Catheter-directed ultrasound-accelerated thrombolysis for the treatment of acute pulmonary embolism

Background

Systemic thrombolysis rapidly improves right ventricular (RV) dysfunction in patients with acute pulmonary embolism (PE) but is associated with major bleeding complications in up to 20%. The efficacy of low-dose, catheter-directed ultrasound-accelerated thrombolysis (USAT) on the reversal of RV dysfunction is unknown.

Materials and methods

We performed a retrospective analysis of 24 PE patients (60 ± 16 years) at intermediate (n = 19) or high risk (n = 5) from the East Jefferson General Hospital who were treated with USAT (mean rt-PA dose 33.5 ± 15.5 mg over 19.7 hours) and received multiplanar contrast-enhanced chest computed tomography (CT) scans at baseline and after USAT at 38 ± 14 hours. All CT measurements were performed by an independent core laboratory.

Results

The right-to-left ventricular dimension ratio (RV/LV ratio) from reconstructed CT four-chamber views at baseline of 1.33 ± 0.24 was significantly reduced to 1.00 ± 0.13 at follow-up by repeated-measures analysis of variance (p < 0.001). The CT-angiographic pulmonary clot burden as assessed by the modified Miller score was significantly reduced from 17.8 ± 5.3 to 8.7 ± 5.1 (p < 0.001). All patients were discharged alive, and there were no systemic bleeding complications but four major access site bleeding complications requiring transfusion and one suspected recurrent massive PE event.

Conclusions

In patients with intermediate and high risk PE, low-dose USAT rapidly reverses right ventricular dilatation and pulmonary clot burden.     

Association of elevated NTproBNP with recurrent thromboembolic events after acute pulmonary embolism

Introduction

N-Terminal Pro-Brain Natriuretic Peptide (NTproBNP) is a predictor of adverse short-term clinical outcomes in patients with acute pulmonary embolism (APE), but its long-term prognostic value remains largely undefined. The aim of this study was to assess the value of plasma NTproBNP with regard to recurrent venous thromboembolism (VTE).

Materials and Methods

NTproBNP levels were measured in 224 consecutive patients with the first episode of acute pulmonary embolism occurring from January 2005 through October 2010. Patients were categorized into two groups by NTproBNP reference range. Follow-ups were performed at 3, 6, and 12 months and yearly thereafter. The primary end point was symptomatic, recurrent fatal or nonfatal VTE.

Results

NTproBNP was elevated in 158 (70.5%) patients and not elevated in 66 (29.5%) patients. After a mean follow-up period of 31.0 ± 19.4 months, patients with elevated NTproBNP showed an increased risk of recurrent VTE (20 patients, 12.7%) compared to those without elevated NTproBNP (only 1 patient, 1.5%) (P = 0.009). Of the 7 deaths related to pulmonary embolism, 6 occurred in patients with elevated NTproBNP compared to patients with normal NTproBNP (1 of 7 deaths). In a multivariate analysis stratified by oral anticoagulant treatment duration, elevated NTproBNP was an independent predictor of recurrent VTE (hazard ratio, 9.32; P = 0.02).

Conclusions

Elevated NTproBNP is associated with recurrent VTE in acute pulmonary embolism patients.     

Impaired spontaneous thrombolytic activity measured by global thrombosis test in males with metabolic syndrome

Introduction

In patients with metabolic syndrome (MetS), activity of the fibrinolytic system is generally surmised to be decreased through increased plasminogen activator inhibitor-1 (PAI-1) generation. However, there have been no detailed reports describing whether the clot lysis activity is more dominant than increased clot formation activity for production of the thrombotic state in MetS.

Methods

The global thrombosis test (GTT) is a novel method designed to test both clot formation and clot lysis activities under physiological conditions by using non-anticoagulated blood samples in vitro. We used the GTT to examine the thrombotic or thrombolytic states in males with MetS.

Results

Lysis time, which reflects spontaneous clot lysis activity, was significantly longer in MetS subjects (median, 1494 s; range, 865-3596 s; n = 30) than in control subjects (median 1246 s; range, 667-2239 s; n = 53). There was no significant difference between the two groups in occlusion time, which reflects platelet function. The mean level of PAI-1 was significantly higher in MetS subjects than in controls (mean ± SE, 8.7 ± 1.1 and 5.0 ± 0.5 ng/mL, respectively). PAI-1 level and lysis time were significantly correlated (r = 0.400, P < 0.01).

Conclusion

These results suggest that male patients with MetS are more likely than controls to experience a thrombotic state through decreased fibrinolytic activity due to increased PAI-1 generation, and that the GTT is useful for evaluating fibrinolytic activity in vitro.     

Microscopic clot fragment evidence of biochemo-mechanical degradation effects in thrombolysis

Introduction

Although fibrinolytic treatment has been used for decades, the interactions between the biochemical mechanisms and the mechanical forces of the streaming blood remain incompletely understood. Analysis of the blood clot surface in vitro was employed to study the concomitant effect of blood plasma flow and recombinant tissue plasminogen activator (rt-PA) on the degradation of retracted, non-occlusive blood clots. Our hypothesis was that a faster tangential plasma flow removed larger fragments and resulted in faster overall thrombolysis.

Materials and Methods

Retracted model blood clots were prepared in an optical microscopy chamber and connected to an artificial perfusion system with either no-flow, or plasma flow with a velocity of 3 cm/s or 30 cm/s with or without added rt-PA at 2 µg/ml. The clot surface was dynamically imaged by an optical microscope for 30 min with 15 s intervals.

Results

The clot fragments removed during rt-PA mediated thrombolysis ranged in size from that of a single red blood cell to large agglomerates composed of more than a thousand red blood cells bound together by partly degraded fibrin. The average and the largest discrete clot area change between images in adjacent time frames were significantly higher with the faster flow than with the slow flow (14,000 μm² and 160,000 μm² vs. 2200 μm² and 10,600 μm²).

Conclusions

On the micrometer scale, thrombolysis consists of sequential removal of clot fragments from the clot surface. With increasing tangential plasma flow velocity, the size of the clot fragments and the overall rate of thrombolysis increases.     

Ambient temperature affects thrombotic potential at rest and following exercise

Introduction

During exercise, ischemic risk increases, possibly due to changes in coagulation and fibrinolytic activity. Previous research suggests ambient temperature affects resting thrombotic potential, but the effect of heat and cold on hemostasis during exercise is unknown. The purpose of this study was to assess changes in coagulation and fibrinolysis during maximal exercise in hot and cold temperatures, and to compare those responses to exercise under temperate conditions.

Materials & Methods

Fifteen healthy men completed maximal exercise tests in hot (30 °C), temperate (20 °C) and cold (5° - 8 °C) temperatures. Blood samples were obtained before and immediately after exercise and analyzed for concentrations of thrombin-antithrombin III (TAT), active tissue plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1). Results were analyzed by ANOVA.

Results

A main effect of time was observed for TAT (temperate = 1.71 ± 0.82 - 2.61 ± 0.43 ng/ml, hot = 1.81 ± 0.73 - 2.62 ± 0.67 ng/ml, cold = 2.33 ± 0.65 - 2.89 ± 0.81 ng/ml, PRE to POST, respectively) and tPA activity (temperate = 0.72 ± 0.44 - 2.71 ± 0.55 IU/ml, hot = 0.72 ± 0.38 - 2.64 ± 0.61 IU/ml, cold = 0.86 ± 0.45 - 2.65 ± 0.77 IU/ml, PRE to POST, respectively). A trend was observed for the PAI-1 response to exercise (temperate = 14.5 ± 23.7 - 12.3 ± 20.2 IU/ml, hot = 15.1 ± 26.5 - 10.0 ± 15.1 IU/ml, cold = 10.5 ± 10.4 - 7.9 ± 9.7 IU/ml, PRE to POST, respectively, p = 0.08). TAT concentrations were significantly higher in cold compared to temperate and hot conditions.

Conclusion

Coagulation potential is elevated during exposure to cold temperatures. These data suggest that risk of an ischemic event may be elevated in the cold.     

Exploring soluble urokinase plasminogen activator receptor and its relationship with arterial stiffness in a bi-ethnic population: the SAfrEIC-study

Introduction

Elevated soluble urokinase-type plasminogen activator receptor (suPAR) indicates an inflammatory state caused by conditions such as HIV and cancer. Recently suPAR was identified as an indicator of cardiovascular disease (CVD). CVD is highly prevalent in black South Africans, but the potential role of suPAR is unknown. We investigated suPAR as a possible marker of arterial stiffness in Africans and Caucasians.

Methods

This study involved 207 Africans and 314 Caucasians (aged 20-70 yrs). C-reactive protein (CRP) and suPAR were determined in fasting blood samples. We measured blood pressure, pulse wave velocity (PWV) and Windkessel arterial compliance (Cwk).

Results

Africans displayed higher suPAR, CRP, PWV and lower Cwk (p < 0.001) compared to Caucasians. SuPAR was elevated in Africans irrespective of gender and smoking. We found strong relationships between PWV and suPAR (r = 0.27; p < 0.001) and Cwk and suPAR (r = − 0.39; p < 0.001) in the whole group, but found no independent relationship of any arterial stiffness measure and suPAR in Africans after adjustment for confounders. Caucasian men indicated a weak significant independent association between Cwk and suPAR (β = − 0.09; p = 0.028).

Conclusion

Africans had higher levels of suPAR and arterial stiffness than Caucasians (p < 0.001), but there was no independent relationship between arterial stiffness and suPAR in the Africans. It is speculated that due to the inflammatory role of suPAR, it will have stronger relationships with atherosclerosis, which has not yet manifested in this relatively young population group. SuPAR may therefore not be an ideal early marker of cardiovascular dysfunction, but may rather indicate established CVD.     

Tissue plasminogen activator (tPA) activity is a novel and early marker of asymptomatic LEAD in type 2 diabetes

Introduction

Lower extremity arterial disease (LEAD) is often one of the first signs of a generalized atherosclerotic disease in type 1 and type 2 diabetic subjects.

Materials and methods

We studied 143 diabetic subjects at 30-70 years of age, M/F 69/74, 74 with type 1 and 69 with type 2 diabetes, without previously known or suspected lower extremity arterial disease. The relationship between early asymptomatic lower extremity arterial disease and blood levels of HbA1c, lipids and fibrinolysis markers (tPA-activity, tPA mass, PAI-1 activity, tPA-PAI-1 complex) was assessed. In parallel, a group with non-diabetic subjects (n = 80) was studied.

Results

35 (24%) diabetic subjects were classified as having sign(s) of LEAD, defined as having at least one reduced peripheral blood pressure measurement, 28% in type 1 vs 20% in type 2 diabetic subjects (p = NS). In univariate logistic regression analyses age, glycemic level (HbA1c), male gender (only in type 1 diabetic subjects), hypertension and tPA activity (only in type 2 diabetic subjects) were positively associated with LEAD. When markers of fibrinolysis were entered into a multivariate model adjusting for age, hypertension, and HbA1c, only tPA activity remained independently associated with LEAD (p = 0.01) and this was also found in type 2 diabetic subjects (p = 0.05). In type 1 diabetic subjects the increase in odds ratio was non-significant.

Conclusions

Tissue plasminogen activator (tPA) activity may be an independent and early marker for asymptomatic lower extremity arterial disease in diabetic subjects, particularly in type 2 diabetes. Thus an altered fibrinolytic activity could be an early marker of atherosclerosis development in the lower extremities but the cause-effect relationship remains unclear.     

Asymmetric dimethylarginine impairs fibrinolytic activity in human umbilical vein endothelial cells via p38 MAPK and NF-κB pathways

Introduction

Asymmetric dimethylarginine (ADMA) is a potent endogenous inhibitor of nitric oxide (NO) synthase. An increased synthesis and/or a reduced catabolism of ADMA might contribute to the onset and progression of thrombosis. The present study was designed to evaluate the effect of ADMA on fibrinolytic factors in endothelial cells, and to investigate the cellular mechanisms.

Materials and Methods

Human umbilical vein endothelial cells (HUVECs) were treated with different concentrations of ADMA for various periods; Then HUVECs were preincubated with NO precursor (L-arginine), MAPK inhibitors, or NF-κB inhibitor (PDTC) before ADMA treatment to repeat the experiment. Protein levels of tissue plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1), and NF-κB activity were measured by ELISA; mRNA levels of tPA and PAI-1 were assayed by qRT-PCR; The activation of MAPK was characterized by western blot analysis.

Results

(1) ADMA decreased tPA antigen levels in time- and concentration-dependent manners, with the maximum effect of 30 μmol/L ADMA for 48 h (control 109.01 ± 4.15 ng/ml vs ADMA 86.76 ± 5.95 ng/ml, p < 0.01); (2) 30 μmol/L ADMA elevated antigen levels of PAI-1 in a time-dependent manner, with the maximum effect of 30 μmol/L ADMA for 48 h (control 2721.12 ± 278.02 ng/ml vs ADMA 3435.78 ± 22.33 ng/ml, p < 0.05); (3) ADMA reduced tPA mRNA levels and increased PAI-1 mRNA levels; (4) L-arginine, SB203580 (p38 MAPK inhibitor) and PDTC attenuated the effects of ADMA on tPA and PAI-1 significantly. (5) ADMA induced a rapid phosphorylation of p38 MAPK, and stimulated NF-κB activity greatly.

Conclusions

ADMA may accelerate thrombosis development by impairing fibrinolytic activity in vascular via inhibiting nitric oxide production and then activating its downstream p38 MAPK and NF-κB pathways.     

A porcine model of massive, totally occlusive, pulmonary embolism

Background

A reliable, animal model of massive, totally occlusive, pulmonary embolism (PE) is lacking.

Objectives

To design an animal model of totally occlusive PE and to challenge the model by a plasminogen activator.

Methods

In eight anaesthetized pigs (~ 90 kg) a massive preformed autologous thrombus was injected into the caval vein. One animal was autopsied to assess the extent of injected clot, whereas in the other animals extracorporeal life support (ECLS) was initiated and continued for three hours. These animals received 100 mg rt-PA. Blood gases, coagulation tests, creatine kinase (CK), lactate dehydrogenase (LDH), end-tidal CO2, systemic and pulmonary artery blood pressures and flow were registered.

Results

All animals went into circulatory arrest within 2 minutes after injection of the thrombus. In the animal where ECLS was not started, autopsy relieved a totally occlusive embolus of the pulmonary artery. The ECLS maintained a systemic blood flow of 6-8 L/min with adequate oxygenation and CO2-removal. However, lactate increased and base-excess became negative. Ddimer increased, fibrinogen decreased, and CK and LDH increased. All seven animals were weaned from ECLS. Despite the rt-PA treatment, the animals had at that time low end tidal CO2/PaCO2 ratio and increased mean pulmonary arterial pressure, suggesting a significant amount of embolic material remaining in the pulmonary artery.

Conclusion

This model of massive, totally occlusive, pulmonary embolism mimics well fatal PE seen in the clinic, and has the potential for use in testing of new therapeutic interventions.     

Active search for chronic thromboembolic pulmonary hypertension does not appear indicated after acute pulmonary embolism

Introduction

Chronic thromboembolic pulmonary hypertension (CTEPH) is a life threatening but often, by pulmonary endarterectomy, curable disease. The incidence of CTEPH after an acute pulmonary embolism (PE) appears to be much higher than previously thought. Systematic follow-up of patients after PE might increase the number of diagnosed CTEPH patients.

Aim

To study whether, compared to current clinical practice, a systematic search for CTEPH in patients after acute PE would increase the number of patients diagnosed with symptomatic, potentially treatable CTEPH.

Methods

Consecutive patients with a prior diagnosis of acute PE were presented with a questionnaire, designed to establish the presence of either new or worsened dyspnea after the acute PE episode. If so, patients were evaluated for the presence of CTEPH.

Results

PE patients (n = 110; 56 ± 18 years) were included after a median follow-up of three years. Overall mortality was 34% (37 patients); 1 patient had died due to CTEPH.In total 62 out of 69 questionnaires were returned; 23 patients reported new or worsened dyspnea related to the PE episode, and qualified for additional testing. In 2 patients, CTEPH was already diagnosed prior to this study. None of the remaining patients met the criteria for the diagnosis of CTEPH. The overall incidence of 2.7% (3/110; 95%CI 0.6-7.8%) is in agreement with earlier reported incidences.

Conclusion

Our findings do not point to a role for a systematic search and pro-active approach towards patients with a recent history of pulmonary embolism to increase the number of patients diagnosed with potentially treatable CTEPH.     

Thrombolytic efficacy of tissue plasminogen activator-loaded echogenic liposomes in a rabbit thrombus model

Introduction

Ultrasound (US)-enhanced thrombolytic treatment protocols currently in clinical trials for stroke applications involve systemic administration of tissue plasminogen activator (tPA; Alteplase), which carries a risk of adverse bleeding events. The present study aimed to compare the thrombolytic efficacy of a tPA-loaded echogenic liposome (ELIP) formulation with insonification protocols causing rapid fragmentation or acoustically-driven diffusion.

Materials and Methods

Thrombi were induced in the abdominal aortas of male New Zealand white rabbits (2-3 kg) using thrombin and a sclerosing agent (sodium ricinoleate) after aortic denudation with a balloon catheter. Thrombolytic and cavitation nucleation agents (200 μg of tPA alone, tPA mixed with 50 μg of a microbubble contrast agent, or tPA-loaded ELIP) were bolus- injected proximal to the clot through a catheter introduced into the abdominal aorta from the carotid artery. Clots were exposed to transabdominal color Doppler US (6 MHz) for 30 minutes at a low mechanical index (MI = 0.2) to induce sustained bubble activity (acoustically-driven diffusion), or for 2 minutes at an MI of 0.4 to cause ELIP fragmentation. Degree of recanalization was determined by Doppler flow measurements distal to the clots.

Results

All treatments showed thrombolysis, but tPA-loaded ELIP was the most efficacious regimen. Both US treatment strategies enhanced thrombolytic activity over control conditions.

Conclusions

The thrombolytic efficacy of tPA-loaded ELIP is comparable to other clinically described effective treatment protocols, while offering the advantages of US monitoring and enhanced thrombolysis from a site-specific delivery agent.     

The effect of clopidogrel besylate and clopidogrel hydrogensulfate on platelet aggregation in patients with coronary artery disease: a retrospective study

Background

Recently several alternative forms of the original clopidogrel hydrogensulfate (CHS) were spread worldwide. A large amount of such drugs turned out to be clopidogrel besylate (CB). Only three studies, involving healthy volunteers, investigated the antiplatelet effect of CB, whereas its attribute remained unexplored in the case of patients with cardiovascular diseases. This retrospective study aimed to evaluate the difference between the antiplatelet effects of two clopidogrel formulas, CHS and CB, on patients with coronary artery diseases.

Methods

Data of 150 patients with previous CHS treatment were investigated. According to the documentations, the CHS therapy was shifted to CB. 94 patients of the selected population received dual antiplatelet therapy, clopidogrel and aspirin. The antiplatelet effects of CHS and CB were compared by ADP induced platelet aggregation measurements using light transmission aggregometry.

Results

Irrespective of the therapeutic combinations the performed statistical investigations failed to show significant difference (p = 0.30) between the effect of CB (AGGmax_CB: 27.6 ± 13.7%) or CHS (AGGmax_CHS: 29.0 ± 15.3%) on the ADP induced platelet aggregation. Insignificant deviations were found in both forms of clopidogrel salts, either in the lack (AGGmax_CB : 32.5 ± 14,2%; AGGmax_CHS: 34,0 ± 16,1%; p = 0,29) or in the presence of aspirin (AGGmax_CB: 24.7 ± 12,5%; AGGmax_CHS: 26,0 ± 14,1%; p = 0,31).

Conclusion

Our results indicated that both CB and CHS had an identical inhibitory effect on ADP induced platelet aggregation in patients with cardiovascular diseases. Moreover their efficiency showed no overall significant difference in the case of dual antiplatelet therapy with aspirin as well. However there might be an inter- and intraindividual variability between the two clopidogrel formulas.     

Central thromboembolism is a possible predictor of right heart dysfunction in normotensive patients with acute pulmonary embolism

Background

Right heart dysfunction is a crucial factor in risk stratification of normotensive patients with pulmonary embolism. Apart from biomarkers, determinants of right heart dysfunction in this group of patients are not yet well established.

Aim and method

In order to identify such determinants, we analysed data of 252 patients with acute pulmonary embolism admitted to our hospital in 2008.

Results

69 out of 140 patients showed right heart dysfunction by echocardiography within 24 hours after diagnosis, 71 did not. Right ventricular dysfunction was significantly more frequent in patients with central clots on computed tomography (p = 0.004), a history of syncope (p < 0.001) and among women on oral contraceptives (p = 0.003). In multiple regression analysis, only central thromboembolism (p < 0.001) was identified as individual predictor of right ventricular dysfunction. Age, gender, body mass index, idiopathic or recurrent thromboembolism, duration of symptoms, preceding surgery, room air oxygen saturation, carcinoma, hypertension, diabetes, renal disease, congestive left heart failure and concomitant lung disease were equally distributed. In comparison with NT-pro brain natriuretic peptide (PPV 67%, NPV 75%, p = 0.782) and troponin I (PPV 76%, NPV 62%, p = 0.336), central thromboembolism has shown to have a greater statistical power in predicting right heart dysfunction in normotensive patients with pulmonary embolism (PPV 78%, NPV 88%, p < 0.001).

Conclusion

Among normotensive patients with acute pulmonary embolism, those with central clots seem to be at greater risk for echocardiographically evaluated right ventricular dysfunction.     

Echocardiographic assessment of pulmonary arterial pressure in the follow-up of patients with pulmonary embolism

Background

Echocardiography remains a clinically useful screening test for chronic thromboembolic pulmonary hypertension (CTEPH) in patients with a history of pulmonary embolism (PE). To devise an effective screening strategy, the definition of a high-risk group is necessary.

Methods

We examined a total of 744 patients with acute symptomatic pulmonary embolism (PE) who were enrolled in a Spanish multicenter study. Patients were monitored every 6 months during the first two years, and then once a year thereafter. Transthoracic echocardiography was used to screen patients with a clinical suspicion of CTEPH during follow-up. Pulmonary arterial hypertension was defined as an estimated pulmonary artery systolic pressure (PAP) > 50 mm Hg. The index thromboembolic episode was considered severe if: (a) the patient was immobilized for medical reasons; or (b) systolic blood pressure was less than 90 mm Hg; or (c) troponin T values were above the reference range.

Results

The incidence of PAP > 50 mm Hg at 36 months was 8.3% (95% confidence interval = 4.6%-14.5%). Statistical analysis showed a highly significant association between a severe index thromboembolic episode and the subsequent detection of PAP > 50 mm Hg on echocardiography, with high positive likelihood ratio (2.40) and negative predictive value (> 0.97).

Conclusions

Patients with a severe index thromboembolic episode would constitute a high-risk group for the development of CTEPH. This group of patients should be subjected to a strict follow-up protocol.     

Bolus tenecteplase for right ventricle dysfunction in hemodynamically stable patients with pulmonary embolism

Introduction

The clinical benefit of thrombolytic treatment over heparin in patients with pulmonary embolism without hemodynamic compromise remains controversial. In these patients bolus tenecteplase has the potential to provide an effective and safe thrombolysis.

Methods

We evaluated the effect of tenecteplase on right ventricle dysfunction (RVD) assessed by echocardiography in hemodynamically stable patients with PE in a multicenter, randomized, double-blind, placebo-controlled study. RVD was defined as right/left ventricle end-diastolic dimension ratio > 1 in the apical 4-chamber view. Patients were randomized to receive weight-adjusted single-bolus tenecteplase or placebo. All patients received unfractionated heparin. Reduction of RVD at 24 hours was the primary efficacy end-point and was evaluated by an independent committee unaware of treatment allocation.

Results

Overall, 58 patients were randomized. Echocardiograms were adequate for efficacy analysis in 51 patients, 23 randomized to tenecteplase and 28 to placebo. The reduction of right to left ventricle end-diastolic dimension ratio at 24 hours was 0.31 ± 0.08 in patients randomized to tenecteplase as compared to 0.10 ± 0.07 in patients randomized to placebo (p = 0.04). One patient randomized to tenecteplase suffered a clinical event (recurrent pulmonary embolism) in comparison to three patients randomized to placebo (1 recurrent pulmonary embolism; 1 clinical deterioration and 1 non pulmonary embolism-related death). Two non fatal major bleedings occurred with tenecteplase (1 intracranial) and one with placebo.

Conclusion

In hemodynamically stable patients with PE, treatment with single bolus tenecteplase is feasible at the same dosages used for acute myocardial infarction and is associated with reduction of RVD at 24 hours. Whether this benefit is associated with an improved clinical outcome without excessive bleeding is currently explored in a large clinical trial.     

Activated inflammatory cells participate in thrombus size through tissue factor and plasminogen activator inhibitor-1 in acute coronary syndrome: Immunohistochemical analysis

Introduction

Recent studies have suggested that circulating inflammatory cells augment the growth of thrombus in acute coronary syndrome (ACS). We therefore immunohistochemically analyzed thrombi in aspirates obtained from patients immediately after the onset of ACS.

Materials and Methods

Two hundred twenty samples were studied. Total thrombus area, white thrombus area, and red thrombus area were measured. As antibodies in immunohistochemical staining, myeloperoxidase (MPO), CD66b, CD68, p-selectin, tissue factor (TF) and PAI-1were employed respectively.

Results

The ratios of areas of red and white thrombi correlated with whole sample areas of enlarged thrombi (r = 0.48, p < 0.001). The immunohistochemical findings revealed granulocytes and macrophages aggregated around p-selectin-positive platelets that shared the boundary between white and red thrombi, a region where MPO and CD66b expression was abundant in neutrophils. The ratios (%) of MPO- and CD66b-positive cells significantly correlated with whole sample areas (r = 0.50; p < 0.001 and r = 0.49; p < 0.001, respectively). Neutrophils and macrophages within thrombi were positive for TF and PAI-1. Along the boundary between red and white thrombi, TF and PAI-1 positivity coincided with MPO-, CD66b- and CD68-positive cells. The ratios of cells positive for both TF and PAI-1 in this area significantly correlated with the whole sample area (r = 0.43, p < 0.001 and r = 0.60, p < 0.001, respectively).

Conclusions

These results suggested that enhanced activation of peripheral neutrophils together with increased TF and PAI-1 expression might comprise a considerable portion of thrombus enlargement.     

Haemostatic system, biochemical profiles, kynurenines and the prevalence of cardiovascular disease in peritoneally dialyzed patients

Introduction

The haemostatic and biochemical abnormalities participate in the progression of cardiovascular disease (CVD) in peritoneally dialysed (PD) patients. Recently, the role of kynurenine (KYN) pathway of tryptophan (TRP) degradation in the development of CVD has been postulated.

Materials and methods

The present study was undertaken to investigate haemostatic parameters, biochemical profiles and kynurenines in PD patients both with and without CVD compared to age- and sex-matched healthy controls.

Results

The multiple biochemical abnormalities were present in PD patients, particularly in those with CVD. Tissue factor (TF), its inhibitor (TFPI), prothrombin fragment 1 + 2 (F_1 + 2)_, urokinase-type plasminogen activator (uPA), its soluble receptor (suPAR), plasmin/antiplasmin (PAP) complexes, KYN, kynurenic (KYNA) and quinolinic (QA) acids levels were significantly higher, whereas TRP was significantly lower in the PD patients than in the controls. Tissue-type plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1) were higher in the patients with CVD than in the patients without CVD and controls. PD patients with CVD had higher F_1 + 2, and they had lower suPAR and KYNA levels compared with PD patients without CVD. KYNA was positively associated with TFPI, whereas its was inversely associated with F_1 + 2 both in the whole PD group and in CVD patients. Logistic regression analysis showed that low KYNA, high glucose, low HDL-cholesterol levels and the duration of dialysis treatment were independently associated with the presence of CVD in PD patients.

Conclusions

The present study suggests a relationship between kynurenine pathway of tryptophan degradation, haemostatic and biochemical disturbances and CVD prevalence in peritoneally dialyzed patients.     

Impaired glucose metabolism and type 2 diabetes are associated with hypercoagulability: potential role of central adiposity and low-grade inflammation--the Hoorn Study

Introduction

Type 2 diabetes (DM2) is associated with greater risk for cardiovascular disease (CVD), which may, at least partially, be explained by prothrombotic alterations. We therefore investigated; first, the extent to which individuals with impaired glucose metabolism (IGM) and/or DM2 had greater levels of thrombin generation than those with normal glucose metabolism (NGM); and second, whether any differences were independent of other cardiovascular risk factors, such as smoking, hypertension, dyslipidaemia, (micro)albuminuria, glycemic control and (central) adiposity, and/or were potentially ‘mediated’ by low-grade inflammation (high-sensitivity C-reactive protein (hsCRP)).

Materials and methods

We studied 744 individuals from the Hoorn Study (275 NGM, 176 IGM and 293 DM2, mean age 68.6 ± 7.1 years). Thrombin generation in platelet-poor plasma was measured using the Calibrated Automated Thrombogram and three parameters were derived: lag time, peak height and endogenous thrombin potential (ETP). Data were analyzed with multiple linear regression analyses.

Results

After adjustment for age, sex, prior CVD and smoking status, individuals with IGM or DM2 had a longer lag time [ß = 0.14 min (95% CI: 0.02; 0.26)], higher peak height [ß = 7.29 nM (− 1.33; 15.91)] and ETP [ß = 35.65nM*min (0.97; 70.34)] than those with NGM. These differences were attenuated to ß = 0.06 min (− 0.07; 0.19), 3.82 nM (− 5.46; 13.10) and 16.34 nM*min (− 20.92; 53.59), respectively, when further adjusted for waist circumference and hsCRP.

Conclusion

Individuals with IGM or DM2 had up to 4% higher thrombin generation compared with NGM, which may be explained, to a great extent, by the greater levels of central adiposity and related low-grade inflammation characterizing these individuals.     

Safety assessment and pharmacodynamics of a novel ultra low molecular weight heparin (RO-14) in healthy volunteers--a first-time-in-human single ascending dose study

Introduction

RO-14 is a novel ultra low molecular heparin. The purpose of this study was to evaluate the safety and pharmacodynamic profile of RO-14 in healthy males.

Materials and methods

We conducted a two-stage, single-center, open-label, randomized study. Two cohorts of 6 volunteers were randomly assigned to 12 single, ascending subcutaneous doses (1750-19950 IU of anti-FXa activity) in an alternating crossover fashion. Safety was assessed by spontaneous/elicited adverse events, medical examination and laboratory tests. Anti-FXa activity and anti-FIIa activity were assessed throughout the 24 hours after dosing. Dose proportionality and linearity of the anti-FXa activity were evaluated.

Results

All doses were well tolerated and there were no bleeding events. At the lowest dose, anti-FXa activity A_max was 0.16 (± 0.02) IU/mL and AUC_0-24 was 1.11 (± 0.24) IU*h/mL, At the highest dose anti-FXa activity A_max was 1.67 (± 0.15) IU/mL; AUC_0-24 was 21.48 (± 4.46) IU*h/mL and t½ was 8.05 h. Mean T_max (all doses) was 2.86 (± 0.39) h. RO-14 showed proportional and linear pharmacodynamics [normalized A_max among doses (p = 0.594) and normalized AUC_0-24 (p = 0.092), correlations between A_max-dose (R² = 0.89, p < 0.001) and AUC_0-24-dose (R² = 0.86, p < 0.001)]. Anti-FIIa activity was below the detection limit (0.1 IU/ml) at all dose levels. No clinically significant changes were observed in the platelet count, APTT, PT, TT, fibrinogen and antithrombin.

Conclusions

In this phase I study, RO-14 exhibited a good safety profile, anti-FXa activity for either prophylaxis or treatment of venous thromboembolism, linear pharmacodynamics, a longer elimination half-life than currently marketed low molecular weight heparin and no anti-FIIa activity.     

Platelet glycoprotein VI (GPVI) for early identification of acute coronary syndrome in patients with chest pain

Background

Platelet glycoprotein VI (GPVI) is elevated in patients with acute coronary syndrome (ACS), stroke and associated with acute coronary events. GPVI may be helpful to distinguish an imminent ACS from non-coronary (NC) causes in patients with chest pain who were transferred to chest pain unit, before the myocardial necrosis is evident with classical biomarkers.

Methods

Based on the findings of our previous studies, we consecutively examined 1004 patients with chest pain in a prospective study design. ACS was found in 416 (41.4%), stable angina pectoris (SAP) in 233 (23.2%), and NC causes of chest pain (hypertension, musculoskeletal disease, pulmonary embolism, myocarditis, cardiophobia) in 355 patients (35.4%). Platelet surface expression of GPVI was measured by flow cytometry.

Results

Patients with ACS showed significantly enhanced GPVI expression levels compared to patients with SAP or NC causes of chest pain (ACSvs.SAP(mean fluorescence intensity (MFI) ± SD):18.9 ± 7.4vs.17.9 ± 9.5;P = 0.028;ACSvs.NC:15.4 ± 6.9;P = 0.002). Elevated GPVI expression was associated with ACS independent of markers of myocardial necrosis like troponin and creatine kinase-MB. Patients with an elevated GPVI expression (MFI ≥ 18.6) had a poorer clinical outcome than patients with baseline GPVI expression in regard to composite cumulative survival that included myocardial infarction, stroke, and cardiovascular death at three months (Log rank;P = 0.025).

Discussion

Platelet GPVI surface expression is enhanced in patients at risk for an ACS and is an early marker for imminent acute coronary events in patients with chest pain.