation by TF in the presence of factor VII (FVII)/FVIIa. This requires long incubation, which may ...result in TF‐independent activity of FX and inaccurate measurement of TF activity.
Aim
To develop a sensitive and specific TF activity assay, which does not register a non‐specific TF activity, using commercial coagulation factors.
Methods
Tissue factor activity was measured based on the ability of TF to accelerate the activation of FX by FVIIa in the presence of factor V (FV)/Va, prothrombin, and phospholipids. Following 4 min incubation at 37°C, TF activity was quantified in test samples of different nature by thrombin generation using a chromogenic substrate.
Results
The TF activity assay proved high sensitivity (low fM range) and specificity, assessed by neutralization of TF activity by anti‐TF antibody and the use of FVIIai. TF activity was detected in extracellular vesicles (EVs) derived from HAP1‐TF+cells, while no activity was measured in EVs from HAP1‐TF/KO cells. The assay was applicable for measurement of TF activity on the surface of live endothelial cells and monocytes activated in vitro, and cell lysates. Infusion of low dose lipopolysaccharide (2 ng/kg bodyweight endotoxin) caused a transient 8‐fold increase (peaked at 4 h) in TF activity in EVs isolated from plasma of healthy volunteers.
Conclusion
Our assay provides a fast, sensitive, and specific measurement of TF activity. It reliably quantifies TF activity on cell surface, cell lysate, and isolated EVs. The assay can be used for laboratory and clinical research.
Background
Plasminogen activator inhibitor‐1 (PAI‐1), the main inhibitor of fibrinolysis, is frequently elevated in obesity and could potentially mediate the risk of venous thromboembolism (VTE) in ...obese subjects. However, whether PAI‐1 is associated with VTE remains uncertain.
Objective
To investigate the association between plasma PAI‐1 levels and risk of future incident VTE and whether PAI‐1 could mediate the VTE risk in obesity.
Methods
A population‐based nested case‐control study, comprising 383 VTE cases and 782 age‐ and sex‐matched controls, was derived from the Tromsø Study cohort. PAI‐1 antigen levels were measured in samples collected at cohort inclusion. Logistic regression was used to calculate odds ratios (ORs) with 95% confidence intervals (CIs) for VTE across PAI‐1 tertiles.
Results
The VTE risk increased dose‐dependently across PAI‐1 tertiles (P for trend <.001) in the age‐ and sex‐adjusted model. The OR of VTE for the highest versus lowest tertile was 1.73 (95% CI 1.27–2.35), and risk estimates were only slightly attenuated with additional stepwise adjustment for body mass index (BMI; OR 1.59, 95% CI 1.16–2.17) and C‐reactive protein (CRP; OR 1.54, 95% CI 1.13–2.11). Similar results were obtained for provoked/unprovoked events, deep vein thrombosis, and pulmonary embolism. In obese subjects (BMI of ≥30 kg/m2 vs. <25 kg/m2), PAI‐1 mediated 14.9% (95% CI 4.1%‐49.4%) of the VTE risk in analysis adjusted for age, sex, and CRP.
Conclusion
Our findings indicate that plasma PAI‐1 is associated with increased risk of future incident VTE and has the potential to partially mediate the VTE risk in obesity.
Identification of individuals with ischemic stroke at particularly high risk of venous thromboembolism (VTE) is crucial for targeted thromboprophylaxis. To guide clinical decision‐making and ...development of risk prediction models, increased knowledge on risk factors and biomarkers is needed. Therefore, we set out to identify risk factors and predictors for VTE in people with ischemic stroke by conducting a systematic review of the literature. Medline and Embase were searched from January 1990 and onwards. Studies investigating demographic, clinical, and/or laboratory factors for stroke‐related VTE were considered. Two reviewers screened all retrieved records, independently and in duplicate. Risk of bias assessments were guided by a structured framework (PROSPERO‐ID: CRD42020176361). Of 4674 identified records, 26 studies were included. Twenty‐six demographic, clinical, and laboratory factors associated with increased risk of stroke‐related VTE after multivariable adjustments were identified. The following factors were reported by ≥2 studies: prior VTE, cancer, prestroke disability, leg weakness, increasing lesion volume of the brain infarct, infection, low Barthel Index, increasing length of hospital stay, biochemical indices of dehydration, as well as elevated levels of D‐dimer, C‐reactive protein, and homocysteine. The majority of the studies were of poor quality with moderate or high risk of bias. In conclusion, this systematic review informs on several potential risk factors and predictors for VTE in people with ischemic stroke. To improve risk stratification and guide development of risk prediction models, further confirmation is needed because there were few high‐quality studies on each factor.
Background
Although venous thromboembolism (VTE) is a leading cause of morbidity and mortality, and socioeconomic status (SES) affects human health and health behavior, few studies have examined the ...association between SES and VTE.
Objectives
We aimed to investigate the association between SES, assessed individually and in a composite score by levels of education, income, and employment status, and incident VTE.
Methods
We used Danish national registries to identify 51 350 persons aged 25–65 years with incident VTE during 1995–2016. For each case, we used incidence density sampling to select five age‐, sex‐, and index‐year‐matched controls from the general Danish population (n = 256 750). SES indicators, including education, income, and employment status, were assessed 1 and 5 years before the VTE. We used conditional logistic regression to compute odds ratios (ORs) with 95% confidence intervals (CIs) for VTE according to individual SES indicators and a composite SES score in analyses adjusted for age, sex, and comorbidities.
Results
Compared with low levels, high educational level (OR 0.74; 95% CI 0.71–0.77), high income (OR 0.70; 95% CI 0.68–0.72), and high employment status (OR 0.66; 95% CI 0.64–0.68) were associated with decreased risk of VTE, even after adjusting for comorbidities. A composite SES score was superior to the individual indicators in assessing VTE risk (OR for high vs. low score: 0.61; 95% CI 0.59–0.63). In sensitivity analysis with SES indicators measured 5 years before the VTE, the risk estimates remained essentially the same.
Conclusion
High levels of both individual SES indicators and a composite SES score were associated with decreased VTE risk.
Background
Microvesicles (MVs) are small double‐membrane encapsulated particles shed from cells. Case‐control studies have reported elevated plasma levels of platelet‐derived MVs (PDMVs) in patients ...with venous thromboembolism (VTE). However, it is not known whether high PDMV levels is a risk factor or a consequence of the acute VTE event.
Objectives
To investigate the association between PDMVs in plasma and risk of future incident VTE.
Methods
We performed a population‐based nested case‐control study with 314 VTE cases and 705 age‐ and sex‐matched controls (from The Tromsø Study) to investigate the association between the proportion of PDMVs (PDMVs%) in plasma and risk of future incident VTE. MVs isolated from plasma sampled at baseline (i.e., before VTE) were stained for platelet markers and analyzed by flow cytometry. PDMVs% were defined as the number of PDMVs divided by the total number of MVs. Odds ratios (ORs) with 95% confidence intervals (CI) for VTE risk were estimated across quartiles of PDMVs%.
Results
Subjects with PDMVs% in the highest quartile had an OR for VTE of 1.78 (95% CI: 1.21–2.64) and 1.99 (95% CI: 1.24–3.26) for provoked VTE, compared to those in the lowest quartile. The association was moderately affected by multivariable adjustment for age, sex, body mass index, C‐reactive protein, platelet count, and cancer. The OR for VTE was higher when the time between blood sampling and event was shorter.
Conclusions
Our results show that high proportions of PDMVs are associated with future risk of incident VTE and imply a role of platelet activation in the pathogenesis of VTE.
Background
Several hemostatic factors and inflammatory markers are associated with the risk of incident venous thromboembolism (VTE), however, most existing data are from case‐control studies in ...Caucasian populations.
Objectives
We aimed to prospectively confirm previous findings and explore less studied biomarkers in relation to VTE risk in a multi‐racial/multi‐ethnic cohort.
Methods
Circulating levels of factor VIII, fibrinogen, D‐dimer, plasmin‐antiplasmin complex (PAP), C‐reactive protein (CRP), and interleukin‐6 (IL‐6) were measured at baseline (2000–2002) in 6706 participants of the Multi‐Ethnic Study of Atherosclerosis. Incident VTE was identified using hospitalization discharge codes from baseline to December 31, 2015. Hazard ratios (HRs) of VTE were estimated in Cox regression models.
Results
There were 227 events during a median of 14 years of follow‐up. Compared with participants in the lowest quartile, the HRs for those above the 95th percentile and p for trend across categories were 3.50 (95% confidence interval CI 1.98–6.19; p < .001) for D‐dimer, 1.49 (95% CI 0.84–2.63; p = .02) for factor VIII, 1.32 (95% CI 0.76–2.28; p = .99) for fibrinogen, 1.92 (95% CI 1.08–3.42; p = .15) for PAP, 1.68 (95% CI 0.81–3.48; p = .08) for CRP, and 2.55 (95% CI 1.15–5.66; p = .07) for IL‐6, after adjustment for demographics and body mass index. For CRP and IL‐6, follow‐up was restricted to 10 years because of violations of the proportional hazards assumption. No significant interactions by age/ethnicity were observed.
Conclusions
We demonstrated a fairly novel association between PAP and risk of incident VTE, and contributed further prospective confirmation regarding the associations of D‐dimer, factor VIII, and IL‐6 with VTE.
Patients with acute venous thromboembolism (VTE) require anticoagulant therapy to prevent recurrent VTE and death, which exposes them to an inherent increased risk of bleeding. Identification of ...patients at high risk of bleeding, and mitigating this risk, is an essential component of the immediate and long‐term therapeutic management of VTE. The bleeding risk can be estimated by either implicit judgment, weighing individual predictors (clinical variables or biomarkers), or by risk prediction tools developed for this purpose. Management of bleeding risk in clinical practice is, however, far from standardized. International guidelines are contradictory and lack clear and consistent guidance on the optimal management of bleeding risk. This report of the ISTH subcommittee on Predictive and Diagnostic Variables in Thrombotic Disease summarizes the evidence on the prediction of bleeding in VTE patients. We systematically searched the literature and identified 34 original studies evaluating either predictors or risk prediction models for prediction of bleeding risk on anticoagulation in VTE patients. Based on this evidence, we provide recommendations for the standardized management of bleeding risk in VTE patients.
Background
The role of combined prothrombotic genotypes in cancer‐related venous thromboembolism (VTE) is scarcely studied. We aimed to investigate the impact of a 5‐single nucleotide polymorphism ...(SNP) score on the risk of VTE in patients with and without cancer using a population‐based case‐cohort.
Methods
Cases with a first VTE (n = 1493) and a subcohort (n = 13 072) were derived from the Tromsø Study (1994‐2012) and the Nord‐Trøndelag Health Study (1995‐2008). Five SNPs previously reported as a risk score were genotyped: ABO (rs8176719), F5 (rs6025), F2 (rs1799963), FGG (rs2066865), and F11 (rs2036914). Hazard ratios (HRs) for VTE were estimated according to cancer status and the number of risk alleles in the 5‐SNP score (0‐1, 2‐3, and ≥4 alleles).
Results
During a median follow‐up of 12.3 years, 1496 individuals were diagnosed with cancer, of whom 232 experienced VTE. The VTE risk increased with the number of risk alleles in the 5‐SNP score among subjects without and with cancer. In cancer‐free subjects, the HR was 2.17 (95% confidence interval CI 1.79‐2.62) for ≥4 versus 0‐1 risk alleles. In cancer patients, the corresponding HR was 1.93 (95% CI 1.28‐2.91). The combination of cancer and ≥4 risk alleles yielded a 17‐fold (HR 17.1, 95% CI 12.5‐23.4) higher risk of VTE compared with cancer‐free subjects with 0‐1 risk alleles.
Conclusion
The risk of VTE increases with the number of prothrombotic risk alleles in subjects with and without cancer, and the combination of prothrombotic risk alleles and cancer leads to a highly elevated risk of VTE.
Background
Cardiorespiratory fitness (CRF) is a strong predictor of future arterial cardiovascular disease and premature mortality. However, there are limited data on the association between CRF and ...the risk of incident venous thromboembolism (VTE).
Objectives
To investigate whether estimated CRF (eCRF) was associated with the risk of incident VTE in a cohort recruited from the general population.
Methods
Participants (n = 10 393) from the sixth survey of the Tromsø Study (2007—08) were included, and incident VTEs were recorded up to 31 December 2016. CRF was estimated in sex‐specific algorithms based on age, waist circumference, resting heart rate, and self‐reported physical activity. Hazard ratios (HRs) with 95% confidence intervals (CIs) of VTE according to categories of eCRF were estimated in Cox regression models adjusted for sex with age as timescale. The impact of weight status was evaluated in analyses stratified by weight category.
Results
There were 176 incident VTEs during follow‐up. Compared with individuals with eCRF < 85% of age‐predicted, those with eCRF of 85% to 100% and >100% of age‐predicted had 46% (HR 0.54; 95% CI 0.39‐0.77) and 67% (HR 0.33; 95% CI 0.20‐0.54) lower VTE risk, respectively. Compared with overweight/obese individuals with eCRF < 85% of age‐predicted, overweight/obese individuals with eCRF ≥ 85% had 50% (HR 0.50, 95% CI 0.35‐0.74) lower risk, and normal weight individuals with eCRF ≥ 85% had 55% (HR 0.45, 95% CI 0.30‐0.68) lower risk.
Conclusions
Higher eCRF was associated with lower risk of incident VTE. The association was independent of weight categories, suggesting that higher eCRF may modify the association between obesity and VTE.
A comprehensive dissection of the role of microRNAs (miRNAs) in gene regulation and subsequent cell functions requires a specific and efficient knockdown or overexpression of the miRNA of interest; ...these are achieved by transfecting the cell of interest with a miRNA inhibitor or a miRNA mimic, respectively. Inhibitors and mimics of miRNAs with a unique chemistry and/or structural modifications are available commercially and require different transfection conditions. Here, we aimed to investigate how various conditions affect the transfection efficacy of two miRNAs with high and low endogenous expression, miR-15a-5p and miR-20b-5p respectively, in human primary cells.
MiRNA inhibitors and mimics from two commonly used commercial vendors were employed, i.e., mirVana (Thermo Fisher Scientific) and locked nucleic acid (LNA) miRNA (Qiagen). We systematically examined and optimized the transfection conditions of such miRNA inhibitors and mimics to primary endothelial cells and monocytes using either a lipid-based carrier (lipofectamine) for delivery or an unassisted uptake. Transfection of LNA inhibitors with either phosphodiester (PE)- or phosphorothioate (PS)-modified nucleotide bonds, delivered using a lipid-based carrier, efficiently downregulated the expression levels of miR-15a-5p already 24 h following transfection. MirVana miR-15a-5p inhibitor displayed a less efficient inhibitory effect, which was not improved 48 h following a single transfection or two consecutive transfections. Interestingly, LNA-PS miR-15a-5p inhibitor efficiently reduced the levels of miR-15a-5p when delivered without a lipid-based carrier in both ECs and monocytes. When using a carrier, mirVana and LNA miR-15a-5p and miR-20b-5p mimics showed similar efficiency 48 h following transfection to ECs and monocytes. None of the miRNA mimics effectively induced overexpression of the respective miRNA when given to primary cells without a carrier.
LNA miRNA inhibitors efficiently downregulated the cellular expression of miRNA, such as miR-15a-5p. Furthermore, our findings suggest that LNA-PS miRNA inhibitors can be delivered in the absence of a lipid-based carrier, whereas miRNA mimics need the aid of a lipid-based carrier to achieve sufficient cellular uptake.
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