RESUMO
BACKGROUND: Circulating histones are released by extensive tissue injury or cell death and play important pathogenic roles in critical illnesses. Their interaction with circulating plasma components and the potential roles in the clinical setting are not fully understood. OBJECTIVES: We aimed to characterize the interaction of histones with fibrinogen and explore its roles in vitro, in vivo, and in patient samples. METHODS: Histone-fibrinogen binding was assessed by electrophoresis and enzyme-linked immunosorbent assay-based affinity assay. Functional significance was explored using washed platelets and endothelial cells in vitro and histone-infusion mouse models in vivo. To determine clinical translatability, a retrospective single-center cohort study was conducted on patients requiring intensive care admission (n = 199) and validated in a cohort of hospitalized patients with COVID-19 (n = 69). RESULTS: Fibrinogen binds histones through its D-domain with high affinity (calf thymus histones, KD = 18.0 ± 5.6 nM; histone 3, KD = 2.7 ± 0.8 nM; and histone 4, KD = 2.0 ± 0.7 nM) and significantly reduces histone-induced endothelial damage and platelet aggregation in vitro and in vivo in a histone-infusion mouse model. Physiologic concentrations of fibrinogen can neutralize low levels of circulating histones and increase the cytotoxicity threshold of histones to 50 µg/mL. In a cohort of patients requiring intensive care, a histone:fibrinogen ratio of ≥6 on admission was associated with moderate-severe thrombocytopenia and independently predicted mortality. This finding was validated in a cohort of hospitalized patients with COVID-19. CONCLUSION: Fibrinogen buffers the cytotoxic properties of circulating histones. Detection and monitoring of circulating histones and histone:fibrinogen ratios will help identify critically ill patients at highest risk of adverse outcomes who might benefit from antihistone therapy.
Assuntos
COVID-19 , Fibrinogênio , Histonas , Ligação Proteica , Humanos , Fibrinogênio/metabolismo , Histonas/sangue , Histonas/metabolismo , COVID-19/sangue , Animais , Masculino , Feminino , Pessoa de Meia-Idade , Estudos Retrospectivos , Idoso , Camundongos , Agregação Plaquetária/efeitos dos fármacos , Plaquetas/metabolismo , SARS-CoV-2 , Células Endoteliais/metabolismo , AdultoRESUMO
BACKGROUND: Coagulopathy and associated bleeding and deep vein thrombosis (DVT) are major causes of morbidity and mortality in patients with acute leukemia. The underlying mechanisms of these complications have not been fully elucidated. OBJECTIVES: To evaluate the associations between biomarker levels and bleeding and DVT in acute leukemia patients. METHODS: We examined plasma levels of activators, inhibitors, and biomarkers of the coagulation and fibrinolytic pathways in patients aged ≥18 years with newly diagnosed acute leukemia compared with those of normal controls. Multivariable regression models were used to examine the association of biomarkers with bleeding and DVT in acute leukemia patients. The study included 358 patients with acute leukemia (29 with acute promyelocytic leukemia [APL], 253 with non-APL acute myeloid leukemia, and 76 with acute lymphoblastic leukemia) and 30 normal controls. RESULTS: Patients with acute leukemia had higher levels of extracellular vesicle tissue factor (EVTF) activity, phosphatidylserine-positive extracellular vesicles, plasminogen activator inhibitor-1, plasmin-antiplasmin complexes, and cell-free DNA and lower levels of citrullinated histone H3-DNA complexes compared with normal controls. APL patients had the highest levels of EVTF activity and the lowest levels of tissue plasminogen activator among acute leukemia patients. There were 41 bleeding and 23 DVT events in acute leukemia patients. High EVTF activity was associated with increased risk of bleeding (subdistribution hazard ratio, 2.30; 95% CI, 0.99-5.31), whereas high levels of plasminogen activator inhibitor-1 were associated with increased risk of DVT (subdistribution hazard ratio, 3.00; 95% CI, 0.95-9.47) in these patients. CONCLUSION: Our study shows alterations in several biomarkers in acute leukemia and identifies biomarkers associated with risk of bleeding and DVT.
Assuntos
Biomarcadores , Coagulação Sanguínea , Hemorragia , Leucemia Mieloide Aguda , Tromboembolia Venosa , Humanos , Masculino , Pessoa de Meia-Idade , Feminino , Adulto , Hemorragia/sangue , Hemorragia/diagnóstico , Tromboembolia Venosa/sangue , Tromboembolia Venosa/diagnóstico , Tromboembolia Venosa/etiologia , Biomarcadores/sangue , Estudos de Casos e Controles , Idoso , Leucemia Mieloide Aguda/sangue , Leucemia Mieloide Aguda/complicações , Leucemia-Linfoma Linfoblástico de Células Precursoras/sangue , Leucemia-Linfoma Linfoblástico de Células Precursoras/complicações , Fatores de Risco , Leucemia Promielocítica Aguda/sangue , Leucemia Promielocítica Aguda/complicações , Trombose Venosa/sangue , Trombose Venosa/diagnóstico , Trombose Venosa/etiologia , Histonas/sangue , Inibidor 1 de Ativador de Plasminogênio/sangue , Tromboplastina/metabolismo , Tromboplastina/análise , Adulto Jovem , Fosfatidilserinas/sangueRESUMO
BACKGROUND: Neutrophil extracellular traps (NETs) occur when neutrophil chromatin is decondensed and extruded into the extracellular space in a web-like structure. Originally described as an anti-microbial function, this process has been implicated in the pathogenesis of pancreatic disease. In addition, NETs are upregulated during physiologic wound-healing and coagulation. This study evaluated how the inflammatory response to pancreatic surgery influences NET formation. METHODS: For this study, 126 patients undergoing pancreatectomy gave consent before participation. Plasma was collected at several time points (preoperatively and through the postoperative outpatient visit). Plasma levels of NET markers, including cell-free DNA (cfDNA), citrullinated histone H3 (CitH3), interleukin (IL)-8, IL-6, and granulocyte colony-stimulating factor (G-CSF) were measured using enzyme-linked immunosorbent assay (ELISA). Patient clinical data were retrospectively collected from a prospectively maintained database. RESULTS: After pancreatic resection, NET markers (cfDNA and CitH3) were elevated, peaking on postoperative days 3 and 4. This increase in NETs was due to an inherent change in neutrophil biology. Postoperatively, NET-inducing cytokines (IL-8, IL-6, and G-CSF) were increased, peaking early in the postoperative course. The patients undergoing the robotic approach had a reduction in NETs during the postoperative period compared with those who underwent the open approach. The patients who experienced a pancreatic leak had an increase in NET markers during the postoperative period. CONCLUSIONS: Pancreatectomy induces cancer-promoting NET formation. The minimally invasive robotic approach may induce fewer NETs, although the current analysis was limited by selection bias. Pancreatic leak resulted in increased NETs. Further study into the potential for NET inhibition during the perioperative period is warranted.
Assuntos
Armadilhas Extracelulares , Pancreatectomia , Neoplasias Pancreáticas , Humanos , Armadilhas Extracelulares/metabolismo , Pancreatectomia/efeitos adversos , Neoplasias Pancreáticas/cirurgia , Neoplasias Pancreáticas/patologia , Masculino , Feminino , Pessoa de Meia-Idade , Idoso , Seguimentos , Neutrófilos/patologia , Neutrófilos/metabolismo , Estudos Retrospectivos , Prognóstico , Ácidos Nucleicos Livres/sangue , Estudos Prospectivos , Adulto , Histonas/metabolismo , Histonas/sangue , Fator Estimulador de Colônias de Granulócitos/sangue , Interleucina-6/sangue , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/metabolismoRESUMO
OBJECTIVES: Serum citrullinated histone H3 (CitH3) levels in humans with rheumatoid arthritis (RA) were examined and the associations between CitH3 levels and disease variables were investigated. METHODS: Serum CitH3 levels were measured using an enzyme-linked immunosorbent assay in 151 RA patients (69 with highly, 32 with moderately, and 20 with mildly active RA and 30 with RA in remission) and 56 healthy controls. Receiver operating characteristic curve analysis was performed to evaluate the discriminant capacity of CitH3 between highly/moderately active RA and RA in remission/mild activity. Furthermore, machine-learning methods were applied to construct a predictive model. RESULTS: CitH3 concentration was more upregulated in patients with highly and moderately active RA than in those with mild activity and remission. The area under the curve for CitH3 was 0.825 for discriminating between highly and mildly active RA, 0.840 for discriminating between highly active RA and RA in remission, 0.789 for discriminating between moderately and mildly active RA, and 0.829 for discriminating between moderately active RA and RA in remission. The correlation analysis revealed that serum CitH3 levels were positively associated with Disease Activity Score 28-Erythrocyte Sedimentation Rate (DAS28-ESR), ESR, C-reactive protein, rheumatoid factor, and some routine blood parameters (WBC, RDW, PLT, N, and N%), while negatively associated with haemoglobin, lymphocyte percentage, and thyroid-stimulating hormone. Through machine learning, the optimal predictive model was selected and had high performance. CONCLUSIONS: CitH3 is significantly associated with disease activity and could serve as a useful candidate biomarker to assess disease activity in patients with RA.
Assuntos
Artrite Reumatoide , Histonas , Humanos , Artrite Reumatoide/sangue , Estudos de Casos e Controles , Histonas/sangue , Fator Reumatoide/sangue , Proteína C-Reativa , Biomarcadores , ELISPOTRESUMO
OBJECTIVE: Exploration of biomarkers to predict the severity of COVID-19 is important to reduce mortality. Upon COVID-19 infection, neutrophil extracellular traps (NET) are formed, which leads to a cytokine storm and host damage. Hence, the extent of NET formation may reflect disease progression and predict mortality in COVID-19. METHODS: We measured 4 NET parameters - cell-free double stranded DNA (cell-free dsDNA), neutrophil elastase, citrullinated histone H3 (Cit-H3), and histone - DNA complex - in 188 COVID-19 patients and 20 healthy controls. Survivors (n=166) were hospitalized with or without oxygen supplementation, while non-survivors (n=22) expired during in-hospital treatment. RESULTS: Cell-free dsDNA was significantly elevated in non-survivors in comparison with survivors and controls. The survival rate of patients with high levels of cell-free dsDNA, neutrophil elastase, and Cit-H3 was significantly lower than that of patients with low levels. These three markers significantly correlated with inflammatory markers (absolute neutrophil count and C-reactive protein). CONCLUSION: Since the increase in NET parameters indicates the unfavourable course of COVID-19 infection, patients predisposed to poor outcome can be rapidly managed through risk stratification by using these NET parameters.
Assuntos
COVID-19 , Armadilhas Extracelulares , Biomarcadores/metabolismo , COVID-19/diagnóstico , Ácidos Nucleicos Livres/sangue , Ácidos Nucleicos Livres/metabolismo , Armadilhas Extracelulares/metabolismo , Histonas/sangue , Histonas/metabolismo , Humanos , Elastase de Leucócito/sangue , Elastase de Leucócito/metabolismo , Neutrófilos/metabolismo , PrognósticoRESUMO
BACKGROUND: Premature ovarian insufficiency (POI) plagues 1% of women under 40, while quite a few remain an unknown cause. The development of sequencing has helped find pathogenic genes and reveal the relationship between DNA repair and ovarian reserve. Through the exome sequencing, our study targets screening out the possible POI pathogenic gene and variants in a Chinese family and 20 sporadic POI patients, preliminarily exploring the functional impact and finding out potential linkages between the gene and POI. RESULTS: The whole exome sequencing suggested a novel FMN2 heterozygous variant c.1949C > T (p.Ser650Leu) carried by all three patients in a Chinese family and another c.1967G > A(p.Arg656His) variant in a sporadic case. Since no FMN2 missense mutation is reported for causing human POI, we preliminarily assessed p.Ser650Leu variant via cross-species alignment and 3D modeling and found it possibly deleterious. A series of functional evidence was consistent with our hypothesis. We proved the expression of FMN2 in different stages of oocytes and observed a statistical difference of chromosomal breakages between the POI patient carrying p.Arg656His variant and the health control (p = 0.0013). Western Blot also suggested a decrease in FMN2 and P21 in the mutant type and an associated increase in H2AX. The p.Arg656His variant with an extremely low frequency also indicated that the gene FMN2 might play an essential role in the genetic etiology of POI. To the best of our knowledge, this is the first POI report on missense variants of FMN2. CONCLUSION: This finding indicates a novel gene possibly related to POI and sheds lights on the study of FMN2.
Assuntos
Forminas/genética , Forminas/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Insuficiência Ovariana Primária/genética , Adulto , Reparo do DNA/genética , Feminino , Feto/metabolismo , Heterozigoto , Histonas/sangue , Humanos , Linfócitos/metabolismo , Estrutura Molecular , Mutação de Sentido Incorreto , Ovário/metabolismo , Linhagem , Insuficiência Ovariana Primária/sangue , Sequenciamento do ExomaRESUMO
Burn injuries elicit a unique and dynamic stress response which can lead to burn injury progression. Though neutrophils represent crucial players in the burn-induced immunological events, the dynamic secretion pattern and systemic levels of neutrophil-derived factors have not been investigated in detail so far. Serum levels of neutrophil elastase (NE), myeloperoxidase (MPO), citrullinated histone H3 (CitH3), and complement factor C3a were quantified in burn victims over 4 weeks post injury. Furthermore, the potential association with mortality, degree of burn injury, and inhalation trauma was evaluated. In addition, leukocyte, platelet, neutrophil, and lymphocyte counts were assessed. Lastly, we analyzed the association of neutrophil-derived factors with clinical severity scoring systems. Serum levels of NE, MPO, CitH3, and C3a were remarkably elevated in burn victims compared to healthy controls. Leukocyte and neutrophil counts were significantly increased on admission day and day 1, while relative lymphocytes were decreased in the first 7 days post burn trauma. Though neutrophil-derived factors did not predict mortality, patients suffering from 3rd degree burn injuries displayed increased CitH3 and NE levels. Accordingly, CitH3 and NE were elevated in cases with higher abbreviated burn severity indices (ABSI). Taken together, our data suggest a role for neutrophil activation and NETosis in burn injuries and burn injury progression. Targeting exacerbated neutrophil activation might represent a new therapeutic option for severe cases of burn injury.
Assuntos
Queimaduras/imunologia , Ativação de Neutrófilo , Neutrófilos/imunologia , Adulto , Idoso , Biomarcadores/sangue , Queimaduras/sangue , Queimaduras/diagnóstico , Queimaduras/mortalidade , Estudos de Casos e Controles , Citrulinação , Complemento C3/metabolismo , Feminino , Histonas/sangue , Humanos , Contagem de Leucócitos , Elastase de Leucócito/sangue , Masculino , Pessoa de Meia-Idade , Neutrófilos/metabolismo , Peroxidase/sangue , Valor Preditivo dos Testes , Prognóstico , Processamento de Proteína Pós-Traducional , Índice de Gravidade de Doença , Fatores de Tempo , Adulto JovemRESUMO
BACKGROUND/AIM: Inflammation is a hallmark of cancer, and the role of neutrophils and neutrophil extracellular traps (NETs) in cancer and cancer-associated thrombosis has attracted a lot of interest. The NET-specific marker H3Cit has been found to be elevated in the plasma of patients with malignancies, suggesting NETs markers as novel cancer biomarkers. This study aimed to determine the levels of NETs markers (H3Cit and dsDNA) in the plasma of women with adnexal masses. PATIENTS AND METHODS: Peripheral blood samples were obtained from 199 patients admitted for primary surgery of adnexal masses. Patients were grouped according to tumor type and stage. Plasma levels of H3Cit-DNA, dsDNA, and CA125 were quantified. RESULTS: Plasma levels of H3Cit-DNA and dsDNA were not elevated in women with borderline or malignant ovarian tumors compared with those of the benign group. Increased levels of CA125 were found in the borderline and ovarian cancer group (ptrend<0.001). In Cox regression analysis, CA125 levels dichotomized at 326 IU/ml (median) were associated with worse overall survival (HR=1.9; 95%CI=1.03-3.36; p=0.038). No differences were found in the survival analyses of malignant ovarian tumors by analyzing the dsDNA and H3Cit-DNA levels. CONCLUSION: There is no association between NETs markers and ovarian tumors.
Assuntos
Armadilhas Extracelulares/metabolismo , Neutrófilos/metabolismo , Neoplasias Ovarianas/sangue , Adulto , Idoso , Biomarcadores Tumorais/sangue , Antígeno Ca-125/sangue , DNA/sangue , Feminino , Histonas/sangue , Humanos , Proteínas de Membrana/sangue , Pessoa de Meia-Idade , Neoplasias Ovarianas/mortalidade , Neoplasias Ovarianas/patologia , Análise de SobrevidaRESUMO
OBJECTIVES: Pulmonary thrombus formation is a hallmark of coronavirus disease 2019 (COVID-19). A dysregulated immune response culminating in thromboinflammation has been described, but the pathomechanisms remain unclear. METHODS: We studied 41 adult COVID-19 patients with positive results on reverse-transcriptase polymerase-chain-reaction assays and 37 sex- and age-matched healthy controls. Number and surface characteristics of extracellular vesicles (EVs) and citrullinated histone H3 levels were determined in plasma upon inclusion by flow cytometry and immunoassay. RESULTS: In total, 20 patients had severe and 21 nonsevere disease. The number of EV (median [25th, 75th percentile]) was significantly higher in patients compared with controls (658.8 [353.2, 876.6] vs. 435.5 [332.5, 585.3], geometric mean ratio [95% confidence intervals]: 2.6 [1.9, 3.6]; p < 0.001). Patients exhibited significantly higher numbers of EVs derived from platelets, endothelial cells, leukocytes, or neutrophils than controls. EVs from alveolar-macrophages and alveolar-epithelial cells were detectable in plasma and were significantly higher in patients. Intercellular adhesion molecule-1-positive EV levels were higher in patients, while no difference between tissue factor-positive and angiotensin-converting enzyme-positive EV was seen between both groups. Levels of EV did not differ between patients with severe and nonsevere COVID-19. Citrullinated histone H3 levels (ng/mL, median [25th, 75th percentile]) were higher in patients than in controls (1.42 [0.6, 3.4] vs. 0.31 [0.1, 0.6], geometric mean ratio: 4.44 [2.6, 7.7]; p < 0.001), and were significantly lower in patients with nonsevere disease compared with those with severe disease. CONCLUSION: EV and citrullinated histone H3 are associated with COVID-19 and could provide information regarding pathophysiology of the disease.
Assuntos
COVID-19/sangue , Vesículas Extracelulares/patologia , Histonas/sangue , SARS-CoV-2 , Adulto , Idoso , Biomarcadores/sangue , Plaquetas/patologia , COVID-19/complicações , Estudos de Casos e Controles , Citrulinação , Armadilhas Extracelulares/metabolismo , Feminino , Histonas/química , Humanos , Masculino , Pessoa de Meia-Idade , Pandemias , Índice de Gravidade de Doença , Tromboinflamação/sangue , Tromboinflamação/etiologiaRESUMO
BACKGROUND: Although appendicitis is one of the most frequently occurring pediatric surgery emergencies, current biomarkers for diagnosis are unspecific and have low predictive values. Neutrophils are an essential component of the innate immune system involved during appendicitis. Thus, the current study aimed to evaluate neutrophils and their activation markers in a prospective cohort study. METHODS: The study population included all children with acute abdominal pain who presented to the pediatric surgery department of 2 large clinics between July 2018 and December 2019. All enrolled subjects underwent blood sample collection with an assessment of white blood cell count, C-reactive protein, cell-free DNA, neutrophil elastase, myeloperoxidase, and citrullinated histone H3. If an appendectomy was performed, the appendix was stained for myeloperoxidase, neutrophil elastase, and citrullinated histone H3 using immunofluorescence. RESULTS: In total, 198 subjects were included in the study, of whom 133 had histological verified appendicitis. In those with appendicitis, white blood cell count and C-reactive protein showed a moderate diagnostic value for (noncomplicated and complicated) appendicitis. However, cell-free DNA (area under the curve .87) and citrullinated histone H3 (area under the curve .88) demonstrated excellent predictive power for appendicitis. Most notably, citrullinated histone H3 was able to distinguish (1) noncomplicated from complicated appendicitis, and (2) predict patient outcome. Moreover, the examined biomarkers appear to reflect tissue expression and disease severity. CONCLUSION: Markers of neutrophil activation and extracellular trap formation are excellent biomarkers for appendicitis. In particular, citrullinated histone H3 may be used to identify children with an increased risk of developing complications after appendicitis.
Assuntos
Apendicite/diagnóstico , Apendicite/patologia , Armadilhas Extracelulares , Ativação de Neutrófilo , Abdome Agudo/etiologia , Apendicite/sangue , Biomarcadores/sangue , Contagem de Células Sanguíneas , Proteína C-Reativa/metabolismo , Ácidos Nucleicos Livres/sangue , Criança , Citrulinação , Feminino , Histonas/sangue , Humanos , Masculino , Estudos Prospectivos , Índice de Gravidade de DoençaRESUMO
Purpose: Acute pancreatitis (AP) is an inflammatory disease. AP starts with sterile inflammation and is often complicated with critical local or systemic infection or sepsis in severe cases. Septic AP activates peptidyl arginine deiminase (PAD) and citrullinates histone H3 (CitH3), leading to neutrophil extracellular trap (NET) formation. Investigating the role of NETs and underlying mechanisms in septic AP may facilitate developing diagnostic and therapeutic approaches. In this study, we sought to identify the expression of CitH3 in septic AP patients and to analyze the correlation of CitH3 concentration with NET components as well as clinical outcomes. Methods: Seventy AP patients with or without sepsis (40 septic cases, 30 nonseptic cases) and 30 healthy volunteers were recruited in this study. Concentration of NET components (CitH3 and double-strain DNA) and key enzymes (PAD2/4) were measured. Clinical and laboratory characteristics of patients were recorded and analyzed. Results: Levels of CitH3 were elevated significantly in septic AP patients compared with those in nonseptic AP and healthy volunteers. The area under the curve (AUC, 95% confidence interval) for diagnosing septic AP was 0.93 (0.86-1.003), and the cutoff was 43.05 pg/ml. Among septic AP cases (n = 40), the concentration of CitH3 was significantly increased in those who did not survive or were admitted to the intensive care unit, when compared with that in those who survived or did not require intensive care unit. Association analysis revealed that CitH3 concentration was positively correlated with PAD2, PAD4, dsDNA concentration, and Sequential Organ Failure Assessment scores. Conclusion: CitH3 concentration increased in septic AP patients and was closely correlated with disease severity and clinical outcomes. CitH3 may potentially be a diagnostic and prognostic biomarker of septic AP.
Assuntos
Histonas/sangue , Pancreatite/sangue , Sepse/sangue , Adulto , Idoso , Biomarcadores/sangue , Citrulinação , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pancreatite/diagnóstico , Prognóstico , Proteína-Arginina Desiminase do Tipo 2/sangue , Proteína-Arginina Desiminase do Tipo 4/sangue , Sepse/diagnósticoRESUMO
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.
Assuntos
Carcinoma Hepatocelular/imunologia , Cirrose Hepática/imunologia , Neoplasias Hepáticas/imunologia , Fígado/imunologia , Neutrófilos/imunologia , Trombose/imunologia , Idoso , Antitrombina III/imunologia , Biomarcadores/sangue , Carcinoma Hepatocelular/sangue , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/patologia , Estudos de Casos e Controles , Citrulinação , DNA/sangue , Armadilhas Extracelulares/imunologia , Feminino , Histonas/sangue , Humanos , Inflamação , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/sangue , Cirrose Hepática/diagnóstico , Cirrose Hepática/patologia , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/patologia , Masculino , Pessoa de Meia-Idade , Neutrófilos/patologia , Peptídeo Hidrolases/sangue , Peptídeo Hidrolases/imunologia , Peroxidase/sangue , Trombose/sangue , Trombose/diagnóstico , Trombose/patologiaRESUMO
Histones play a key role in chromatin remodeling and gene transcription. Further, free histones in the blood act as damage-associated molecules. Administration of histones to animals results in systemic inflammatory and toxic effects. Myelin basic protein is the principal constituent element of the myelin-proteolipid sheath of axons. Abzymes (antibodies with catalytic activities) are the original features of some autoimmune diseases. In this study, electrophoretically homogeneous IgGs against H1, H2A, H2B, H3, and H4 histones and myelin basic protein (MBP) were isolated from the blood sera of multiple sclerosis (MS) patients by several affinity chromatographies. Using MALDI mass spectrometry, the sites of H1 histone cleavage by IgGs against H1, H2A, H2B, H3, H4, and MBP were determined. It was shown that IgGs against H1 split H1 at 12 sites, while the number of cleavage sites by abzymes against other histones was lower: H2A (9), H2B (7), H3 (3), and H4 (3). The minimum rate of H1 hydrolysis was observed for antibodies against H3 and H4. A high rate of hydrolysis and the maximum number of H1 hydrolysis sites (17) were found for antibodies against MBP. Only a few sites of H1 hydrolysis by anti-H1 antibodies coincided with those for IgGs against H2A, H2B, H3, H4, and MBP. Thus, the polyreactivity of complexation and the enzymatic cross-activity of antibodies against H1, four other histones, and MBP have first been shown. Since histones act as damage molecules, abzymes against histones and MBP can play a negative role in the pathogenesis of MS and probably other different diseases as well.
Assuntos
Anticorpos Catalíticos/química , Autoanticorpos/química , Histonas/química , Imunoglobulina G/química , Esclerose Múltipla/sangue , Proteína Básica da Mielina/química , Sequência de Aminoácidos , Anticorpos Catalíticos/sangue , Anticorpos Catalíticos/isolamento & purificação , Autoanticorpos/sangue , Autoanticorpos/isolamento & purificação , Sítios de Ligação , Cromatografia de Afinidade , Histonas/sangue , Histonas/imunologia , Humanos , Hidrólise , Imunoglobulina G/sangue , Imunoglobulina G/isolamento & purificação , Esclerose Múltipla/imunologia , Esclerose Múltipla/patologia , Proteína Básica da Mielina/sangue , Proteína Básica da Mielina/imunologia , Ligação Proteica , Isoformas de Proteínas/sangue , Isoformas de Proteínas/química , Isoformas de Proteínas/imunologia , Proteólise , Especificidade por SubstratoRESUMO
Coronavirus disease 19 (COVID-19) presents with disease severities of varying degree. In its most severe form, infection may lead to respiratory failure and multi-organ dysfunction. Here we study the levels and evolution of the damage associated molecular patterns (DAMPS) cell free DNA (cfDNA), extracellular histone H3 (H3) and neutrophil elastase (NE), and the immune modulators GAS6 and AXL in relation to clinical parameters, ICU scoring systems and mortality in patients (n = 100) with severe COVID-19. cfDNA, H3, NE, GAS6 and AXL were increased in COVID-19 patients compared to controls. These measures associated with occurrence of clinical events and intensive care unit acquired weakness (ICUAW). cfDNA and GAS6 decreased in time in patients surviving to 30 days post ICU admission. A decrease of 27.2 ng/mL cfDNA during ICU stay associated with patient survival, whereas levels of GAS6 decreasing more than 4.0 ng/mL associated with survival. The presence of H3 in plasma was a common feature of COVID-19 patients, detected in 38% of the patients at ICU admission. NETosis markers cfDNA, H3 and NE correlated well with parameters of tissue damage and neutrophil counts. Furthermore, cfDNA correlated with lowest p/f ratio and a lowering in cfDNA was observed in patients with ventilator-free days.
Assuntos
Biomarcadores/sangue , COVID-19/patologia , Idoso , COVID-19/mortalidade , COVID-19/virologia , Ácidos Nucleicos Livres/sangue , Estado Terminal , Feminino , Histonas/análise , Histonas/sangue , Humanos , Unidades de Terapia Intensiva , Peptídeos e Proteínas de Sinalização Intercelular/sangue , Estimativa de Kaplan-Meier , Elastase de Leucócito/sangue , Masculino , Pessoa de Meia-Idade , Prognóstico , SARS-CoV-2/isolamento & purificaçãoRESUMO
Retinal vein occlusion (RVO) is the second most common retinal vascular disorders and causes visual damage in a large population. Neutrophil extracellular traps (NETs) formation (NETosis) is an important cause of vascular diseases, however, the association between NETs related biomarkers and RVO development remained unclear. In this pilot study, a total of 77 RVO cases and 48 controls were included between Jan 2020 and July 2020. Besides, the circulating levels of three NETs related markers, cell-free DNA (cfDNA), myeloperoxidase (MPO)-DNA and citrullinated histone H3 (H3Cit), were detected in all the participants and thus the association between NETosis and RVO incidence was analyzed. Advanced assays were conducted to investigate the inflammation and thrombosis related biomarkers in RVO cases with higher or lower NETs biomarkers. When the results were considered, it was found that NETs biomarkers, including cfDNA, MPO-DNA and H3Cit, were increased in the RVO cases comparing with the controls (P < 0.05). Through the receiver operating characteristic (ROC) analyses, we found that circulating NETs related biomarkers demonstrated potential diagnostic effects for RVO and the AUCs of plasma cfDNA, MPO-DNA and H3Cit were 0.859, 0.871 and 0.928, respectively (P < 0.001). Through analyzing the correlations between circulating NETs markers and RVO stages and durations, inflammatory markers as well as thrombotic indexes, it was found that NETs were related with the RVO subtypes, inflammatory status and thrombus formation. In conclusion, the plasma NETs remnants are significantly increased in RVO cases. Besides, advanced studies demonstrate that inflammation as well as thrombus formation might be involved in this association.
Assuntos
Biomarcadores/sangue , Armadilhas Extracelulares/metabolismo , Oclusão da Veia Retiniana/epidemiologia , Idoso , Estudos de Casos e Controles , DNA/análise , Feminino , Histonas/sangue , Humanos , Incidência , Inflamação/metabolismo , Masculino , Pessoa de Meia-Idade , Peroxidase/genética , Projetos Piloto , Oclusão da Veia Retiniana/sangueRESUMO
OBJECTIVE: We want to explore the changing law of circulating histones in the acute stage of urosepsis and to find more sensitive and specific biomarkers for diagnosing urosepsis as early as possible. METHODS: Twenty healthy male New Zealand rabbits were randomly divided into 4 groups (N = 5): the control group, sham group, model group of LPS 600 µg/kg, and model group of LPS 1000 µg/kg. Heart rate (HR), respiration rate (RR), rectal temperature (T), and mean arterial pressure (MAP) were examined at 1, 3, 6, 12, and 24 hours after operation. Besides, peripheral blood cell counts (RBC, WBC, PLT, and Hb) and C reaction protein (CRP) were tested at 1, 3, and 6 hours after operation, while the levels of histone H3, MMP-9, TIMP-1, and procalcitonin (PCT) in the serum were tested at 1, 3, and 6 hours after operation by ELISA. The heart, left lung, liver, and left kidney were harvested for HE stain and observed to research the pathological change of these tissues. RESULTS: (1) The general status of rabbits: rabbits in the control and sham groups came out in 2 h after operation and regain to drink and eat in 12-24 h after operation. State of the rabbits in the control group was better than that in the sham group. Rabbits in the model groups were languid after operation and stopped to drink and eat. (2) Vital signs of rabbits: there was no statistic difference in HR (P = 0.238) and RR (P = 0.813) among all groups. MAP of the model groups decreased at 3 h postoperative, but transient (P < 0.001). The T of the LPS 1000 group decreased at 6 h postoperative (P = 0.003). (3) The change of biomarkers: H3 level of the LPS groups in the serum increased at 1 h postoperative (P < 0.01); MMP-9 of the LPS 1000 group increased at 1 h postoperative (P < 0.01); WBC of the model groups decreased at 3 h postoperative (P < 0.05); PLT of the LPS 1000 group is significantly increased at 1 h postoperative (P < 0.05); no statistic difference was found in CRP, PCT, and TIMP-1 among all groups. (4) Pathological sections: no abnormal performance was found in the control and sham groups. Glomerulus of the model groups was out of shape and necrosis with obvious renal tubule expansion. Pulmonary pathology showed alveolar septum diffuse increased and inflammatory infiltrate. Change of the LPS 1000 group was more serious than that of the LPS 600 group. CONCLUSIONS: Ligating the ureter after an injection of 1000 µg/kg LPS into the ureter of the rabbit can establish the animal model of urosepsis. Histone H3 increase immediately at 1 h postoperative and are promised to be biomarkers of urosepsis, which are more effective than WBC, CRP, and PCT.
Assuntos
Diagnóstico Precoce , Histonas/sangue , Sepse/sangue , Sepse/diagnóstico , Animais , Pressão Arterial , Temperatura Corporal , Proteína C-Reativa/metabolismo , Modelos Animais de Doenças , Contagem de Leucócitos , Lipopolissacarídeos , Masculino , Metaloproteinase 9 da Matriz/metabolismo , Especificidade de Órgãos , Contagem de Plaquetas , Pró-Calcitonina/sangue , Curva ROC , Coelhos , Sensibilidade e Especificidade , Sepse/patologia , Sepse/fisiopatologia , Inibidor Tecidual de Metaloproteinase-1/sangue , Sinais VitaisRESUMO
The outcome of intravenous thrombolysis using recombinant tissue plasminogen activator (rt-PA) is only favorable in ≈ 40% of acute ischemic stroke (AIS) patients. Moreover, in ≈ 6-8% of cases, intracerebral hemorrhage (ICH) develops. We tested whether a modification of clot lysis assay (CLA), might predict therapy outcomes and safety. In this prospective observational study, blood samples of 231 AIS patients, all receiving intravenous rt-PA, were taken before thrombolysis. Cell-free DNA (cfDNA), CLA and CLA supplemented with cfDNA and histones (mCLA) were determined from the blood samples. Stroke severity was determined by NIHSS on admission. ICH was classified according to ECASSII. Short- and long-term outcomes were defined at 7 and 90 days post-event according to ΔNIHSS and by the modified Rankin Scale, respectively. Stroke severity demonstrated a step-wise positive association with cfDNA levels, while a negative association was found with the time to reach 50% lysis (50%CLT) parameter of CLA and mCLA. ROC analysis showed improved diagnostic performance of the mCLA. Logistic regression analysis proved that 50%CLT is a predictor of short-term therapy failure, while the AUC parameter predicts ICH occurrence. A modified CLA, supplemented with cfDNA and histones, might be a promising tool to predict short-term AIS outcomes and post-lysis ICH.
Assuntos
Fibrinólise , Fibrinolíticos/uso terapêutico , AVC Isquêmico/tratamento farmacológico , Terapia Trombolítica , Trombose/tratamento farmacológico , Ativador de Plasminogênio Tecidual/uso terapêutico , Administração Intravenosa , Idoso , Ácidos Nucleicos Livres/sangue , Hemorragia Cerebral/diagnóstico , Hemorragia Cerebral/etiologia , Feminino , Fibrinolíticos/administração & dosagem , Histonas/sangue , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Proteínas Recombinantes/uso terapêutico , Índice de Gravidade de Doença , Terapia Trombolítica/efeitos adversos , Ativador de Plasminogênio Tecidual/administração & dosagem , Resultado do TratamentoRESUMO
OBJECTIVE: Beyond reducing inflammation and troponin T (TnT) release, the interleukin-6 receptor antagonist tocilizumab reduces neutrophil counts in patients with non-ST segment elevation myocardial infarction (NSTEMI). It is unclear if this is related to formation of neutrophil extracellular traps (NETs), carrying inflammatory and thrombotic properties. METHODS: In a placebo-controlled trial, 117 patients with NSTEMI were randomised to a single dose of tocilizumab (n=58) or placebo (n=59) before coronary angiography. The NETs related markers double-stranded DNA (dsDNA), myloperoxidase-DNA (MPO-DNA) and citrullinated histone 3 (H3Cit) were measured at five consecutive time points during hospitalisation (days 1-3). RESULTS: Our major findings were: (1) H3Cit levels were significantly higher in the tocilizumab compared with the placebo group at all time points (all p<0.05), and H3Cit area under the curve (AUC) was 2.3 fold higher in the tocilizumab compared with placebo group (p<0.0001). (2) MPO-DNA and dsDNA did not differ between the groups. (3) In both treatment arms, dsDNA AUC was associated with TnT AUC. (4) Neutrophil count AUC correlated inversely to H3Cit AUC (p=0.015) in the total population. CONCLUSIONS: In patients with NSTEMI, treatment with tocilizumab is associated with increased circulating H3Cit levels, suggesting that tocilizumab enhances NETosis. Further studies should clarify whether NETosis is a relevant side effect of tocilizumab. Regardless of tocilizumab, dsDNA associated with TnT release, indicating a link between extracellular nuclear material and myocardial injury.
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Histonas/sangue , Infarto do Miocárdio sem Supradesnível do Segmento ST/tratamento farmacológico , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores/sangue , Relação Dose-Resposta a Droga , Método Duplo-Cego , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio sem Supradesnível do Segmento ST/sangue , Troponina T/sangue , Adulto JovemRESUMO
Sepsis is defined as a life-threatening organ dysfunction caused by a dysregulated host response to infection and is associated with high morbidity and mortality. Circulating histones (CHs), a group of damage-associated molecular pattern molecules mainly derived from neutrophil extracellular traps, play a crucial role in sepsis by mediating inflammation response, organ injury and death through Toll-like receptors or inflammasome pathways. Herein, we first elucidate the molecular mechanisms of histone-induced inflammation amplification, endothelium injury and cascade coagulation activation, and discuss the close correlation between elevated level of CHs and disease severity as well as mortality in patients with sepsis. Furthermore, current state-of-the-art on anti-histone therapy with antibodies, histone-binding proteins (namely recombinant thrombomodulin and activated protein C), and heparin is summarized to propose promising approaches for sepsis treatment.
Assuntos
Alarminas/sangue , Anti-Inflamatórios/farmacologia , Histonas/sangue , Sepse/diagnóstico , Alarminas/antagonistas & inibidores , Alarminas/metabolismo , Animais , Anti-Inflamatórios/uso terapêutico , Biomarcadores/sangue , Biomarcadores/metabolismo , Coagulação Sanguínea/efeitos dos fármacos , Coagulação Sanguínea/imunologia , Modelos Animais de Doenças , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/imunologia , Endotélio Vascular/patologia , Armadilhas Extracelulares/efeitos dos fármacos , Armadilhas Extracelulares/imunologia , Armadilhas Extracelulares/metabolismo , Heparina/farmacologia , Heparina/uso terapêutico , Histonas/antagonistas & inibidores , Histonas/metabolismo , Humanos , Terapia de Alvo Molecular/métodos , Prognóstico , Proteína C/farmacologia , Proteína C/uso terapêutico , Proteínas Recombinantes/farmacologia , Proteínas Recombinantes/uso terapêutico , Sepse/sangue , Sepse/tratamento farmacológico , Sepse/imunologia , Índice de Gravidade de Doença , Transdução de Sinais/imunologia , Trombomodulina/uso terapêuticoRESUMO
Alteration of epigenetic modifications plays an important role in human cancer. Notably, the dysregulation of histone post-translational modifications (PTMs) has been associated with several cancers including colorectal cancer (CRC). However, the signature of histone PTMs on circulating nucleosomes is still not well described. We have developed a fast and robust enrichment method to isolate circulating nucleosomes from plasma for further downstream proteomic analysis. This method enabled us to quantify the global alterations of histone PTMs from 9 CRC patients and 9 healthy donors. Among 54 histone proteoforms identified and quantified in plasma samples, 13 histone PTMs were distinctive in CRC. Notably, methylation of histone H3K9 and H3K27, acetylation of histone H3 and citrullination of histone H2A1R3 were upregulated in plasma of CRC patients. A comparative analysis of paired samples identified 3 common histone PTMs in plasma and tumor tissue including the methylation and acetylation state of lysine 27 of histone H3. Moreover, we highlight for the first time that histone H2A1R3 citrulline is a modification upregulated in CRC patients. This new method presented herein allows the detection and quantification of histone variants and histone PTMs from circulating nucleosomes in plasma samples and could be used for biomarker discovery of cancer.