Prothrombotic State In Patients Research Articles

Low extracellular vesicle\u2013associated tissue factor activity in patients with persistent lupus anticoagulant and a history of thrombosis

Lupus anticoagulants (LA) are a heterogeneous group of antiphospholipid antibodies (aPLAs) that promote thrombosis. Tissue factor (TF)–bearing extracellular vesicles (EVs) might contribute to the prothrombotic state of patients with persistent LA and a history of thrombosis. To investigate if EV-associated TF activity is elevated in a well-defined group of LA-positive patients with a history of thrombosis in comparison to that of healthy controls. Adult patients (n = 94, median age 40.1 years, interquartile range (IQR) 29.9–53.4; 87% females) positive for LA and a history of thrombosis (78% venous thrombosis, 17% arterial thrombosis, 5% venous thrombosis and arterial thrombosis) and healthy age- and sex-matched controls (n = 30, median age 42.9 years, IQR 38.6–45.8, 77% females) were included in this study. EV-TF activity was determined with a factor Xa generation assay and anti-β2-glycoprotein (anti-β2GPI) and anticardiolipin (aCL) antibodies by enzyme-linked immunoassays. EV-TF activity did not differ between 94 LA-positive patients with a history of thrombosis (median 0.05 pg/mL, IQR 0.00–0.14) and 30 healthy controls (median 0.06, IQR 0.00–0.11, p = 0.7745). No correlation was found between EV-TF activity and lupus-sensitive activated partial thromboplastin time (aPTT-LA) (rho = 0.034), Rosner index (rho = − 0.056), anti-β2GPI IgG (rho = 0.05), anti-β2GPI IgM (rho = − 0.08), aCL IgG (rho = 0.12), and aCL IgM (rho = − 0.11) in LA-positive patients. We found low EV-TF activity levels in LA-positive patients and a history of thrombosis and no correlation with analyzed aPLAs. Our data indicate that circulating TF-bearing EVs do not contribute to the prothrombotic state of patients with LA.

Left Atrial Strain Predicts Pro-Thrombotic State in Patients with Non-Valvular Atrial Fibrillation.

Atrial fibrillation (AF) has a high prevalence in the population and it is responsible for up to the 25% of the strokes in elderly people. The aim of our study was to assess the correlations of left atrial (LA) functional parameter, global peak atrial longitudinal strain (PALS), derived from speckle tracking echocardiography (STE), with transesophageal echocardiography (TEE) findings in patients with persistent AF undergoing TEE before electrical cardioversion or ablation procedures. 79 patients (58 males, 21 females) with persistent AF waiting for cardioversion were included in the study. The patients underwent conventional two-dimensional (2D) and 2D speckle tracking echocardiogram. PALS were measured in all subjects. Patients were divided into two groups according to the presence of reduced LA appendage (LAA) emptying velocity (<25 cm/s) and/or thrombus in the LAA at TEE examination. Patients with reduced LAA emptying velocity and/or thrombus at TEE examination showed a significantly higher LA volume and increased E/E' ratio. 4-chamber, 2-chamber and global PALS were significantly lower in patients with reduced LAA emptying velocity and/or thrombus (6.8 ± 2.0% vs. 27.5 ± 5.4%, P < 0.0001; 8.6 ± 3.5% vs. 29.4 ± 7.1%, P < 0.0001; 7.9 ± 3.2% vs. 28.5 ± 6.1%, P <0.0001, respectively). Among all variables analyzed, global PALS demonstrated the highest diagnostic accuracy (AUC of 0.92) and, with a cut-off value less than 8.1%, good sensitivity and specificity of 87% and 94%, respectively, to predict LAA thrombus and/or reduced LAA emptying velocity.

IL-33 stimulates the release of procoagulant microvesicles from human monocytes and differentially increases tissue factor in human monocyte subsets.

Monocytes and monocyte-derived microvesicles (MVs) are the main source of circulating tissue factor (TF). Increased monocyte TF expression and increased circulating levels of procoagulant MVs contribute to the formation of a prothrombotic state in patients with cardiovascular disease. Interleukin (IL)-33 is a pro-inflammatory cytokine involved in atherosclerosis and other inflammatory diseases, but its role in regulating thrombosis is still unclear. The aim of the present study was to investigate the effects of IL-33 on the procoagulant properties of human monocytes and monocyte-derived MVs. IL-33 induced a time- and concentration-dependent increase of monocyte TF mRNA and protein levels via binding to the ST2-receptor and activation of the NF-κB-pathway. The IL-33 treated monocytes also released CD14+TF+ MVs and IL-33 was found to increase the TF activity of both the isolated monocytes and monocyte-derived MVs. The monocytes were classified into subsets according to their CD14 and CD16 expression. Intermediate monocytes (IM) showed the highest ST2 receptor expression, followed by non-classical monocytes (NCM), and classical monocytes (CM). IL-33 induced a significant increase of TF only in the IM (p<0.01), with a tendency in NCM (p=0.06), but no increase was observed in CM. Finally, plasma levels of IL-33 were positively correlated with CD14+TF+ MVs in patients undergoing carotid endarterectomy (r=0.480; p=0.032; n=20). We hereby provide novel evidence that the proinflammatory cytokine IL-33 induces differential TF expression and activity in monocyte subsets, as well as the release of procoagulant MVs. In this manner, IL-33 may contribute to the formation of a prothrombotic state characteristic for cardiovascular disease.

Early Low Level Hemolysis Is Associated with Subsequent Device Thrombosis and Ischemic Stroke during Continuous Flow Left Ventricular Assist Device Support By the Heart Mate II

Background: Heart failure affects more than 7 million people in the United States with over 250,000 patients in advanced stages with a dismal 1 year survival of less than 20%. For such patients, mechanical circulatory support (MCS) with continuous flow left ventricular assist devices (CF LVADs) and 1-year survival rates of >80%, has become a viable treatment option, either as a bridge to heart transplantation or as destination therapy. Although bleeding is a known major problem during CF LVAD support, a more devastating sequela is the rising incidence of thrombosis. The etiology of thrombosis remains uncertain. Hemolysis is associated with a prothrombotic state in patients with paroxysmal nocturnal hemoglobinuria and in individuals with hemoglobinopathies, and is a significant complication of CF LVADs. We hypothesized that early low level hemolysis, as assessed by LDH at the time of discharge, could be related to subsequent thrombosis and might serve as a predictive biomarker.Methods: A retrospective single center review of all patients supported for at least 30 days by a Heart Mate (HM) II CF LVAD from January 2010 to July 2015 was conducted. All patients were on the same a priori antithrombotic regimen of aspirin 325 mg daily and warfarin therapy with a target INR of 2-3. Major demographics and LDH levels from the time of discharge of the HM II implant hospitalization were retrieved. Present, but low level hemolysis was defined by LDH levels of >400 U/L (normal range: 125-305 U/L). Cumulative survival free from a combined endpoint of confirmed device thrombosis and ischemic stroke (iCVA) was compared by using Kaplan Meier analysis. Device thrombosis was suspected by aberrant device parameters/malfunction and confirmed by device inspection after surgical exchange. Ischemic CVA was suspected based on neurologic symptoms and confirmed on Brain CT scan.Results: Overall, 123 patients were supported for 428±347 days by the HM II. Twelve (9.7%) had a thrombotic event from either a confirmed device thrombus (n=6) or an iCVA (n=6). There were no significant differences in age (thrombosis: 48±13yrs, no thrombosis: 54±12yrs, p=0.1) and gender (thrombosis: 16% female, no thrombosis: 23% female, p=0.9). Patients with thrombosis had a higher LDH at discharge (504 ±307 U/L) in comparison to those without thrombosis (350 ±110 U/L, p<0.0001). Patients with a LDH >400 U/L (n=88) at discharge had an over 6-fold higher risk of subsequent device thrombosis or iCVA in comparison to those with LDH levels ≤400 U/L (n=35, HR: 6.2, 95% CI: 1.7-22.3, p=0.002, Figure 1). Despite salvage interventions after thrombosis such as device exchange or heart transplantation, there was a trend towards worse overall survival in patients with a discharge LDH >400 U/L (HR: 1.5, 95% CI: 0.92-2.5, p=0.07).Conclusion: Low level hemolysis, as assessed by increased LDH levels at discharge during HM II support, is associated with thrombosis. Further studies to evaluate LDH as a predictive marker for thrombosis and investigations into the mechanisms of a hemolysis-induced pro-thrombotic state during CF LVAD support are warranted. [Display omitted] DisclosuresGoldstein:St. Judes Medical: Consultancy. Jorde:St. Judes Medical: Consultancy.

Prothrombotic State in Patients With a Left Atrial Appendage Thrombus of Unknown Origin and Cerebrovascular Events.

We hypothesized that formation of left atrial appendage (LAA) thrombi of unknown origin is associated with altered fibrin clot properties and blood hypercoagulability. In a case-control study, we investigated 32 patients with a history of LAA thrombus after successful anticoagulant treatment versus 32 control subjects matched for age, sex, and diabetes mellitus. All subjects had previous ischemic stroke, transient ischemic attack, or migraine associated with patent foramen ovale. Patients with documented atrial fibrillation were excluded. We determined plasma fibrin clot permeability, fibrinolytic efficiency, thrombin generation, platelet and endothelial markers. Stroke or transient ischemic attack were assessed during a median follow-up of 74 (range 19-98) months. Compared with controls, patients with LAA thrombus more frequently were smokers (43.8% versus 18.8%) and had 20% prolonged clot lysis time, lower plasminogen (-14%), and higher plasminogen activator inhibitor-1 (+17%), thrombin-antithrombin complexes (+17%), CD40 ligand (+30%), P-selectin (+29%), and von Willebrand factor (+30%, all P<0.05). Occurrence of LAA thrombus was predicted by von Willebrand factor (β=0.038, P=0.004), plasminogen (β=-0.048, P=0.01), plasminogen activator inhibitor-1 (β=-0.161, P=0.03), and clot permeability (β=-1.076, P=0.03). During follow-up, cerebrovascular events occurred in 10 (33.33%) of the 30 available patients in the LAA thrombus group, including 7 (23.3%) with recurrent LAA thrombus and 4 (13.33%) with documented atrial fibrillation. Recurrent LAA thrombus was associated with lower baseline Ks and higher thrombin generation, fibrinogen, plasminogen activator inhibitor-1, and soluble CD40 ligand (all P<0.05). Prothrombotic blood alterations could be involved in the LAA thrombus formation in patients without documented atrial fibrillation and are associated with increased risk of stroke or transient ischemic attack during follow-up.

Coagulation, Fibrinolysis, and Platelet Activation Following Open Surgical or Percutaneous Angioplasty Revascularization for Symptomatic Lower Limb Chronic Ischemia.

Critical limb ischemia (CLI) is associated with a prothrombotic diathesis that involves a complex balance between the coagulation and fibrinolytic systems. Knowledge of this is essential when considering revascularization procedures but is often overlooked. The aim of this review is to summarize the available literature and provide an overview of the effects of lower limb angioplasty and open surgical revascularization on coagulation, fibrinolysis, and platelet activation. A MEDLINE and EMBASE search was conducted between 1973 and 2014 for articles relating to the effects of revascularization for patients with CLI on the fibrinolytic and coagulation pathways. Studies with a small cohort of patients (<5) were rejected. Many of the studies included in this analysis had small cohorts. Multiple markers were assessed across the published literature including von Willebrand factor, tissue factor, prothrombin fragments 1 and 2, platelets, soluble platelet selectin, plasminogen activator inhibitor 1, tissue plasminogen activator, and thrombin-antithrombin complex. Percutaneous intervention causes an exaggerated prothrombotic and a disturbed fibrinolytic effect. Surgery seems to cause a similar prothrombotic derangement with reduced fibrinolysis and platelet hyperactivity, but this appears to be maintained for a considerable amount of time postoperatively. There is a sparse amount published on the effects of the coagulation and fibrinolytic systems in patients undergoing intervention for CLI. Much of these studies are small, historical, and completely heterogeneous, making it difficult to draw meaningful conclusions. The literature does identify a prothrombotic state in patients with CLI, which appears to be exacerbated by any form of intervention and prolonged in those having surgery. Understanding this may allow us to tailor the intervention offered to patients and prevent limb loss.

Prothrombotic state in patients with severe and prednisolone-dependent asthma

Epidemiologic studies have shown that asthmatic patients, in particular those with severe disease, have increased risk of pulmonary embolism. It is unknown whether these patients have a prothrombotic state under stable conditions. We sought to compare coagulation and fibrinolysis parameters between healthy subjects and patients with mild, severe, and prednisolone-dependent asthma under stable conditions and to investigate whether hemostatic markers correlate with airway inflammation. In 126 adults (33 healthy control subjects, 31 patients with mild asthma, 32 patients with severe asthma, and 30 patients with prednisolone-dependent asthma) parameters of inflammation (peripheral blood eosinophils and neutrophils) and markers of hemostasis (endogenous thrombin potential [ETP], thrombin-antithrombin complex, plasmin-α2-antiplasmin complex, plasminogen activator inhibitor type 1 [PAI-1], D-dimer, and von Willebrand factor [vWF]) were measured in plasma. One-way ANOVA with the post hoc Bonferroni test was used for group comparison, and linear regression analysis was used for correlations. We observed increased ETP (121% vs 99%, overall P<.01), plasmin-α2-antiplasmin complex (520 vs 409μg/L, overall P=.04), PAI-1 (10 vs 7ng/mL, overall P=.02), and vWF (142% vs 87%, overall P<.01) levels in asthmatic patients compared with healthy control subjects. ETP, PAI-1, and vWF levels increased with increasing asthma severity. In addition, we found a correlation between ETP and vWF with neutrophil but not eosinophil counts. Asthmatic patients have a prothrombotic state that increases with asthma severity. This might explain why patients with asthma, in particular those with severe disease, have an increased risk of venous thromboembolism.

Metabolic Syndrome and Deep Vein Thrombosis After Total Knee and Hip Arthroplasty

BackgroundMetabolic syndrome (MS), defined as obesity, hypertension, hyperglycemia, and dyslipidemia, is prevalent among patients undergoing total joint arthroplasty (TJA). MS has proven to promote a proinflammatory and prothrombotic state in patients. Venous thromboembolism is one of the major complications of TJA. The purpose of this retrospective study is to identify whether MS and its components increase the risk of deep vein thrombosis (DVT) after TJA. MethodsWe retrospectively reviewed 1553 patients undergoing primary unilateral TJA from 2007 to 2014. MS was diagnosed based on the World Health Organization criteria. All subjects received venography after operation to screen for DVT. Symptomatic DVT events after TJA were also recorded. Univariate analysis and multivariate logistic regression analysis were used to identify the association of MS and its components with postoperative DVT. ResultsThe prevalence of MS in patients undergoing TJA was 5.1% (n = 79). A total of 335 patients (21.6%) were diagnosed with DVT by venography. Seventy-eight patients (5.0%) developed symptomatic DVT. In the total knee arthroplasty group, MS and obesity were related to postoperative DVT. MS alone was found to be associated with symptomatic DVT. In the total hip arthroplasty group, MS increased the risk of symptomatic DVT. However, obesity, rather than MS, was associated with total DVT after total hip arthroplasty. ConclusionsMS was a significant risk factor for DVT after TJA. Strategies to minimize the adverse effect of MS should be considered for these patients.

Pre-Existence of Prothrombotic State in Patients with Atrial Fibrillation Despite Therapy with New and Traditional Anti-Coagulant Drugs

Background: Oral anticoagulants such as warfarin (W) have been conventionally used for the management of atrial fibrillation (AF). Despite the effectiveness of W, its use in AF patients requiring anticoagulation is suboptimal with an even greater underuse seen in elderly patients who are at higher risk of stroke. New oral anticoagulants such as rivaroxaban (R) and apixaban (A) have been approved to manage thrombotic and cardiovascular disorders including AF. The newer anticoagulants do not require continuous monitoring like W does and are much more convenient for patients with AF.Objective: To profile the baseline level of circulating thrombogenic biomarkers von Willebrand Factor (vWF), prothrombin fragment 1.2 (F1+2), microparticle bound tissue factor (MP-TF) and plasminogen activator inhibitor (PAI-1) in patients with AF. Additionally, the effect of both newer (R and A) and traditional (W) anticoagulants on the levels of thrombogenic biomarkers in patients with AF will be assessed.Materials: Citrated blood was drawn from thirty AF patients prior to ablation surgery and spun at 3000 rpm to obtain platelet poor plasma. Normal plasma samples from healthy controls were purchased from a commercial source (George King Biomedical, Overland Park, KS). The plasma samples were analyzed using a biochip array (Randox, London, UK) for metabolic syndrome biomarkers including PAI-1 and ELISA kits for vWF, MP-TF (Hyphen BioMed, Nueville-Sur-Oise, France) and prothrombin F1+2 (Siemens, Newark, DE).Results: Circulating levels of vWF, MP-TF and PAI-1 were statistically increased in patients with AF compared to normal (P<0.0001, P<0.0001, and P=0.0014, respectively). Circulating levels of prothrombin F1+2 showed no difference between the AF and normal group (P=0.2696). AF patients (n=30) were divided into two groups based on their usage (Group 1, n=21) and non-usage (Group 2, n=9) of any anticoagulant. Furthermore, those on anticoagulants were divided based on their use of newer (R and A, Group 3, n=16) or traditional (W, Group 4, n=4) anticoagulants. A statistical increase in vWF (P<0.0001), MP-TF (P<0.0001) and PAI-1 (P=0.011) remained in Group 1 compared to normal while a statistical increase in prothrombin F1+2 (P=0.0343) and PAI-1 (P=0.0040) were noted in Group 2 compared to normal. vWF (P=0.0036) and MP-TF (P=0.0059) were elevated in Group 1 compared to Group 2 while prothrombin F1+2 (P=0.0697) and PAI-1 (P=0.4548) showed no difference between the two groups. Furthermore, there was no statistical difference in the level of any thrombogenic biomarker in AF patients between Group 3 (R and A) and Group 4 (W). (Table 1)Discussion: Elevated levels of vWF, MP-TF and PAI-1 seen in AF patients compared to normal provide insight into an additional risk of thrombogenesis associated with AF which is not targeted by current anticoagulant medications. Most patients are assessed using a stroke risk stratification scale (CHA2DS2VASc, CHADS2, CHADS-VASC, or CHADS) to determine if anti-coagulants should be used to prevent stroke associated with AF. Of the 30 patients examined in this study, 8/9 (89%) patients who were not on anticoagulants had a stroke risk stratification score of 0 while 20/21(95%) patients who were on anticoagulants had a score of >1. This data supports studies which suggest that adding levels of prothrombotic biomarkers to current risk stratification scales could be more effective in assessing the risk of stroke of patients with AF. This data also suggests that although very effective in lowering prothrombin F1+2 levels in AF, the newer anticoagulants, R and A, and the traditional anticoagulant, W, still leave additional prothrombotic biomarkers unaffected. These unaffected biomarkers could be the potential target of future drug therapies which could lower the risk of stroke in patients with AF even more than the use of newer/traditional anticoagulants alone. Table 1Biomarkers of Thrombogenesis in AF and Normal GroupsGroupvWF (concentration %)Prothrombin F1+2 (pmol/L)MP-TF (pg/mL)PAI-1 (ng/mL)Normal4140 ± 919 n=46106.1 ± 52.7 n=500.38 ± 0.25 n=483.21 ± 4.13 n=25AFGroup 1 Anticoagulant n=216616 ± 1173107.9 ± 61.20.93 ± 0.715.69 ± 4.15Group 2 Non-Anticoagulant n=94788 ± 1338162.5 ± 93.30.51 ± 0.136.49 ± 3.14Group 3 New Anticoagulants (R and A) n=166721 ± 1127103.5 ± 37.60.76 ± 0.345.68 ± 4.53Group 4 Traditional Anticoagulants (W) n=46387 ± 158075.2 ± 52.60.94 ± 0.335.63 ± 3.52 DisclosuresNo relevant conflicts of interest to declare.

Association of the CHA2DS2-VASc score with left atrial spontaneous echo contrast: a cross-sectional study of patients with rheumatic mitral stenosis in sinus rhythm.

Mitral valve stenosis is a common manifestation of chronic rheumatic heart disease. In rheumatic mitral valve stenosis (RMVS) patients, left atrial spontaneous echo contrast (LASEC) is an independent predictor of thromboembolism risk. While the anticoagulant therapy algorithm for atrial fibrillation patients is clear, the clinical tools determining high-risk patients in sinus rhythm are insufficient. Our aim is to examine the relationship between CHA2DS2-VASc score in RMVS patients in sinus rhythm and the presence of LASEC. The patients with RMVS upon presentation to the cardiology polyclinic were included in this cross-sectional study consecutively, and CHA2DS2-VASc scores were calculated. All patients were evaluated with transthoracic and transesophageal echocardiography and were divided into two groups as those with and without LASEC. The total number of patients was 265, with LASEC determined in 97 (36.6%) and not determined in 168 (63.4%). No significant differences in terms of age, gender, and body mass index were found between the groups. Patients with LASEC had higher mean CHA2DS2-VASc score than patients without LASEC (2.10±1.21 vs. 1.11±0.7, respectively; p<0.001). In the multivariate logistic regression analysis, it has been determined that there is an independent association between the existence of LASEC and CHA2DS2-VASc score (OR 3.176, CI 1.937-5.206; p<0.001). The ROC analysis revealed that CHA2DS2-VASc score 2 or more predicted presence of LASEC with a sensitivity of 71% and a specificity of 82% (AUC 0.746, 95% CI 0.682-0.810). The CHA2DS2-VASc score could be useful marker to detect prothrombotic state in patients with RMVS in sinus rhythm.

Anticoagulation in patients with pulmonary arterial hypertension: An update on current knowledge

Pulmonary hypertension is a severe clinical condition characterized by molecular and anatomic changes in pulmonary circulation. It is associated with increased pulmonary vascular resistance, which leads to right-sided heart failure if left untreated and, ultimately, death. Treatment of patients with pulmonary arterial hypertension (PAH) involves a complex strategy that takes into consideration disease severity, general and supportive measures, and combination drug regimens. Abnormalities of blood coagulation factors, anti-thrombotic factors, and the fibrinolytic system may contribute to a prothrombotic state in patients with idiopathic PAH. These physiologic changes, in concert with the presence of non-specific risk factors for venous thromboembolism such as heart failure and immobility, are thought to be the basis for oral anticoagulation in PAH. Several observational studies provide helpful information in favor of anticoagulation use in idiopathic PAH but not in other pulmonary hypertension etiologies. Guideline recommendations are based on the lack of prospective comparative trials in this regard. For that reason, large differences exist in the use of anticoagulants in different countries and centers. More studies should be carried out to clarify the risks and the potential benefits of anticoagulant use in a heterogeneous population of patients who are already at considerable life risk.

Prothrombotic state of patients with unexplained recurrent spontaneous abortion.

To determine the effect of aspirin and low-molecular-weight heparin (LMWH) for women affected by recurrent spontaneous abortion (RSA). The present prospective observational study included women with RSA (≥ 2 previous spontaneous abortions) and those without a history of RSA attending Ren Ji Hospital, Shanghai, China, between March 2011 and March 2014. D-dimer levels, auto-antibodies, and platelet aggregation in response to arachidonic acid (AA) and adenosine diphosphate (ADP) were monitored before pregnancy. Women with unexplained RSA and increased platelet aggregation received low-dose aspirin, and those with elevated D-dimer levels (> 0.75 μg/mL) received LMWH. Platelet aggregation in response to AA and ADP, and D-dimer levels were monitored during pregnancy. Among 517 participants with RSA, 325 had unexplained RSA. Compared with 63 control patients, those with unexplained RSA and two, three, four, or at least five previous spontaneous abortions had increased platelet aggregation in response to AA (P < 0.05 for all comparisons) but not to ADP. Among 37 women with unexplained RSA who became pregnant, aspirin reduced platelet aggregation in response to AA (P < 0.001). LMWH reduced D-dimer levels during pregnancy (P < 0.001). Among 37 women with unexplained RSA who became pregnant and completed the study, 33 (89.2%) had a live birth. Aspirin and LMWH maintained a lower thrombotic state and led to a high live birth rate among patients with unexplained RSA.

Thrombin generation and factor X assays for the assessment of warfarin anticoagulation in thrombotic antiphospholipid syndrome

Monitoring warfarin anticoagulation in patients with thrombotic antiphospholipid syndrome (APS) may be complicated by the sensitivity of different thromboplastins to lupus anticoagulant. The aim of this study was to compare the degree of anticoagulation intensity in thrombotic APS and non-APS patients (50 in each group) on long-term warfarin, by measurement of the INR with two widely available thromboplastins with instrument-specific ISI values, and to investigate the potential role of amidolytic FX levels and thrombin generation (TG) testing in the assessment of anticoagulant intensity in thrombotic APS patients. There were no overall differences in INR between reagents or patient groups, but 20% (10/50) of APS patients showed ≥0.5 INR unit difference between reagents, which would have resulted in altered clinical management in some patients. FX levels were useful in assessing anticoagulation intensity for INR 2.0-3.0, but showed poor utility at INR ≥3.5 where the lowest measured FX level was 12IU/dL. In contrast, ETP and peak thrombin showed significant inverse correlations with the INR, suggesting that TG testing may be helpful in the determination of true anticoagulant intensity in APS patients, including those with ≥3.5 INR. TG testing also highlighted a subgroup of APS patients with increased peak thrombin relative to the intensity of anticoagulation as assessed by INR and FX, suggesting that TG testing may be useful in identifying an ongoing prothrombotic state in patients with apparently adequate anticoagulation intensity as assessed by INR.

Current antiplatelet treatment strategy in patients with diabetes mellitus.

Patients with diabetes mellitus (DM) have accelerated atherosclerosis with an increased risk for atherothrombotic cardiovascular complications. A state of high platelet reactivity and activation, hypercoagulability (prothrombotic state) and a subdued response to standard antiplatelet agents may explain high rate of adverse cardiovascular events in patients with DM. Several antithrombotic treatment strategies have been developed to control the prothrombotic state in patients with DM: dose modification of commonly used agents; use of potent agents; and addition of a third antithrombotic drug (triple therapy) to commonly prescribed dual antiplatelet therapy of aspirin and a P2Y12 inhibitor. The present review aims to provide an overview of the current knowledge on platelet abnormalities in patients with DM, focusing on the challenges and perspectives of antiplatelet treatment strategies in this population.

The coagulation system in patients with end-stage liver disease.

In patients with cirrhosis, routine laboratory tests for primary hemostasis and coagulation usually show anomalies that are associated with excess bleeding in other settings, in particular low platelet counts and prolonged prothrombin time. However, under conditions similar to those in vivo, primary hemostasis and thrombin production do not appear to be decreased in patients with cirrhosis, particularly when the platelet count is above 75,000/μl. Furthermore, there is laboratory and epidemiological evidence of a mild procoagulant and prothrombotic state in patients with cirrhosis. Bleeding is mainly because of portal hypertension rather than defective hemostasis. There is some evidence that anticoagulation therapy is not associated with an excess of severe bleeding and that it could improve the outcome in patients without portal vein thrombosis. At present, there is no clear evidence that portal vein thrombosis is responsible for the progression of liver disease and that anticoagulation therapy would improve the outcome of patients with portal vein thrombosis.

Case Report: Autoimmune Hemolytic Anemia with Venous Thromboembolism, a Common Complication of a Rare Disease

IntroductionAutoimmune hemolytic anemia (AIHA) is accelerated erythrocyte destruction by binding of IgG (80%) or IgM (20%) antibodies with red blood cell (RBC) antigens, leading to formation of spherocytes and eventual phagocytosis in the spleen. Its incidence is 1 in 80,000 to 100,000 of a given population/year in the Caucasians. Its peak incidence is in 60-70 years of age with male: female ratio of 40:60. Although rare in incidence, AIHA is the most common cause of acquired hemolytic anemia. Multiple triggering factors have been described for AIHA that includes idiopathic, genetic, infection, inflammatory disorders, drugs and lymphoproliferative disorders. Venous thromboembolism (VTE) is a more serious, acute, overlooked and often-lethal complication of AIHA. Several studies including a review of 47 patients with AIHA supporting the existence of a prothrombotic state in patients with AIHA revealed that pulmonary embolism was the most common cause of death. It is essential to recognize and treat VTE, and thromoboprophylaxis measures should be taken to prevent VTE associated morbidity and mortality.Case summaryPatient is a 76 year-old-Caucasian-female with medical history significant for peptic ulcer disease and hypertension presented with 1-month history of dyspnea on moderate exertion and a near syncopal episode a week prior. Vital Signs revealed tachycardia at 100/min. Physical examination depicted marked pallor in the conjunctiva and on palmar creases with no lymphadenopathy, murmurs, hepatosplenomegaly, petechiae, purpura and peripheral edema. Lab test revealed hemoglobin 3.8 g/dl, platelets 205 X 109/L, reticulocytes 10.3%, absolute reticulocyte count 3.8, haptoglobin 142 and LDH 337 U/L. Peripheral smear revealed spherocytes, and schistocytes with unremarkable platelets and leukocytes. Direct coombs test revealed strongly positive IgG. Hemoglobin was stabilized after 4 units of packed red blood cells transfusion, started on oral methyl-prednisolone at 1 mg/Kg/d with a prolonged taper course and discharged upon clinical improvement. After 2 weeks, she presented with progressively increasing swelling of both legs and increasing tiredness. Vitals signs were stable. Physical examination was remarkable for bilateral mid-shin level pitting edema with normal distal pulsations. Lab values revealed low hemoglobin, slightly increased LDH and mildly high absolute reticulocyte count. Bilateral lower extremity venous doppler showed extensive deep vein thrombosis (DVT) of the right and left common femoral veins, right proximal femoral vein, left femoral vein and left popliteal vein. She was started on IV heparin and coumadin, continued oral methyl-prednisolone at 1 mg/Kg/d and added rituximab scheduled infusion. She was clinically improved on discharge.DiscussionThere are various proposed mechanisms of VTE formation in AIHA such as loss of RBC membrane with exposure of phosphotidyl serine and subsequent formation of tenase and prothrombinase complexes, increased tissue factor expression by the endothelial cells, and sequestration of nitric oxide released from RBC. In a recent review, it was found that 27% of AIHA patients suffered from an episode of VTE. Out of 8 patients with VTE, 5 had lupus anticoagulant antibody (LA) and 4 had anti-cardiolipin antibody (ACC). Thrombosis also occurred in 15% of AIHA with no LA and ACC, so multifactorial interplay seems more likely. In an audit of prophylactic anticoagulation in acute exacerbation of severe AIHA, VTE occurred in 5 of 15 patients without anticoagulant prophylaxis. VTE occurred in only 1 of 21 patients with anticoagulant prophylaxis. In conclusion, it is essential to recognize this acute, serious and deadly complication. It is premature to recommend routine use of thromboprophylaxis for all patients with hemolytic episodes from AIHA. Based on the studies, coexisting ACC and LA are strong predictive risk factors for VTE in AIHA and can benefit from thromboprophylaxis. We need more clinical investigations to develop core clinical and laboratory criteria to stratify VTE risk in AIHA patients. DisclosuresNo relevant conflicts of interest to declare.

OP-098 Both Carvedilol and Nebivolol may Improve Platelet Function and Prothrombotic State in Patients with Non-Ischemic Heart Failure

OP-098 Both Carvedilol and Nebivolol may Improve Platelet Function and Prothrombotic State in Patients with Non-Ischemic Heart Failure

Selective serotonin reuptake inhibitors and risk for gastrointestinal bleeding

The most of the known effects of selective serotonin reuptake inhibitors, beneficial or harmful, are associated with the inhibitory action of the serotonin reuptake transporter. This mechanism is present not only in neurons, but also in other cells such as platelets. Serotoninergic mechanism seems to have an important role in hemostasis, which has long been underestimated. Abnormal activation may lead to a prothrombotic state in patients treated with selective serotonin reuptake inhibitors. On one hand there may be an increased risk of bleeding, and on the other hand reduction in thrombotic risk may be possible. Serotonin is critical to maintain a platelet haemostatic function, such as platelet aggregation. Evidences from the studies support the hypothesis that antidepressants with a relevant blockade of action of serotonin reuptake mechanism may increase the risk of bleeding, which can occur anywhere in the body. Epidemiological evidences are, however, the most robust for upper gastrointestinal bleeding. It is estimated that this bleeding can occur in 1 in 100 to 1 in 1.000 patient-years of exposure to the high-affinity selective serotonin reuptake inhibitors, with very old patients at the highest risk. The increased risk may be of particular relevance when selective serotonin reuptake inhibitors are taken simultaneously with nonsteroidal anti-inflammatory drugs, low dose of aspirin or warfarin.

Specific Gtpase Dynamin Isoforms Regulate Megakaryocyte Membrane Remodeling and The Formation Of Multivesicular Bodies and Microparticles

Cdc42 interacting protein 4 (CIP4) is a membrane-associated BAR protein, which also forms a complex via its SH3 domain with the dynamins (DNMs) and Wiskott-Aldrich Syndrome (WAS) protein. Thus, CIP4 remodels the plasma membrane and cortical actin cytoskeleton. To determine its physiological function, we generated CIP4-null mice. They displayed thrombocytopenia similar to that of WAS-null mice and have abnormal megakaryocytes (MKs) with decreased proplatelet formation and underdeveloped demarcation membrane system (DMS) (Chen et al, Blood 2013). The DMS is an extensive network of membrane tubules which serves as a membrane reservoir for proplatelet formation. The membranes are enriched for polyphosphoinositides that are docking sites for BAR proteins and for pleckstrin homology domain-containing proteins such as the dynamins. Still, the formation of the DMS is poorly understood. Dynamins are cell vesicle trafficking proteins that possess a GTPase domain. They induce neck vesicle constriction and scission from the plasma membrane. When the GTPase activity is abrogated, vesicle scission does not occur; instead, the plasma membrane invagination induced by the BAR proteins results in deep plasma membrane tubulations. Of the three dynamin isoforms, DNM3 participates in MK development including DMS formation (Reems et al, Exp Hematol 2008; Wang et al, Stem Cells Dev 2011). Moreover, a recent genome-wide association study suggested that an MK-specific DNM3 isoform might play a role in human platelet size determination (Nürnberg et al, Blood 2012). However the exact mechanism for dynamin's participation in DMS formation is unclear. A double knockout for dynamin 1 and dynamin 3 in neurons causes accumulation of long invaginations from the plasma membrane (Ferguson and De Camilli, Nat Rev Mol Cell Biol 2012). We initially hypothesized that CIP4's association with DNM3 contributes to the DMS development during platelet biogenesis and wanted to test for functional redundancy with other dynamins present in MKs and platelets. To determine if CIP4 interacts with dynamin in the MK lineage, we found that following either phorbol ester (PMA) or fibronectin stimulation in the human MK cell line CHRF-288, CIP4 co-precipitated with DNM3 and colocalized by confocal microscopy. To determine dynamin's effect on membrane biophysical properties, we measured the fluorescence anisotropy, which reflects the disorder of membrane lipids due to movement and indicate membrane rigidity. Compared with controls in CHRF-288 cells, shRNA-mediated knockdown (KD) of DNM2 or DNM3 resulted in higher membrane rigidity in response to PMA. The strongest effect was seen in double KD cells with decreased fluidity by 2.6 ± 0.3%, which is similar to what was observed with CIP4 KD and is physiologically significant (Chen et al Blood 2013). KD of DNM2 resulted in aberrant morphology, greater cell diameter, and electron microscopy (EM) showed formation of new multivesicular bodies (MVBs) which are sorting compartments during α- and dense granules formation. Single DNM3 KD cells had no observable phenotype. EM imaging of DNM2 and DNM3 double KD cells revealed plasma membrane tubulation that resembles the DMS. While control CHRF-288 cells, with high DNM3 protein expression, do not have a DMS at baseline, MK cell lines Meg-01 and L8057, with respectively lower or no dynamin-3 protein expression, both have a DMS (Battinelli et al PNAS 2001; Ishida Y et al, Exp Hematol 1993). Platelet microparticles (MPs) are known to mediate a prothrombotic state in patients. Having previously found that CIP4-null mice show reduced levels of platelet MPs, we measured MPs in dynamin knockdown cell supernatant by flow cytometry and CD41/Annexin V staining. Surprisingly, we found that microparticle levels were increased 2.9-fold in DNM2 KD cells and 3.8-fold in double DNM2 and DNM3 KD cells. Our findings suggest that: 1) there is only partial functional redundancy between DNM2 and DNM3 in platelet biogenesis, 2) DNM2 controls MVB formation and MP release in MK cells, and 3) the CIP4-dynamin pathway contributes to DMS formation. It is possible that CIP4's interaction with dynamins restrains their spatial and temporal activity to allow for long invaginations to accumulate in the DMS. Dynamin depletion might also increase surface membrane availability for MP formation. Dynamins are thus potential targets to modulate thrombotic state and platelet biogenesis. Disclosures:No relevant conflicts of interest to declare.

Prognostic role of plasma von Willebrand factor and soluble E‐selectin levels for future cardiovascular events in a ‘real‐world’ community cohort of patients with atrial fibrillation

Endothelial damage/dysfunction may contribute to a prothrombotic state in patients with atrial fibrillation (AF) and the increased risk of thromboembolism and cardiovascular events. Raised plasma von Willebrand factor (vWf), an established marker of endothelial damage/dysfunction, has been associated with stroke and vascular events, at least in a clinical trial population. Soluble E-selectin (sE-sel) is another biomarker of endothelial activation/dysfunction, with more limited data on prognostic outcomes in AF. To assess the relationship between the levels of vWf, sE-sel and clinical adverse outcomes (including stroke, MI and all-cause mortality) in a 'real-world' community cohort of patients with AF. We studied 423 patients (mean age 72·7 ± 8·4 years, 55·6% male) with nonvalvular AF, with a median follow-up of 19 (9-31) months. Plasma vWf and sE-sel levels were measured using enzyme-linked immunosorbent assay (ELISA). There were 94 clinical adverse events (22·2%) observed during a median follow-up of 19 months. Patients with clinical events had significantly higher vWf (P < 0·001) and sE-sel levels at baseline (P < 0·001) compared with those who were event free. Kaplan-Meir analyses demonstrated that more clinical adverse events occurred in the upper tertile of vWf [upper vs. lowest tertile, RR 3·8, 95% CI (2·63-5·57), P < 0·001; upper vs. middle tertile, RR 10·5, 95% CI (5·33-20·60), P < 0·001]. Similarly, the highest tertile of sE-sel was associated with more adverse events [upper vs. lowest tertile, RR 3·7, 95% CI (2·51-5·31), P < 0·001; upper vs. middle tertile, RR 6·5, 95% CI (3·56-11·91), P < 0·001]. High plasma vWf and soluble E-selectin levels are associated with an increased risk of clinical adverse events (acute myocardial infarction, ischaemic stroke and all-cause mortality) in 'real-world' patients with AF. These soluble biomarkers may potentially aid clinical risk stratification in this common arrhythmia.