mocetinostat and Thrombosis

The intention of our study was to establish the prevalence of low birth weight (LBW) as well as risk factors for LBW in infants born to a convenience sample of women enrolled in a home visitation maternal care program associated with the Center for Human Development in Southwest Trifinio, Guatemala.. This is an observational study analyzing self-reported data from a quality improvement database. We recorded the distribution of birthweights of infants born to women enrolled in Madres Sanas that delivered between October 2018 and December 2019. We grouped women by LBW (<2500g ) and adequate birthweight (≥2500g) infants, and performed bivariate comparisons using sociodemographic, obstetric, and intrapartum data. Using the independent variables shown to have an association with LBW, we then performed a multivariable analysis.. Two significant findings emerged from our analysis: LBW infants were more commonly born to women who were younger in age and who had received fewer than 4 prenatal visits. These findings are consistent with existing literature on LBW in Latin America. Our study helps to strengthen the data around these associations and gives credence to programming and policy efforts in Latin America that support adequate prenatal care for all and youth education about reproductive health and contraceptive access.

Topics: Adult; Antioxidants; Chromatography, High Pressure Liquid; Dried Blood Spot Testing; Epigastric Arteries; Ginsenosides; Gold; Horseradish Peroxidase; Humans; Hydrogen Peroxide; Leukemia, Myeloid, Acute; Mammaplasty; Mass Spectrometry; Metal Nanoparticles; Myelodysplastic Syndromes; Panax; Perforator Flap; Peroxidase; Postoperative Complications; Quality Control; Reproducibility of Results; Retrospective Studies; Risk Factors; Saponins; Sarcopenia; Sleep Aids, Pharmaceutical; Spectrometry, Mass, Electrospray Ionization; Tandem Mass Spectrometry; Thromboembolism; Thrombosis

Atherosclerosis is the main pathophysiological process underlying coronary artery disease (CAD). Acute complications of atherosclerosis, such as myocardial infarction, are caused by the rupture of vulnerable atherosclerotic plaques, which are characterized by thin, highly inflamed, and collagen-poor fibrous caps. Several lines of evidence mechanistically link the heme peroxidase myeloperoxidase (MPO), inflammation as well as acute and chronic manifestations of atherosclerosis. MPO and MPO-derived oxidants have been shown to contribute to the formation of foam cells, endothelial dysfunction and apoptosis, the activation of latent matrix metalloproteinases, and the expression of tissue factor that can promote the development of vulnerable plaque. As such, detection, quantification and imaging of MPO mass and activity have become useful in cardiac risk stratification, both for disease assessment and in the identification of patients at risk of plaque rupture. This review summarizes the current knowledge about the role of MPO in CAD with a focus on its possible roles in plaque rupture and recent advances to quantify and image MPO in plasma and atherosclerotic plaques.

Topics: Apoptosis; Biomarkers; Coronary Artery Disease; Endothelium, Vascular; Humans; Lipoproteins, HDL; Lipoproteins, LDL; Luminescent Measurements; Luminol; Molecular Imaging; Peroxidase; Plaque, Atherosclerotic; Thrombosis

Myeloperoxidase (MPO) is a microbicidal and reactive species-generating enzyme. It traditionally is considered to be stored mostly within polymorphonuclear leukocytes and is strongly implicated in the pathogenesis of numerous diseases. MPO also has been studied for at least 20 years as a marker of hemodialysis procedure biocompatibility and oxidative stress generation; research yielded discordant and inconclusive results. In this review, a novel and growing body of evidence indicating that MPO also is a potent blood vessel-bound enzyme that can be mobilized rapidly and extensively into circulating blood by exogenous heparin is discussed. Beneficial consequences of such evoked arterial wall MPO depletion that may be counterbalanced in part by the harmful effects of circulating MPO on polymorphonuclear leukocyte activation and thus atherosclerosis propagation also are presented. Potential clinical implications of these undervalued phenomena in commonly atherosclerotic maintenance hemodialysis patients regularly administered large doses of heparin for temporary blood anticoagulation (frequently over years) are stressed, including the challenging issue of morbidity and mortality. In view of the plausible clinical importance of the novel MPO-oxidative stress-heparin interaction in this population, the need for additional studies assessing different dialyzer membranes, various heparin types (unfractionated heparin versus low-molecular-weight heparins versus pentasaccharides), as well as different anticoagulation regimens, is emphasized.

Topics: Anticoagulants; Atherosclerosis; Benzamidines; Biocompatible Materials; Biomarkers; Blood Vessels; Enzyme Activation; Guanidines; Heparin; Heparin, Low-Molecular-Weight; Humans; Membranes, Artificial; Neutrophils; Oxidative Stress; Peroxidase; Renal Dialysis; Thrombosis

Topics: 5-Lipoxygenase-Activating Proteins; Animals; Arteriosclerosis; C-Reactive Protein; Carrier Proteins; CD40 Antigens; Cholesterol, HDL; Coronary Disease; Diagnostic Imaging; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Inflammation; Membrane Proteins; Peroxidase; Peroxisome Proliferator-Activated Receptors; Risk Assessment; Thrombosis

In patients with acute coronary syndromes (ACS), early therapy with high-dose statins may reduce short-term adverse clinical outcomes. The mechanisms responsible are not known but could involve anti-inflammatory or anti-thrombotic effects. Compelling evidence from experimental models and clinical studies suggests that the interplay between inflammatory and thrombotic systems, typified by platelet-monocyte and platelet-neutrophil interactions, might be a key regulator of ischemic vascular events. The study sought to determine if early, high-dose administration of the HMG-CoA reductase inhibitor rosuvastatin in the setting of ACS exerts beneficial vascular effects by reducing, and inhibiting biomarkers of thromboinflammation, such as platelet-monocyte and platelet-neutrophil interactions, and biomarkers of myocardial necrosis. A total of 54 patients presenting with ACS within 8 h of symptom onset were randomized to rosuvastatin 40 mg or placebo. Rosuvastatin significantly reduced interactions between platelets and circulating neutrophils (P = 0.015) and monocytes (P = 0.009) within 24 h. No significant effects were observed on platelet aggregation or plasma levels of PF4, sP-selectin, or sCD40L, whereas significant reductions of RANTES occurred over time in both treatment groups. Plasma levels of myeloperoxidase (MPO) declined more rapidly with rosuvastatin therapy than placebo. In a subset of patients with normal cardiac necrosis biomarkers at randomization, rosuvastatin therapy was associated with less myocardial damage as measured by troponin-I or CK-MB. Early administration of high-dose statin therapy in patients with ACS appears to improve biomarkers of inflammation within 8 h, which may translate into fewer ischemic events.

Topics: Acute Coronary Syndrome; Adult; Aged; Biomarkers; Blood Platelets; CD40 Ligand; Cell Communication; Creatine Kinase, MB Form; Dose-Response Relationship, Drug; Early Medical Intervention; Female; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Inflammation; Male; Middle Aged; Monocytes; Neutrophils; P-Selectin; Peroxidase; Platelet Factor 4; Rosuvastatin Calcium; Thrombosis; Treatment Outcome; Troponin I

The biocompatibility of cardiopulmonary bypass surfaces has been improved by heparin and polymer surface modifications. The present study compared the effect of two such coatings on the inflammatory reactions after open heart surgery.. Thirty patients undergoing elective heart surgery were randomly assigned to receive one of two types of coated circuits: Bioline (n=15) or phosphorylcholine (Phisio, n=15). The platelet and leukocyte counts, neutrophil activation (myeloperoxidase), complement activation (C3a and TCC), concentrations of lactate dehydrogenase, 27 cytokines (including interleukins, chemokines and growth factors), thrombin-antithrombin complexes, and the endothelial cell marker syndecan-1 were analyzed at five predetermined time points until 24 hrs post operatively.. Most measurements were comparable in both groups. However, myeloperoxidase was significantly higher in the Bioline group (p < 0.001). Postoperative lactate dehydrogenase concentrations were significantly higher in the Phisio group (p<0.01) and the maximal concentration of thrombin-antithrombin complexes 2 hours postoperatively tended to be higher in the Phisio group (p=0.08), consistent with a longer aortic cross-clamp and cardiopulmonary bypass time.. The two circuits exhibited a comparable degree of in vivo biocompatibility.

Topics: Aged; Anticoagulants; Antithrombin III; Cardiac Surgical Procedures; Cardiopulmonary Bypass; Coated Materials, Biocompatible; Complement C3a; Cytokines; Female; Hemoglobins; Heparin; Humans; Inflammation; L-Lactate Dehydrogenase; Leukocyte Count; Male; Middle Aged; Peptide Hydrolases; Peptides; Peroxidase; Phosphorylcholine; Platelet Count; Syndecan-1; Thrombosis

Pueraria lobata (Willd.) Ohwi and Pueraria lobata var. thomsonii (Benth.) Maesen are nutritious medicine food homology plants that are widely used in the food and health products industry and are excellent natural materials for the development of new health foods, with great potential for domestic and foreign markets. Clinically, P. lobata and P. thomsonii are used to treat coronary heart disease, atherosclerosis, cerebral infarction and other cardiovascular diseases, and antithrombotic actions may be their core effect in the treatment of thrombotic diseases. However, the underlying mechanisms of the antithrombotic properties of P. lobata and P. thomsonii have not been clarified.. First, P. lobata and P. thomsonii were identified by high-performance liquid chromatography (HPLC). An arteriovenous bypass thrombosis rat model was established. Thrombus dry‒wet weight, platelet accumulation rate and the four coagulation indices, including activated partial thromboplastin time (APTT), prothrombin time (PT), thrombin time (TT) and fibrinogen (FIB), were detected in plasma to manifest the P. lobata and P. thomsonii antithrombotic function. Network pharmacology and molecular docking methods were used to obtain key targets and verify reliability. David 6.8 was used for GO and KEGG analyses to explore pathways and potential targets for P. lobata and P. thomsonii antithrombotic functions. Prostaglandin I2 (PGI2), thromboxane A2 (TXA2), cyclooxygenase 2 (COX-2), myeloperoxidase (MPO) and endothelial nitric oxide synthase (eNOS) were tested by enzyme-linked immunosorbent assay (ELISA).. The results indicated that P. lobata and P. thomsonii can reduce thrombus dry‒wet weight and platelet accumulation in rats and inhibit TT, APTT, FIB, and PT. A comprehensive network pharmacology approach successfully identified 9 active ingredients in P. lobata and P. thomsonii. The main active ingredients include polyphenols, amino acids and flavonoids. A total of 15 antithrombotic function targets were obtained, including 3 key targets (PTGS2, NOS3, MPO). Pathway analysis showed 10 significant related pathways and 29 biological processes. P. lobata and P. thomsonii inhibited platelet aggregation by upregulating PGI2 and downregulating TXA2, inhibited PTGS2 to reduce inflammation, and increased the level of eNOS to promote vasodilation. In addition, P. lobata and P. thomsonii alleviated oxidative stress by increasing SOD levels and significantly decreasing MDA contents.. The results of the study further clarify the antithrombotic mechanism of action of P. lobata and P. thomsonii, which provides a scientific basis for the development of new drugs for thrombogenic diseases and lays the foundation for the development of P. lobata and P. thomsonii herbal resources and P. lobata and P. thomsonii health products.

Topics: Amino Acids; Animals; Cyclooxygenase 2; Epoprostenol; Fibrinogen; Fibrinolytic Agents; Flavonoids; Molecular Docking Simulation; Network Pharmacology; Nitric Oxide Synthase Type III; Peroxidase; Pueraria; Rats; Reproducibility of Results; Superoxide Dismutase; Thrombosis; Thromboxane A2

Due to its unique properties and high biomedical relevance fibrinogen is a promising protein for the development of various matrixes and scaffolds for biotechnological applications. Fibrinogen molecules may form extensive clots either upon specific cleavage by thrombin or in thrombin-free environment, for example, in the presence of different salts. Here, we report the novel type of non-conventional fibrinogen clot formation, which is mediated by myeloperoxidase and takes place even at low fibrinogen concentrations (<0.1 mg/ml). We have revealed fibrillar nature of myeloperoxidase-mediated fibrinogen clots, which differ morphologically from fibrin clots. We have shown that fibrinogen clotting is mediated by direct interaction of myeloperoxidase molecules with the outer globular regions of fibrinogen molecules followed by fibrinogen unfolding from its natural trinodular to a fibrillar structure. We have demonstrated a major role of the Debye screening effect in regulating of myeloperoxidase-induced fibrinogen clotting, which is facilitated by small ionic strength. While fibrinogen in an aqueous solution with myeloperoxidase undergoes changes, the enzymatic activity of myeloperoxidase is not inhibited in excess of fibrinogen. The obtained results open new insights into fibrinogen clotting, give new possibilities for the development of fibrinogen-based functional biomaterials, and provide the novel concepts of protein unfolding.

Topics: Blood Coagulation; Fibrin; Fibrinogen; Humans; Peroxidase; Thrombin; Thrombosis

During the COVID-19 pandemic, vaccination is the most important countermeasure. Pharmacovigilance concerns however emerged with very rare, but potentially disastrous thrombotic complications following vaccination with ChAdOx1. Platelet factor-4 antibody mediated vaccine-induced immune thrombotic thrombocytopenia (VITT) was described as an underlying mechanism of these thrombotic events. Recent work moreover suggests that mechanisms of immunothrombosis including neutrophil extracellular trap (NET) formation might be critical for thrombogenesis during VITT. In this study, we investigated blood and thrombus specimens of a female patient who suffered severe stroke due to VITT after vaccination with ChAdOx1 in comparison to 13 control stroke patients with similar clinical characteristics. We analyzed cerebral thrombi using histological examination, staining of complement factors, NET-markers, DNase and LL-37. In blood samples at the hyper-acute phase of stroke and 7 days later, we determined cell-free DNA, myeloperoxidase-histone complexes, DNase activity, myeloperoxidase activity, LL-37 and inflammatory cytokines. NET markers were identified in thrombi of all patients. Interestingly, the thrombus of the VITT-patient exclusively revealed complement factors and high amounts of DNase and LL-37. High DNase activity was also measured in blood, implying a disturbed NET-regulation. Furthermore, serum of the VITT-patient inhibited reactive oxygen species-dependent NET-release by phorbol-myristate-acetate to a lesser degree compared to controls, indicating either less efficient NET-inhibition or enhanced NET-induction in the blood of the VITT-patient. Additionally, the changes in specific cytokines over time were emphasized in the VITT-patient as well. In conclusion, insufficient resolution of NETs, e.g. by endogenous DNases or protection of NETs against degradation by embedded factors like the antimicrobial peptide LL-37 might thus be an important factor in the pathology of VITT besides increased NET-formation. On the basis of these findings, we discuss the potential implications of the mechanisms of disturbed NETs-degradation for diagnostic and therapeutic approaches in VITT-related thrombogenesis, other auto-immune disorders and beyond.

Topics: COVID-19; Deoxyribonuclease I; Deoxyribonucleases; Extracellular Traps; Female; Humans; Neutrophils; Pandemics; Peroxidase; Platelet Factor 4; Purpura, Thrombocytopenic, Idiopathic; Stroke; Thrombocytopenia; Thrombosis; Vaccines

Neutrophil extracellular traps (NETs) are web-like structures consisting of DNA, histones and granule proteins, released from neutrophils in thrombus formation, inflammation, and cancer. We asked if plasma levels of the NET markers myeloperoxidase (MPO)-DNA and citrullinated histone H3 (H3Cit)-DNA, are elevated in liver cirrhosis and hepatocellular carcinoma (HCC) and if the levels correlate with clinical parameters. MPO-DNA, H3Cit-DNA, and thrombin-antithrombin (TAT) complex, as a marker of coagulation activity, were measured using ELISA in plasma from 82 patients with HCC, 95 patients with cirrhosis and 50 healthy controls. Correlations were made to clinical parameters and laboratory data and patients were followed for a median of 22.5 months regarding thrombosis development. H3Cit-DNA was significantly (p < 0.01) elevated in plasma from cirrhosis (66.4 ng/mL) and HCC (63.8 ng/mL) patients compared to healthy controls (31.8 ng/mL). TAT levels showed similar pattern (3.1, 3.7, and 0.0 µg/mL respectively, p < 0.01). MPO-DNA was significantly (p < 0.01) elevated in cirrhosis patients (0.53 O.D.) as compared to controls (0.33 O.D.). Levels of MPO-DNA and H3Cit-DNA correlated positively with Child-Pugh and MELD score. TAT was increased in all Child-Pugh and MELD groups. In multivariable logistic regression, Child B and C liver cirrhosis were independent predictors of elevated H3Cit-DNA in plasma. Levels of MPO-DNA and H3Cit-DNA were similar in patients with or without history of thrombosis, or thrombus formation during follow-up. In conclusion, plasma markers of NET formation are elevated in liver cirrhosis and correlate to the degree of liver dysfunction in patients with liver cirrhosis and/or HCC. The presence of HCC did not further increase the plasma levels of NET markers as compared to patients with cirrhosis only.

Topics: Aged; Antithrombin III; Biomarkers; Carcinoma, Hepatocellular; Case-Control Studies; Citrullination; DNA; Extracellular Traps; Female; Histones; Humans; Inflammation; Liver; Liver Cirrhosis; Liver Neoplasms; Male; Middle Aged; Neutrophils; Peptide Hydrolases; Peroxidase; Thrombosis

Myeloperoxidase antineutrophil cytoplasmic antibody (MPO-ANCA) associated vasculitis is an autoimmune disease usually with severe multiple dysfunction syndrome, especially prominent acute renal failure. A 65-year-old woman was admitted with progressive dyspnoea for six months and fever, sputum with blood, pain of the lower extremities and intermittent claudication for two days, indicating multiple organ involvement (respiratory system, blood vessels). The renal involvement was relatively mild, presenting with microscopic haematuria. The chest computed tomography demonstrated multiple pulmonary embolisms. Ultrasound and computed tomography angiography for the lower extremity vessels showed venous and arterial thrombosis. Exclusion of other diseases that can cause multiple organ damage and thrombosis, the positive perinuclear ANCA and MPO-ANCA strongly support the diagnosis of MPO-ANAC-associated vasculitis. The patient's physical condition has been greatly improved by treatment with corticosteroids and anticoagulation.

Topics: Aged; Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis; Female; Humans; Lower Extremity; Peroxidase; Pulmonary Embolism; Thrombosis

Myeloperoxidase (MPO) secreted by neutrophils is the enzyme that kills bacteria and other pathogens. Acute myocardial infarction (AMI) is usually caused by thrombosis in response to vulnerable plaque rupture. Circulating MPO was found to be associated with increased mortality in AMI patients. However, the relationship between MPO in thrombi and the prognosis of AMI patients remains unknown.. MPO expression in thrombi is associated with the prognosis of patients who underwent primary percutaneous coronary intervention (PCI) after AMI.. This study included 41 consecutive patients with acute ST-elevation myocardial infarction, who successfully underwent primary PCI, during which we collected thrombi remaining in the culprit artery using aspiration catheters. These thrombus samples were fixed, and immunohistochemical staining against MPO and heme oxygenase-1 (HO-1) was conducted. Enrolled patients were divided into two groups based on the induction of thrombotic MPO, which was quantified using Image J software.. We observed that increased MPO was associated with lower left ventricular ejection fraction (LVEF) and worse LV remodeling in AMI patients. Instead, patients with decreased thrombotic MPO induction had a considerable improvement in LVEF 1 month after discharge (54.4 ± 2.0% vs. 61.1 ± 2.3%, p < 0.01). In the MPO group, a reduction in the thrombotic HO-1 level contributed to the development of adverse LV remodeling. Logistic regression showed that MPO was a considerable risk factor for adverse LV remodeling (adjusted OR 3.70, p < 0.05).. MPO expression in thrombi is associated with reduced LVEF and deteriorated LV remodeling in AMI patients, which may be due to HO-1 suppression in thrombi.

Topics: Heme Oxygenase-1; Humans; Percutaneous Coronary Intervention; Peroxidase; ST Elevation Myocardial Infarction; Stroke Volume; Thrombosis; Ventricular Function, Left; Ventricular Remodeling

Saccular aneurysms are thought to have a worse prognosis than fusiform aneurysms in humans, due to hemodynamic reasons. However, data comparing hemodynamic and biology in saccular and fusiform aneurysms are lacking. The main objective was to evaluate the impact of aneurysm morphology on intra-luminal thrombus (ILT) formation and activity.. Forty Lewis rats were ran-domly divided into 2 groups of 20: "saccular" (Group A) and "fusiform" (Group B) aneurysms. Decellularized thoracic aortas from guinea pigs were xenografted to create saccular or fusiform aneurysms. Final imaging evaluation of the aneurysms was carried out during the third week, by quantitative Doppler ultrasound and magnetic resonance imaging. Assays of myeloperoxidase (MPO), platelet factor 4 (PF4), advanced oxidation protein products (AOPPs) iron and matrix metallopeptidase-9 (MMP-9) were performed as biological criteria.. Quantitatively, saccular aneurysms are characterized by a more thicker ILT, lower inflow velocities and more important relative backflow velocities as compared to fusiform aneurysms. Compared to fusiform, saccular aneurysms released significantly more MPO (p = 0.004), PF4 (p = 0.02), AOPPs (p < 0.002), iron (p < 0.0001) and MMP-9 (p < 0.04).. Experimental saccular and fusiform aneurysms show differential specific hemodynamics, which seem to impact the histology and the biology of the ILT in each type of aneurysm.

Topics: Advanced Oxidation Protein Products; Animals; Aorta, Abdominal; Aortic Aneurysm, Abdominal; Disease Models, Animal; Guinea Pigs; Hemodynamics; Iron; Male; Matrix Metalloproteinase 9; Peroxidase; Platelet Factor 4; Rats, Inbred Lew; Thrombosis; Time Factors

COVID-19 affects millions of patients worldwide, with clinical presentation ranging from isolated thrombosis to acute respiratory distress syndrome (ARDS) requiring ventilator support. Neutrophil extracellular traps (NETs) originate from decondensed chromatin released to immobilize pathogens, and they can trigger immunothrombosis. We studied the connection between NETs and COVID-19 severity and progression. We conducted a prospective cohort study of COVID-19 patients (n = 33) and age- and sex-matched controls (n = 17). We measured plasma myeloperoxidase (MPO)-DNA complexes (NETs), platelet factor 4, RANTES, and selected cytokines. Three COVID-19 lung autopsies were examined for NETs and platelet involvement. We assessed NET formation ex vivo in COVID-19 neutrophils and in healthy neutrophils incubated with COVID-19 plasma. We also tested the ability of neonatal NET-inhibitory factor (nNIF) to block NET formation induced by COVID-19 plasma. Plasma MPO-DNA complexes increased in COVID-19, with intubation (P < .0001) and death (P < .0005) as outcome. Illness severity correlated directly with plasma MPO-DNA complexes (P = .0360), whereas Pao2/fraction of inspired oxygen correlated inversely (P = .0340). Soluble and cellular factors triggering NETs were significantly increased in COVID-19, and pulmonary autopsies confirmed NET-containing microthrombi with neutrophil-platelet infiltration. Finally, COVID-19 neutrophils ex vivo displayed excessive NETs at baseline, and COVID-19 plasma triggered NET formation, which was blocked by nNIF. Thus, NETs triggering immunothrombosis may, in part, explain the prothrombotic clinical presentations in COVID-19, and NETs may represent targets for therapeutic intervention.

Topics: Adult; Aged; Betacoronavirus; Blood Platelets; Blood Proteins; Coronavirus Infections; COVID-19; Extracellular Traps; Female; Humans; Male; Middle Aged; Neutrophil Infiltration; Neutrophils; Pandemics; Peroxidase; Pneumonia, Viral; Prospective Studies; SARS-CoV-2; Thrombosis

Topics: Antibodies, Antineutrophil Cytoplasmic; Humans; Necrosis; Peroxidase; Thrombosis; Vasculitis

Cancer patients are at increased risk of developing thrombosis, comorbidity that has been associated with increased neutrophil counts and the formation of neutrophil extracellular traps (NETs). Interleukin-1β (IL-1β) modulates the expression of granulocyte colony-stimulating factor (G-CSF), a cytokine that promotes cancer-associated neutrophilia and NET generation. Herein, we combined a murine breast cancer model with a flow-restriction thrombosis model to evaluate whether the IL-1β blockade could interfere with cancer-associated thrombosis. Mice bearing metastatic 4T1 tumors exhibited high neutrophil counts as well as elevated expression of G-CSF and IL-1β in their tumors. On the other hand, mice bearing non-metastatic 67NR tumors showed no elevation in neutrophil counts and displayed low expression levels of G-CSF and IL-1β in their tumors. 4T1 tumor-bearing mice but not 67NR tumor-bearing mice exhibited a NET-dependent prothrombotic state. Pharmacological blockade of IL-1 receptor (IL-1R) decreased the primary growth of 4T1 tumors and reduced the systemic levels of myeloperoxidase, cell-free DNA (cfDNA) and G-CSF, without interfering with the neutrophil counts. Most remarkably, the blockade of IL-1R abolished the prothrombotic state observed in 4T1 tumor-bearing mice. Overall, our results demonstrate that IL-1β might be a feasible target to attenuate cancer-associated thrombosis, particularly in cancer types that rely on increased G-CSF production and involvement of NET formation.

Topics: Animals; Breast Neoplasms; Cell Line, Tumor; Extracellular Traps; Female; Granulocyte Colony-Stimulating Factor; Humans; Interleukin 1 Receptor Antagonist Protein; Interleukin-1beta; Leukocyte Count; Mammary Neoplasms, Experimental; Mice, Inbred BALB C; Neutrophils; Peroxidase; Receptors, Interleukin-1; Thrombosis; Tumor Burden

Type 2 diabetes mellitus (T2DM) is associated with a hypercoagulable state and increased neutrophil extracellular traps formation (NETosis). We investigated predictors of NETosis and cell death markers in circulating blood and their association with a prothrombotic state in T2DM.. In a cross-sectional study involving 113 T2DM patients aged 63.7 ± 8.2 years, we investigated citrullinated histone H3 (H3Cit), cell-free deoxyribonucleic acid (cfDNA), myeloperoxidase, neutrophil elastase, and inflammation markers, along with thrombin generation (TG), plasma clot lysis time (CLT), clot permeability (K. On multivariate logistic regression analysis adjusted for age and gender, predictors of high H3Cit (≥ 7.36 ng/mL, upper quartile) were: glycated hemoglobin (HbA1c) ≥ 7.0% and interleukin-6. Interleukin-6 was also found to be a predictor of high cfDNA (≥ 2.84 µg/mL, upper quartile) along with glucose. Citrullinated histone H3 and cfDNA correlated positively with CLT and inversely with K. In T2DM, NETosis detectable in circulating blood is associated with inflammatory state and a prothrombotic state, especially hypofibrinolysis.

Topics: Aged; Biomarkers; Blood Glucose; Cell-Free Nucleic Acids; Citrullination; Cross-Sectional Studies; Diabetes Mellitus, Type 2; DNA; Extracellular Traps; Female; Fibrinolysis; Glycated Hemoglobin; Histones; Humans; Inflammation Mediators; Leukocyte Elastase; Male; Middle Aged; Peroxidase; Risk Factors; Thrombin; Thrombosis

Hypochlorite (HOCl), a strong oxidant and antimicrobial agent, has been proposed to be associated with hemostatic abnormalities during inflammatory response. However, its complex impact on hemostasis is not completely understood. In this report we studied the effect of clinically relevant (micromolar) HOCl concentrations on thrombus formation under flow, kinetics of platelet-fibrin clot formation, its architecture, retraction, and lysis. We found that HOCl (up to 500 µM) did not affect kinetics of coagulation measured in whole blood. HOCl (500-1000 µM) markedly diminished thrombus formation under flow. Clot retraction rate was reduced by HOCl dose-dependently (50-500 µM). HOCl (125-500 µM) inhibited fibrinolysis in whole blood and in platelet-depleted plasma, dose-dependently. Activity of plasmin was reduced by HOCl at concentrations started from 500 µM. HOCl (up to 500 µM) did not reduce plasminogen binding to fibrin under flow. HOCl (125-500 µM) modulated architecture of fibrin- and platelet-fibrin clots towards structures made of thin and densely packed fibers. Exposure of pure fibrinogen to HOCl (10-1000 µM) resulted in formation of dityrosine and was associated with altered fibrin structure derived from such modified fibrinogen. HOCl-altered fibrin net structure was not related with modulation of platelet procoagulant response, thrombin generation, and factor XIII activity. We conclude that, in human blood, clinically relevant HOCl concentrations may inhibit thrombus formation under flow, clot retraction and fibrinolysis. Fibrinolysis and clot retraction seem to be the most sensitive to HOCl-evoked inhibition. HOCl-modified fibrinogen and altered clot structure associated with it are likely to be primary sources of attenuated fibrinolysis.

Topics: Blood Coagulation; Blood Platelets; Clot Retraction; Factor XIII; Fibrin; Fibrinogen; Fibrinolysis; Hemostasis; Humans; Hypochlorous Acid; Inflammation; Peroxidase; Thrombin; Thrombosis

Neutrophil extracellular traps (NETs) are networks of extracellular fibres produced from neutrophil DNA with a pathogenic role in infection, thrombosis and other conditions. Reliable assays for measuring NETs are desirable as novel treatments targeting NETs are being explored for the treatment of these conditions. We compare a whole blood flow cytometry method with serum assays to measure NETs-associated markers in patients with sepsis and thrombosis.. Patients with deep venous thrombosis (n = 25), sepsis (n = 21) and healthy controls (n = 23) were included in the study. Neutrophil surface NETs markers were determined by flow cytometry on whole blood samples by gating of neutrophils stained for surface citrullinated histone (H3cit) and myeloperoxidase (MPO). Serum double-stranded (ds) DNA, MPO, myeloid-related protein, nucleosomes, DNAse, elastase, human high-mobility group box 1 and MPO-DNA complexes were quantified as circulating markers of NETs.. Neutrophil NETs markers by flow cytometry and serum NETs markers were significantly higher in patients with thrombosis and sepsis compared with healthy controls. Neutrophil NETs markers significantly correlated with the serum marker dsDNA.. Flow cytometry detection of neutrophil NETs markers is feasible in whole blood and correlates with serum markers of NETs. We propose the flow cytometry detection of MPO/H3cit positive neutrophils and serum dsDNA as simple methods to quantify cellular and extracellular NET markers in patients with thrombosis and sepsis.

Topics: Biomarkers; Case-Control Studies; DNA; Extracellular Traps; Flow Cytometry; Histones; Humans; Peroxidase; Sepsis; Thrombosis

Acute coronary syndromes can be initiated by either atherosclerotic fibrous cap ruptures, superficial plaque erosions or intraplaque haemorrhages (IPHs). Since neutrophil extracellular traps (NETs) display pro-inflammatory and pro-thrombotic properties, we investigated the presence, extent and distribution of neutrophils and NETs in different types of plaque complications in relation to the age of overlying thrombus mass or haemorrhage. Sixty-four paraffin-embedded coronary plaque segments of 30 acute myocardial infarction patients were retrieved from the autopsy archives, which contained 44 complicated plaques (17 IPHs, 9 erosions and 18 ruptures) and 20 intact plaques. Complicated plaques were further categorized according to the histological age of thrombus or haemorrhage. Immunohistochemistry was performed to visualize neutrophils (anti-myeloperoxidase, anti-elastase and anti-CD177) and NETs (anti-citrullinated histone-3 and anti-peptidyl-arginine-deiminase-4). The results were scored semi-quantitatively. Neutrophils and NETs were abundantly present in all types of complicated, but not in intact, plaques (

Topics: Coronary Artery Disease; Disease Progression; Endocytosis; Extracellular Traps; Hemorrhage; Histones; Humans; Immunohistochemistry; Myocardial Infarction; Neutrophil Infiltration; Neutrophils; Paraffin Embedding; Peroxidase; Plaque, Atherosclerotic; Thrombosis

This study aimed to explore the effects of ticagrelor (a P2Y12 receptor inhibitor) on interleukin (IL)-17 and myeloperoxidase (MPO) expression in coronary thrombus as well as on the coronary blood flow in ST-segment elevation myocardial infarction (STEMI) patients following percutaneous coronary intervention (PCI).. Forty STEMI patients who were admitted to the First Affiliated Hospital of Harbin Medical University between August 1, 2014 and December 30, 2014 were enrolled in this study according to a set inclusion criteria. They were randomized to ticagrelor and clopidogrel groups and treated with 180 mg ticagrelor and 600 mg clopidogrel before PCI, respectively. Intracoronary thrombus aspiration was performed by a physician during PCI. Immunohistochemistry and Western blot analysis were carried out to detect the expression of IL-17 and MPO in the thrombus. Corrected thrombolysis in myocardial infarction frame count (CTFC) was used to evaluate blood flow after PCI.. Immunohistochemistry results showed that the average positive staining area percentage of IL-17 and MPO in the clopidogrel group was significantly higher than that in the ticagrelor group. Western blot analysis also showed similar results for IL-17 (clopidogrel 0.71 ± 0.036, ticagrelor 0.50 ± 0.56) and MPO (clopidogrel 0.50 ± 0.040; ticagrelor 0.38 ± 0.06). CTFC was lower in the ticagrelor group than that in the clopidogrel group (P < 0.05).. Ticagrelor is more effective than clopidogrel in reducing inflammation thrombosis and improving postprocedural PCI blood flow in STEMI patients. This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

Topics: Adolescent; Adult; Aged; Clopidogrel; Coronary Circulation; Female; Humans; Interleukin-17; Middle Aged; Peroxidase; Platelet Aggregation Inhibitors; ST Elevation Myocardial Infarction; Thrombosis; Ticagrelor; Young Adult

Neutrophil extracellular traps (NETs) are extracellular DNA filaments formed during neutrophil activation. This process, called netosis, was originally associated with neutrophil antibacterial properties. However, several lines of evidence now suggest a major role for netosis in thrombosis, autoimmune diseases, and cancer. We demonstrate here that highly purified human blood monocytes are also capable of extracellular trap (ET) release in response to several stimuli. Monocyte ETs display a morphology analogous to NETs and are associated with myeloperoxidase (MPO), lactoferrin (LF), citrullinated histones, and elastase. Monocyte ET release depends on oxidative burst but not on MPO activity, in contrast to neutrophils. Moreover, we demonstrate procoagulant activity for monocyte ETs, a feature that could be relevant to monocyte thrombogenic properties. This new cellular mechanism is likely to have implications in the multiple pathologic contexts where monocytes are implicated, such as inflammatory disorders, infection, or thrombosis.

Topics: Extracellular Traps; Histones; Humans; Infections; Inflammation; Lactoferrin; Monocytes; Pancreatic Elastase; Peroxidase; Thrombosis

Hypoxia preconditioning has been proven to be an effective method to enhance the therapeutic action of mesenchymal stem cells (MSCs). However, the beneficial effects of hypoxic MSCs in ischemia/reperfusion (I/R) lung injury have yet to be investigated. In this study, we hypothesized that the administration of hypoxic MSCs would have a positive therapeutic impact on I/R lung injury at molecular, cellular, and functional levels.. I/R lung injury was induced in isolated and perfused rat lungs. Hypoxic MSCs were administered in perfusate at a low (2.5×105 cells) and high (1×106 cells) dose. Rats ventilated with a low tidal volume of 6 ml/kg served as controls. Hemodynamics, lung injury indices, inflammatory responses and activation of apoptotic pathways were determined.. I/R induced permeability pulmonary edema with capillary leakage and increased levels of reactive oxygen species (ROS), pro-inflammatory cytokines, adhesion molecules, cytosolic cytochrome C, and activated MAPK, NF-κB, and apoptotic pathways. The administration of a low dose of hypoxic MSCs effectively attenuated I/R pathologic lung injury score by inhibiting inflammatory responses associated with the generation of ROS and anti-apoptosis effect, however this effect was not observed with a high dose of hypoxic MSCs. Mechanistically, a low dose of hypoxic MSCs down-regulated P38 MAPK and NF-κB signaling but upregulated glutathione, prostaglandin E2, IL-10, mitochondrial cytochrome C and Bcl-2. MSCs infused at a low dose migrated into interstitial and alveolar spaces and bronchial trees, while MSCs infused at a high dose aggregated in the microcirculation and induced pulmonary embolism.. Hypoxic MSCs can quickly migrate into extravascular lung tissue and adhere to other inflammatory or structure cells and attenuate I/R lung injury through anti-oxidant, anti-inflammatory and anti-apoptotic mechanisms. However, the dose of MSCs needs to be optimized to prevent pulmonary embolism and thrombosis.

Topics: Animals; Antioxidants; Apoptosis; Biomarkers; Bronchoalveolar Lavage Fluid; Capillaries; Caspase 3; Cell Hypoxia; Cytochromes c; Cytosol; Glutathione; Hemodynamics; Hydrogen Peroxide; Intercellular Adhesion Molecule-1; Leukocyte Count; Lung Injury; Male; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mitogen-Activated Protein Kinases; NF-kappa B; Organ Size; Peroxidase; Protein Carbonylation; Proto-Oncogene Proteins c-bcl-2; Pulmonary Embolism; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction; Thiobarbituric Acid Reactive Substances; Thrombosis; Vascular Cell Adhesion Molecule-1

Myeloperoxidase (MPO) is a cationic heme enzyme stored in neutrophilic polymorphonuclear leukocytes (PMNs) that has recently been implicated in inflammatory cell signaling and tissue damage. Although PMNs play a critical role in both innate immunity and vascular thrombosis, no previous study has systematically investigated the effect of MPO on blood coagulation. Here, we show that PMN-derived MPO inhibits the procoagulant activity (PCA) of lipidated recombinant human tissue factor (rhTF) in a time- and concentration-dependent manner that involves, but is not entirely dependent on the enzyme's catalytic activity. Similarly, MPO together with its substrate, H2O2, inhibited the PCA of plasma microvesicles isolated from lipopolysaccharide (LPS)-stimulated whole blood, an effect additive to that of a function blocking TF antibody. Treatment of whole blood with LPS or phorbol-myristate-acetate dramatically increased MPO plasma levels, and co-incubation with 4-ABAH, a specific MPO inhibitor, significantly enhanced the PCA in plasma supernatants. MPO and MPO/H2O2 also inhibited the PCA of activated platelets and purified phospholipids (PLs), suggesting that modulation of negatively charged PLs, i.e., phosphatidylserine, rather than direct interference with the TF/FVIIa initiation complex was involved. Consistently, pretreatment of activated platelets with MPO or MPO/H2O2 attenuated the subsequent binding of lactadherin, which specifically recognizes procoagulant PS on cell membranes. Finally, endogenously released MPO regulated the PCA of THP1 cells in an autocrine manner dependent on the binding to CD11b/CD18 integrins. Collectively, these findings indicate that MPO is a negative regulator of PL-dependent coagulation and suggest a more complex role of activated PMNs in haemostasis and thrombosis.

Topics: Blood Coagulation; Factor VIIa; HL-60 Cells; Humans; Hydrogen Peroxide; Lipopolysaccharides; Neutrophils; Peroxidase; Phosphatidylserines; Phospholipids; Protein Binding; Secretory Vesicles; THP-1 Cells; Thromboplastin; Thrombosis

ADAMTS13 is a plasma metalloproteinase that cleaves large multimeric forms of von Willebrand factor (VWF) to smaller, less adhesive forms. ADAMTS13 activity is reduced in systemic inflammatory syndromes, but the cause is unknown. Here, we examined whether neutrophil-derived oxidants can regulate ADAMTS13 activity. We exposed ADAMTS13 to hypochlorous acid (HOCl), produced by a myeloperoxidase-H2O2-Cl(-) system, and determined its residual proteolytic activity using both a VWF A2 peptide substrate and multimeric plasma VWF. Treatment with 25 nm myeloperoxidase plus 50 μm H2O2 reduced ADAMTS13 activity by >85%. Using mass spectrometry, we demonstrated that Met(249), Met(331), and Met(496) in important functional domains of ADAMTS13 were oxidized to methionine sulfoxide in an HOCl concentration-dependent manner. The loss of enzyme activity correlated with the extent of oxidation of these residues. These Met residues were also oxidized in ADAMTS13 exposed to activated human neutrophils, accompanied by reduced enzyme activity. ADAMTS13 treated with either neutrophil elastase or plasmin was inhibited to a lesser extent, especially in the presence of plasma. These observations suggest that oxidation could be an important mechanism for ADAMTS13 inactivation during inflammation and contribute to the prothrombotic tendency associated with inflammation.

Topics: ADAM Proteins; ADAMTS13 Protein; Chromatography, Liquid; Fibrinolysin; Gene Expression Regulation, Enzymologic; Humans; Hydrogen Peroxide; Hypochlorous Acid; Inflammation; Leukocyte Elastase; Mass Spectrometry; Methionine; Neutrophils; Oxidants; Oxygen; Peroxidase; Protein Structure, Tertiary; Thrombosis; von Willebrand Factor

Myeloperoxidase is a proinflammatory protein that appears as a result of increased oxidative stress. It plays an important role in the promotion and progression of atherosclerosis. The aim of this study was to determine the importance of MPO as a predictive parameter for thrombosis of arteriovenous fistula (AVF). The study involved monitoring patients with AVFs for hemodialysis over a period of 2 years. There were 41 patients, 19 (46%) men and 22 (54%) women, with mean age of 65 ± 12.7 years. Routine laboratory analyses were carried out in all respondents, including determination of MPO concentration. Gender, demographic and anthropometrical characteristics, smoking, alcohol consumption, as well as the presence of diabetic nephropathy, as an etiological factor of kidney disease, were recorded. The group of patients who developed initial thrombosis of the AVFs had significantly different values for leukocytes (8.5 ± 3.8 vs. 7.3 ± 2.1, P = 0.024), erythrocytes (2.8 ± 0.27 vs. 3.2 ± 0.65; P = 0.019), hemoglobin (88.5 ± 81 vs. 99.1 ± 6.02; P = 0.041), and myeloperoxidase (19.3 ± 4.67 vs. 11.1 ± 4.43; P = 0.007) when compared with the group without fistula thrombosis. Diabetic nephropathy (P = 0.02) characterized the group of patients with thrombosis of the fistula. Diabetic nephropathy (B = 2.53, P = 0.049) and MPO (B = 0.03, P = 0.029) were statistically significant predictors of fistula thrombosis. In our study, MPO and diabetic nephropathy were predictors of thrombosis of the AVF.

Topics: Aged; Arteriovenous Fistula; Diabetic Nephropathies; Female; Humans; Male; Oxidative Stress; Peroxidase; Predictive Value of Tests; Renal Dialysis; Thrombosis

An association of intraluminal thrombus (ILT) with abdominal aortic aneurysm (AAA) growth has been suggested. Previous in vitro experiments have demonstrated that aneurysm-associated thrombus may secrete proteolytic enzymes and may develop local hypoxia that might lead to the formation of tissue-damaging reactive oxygen species. In this study, we assessed the hypothesis that ventral ILT thickness is associated with markers of proteolysis and with lipid oxidation in the underlying AAA vessel wall.. Ventral AAA tissue was collected from asymptomatic patients at the site of maximal diameter during open aneurysm repair. Segments were divided, one part for biochemical measurements and one for histologic analyses. We measured total cathepsin B, cathepsin S levels, and matrix metalloproteinase (MMP)-2 and MMP-9 activity. Myeloperoxidase and thiobarbituric acid reactive substances were determined as measures of lipid oxidation. Histologic segments were analyzed semiquantitatively for the presence of collagen, elastin, vascular smooth muscle cells (VSMCs), and inflammatory cells. Preoperative computed tomography angiography scans of 83 consecutive patients were analyzed. A three-dimensional reconstruction was obtained, and a center lumen line of the aorta was constructed. Ventral ILT thickness was measured in the anteroposterior direction at the level of maximal aneurysm diameter on the orthogonal slices.. Ventral ILT thickness was positively correlated with aortic diameter (r=0.25; P=.02) and with MMP-2 levels (r=0.27; P=.02). No biochemical correlations were observed with MMP-9 activity or cathepsin B and S expression. No correlation between ventral ILT thickness and myeloperoxidase or thiobarbituric acid reactive substances was observed. Ventral ILT thickness was negatively correlated with VSMCs (no staining, 18.5 [interquartile range, 12.0-25.5] mm; minor, 17.6 [10.7-22.1] mm; moderate, 14.5 [4.6-21.7] mm; and heavy, 8.0 [0.0-12.3] mm, respectively; P=.01) and the amount of elastin (no staining, 18.6 [12.2-30.0] mm; minor, 16.5 [9.0-22.1] mm; moderate, 11.7 [2.5-15.3] mm; and heavy 7.7 [0.0-7.7] mm, respectively; P=.01) in the medial aortic layer.. ILT thickness appeared to be associated with VSMCs apoptosis and elastin degradation and was positively associated with MMP-2 concentrations in the underlying wall. This suggests that ILT thickness affects AAA wall stability and might contribute to AAA growth and rupture. ILT thickness was not correlated with markers of lipid oxidation.

Topics: Aged; Aorta, Abdominal; Aortic Aneurysm, Abdominal; Aortic Rupture; Aortography; Apoptosis; Biopsy; Cathepsin B; Cathepsins; Collagen; Elastin; Female; Humans; Inflammation; Linear Models; Lipid Peroxidation; Logistic Models; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Multivariate Analysis; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Observer Variation; Peroxidase; Predictive Value of Tests; Reproducibility of Results; Thiobarbituric Acid Reactive Substances; Thrombosis; Tomography, X-Ray Computed

Neutrophils are important cellular sources of interleukin (IL) 17A and -F. Moreover, upon activation neutrophils are able to excrete chromatin embedded with components from their cytoplasmic granules to form 'neutrophil extracellular traps' (NETs). Recent studies suggested that NETs contribute to thrombosis by promoting fibrin deposition and platelet aggregation. IL17A may also promote thrombosis by enhancing platelet aggregation. In the present study we investigated the presence of neutrophils, NETs and IL17A and -F in coronary thrombosuction specimens obtained from patients after acute myocardial infarction. Neutrophils and NETs were identified using histochemical (HE, Feulgen procedure) and immunohistochemical stainings (Histone H1, myeloperoxidase, neutrophil elastase) in 15 fresh, 15 lytic and 15 organised thrombi. The presence and distribution of IL17A and -F was studied using (immuno)histochemical double staining and spectral image analysis, rtPCR and Western blot. High numbers of neutrophils are present (10-30% of the thrombus mass) in fresh and lytic, but not in organized thrombus. NETs were frequently observed in fresh (4/15) and lytic (12/15), but never in organised thrombus specimens. Double staining combining the Feulgen reaction with Histone-H1, MPO or neutrophil elastase confirmed colocalisation with DNA. Cytoplasmatic IL17A/F staining was found in the majority of the neutrophils, extracellularly and in NETs. Western blotting confirmed the presence of IL17A and IL17F in thrombus specimens. In conclusion, a large burden of neutrophils, neutrophil extracellular traps and IL17A and -F are important constituents of fresh and lytic thrombus after acute myocardial infarction. The specific colocalisation of these indicates a role during thrombus stabilisation and growth.

Topics: Biomarkers; Blotting, Western; Histones; Humans; Immunohistochemistry; Interleukin-17; Leukocyte Elastase; Myocardial Infarction; Neutrophil Activation; Neutrophils; Peroxidase; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Rosaniline Dyes; Thrombectomy; Thrombosis

Experiments were designed to investigate the effects of ethyl pyruvate (EP) in a murine model of hind-limb ischemia-reperfusion (IR) injury.. C57BL6 mice underwent 90 minutes of unilateral ischemia followed by 24 hours of reperfusion using two treatment protocols. For the preischemic treatment (pre-I) protocol, mice (n=6) were given 300 mg/kg EP before ischemia, followed by 150 mg/kg of EP just before reperfusion and at 6 hours and 12 hours after reperfusion. In a postischemic treatment (post-I) protocol, mice (n=7) were treated with 300 mg/kg EP at the end of the ischemic period, then 15 minutes later, and 2 hours after reperfusion and 150 mg/kg of EP at 4 hours, 6 hours, 10 hours, 16 hours, and 22 hours after reperfusion. Controls mice for both protocols were treated with lactated Ringers alone at time intervals identical to EP. Skeletal muscle levels of adenosine triphosphate (ATP), interleukin-1β, keratinocyte chemoattractant protein, and thrombin antithrombin-3 complex were measured. Skeletal muscle architectural integrity was assessed microscopically.. ATP levels were higher in mice treated with EP compared with controls under the both treatment protocols (p=0.02). Interleukin-1β, keratinocyte chemoattractant protein, thrombin antithrombin-3 complex (p<0.05), and the percentage of injured fibers (p<0.0001) were significantly decreased in treated versus control mice under the both protocols.. Muscle fiber injury and markers of tissue thrombosis and inflammation were reduced, and ATP was preserved with EP in pre-I and post-I protocols. Further investigation of the efficacy of EP to modulate IR injury in a larger animal model of IR injury is warranted.

Topics: Adenosine Triphosphate; Animals; Antithrombin III; Disease Models, Animal; Inflammation; Interleukin-1; Lactates; Mice; Mice, Inbred C57BL; Muscle, Skeletal; Peptide Hydrolases; Peroxidase; Pyruvates; Reperfusion Injury; Thrombosis

Platelet and leukocyte products are involved in atherothrombosis. However, the determinants of platelet and leukocyte markers assessed by flow cytometry have not been documented in a population-based sample.. We performed flow cytometry on blood from participants (n=1894) in the Atherosclerosis Risk in Communities (ARIC) Carotid MRI Study. Cellular aggregates and multiple platelet and leukocyte markers, such as myeloperoxidase in granulocytes and toll-like receptor-4, CD14, and CD45 in monocytes, were quantified. Their cross-sectional associations with demographic and risk factors were assessed using multiple linear regression. Mean values of most cellular markers and aggregates were considerably higher in blacks than whites (p<0.01). There were some differences in cellular markers between men and women, but little association with age. LDL-cholesterol was associated positively with several markers (toll-like receptor-4 and myeloperoxidase in granulocytes and CD162 in lymphocytes). Cholesterol-lowering therapy tended to show opposite associations. Smokers had much higher granulocyte myeloperoxidase than nonsmokers. However, most other correlations between risk factors and cellular markers were nonsignificant.. Race/ethnicity, sex, and to a lesser degree LDL-cholesterol and cholesterol-lowering therapy, but few other risk factors, were correlated with markers of cellular activation in this population-based study.

Topics: Blood Platelets; Cholesterol, LDL; Female; Flow Cytometry; Granulocytes; Humans; Leukocytes; Lymphocytes; Male; Membrane Glycoproteins; Peroxidase; Risk; Risk Factors; Thrombosis; Toll-Like Receptor 4

Activation of extravascular coagulation has been reported in acute lung injury models of sepsis and acute respiratory distress syndrome. Thrombin, the main effector protease of extravascular coagulation, activates proinflammatory cell types, including macrophages, endothelial cells, and neutrophils, each of which participates in lung ischemia-reperfusion injury. We used hirudin, a potent, specific direct thrombin inhibitor, to define the role of thrombin in lung ischemia-reperfusion injury.. Rats were pretreated with hirudin 30 minutes before warm, in situ left lung ischemia and reperfusion. Multiple in vivo assessments of lung injury were determined, and mechanistic studies assessed transcriptional regulation early in reperfusion and proinflammatory protein secretion late in reperfusion. Immunohistochemistry localized thrombin activation.. Thrombin localized to macrophages and endothelial and epithelial cells early in reperfusion. Hirudin significantly limited lung ischemia-reperfusion injury-induced derangements in vascular permeability and intraalveolar inflammatory cell sequestration, resulting in improved arterial oxygenation after ischemia and 4 hours of reperfusion. The protection was transcriptionally mediated by attenuated activator protein-1 and early growth response-1 transactivation, but not nuclear factor kappa B transactivation. This was associated with reduced chemokine, but not tumor necrosis factor alpha, secretion late in reperfusion.. Thrombin promotes lung ischemia-reperfusion injury, as hirudin protected against experimental acute lung injury. Hirudin conferred protection through a mechanism independent of nuclear factor kappa B and tumor necrosis factor alpha, suggesting that its effects may be mediated by a parallel, synergistic inflammatory pathway through activator protein-1 and early growth response-1.

Topics: Animals; Capillary Permeability; Disease Models, Animal; Humans; Inflammation; Ischemia; Peroxidase; Pulmonary Circulation; Rats; Rats, Long-Evans; Reperfusion; Reperfusion Injury; Respiratory Function Tests; Thrombosis

Recombinant human G-CSF (rHuG-CSF) is used for hematopoietic progenitor cells (HPC) mobilization and collection. Activation of polymorphonuclear leukocytes (PMN) is present during rHuG-CSF treatment and is associated with endothelial cell dysfunction and hypercoagulation. We evaluated whether PMN activation by rHuG-CSF may alter the blood oxidative status and subsequently affect the vascular cell function. Fourteen healthy individuals received rHuG-CSF for HPC harvesting. Blood was drawn before starting rHuG-CSF (T0), on the last day of rHuG-CSF (T1) and 1 week after stopping rHuG-CSF (T2). Levels of CD11b, myeloperoxidase (MPO), hydroperoxides, nitric oxide (NO), and soluble endothelium (sES), leukocyte (sLS), and platelet (sPS) selectins were measured. During rHuG-CSF, CD11b, MPO and hydroperoxides significantly increased, while NO levels significantly decreased, compared with T0. At T2 all these markers returned to baseline values. Significant increments of all selectins were observed during rHuG-CSF. At T2 sES and sEP significantly decreased back to pre-treatment values, whereas sLS remained significantly high. These data show that rHuG-CSF induces a transient inflammatory status characterized by circulating activated PMN, which release reactive oxygen species and intracellular proteases, promoting the onset of an abnormal oxidative status. This process may modify the hemostatic balance towards a pro-thrombotic state.

Topics: Adolescent; Adult; Aged; Blood Donors; CD11b Antigen; Child; Female; Granulocyte Colony-Stimulating Factor; Hematopoietic Stem Cell Mobilization; Humans; Hydrogen Peroxide; Male; Middle Aged; Neutrophil Activation; Nitric Oxide; Peroxidase; Recombinant Proteins; Selectins; Thrombosis

Reperfusion injury and hepatic artery thrombosis are major causes of graft failure after liver transplantation. The magnitude of oxidative stress increases after reperfusion and the appearance of an arterial thrombosis presents a higher risk for the graft and patient survival.. The aim of the study was to detect the level of oxidative stress in the perioperative period of transplantation.. Clinical documentations of 32 patients were investigated and the level of myeloperoxidase (MPO) was measured for the monitoring of the oxidative stress.. The mean age of the patients was 43 years and hepatitis C cirrhosis was the most common indication (14 cases, 43%). Two retransplantations were done. In 24 cases (75%) the primary graft functions and patient survival were good. Eight patients died, in two cases because of acute liver failure, in two cases due to primary non function and in four cases due to late complications. The incidence of hepatic artery thrombosis was 11% (4 cases) and the incidence of acute rejection was 35% (12 cases). The level of MPO was higher (65 ng/ml) in all patients before operation. After the first 48 hours this level increased significantly (p < 0.0001) up to the mean level of 123 ng/ml and decreased after one week. In the cases with acute liver failure and hepatic artery thrombosis high levels of MPO were measured.. This study provides evidence of increased oxidative stress before liver transplantation. The magnitude of these changes increased after operation, mostly in cases with acute liver failure and hepatic artery thrombosis. Reducing the reperfusion injury and performing an "ideal" arterial supply for the liver-graft present better survival.

Topics: Adolescent; Adult; Child; Female; Graft Rejection; Hepatic Artery; Humans; Hungary; Incidence; Liver; Liver Diseases; Liver Failure, Acute; Liver Transplantation; Male; Middle Aged; Oxidative Stress; Peroxidase; Prospective Studies; Thrombosis; Time Factors

Platelet-activating factor (PAF) is involved in many pathologic conditions through its potent proinflammatory and vasoactive effects. Using a specific PAF antagonist, SM-12502, we investigated the role of PAF in rat experimental glomerular thrombosis. In this model, sequential injections of nephrotoxic serum (NTS) and lipopolysaccharide (LPS) selectively induce glomerular fibrin deposition accompanied by neutrophil accumulation. SM-12502, when injected simultaneously with either NTS or LPS, strongly inhibited glomerular fibrin deposition in a dose-dependent manner. In contrast, neutrophil invasion was similar in both SM-12502-injected and uninjected rats, suggesting that the antithrombotic effect was not mediated by inhibition of neutrophil migration. However, serum myeloperoxidase activity, a marker of neutrophil activation, was significantly suppressed by treatment with SM-12502. From a previous finding supporting the indispensable role of neutrophils in this model and the current observations, SM-12502 is suggested to attenuate glomerular thrombosis by inhibiting neutrophil activation. Thus, the present findings suggest an involvement of PAF in this glomerular thrombosis model.

Topics: Animals; Fibrin; Kidney Glomerulus; Lipopolysaccharides; Male; Neutrophil Activation; Peroxidase; Platelet Activating Factor; Rats; Rats, Wistar; Thiazoles; Thiazolidines; Thrombosis

The present investigation was undertaken to study the alterations in free radical generation and release of nitric oxide (NO) from polymorphonuclear leukocytes (PMNLs) following thrombosis. Thrombosis was induced in rats by intravenous injection of collagen and adrenaline. PMNLs were separated from rat blood by using dextran sedimentation and Ficoll-Hypaque. Arachidonic acid (AA), formyl methionine leucine phenylalanine (FMLP) and opsonized zymosan (OZ) induced free radical generation was estimated as luminol (LCL) and Lucigenin (LUCDCL) dependent chemiluminescence. PMNLs nitric oxide synthase (NOS) activity and NO release were measured by using [14C] L-Arginine (L-Arg) and oxy-hemoglobin respectively. LCL and LUCDCL responses in rat PMNLs were significantly attenuated following thrombosis. There was no change in the release of myeloperoxidase enzyme (MPO) from PMNLs obtained following thrombosis. PMNLs NOS activity and NO release were also found to be increased after thrombosis. Pretreatment of rat PMNLs with 10 mM L-NAME (NO precursor) or 100 microM sodium nitroprusside (NO donor), resulted in significant reduction of AA induced LCL response. Results obtained indicate that NO release form PMNLs was augmented while free radical generation response was attenuated after the induction of thrombosis.

Topics: Animals; Arachidonic Acid; Cell Hypoxia; Cells, Cultured; Collagen; Epinephrine; Free Radicals; Luminescent Measurements; Male; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitroprusside; Peroxidase; Rats; Rats, Sprague-Dawley; Thrombosis; Zymosan

This solid-phase colorimetric microtiter-plate clotting assay is much more sensitive than standard clotting tests. In enzyme-linked coagulation assay (ELCA), enzyme-labeled fibrinogen and solid-phase fibrinogen are the substrate for thrombin generated in the clotting cascade. We used this assay to measure the factors of the extrinsic pathway by an extrinsic pathway-specific assay (EP-ELCA) and to determine the individual factors of the extrinsic pathway (VII, X, V, II) in plasmas of coumadin-treated and heparin-treated patients, with prothrombin time (PT) values used as a reference. In the ELCA method, samples and controls are incubated on the same plate, eliminating the requirement for pre-standardization of the substrate "plasma" before the factor assay is done. Concentrations of factors are determined by serially diluting sample and control plasmas to yield equivalent activity at given dilutions, a more direct approach for measuring specific factors than determining log concentrations vs log clotting time. Changes in the concentrations of clotting factors are seen before changes are apparent by PT. For coumadin-treated patients, all vitamin K-dependent factors were significantly (P less than or equal to 0.001) less than in normal controls, whereas factor V concentrations were normal, as expected. For patients treated with heparin, concentrations of factors X and VII were less than in normal controls (P less than 0.01) and results for EP-ELCA, II, and V assays were normal. This methodology can readily be automated.

Topics: Adult; Aged; Aged, 80 and over; Blood Coagulation Factors; Blood Coagulation Tests; Disseminated Intravascular Coagulation; Factor V; Factor VII; Factor X; Fibrinogen; Heparin; Humans; Male; Peroxidase; Prothrombin; Prothrombin Time; Thrombosis; Vitamin K; Warfarin

Using an automated cytochemical analyzer used for routine differential counts, we have been able to demonstrate acquired myeloperoxidase deficiency in 102 patients at our institution. Clinical and laboratory data on these patients showed a high incidence of diabetes mellitus (25.5%) and thrombotic diseases (24.5%), as well as a strikingly constant hyperfibrinogenemia (mean = 635 mg/100 ml; range = 360-1015 mg/100 ml). In 4 additional acute leukemia patients in complete remission, a close time correlation was noted between acquired MPO deficiency, diffuse intravascular coagulation and relapse. These findings indicate the importance of the relationships between neutrophil granulocytes and blood coagulation, and suggest that similar changes in neutrophil MPO activity may represent an early morphological indicator of subclinical activation of blood coagulation.

Topics: Adolescent; Adult; Aged; Blood Coagulation; Child; Child, Preschool; Diabetes Mellitus; Disseminated Intravascular Coagulation; Female; Fibrinogen; Humans; Infant; Infant, Newborn; Leukemia, Myeloid, Acute; Male; Middle Aged; Neutrophils; Peroxidase; Peroxidases; Thrombosis