sTM Concentration Research Articles

Value of Serum Thrombomodulin as a Marker and Predictor in Patients with Sepsis-Associated Acute Kidney Injury.

To investigate the serum soluble thrombomodulin (sTM) concentration in patients with sepsis-associated acute kidney injury (AKI) and to determine the value of sTM in predicting AKI and mortality in sepsis patients. This prospective observational study was conducted on 71 patients diagnosed with sepsis according to Sepsis 3 at the Intensive Care Unit, Hue Central Hospital, Vietnam, from September 2021 to February 2023. Among 71 sepsis patients, there were 38 (53.5%) AKI cases, including 16 (22.5%) cases of stage 1 AKI, 14 (19.7%) cases of stage 2 AKI, 8 (11.3%) cases of stage 3 AKI, 16 (22.5%) cases of renal replacement therapy, 28 (39.4%) cases of septic shock, and 21 (29.6%) cases of mortality within 28 days. The concentrations of lactate and IL-6 in the AKI and mortality groups were statistically significantly greater than those in the non-AKI and survival groups (p < 0.05). The serum sTM concentration was 4.33 ng/mL, the serum sTM level in the AKI group was statistically significantly higher than that in the non-AKI group (sTM [4.71 vs 2.54 ng/mL, p < 0.001]), and the serum sTM level in the mortality group was statistically significantly higher than the survival group (sTM [4.78 vs 3.87 ng/mL, p < 0.001]). The AUC of sTM for predicting AKI was 0.864; the AUCs of sTM, IL-6, SOFA, and APACHE II for predicting mortality were 0.811, 0.671, 0.816, and 0.705, respectively. AKI was a prevalent complication among sepsis patients at the ICU. In the AKI and mortality groups, sTM concentration was statistically significantly higher than that in the non-AKI and survival groups. sTM was the predictor of acute kidney injury and mortality in patients with sepsis.

Endothelial dysfunction markers and immune response indices in cosmonauts’ blood after long-duration space flights

Space flight factors are known to cause a malfunction in the human immune system and lead to damage to blood vessels. The hemostatic function of endothelium during space missions and its interaction with human immunity has not been determined so far. In this work, we investigated the markers of endothelial activation and damage (plasma concentrations of soluble thrombomodulin fraction (sTM), von Willebrand factor (vWF), highly sensitive C-reactive protein (hs-CRP)), as well as the level of D-dimer and compared them to the immunological parameters characterizing the state of human humoral and cellular immunity. The immune status of long-duration ISS crewmembers was assessed by whole-blood testing, and comprehensive postflight immune assessment included the analysis of leukocyte distribution. Flow cytometry was applied to determine the absolute counts and the percentage of lymphocyte subsets: B cells (CD19+), T cells (CD3+, CD3+CD4+, CD3+CD8+), NK cells (CD3−CD16+CD56+, CD11b+CD56+), and activated subsets (CD3+CD25+ and CD3+HLA-DR+). The in vitro basal cytokine production was investigated in whole blood cell culture. The cytokines IFN-gamma, IL-1-beta, IL-4, IL-6, IL-10, IL-18, and TNF-alpha were measured in plasma and the 24-h supernatants by a sensitive enzyme-linked immunosorbent assay. A significant increase in the plasma levels of vWF and hs-CRP and a decrease in the concentration of sTM after spaceflights were detected. Divergent changes in the parameters characterizing the state of the immune system were observed. We propose that the changes revealed may lead to an increase in the procoagulant activity of blood plasma, suppression of protein C activation and thrombin inhibition, as well as to an increase in the adhesive-aggregate potential of platelets, especially in case of changes in the rheological characteristics of blood flow during re-adaptation to ground conditions. We also speculate that the immune system might play an important role in vessel damage during long-duration missions.

Endothelial Markers: Thrombomodulin and Von Willebrand Factor and Risk of Kidney Thrombosis After Transplantation

Endothelial disturbance is well known as one of the causes of thrombosis. This study measured von Willebrand factor (vWF) and soluble thrombomodulin (sTM) in renal vein blood to evaluate for the risk of thrombosis after kidney transplantation. A cross-sectional study that included 61 consecutive recipients of kidney transplant. The sTM and activity of vWF were evaluated in blood of the renal vein at the time of reperfusion. The renal vein blood had higher values of vWF activity and sTM concentration than the peripheral blood. In the first minutes of reperfusion, the concentration of thrombomodulin was the highest but activity of vWF was the lowest. As the reperfusion continued, thrombomodulin gradually decreased, but vWF increased. The strong correlations of TMs and vWF with warm ischemia were observed (r=0.5577 and r=0.3429, respectively). Thrombosis was found in about 10% of all recipients. However, other complications, such as delayed graft function or ureter necrosis, were associated with high values of vWF and sTM. They were correlated with increased sTM concentration and activity of vWF (P < .006 and P < .05, respectively). This was confirmed by analysis of the receiver operator characteristic curve. The area under the curve values for TMs and vWF were 0.762 and 0.602, respectively (P < .0001 and P < .015, respectively). The cutoff points for sTM and vWF were 14.89 ng/mL and 129.89%, respectively. Positive prediction value sTM and vWF were 76% and 66% and negative prediction value 69% and 59%, respectively. The endothelium of a transplanted kidney could be involved in the pathogenesis of renal thrombosis. Endothelial protection during harvesting can greatly contribute to the improvement of transplantation outcomes. The renal pool of sTM and vWF could be a useful marker of the risk of renal thrombosis.

Bacillus subtilis PB6 Supplementation in Weaned Holstein Steers During an Experimental Salmonella Challenge.

To evaluate the effects of a patented Bacillus subtilis probiotic, weaned Holstein steers, not shedding Salmonella (n = 40; ∼90 kg), were supplemented (CLO) or not (CON) with CLOSTAT® (13 g/hd per day; Kemin Industries, Des Moines, IA) in a starter ration for 35 d. The calves were assigned to one of four treatments in a 2 × 2 factorial design with CLO and CON calves that were orally administered Salmonella (STM) or not (NoSTM). Calves were challenged with 1.6 × 106 colony-forming unit (CFU) Salmonella Typhimurium (resistant to 50 μg/mL nalidixic acid) in 1 L of milk replacer on day 0. Blood samples were collected through jugular catheters every 6 h for 96 h, and body temperature was measured every 5 min through indwelling rectal temperature recording devices. Five calves from each treatment were harvested 48 h postchallenge, and the remaining calves were harvested 96 h postchallenge. During necropsy, tissues were collected for the isolation and quantification of the inoculated STM from various tissues. The CLOSTM group had reduced STM concentrations in the jejunum, ileum, and transverse colon 48 h after the challenge (p ≤ 0.03), but were not different 96 h postchallenge (p > 0.05). Decreased (p < 0.01) pyrexia was observed after the challenge in CLOSTM calves when compared with CONSTM calves. White blood cells and lymphocyte counts were increased (p ≤ 0.05) in CLOSTM calves after the challenge in comparison with other treatments. In calves given STM, the CLO group had greater feed intake before and after the challenge (p < 0.01) compared with the CON group. Increased serum IL-6 and IFN-γ concentrations were observed in the CONSTM group compared with other treatments. Overall, CLO reduced Salmonella presence and concentrations in gastrointestinal tissues while simultaneously reducing the severity of the challenge as indicated by blood parameters and the reduced febrile response.

Biomarkers in patients with Takotsubo cardiomyopathy compared to patients with acute anterior ST-elevation myocardial infarction

Background: Takotsubo cardiomyopathy (TTC) is a syndrome of acute non-coronary heart failure with similar symptoms and electrocardiograms to acute anterior ST-elevation myocardial infarction (STEMI). Little is known about the pathophysiology of TTC. We assessed admission plasma concentrations of biomarkers reflecting neuroendocrine response (copeptin, mid-regional-pro-adrenomedullin, pro-atrial-natriuretic-peptide, soluble thrombomodulin (sTM), syndecan-1) and inflammation (suppression-of-tumorigenicity 2 (ST2), high-sensitive C-reactive-protein) in TTC patients and compared to patients with acute anterior STEMI.Materials and methods: Twenty TTC patients were matched with 40 STEMI patients by age, gender and left ventricular ejection fraction. Blood was sampled upon hospital admission immediately before acute coronary angiography.Results: The groups had similar comorbidities. TTC patients had higher plasma concentrations of sTM: 7.94 (5.89;9.61) vs. 6.42 (5.50;7.82)ng/ml, p = 0.04 and ST2 (53 (32;157) vs. 45 (31;55)ng/ml, p = 0.008) and higher heart rate: 101 (33) vs. 76(14)bpm, p = 0.0001, but lower concentrations of copeptin (10.4 (7.6;39) vs. 92.3 (13;197)pmol/l, p < 0.05) and troponin T (348 (98;759) vs. 1190 (261;4105)ng/l, p = 0.04).Conclusion: TTC patients had higher plasma concentrations of sTM and ST2, higher heart rate and lower copeptin and troponin T concentrations compared to acute anterior STEMI patients. This study contributes to the hypothesis that TTC patients have endothelial cell damage and are hemodynamically more stable than patients with acute anterior STEMI on admission.

Changes in Biomarkers of Coagulation, Fibrinolytic, and Endothelial Functions for Evaluating the Predisposition to Venous Thromboembolism in Patients With Hereditary Thrombophilia.

The changes in the coagulation, fibrinolytic, and endothelial functions are correlated with the pathophysiology of the thromboembolic diseases during acute illness. However, these changes in patients with hereditary thrombophilia who were not in the acute stage of venous thromboembolism (VTE) are unclear. A panel of 4 biomarkers, including thrombin–antithrombin complex (TAT), plasmin-α2-plasmin inhibitor complex (PIC), tissue-type plasminogen activator/plasminogen activator inhibitor-1 complex (t-PAIC), and soluble thrombomodulin (sTM), were assayed in 100 healthy controls and 100 patients with thrombophilia. Although significantly higher concentrations of TAT, PIC, t-PAIC, and sTM were observed in patients with thrombophilia than in healthy controls, 70 patients showed absolutely normal levels of the above 4 biomarkers. Among the other 30 patients who had at least 1 biomarker out of the corresponding reference interval, 26 of them presented elevated PIC with or without increased TAT. Except for sTM, other 3 biomarkers did not show significant differences in patients with previous VTE compared to those without. Patients with single episode of VTE had obviously lower t-PAIC than those with multiple episodes of VTE, whereas the levels of TAT, PIC, and sTM were unassociated with the number of thrombosis episodes. Most thrombophilia patients who were not in the acute stage of VTE showed normal coagulation, fibrinolytic, and endothelial functions. Thus, we were unable to show that the one-time response of this panel was clinically helpful in determining thrombosis risk in thrombophilia individuals. Future studies should focus on the dynamic monitoring during the chronic phase of VTE to offer special advantages for patients with thrombophilia.

The endothelial status reflected by circulating endothelial cells, circulating endothelial progenitor cells and soluble thrombomodulin in patients with mild and resistant hypertension

BackgroundThe aim of the present study was to evaluate endothelial status by measuring the concentration of novel markers of endothelial dysfunction (ED): a number of circulating endothelial cells (CECs), circulating endothelial progenitor cells (CEPCs) and their ratio (CEPCs/CECs) as well as a traditional parameter - soluble thrombomodulin (sTM) in patients with resistant (RH) and mild hypertension (MH). Materials and methodsThirty patients with MH and thirty subjects with RH were involved in the study. The control group included thirty-three age and sex-matched normotensive volunteers. We used multicolor flow cytometry for CECs and CEPCs analysis and the commercial human sTM ELISA kit to measure plasma sTM concentration. ResultsAn elevated CECs number and a decreased CEPCs/CECs ratio was found in MH as well as in RH patients in comparison with normotensive volunteers. CECs correlated positively with an increased triglycerides in MH patients and an elevated LDL-cholesterol and hsCRP in RH group. Positive correlation between CEPCs and LDL-cholesterol level was observed in both types of hypertension. ConclusionsThe results of the present study suggest that an endothelial alteration accompanies hypertension. The number of CECs reflecting the extent of endothelial damage does not appear to be related to the severity of disease. The drastically decreased ratio between CEPCs and CECs observed in both groups of patients suggests an inadequate process of endothelial regeneration. Among analyzed factors inflammation and lipid abnormalities may have significant contribution in endothelial pathology in hypertension.

Coexistence of endothelial dysfunction, fibrinolysis activation and diminished interleukin 6 level in morbid obesity

ABSTRAKTWstęp: Celem pracy była ocena wybranych parametrów dysfunkcji śródbłonka i hemostazy u chorych z otyłością. Badania dotyczące zaburzeń komórek śródbłonka w otyłości ogromnej są w literaturze spotykane zaskakująco rzadko.Materiały i metody: Badanie przeprowadzono u 74 chorych z otyłością ze wskaźnikiem masy ciała (BMI) &gt;40 kg/m2 oraz u 30 zdrowych ochotników z prawidłowym BMI. W osoczu krwi chorych przeprowadzono testy stężenia: rozpuszczalnej formy trombomoduliny (sTM), antygenu czynnika von Willebranda (vWF:Ag), antygenu tkankowego aktywatora plazminogenu (t-PA:Ag) i antygenu inhibitora aktywatora plazminogenu typu 1 (PAI-1:Ag) oraz rozpuszczalnej frakcji selektyny E i P (sE-selektyny i sP-selektyny). Dokonano także pomiaru stężenia interleukiny 6 (Il-6).Wyniki: Wyniki badań wskazały znaczne zwiększenie stężenia sTM, t-PA:Ag, PAI-1:Ag i sE-selektyny oraz istotne zmniejszone stężenie vWF:Ag i sP-selektyny w otyłości ogromnej w porównaniu z grupą kontrolną. Wykazano także w otyłości ogromnej obniżenie stężenia Il-6.Wnioski: W otyłości ogromnej dochodzi do uszkodzenia śródbłonka, co zwiększa ryzyko ewentualnych powikłań zakrzepowo-zatorowych. Zwiększenie składowych układu fibrynolizy oraz hamowanie aktywacji płytek można traktować jako mechanizmy kompensacyjne zmniejszające ryzyko powikłań. Spadek stężenia Il-6 może być wyrazem ograniczania procesu zapalnego, ale równocześnie może to być przyczyna zwiększonej podatności na choroby infekcyjne.

Plasma concentration of selected biochemical markers of endothelial dysfunction in women with various severity of chronic venous insufficiency (CVI)-A pilot study.

BackgroundAlthough the endothelial dysfunction is considered to be implicated in the pathogenesis of chronic venous insufficiency (CVI) the endothelial status in patients with venous disorders is still not fully evaluated. Therefore the aim of the study was to measure the concentration of selected markers of endothelial dysfunction: von Willebrand factor (vWf), soluble P-selectin (sP-selectin), soluble thrombomodulin (sTM) and soluble VE-cadherin (sVE-cadherin) in CVI women who constitute the most numerous group of patients suffering from venous disease.Materials and methodsForty four women with CVI were involved in the study and divided into subgroups based on CEAP classification. Concentration of vWf, sP-selectin, sTM and sVE-cadherin were measured and compared with those obtained in 25 healthy age and sex-matched women.ResultsIt was found that the concentration of sTM increased and sVEcadherin decreased along with disease severity in CVI women. A significant rise of sTM was observed especially in CVI women, with the highest inflammation status reflected by hsCRP or elastase concentration, and in CVI women with a high oxidative stress manifested by an increased plasma MDA. A significant fall of circulating sVE-cadherin was reported in CVI women with moderate to highest intensity of inflammation and oxidative stress. There was no change in vWF and sP-selectin concentration at any stage of CVI severity.ConclusionsThe results of the present study demonstrate the presence of endothelial dysfunction in women suffering from CVI which seems to progress with the disease severity and may be associated with inflammation and enhanced oxidative stress.

Clinical usefulness of videocapillaroscopy and selected endothelial cell activation markers in people with Type 1 diabetes mellitus complicated by microangiopathy

PurposeMicrovascular abnormalities are one of the most important causes of persistent diabetic complications. The aim of our study was to compare microvascular changes examined by nailfold videocapillaroscopy (NVC) examination with serum concentrations of vascular endothelial growth factor (VEGF), soluble thrombomodulin (sTM) and endothelin-1 (ET-1) in people with Type 1 diabetes with and without microangiopathy. Material/methodsThe study included 106 people with Type 1 diabetes and 40 healthy controls. All participants were evaluated by extensive clinical, laboratory and capillaroscopic studies. NVC was performed using a stereomicroscope SZ 4045 (Olympus, Germany). The intensity of morphological changes was graded from 0 to 3. Serum levels of VEGF, sTM and ET-1 were determined by an enzyme-linked immunosorbent assay (ELISA). ResultsMorphological changes were observed by NVC in 86 out of 106 (81%) people with Type 1 diabetes mellitus. Severe capillaroscopic changes (score 3) were seen in 32 out of 54 (59%) people with microangiopathy, but in only seven out of 52 (13%) individuals without microangiopathy. Higher serum concentration of VEGF (p<0.001), ET-1 (p<0.001) and sTM (p<0.05) were demonstrated in people with diabetes complicated with microangiopathy compared to healthy controls. Moreover, comparison between people with and without microangiopathic complications showed a significantly higher capillaroscopic score and sTM serum concentration in the group with retinopathy (p<0.001) nephropathy (p<0.001) and neuropathy (p<0.01). ConclusionsOur results suggest that abnormalities in NVC may reflect the extent of microvascular involvement and associated with higher VEGF, sTM and ET-1 serum levels, as well as with microangiopathic complications in diabetic people.

Pharmacokinetics of Standard- and Reduced-Dose Recombinant Human Soluble Thrombomodulin in Patients with Septic Disseminated Intravascular Coagulation during Continuous Hemodiafiltration.

Recombinant human soluble thrombomodulin (rTM) is reportedly excreted by the kidneys; therefore, the recommended dose for patients with renal impairment is one-third of the standard dose. The aim of this study was to evaluate whether this reduced dose of rTM achieves effective drug concentrations that are comparable to those of the standard dose in treating sepsis-induced disseminated intravascular coagulation (DIC) during continuous hemodiafiltration (CHDF). Eight patients in an intensive care unit were randomized to receive either reduced-dose (0.02 mg/kg, n = 4) or standard-dose (0.06 mg/kg, n = 4) rTM. We evaluated the effect of standard dose in comparison to that of reduced dose on the pharmacokinetics (PKs) of rTM for the sepsis-induced DIC patients receiving CHDF. Patients received rTM during a 30-min infusion for six consecutive days. PK parameters of rTM were analyzed using the one-compartment model. The elimination half-life, clearance (T1/2), and distribution volume of sTM were similar between the reduced and standard doses. The maximum concentration (Cmax) and area under the concentration-time curve (AUC) of sTM were approximately 2.5 times higher with standard-dose daily infusions than that with reduced-dose drip infusions (p = 0.041 and 0.062, respectively). The time when the blood concentration of sTM was >500 ng/mL, i.e., the holding time, was significantly longer with standard-dose infusions than those with reduced dose (p = 0.039). rTM displayed dose-dependent PK behavior at clinically relevant doses. During CHDF, effective blood concentration of rTM was not achieved with the reduced dose, and rTM was found to not bioaccumulate. Therefore, this pilot study suggests that reducing the rTM dose is unnecessary, even in sepsis-induced DIC patients who require CHDF. However, we need to perform a definitive study to determine the dosage of rTM for the case.

The Association of Serum Thrombomodulin with Endothelial Injuring Factors in Abdominal Aortic Aneurysm.

Background. The aim of the present study was to evaluate the concentration of serum thrombomodulin (sTM) in the AAA patients and to examine its correlation with various factors which may potentially participate in the endothelial injury. Materials and Methods. Forty-one patients with AAA were involved and divided into subgroups based on different criteria. Concentration of sTM was measured using enzyme-linked-immunosorbent assay (ELISA). The results were compared with those obtained in 30 healthy age- and sex-matched volunteers. Results. The higher concentration of sTM was observed in AAA patients compared with those in controls volunteers [2.37 (1.97–2.82) ng/mL versus 3.93 (2.43–9.20) ng/mL, P < 0.001]. An elevated sTM associated significantly with increased triglycerides (TAG) [P = 0.022], cholesterol [P = 0.029], hsCRP [P = 0.031], and advanced glycation end products (AGEs) [P = 0.033]. Conclusions. The elevation of serum sTM level suggests that endothelial damage occurs in AAA pathogenesis. The correlations observed indicate that lipids abnormalities, inflammation, and oxidative stress may be involved in this destructive process.

Emission patterns of acrylonitrile and styrene around an industrial wastewater treatment plant in Iran

The uncontrolled releases of volatile organic compounds (VOCs) from wastewater treatment plants (WWTPs) have been highly concerned due to the associated public health risks. In petrochemical industries, WWTPs are responsible for various organic compound emissions into the atmosphere, which can considered as the main source of VOCs emission in such industries. The typical high-strength petrochemical wastewater is generated from an acrylonitrile–butadiene–styrene (ABS) resin manufacturing plant that usually needs pretreatment before discharging to the main WWTP. The objective of this study was to investigate the emissions and fates of acrylonitrile (ACN) and styrene (STM) through wastewater pretreatment units operated in an ABS manufacturing plant. In this study, the emission rates of ACN and STM were estimated by means of EPA’s Water9 emission model. Subsequently, the emission rates were used as the input data of AERMOD model to simulate the atmospheric behaviors of emitted ACN and STM. The results of Water9 model showed that 57 and 81 % of influent ACN and STM are emitted to the air through pretreatment units, respectively. For both of them, the equalization basin had the major portion of emission to the atmosphere. The concentration distribution profiles of ACN and STM resulted from AERMOD model indicted that the concentration of STM was lower than EPA reference concentration (RfC); however, the higher concentration of ACN (higher than RfC) occurred near the WWTP as well as the neighbor ambient.

Plasma homocysteine involved in methylation and expression of thrombomodulin in cerebral infarction

Homocysteine (Hcy) regulates endothelial injury and methylation status of key genes in cerebral ischemia. Thrombomodulin (TM) may be protective against cerebral ischemia by downregulating coagulation. However, it remains unclear whether Hcy involved in methylation and expression of TM in cerebral infarction (CI). Here, we find patients with cerebral infarction had a higher TM methylation level than controls (74.2% vs 47.5%, X2 = 14.724, P = 0.00), which are positively correlated with plasma levels of tHcy (r = 0.701, P = 0.00) and negatively related to mRNA expression of TM (r = −0.711, P = 0.00). Plasma levels of tHcy (t = 7.566, P = 0.00) and sTM (t = 17.268, P = 0.00) are significantly higher in cases than in controls. Our data indicate hyperhomocysteine leads to hypermethylation of the TM gene and further induces TM gene silencing, which may play an important role in the occurrence and development of CI. Plasma higher concentrations of sTM in cases are not caused by TM expression and may be only a result of Hcy induced endothelial injury.

Long-Range Allosteric Linkage Between Exosites Reciprocally Regulates the Zymogenicity of Prothrombin Derivatives

The binding of ligands to anion binding exosite I (ABEI) and exosite II (ABEII) on prothrombin (II) derivatives plays an integral role in regulating their function. These exosites are located on opposite faces of the proteinase domain and exhibit unique binding specificities. Fragment 1.2 (F12) binds to ABEII and acts as a zymogen-promoting allosteric ligand. Conversely, Na+ and active site ligands stabilize the proteinase state. Here, we investigated the allosteric linkage between ABEI and the Na+ binding site, active site or ABEII using multiple ABEI ligands and prothrombin derivatives differentially poised along the zymogen to proteinase continuum. Prethrombin 2 (P2) represents the most zymogen-like state that differs from thrombin (IIa) because it is not cleaved at the R320 site. To mimic the zymogen-like character of P2 in a cleaved IIa molecule, residues responsible for N-terminal insertion and proteinase formation (IVE) were swapped with TAT to produce IIaTAT, a mutant with vastly diminished proteolytic activity. Alanine was substituted with the catalytic serine residue in IIa (IIaS195A) to represent the proteinase without a ligand at its active site. The thermodynamics of interactions between the thrombin inhibitor and ABEI ligand hirugen (Hir) and the various reference states was assessed using isothermal titration calorimetry (ITC). Titration of Hir into P2, IIaTAT or IIaS195A revealed thermodynamically more favorable binding to proteinase-like IIaS195A in comparison to zymogen-like P2 or IIaTAT. Binding of Hir to IIaS195A was affected by the concentration of Na+ at constant ionic strength. Global analysis done in the presence of increasing concentrations of Na+ revealed a 5-fold increase in Hir binding affinity when IIaS195A is ligated with Na+; demonstrating positive allosteric linkage between ABEI and Na+ binding. Using a truncated Staphylocoagulase variant that binds ABEI without N-terminus insertion (SC13-325), we found that SC13-325 binding alone promoted active site opening and fluorescent inhibitor incorporation in zymogen-like P2. These data reinforce the observation that ABEI ligands promote a proteinase-like state in prothrombin derivatives through positive allosteric linkage with the Na+ binding and active sites. Interestingly, inhibitor incorporation and ITC studies both showed that SC13-325 interacts poorly with II, but strongly with P2 despite both species being zymogens. These findings imply that the ABEII ligand F12, which is produced upon cleavage of II at R271 to form P2, may display negative allosteric linkage with ABEI. Titration of SC13-325 into pre-formed complexes of P2/F12 revealed a drastic reduction in affinity of SC13-325 for P2 when F12 is bound to ABEII. Thus, F12 binding at ABEII negatively affects ABEI binding. Further studies used soluble thrombomodulin (sTM) as the most physiologically pertinent ligand for ABEI. ITC and global analysis of the binding of F12 to P2 in the presence of different concentrations of sTM revealed a ~1200-fold decrease in binding affinity and enthalpy for either the binding of F12 to P2 bound to sTM or the binding of sTM to P2 bound to F12. These data illustrate the strong negative linkage associated with the binding of protein ligands to ABEI and ABEII which yields the appearance of competitive and mutually exclusive binding at the two sites despite the fact that they are on opposite sides of the proteinase domain. Allosteric linkage between ligand binding at the two exosites is centered on the ability of F12 binding to ABEII and favor zymogen-like forms and ligand binding to ABEI to favor proteinase-like forms. Thus, allosteric linkage among exosites is vital to the interconversion of prothrombin species along the zymogen to proteinase spectrum. These ligand-dependent conformational shifts and associated changes in function are likely to greatly contribute to the dynamic roles that IIa plays during coagulation. DisclosuresNo relevant conflicts of interest to declare.

Endothelial Injury Associated with Cold or Warm Blood Cardioplegia during Coronary Artery Bypass Graft Surgery.

The aim of this investigation was to analyze the impact of intermittent cold blood cardioplegia (ICC) and intermittent warm blood cardioplegia (IWC) on endothelial injury in patients referred to elective on-pump coronary artery bypass graft (CABG) surgery. Patients undergoing CABG procedures were randomized to either ICC or IWC. Myocardial injury was assessed by CK-MB and cardiac troponin T (cTnT). Endothelial injury was quantified by circulating endothelial cells (CECs), von Willebrand factor (vWF), and soluble thrombomodulin (sTM). Perioperative myocardial injury (PMI) and major adverse cardiac events (MACE) were recorded. Demographic data and preoperative risk profile of included patients (ICC: n = 32, IWC: n = 36) were comparable. No deaths, PMI, or MACE were observed. Levels of CK-MB and cTnT did not show intergroup differences. Concentrations of CECs peaked at 6 h postoperatively with significantly higher values for IWC-patients at 1 h (ICC: 10.1 ± 3.9/mL; IWC: 18.4 ± 4.1/mL; P = 0.012) and 6 h (ICC: 19.3 ± 6.2/mL; IWC: 29.2 ± 6.7/mL; P < 0.001). Concentrations of vWF (ICC: 178.4 ± 73.2 U/dL; IWC: 258.2 ± 89.7 U/dL; P < 0.001) and sTM (ICC: 3.2 ± 2.1 ng/mL; IWC: 5.2 ± 2.4 ng/mL; P = 0.011) were significantly elevated in IWC-group at 1 h postoperatively. This study shows that the use of IWC is associated with a higher extent of endothelial injury compared to ICC without differences in clinical endpoints.

High levels of soluble thrombomodulin may be a marker of arterial disease and peripheral ischemia in Egyptian patients with diabetes mellitus

Background Thrombomodulin is an endothelial cell membrane protein acting as a cofactor for the activation of plasma protein C. It was found that soluble forms of thrombomodulin (sTM) exist in plasma and is considered a marker of endothelial dysfunction. Materials and methods The study included 160 individuals: 40 healthy controls and 120 diabetic patients (40 patients with type 1 diabetes and 80 patients with type 2 diabetes). sTM concentrations were measured by an enzyme-linked immunosorbent assay-based assay employing monoclonal antithrombomodulin antibodies. Results The diabetic patients had highly significantly increased plasma sTM concentrations compared with the control group (P Conclusion We recommend including sTM in the follow-up panel of diabetic patients because of its potential ability to predict the risk for ischemic manifestations.

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.

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.

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.