RESUMO
BACKGROUND: Reactive oxygen species (ROS) contribute to platelet hyperactivation during aging. Several oxidative pathways and antioxidant enzymes have been implicated; however, their mechanistic contributions during aging remain elusive. We hypothesized that mitochondria are an important source of platelet ROS and that mitochondrial SOD2 (superoxide dismutase) protects against mitochondrial ROS-driven platelet activation and thrombosis during aging. METHODS: We studied littermates of platelet-specific SOD2-knockout (SOD2fl/flPf4Cre, pSOD2-KO) and control (SOD2fl/fl) mice at young (4-5 months) or old (18-20 months) ages. We examined agonist-induced platelet activation, platelet-dependent thrombin generation potential, and susceptibility to in vivo thrombosis. RESULTS: Platelet αIIbß3 activation, aggregation, and adhesion were increased to similar extents in aged mice of both genotypes compared with young mice. In contrast, the age-dependent increases in mitochondrial and total cellular ROS, calcium elevation, and phosphatidylserine exposure were augmented in platelets from pSOD2-KO mice compared with control mice. Aged pSOD2-KO mice showed increased platelet-dependent thrombin generation compared with aged control mice. In vivo, aged pSOD2-KO mice exhibited enhanced susceptibility to carotid artery and pulmonary thrombosis compared to aged control mice. Adoptive transfer of platelets from aged pSOD2-KO but not aged control mice increased thrombotic susceptibility in aged host mice, suggesting a prothrombotic effect of platelet pSOD2 deficiency. Treatment with avasopasem manganese (GC4419), a SOD mimetic, decreased platelet mitochondrial pro-oxidants, cellular ROS levels, and inhibited procoagulant platelet formation and arterial thrombosis in aged mice. CONCLUSIONS: Platelet mitochondrial ROS contributes to age-related thrombosis and endogenous SOD2 protects from platelet-dependent thrombin generation and thrombosis during aging.
Assuntos
Trombina , Trombose , Camundongos , Animais , Trombina/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Camundongos Knockout , Plaquetas/metabolismo , Trombose/genética , Trombose/prevenção & controle , Trombose/induzido quimicamente , Superóxido Dismutase/metabolismo , Superóxido Dismutase-1/metabolismo , Envelhecimento/metabolismoRESUMO
We present a case of heavy lone coronary thrombosis in the setting of COVID-19 infection. We highlight the special angiographic, ultrasonographic, and histological features of this thrombus, and we describe the application of carotid stent retriever for its removal.
Assuntos
COVID-19 , Trombose Coronária , Trombose Coronária/diagnóstico por imagem , Trombose Coronária/etiologia , Trombose Coronária/terapia , Vasos Coronários , Humanos , SARS-CoV-2 , Stents , Trombectomia , Resultado do TratamentoRESUMO
Platelet-derived extracellular vesicles (PEVs) play important roles in hemostasis and thrombosis. There are three major types of PEVs described based on their size and characteristics, but newer types may continue to emerge owing to the ongoing improvement in the methodologies and terms used to define various types of EVs. As the literature on EVs is growing, there are continuing attempts to standardize protocols for EV isolation and reach consensus in the field. This review provides information on mechanisms of PEV production, characteristics, cellular interaction, and their pathological role, especially in autoimmune and infectious diseases. We also highlight the mechanisms through which PEVs can activate parent cells in a feedback loop.
Assuntos
Vesículas Extracelulares , Trombose , Plaquetas , Hemostasia , HumanosRESUMO
Platelets are small, anucleated effector cells that play an important role in linking the hemostatic and inflammatory processes in the body. Platelet function is known to be altered under various inflammatory conditions including aging. A gain in platelet function during aging can increase the risk of thrombotic events, such as stroke and acute myocardial infarction. Anti-platelet therapy is designed to reduce risk of serious cerebrovascular and cardiovascular events, but the adverse consequences of therapy, such as risk for bleeding increases with aging as well. Age-associated comorbidities such as obesity, diabetes, and hyperlipidemia also contribute to increased platelet activity and thus can enhance the risk of thrombosis. Therefore, identification of unique mechanisms of platelet dysfunction in aging and in age-associated comorbidities is warranted to design novel antiplatelet drugs. This review outlines some of the current areas of research on aging-related mechanisms of platelet hyperactivity and addresses the clinical urgency for designing anti-platelet therapies toward novel molecular targets in the aging population.
Assuntos
Envelhecimento/efeitos dos fármacos , Transtornos Plaquetários/tratamento farmacológico , Plaquetas/efeitos dos fármacos , Ativação Plaquetária/efeitos dos fármacos , Trombose/complicações , Idoso , Envelhecimento/imunologia , Envelhecimento/metabolismo , Animais , Plaquetas/imunologia , Plaquetas/metabolismo , Comorbidade , Humanos , Inflamação/imunologia , Estresse Oxidativo , Ativação Plaquetária/imunologia , Fatores de Risco , Transdução de Sinais/genética , Trombose/fisiopatologiaRESUMO
INTRODUCTION: Rural and critical access hospitals rely on the "drip and ship" practice using helicopter emergency medical services (HEMS). But those helicopter flights are an unusual environment with physical factors such as vibration and accelerations that could potentially affect the stability, and pharmacological properties of IV rtPA, an issue that has not been previously addressed. MATERIALS AND METHODS: This was a prospective cohort study of consecutive acute ischemic stroke patients receiving IV rtPA through a Comprehensive Stroke Center from November 2015 to February 2017 to measure the effects of HEMS on the integrity and activity of rtPA by collecting residual medication left in the vial. CLINICAL TRIAL REGISTRATION: http://www.clinicaltrials.gov. NCT02752256 RESULTS: A total of 33 patients and rtPA samples were included; 18 patients who presented directly to the Comprehensive Stroke Center emergency department and 15 patients who received rtPA during air ambulance transfer. The median rtPA antigen concentration in the residual medication vial was 3.04 mg/mL (IQR: 1.24-3.87) in the HEMS group and 1.91 mg/mL (IQR: 1.33-2.60) in the controls (P = .168). There were no significant differences in rtPA activity or specific activity between the HEMS and control groups and there was no association between total HEMS flight time on overall rtPA specific activity. CONCLUSIONS: In summary, this study provides supportive evidence of the lack of a detrimental effect of the HEMS physical environment on the integrity of rtPA, therefore endorsing current drip and ship practices without infusion adjustments.
Assuntos
Resgate Aéreo , Isquemia Encefálica/tratamento farmacológico , Serviços Médicos de Emergência/métodos , Fibrinolíticos/administração & dosagem , Acidente Vascular Cerebral/tratamento farmacológico , Terapia Trombolítica/métodos , Ativador de Plasminogênio Tecidual/administração & dosagem , Isquemia Encefálica/diagnóstico , Isquemia Encefálica/fisiopatologia , Estabilidade de Medicamentos , Estabilidade Enzimática , Fibrinolíticos/efeitos adversos , Fibrinolíticos/química , Humanos , Infusões Intravenosas , Estudos Prospectivos , Proteínas Recombinantes/administração & dosagem , Acidente Vascular Cerebral/diagnóstico , Acidente Vascular Cerebral/fisiopatologia , Terapia Trombolítica/efeitos adversos , Fatores de Tempo , Tempo para o Tratamento , Ativador de Plasminogênio Tecidual/efeitos adversos , Ativador de Plasminogênio Tecidual/química , Resultado do TratamentoRESUMO
OBJECTIVE: Clinical evidence suggests an association between oxidative stress and vascular disease, and in vitro studies have demonstrated that reactive oxygen species can have prothrombotic effects on vascular and blood cells. It remains unclear, however, whether elevated levels of reactive oxygen species accelerate susceptibility to experimental thrombosis in vivo. APPROACH AND RESULTS: Using a murine model with genetic deficiency in superoxide dismutase-1 (SOD1), we measured susceptibility to carotid artery thrombosis in response to photochemical injury. We found that SOD1-deficient (Sod1(-/-)) mice formed stable arterial occlusions significantly faster than wild-type (Sod1(+/+)) mice (P<0.05). Sod1(-/-) mice also developed significantly larger venous thrombi than Sod1(+/+) mice after inferior vena cava ligation (P<0.05). Activation of protein C by thrombin in lung was diminished in Sod1(-/-) mice (P<0.05 versus Sod1(+/+) mice), and generation of activated protein C in response to infusion of thrombin in vivo was decreased in Sod1(-/-) mice (P<0.05 versus Sod1(+/+) mice). SOD1 deficiency had no effect on the expression of thrombomodulin, endothelial protein C receptor, or tissue factor in lung or levels of protein C in plasma. Exposure of human thrombomodulin to superoxide in vitro caused oxidation of multiple methionine residues, including critical methionine 388, and a 40% decrease in thrombomodulin-dependent activation of protein C (P<0.05). SOD and catalase protected against superoxide-induced methionine oxidation and restored protein C activation in vitro (P<0.05). CONCLUSIONS: SOD prevents thrombomodulin methionine oxidation, promotes protein C activation, and protects against arterial and venous thrombosis in mice.
Assuntos
Lesões das Artérias Carótidas/enzimologia , Proteína C/metabolismo , Superóxido Dismutase/deficiência , Trombose/enzimologia , Trombose Venosa/enzimologia , Animais , Lesões das Artérias Carótidas/genética , Lesões das Artérias Carótidas/patologia , Artéria Carótida Primitiva/enzimologia , Artéria Carótida Primitiva/patologia , Catalase/metabolismo , Modelos Animais de Doenças , Ativação Enzimática , Humanos , Camundongos da Linhagem 129 , Camundongos Transgênicos , Oxirredução , Estresse Oxidativo , Superóxido Dismutase/genética , Superóxido Dismutase-1 , Superóxidos/metabolismo , Trombomodulina/metabolismo , Trombose/genética , Trombose/patologia , Veia Cava Inferior/enzimologia , Veia Cava Inferior/patologia , Trombose Venosa/patologiaRESUMO
BACKGROUND: The incidence of thrombotic events increases during aging, but the mechanisms are not well understood. To investigate the prothrombotic role of oxidative stress during aging, we tested the hypothesis that aged mice overexpressing the antioxidant enzyme glutathione peroxidase-1 (Gpx1) are protected from experimental thrombosis. METHODS AND RESULTS: Susceptibility to carotid artery thrombosis was first examined in wild-type C57BL/6J mice. After photochemical injury of the carotid artery, the time to stable occlusion was significantly shorter in 12- and 18-month-old mice compared with 4-month-old mice (P<0.01). Unlike wild-type mice, transgenic mice overexpressing Gpx1 (Gpx1 Tg) did not exhibit shortened times to occlusion of the carotid artery at 12 or 18 months of age. Wild-type mice also exhibited increased susceptibility to venous thrombosis after inferior vena cava ligation at 12 or 18 months of age (P<0.05 versus 4 months of age). Gpx1 Tg mice were protected from this aging-related enhanced susceptibility to venous thrombosis. Age-dependent platelet hyperactivation, evidenced by increased hydrogen peroxide, fibrinogen binding, and activation of fibrinogen receptor αIIbß3, was observed in thrombin-activated platelets from wild-type but not Gpx1 Tg mice (P<0.05). Enhanced platelet activation responses in aged mice were also prevented by polyethylene glycol-catalase or apocynin, an inhibitor of NADPH oxidase. Aged mice displayed increased intraplatelet expression of p47(phox) and superoxide dismutase-1, suggesting a mechanistic pathway for increased hydrogen peroxide generation. CONCLUSIONS: Our findings demonstrate that hydrogen peroxide is a key mediator of platelet hyperactivity and enhanced thrombotic susceptibility in aged mice.
Assuntos
Envelhecimento/metabolismo , Plaquetas/metabolismo , Peróxido de Hidrogênio/metabolismo , Ativação Plaquetária/fisiologia , Trombose/epidemiologia , Trombose/metabolismo , Acetofenonas/farmacologia , Animais , Catalase/farmacologia , Feminino , Glutationa Peroxidase/genética , Glutationa Peroxidase/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Modelos Animais , NADPH Oxidases/antagonistas & inibidores , NADPH Oxidases/metabolismo , Estresse Oxidativo/fisiologia , Ativação Plaquetária/efeitos dos fármacos , Polietilenoglicóis/farmacologia , Fatores de Risco , Superóxido Dismutase/metabolismo , Superóxido Dismutase-1 , Trombose/fisiopatologia , Glutationa Peroxidase GPX1RESUMO
Hyperhomocysteinemia confers a high risk for thrombotic vascular events, but homocysteine-lowering therapies have been ineffective in reducing the incidence of secondary vascular outcomes, raising questions regarding the role of homocysteine as a mediator of cardiovascular disease. Therefore, to determine the contribution of elevated homocysteine to thrombosis susceptibility, we studied Cbs(-/-) mice conditionally expressing a zinc-inducible mutated human CBS (I278T) transgene. Tg-I278T Cbs(-/-) mice exhibited severe hyperhomocysteinemia and endothelial dysfunction in cerebral arterioles. Surprisingly, however, these mice did not display increased susceptibility to arterial or venous thrombosis as measured by photochemical injury in the carotid artery, chemical injury in the carotid artery or mesenteric arterioles, or ligation of the inferior vena cava. A survey of hemostatic and hemodynamic parameters revealed no detectible differences between control and Tg-I278T Cbs(-/-) mice. Our data demonstrate that severe elevation in homocysteine leads to the development of vascular endothelial dysfunction but is not sufficient to promote thrombosis. These findings may provide insights into the failure of homocysteine-lowering trials in secondary prevention from thrombotic vascular events.
Assuntos
Modelos Animais de Doenças , Hiper-Homocisteinemia/complicações , Hiper-Homocisteinemia/patologia , Camundongos , Trombose/etiologia , Animais , Cistationina beta-Sintase/genética , Feminino , Testes Hematológicos , Hemodinâmica/genética , Hemodinâmica/fisiologia , Humanos , Hiper-Homocisteinemia/sangue , Hiper-Homocisteinemia/genética , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , Fatores de Risco , Índice de Gravidade de DoençaRESUMO
BACKGROUND: Alcoholic steatohepatitis (ASH) is caused in part by the effects of ethanol (EtOH) on hepatic methionine metabolism. METHODS: To investigate the phenotypic and epigenetic consequences of altered methionine metabolism in this disease, we studied the effects of 4-week intragastric EtOH feeding with and without the methyl donor betaine in cystathionine beta synthase (CßS) heterozygous C57BL/6J mice. RESULTS: The histopathology of early ASH was induced by EtOH feeding and prevented by betaine supplementation, while EtOH feeding reduced and betaine supplementation maintained the hepatic methylation ratio of the universal methyl donor S-adenosylmethionine (SAM) to the methyltransferase inhibitor S-adenosylhomocysteine (SAH). MethylC-seq genomic sequencing of heterozygous liver samples from each diet group found 2 to 4% reduced methylation in gene bodies, but not promoter regions of all autosomes of EtOH-fed mice, each of which were normalized in samples from mice fed the betaine-supplemented diet. The transcript levels of nitric oxide synthase (Nos2) and DNA methyltransferase 1 (Dnmt1) were increased, while those of peroxisome proliferator receptor-α (Pparα) were reduced in EtOH-fed mice, and each was normalized in mice fed the betaine-supplemented diet. DNA pyrosequencing of CßS heterozygous samples found reduced methylation in a gene body of Nos2 by EtOH feeding that was restored by betaine supplementation and was correlated inversely with its expression and positively with SAM/SAH ratios. CONCLUSIONS: The present study has demonstrated relationships among EtOH induction of ASH with aberrant methionine metabolism that was associated with gene body DNA hypomethylation in all autosomes and was prevented by betaine supplementation. The data imply that EtOH-induced changes in selected gene transcript levels and hypomethylation in gene bodies during the induction of ASH are a result of altered methionine metabolism that can be reversed through dietary supplementation of methyl donors.
Assuntos
Betaína/uso terapêutico , Metilação de DNA/efeitos dos fármacos , Etanol/farmacologia , Fígado Gorduroso Alcoólico/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Homocistinúria/metabolismo , Animais , DNA (Citosina-5-)-Metiltransferase 1 , DNA (Citosina-5-)-Metiltransferases/análise , Suplementos Nutricionais , Fígado/química , Fígado/efeitos dos fármacos , Metionina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico Sintase Tipo II/análise , PPAR alfa/análise , S-Adenosil-Homocisteína/metabolismo , S-Adenosilmetionina/metabolismoRESUMO
PURPOSE: Cisplatin contributes to acute kidney injury (AKI) and chronic kidney disease (CKD) that occurs with greater frequency and severity in older patients. Age-associated cisplatin sensitivity in human fibroblasts involves increased mitochondrial superoxide produced by older donor cells. EXPERIMENTAL DESIGN: Young and old C57BL/6 J murine models of cisplatin-induced AKI and CKD were treated with the SOD mimetic avasopasem manganese to investigate the potential antioxidant and anti-inflammatory effects. Adverse event reporting from a phase 2 and a phase 3 randomized clinical trial (NCT02508389 and NCT03689712) conducted in patients treated with cisplatin and AVA was determined to have established the incidence and severity of AKI. RESULTS: Cisplatin-induced AKI and CKD occurred in all mice, however, was more pronounced in older mice. AVA reduced cisplatin-induced mortality, AKI, and CKD, in older animals. AVA also alleviated cisplatin-induced alterations in mitochondrial electron transport chain (ETC) complex activities and NADPH Oxidase 4 (NOX4) and inhibited the increased levels of the inflammation markers, TNFα, IL1, ICAM-1, and VCAM-1. Analysis of age-stratified subjects treated with cisplatin from clinical trials (NCT02508389, NCT03689712) also supported that the incidence of AKI increased with age and AVA reduced age-associated therapy-induced adverse events (AE), including hypomagnesemia, increased creatinine, and AKI. CONCLUSIONS: Older mice and humans are more susceptible to cisplatin-induced kidney injury, and treatment with AVA mitigates age-associated damage. Mitochondrial ETC and NOX4 activities represent sources of superoxide production contributing to cisplatin-induced kidney injury, and pro-inflammatory cytokine production and endothelial dysfunction may also be increased by superoxide formation.
Assuntos
Injúria Renal Aguda , Compostos Organometálicos , Insuficiência Renal Crônica , Humanos , Camundongos , Animais , Idoso , Cisplatino/farmacologia , Antioxidantes/farmacologia , Antioxidantes/uso terapêutico , Superóxidos , Camundongos Endogâmicos C57BL , Rim , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/tratamento farmacológico , Anti-Inflamatórios/farmacologiaRESUMO
BACKGROUND: COVID-19 can cause profound inflammation and coagulopathy, and while many mechanisms have been proposed, there is no known common pathway leading to a prothrombotic state. OBJECTIVES: From the beginning of the COVID-19 pandemic, elevated levels of extracellular histones have been found in plasma of patients infected with SARS-CoV-2. We hypothesized that platelet activation triggered by extracellular histones might represent a unifying mechanism leading to increased thrombin generation and thrombosis. METHODS: We utilized blood samples collected from an early clinical trial of hospitalized COVID-19 patients (NCT04360824) and recruited healthy subjects as controls. Using plasma samples, we measured the procoagulant and prothrombotic potential of circulating extracellular histones and extracellular vesicles (EVs). Platelet prothrombotic activity was assessed via thrombin generation potential and platelet thrombus growth. Circulating EVs were assessed for thrombin generation potential in vitro in plasma and enhancement of thrombotic susceptibility in vivo in mice. RESULTS: Compared with controls, COVID-19 patients had elevated plasma levels of citrullinated histone H3, cell-free DNA, nucleosomes, and EVs. Plasma from COVID-19 patients promoted platelet activation, platelet-dependent thrombin generation, thrombus growth under venous shear stress, and release of platelet-derived EVs. These prothrombotic effects of COVID-19 plasma were inhibited by an RNA aptamer that neutralizes both free and DNA-bound histones. EVs isolated from COVID-19 plasma enhanced thrombin generation in vitro and potentiated venous thrombosis in mice in vivo. CONCLUSION: We conclude that extracellular histones and procoagulant EVs drive the prothrombotic state in COVID-19 and that histone-targeted therapy may prove beneficial.
Assuntos
Plaquetas , COVID-19 , Vesículas Extracelulares , Histonas , Ativação Plaquetária , Trombose , Adulto , Animais , Feminino , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Coagulação Sanguínea , Plaquetas/metabolismo , Estudos de Casos e Controles , Citrulinação , COVID-19/sangue , COVID-19/complicações , Vesículas Extracelulares/metabolismo , Histonas/sangue , Camundongos Endogâmicos C57BL , Nucleossomos/metabolismo , Trombina/metabolismo , Trombose/sangue , Trombose/etiologia , Ensaios Clínicos Fase IV como AssuntoRESUMO
The University of North Carolina Symposia on Hemostasis began in 2002, with The First Symposium on Hemostasis with a Special Focus on FVIIa and Tissue Factor. They have occurred biannually since and have maintained the primary goal of establishing a forum for the sharing of outstanding advances made in the basic sciences of hemostasis. The 2024 11th Symposium on Hemostasis will bring together leading scientists from around the globe to present and discuss the latest research related to coagulation factors and platelet biology. In keeping with the tradition of the conference, we expect novel cross-disciplinary collaborations to result from bringing together fundamental scientists and physician-scientists from different backgrounds and perspectives. The aim of these collaborations is to springboard the next generation of important advances in the field. This year's program was designed to discuss Coagulation and Platelet Biology at the Intersection of Health and Disease. The goal is to develop a better understanding of the pathophysiologic mechanisms leading to hemostatic and thrombotic disorders as this understanding is critical for the continued development of safe and efficacious therapeutics. Included in this review article are illustrated capsules provided by our speakers that highlight the main conclusions of the invited talks.
RESUMO
Ischemic thrombotic disease, characterized by the formation of obstructive blood clots within arteries or veins, is a condition associated with life-threatening events, such as stroke, myocardial infarction, deep vein thrombosis, and pulmonary embolism. The conventional therapeutic strategy relies on treatments with anticoagulants that unfortunately pose an inherent risk of bleeding complications. These anticoagulants primarily target clotting factors, often overlooking upstream events, including the release of neutrophil extracellular traps (NETs). Neutrophils are integral components of the innate immune system, traditionally known for their role in combating pathogens through NET formation. Emerging evidence has now revealed that NETs contribute to a prothrombotic milieu by promoting platelet activation, increasing thrombin generation, and providing a scaffold for clot formation. Additionally, NET components enhance clot stability and resistance to fibrinolysis. Clinical and preclinical studies have underscored the mechanistic involvement of NETs in the pathogenesis of thrombotic complications, since the clots obtained from patients and experimental models consistently exhibit the presence of NETs. Given these insights, the inhibition of NETs or NET formation is emerging as a promising therapeutic approach for ischemic thrombotic diseases. Recent investigations also implicate a role for the nucleotide-binding oligomerization domain (NOD)-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome as a mediator of NETosis and thrombosis, suggesting that NLRP3 inhibition may also hold potential for mitigating thrombotic events. Therefore, future preclinical and clinical studies aimed at identifying and validating NLRP3 inhibition as a novel therapeutic intervention for thrombotic disorders are imperative.
Assuntos
Armadilhas Extracelulares , Trombose , Humanos , Proteína 3 que Contém Domínio de Pirina da Família NLR , Trombose/patologia , Neutrófilos , Armadilhas Extracelulares/fisiologia , AnticoagulantesRESUMO
Hyperhomocysteinemia has long been associated with atherosclerosis and thrombosis and is an independent risk factor for cardiovascular disease. Its causes include both genetic and environmental factors. Although homocysteine is produced in every cell as an intermediate of the methionine cycle, the liver contributes the major portion found in circulation, and fatty liver is a common finding in homocystinuric patients. To understand the spectrum of proteins and associated pathways affected by hyperhomocysteinemia, we analyzed the mouse liver proteome of gene-induced (cystathionine beta-synthase (CBS)) and diet-induced (high methionine) hyperhomocysteinemic mice using two-dimensional difference gel electrophoresis and Ingenuity Pathway Analysis. Nine proteins were identified whose expression was significantly changed by 2-fold (p < or = 0.05) as a result of genotype, 27 proteins were changed as a result of diet, and 14 proteins were changed in response to genotype and diet. Importantly, three enzymes of the methionine cycle were up-regulated. S-Adenosylhomocysteine hydrolase increased in response to genotype and/or diet, whereas glycine N-methyltransferase and betaine-homocysteine methyltransferase only increased in response to diet. The antioxidant proteins peroxiredoxins 1 and 2 increased in wild-type mice fed the high methionine diet but not in the CBS mutants, suggesting a dysregulation in the antioxidant capacity of those animals. Furthermore, thioredoxin 1 decreased in both wild-type and CBS mutants on the diet but not in the mutants fed a control diet. Several urea cycle proteins increased in both diet groups; however, arginase 1 decreased in the CBS(+/-) mice fed the control diet. Pathway analysis identified the retinoid X receptor signaling pathway as the top ranked network associated with the CBS(+/-) genotype, whereas xenobiotic metabolism and the NRF2-mediated oxidative stress response were associated with the high methionine diet. Our results show that hyperhomocysteinemia, whether caused by a genetic mutation or diet, alters the abundance of several liver proteins involved in homocysteine/methionine metabolism, the urea cycle, and antioxidant defense.
Assuntos
Antioxidantes/metabolismo , Cistationina beta-Sintase/genética , Dieta/efeitos adversos , Homocisteína/metabolismo , Hiper-Homocisteinemia/enzimologia , Fígado/enzimologia , Metionina/metabolismo , Nutrigenômica , Ureia/metabolismo , Adenosil-Homocisteinase/metabolismo , Animais , Betaína-Homocisteína S-Metiltransferase/metabolismo , Glicina N-Metiltransferase/metabolismo , Hiper-Homocisteinemia/induzido quimicamente , Hiper-Homocisteinemia/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Peroxirredoxinas/metabolismo , ProteômicaRESUMO
Aging is intrinsically linked with physiologic decline and is a major risk factor for a broad range of diseases. The deleterious effects of advancing age on the vascular system are evidenced by the high incidence and prevalence of cardiovascular disease in the elderly. Reactive oxygen species are critical mediators of normal vascular physiology and have been shown to gradually increase in the vasculature with age. There is a growing appreciation for the complexity of oxidant and antioxidant systems at the cellular and molecular levels, and accumulating evidence indicates a causal association between oxidative stress and age-related vascular disease. Herein, we review the current understanding of mechanistic links between oxidative stress and thrombotic vascular disease and the changes that occur with aging. While several vascular cells are key contributors, we focus on oxidative changes that occur in platelets and their mediation in disease progression. Additionally, we discuss the impact of comorbid conditions (i.e., diabetes, atherosclerosis, obesity, cancer, etc.) that have been associated with platelet redox dysregulation and vascular disease pathogenesis. As we continue to unravel the fundamental redox mechanisms of the vascular system, we will be able to develop more targeted therapeutic strategies for the prevention and management of age-associated vascular disease.
RESUMO
Background Human aging is associated with increased risk of thrombosis, but the mechanisms are poorly defined. We hypothesized that aging induces peroxide-dependent release of neutrophil extracellular traps that contribute to thrombin generation and thrombosis. Methods and Results We studied C57BL6J mice and littermates of glutathione peroxidase-1 transgenic and wild-type mice at young (4 month) and old (20 month) ages and a healthy cohort of young (18-39 years) or middle-aged/older (50-72 years) humans. In plasma, we measured thrombin generation potential and components of neutrophil extracellular traps (cell-free DNA and citrullinated histone). Aged wild-type mice displayed a significant increase in thrombin generation that was decreased in aged glutathione peroxidase-1 transgenic mice. Both aged wild-type and aged glutathione peroxidase-1 transgenic mice demonstrated similar elevation of plasma cell-free DNA compared with young mice. In contrast, plasma levels of citrullinated histone were not altered with age or genotype. Release of neutrophil extracellular traps from neutrophils in vitro was also similar between young and aged wild-type or glutathione peroxidase-1 transgenic mice. Treatment of plasma or mice with DNase 1 decreased age-associated increases in thrombin generation, and DNase 1 treatment blocked the development of experimental venous thrombi in aged C57BL6J mice. Similarly, thrombin generation potential and plasma cell-free DNA, but not citrullinated histone, were higher in middle-aged/older humans, and treatment of plasma with DNase 1 reversed the increase in thrombin generation. Conclusions We conclude that DNase 1 limits thrombin generation and protects from venous thrombosis during aging, likely by hydrolyzing cell-free DNA.
Assuntos
Ácidos Nucleicos Livres , Trombose , Trombose Venosa , Idoso , Envelhecimento , Animais , Estudos Transversais , Desoxirribonucleases , Glutationa Peroxidase , Histonas , Humanos , Camundongos , Pessoa de Meia-Idade , Neutrófilos/metabolismo , Trombina/metabolismo , Trombose Venosa/genética , Trombose Venosa/prevenção & controleRESUMO
UNLABELLED: We tested the hypothesis that the pathogenesis of alcoholic liver injury is mediated by epigenetic changes in regulatory genes that result from the induction of aberrant methionine metabolism by ethanol feeding. Five-month-old cystathionine beta synthase heterozygous and wild-type C57BL/6J littermate mice were fed liquid control or ethanol diets by intragastric infusion for 4 weeks. Both ethanol-fed groups showed typical histopathology of alcoholic steatohepatitis, with reduction in liver S-adenosylmethionine (SAM), elevation in liver S-adenosylhomocysteine (SAH), and reduction in the SAM/SAH ratio with interactions of ethanol and genotype effects. Hepatic endoplasmic reticulum stress signals including glucose-regulated protein-78 (GRP78), activating transcription factor 4, growth arrest and DNA damage-inducible gene 153 (GADD153), caspase 12, and transcription factor sterol response element binding protein-1c (SREBP-1c) were up-regulated in ethanol-fed mice with genotype interactions and negative correlations with the SAM/SAH ratio. Immunohistochemical staining showed reduction in trimethylated histone H3 lysine-9 (3meH3K9) protein levels in centrilobular regions in both ethanol groups, with no changes in trimethylated histone H3 lysine-4 levels. The chromatin immunoprecipitation assay revealed a decrease in levels of suppressor chromatin marker 3meH3K9 in the promoter regions of GRP78, SREBP-1c, and GADD153 in ethanol-treated heterozygous cystathionine beta synthase mice. The messenger RNA expression of the histone H3K9 methyltransferase EHMT2 (G9a) was selectively decreased in ethanol-fed mice. CONCLUSION: The pathogenesis of alcoholic steatohepatitis is mediated in part through the effects of altered methionine metabolism on epigenetic regulation of pathways of endoplasmic reticulum stress relating to apoptosis and lipogenesis.
Assuntos
Retículo Endoplasmático/genética , Epigênese Genética , Fígado Gorduroso Alcoólico/etiologia , Homocistinúria/genética , Homocistinúria/metabolismo , Fígado/ultraestrutura , Estresse Fisiológico/genética , Animais , Chaperona BiP do Retículo Endoplasmático , Etanol/administração & dosagem , Fígado Gorduroso Alcoólico/genética , Fígado Gorduroso Alcoólico/metabolismo , CamundongosRESUMO
BACKGROUND: Thromboembolism affects up to 30% of children undergoing treatment for acute lymphoblastic leukemia (ALL). Increased thrombin generation has been reported in ALL, but the mechanisms remain elusive. OBJECTIVE: We aimed to show that extracellular traps and cell-free DNA (cfDNA) promote thrombin generation in pediatric ALL. METHODS: In a longitudinal single-center study, we recruited 17 consecutive pediatric ALL patients. Serial blood samples were collected at diagnosis and weekly during the 4-week induction phase of antileukemic chemotherapy. Healthy children (n = 14) and children with deep vein thrombosis (DVT; n = 7) or sepsis (n = 5) were recruited as negative and positive controls, respectively. In plasma, we measured endogenous thrombin generation potential (ETP) and components of extracellular traps, including cfDNA. RESULTS: In patients with ALL, ETP was increased at baseline and remained significantly elevated throughout the induction therapy. Plasma levels of cfDNA were increased at baseline and during the first 3 weeks of induction therapy. The extent of enhancement of ETP and plasma cfDNA in patients with ALL was similar to that seen in patients with DVT or sepsis. Treatment of plasma with DNase 1 lowered ETP in patients with ALL at each time point but did not affect ETP in healthy controls. CONCLUSION: We conclude that childhood ALL is associated with a prothrombotic milieu at the time of diagnosis that continues during induction chemotherapy, and cfDNA contributes to increased thrombogenic potential.
RESUMO
BACKGROUND: Coronavirus disease 2019 (COVID-19) is associated with coagulopathy but the optimal prophylactic anticoagulation therapy remains uncertain and may depend on COVID-19 severity. OBJECTIVE: To compare outcomes in hospitalized adults with severe COVID-19 treated with standard prophylactic versus intermediate dose enoxaparin. METHODS: We conducted a multi-center, open-label, randomized controlled trial comparing standard prophylactic dose versus intermediate dose enoxaparin in adults who were hospitalized with COVID-19 and admitted to an intensive care unit (ICU) and/or had laboratory evidence of coagulopathy. Patients were randomly assigned in a 1:1 ratio to receive standard prophylactic dose enoxaparin or intermediate weight-adjusted dose enoxaparin. The primary outcome was all-cause mortality at 30 days. Secondary outcomes included arterial or venous thromboembolism and major bleeding. RESULTS: A total of 176 patients (99 males and 77 females) underwent randomization. In the intention-to-treat population, all-cause mortality at 30 days was 15% for intermediate dose enoxaparin and 21% for standard prophylactic dose enoxaparin (odds ratio, 0.66; 95% confidence interval, 0.30-1.45; P = .31 by Chi-square test). Unadjusted Cox proportional hazards modeling demonstrated no significant difference in mortality between intermediate and standard dose enoxaparin (hazard ratio, 0.67; 95% confidence interval, 0.33-1.37; P = .28). Arterial or venous thrombosis occurred in 13% of patients assigned to intermediate dose enoxaparin and 9% of patients assigned to standard dose enoxaparin. Major bleeding occurred in 2% of patients in each arm. CONCLUSION: In hospitalized adults with severe COVID-19, standard prophylactic dose and intermediate dose enoxaparin did not differ significantly in preventing death or thrombosis at 30 days.