Background: Despite numerous studies on cardioprotective effects of omega-3 polyunsaturated fatty acids (n-3 PUFAs), there is limited evidence for n-3 PUFA-mediated effects, especially at its higher dose, on cardiovascular risk in patients with type 2 diabetes (DM2) and established atherosclerosis.Purpose: To investigate the effect of daily treatment with a higher dose (2 g) of n-3 PUFAs on platelet function, coagulation parameters, fibrin clot properties, markers of systemic inflammation and metabolic status, in patients with atherosclerotic vascular disease and DM2 who receive optimal medical therapy.Methods: We conducted a prospective, double-blind, placebo-controlled, randomized, double-center study, in which thrombin generation (plasma thrombogenic potential from automated thrombogram), fibrin clot properties (plasma fibrin clot permeability; lysis time), platelet aggregation (light transmission aggregometry with adenosine diphosphate and arachidonic acid used as agonists), HbA1c, insulin level, lipid profiles, leptin and adiponectin levels, as well as markers of systemic inflammation (i.e., hsCRP, IL-6, TNF-α, ICAM-1, VCAM-1, and myeloperoxidase) were determined at baseline and at 3 months after treatment with 2 g/day of n-3 PUFAs (n = 36) or placebo (n = 38). Moreover, we assessed serum fatty acids of the phospholipid fraction by gas chromatography both at baseline and at the end of the study.Results: Majority of patients were treated with optimal medical therapy and achieved recommended treatment targets. Despite higher serum levels of eicosapentaenoic acid (EPA) (by 204%; p < 0.001) and docosahexaenoic acid (DHA) (by 62%; p < 0.0001) in n-3 PUFA group at the end of treatment no changes in platelet aggregation, thrombin generation, fibrin clot properties or markers of systemic inflammation were observed. No intergroup differences in the insulin, HbA1c and lipid levels were found at the end of the study. There was no change in adiponectin and leptin in interventional group, however leptin increased in control group (p = 0.01), therefore after study period leptin levels were lower in the interventional group (p = 0.01). Additionally, resolvin D1 did not differ between interventional and control group.Conclusions: In conclusion, our study demonstrated that in patients with long-standing, well-controlled DM2 and atherosclerotic disease the treatment with a high dose of n-3 PUFAs (namely, 1 g/day of EPA and 1 g/day of DHA for 3 months) does not improve coagulation, metabolic, and inflammatory status when measured with the specified tests.The study was registered in ClinicalTrials.gov; identifier: NCT02178501. Registration date: April 12, 2014

There are inconsistent data about the role of serum phospholipid fatty acid composition in patients with type 2 diabetes (T2DM) and atherosclerotic cardiovascular disease (ASCVD). The aim of the study was to investigate the relationship between serum phospholipid fatty acid composition, systemic low-grade inflammation, and glycemic control in high-risk T2DM patients. Seventy-four patients (26% women, mean age 65.6 ± 6.8 years) with T2DM (median diabetes duration 10 years) and documented ASCVD (74 with coronary artery disease, 26 with peripheral arterial disease) were enrolled in the study. Baseline HbA1c was estimated using turbidimetric inhibition immunoassay. According to the median value of HbA1c the patients were grouped into those with HbA1c < 7.0% (< 53 mmol/mol) (n = 38) and those with HbA1c ≥ 7.0% (≥ 53 mmol/mol) (n = 36). Serum phospholipid fatty acids were measured with gas chromatography. Patients with HbA1c ≥ 7.0%, compared with those with HbA1c < 7.0% had similar composition of saturated and monounsaturated fatty acids in serum phospholipids, but had higher concentrations of linoleic acid (LA) and higher n-6/n-3 polyunsaturated fatty acid (PUFA) ratio as well as lower levels of eicosapentaenoic acid (EPA), total n-3 PUFAs, and the EPA/arachidonic acid ratio. We found that LA (r = 0.25; p = 0.03) and n-6/n-3 PUFA ratio (r = 0.28; p = 0.02) were positively correlated with HbA1c. Multivariate logistic regression analysis showed that n-6/n-3 PUFA ratio, hsCRP and T2DM duration were independent predictors of worse glycemic control in patients with T2DM and ASCVD. This study showed that glycemic control in high-risk T2DM patients with ASCVD was significantly associated with unfavorable serum phospholipid n-6/n-3 PUFA ratio and greater systemic inflammation.

Heart failure (HF) is a clinical syndrome that is divided into 3 subtypes based on the left ventricular ejection fraction. Every subtype has specific clinical characteristics and concomitant diseases, substantially increasing risk of thromboembolic complications, such as stroke, peripheral embolism and pulmonary embolism. Despite the annual prevalence of 1% and devastating clinical consequences, thromboembolic complications are not typically recognized as the leading problem in patients with HF, representing an underappreciated clinical challenge. Although the currently available data do not support routine anticoagulation in patients with HF and sinus rhythm, initial reports suggest that such strategy might be beneficial in a subset of patients at especially high thromboembolic risk. Considering the existing evidence gap, we aimed to review the currently available data regarding coagulation disorders in acute and chronic HF based on the insight from preclinical and clinical studies, to summarize the evidence regarding anticoagulation in HF in special-case scenarios and to outline future research directions so as to establish the optimal patient-tailored strategies for antiplatelet and anticoagulant therapy in HF. In summary, we highlight the top 10 pearls in the management of patients with HF and no other specific indications for oral anticoagulation therapy. Further studies are urgently needed to shed light on the pathophysiological role of platelet activation in HF and to evaluate whether antiplatelet or antithrombotic therapy could be beneficial in patients with HF. LAY SUMMARY: Heart failure (HF) is a clinical syndrome divided into 3 subtypes on the basis of the left ventricular systolic function. Every subtype has specific clinical characteristics and concomitant diseases, substantially increasing the risk of thromboembolic complications, such as stroke, peripheral embolism and pulmonary embolism. Despite the annual prevalence of 1% and devastating clinical consequences, thromboembolic complications are not typically recognized as the leading problem in patients with HF, representing an underappreciated clinical challenge. Although the currently available data do not support routine anticoagulation in patients with HF and no atrial arrhythmia, initial reports suggest that such a strategy might be beneficial in a subset of patients at especially high risk of thrombotic complications. Considering the existing evidence gap, we aimed to review the currently available data regarding coagulation problems in stable and unstable patients with HF based on the insight from preclinical and clinical studies, to summarize the evidence regarding anticoagulation in HF in specific patient groups and to outline future research directions to establish the optimal strategies for antiplatelet and anticoagulant therapy in HF, tailored to the needs of an individual patient. In summary, we highlight the top 10 pearls in the management of patients with HF and no other specific indications for oral anticoagulation therapy.

Scanning Electron Microscopy (SEM) is a powerful, high-resolution imaging technique widely used to analyze the structure of fibrin networks. Currently, structural features, such as fiber diameter, length, density, and porosity, are mostly analyzed manually, which is tedious and may introduce user bias. A reliable, automated structural image analysis method would mitigate these drawbacks. We evaluated the performance of DiameterJ (an ImageJ plug-in) for analyzing fibrin fiber diameter by comparing automated DiameterJ outputs with manual diameter measurements in four SEM data sets with different imaging parameters. We also investigated correlations between biophysical fibrin clot properties and diameter, and between clot permeability and DiameterJ-determined clot porosity. Several of the 24 DiameterJ algorithms returned diameter values that highly correlated with and closely matched the values of the manual measurements. However, optimal performance was dependent on the pixel size of the images—best results were obtained for images with a pixel size of 8–10 nm (13–16 pixels/fiber). Larger or smaller pixels resulted in an over- or underestimation of diameter values, respectively. The correlation between clot permeability and DiameterJ-determined clot porosity was modest, likely because it is difficult to establish the correct image depth of field in this analysis. In conclusion, several DiameterJ algorithms (M6, M5, T3) perform well for diameter determination from SEM images, given the appropriate imaging conditions (13–16 pixels/fiber). Determining fibrin clot porosity via DiameterJ is challenging.

Little is known about factors that affect the composition of contracted blood clots in specific diseases. We investigated the content of polyhedral erythrocytes (polyhedrocytes) formed in blood clots and its determinants in type 2 diabetes (T2D) patients. In 97 patients with long-standing T2D [median HbA1c, 6.4% (interquartile range 5.9–7.8)], we measured in vitro the composition of blood clots, including a clot area covered by polyhedrocytes using scanning electron microscopy and the erythrocyte compression index (ECI), defined as a ratio of the mean polyhedrocyte area to the mean native erythrocyte area. Moreover, plasma fibrin clot permeability (Ks), clot lysis time (CLT), thrombin generation, oxidative stress [total protein carbonyl (total PC), total antioxidant capacity and thiobarbituric acid reactive substances (TBARS)], and platelet activation markers were determined. The impact of glucose concentration on polyhedrocytes formation was assessed in vitro. Polyhedrocytes content in contracted clots was positively correlated with glucose (r = 0.24, p = 0.028), glycated hemoglobin (r = 0.40, p = 0.024), total cholesterol (r = 0.22, p = 0.044), TBARS (r = 0.60, p = 0.0027), P-selectin (r = 0.54, p = 0.0078) and platelet factor-4, PF4 (r = 0.59, p = 0.0032), but not with thrombin generation, platelet count, Ks or CLT. Patients who formed more polyhedrocytes (≥ 10th percentile) (n = 83, 85.6%) had higher glucose (+ 15.7%, p = 0.018), fibrinogen (+ 16.6%, p = 0.004), lower red blood cell distribution width (RDW, − 8.8%, p = 0.034), reduced plasma clot density (− 21.8% Ks, p = 0.011) and impaired fibrinolysis (+ 6.5% CLT, p = 0.037) when compared to patients with lesser amount of polyhedrocytes (< 10th percentile). ECI and the content of polyhedrocytes were strongly associated with total PC (r = 0.79, p = 0.036 and r = 0.67, p = 0.0004, respectively). In vitro an increase of glucose concentration by 10 mmol/L was associated with 94% higher polyhedrocytes content (p = 0.033) when compared to the baseline (7.1 mM). After adjustment for age, sex and fibrinogen, multiple regression analysis showed that RDW was the only independent predictor of polyhedrocytes content in T2D (OR = 0.61, 95% CI 0.39–0.92). Poor glycemic control, together with enhanced platelet activation and oxidative stress, increase the content of polyhedrocytes in blood clots generated in T2D patients.