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1.
Int J Mol Sci ; 22(16)2021 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-34445374

RESUMO

Angiotensin II (Ang II) induces hypertension and endothelial dysfunction, but the involvement of thrombin in these responses is not clear. Here, we assessed the effects of the inhibition of thrombin activity by dabigatran on Ang II-induced hypertension and endothelial dysfunction in mice with a particular focus on NO- and 20-HETE-dependent pathways. As expected, dabigatran administration significantly delayed thrombin generation (CAT assay) in Ang II-treated hypertensive mice, and interestingly, it prevented endothelial dysfunction development, but it did not affect elevated blood pressure nor excessive aortic wall thickening. Dabigatran's effects on endothelial function in Ang II-treated mice were evidenced by improved NO-dependent relaxation in the aorta in response to acetylcholine in vivo (MRI measurements) and increased systemic NO bioavailability (NO2- quantification) with a concomitant increased ex vivo production of endothelium-derived NO (EPR analysis). Dabigatran treatment also contributed to the reduction in the endothelial expression of pro-inflammatory vWF and ICAM-1. Interestingly, the fall in systemic NO bioavailability in Ang II-treated mice was associated with increased 20-HETE concentration in plasma (UPLC-MS/MS analysis), which was normalised by dabigatran treatment. Taking together, the inhibition of thrombin activity in Ang II-induced hypertension in mice improves the NO-dependent function of vascular endothelium and normalises the 20-HETE-depedent pathway without affecting the blood pressure and vascular remodelling.


Assuntos
Angiotensina II/efeitos adversos , Antitrombinas/administração & dosagem , Dabigatrana/administração & dosagem , Ácidos Hidroxieicosatetraenoicos/sangue , Hipertensão/metabolismo , Remodelação Vascular/efeitos dos fármacos , Animais , Antitrombinas/farmacologia , Cromatografia Líquida , Dabigatrana/farmacologia , Modelos Animais de Doenças , Hipertensão/sangue , Hipertensão/induzido quimicamente , Molécula 1 de Adesão Intercelular/metabolismo , Masculino , Camundongos , Óxido Nítrico/metabolismo , Espectrometria de Massas em Tandem , Fator de von Willebrand/metabolismo
2.
Pan Afr Med J ; 39: 13, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34394804

RESUMO

Introduction: von Willebrand Disease (vWD) is the most prevalent bleeding disorder. Women are more likely to manifest abnormal bleeding symptoms due to physiologic events and menorrhagia is the most common presenting symptom. Methods: this case-control study included 168 women aged between 18 and 45. The cases had menorrhagia whilst the controls did not. Blood grouping, activated partial thromboplastin time and von Willebrand factor activity tests were performed on samples collected from consenting study participants. Results: the mean age was 29.96 ± 7.37. Mean vWF activity of cases was 66.6% and of controls 97.8%. The mean activated Partial ThromboplastinTime (aPTT) of cases was 31.09s and of controls was 30.40s. There was no difference in the vWF activity between blood group O (86.3%) and non-blood group O (88.0%) participants. Eight women were diagnosed with von Willebrand disease, 6 cases and 2 controls. Higher odds of von Willebrand disease were seen in the cases (OR = 6.6). Epistaxis, von Willebrand and factor activity levels and family history of menorrhagia were associated with an increased risk for menorrhagia. Conclusion: von Willebrand factor activity levels were associated with menorrhagia while activated partial thromboplastin time was not. vWF activity levels did not depend on any specific blood group. The prevalence of von Willebrand disease was significantly higher in participants with menorrhagia and repeated epistaxis and family history of menorrhagia pointed to a higher risk of menorrhagia.


Assuntos
Hemorragia/etiologia , Menorragia/etiologia , Doenças de von Willebrand/diagnóstico , Fator de von Willebrand/metabolismo , Adolescente , Adulto , Biomarcadores/metabolismo , Estudos de Casos e Controles , Epistaxe/epidemiologia , Feminino , Humanos , Pessoa de Meia-Idade , Tempo de Tromboplastina Parcial , Prevalência , Adulto Jovem , Zâmbia , Doenças de von Willebrand/complicações
3.
Clin Immunol ; 229: 108794, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34245915

RESUMO

C3 glomerulopathy (C3G) is a rare renal disease characterized by predominant glomerular C3 staining. Complement alternative pathway dysregulation due to inherited complement defects is associated with C3G. To identify novel C3G-related genes, we screened 86 genes in the complement, coagulation and endothelial systems in 35 C3G patients by targeted genomic enrichment and massively parallel sequencing. Surprisingly, the most frequently mutated gene was VWF. Patients with VWF variants had significantly higher proteinuria levels, higher crescent formation and lower factor H (FH) levels. We further selected two VWF variants to transiently express the von Willebrand factor (vWF) protein, we found that vWF expression from the c.1519A > G variant was significantly reduced. In vitro results further indicated that vWF could regulate complement activation, as it could bind to FH and C3b, act as a cofactor for factor I-mediated cleavage of C3b. Thus, we speculated that vWF might be involved in the pathogenesis of C3G.


Assuntos
Complemento C3/metabolismo , Glomerulonefrite Membranoproliferativa/genética , Glomerulonefrite/genética , Fator de von Willebrand/genética , Adolescente , Adulto , Estudos de Casos e Controles , China , Estudos de Coortes , Complemento C3b/metabolismo , Fator H do Complemento/metabolismo , Via Alternativa do Complemento , Feminino , Variação Genética , Glomerulonefrite/imunologia , Glomerulonefrite/patologia , Glomerulonefrite Membranoproliferativa/imunologia , Glomerulonefrite Membranoproliferativa/patologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Técnicas In Vitro , Glomérulos Renais/imunologia , Glomérulos Renais/patologia , Masculino , Pessoa de Meia-Idade , Modelos Imunológicos , Simulação de Dinâmica Molecular , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Análise de Sequência de DNA , Adulto Jovem , Fator de von Willebrand/química , Fator de von Willebrand/metabolismo
5.
Folia Histochem Cytobiol ; 59(2): 108-113, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34003485

RESUMO

INTRODUCTION: Endothelial dysfunction is a critical part of heart failure (HF) pathophysiology. It is not clear, however, whether it is present at the similar level in the early and late HF stages. MATERIAL AND METHODS: von Willebrand factor (vWF) and its mRNA levels in biopsies of non-ischemic patients with HF secondary to dilated cardiomyopathy were studied. Consecutive patients with HF were divided into two groups: group A with disease duration ≤ 12 months (n = 59) and group B with disease duration > 12 months (n = 68). The immunoreactivity of the vWF was compared with autopsy sections of 19 control cases. Tissue vWF gene expression was analyzed at the mRNA level by RT-PCR. RESULTS: In the group A, there was lower vWF immunoreactivity in the coronary microvessels compared to the group B [1.5 (1.0-2.0) vs. 2.0 (1.5-2.4), P = 0.001]. In the control group, only weak vWF expression was observed. Protein expression was not accompanied by vWF mRNA whose levels were significantly higher in the Group A as compared to the Group B [14671 (4932-51561) vs. 3643 (185.3-9030.8), P = 0.005]. Protein vWF expression was inversely associated with its mRNA levels (r = -0.34, P = 0.04). CONCLUSIONS: High myocardial protein expression of vWF in patients with long-lasting HF symptoms may highlight the persistent nature of endothelial dysfunction in such a cohort of patients.


Assuntos
Endotélio Vascular/metabolismo , Insuficiência Cardíaca/metabolismo , Miocárdio/metabolismo , Fator de von Willebrand/metabolismo , Adulto , Vasos Coronários/metabolismo , Progressão da Doença , Feminino , Humanos , Imuno-Histoquímica , Masculino , Microvasos/metabolismo , Pessoa de Meia-Idade , Regulação para Cima
6.
Am J Hematol ; 96(8): 1049-1055, 2021 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-33991361

RESUMO

The metalloproteinase ADAMTS13 (a disintegrin with a thrombospondin type 1 motif, member 13), also known as VWF (von Willebrand factor) protease, may be assessed in a vast array of clinical conditions. Notably, a severe deficiency of ADAMTS13 characterizes TTP (thrombotic thrombocytopenic purpura), a rare but potentially fatal disorder associated with thrombosis due to accumulation of prothrombotic ultra-large VWF multimers. Although prompt identification/exclusion of TTP can be facilitated by rapid ADAMTS13 testing, the most commonly utilized assays are based on ELISA (enzyme linked immunosorbent assay) and require long turnaround time and have relatively limited throughput. Nevertheless, several rapid ADAMTS13 assays are now available, at least in select geographies. The current mini-review discusses these issues, as well as the potential utility of ADAMTS13 testing in a range of other conditions, including coronavirus disease 2019 (COVID-19).


Assuntos
Proteína ADAMTS13/sangue , COVID-19/complicações , Púrpura Trombocitopênica Trombótica/diagnóstico , SARS-CoV-2 , Proteína ADAMTS13/deficiência , Proteína ADAMTS13/imunologia , Autoanticorpos/sangue , Autoanticorpos/imunologia , COVID-19/sangue , Ensaio de Imunoadsorção Enzimática , Feminino , Transferência Ressonante de Energia de Fluorescência , Humanos , Medições Luminescentes , Masculino , Estudos Multicêntricos como Assunto , Pré-Eclâmpsia/diagnóstico , Pré-Eclâmpsia/enzimologia , Valor Preditivo dos Testes , Gravidez , Púrpura Trombocitopênica Trombótica/sangue , Púrpura Trombocitopênica Trombótica/enzimologia , Púrpura Trombocitopênica Trombótica/etiologia , Proteínas Recombinantes/metabolismo , Sensibilidade e Especificidade , Doenças de von Willebrand/diagnóstico , Doenças de von Willebrand/enzimologia , Fator de von Willebrand/metabolismo
7.
Microvasc Res ; 137: 104188, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34022205

RESUMO

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been led to a pandemic emergency. So far, different pathological pathways for SARS-CoV-2 infection have been introduced in which the excess release of pro-inflammatory cytokines (such as interleukin 1 ß [IL-1ß], IL-6, and tumor necrosis factor α [TNFα]) has earned most of the attentions. However, recent studies have identified new pathways with at least the same level of importance as cytokine storm in which endothelial cell (EC) dysfunction is one of them. In COVID-19, two main pathologic phenomena have been seen as a result of EC dysfunction: hyper-coagulation state and pathologic angiogenesis. The EC dysfunction-induced hypercoagulation state seems to be caused by alteration in the levels of different factors such as plasminogen activator inhibitor 1 (PAI-1), von Willebrand factor (vWF) antigen, soluble thrombomodulin, and tissue factor pathway inhibitor (TFPI). As data have shown, these thromboembolic events are associated with severity of disease severity or even death in COVID-19 patients. Other than thromboembolic events, pathologic angiogenesis is among the recent findings. Furthermore, over-expression/higher levels of different proangiogenic factors such as vascular endothelial growth factor (VEGF), hypoxia-inducible factor 1 α (HIF-1α), IL-6, TNF receptor super family 1A and 12, and angiotensin-converting enzyme 2 (ACE2) have been found in the lung biopsies/sera of both survived and non-survived COVID-19 patients. Also, there are some hypotheses regarding the role of nitric oxide in EC dysfunction and acute respiratory distress syndrome (ARDS) in SARS-CoV-2 infection. It has been demonstrated that different pathways involved in inflammation are generally common with EC dysfunction and angiogenesis. Altogether, considering the common possible upstream pathways in cytokine storm, pathologic angiogenesis, and EC dysfunction, it seems that targeting these molecules (such as nuclear factor κB) could be more effective in the management of patients with COVID-19.


Assuntos
COVID-19/sangue , COVID-19/fisiopatologia , Células Endoteliais/metabolismo , Neovascularização Patológica , Enzima de Conversão de Angiotensina 2/metabolismo , Biomarcadores/metabolismo , Coagulação Sanguínea , Síndrome da Liberação de Citocina , Humanos , Inflamação , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Lipoproteínas/metabolismo , Óxido Nítrico/metabolismo , Inibidor 1 de Ativador de Plasminogênio/metabolismo , SARS-CoV-2 , Fator de Necrose Tumoral alfa/metabolismo , Fator de von Willebrand/metabolismo
8.
Angiogenesis ; 24(3): 407-411, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33974165

RESUMO

BACKGROUND: Microthrombosis is a hallmark of COVID-19. We previously described von willebrand factor (VWF) and their high molecular weight multimers (HMWMs) as potential trigger of microthrombosis. OBJECTIVES: Investigate VWF activity with collagen-binding assay and ADAMTS13 in COVID-19. METHODS AND RESULTS: Our study enrolled 77 hospitalized COVID-19 patients including 37 suffering from a non-critical form and 40 with critical form. Plasma levels of VWF collagen-binding ability (VWF:CB) and ADAMTS13 activity (ADAMTS13:Act) were measured in the first 48 hours following admission. VWF:CB was increased in critical (631% IQR [460-704]) patients compared to non-critical patients (259% [235-330], p < 0.005). VWF:CB was significantly associated (r = 0.564, p < 0.001) with HMWMs. Moreover, median ADAMTS13:Act was lower in critical (64.8 IU/dL IQR 50.0-77.7) than non-critical patients (85.0 IU/dL IQR 75.8-94.7, p < 0.001), even if no patients displayed majors deficits. VWF:Ag-to-ADAMTS13:Act ratio was highly associated with VWF:CB (r = 0.916, p < 0.001). Moreover, VWF:CB level was highly predictive of COVID-19 in-hospital mortality as shown by the ROC curve analysis (AUC = 0.92, p < 0.0001) in which we identified a VWF:CB cut-off of 446% as providing the best predictor sensitivity-specificity balance. We confirmed this cut-off thanks to a Kaplan-Meier estimator analysis (log-rank p < 0.001) and a Cox-proportional Hazard model (HR = 49.1, 95% CI 1.81-1328.2, p = 0.021) adjusted on, BMI, C-reactive protein, and D-dimer levels. CONCLUSION: VWF:CB levels could summarize both VWF increased levels and hyper-reactivity subsequent to ADAMTS13 overflow and, therefore, be a valuable and easy to perform clinical biomarker of microthrombosis and COVID-19 severity.


Assuntos
Proteína ADAMTS13/sangue , COVID-19/sangue , COVID-19/mortalidade , Pandemias , SARS-CoV-2 , Fator de von Willebrand/metabolismo , Idoso , Biomarcadores/sangue , Colágeno/metabolismo , Estudos Transversais , Feminino , Mortalidade Hospitalar , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Paris/epidemiologia , Modelos de Riscos Proporcionais , Ligação Proteica , Índice de Gravidade de Doença
9.
Proc Natl Acad Sci U S A ; 118(11)2021 03 16.
Artigo em Inglês | MEDLINE | ID: mdl-33836597

RESUMO

Thrombomodulin (TM) is a thrombin receptor on endothelial cells that is involved in promoting activation of the anticoagulant protein C pathway during blood coagulation. TM also exerts protective anti-inflammatory properties through a poorly understood mechanism. In this study, we investigated the importance of TM signaling to cellular functions by deleting it from endothelial cells by CRISPR-Cas9 technology and analyzed the resultant phenotype of TM-deficient (TM -/- ) cells. Deficiency of TM in endothelial cells resulted in increased basal permeability and hyperpermeability when stimulated by thrombin and TNF-α. The loss of the basal barrier permeability function was accompanied by increased tyrosine phosphorylation of VE-cadherin and reduced polymerization of F-actin filaments at cellular junctions. A significant increase in basal NF-κB signaling and expression of inflammatory cell adhesion molecules was observed in TM -/- cells that resulted in enhanced adhesion of leukocytes to TM -/- cells in flow chamber experiments. There was also a marked increase in expression, storage, and release of the von Willebrand factor (VWF) and decreased storage and release of angiopoietin-2 in TM -/- cells. In a flow chamber assay, isolated platelets adhered to TM -/- cells, forming characteristic VWF-platelet strings. Increased VWF levels and inflammatory foci were also observed in the lungs of tamoxifen-treated ERcre-TMf/f mice. Reexpression of the TM construct in TM -/- cells, but not treatment with soluble TM, normalized the cellular phenotype. Based on these results, we postulate cell-bound TM endows a quiescent cellular phenotype by tightly regulating expression of procoagulant, proinflammatory, and angiogenic molecules in vascular endothelial cells.


Assuntos
Células Endoteliais/metabolismo , Endotélio Vascular/metabolismo , Trombomodulina/metabolismo , Angiopoietina-2/metabolismo , Animais , Plaquetas/citologia , Permeabilidade Capilar , Adesão Celular , Células Endoteliais/citologia , Endotélio Vascular/citologia , Humanos , Inflamação , Leucócitos/citologia , Pulmão/metabolismo , Camundongos , Receptor PAR-1/metabolismo , Trombina/metabolismo , Trombomodulina/deficiência , Trombomodulina/genética , Fator de von Willebrand/metabolismo
10.
Nat Commun ; 12(1): 2360, 2021 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-33883551

RESUMO

Von Willebrand factor (VWF) activates in response to shear flow to initiate hemostasis, while aberrant activation could lead to thrombosis. Above a critical shear force, the A1 domain of VWF becomes activated and captures platelets via the GPIb-IX complex. Here we show that the shear-responsive element controlling VWF activation resides in the discontinuous autoinhibitory module (AIM) flanking A1. Application of tensile force in a single-molecule setting induces cooperative unfolding of the AIM to expose A1. The AIM-unfolding force is lowered by truncating either N- or C-terminal AIM region, type 2B VWD mutations, or binding of a ristocetin-mimicking monoclonal antibody, all of which could activate A1. Furthermore, the AIM is mechanically stabilized by the nanobody that comprises caplacizumab, the only FDA-approved anti-thrombotic drug to-date that targets VWF. Thus, the AIM is a mechano-regulator of VWF activity. Its conformational dynamics may define the extent of VWF autoinhibition and subsequent activation under force.


Assuntos
Fator de von Willebrand/química , Fator de von Willebrand/metabolismo , Anticorpos Monoclonais/farmacologia , Fenômenos Biomecânicos , Cristalografia por Raios X , Humanos , Técnicas In Vitro , Modelos Moleculares , Mutação , Agregação Plaquetária/efeitos dos fármacos , Conformação Proteica , Domínios Proteicos , Estabilidade Proteica , Desdobramento de Proteína , Ristocetina/farmacologia , Imagem Individual de Molécula , Anticorpos de Domínio Único/farmacologia , Resistência à Tração , Fator de von Willebrand/genética
11.
Commun Biol ; 4(1): 453, 2021 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-33846500

RESUMO

The Staphylococcus aureus cell wall-anchored adhesin ClfA binds to the very large blood circulating protein, von Willebrand factor (vWF) via vWF-binding protein (vWbp), a secreted protein that does not bind the cell wall covalently. Here we perform force spectroscopy studies on living bacteria to unravel the molecular mechanism of this interaction. We discover that the presence of all three binding partners leads to very high binding forces (2000 pN), largely outperforming other known ternary complexes involving adhesins. Strikingly, our experiments indicate that a direct interaction involving features of the dock, lock and latch mechanism must occur between ClfA and vWF to sustain the extreme tensile strength of the ternary complex. Our results support a previously undescribed mechanism whereby vWbp activates a direct, ultra-strong interaction between ClfA and vWF. This intriguing interaction represents a potential target for therapeutic interventions, including synthetic peptides inhibiting the ultra-strong interactions between ClfA and its ligands.


Assuntos
Aderência Bacteriana , Proteínas de Transporte/metabolismo , Coagulase/metabolismo , Staphylococcus aureus/fisiologia , Fator de von Willebrand/metabolismo , Análise Espectral
12.
Biochemistry (Mosc) ; 86(2): 123-131, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33832411

RESUMO

Here, we propose a new approach for quantitative estimation of von Willebrand factor (vWF) exposed on the surface of endothelial cells (ECs) using the ARC1779 aptamer that interacts with the vWF A1 domain. To visualize complex formation between vWF and the aptamer, the latter was conjugated with the Cy5 fluorescent label. Cultured human umbilical vein endothelial cells (HUVEC) were stained with the ARC1779-Cy5 conjugate and imaged with a fluorescence microscope. The images were analyzed with the CellProfiler software. vWF released from the Weibel-Palade bodies was observed as bright dot-like structures of round and irregular shape, the number of which increased several times after HUVEC exposure to histamine or thrombin. Staining with ARC1779-Cy5 also revealed long filamentous vWF structures on the surface of activated HUVEC. vWF secretion by ECs is activated by the second messengers cAMP and Ca2+. There is evidence that hydrogen peroxide also acts as a second messenger in ECs. In addition, exogenous H2O2 formed in leukocytes can enter ECs. The aim of our study was to determine the effect of H2O2 on the vWF exposure at the surface of HUVEC using the proposed method. It was shown that hydrogen peroxide at concentration 100 µM, which is lower than the cytotoxicity threshold of H2O2 for cultured HUVEC, increased several times the number of dot-like structures and total amount of vWF exposed on plasma membrane of HUVEC, which suggest that H2O2 acts as a mediator that activates exocytosis of Weibel-Palade bodies and vWF secretion in the vascular endothelium during inflammation and upon elevated generation of endogenous reactive oxygen species in ECs.


Assuntos
Aptâmeros de Nucleotídeos/química , Exocitose , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Peróxido de Hidrogênio/farmacologia , Fator de von Willebrand/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/fisiologia , Humanos , Corpos de Weibel-Palade , Fator de von Willebrand/química
13.
Neurology ; 96(20): e2481-e2487, 2021 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-33795393

RESUMO

OBJECTIVE: To assess support for a causal relationship between hemostatic measures and migraine susceptibility using genetic instrumental analysis. METHODS: Two-sample Mendelian randomization instrumental analyses leveraging available genome-wide association study (GWAS) summary statistics were applied to hemostatic measures as potentially causal for migraine and its subtypes, migraine with aura (MA) and migraine without aura (MO). Twelve blood-based measures of hemostasis were examined, including plasma level or activity of 8 hemostatic factors and 2 fibrinopeptides together with 2 hemostasis clinical tests. RESULTS: There were significant instrumental effects between increased coagulation factor VIII activity (FVIII; odds ratio [95% confidence interval] 1.05 [1.03, 1.08]/SD, p = 6.08 × 10-05), von Willebrand factor level (vWF; 1.05 [1.03, 1.08]/SD, p = 2.25 × 10-06), and phosphorylated fibrinopeptide A level (1.13 [1.07, 1.19]/SD, p = 5.44 × 10-06) with migraine susceptibility. When extended to migraine subtypes, FVIII, vWF, and phosphorylated fibrinopeptide A showed slightly stronger effects with MA than overall migraine. Fibrinogen level was inversely linked with MA (0.76 [0.64, 0.91]/SD, p = 2.32 × 10-03) but not overall migraine. None of the hemostatic factors was linked with MO. In sensitivity analysis, effects for fibrinogen and phosphorylated fibrinopeptide A were robust, whereas independent effects of FVIII and vWF could not be distinguished, and FVIII associations were potentially affected by pleiotropy at the ABO locus. Causal effects from migraine to the hemostatic measures were not supported in reverse Mendelian randomization. However, MA was not included due to lack of instruments. CONCLUSIONS: The findings support potential causality of increased FVIII, vWF, and phosphorylated fibrinopeptide A and decreased fibrinogen in migraine susceptibility, especially for MA, potentially revealing etiologic relationships between hemostasis and migraine.


Assuntos
Hemostasia/genética , Enxaqueca com Aura/genética , Enxaqueca sem Aura/genética , Estudos de Casos e Controles , Fator VII/metabolismo , Fator VIII/metabolismo , Fator XI/metabolismo , Produtos de Degradação da Fibrina e do Fibrinogênio/metabolismo , Fibrinogênio/metabolismo , Fibrinopeptídeo A/metabolismo , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Coeficiente Internacional Normatizado , Análise da Randomização Mendeliana , Transtornos de Enxaqueca/sangue , Transtornos de Enxaqueca/epidemiologia , Transtornos de Enxaqueca/genética , Enxaqueca com Aura/sangue , Enxaqueca com Aura/epidemiologia , Enxaqueca sem Aura/sangue , Enxaqueca sem Aura/epidemiologia , Tempo de Tromboplastina Parcial , Inibidor 1 de Ativador de Plasminogênio/sangue , Tempo de Protrombina , Ativador de Plasminogênio Tecidual/sangue , Fator de von Willebrand/metabolismo
14.
Semin Thromb Hemost ; 47(4): 400-418, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33893632

RESUMO

von Willebrand factor (VWF) is a large adhesive multimeric protein involved in hemostasis. The larger the size (or number of VWF multimers), the greater the functionality of the protein. A deficiency or defect of VWF can lead to von Willebrand disease (VWD) and cause bleeding. Conversely, an increase in VWF may create an environment that promotes thrombosis. ADAMS-13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13), sometimes called VWF-cleaving protease, is primarily responsible for controlling the size of VWF. The most severe deficiency (<10% of normal levels) of ADAMTS-13 arises in thrombotic thrombocytopenic purpura, a condition characterized by the presence of ultralarge VWF and clinically resulting in enhanced risk of thrombosis. However, ADAMTS-13 deficiency may result from other pathological processes. Of relevance is the recent finding that COVID-19 (coronavirus disease 2019) is associated with both increased levels and activity of VWF as well as generally decreased (or occasionally normal) activity levels of ADAMTS-13. Thus, in COVID-19 there is an alteration in the VWF/ADAMTS-13 axis, most often described by increased VWF/ADAMTS-13 ratio (or reduced ADAMTS-13/VWF ratio). COVID-19 is also associated with high prothrombotic risk. Thus, the imbalance of VWF and ADAMTS-13 in COVID-19 may be providing a milieu that promotes (micro)thrombosis, in a clinical picture resembling a secondary thrombotic microangiopathy in some patients. This review therefore assesses the literature on VWF, ADAMTS-13, and COVID-19. Whenever reported in COVID-19, VWF has always been identified as raised (compared with normal reference ranges or control populations). Reports have included VWF level (i.e., VWF antigen) and in some cases one or more VWF "activity" (e.g., collagen binding; platelet glycoprotein Ib [GPIb] binding, using ristocetin cofactor or more modern versions including VWF:GPIbR [recombinant] and VWF:GPIbM [mutant]). Whenever reported, ADAMTS-13 has been reported as "normal" or reduced; however, it should be recognized that "normal" levels may still identify a relative reduction in individual cases. Some reports also discuss the raised VWF/ADAMTS-13 (or reduced ADAMTS-13/VWF) ratio, but very few provide actual numerical data.


Assuntos
Proteína ADAMTS13/sangue , COVID-19 , SARS-CoV-2/metabolismo , Trombose , Fator de von Willebrand/metabolismo , COVID-19/sangue , COVID-19/complicações , COVID-19/mortalidade , Humanos , Trombose/sangue , Trombose/etiologia , Trombose/mortalidade
15.
Clin Appl Thromb Hemost ; 27: 1076029621999099, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33835872

RESUMO

Among COVID-19 hospitalized patients, high incidence of alterations in inflammatory and coagulation biomarkers correlates with a poor prognosis. Comorbidities such as chronic degenerative diseases are frequently associated with complications in COVID-19 patients. The aim of this study was to evaluate inflammatory and procoagulant biomarkers in COVID-19 patients from a public hospital in Mexico. Blood was sampled within the first 48 h after admission in 119 confirmed COVID-19 patients that were classified in 3 groups according to oxygen demand, evolution and the severity of the disease as follows: 1) Non severe: nasal cannula or oxygen mask; 2) Severe: high flow nasal cannula and 3) Death: mechanical ventilation eventually leading to fatal outcome. Blood samples from 20 healthy donors were included as a Control Group. Analysis of inflammatory and coagulation biomarkers including D-dimer, interleukin 6, interleukin 8, PAI-1, P-selectin and VWF was performed in plasma. Routine laboratory and clinical biomarkers were also included and compared among groups. Concentrations of D-dimer (14.5 ± 13.8 µg/ml) and PAI-1 (1223 ± 889.6 ng/ml) were significantly elevated in severe COVID-19 patients (P < 0.0001). A significant difference was found in interleukin-6, PAI-1 and P-selectin in non-severe and healthy donors when compared to Severe COVID-19 and deceased patients (P < 0.001). VWF levels were also significantly different between severe patients (153.5 ± 24.3 UI/dl) and non-severe ones (133.9 ± 20.2 UI/dl) (P < 0.0001). WBC and glucose levels were also significantly elevated in patients with Severe COVID-19. Plasma concentrations of all prothrombotic biomarkers were significantly higher in patients with a fatal outcome.


Assuntos
Biomarcadores/sangue , COVID-19/sangue , Mediadores da Inflamação/sangue , SARS-CoV-2 , Adulto , Idoso , COVID-19/complicações , COVID-19/epidemiologia , Estudos de Casos e Controles , Feminino , Produtos de Degradação da Fibrina e do Fibrinogênio/metabolismo , Hospitalização , Humanos , Interleucina-6/sangue , Masculino , México/epidemiologia , Pessoa de Meia-Idade , Selectina-P/sangue , Pandemias , Inibidor 1 de Ativador de Plasminogênio/sangue , Prognóstico , Índice de Gravidade de Doença , Trombose/sangue , Trombose/etiologia , Fator de von Willebrand/metabolismo
16.
Int J Mol Sci ; 22(8)2021 Apr 18.
Artigo em Inglês | MEDLINE | ID: mdl-33919627

RESUMO

The antiphospholipid syndrome (APS) is characterized by thrombosis and/or pregnancy morbidity with the persistent presence of antiphospholipid antibodies (aPLs). Laboratory criteria for the classification of APS include the detection of lupus anticoagulant (LAC), anti-cardiolipin (aCL) antibodies and anti-ß2glycoprotein I (aß2GPI) antibodies. Clinical criteria for the classification of thrombotic APS include venous and arterial thrombosis, along with microvascular thrombosis. Several aPLs, including LAC, aß2GPI and anti-phosphatidylserine/prothrombin antibodies (aPS/PT) have been associated with arterial thrombosis. The Von Willebrand Factor (VWF) plays an important role in arterial thrombosis by mediating platelet adhesion and aggregation. Studies have shown that aPLs antibodies present in APS patients are able to increase the risk of arterial thrombosis by upregulating the plasma levels of active VWF and by promoting platelet activation. Inflammatory reactions induced by APS may also provide a suitable condition for arterial thrombosis, mostly ischemic stroke and myocardial infarction. The presence of other cardiovascular risk factors can enhance the effect of aPLs and increase the risk for thrombosis even more. These factors should therefore be taken into account when investigating APS-related arterial thrombosis. Nevertheless, the exact mechanism by which aPLs can cause thrombosis remains to be elucidated.


Assuntos
Síndrome Antifosfolipídica/imunologia , Síndrome Antifosfolipídica/metabolismo , Plaquetas/metabolismo , Trombose/metabolismo , Fator de von Willebrand/metabolismo , Animais , Anticorpos Antifosfolipídeos/imunologia , Anticorpos Antifosfolipídeos/metabolismo , Feminino , Humanos , Masculino , Trombose/imunologia
17.
Mar Drugs ; 19(4)2021 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-33806251

RESUMO

The von Willebrand factor type D (VWD) domain in vitellogenin has recently been found to bind tetrodotoxin. The way in which this protein domain associates with tetrodotoxin and participates in transporting tetrodotoxin in vivo remains unclear. A cDNA fragment of the vitellogenin gene containing the VWD domain from pufferfish (Takifugu flavidus) (TfVWD) was cloned. Using in silico structural and docking analyses of the predicted protein, we determined that key amino acids (namely, Val115, ASP116, Val117, and Lys122) in TfVWD mediate its binding to tetrodotoxin, which was supported by in vitro surface plasmon resonance analysis. Moreover, incubating recombinant rTfVWD together with tetrodotoxin attenuated its toxicity in vivo, further supporting protein-toxin binding and indicating associated toxicity-neutralizing effects. Finally, the expression profiling of TfVWD across different tissues and developmental stages indicated that its distribution patterns mirrored those of tetrodotoxin, suggesting that TfVWD may be involved in tetrodotoxin transport in pufferfish. For the first time, this study reveals the amino acids that mediate the binding of TfVWD to tetrodotoxin and provides a basis for further exploration of the molecular mechanisms underlying the enrichment and transfer of tetrodotoxin in pufferfish.


Assuntos
Proteínas de Peixes/metabolismo , Takifugu/metabolismo , Tetrodotoxina/metabolismo , Vitelogeninas/metabolismo , Fator de von Willebrand/metabolismo , Animais , Proteínas de Peixes/genética , Simulação de Acoplamento Molecular , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Vitelogeninas/genética , Fator de von Willebrand/genética
18.
J Mol Biol ; 433(13): 166954, 2021 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-33771572

RESUMO

Von Willebrand Factor (vWF), a 300-kDa plasma protein key to homeostasis, is cleaved at a single site by multi-domain metallopeptidase ADAMTS-13. vWF is the only known substrate of this peptidase, which circulates in a latent form and becomes allosterically activated by substrate binding. Herein, we characterised the complex formed by a competent peptidase construct (AD13-MDTCS) comprising metallopeptidase (M), disintegrin-like (D), thrombospondin (T), cysteine-rich (C), and spacer (S) domains, with a 73-residue functionally relevant vWF-peptide, using nine complementary techniques. Pull-down assays, gel electrophoresis, and surface plasmon resonance revealed tight binding with sub-micromolar affinity. Cross-linking mass spectrometry with four reagents showed that, within the peptidase, domain D approaches M, C, and S. S is positioned close to M and C, and the peptide contacts all domains. Hydrogen/deuterium exchange mass spectrometry revealed strong and weak protection for C/D and M/S, respectively. Structural analysis by multi-angle laser light scattering and small-angle X-ray scattering in solution revealed that the enzyme adopted highly flexible unbound, latent structures and peptide-bound, active structures that differed from the AD13-MDTCS crystal structure. Moreover, the peptide behaved like a self-avoiding random chain. We integrated the results with computational approaches, derived an ensemble of structures that collectively satisfied all experimental restraints, and discussed the functional implications. The interaction conforms to a 'fuzzy complex' that follows a 'dynamic zipper' mechanism involving numerous reversible, weak but additive interactions that result in strong binding and cleavage. Our findings contribute to illuminating the biochemistry of the vWF:ADAMTS-13 axis.


Assuntos
Proteína ADAMTS13/metabolismo , Processamento de Proteína Pós-Traducional , Fator de von Willebrand/química , Fator de von Willebrand/metabolismo , Reagentes para Ligações Cruzadas/química , Humanos , Cinética , Modelos Moleculares , Peptídeos/química , Ligação Proteica , Soluções , Fator de von Willebrand/isolamento & purificação
20.
Eur J Pharmacol ; 898: 173994, 2021 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-33675784

RESUMO

Disintegrin and metalloproteinase 28 (ADAM28) is a member of the disintegrin and metalloprotease domain (ADAM) family. It is associated with the growth and metastasis of various malignancies in vivo, but its role in gastric cancer remains unclear. The purpose of this study was to investigate the effect of ADAM28 derived from gastric cancer and endothelium on gastric cancer cells and its related mechanisms. In this study, Western blot analysis and q-PCR results showed that ADAM28 was up-regulated in gastric cancer cell lines. The TCGA database showed that patients with high ADAM28 expression had significantly shorter overall survival than those with low ADAM28 expression. By MTT analysis, wound healing assay, and flow cytometry, we found that overexpression/knockdown of ADAM28 expression in gastric cancer cells can regulate cell proliferation, apoptosis and migration in vitro. In addition, overexpression/knockdown of ADAM28 in human umbilical vein endothelial cells (HUVECs) in the upper ventricle can regulate the apoptosis of lower ventricular gastric cancer cells in the co-culture system. Furthermore, ELISA demonstrated that knockdown of ADAM28 from endothelial cells increased the expression of von Willebrand Factor (vWF) in the supernatant. We found that ADAM28 both from gastric cancer cells and HUVECs eliminated vWF-induced apoptosis of gastric cancer cells by cleaving vWF, and the addition of the vWF knockdown plasmid eliminated the increase of integrin ß3, p-TP53 and c-Casp3 caused by ADAM28 knockdown. In conclusion, ADAM28 from endothelium and gastric cancer may cleave vWF to eliminate vWF-induced apoptosis of gastric cancer cells and play an pro-metastasis effect.


Assuntos
Proteínas ADAM/metabolismo , Apoptose , Células Endoteliais da Veia Umbilical Humana/enzimologia , Comunicação Parácrina , Neoplasias Gástricas/enzimologia , Fator de von Willebrand/metabolismo , Proteínas ADAM/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Técnicas de Cocultura , Regulação Neoplásica da Expressão Gênica , Humanos , Invasividade Neoplásica , Transdução de Sinais , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia
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