Soluble Thrombomodulin Levels Research Articles

Prognostic value of plasma levels of thrombomodulin and von Willebrand factor in Egyptian children with acute lymphoblastic leukemia

Acute lymphoblastic leukemia is the most common childhood malignancies, representing nearly one-third of all pediatric cancers. Thrombomodulin is a membrane glycoprotein in the vascular endothelium. Its plasma level depends on the integrity of the endothelium. Soluble thrombomodulin is derived from injured endothelial cells or proteolytically cleaved from thrombomodulin by proteases. Von Willebrand factor is a blood glycoprotein involved in homeostasis. Its plasma level increases in neoplastic diseases and arises from adverse changes in the endothelium. Severe endothelial dysfunction is present during the acute phase of acute lymphoblastic leukemia. Plasma levels of von Willebrand factor and soluble thrombomodulin have been used as indexes of endothelial dysfunction. The aim of this study was to assess serum soluble thrombomodulin and von Willebrand factor levels in children with acute lymphoblastic leukemia during acute phase of the disease to assess their potential prognostic value. Forty patients with acute lymphoblastic leukemia included 26 males and 14 females with their ages ranged from 2 to 10 years and 20 healthy children of matched age and sex were included in this study. We analyzed serum soluble thrombomodulin and von Willebrand factor levels by enzyme-linked immunosorbent assay. In children with acute lymphoblastic leukemia, there was a significant increase in soluble thrombomodulin, and von Willebrand factor levels during the acute phase of the disease. Children with an unfavorable outcome had higher levels of thrombomodulin and von Willebrand factor levels. Serum thrombomodulin and von Willebrand factor levels as a parameter of endothelial dysfunction during the acute phase of acute lymphoblastic leukemia might represent an additional prognostic marker in childhood acute lymphoblastic leukemia.

Soluble Thrombomodulin and Endothelial Cell Protein C Receptor Levels in Patients with Cerebral Venous and Sinus Thrombosis

Introduction: Cerebral venous and sinus thrombosis (CVST) is a potentially serious disorder. In search for underlying coagulopathy in CVST patients, we measured plasma levels of soluble thrombomodulin (sTM) and soluble endothelial protein C receptor (sEPCR) in patients with CVST as candidate biomarkers of thrombosis. Materials and Methods: The concentrations of sTM and sEPCR in 19 CVST patients without protein C and protein S deficiency were measured by ELISA and compared with 53 healthy controls. We also examined the association between those plasma markers and the risk of CVST. Results: Significant difference was identified in plasma sTM level between CVST patients and controls (0.95 ± 0.09 vs. 1.26 ± 0.12 ng/ml, respectively; p = 0.038). The plasma level of sEPCR was not different between groups, and no significant quantitative association was identified between the risk of CVST and the levels of sEPCR. The adjusted odds ratio for CVST associated with low (<10th percentile) levels of sTM was 2.3 (95% CI: 1.29-20.08; p = 0.012) after adjustment for confounding factors. Conclusion: Decreased plasma sTM concentration may be associated with an increased risk of CVST.

Clot lysis phenotype and response to recombinant factor VIIa in plasma of haemophilia A inhibitor patients

Recombinant activated factor VII (rFVIIa) is a haemostatic agent that is used for the treatment of haemophilia A patients with inhibitors. However, clinical response to rFVIIa is variable and unpredictable with currently available assays. We investigated the anti-fibrinolytic effects of rFVIIa in relation to thrombin generation (TG) and other haemostatic parameters in haemophilia A patients with inhibitors. After addition of rFVIIa to plasma, the clot-lysis assay, TF-dependent TG, TF-independent TG and parameters involved in coagulation, anticoagulation and fibrinolysis were assessed. The clot-lysis test distinguished two groups of patients: a group with a normal and a group with impaired anti-fibrinolytic response to rFVIIa. Our results showed a dose-dependent increase in TF-dependent TG and TF-independent TG in all individuals. There was a significant difference in TF-independent TG parameters between the normal and impaired response groups. In addition, there was a difference between the normal and impaired response group in prothrombin time, which could be explained by significantly higher levels of coagulation factors in the normal response group, and soluble thrombomodulin. In conclusion, we observed different in vitro responses following rFVIIa addition in plasma of patients with haemophilia A and inhibitors, which could be partially attributed to levels of procoagulant proteins and soluble thrombomodulin.

Normal prothrombinase activity, increased systemic thrombin activity, and lower antithrombin levels in patients with disseminated intravascular coagulation at an early phase of trauma: Comparison with acute coagulopathy of trauma-shock

We tested the hypotheses that an increase in systemic thrombin activity occurs in both disseminated intravascular coagulation (DIC) with the fibrinolytic phenotype and in acute coagulopathy of trauma shock (ACoTS), and that the patients diagnosed as having ACoTS overlap or are identical with those diagnosed as having DIC. We made a prospective study of 57 trauma patients, including 30 patients with DIC and 27 patients without DIC. Patients with ACoTS, defined as a prothrombin time ratio >1.2, were also investigated. We included 12 healthy volunteers as controls. The levels of soluble fibrin, antithrombin, prothrombinase activity, soluble thrombomodulin, and markers of fibrin(ogen)olysis were measured on days 1 and 3 after the trauma. The systemic inflammatory response syndrome and the Sequential Organ Failure Assessment were scored to evaluate the extent of inflammation and organ dysfunction. Patients with DIC showed more systemic inflammation and greater Sequential Organ Failure Assessment scores and were transfused with more blood products than the patients without DIC. On day 1, normal prothrombinase activity, increased soluble fibrin, lesser levels of antithrombin, and increased soluble thrombomodulin were observed in patients with DIC in comparison with controls and non-DIC patients. These changes were more prominent in patients with DIC who met the overt criteria for DIC established by the International Society on Thrombosis and Haemostasis. Multiple regression analysis showed that antithrombin is an independent predictor of high soluble fibrin in DIC patients. Greater levels of fibrin and fibrinogen degradation products, D-dimer, and the fibrin and fibrinogen degradation products/D-dimer ratio indicated increased fibrin(ogen)olysis in DIC patients. Almost all ACoTS patients overlapped with the DIC patients. The changes in the measured variables in ACoTS patients coincided with those in DIC patients. Normal prothrombinase activity and insufficient control of coagulation give rise to systemic increase in thrombin generation and its activity in patients with DIC with the fibrinolytic phenotype at an early phase of trauma. The same is true in patients with ACoTS, and shutoff of thrombin generation was not observed.

Microvascular Damage in Type 1 Diabetic Patients Is Reversed in the First Year After Simultaneous Pancreas–Kidney Transplantation

Simultaneous pancreas-kidney transplantation (SPK) is an advanced treatment option for type 1 diabetes mellitus (DM) patients with microvascular disease including nephropathy. Sidestreamdarkfield (SDF) imaging has emerged as a noninvasive tool to visualize the human microcirculation. This study assessed the effect of SPK in diabetic nephropathy (DN) patients on microvascular alterations using SDF and correlated this with markers for endothelial dysfunction. Microvascular morphology was visualized using SDF of the oral mucosa in DN (n = 26) and SPK patients (n = 38), healthy controls (n = 20), DM1 patients (n = 15, DM ≥ 40 mL/min) and DN patients with a kidney transplant (KTx, n = 15). Furthermore, 21 DN patients were studied longitudinally up to 12 months after SPK. Circulating levels of angiopoietin-1 (Ang-1), angiopoietin-2 (Ang-2) and soluble thrombomodulin (sTM) were measured using ELISA. Capillary tortuosity in the DN (1.83 ± 0.42) and DM ≥ 40 mL/min (1.55 ± 0.1) group was increased and showed reversal after SPK (1.31 ± 0.3, p < 0.001), but not after KTx (1.64 ± 0.1). sTM levels were increased in DN patients and reduced in SPK and KTx recipients (p < 0.05), while the Ang-2/Ang-1 ratio was normalized after SPK and not after KTx alone (from 0.16 ± 0.04 to 0.08 ± 0.02, p < 0.05). Interestingly, in the longitudinal study, reversal of capillary tortuosity and decrease in Ang-2/Ang-1 ratio and sTM was observed within 12 months after SPK. SPK is effective in reversing the systemic microvascular structural abnormalities in DN patients in the first year after transplantation.

Association of the Thrombomodulin Gene c.1418C&gt;T Polymorphism With Thrombomodulin Levels and With Venous Thrombosis Risk

To investigate the association of the THBD c.1418C>T polymorphism, which encodes for the replacement of Ala455 by Val in thrombomodulin (TM), with venous thromboembolism (VTE), plasma soluble TM, and activated protein C levels. In addition, human umbilical vein endothelial cells (HUVEC) isolated from 100 umbilical cords were used to analyze the relation between this polymorphism and THBD mRNA and TM protein expression. The THBD c.1418C>T polymorphism was genotyped in 1173 patients with VTE and 1262 control subjects. Levels of soluble TM and activated protein C were measured in 414 patients with VTE (not on oral anticoagulants) and 451 controls. HUVECs were genotyped for the polymorphism and analyzed for THBD mRNA and TM protein expression and for the ability to enhance protein C activation by thrombin. The 1418T allele frequency was lower in patients than in controls (P<0.001), and its presence was associated with a reduced VTE risk, reduced soluble TM levels, and increased circulating activated protein C levels (P<0.001). In cultured HUVEC, the 1418T allele did not influence THBD expression but was associated with increased TM in cell lysates, increased rate of protein C activation, and reduced soluble TM levels in conditioned medium. The THBD 1418T allele is associated with lower soluble TM, both in plasma and in HUVEC-conditioned medium, and with an increase in functional membrane-bound TM in HUVEC, which could explain the increased activated protein C levels and the reduced VTE risk observed in individuals carrying this allele.

Dengue Virus Enhances Thrombomodulin and ICAM-1 Expression through the Macrophage Migration Inhibitory Factor Induction of the MAPK and PI3K Signaling Pathways

Dengue virus (DV) infections cause mild dengue fever (DF) or severe life-threatening dengue hemorrhagic fever (DHF). The mechanisms that cause hemorrhage in DV infections remain poorly understood. Thrombomodulin (TM) is a glycoprotein expressed on the surface of vascular endothelial cells that plays an important role in the thrombin-mediated activation of protein C. Prior studies have shown that the serum levels of soluble TM (sTM) and macrophage migration inhibitory factor (MIF) are significantly increased in DHF patients compared to levels in DF patients or normal controls. In this study, we investigated how MIF and sTM concentrations are enhanced in the plasma of DHF patients and the potential effect of MIF on coagulation through its influence on two factors: thrombomodulin (TM) and intercellular adhesion molecule-1 (ICAM-1) in endothelial cells and monocytes. Recombinant human macrophage migration inhibitory factor (rMIF) was used to treat monocytic THP-1 cells and endothelial HMEC-1 cells or primary HUVEC cells. The subsequent expression of TM and ICAM-1 was assessed by immunofluorescent staining and flow cytometry analysis. Additionally, the co-incubation of THP-1 cells with various cell signaling pathway inhibitors was used to determine the pathways through which MIF mediated its effect. The data provided evidence that severe DV infections induce MIF expression, which in turn stimulates monocytes or endothelial cells to express TM and ICAM-1 via the Erk, JNK MAPK and the PI3K signaling pathways, supporting the idea that MIF may play an important role as a regulator of coagulation.

Soluble thrombomodulin levels in plasma of multiple sclerosis patients and their implication

Thrombomodulin (TM) on the cell-surface of cerebrovascular endothelial cells (CECs) is released into blood upon CEC damage. TM promotes activation of protein C (APC), an anticoagulant, anti-inflammatory, neuroprotective molecule that protects CECs and impedes inflammatory cell migration across the blood–brain barrier (BBB). Multiple sclerosis (MS) is associated with CEC damage and BBB dysfunction. We evaluated soluble TM (sTM) levels as a biomarker of BBB integrity and whether glatiramer acetate (GA) influenced sTM levels in MS patients. sTM levels quantified by 2-site ELISA from sera of healthy controls and systemic lupus erythematosus (SLE) patients (CEC-damage positive control) were compared with levels from patients with relapsing–remitting (RRMS) or secondary-progressive MS (SPMS), stratified as: RRMS/GA/no relapse, RRMS/GA/in relapse, RRMS no GA/no relapse, RRMS/no GA/in relapse; and SPMS/no GA. Additionally, soluble endothelial protein C receptor (sEPCR) levels were assessed in the non-stratified MS group, SLE patients, and controls. sTM levels were highest in RRMS patients taking GA with or without relapse, followed in decreasing order by SLE, RRMS/no GA/in relapse, SPMS, RRMS/no GA/no relapse, healthy controls. sEPCR levels were highest in MS patients, then SLE, then controls. sTM may be a useful biomarker of BBB integrity in RRMS patients. Further evaluation of sEPCR is needed. The finding that the highest sTM levels were in RRMS patients taking GA is interesting and warrants further investigation.

Hemostatic changes after 1 month of thalidomide and dexamethasone therapy in patients with multiple myeloma

Thromboembolic events (TEE) are a serious clinical problem in multiple myeloma (MM) patients receiving thalidomide (T). Thirty-one MM patients were tested on diagnosis and after 2 and 4 weeks of therapy with T alone, or T in combination with dexamethasone (TD). Closure time (CT) in PFA-100 and P-selectin expression were assessed, as well as plasma levels of thrombin-antithrombin complexes (TAT), D-dimer (DD), soluble thrombomodulin (sTM) and von Willebrand factor antigen (vWF:Ag), along with the activity of coagulation factor VII and factor VIII. The concentration of vascular endothelial growth factor and its type 1 and 2 receptors were also assayed. On diagnosis, significantly prolonged median CT with both used cartridges, elevated P-selectin expression, DD concentration, TAT, vWF:Ag and factor VIII and factor VII activity were seen in the patient group as compared to controls. Therapy with these regimens caused marked shortening of CT with both cartridges. Treatment with TD leads to the significant increase in CD62P expression on platelets. Median TAT value increased significantly in relation to baseline after therapy with both regimens. Factor VIII activity exceeded 150 % in all patients after 2 weeks of TD therapy and was markedly elevated compared to baseline. One month of TD therapy significantly increased sTM concentration. These results demonstrate the enhanced platelet and coagulation system activation already present in MM patients on diagnosis, which is further increased by antimyeloma therapy. These changes are more pronounced after TD therapy and may promote TEE. Tested angiogenesis marker levels are elevated already on diagnosis, do not change after therapy and have no significant impact on the coagulation system in patients with MM.

Effect of secondary hyperparathyroidism treatment with cinacalcet on selected adipokines and markers of endothelial injury in hemodialysis patients: a preliminary report

Disturbances in endothelial function, adipokines, and mineral metabolism due to secondary hyperparathyroidism (SHPT) shorten the lifespan in hemodialysis (HD) patients. The aim of the study was to evaluate the effect of SHPT treatment with cinacalcet on selected adipokines and markers of endothelial injury in HD patients. Soluble thrombomodulin (sTM), E-selectin, leptin, and adiponectin levels were measured in patients with SHPT at baseline and at 6 months of cinacalcet treatment. A total of 18 patients completed the study. SHPT treatment with cinacalcet decreased calcium, phosphate, and intact parathormone (iPTH) levels; however, no significant changes in sTM, E-selectin, leptin, or adiponectin were observed. iPTH levels after treatment correlated with age (r = -0.51, P = 0.031), mean cinacalcet dose (r = 0.65, P = 0.004), as well as baseline calcium (r = 0.65 P = 0.003), iPTH (r = 0.59, P = 0.01), E-selectin (r = 0.56, P = 0.016), and leptin (r = -0.49, P = 0.039). Cinacalcet treatment does not affect the markers of endothelial function and selected adipokines. Effectiveness of treatment may be modulated by E-selectin and leptin.

Biochemical predictors of cardiac rhythm at 1year follow-up in patients with non-valvular atrial fibrillation.

C-reactive protein (CRP) is increased in patients with atrial fibrillation (AF) and it might predict the lack of sinus rhythm maintenance in the long term follow-up. Scarce data have been reported relating endothelial dysfunction and/or haemostatic or platelet markers to sinus rhythm maintenance in AF patients. We aimed to determine whether some biochemical markers including soluble thrombomodulin (sTM), soluble P-selectin, thrombin-antithrombin (TAT) complex and CRP levels could predict sinus rhythm maintenance at 1year follow-up in non-valvular AF patients. 130 patients (70 males, mean age 67±13years) with newly diagnosed AF naïve of antithrombotic or antiplatelet therapy were studied. Baseline CRP, P-selectin, sTM and TAT levels were compared to those of 20 matched-healthy subjects in sinus rhythm. AF patients had significantly higher plasma CRP (10.5±2.2 vs 3.25±0.3mg/l, p=0.001), P-selectin (219±141 vs 126ng/ml, p=0.01), and TAT (54±237 vs 2.7±3.3ng/l, p=0.001) levels compared with controls. Soluble TM levels, although higher than controls, did not reach statistical significance. Multivariate regression analyses showed that elevated CRP (OR=4.8, p=0.02) and sTM (OR=1.05, p=0.04) were the only two predictors of lack of sinus rhythm at 1year follow-up. An altered inflammatory, haemostatic, endothelial and platelet environment exists in newly diagnosed AF patients. CRP and sTM levels might be used as biochemical markers to predict the status of cardiac rhythm at 1year follow-up in some AF patients.

Increased Plasma Soluble Thrombomodulin Levels in Cardioembolic Stroke

Soluble thrombomodulin (sTM) has been proposed as a potential marker of ischemic stroke. Results from previous studies remain controversial among different populations. We performed an analysis of plasma levels of sTM in Thai patients with acute ischemic stroke and determined whether sTM levels correlate with stroke subtypes, severity, and risk factors. Ninety-three patients and 76 controls were enrolled. Blood samples were obtained within 24 hours after stroke onset. Plasma sTM levels, measured using quantitative enzyme-linked immunosorbent assay, were significantly higher in patients than controls (P < .005), with the mean ± standard deviation (SD) levels of 3.08 ± 1.05 and 2.57 ± 1.15 ng/mL, respectively. Plasma levels of sTM in patients with cardioembolic subtype were significantly higher than in patients with other stroke subtypes, with the mean ± SD levels of 3.79 ± 1.26, 2.38 ± 0.68 (P < .009), and 2.38 ± 0.44 (P < .05) ng/mL for cardioembolism, large artery atherosclerosis, and small artery occlusion, respectively. Plasma sTM levels were not associated with stroke severity and risk factors of stroke; however, there was a slight relationship between high sTM levels and the presence of atrial fibrillation in the patient group. In conclusion, plasma sTM levels were increased in Thai patients with cardioembolic stroke and may be a potential marker during the acute phase.

Hypoperfusion in Severely Injured Trauma Patients is Associated With Reduced Coagulation Factor Activity

Recent studies have shown that acute traumatic coagulopathy is associated with hypoperfusion, increased plasma levels of soluble thrombomodulin, and decreased levels of protein C but with no change in factor VII activity. These findings led to the hypothesis that acute traumatic coagulopathy is primarily due to systemic anticoagulation, by activated protein C, rather than decreases in serine protease activity. This study was designed to examine the effect of hypoperfusion secondary to traumatic injury on the activity of coagulation factors. Post hoc analysis of prospectively collected data on severely injured adult trauma patients presenting to a single trauma center within 120 minutes of injury. Venous blood was analyzed for activity of factors II, V, VII, VIII, IX, X, and XI. Base deficit from arterial blood samples was used as a marker of hypoperfusion. Seventy-one patients were identified. The activity of factors II, V, VII, IX, X, and XI correlated negatively with base deficit, and after stratification into three groups, based on the severity of hypoperfusion, a statistically significant dose-related reduction in the activity of factors II, VII, IX, X, and XI was observed. Hypoperfusion is also associated with marked reductions in factor V activity levels, but these appear to be relatively independent of the degree of hypoperfusion. The activity of factor VIII did not correlate with base deficit. Hypoperfusion in trauma patients is associated with a moderate, dose-dependent reduction in the activity of coagulation factors II, VII, IX, X, and XI, and a more marked reduction in factor V activity, which is relatively independent of the severity of shock. These findings suggest that the mechanisms underlying decreased factor V activity--which could be due to activated protein C mediated cleavage, thus providing a possible link between the proposed thrombomodulin/thrombin-APC pathway and the serine proteases of the coagulation cascade--and the reductions in factors II, VII, IX, X, and XI may differ. Preservation of coagulation factor activity in the majority of normally and moderately hypoperfused patients suggests that aggressive administration of plasma is probably only indicated in severely hypoperfused patients. Markers of hypoperfusion, such as base deficit, might be better and more readily available predictors of who require coagulation support than international normalized ratio or activated partial thromboplastin time.

Adhesion molecules and high‐sensitivity C‐reactive protein levels in patients with sickle cell beta‐thalassaemia

The primary symptoms of sickle cell disease (SCD) arise from vaso-occlusive crises. The pathogenesis of these crises is complex phenomenon where endothelial activation and damage has a major role. Chronic inflammation also plays an important role in the pathophysiology of SCD. We aimed to investigate endothelial activation in Caucasian Greek patients with SCD by means of measuring adhesion molecules and markers of inflammation. Twenty-eight patients with SCD aged 5-63 years were included in the study. Most of the patients (23/28) were double heterozygotes for sickle cell/beta-thalassaemia, while five patients (5/28) were sickle cell homozygotes. Patients were treated with one/or more of hydroxyurea, therapeutic phlebotomies, blood transfusion or splenectomy. Twenty apparently healthy individuals matched for age and sex formed the control group. Measurements of soluble intercellular adhesion molecule-1, (sICAM-1), soluble vascular cell adhesion molecule-1 (sVCAM-1), P-selectin, E-selectin, soluble thrombomodulin (sTM) and high-sensitivity C-reactive protein (hs-CRP) levels were performed using immunoassays in both patients and healthy individuals. We found that all endothelial adhesion molecules and hs-CRP were significantly increased (P < 0·001) in patients with SCD compared with controls, while sTM levels did not differ significantly (P > 0·05) and this increase was not influenced by the treatment. Our findings demonstrate the high degree of endothelial activation and damage seen in sickle cell patients even in steady-state condition, as well as the important chronic inflammation underlying the pathophysiology of this widespread disease.

SRAGE in diabetic and non-diabetic critically ill patients: effects of intensive insulin therapy

IntroductionHyperglycemia represents an independent prognostic factor in critically ill non-diabetic patients but not in those with diabetes. In this context, there is an ongoing debate on the benefit of an intensive insulin therapy, particularly in diabetic patients. We tested the hypothesis that expression of the receptor for advanced glycation end-products (RAGE), an important signal transduction receptor that elicits long-lasting nuclear factor kappa B (NF-κB) activation, may underlie this difference. RAGE expression is regulated by multiple ligands, including high mobility group box-1 (HMGB-1), and is reflected by its released soluble form (sRAGE).MethodsA predesigned analysis was conducted of prospectively collected samples from 76 hyperglycemic critically ill patients (33 type-2 diabetes, 43 non-diabetes) aged ≥18 years with blood glucose of > 6.1 mmol/L enrolled in a randomized controlled trial comparing intensive insulin therapy with conventional insulin therapy. sRAGE and its ligand HMGB-1 together with IL-6, and soluble thrombomodulin (as markers of inflammation and endothelial cell injury, respectively) were evaluated in ICU, at Days 1, 3, 5 and 7. Plasma samples from 18 healthy subjects were used as controls.ResultsBoth diabetic and non-diabetic hyperglycemic patients showed increased plasma sRAGE, HMGB-1 and soluble thrombomodulin levels at the time of admission to ICU. Plasma IL-6 concentration was only increased in non-diabetic patients. Plasma levels of sRAGE were higher in diabetic compared with non-diabetic patients. Intensive insulin therapy resulted in a significant decrease of sRAGE and thrombomodulin at Day 7, in diabetic but not in non-diabetic patients. Circulating sRAGE levels correlated positively with IL-6 and soluble thrombomodulin levels and inversely with HMGB-1. Multivariate regression analysis demonstrated that sRAGE remains independently correlated with HMGB-1 only in diabetic patients. Neither sRAGE nor any inflammatory markers are associated with mortality.ConclusionsThese findings support the hypothesis that sRAGE release, time-course and response to intensive insulin therapy differ between hyperglycemic diabetic and non-diabetic critically ill patients. Whether this difference underlies the dissimilarity in clinical outcome of hyperglycemia in these two conditions warrants further studies.

Soluble Thrombomodulin Levels are Related to Inflammation after Coronary Bypass Surgery

Background: Soluble Thrombomodulin (sTM) is released from endothelial cells after injury by activated leukocytes in inflammatory states and it has been associated with worse prognosis in inflammatory conditions. The purpose of this study was to analyze the association between levels of sTM and inflammation and to describe the possible explanations about association between sTM and post-operative complications. Methods and Materials: We measured levels of sTM, IL6 and leukocytes in different moments during the first month in 18 patients after coronary by-pass graft surgery. Chisquare test and Fisher test were used for categorical variables. For continuous variables we used either Student t test or Wilcoxon rank-sum test depending on whether the data were normally distributed. Results: The levels of sTM increased during the first post-surgery week, and then decreased to values similar to those recorded pre-operatively. IL6 peaked at after 24 hours, and significantly correlated with white blood cell count (Spearman 0.69, p=<0.0001). There was a significant association between sTM at T3 and white blood cells count at T1 (p=0.01224). Two patients had three or more post-operative complications, and they presented higher levels of sTM. Conclusions: We found a transient increase in sTM during the first week after CABG associated with an inflammatory response and leukocytosis. These changes could imply endothelial dysfunction after surgery and may represent prognostic factors for outcomes.

Efficacy of olmesartan therapy on fibrinolytic capacity in patients with hypertension

The efficacy of olmesartan on fibrinolytic capacity has not been studied yet. Therefore, the aim of the present study was to investigate the efficacy of olmesartan on hemostatic/fibrinolytic status by measuring plasma level of plasminogen activator inhibitor-1 (PAI-1) and soluble thrombomodulin levels in patients with hypertension. Forty-two consecutive, newly diagnosed (25 women and 17 men with a mean age of 48 ± 8 years) patients with untreated essential hypertension were included in the study. Olmesartan medoxomil (20 mg/day) was started and the patients were followed up for 6 months. Baseline biochemical variables, thrombomodulin, and PAI-1 levels were compared with the levels of these variables measured at the end of the 6-month follow-up period. After 6 months of treatment with olmesartan medoxomil, there was a significant reduction in systolic and diastolic blood pressure (from 159.5 ± 10.9 to 134.6 ± 12.7 mmHg and from 98.0 ± 6.3 to 83.9 ± 7.0 mmHg, respectively). Mean plasma PAI-1 and thrombomodulin levels were also significantly decreased (59.73 ± 41.91 vs. 48.60 ± 33.65 ng/ml, P = 0.001 and 8.09 ± 2.29 vs. 6.92 ± 1.42 μg/l, P < 0.001, respectively). Olmesartan medoxomil decreased plasma PAI-1 and thrombomodulin levels after 6 months of therapy, indicating a favorable effect on fibrinolytic capacity in patients with essential hypertension.

Calibrated automated thrombography demonstrates hypercoagulability in patients with idiopathic pulmonary arterial hypertension

Introduction Pathogenesis of idiopathic pulmonary arterial hypertension (iPAH) includes endothelial dysfunction and in situ thrombosis. A hypercoagulable state has also been postulated but never demonstrated. Our objective was to determine whether patients with iPAH had a hypercoagulable state using calibrated automated thrombography (CAT), a new tool to phenotype coagulation in vitro. Patients and methods 16 patients with iPAH and 29 controls were studied. In vitro platelet dependent coagulation phenotyping by CAT monitored the activity of thrombin generation over time. Plasma levels of soluble thrombomodulin, tissue factor pathway inhibitor (TFPI) and von Willebrand factor (VWF) were measured as endothelial biomarkers. Results Endogenous thrombin potential (ETP) in the absence of activated protein C (APC) tended to be increased in patients compared to controls (1769 versus 1656 nM.min; p = 0.053). ETP was higher in the presence of APC 25 nM (ETP-APC) in patients (781 versus 494 nM.min; p = 0.005). Five patients had ETP-APC higher than the 95th centile of controls. Other CAT parameters (lag time, peak thrombin and time to peak) were all consistent with some degree of hypercoagulability in patients. Regarding endothelial plasma biomarkers sTM was lower (28.4 versus 40.6 μg/l, p = 0.0108) in patients; TFPI antigen and activity (respectively: 14.3 versus 10.5 μg/l, p = 0.0167; 1.155 versus 1.070, p = 0.0021) and VWF (1300 versus 976%, p = 0.0108) were higher in patients. Conclusion We have demonstrated that at least some patients with iPAH have a hypercoagulable phenotype.

Impaired anti-platelet effect of aspirin, inflammation and platelet turnover in cardiac surgery☆☆☆

A reduced platelet inhibitory response to acetyl salicylic acid (ASA) has been associated with an increased risk of graft thrombotic occlusion after coronary artery bypass grafting (CABG). We performed a prospective, observational study of 18 patients on 100 mg/day ASA before and after CABG. We assessed antiplatelet response to ASA and its relationship with platelet turnover, inflammatory markers, and soluble thrombomodulin (sTM) levels. All patients showed optimal response to ASA preoperatively but had higher values during follow-up. Platelet aggregation and platelet count in the perioperative period were significantly associated (P=0.05). Platelet turnover was defined as the average daily turnover (ADTO). The lowest inhibitory value (28% of patients > or =6 Omega) was recorded at the same time of the highest platelet turnover (>10% daily in 77.77% of patients), one week after CABG. ADTO >10% was associated with an increased risk of platelet aggregation > or =6 Omega. Levels of sTM were significantly higher one week after CABG (median 13 vs. 3 ng/ml preoperatively, P=0.0011). There is a transient impairment in ASA antiplatelet effect after CABG related to an increased platelet turnover caused by the inflammatory process. This could be responsible for the high risk of occlusive thrombosis.

High Levels of Protein C Are Determinated by PROCR H3 in a Dutch Family.

Abstract 2139Poster Board II-116The natural anticoagulant protein C (PC) circulates in blood at a concentration of about 60 nM. Inter-individual variations in the levels of PC are in part genetically determined, but which loci in the genome are involved is only partially known. In a recent study we identified a locus on chromosome 20 which was associated with high PC levels in a large pedigree from the GENES study (LOD score >5 at 55 cMorgan). Candidate genes related to the PC pathway under the LOD-1 region encoded FOXA2 (previously known as HNF3 beta, a nuclear factor regulating protein C gene transcription), thrombomodulin (THBD,which is key to activation of PC), and the endothelial protein C receptor (PROCR). Here we present data that pinpoint a SNP in PROCRas being responsible for the observed segregation of high PC levels. The pedigree has 218 members and was ascertained through a proband with a family history of venous thrombosis (VT). Classical genetic risk factors for thrombosis (i.e. PC-, PS-, antithrombin deficiency, factor V Leiden and prothrombin G20201A) were not present. Complete medical data, plasma measurements and DNA was available for 161 family members. The mean age was 47±15 (range 15-87) years. The mean PC plasma level was 116±25% (range 72-212). Four family members had experienced VT and 2 had had recurrence. These symptomatic members had normal to high PC levels (66, 82, 114 and 178%).Haplotypes (and genotypes) for PROCR were determined in the family members by TaqMan assay using tag SNPs (single nucleotide polymorphisms) and PROCR H3 was associated with the levels of PC in the family. Furthermore, the promoter, exons, and 3`UTR of the 3 candidate genes were sequenced in 13 individuals, 9 with high and 4 with normal plasma PC levels. Critical SNPs that were encountered during sequencing were genotyped in all family members, namely FOXA2 rs1055080 (3`UTR) and rs2277764 (promoter region). As those 2 SNPs were inherited together in the set of 13 patients and also in the LETS (data not shown), our further analysis used only rs1055080. Plasma soluble endothelial protein C receptor (sEPCR) and soluble thrombomodulin (sTM) levels were measured with an ELISA assay. PC and sEPCR and levels were compared between PROCR H3 and FOXA2 rs1055080 carriers and non carriers by Student's t-test. sTM was analyzed by Mann-Whitney test. Association between PC levels and sEPCR/sTM levels were evaluated using linear regression analysis. Afterwards associations were adjusted for the PROCR H3 and FOXA2 rs1055080 SNP separately to detect their possible confounding effect. DNA sequencing only yielded previously reported SNPs in FOXA2, THBD and PROCR. Only the above mentioned SNPs were associated with PC plasma levels. Linkage analysis for PC levels using the original markers (from Marshifield) and adding the new PROCR and FOXA2 SNPs did not change the LOD score. When the analysis was adjusted for the mentioned markers, the LOD score dropped below 2. sEPCR has a bimodal distribution; mean ± SD was 103±27 ng/ml for the first mode and 262±70 ng/ml for the second mode. Median (range) sTM was 1.2 ng/ml (0.1-4). Linkage analysis for sEPCR levels yielded a high LOD score (above 6) that was accentuated to above 8 when PROCR H3 was included as a marker. For sTM, the LOD score was low with every combination of markers. PC, sEPCR and sTM levels were compared between PROCR H3 carriers and non-carriers and both PC levels and sEPCR levels were influenced by this PROCR haplotype, but not sTM. In conclusion, chromosome 20 harbors a locus which influences PC levels and also the levels of sEPCR, but not the levels of sTM. A detailed analysis with SNPs in PROCR, THBD and FOXA2suggests that the so-called PROCR H3 is directly responsible for the increased PC and sEPCR levels in this family. PROCR H3 is known to represent a g.A6936G substitution leading to a p.Ser219Gly replacement in the transmembrane domain of EPCR. The Gly219 isoform is more sensitive to sheddases (such as the ADAM17 metalloprotease) and is associated with generation of truncated mRNA lacking the transmembrane domain. However, the exact mechanism by which EPCR and sEPCR levels influence the level of PC remains to be determined Disclosures:No relevant conflicts of interest to declare.