RESUMEN
BACKGROUND: Inflammation is a prominent clinical manifestation in type 2 diabetes mellitus (T2DM) patients, often associated with insulin resistance, metabolic dysregulation, and other complications. AIM OF THE STUDY: The present study has been designed to check the serum levels of PAR-1 and correlate with various clinical manifestations and inflammatory cytokines levels in type 2 diabetic subjects. MATERIAL AND METHODS: The study population was divided into two groups, healthy volunteers (n = 15): normal glycated hemoglobin (HbA1c) (4.26 ± 0.55) and type 2 diabetic subjects (n = 30): HbA1c levels (7.80 ± 2.41). The serum levels of PAR-1 (ELISA method) were studied in both groups and correlated with demographic parameters age, weight, body mass index (BMI), and conventional inflammation biomarkers like C-reactive protein (CRP), interleukin 6 (IL-6), interleukin 8 (IL-8), and tumour necrosis factor-alpha (TNF-α). RESULTS: The demographic variables including the body weight (77.38 ± 10.00 vs. controls 55.26 ± 6.99), BMI (29.39 ± 3.61 vs. controls 25.25 ± 4.01), glycemic index HbA1c (7.80 ± 2.41 vs. controls 4.26 ± 0.55) were found to be statistically increased in T2DM subjects than the healthy control group. The levels of various inflammatory biomarkers and PAR-1 were significantly elevated in T2DM groups in comparison to healthy volunteers. The univariate and multivariate regression analysis revealed that elevated PAR-1 levels positively correlated with increased body weight, BMI, HbA1c, and inflammatory cytokines. CONCLUSION: Our findings indicate that the elevated serum PAR-1 levels serve as an independent predictor of inflammation in T2DM subjects and might have prognostic value for determining T2DM progression.
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Diabetes Mellitus Tipo 2 , Receptor PAR-1 , Factor de Necrosis Tumoral alfa , Biomarcadores , Glucemia/metabolismo , Peso Corporal , Proteína C-Reactiva/análisis , Citocinas , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/diagnóstico , Hemoglobina Glucada/análisis , Hemoglobina Glucada/metabolismo , Humanos , Inflamación/complicaciones , Inflamación/diagnóstico , Interleucina-6 , Interleucina-8 , Receptor PAR-1/sangreRESUMEN
OBJECTIVE: The platelet phenotype in certain patients and clinical contexts may differ from healthy conditions. We evaluated platelet activation through specific receptors in healthy men and women, comparing this to patients presenting with ST-segment-elevation myocardial infarction and non-ST-segment-elevation myocardial infarction. Approach and Results: We identified independent predictors of platelet activation through certain receptors and a murine MI model further explored these findings. Platelets from healthy women and female mice are more reactive through PARs (protease-activated receptors) compared with platelets from men and male mice. Multivariate regression analyses revealed male sex and non-ST-segment-elevation myocardial infarction as independent predictors of enhanced PAR1 activation in human platelets. Platelet PAR1 signaling decreased in women and increased in men during MI which was the opposite of what was observed during healthy conditions. Similarly, in mice, thrombin-mediated platelet activation was greater in healthy females compared with males, and lesser in females compared with males at the time of MI. CONCLUSIONS: Sex-specific signaling in platelets seems to be a cross-species phenomenon. The divergent platelet phenotype in males and females at the time of MI suggests a sex-specific antiplatelet drug regimen should be prospectively evaluated.
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Plaquetas/metabolismo , Infarto del Miocardio sin Elevación del ST/sangre , Activación Plaquetaria , Receptor PAR-1/sangre , Infarto del Miocardio con Elevación del ST/sangre , Anciano , Animales , Plaquetas/efectos de los fármacos , Estudios de Casos y Controles , Modelos Animales de Enfermedad , Femenino , Humanos , Masculino , Ratones Endogámicos C57BL , Persona de Mediana Edad , Fenotipo , Activación Plaquetaria/efectos de los fármacos , Factores Sexuales , Transducción de Señal , Trombina/farmacologíaRESUMEN
We tested a novel hypothesis that elevated levels of proteases in the maternal circulation of preeclamptic women activate neutrophils due to their pregnancy-specific expression of protease-activated receptor 1 (PAR-1). Plasma was collected longitudinally from normal pregnant and preeclamptic women and analyzed for MMP-1 and neutrophil elastase. Neutrophils were isolated for culture and confocal microscopy. Omental fat was collected for immunohistochemistry. Circulating proteases were significantly elevated in preeclampsia. Confocal microscopy revealed that tet methylcytosine dioxygenase 2 (TET2), a DNA de-methylase, and p65 subunit of NF-κB were strongly localized to the nucleus of untreated neutrophils of preeclamptic women, but in untreated neutrophils of normal pregnant women they were restricted to the cytosol. Treatment of normal pregnancy neutrophils with proteases activated PAR-1, leading to activation of RhoA kinase (ROCK), which triggered translocation of TET2 and p65 from the cytosol into the nucleus, mimicking the nuclear localization in neutrophils of preeclamptic women. IL-8, an NF-κB-regulated gene, increased in association with TET2 and p65 nuclear localization. Co-treatment with inhibitors of PAR-1 or ROCK prevented nuclear translocation and IL-8 did not increase. Treatment of preeclamptic pregnancy neutrophils with inhibitors emptied the nucleus of TET2 and p65, mimicking the cytosolic localization of normal pregnancy neutrophils. Expression of PAR-1 and TET2 were markedly increased in omental fat vessels and neutrophils of preeclamptic women. We conclude that elevated levels of circulating proteases in preeclamptic women activate neutrophils due to their pregnancy-specific expression of PAR-1 and speculate that TET2 DNA de-methylation plays a role in the inflammatory response.
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Epigénesis Genética , Neutrófilos/metabolismo , Péptido Hidrolasas/sangre , Preeclampsia/sangre , Transducción de Señal , Adulto , Proteínas de Unión al ADN/sangre , Dioxigenasas , Femenino , Humanos , Elastasa de Leucocito/sangre , Estudios Longitudinales , Metaloproteinasa 1 de la Matriz/sangre , FN-kappa B/sangre , Preeclampsia/genética , Embarazo , Proteínas Proto-Oncogénicas/sangre , Receptor PAR-1/sangreRESUMEN
OBJECTIVE: Protease-activated receptor-1 (PAR1) is classically activated by thrombin and is critical in controlling the balance of hemostasis and thrombosis. More recently, it has been shown that noncanonical activation of PAR1 by matrix metalloprotease-1 (MMP1) contributes to arterial thrombosis. However, the role of PAR1 in long-term development of atherosclerosis is unknown, regardless of the protease agonist. APPROACH AND RESULTS: We found that plasma MMP1 was significantly correlated (R=0.33; P=0.0015) with coronary atherosclerotic burden as determined by angiography in 91 patients with coronary artery disease and acute coronary syndrome undergoing cardiac catheterization or percutaneous coronary intervention. A cell-penetrating PAR1 pepducin, PZ-128, currently being tested as an antithrombotic agent in the acute setting in the TRIP-PCI study (Thrombin Receptor Inhibitory Pepducin-Percutaneous Coronary Intervention), caused a significant decrease in total atherosclerotic burden by 58% to 70% (P<0.05) and reduced plaque macrophage content by 54% (P<0.05) in apolipoprotein E-deficient mice. An MMP1 inhibitor gave similar beneficial effects, in contrast to the thrombin inhibitor bivalirudin that gave no improvement on atherosclerosis end points. Mechanistic studies revealed that inflammatory signaling mediated by MMP1-PAR1 plays a critical role in amplifying tumor necrosis factor α signaling in endothelial cells. CONCLUSIONS: These data suggest that targeting the MMP1-PAR1 system may be effective in tamping down chronic inflammatory signaling in plaques and halting the progression of atherosclerosis.
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Enfermedades de la Aorta/enzimología , Aterosclerosis/enzimología , Enfermedades de las Arterias Carótidas/enzimología , Enfermedad de la Arteria Coronaria/enzimología , Metaloproteinasa 13 de la Matriz/metabolismo , Metaloproteinasa 1 de la Matriz/metabolismo , Receptor PAR-1/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Animales , Enfermedades de la Aorta/patología , Enfermedades de la Aorta/prevención & control , Aterosclerosis/patología , Aterosclerosis/prevención & control , Biomarcadores/sangre , Enfermedades de las Arterias Carótidas/patología , Enfermedades de las Arterias Carótidas/prevención & control , Línea Celular , Péptidos de Penetración Celular/farmacología , Ensayos Clínicos Fase II como Asunto , Enfermedad de la Arteria Coronaria/sangre , Enfermedad de la Arteria Coronaria/patología , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Femenino , Fibrinolíticos/farmacología , Células Endoteliales de la Vena Umbilical Humana/enzimología , Humanos , Ácidos Hidroxámicos/farmacología , Lipopéptidos/farmacología , Masculino , Metaloproteinasa 1 de la Matriz/sangre , Inhibidores de la Metaloproteinasa de la Matriz/farmacología , Ratones Endogámicos C57BL , Ratones Noqueados para ApoE , Persona de Mediana Edad , Estudios Multicéntricos como Asunto , Oligopéptidos/farmacología , Placa Aterosclerótica , Ensayos Clínicos Controlados Aleatorios como Asunto , Receptor PAR-1/antagonistas & inhibidores , Receptor PAR-1/sangre , Transducción de Señal , Factor de Necrosis Tumoral alfa/sangre , Estados UnidosRESUMEN
Activated platelets generate an eicosanoid proposed to be 8-hydroxy-9,10-dioxolane A3 (DXA3). Herein, we demonstrate that significant amounts of DXA3 are rapidly attached to phosphatidylethanolamine (PE) forming four esterified eicosanoids, 16:0p, 18:0p, 18:1p and 18:0a/DXA3-PEs that can activate neutrophil integrin expression. These lipids comprise the majority of DXA3 generated by platelets, are formed in ng amounts (24.3±6.1ng/2×108) and remain membrane bound. Pharmacological studies revealed DXA3-PE formation involves cyclooxygenase-1 (COX), protease-activated receptors (PAR) 1 and 4, cytosolic phospholipase A2 (cPLA2), phospholipase C and intracellular calcium. They are generated primarily via esterification of newly formed DXA3, but can also be formed in vitro via co-oxidation of PE during COX-1 co-oxidation of arachidonate. All four DXA3-PEs were detected in human clots. Purified platelet DXA3-PE activated neutrophil Mac-1 expression, independently of its hydrolysis to the free eicosanoid. This study demonstrates the structures and cellular synthetic pathway for a family of leukocyte-activating platelet phospholipids generated on acute activation, adding to the growing evidence that enzymatic PE oxidation is a physiological event in innate immune cells.
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Plaquetas/metabolismo , Dioxolanos/sangre , Integrinas/sangre , Lípidos/sangre , Fosfatidiletanolaminas/sangre , Calcio/sangre , Ciclooxigenasa 1/sangre , Eicosanoides/sangre , Regulación de la Expresión Génica , Humanos , Integrinas/biosíntesis , Antígeno de Macrófago-1/genética , Neutrófilos/metabolismo , Oxidación-Reducción , Fosfolipasas A2 Citosólicas/sangre , Activación Plaquetaria/genética , Receptor PAR-1/sangre , Receptores de Trombina/sangre , Trombina/metabolismo , Fosfolipasas de Tipo C/sangreRESUMEN
Essentials Roles of the two thrombin receptors in platelet signaling are poorly understood. Computational systems biology modeling was used together with continuous flow cytometry. Dual-receptor system has wide-range sensitivity to thrombin and optimal response dynamics. Procoagulant platelet formation is determined by donor-specific activities of the two receptors. SUMMARY: Background Activation of human platelets with thrombin proceeds via two protease-activated receptors (PARs), PAR1 and PAR4, that have identical main intracellular signaling responses. Although there is evidence that they have different cleavage/inactivation kinetics (and some secondary variations in signaling), the reason for such redundancy is not clear. Methods We developed a multicompartmental stochastic computational systems biology model of dual-receptor thrombin signaling in platelets to gain insight into the mechanisms and roles of PAR1 and PAR4 functioning. Experiments employing continuous flow cytometry of washed human platelets were used to validate the model and test its predictions. Activity of PAR receptors in donors was evaluated by mRNA measurement and by polymorphism sequencing. Results Although PAR1 activation produced rapid and short-lived response, signaling via PAR4 developed slowly and propagated in time. Response of the dual-receptor system was both rapid and prolonged in time. Inclusion of PAR1/PAR4 heterodimer formation promoted PAR4 signaling in the medium range of thrombin concentration (about 10 nm), with little contribution at high and low thrombin. Different dynamics and dose-dependence of procoagulant platelet formation in healthy donors was associated with individual variations in PAR1 and PAR4 activities and particularly by the Ala120Thr polymorphism in the F2RL3 gene encoding PAR4. Conclusions The dual-receptor combination is critical to produce a response combining three critical advantages: sensitivity to thrombin concentration, rapid onset and steady propagation; specific features of the protease-activated receptors do not allow combination of all three in a single receptor.
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Plaquetas/metabolismo , Activación Plaquetaria , Biología de Sistemas , Trombina/metabolismo , Adolescente , Adulto , Animales , Plaquetas/citología , Simulación por Computador , Dimerización , Femenino , Humanos , Cinética , Masculino , Agregación Plaquetaria , Polimorfismo Genético , Receptor PAR-1/sangre , Receptores de Trombina/sangre , Receptores de Trombina/genética , Transducción de Señal , Adulto JovenRESUMEN
Despite the use of therapies recommended in practice guidelines for secondary prevention in patients with atherosclerotic coronary artery disease, the residual risk for cardiovascular events remains high. Some of the residual risk is believed to result from incomplete platelet inhibition with current therapy. Vorapaxar is a first-in-class, novel antiplatelet agent that acts by antagonizing the PAR-1 receptor, inhibiting thrombin-mediated platelet activation. Vorapaxar was recently approved by the Food and Drug Administration for secondary prevention of cardiovascular events in patients with a history of myocardial infarction or peripheral artery disease who do not have a history of transient ischemic attack or stroke. We review the data from two key phase III cardiovascular outcome trials with vorapaxar: TRACER and TRA 2P-TIMI 50. We will focus on identifying the key patient populations that should be identified for treatment, highlight practical clinical issues when prescribing vorapaxar, and review unanswered questions. Vorapaxar should be considered in patients at high risk for recurrent ischemic events and low risk of bleeding.
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Plaquetas/efectos de los fármacos , Enfermedad de la Arteria Coronaria/tratamiento farmacológico , Lactonas/uso terapéutico , Infarto del Miocardio/prevención & control , Inhibidores de Agregación Plaquetaria/uso terapéutico , Piridinas/uso terapéutico , Receptor PAR-1/antagonistas & inhibidores , Prevención Secundaria/métodos , Animales , Plaquetas/metabolismo , Enfermedad de la Arteria Coronaria/sangre , Enfermedad de la Arteria Coronaria/complicaciones , Hemorragia/inducido químicamente , Humanos , Lactonas/efectos adversos , Infarto del Miocardio/sangre , Infarto del Miocardio/etiología , Selección de Paciente , Placa Aterosclerótica , Inhibidores de Agregación Plaquetaria/efectos adversos , Piridinas/efectos adversos , Receptor PAR-1/sangre , Recurrencia , Medición de Riesgo , Factores de Riesgo , Transducción de Señal/efectos de los fármacos , Resultado del TratamientoRESUMEN
This article reviews the pharmacology, clinical efficacy, and safety of vorapaxar in reducing cardiovascular risk. Vorapaxar is a tricyclic himbacine-derived reversible inhibitor of platelet surface protease activator receptor-1, which prevents thrombin from activating platelets. Two Phase III clinical trials and multiple subanalyses from the two trials with vorapaxar have been published. In patients with recent acute coronary syndrome, vorapaxar, when added to standard therapy, did not reduce the composite cardiovascular end point. In contrary, in a study of secondary prevention for patients with cardiovascular diseases, vorapaxar reduced the risk of cardiovascular death or ischemic events (myocardial infarction, stroke) in patients with stable atherosclerosis who were receiving standard therapy. Vorapaxar is approved in the US for use with aspirin and/or clopidogrel in the secondary prevention of thrombogenic cardiovascular events in stable patients with peripheral arterial disease or a history of myocardial infarction. Vorapaxar increases risk of bleeding and is contraindicated in patients with previous cerebrovascular events. It is essential to balance individual patient's bleeding risk to any further cardiovascular benefits that they may get. Future investigation is also needed to evaluate use of vorapaxar with newer antiplatelet agents such as ticagrelor and cangrelor, as well as its role as monotherapy.
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Plaquetas/efectos de los fármacos , Enfermedades Cardiovasculares/prevención & control , Lactonas/uso terapéutico , Inhibidores de Agregación Plaquetaria/uso terapéutico , Agregación Plaquetaria/efectos de los fármacos , Piridinas/uso terapéutico , Receptor PAR-1/antagonistas & inhibidores , Animales , Plaquetas/metabolismo , Enfermedades Cardiovasculares/sangre , Enfermedades Cardiovasculares/etiología , Quimioterapia Combinada , Hemorragia/inducido químicamente , Humanos , Lactonas/efectos adversos , Selección de Paciente , Inhibidores de Agregación Plaquetaria/efectos adversos , Piridinas/efectos adversos , Receptor PAR-1/sangre , Medición de Riesgo , Factores de Riesgo , Resultado del TratamientoRESUMEN
OBJECTIVE: Protease-activated receptors (PAR) are G protein coupled receptors and they regulate many biological processes, including coagulation and cell survival and they might be good markers in some types of malignant tumors, providing useful information in diagnosis and prognosis. The objective of this study was to determine the clinical significance of the serum levels of PAR1 in lung cancer patients. PATIENTS AND METHODS: Eighty patients with lung cancer were enrolled into this study. Serum PAR1 levels were determined by the solid-phase sandwich ELISA method. Median age was 58.5-years old, range 36 to 80 years. RESULTS: The majority of the patients had NSCLC (85%) and stage IV disease (56%). The baseline serum PAR1 concentrations of the lung cancer patients were significantly higher than control group (median values 26.45 ng/mL v 0.07 ng/mL, p < 0.001). However, clinical variables including age, gender, histology, stage of disease, and response to chemotherapy were not found to be correlated with serum PAR1 levels (p > 0.05). Moreover, it failed to show any prognostic value on the survival of the lung cancer patients. CONCLUSIONS: The serum levels of PAR1 might have a diagnostic value in lung cancer patients. However, its predictive and prognostic values were not determined.
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Carcinoma de Pulmón de Células no Pequeñas/sangre , Neoplasias Pulmonares/sangre , Receptor PAR-1/sangre , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/sangre , Carcinoma de Pulmón de Células no Pequeñas/diagnóstico , Estudios de Casos y Controles , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Neoplasias Pulmonares/diagnóstico , Masculino , Persona de Mediana Edad , PronósticoRESUMEN
Exocytosis of lysosomal contents from platelets has been speculated to participate in clearance of thrombi and vessel wall remodelling. The mechanisms that regulate lysosomal exocytosis in platelets are, however, still unclear. The aim of this study was to identify the pathways underlying platelet lysosomal secretion and elucidate how this process is controlled by platelet inhibitors. We found that high concentrations of thrombin induced partial lysosomal exocytosis as assessed by analysis of the activity of released N-acetyl-ß-glucosaminidase (NAG) and by identifying the fraction of platelets exposing the lysosomal-associated membrane protein (LAMP)-1 on the cell surface by flow cytometry. Stimulation of thrombin receptors PAR1 or PAR4 with specific peptides was equally effective in inducing LAMP-1 surface expression. Notably, lysosomal exocytosis in response to thrombin was significantly reduced if the secondary activation by ADP was inhibited by the P2Y12 antagonist cangrelor, while inhibition of thromboxane A2 formation by treatment with acetylsalicylic acid was of minor importance in this regard. Moreover, the NO-releasing drug S-nitroso-N-acetyl penicillamine (SNAP) or the cyclic AMP-elevating eicosanoid prostaglandin I2 (PGI2) significantly suppressed lysosomal exocytosis. We conclude that platelet inhibitors that mimic functional endothelium such as PGI2 or NO efficiently counteract lysosomal exocytosis. Furthermore, we suggest that secondary release of ADP and concomitant signaling via PAR1/4- and P2Y12 receptors is important for efficient platelet lysosomal exocytosis by thrombin.
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Adenosina Difosfato/sangre , Plaquetas/metabolismo , Acetilglucosaminidasa/sangre , Adenosina Difosfato/farmacología , Adenosina Monofosfato/análogos & derivados , Adenosina Monofosfato/farmacología , Plaquetas/efectos de los fármacos , Epoprostenol/sangre , Exocitosis/efectos de los fármacos , Humanos , Proteínas de Membrana de los Lisosomas/biosíntesis , Proteínas de Membrana de los Lisosomas/sangre , Lisosomas/efectos de los fármacos , Lisosomas/metabolismo , Antagonistas del Receptor Purinérgico P2Y/farmacología , Receptor PAR-1/sangre , Trombina/farmacologíaRESUMEN
Immune thrombocytopenia (ITP) patients with similarly low platelet counts differ in their tendency to bleed. To determine if differences in platelet function in ITP patients account for this variation in bleeding tendency, we conducted a single-center, cross-sectional study of pediatric patients with ITP. Bleeding severity (assessed by standardized bleeding score) and platelet function (assessed by whole blood flow cytometry) with and without agonist stimulation was evaluated in 57 ITP patients (median age, 9.9 years). After adjustment for platelet count, higher levels of thrombin receptor activating peptide (TRAP)-stimulated percent P-selectin- and activated glycoprotein (GP)IIb-IIIa-positive platelets were significantly associated with a lower bleeding score, whereas higher levels of immature platelet fraction (IPF), TRAP-stimulated platelet surface CD42b, unstimulated platelet surface P-selectin, and platelet forward light scatter (FSC) were associated with a higher bleeding score. Thus, platelet function tests related to platelet age (IPF, FSC) and activation through the protease activated receptor 1 (PAR1) thrombin receptor (TRAP-stimulated P-selectin, activated GPIIb-IIIa, and CD42b), independent of platelet count, are associated with concurrent bleeding severity in ITP. These tests may be useful markers of future bleeding risk in ITP.
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Hemorragia/sangre , Hemorragia/etiología , Recuento de Plaquetas , Pruebas de Función Plaquetaria , Púrpura Trombocitopénica Idiopática/sangre , Púrpura Trombocitopénica Idiopática/complicaciones , Adolescente , Plaquetas/patología , Plaquetas/fisiología , Diferenciación Celular , Micropartículas Derivadas de Células/fisiología , Niño , Preescolar , Estudios Transversales , Femenino , Citometría de Flujo , Humanos , Luz , Masculino , Volúmen Plaquetario Medio , Selectina-P/sangre , Fragmentos de Péptidos/sangre , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/metabolismo , Complejo GPIb-IX de Glicoproteína Plaquetaria/metabolismo , Receptor PAR-1/sangre , Dispersión de RadiaciónRESUMEN
BACKGROUND: The activation of pro-coagulation mechanisms associated with the vascular wall's immune and inflammatory responses wall to injury plays a crucial role in the mechanisms of the induction and progression of atherosclerosis. OBJECTIVES: The aim of this study was to determine the role of protease activated receptors (PAR-1) expressed on the surface of blood platelets in the pathogenesis of chronic peripheral arterial obliterative disease (PAOD) in patients with obliterative atherosclerosis (n = 24) and diabetic macroangiopathy (n = 16), as well as in the controls (n = 12). MATERIAL AND METHODS: In addition to the expression of PAR-1, serum/plasma concentrations of thrombin-antithrombin complex (TAT), the von Willebrand factor (vWF), the platelet-derived growth factor, monocyte chemotactic protein, the soluble form of the platelet endothelial cell adhesion molecule, thrombin activatable fibrinolysis inhibitor and interleukin 6 (IL-6) were determined. RESULTS: Compared to the controls, PAOD patients were characterized by significantly higher levels of PAR-1 expression, vWF, TAT and IL-6. Individuals with diabetic macroangiopathy did not differ significantly from individuals with obliterative atherosclerosis in terms of PAR-1 expression. Upon activation with thrombin receptor antagonist peptide (TRAP), the levels of PAR-1 were comparable in all analyzed groups. In patients with diabetic macroangiopathy, a significant association was observed between the expression of PAR-1 on the surface of the platelet and the serum TAT concentration, as well as between TAT and serum IL-6 concentration. CONCLUSIONS: Enhanced expression of PAR-1 on the thrombocyte surface in chronic PAOD patients occurs equally in cases of diabetic macroangiopathy and in individuals free from this endocrine pathology.
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Plaquetas/química , Enfermedad Arterial Periférica/sangre , Receptor PAR-1/sangre , Antitrombina III , Enfermedad Crónica , Humanos , Persona de Mediana Edad , Péptido Hidrolasas/sangre , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/análisis , Factor de von Willebrand/análisisRESUMEN
AIMS: Platelets are a natural source of growth factors, cytokines and chemokines, that regulate angiogenesis and inflammation. It has been suggested that differential release of pro- and anti-angiogenic growth factors from platelet α-granules by protease-activated receptors (PAR) 1 and 4 may be important for the regulation of angiogenesis. We aimed to compare the releasates of unstimulated platelets with PAR-1- and PAR-4-stimulated platelets. METHODS AND RESULTS: The release of ß-thromboglobulin, platelet factor (PF)-4, thrombospondin, platelet-derived growth factor (PDGF)-A/B, regulated and normal T-cell expressed and secreted (RANTES/CCL5), endostatin, CXCL12, and vascular endothelial growth factor (VEGF) was measured with enzyme-linked immunosorbent assay (ELISA). Mass spectrometry (MS)-based quantitative proteomics identified 93 proteins from platelets stimulated with PAR-1 and PAR-4. A strong correlation between the factors released after either stimulus was observed (Spearman's r 0.94, P < 0.001). Analysis with ELISA showed that stimulation with PAR-1 or PAR-4 lead to non-differential release of ß-thromboglobulin, PF-4, thrombospondin, PDGF-A/B, RANTES/CCL5, endostatin, CXCL12, and VEGF. Release of thrombospondin was slightly lower after PAR-1 stimulation (7.2 µg/mL), compared with PAR-4 induced release (9.8 µg/mL; P < 0.05). CONCLUSIONS: Both ELISA on established α-granule proteins and MS-based quantitative proteomics showed that the most abundant α-granule proteins are released in similar quantities from platelets after stimulation with either PAR-1 or PAR-4. Our findings provide evidence against the hypothesis that PAR-1 and PAR-4 stimulation of platelets trigger differential release of alpha-granule, but further studies are needed to draw conclusions for physiological conditions.
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Proteínas Reguladoras de la Apoptosis/sangre , Plaquetas/fisiología , Receptor PAR-1/sangre , Quimiocina CXCL12/sangre , Ensayo de Inmunoadsorción Enzimática , Fibrinógeno/metabolismo , Humanos , Espectrometría de Masas , Neovascularización Fisiológica , Selectina-P/sangre , Activación Plaquetaria/fisiología , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/metabolismo , Proteómica , Vesículas Secretoras/fisiología , Factor A de Crecimiento Endotelial Vascular/sangre , beta-Tromboglobulina/metabolismoRESUMEN
Platelet activation with subsequent aggregation is a complex process leading to thrombus formation, which remains a key component for atherothrombotic manifestations, in particular myocardial infarction. Therefore, antiplatelet therapies are pivotal for the treatment of these patients. Current oral antiplatelet therapies used for secondary prevention of ischemic recurrences include aspirin and adenosine diphosphate P2Y12 platelet-receptor antagonists. However, despite these therapies, patients who have experienced a myocardial infarction remain at risk for ischemic recurrences. Therefore, more aggressive secondary prevention measures have been an area of research, including identifying additional targets modulating platelet-activation and -aggregation processes. Among these, thrombin-mediated platelet activation via protease-activated receptors (PARs) has been subject to extensive clinical investigation. Several PAR-1 receptor antagonists have been developed. However, vorapaxar is the only one that has completed large-scale clinical investigation. The present manuscript will provide an overview on the role of thrombin-mediated signaling, the impact of PAR-1 blockade with vorapaxar on ischemic and bleeding outcomes, and the potential role for vorapaxar in clinical practice.
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Plaquetas/efectos de los fármacos , Lactonas/uso terapéutico , Infarto del Miocardio/tratamiento farmacológico , Inhibidores de Agregación Plaquetaria/uso terapéutico , Agregación Plaquetaria/efectos de los fármacos , Piridinas/uso terapéutico , Receptor PAR-1/antagonistas & inhibidores , Animales , Plaquetas/enzimología , Hemorragia/inducido químicamente , Humanos , Lactonas/efectos adversos , Lactonas/farmacocinética , Terapia Molecular Dirigida , Infarto del Miocardio/sangre , Infarto del Miocardio/diagnóstico , Infarto del Miocardio/enzimología , Inhibidores de Agregación Plaquetaria/efectos adversos , Inhibidores de Agregación Plaquetaria/farmacocinética , Piridinas/efectos adversos , Piridinas/farmacocinética , Receptor PAR-1/sangre , Factores de Riesgo , Prevención Secundaria , Transducción de Señal/efectos de los fármacos , Resultado del TratamientoRESUMEN
The role of molecular markers in ovarian cancer is still a matter of debate. Protease-activated receptor-1 (PAR1) might be a good marker in some types of malignant tumors and might provide useful information in diagnosis and prognosis. The objective of this study was to evaluate the serum levels of PAR1 in regard to diagnostic, predictive, and prognostic value in epithelial ovarian cancer (EOC) patients. Forty-four EOC patients were enrolled in this study. Serum PAR1 levels were determined by enzyme-linked immunosorbent assay (ELISA) method. Twenty-five age- and sex-matched healthy controls were included in the analysis. The median age of patients was 58 years old, ranging from 22 to 83 years, where most of them had advanced disease (stage III-IV) (n = 40, 91%). The median serum PAR1 values were significantly elevated in patients compared to healthy controls (1.52 ng/ml vs. 1.13 ng/ml) (p = 0.03), whereas any clinical variables including response to chemotherapy did not associate with serum assay (p > 0.05). Progression-free survival (PFS) and overall survival (OS) of patients who did not respond to chemotherapy nor had platinum resistance in relapsed disease were poorer in the analyses. On the other hand, serum PAR1 levels showed no significant adverse effect on either PFS or OS (p = 0.43 and p = 0.49, respectively). These results proved that baseline serum PAR1 levels of patients with EOC were significantly higher than those of healthy people. However, these assays suggested no predictive or prognostic value in this group of patients.
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Neoplasias Glandulares y Epiteliales/diagnóstico , Neoplasias Ováricas/diagnóstico , Receptor PAR-1/sangre , Adulto , Anciano , Anciano de 80 o más Años , Carcinoma Epitelial de Ovario , Supervivencia sin Enfermedad , Femenino , Humanos , Metaloproteinasa 1 de la Matriz/sangre , Persona de Mediana Edad , Clasificación del Tumor , Neoplasias Glandulares y Epiteliales/sangre , Neoplasias Glandulares y Epiteliales/mortalidad , Neoplasias Glandulares y Epiteliales/patología , Neoplasias Ováricas/sangre , Neoplasias Ováricas/mortalidad , Neoplasias Ováricas/patologíaRESUMEN
We studied the association of thrombin generation potential with platelet protease activated receptor (PAR)-1 regulation and platelet activation in 52 stable coronary artery disease patients on continuous therapy with aspirin and clopidogrel (n = 42) or prasugrel (n = 10). Compared to controls, peak thrombin generation potential was elevated in only 11 patients (p > 0.05), while F1.2 was elevated in 26 patients (p < 0.0001). PAR-1 and thrombin inducible P-selectin expression were significantly elevated in patients compared to controls (p < 0.001). There were no significant correlations between levels of thrombin generation potential or F1.2 and PAR-1 regulation. However, there was a significant inverse correlation between levels of peak thrombin generation potential and in vitro thrombin-inducible expression of P-selectin (p = 0.002), suggesting in vivo depletion of platelet alpha granules due to ongoing platelet activation.
Asunto(s)
Aspirina/administración & dosificación , Plaquetas/efectos de los fármacos , Enfermedad de la Arteria Coronaria/tratamiento farmacológico , Selectina-P/sangre , Piperazinas/administración & dosificación , Inhibidores de Agregación Plaquetaria/administración & dosificación , Receptor PAR-1/sangre , Tiofenos/administración & dosificación , Trombina/metabolismo , Ticlopidina/análogos & derivados , Adulto , Anciano , Biomarcadores/sangre , Plaquetas/metabolismo , Estudios de Casos y Controles , Clopidogrel , Enfermedad de la Arteria Coronaria/sangre , Enfermedad de la Arteria Coronaria/diagnóstico , Femenino , Humanos , Masculino , Persona de Mediana Edad , Fragmentos de Péptidos/sangre , Activación Plaquetaria/efectos de los fármacos , Clorhidrato de Prasugrel , Protrombina , Ticlopidina/administración & dosificaciónRESUMEN
Platelets are the key in the pathogenesis of atherothrombotic disease such as acute coronary syndromes, stroke, and peripheral arterial disease. Current anti-platelet treatments are mainly based on inhibition of two important pathways of platelet activation: thromboxane A2 (TXA2) mediated (aspirin) and adenosine diphosphate (ADP)-P2Y12 receptor mediated (clopidogrel, prasugrel, and ticagrelor). Despite the dual anti-platelet therapy with aspirin and P2Y12 inhibitors have reduced ischemic events in patients with acute coronary syndromes (ACS), the rate of recurrent ischemic complication after ACS remains high. Combination of multiple anti-platelet agents is also associated with increased risk of bleeding. Thrombin is a potent platelet agonist and the increase of its activity has been reported in patients with ACS. Platelet effects of thrombin are mediated by protease-activated receptors (PAR), and PAR-1 is the most important receptor in human platelets. Two PAR-1 antagonists, vorapaxar and atopaxar, have undergone clinical investigation. In this review, we will describe the pharmacology of PAR-1 antagonists and will review and discuss results of randomized clinical trials with PAR-1 antagonists.
Asunto(s)
Plaquetas/efectos de los fármacos , Enfermedades Cardiovasculares/tratamiento farmacológico , Inhibidores de Agregación Plaquetaria/uso terapéutico , Receptor PAR-1/antagonistas & inhibidores , Animales , Plaquetas/metabolismo , Enfermedades Cardiovasculares/sangre , Diseño de Fármacos , Humanos , Iminas/uso terapéutico , Lactonas/uso terapéutico , Ligandos , Activación Plaquetaria/efectos de los fármacos , Inhibidores de Agregación Plaquetaria/efectos adversos , Inhibidores de Agregación Plaquetaria/química , Piridinas/uso terapéutico , Receptor PAR-1/sangre , Transducción de Señal/efectos de los fármacos , Trombina/metabolismo , Resultado del TratamientoRESUMEN
Obesity is associated with a prothrombotic state resulting from increased thrombin generation, platelet hyper-reactivity, and decreased fibrinolysis. Data on the influence of obesity on clopidogrel-mediated platelet inhibition are conflicting and limited to platelet function tests. Moreover, there are no data on thrombin-inducible platelet activation in obese patients. We therefore investigated response to clopidogrel therapy and protease activated receptor (PAR)-1 mediated platelet activation in obese and nonobese patients undergoing angioplasty and stenting for cardiovascular disease. The vasodilator-stimulated phosphoprotein (VASP) phosphorylation assay, multiple electrode aggregometry (MEA) with adenosine diphosphate (ADP), and surface expressions of P-selectin and activated glycoprotein (GP) IIb/IIIa in response to ADP and thrombin receptor activating peptide (TRAP)-6 were assessed in 71 obese and 245 nonobese patients. Obesity was independently associated with higher residual platelet reactivity by the VASP assay and MEA ADP, and with platelet surface expressions of P-selectin and activated GPIIb/IIIa in response to ADP (all P ≤ 0.04). Further, high on-treatment residual ADP-inducible platelet reactivity by the VASP assay and by MEA ADP were significantly more frequent in obese patients compared with nonobese patients (both P ≤ 0.04). Finally, PAR-1 mediated platelet activation as assessed by expression of P-selectin and activated GPIIb/IIIa in response to TRAP-6 was significantly more pronounced in obese patients than in patients without obesity (both P ≤ 0.02). In conclusion, obese patients undergoing angioplasty and stenting exhibit a diminished response to clopidogrel and an increased susceptibility to TRAP-6 inducible platelet activation.
Asunto(s)
Obesidad/sangre , Obesidad/complicaciones , Inhibidores de Agregación Plaquetaria/farmacología , Receptor PAR-1/sangre , Trombosis/sangre , Trombosis/etiología , Ticlopidina/análogos & derivados , Adenosina Difosfato/farmacología , Anciano , Angioplastia/efectos adversos , Plaquetas/metabolismo , Estudios de Casos y Controles , Moléculas de Adhesión Celular/sangre , Clopidogrel , Femenino , Humanos , Masculino , Proteínas de Microfilamentos/sangre , Persona de Mediana Edad , Obesidad/tratamiento farmacológico , Selectina-P/sangre , Fosfoproteínas/sangre , Activación Plaquetaria/efectos de los fármacos , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/metabolismo , Stents/efectos adversos , Trombosis/prevención & control , Ticlopidina/farmacología , Investigación Biomédica TraslacionalRESUMEN
Dual antiplatelet therapy with aspirin and adenosine diphosphate (ADP) receptor inhibitors significantly improves the outcome of patients with stable coronary heart disease. However, abundant thrombin generation, which is not influenced by this dual antiplatelet therapy, is a major reason for recurrent thromboembolic disease in these patients. We, therefore, assessed in a hypothesis generating study in patients with stable coronary artery disease specifically the relation of responsiveness of the platelet thrombin receptor protease-activated receptor (PAR)-1 to the magnitude of the inhibition of the ADP receptor. PAR-1 regulation was studied prospectively in 86 consecutive patients with stable coronary artery disease treated with aspirin and clopidogrel (67 patients) or prasugrel (19 patients) and correlated the data to ADP inducible platelet reactivity by impedance aggregometry. PAR-1 expression did not differ between patients on aspirin and clopidogrel vs patients on aspirin and prasugrel (P > 0.5). PAR-1 levels were correlated to P-selectin expression (P < 0.0001). The higher the PAR-1 expression the more profound was the in vitro thrombin-inducible platelet activation. However, neither ex vivo PAR-1 expression nor in vitro thrombin-inducible PAR-1 were correlated to ADP-inducible platelet aggregation (P > 0.5). Thus, like in a real life scenario, patients with stable ischemic heart disease on dual antiplatelet therapy may express high levels of PAR-1, which are associated with profound thrombin-inducible platelet activation. This responsiveness cannot be predicted by the magnitude of ADP responsiveness.