RESUMEN
Migrasomes are organelles that are generated by migrating cells. Here we report the key role of neutrophil-derived migrasomes in haemostasis. We found that a large number of neutrophil-derived migrasomes exist in the blood of mice and humans. Compared with neutrophil cell bodies and platelets, these migrasomes adsorb and enrich coagulation factors on the surface. Moreover, they are highly enriched with adhesion molecules, which enable them to preferentially accumulate at sites of injury, where they trigger platelet activation and clot formation. Depletion of neutrophils, or genetic reduction of the number of these migrasomes, significantly decreases platelet plug formation and impairs coagulation. These defects can be rescued by intravenous injection of purified neutrophil-derived migrasomes. Our study reveals neutrophil-derived migrasomes as a previously unrecognized essential component of the haemostasis system, which may shed light on the cause of various coagulation disorders and open therapeutic possibilities.
Asunto(s)
Coagulación Sanguínea , Plaquetas , Ratones Endogámicos C57BL , Neutrófilos , Neutrófilos/metabolismo , Animales , Humanos , Plaquetas/metabolismo , Ratones , Hemostasis , Movimiento Celular , Activación Plaquetaria , Masculino , Factores de Coagulación Sanguínea/metabolismo , Factores de Coagulación Sanguínea/genéticaRESUMEN
Pro- and anticoagulant factors are core components of hemostasis [...].
Asunto(s)
Factores de Coagulación Sanguínea , Humanos , Factores de Coagulación Sanguínea/metabolismo , Factores de Coagulación Sanguínea/genética , Anticoagulantes/uso terapéutico , Anticoagulantes/farmacología , Coagulación Sanguínea , Hemostasis , AnimalesRESUMEN
BACKGROUND AND OBJECTIVES: Obesity is hypothesized to induce a hypercoagulable state that increases stroke risk. The molecular mechanisms underlying this association are largely uncharacterized. We aimed to apply mendelian randomization to identify whether the association of genetically proxied body mass index (BMI) with cardioembolic stroke risk is mediated by changes in levels of circulating coagulation factors. METHODS: Genetic proxies for BMI and levels of circulating coagulation factors were obtained, respectively, from the Genetic Investigation of ANthropometric Traits consortium (n = 694,649) and deCODE cohort (n = 35,559). Genetic associations with cardioembolic stroke risk were obtained from the GIGASTROKE consortium (10,804 cases and 1,234,804 controls). We performed a two-sample mendelian randomization analysis testing the association of genetically proxied BMI with cardioembolic stroke risk, genetically proxied BMI with levels of coagulation factors, and genetically proxied levels of coagulation factors with cardioembolic stroke risk. These estimates were carried forward to mediation and sensitivity analyses. RESULTS: A 1-SD increase in genetically proxied BMI associated with increased cardioembolic stroke risk (OR of cardioembolic stroke per 1-SD of BMI 1.20, 95% CI 1.08-1.33, p = 8.65 × 10-4) with similar findings in statistical sensitivity analyses more robust to the inclusion of pleiotropic variants. Genetically proxied BMI was further associated with increased levels of Factor VII, Factor Xa, Factor XI, and Protein S (all p < 5.9 × 10-6). Of these factors, genetically proxied levels of Factor XI were associated with cardioembolic stroke risk (OR of cardioembolic stroke per 1-SD increase in Factor XI levels 1.32, 1.19-1.46, p = 6.18 × 10-8). The mediated effect of genetically proxied BMI through Factor XI accounted for 26% (6%-49%) of the total effect of BMI on cardioembolic stroke. DISCUSSION: Human genetic data support increased levels of Factor XI as a mechanistic explanation for how obesity increases cardioembolic stroke risk. The clinical relevance of this association warrants further investigation within ongoing clinical trials of Factor XI inhibition.
Asunto(s)
Índice de Masa Corporal , Análisis de la Aleatorización Mendeliana , Obesidad , Trombofilia , Humanos , Obesidad/genética , Obesidad/complicaciones , Obesidad/sangre , Obesidad/epidemiología , Trombofilia/genética , Trombofilia/sangre , Accidente Cerebrovascular/genética , Accidente Cerebrovascular/epidemiología , Accidente Cerebrovascular/sangre , Femenino , Factores de Coagulación Sanguínea/genética , Masculino , Factores de Riesgo , Accidente Cerebrovascular Embólico/genética , Accidente Cerebrovascular Embólico/epidemiologíaRESUMEN
Previous genetic studies of venous thromboembolism (VTE) have been largely limited to common variants, leaving the genetic determinants relatively incomplete. We performed an exome-wide association study of VTE among 14,723 cases and 334,315 controls. Fourteen known and four novel genes (SRSF6, PHPT1, CGN, and MAP3K2) were identified through protein-coding variants, with broad replication in the FinnGen cohort. Most genes we discovered exhibited the potential to predict future VTE events in longitudinal analysis. Notably, we provide evidence for the additive contribution of rare coding variants to known genome-wide polygenic risk in shaping VTE risk. The identified genes were enriched in pathways affecting coagulation and platelet activation, along with liver-specific expression. The pleiotropic effects of these genes indicated the potential involvement of coagulation factors, blood cell traits, liver function, and immunometabolic processes in VTE pathogenesis. In conclusion, our study unveils the valuable contribution of protein-coding variants in VTE etiology and sheds new light on its risk stratification.
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Tromboembolia Venosa , Humanos , Tromboembolia Venosa/genética , Factores de Riesgo , Factores de Coagulación Sanguínea/genética , Exoma , Estudio de Asociación del Genoma Completo , Factores de Empalme Serina-Arginina/genética , Fosfoproteínas/genéticaRESUMEN
BACKGROUND: The procoagulant phenotype in cancer is linked to thrombosis, cancer progression, and immune response. A novel treatment that reduces the risk of both thrombosis and cancer progression without excess bleeding risk remains to be identified. OBJECTIVES: Here, we aimed to broadly investigate the breast tumor coagulome and its relation to prognosis, treatment response to chemotherapy, and the tumor microenvironment. METHODS: Key coagulation-related genes (n = 35) were studied in a Norwegian cohort with tumor (n = 134) and normal (n = 189) tissue and in the Cancer Genome Atlas (n = 1052) data set. We performed gene set variation analysis in the Norwegian cohort, and in the Cancer Genome Atlas cohort, associations with the tumor microenvironment and prognosis were evaluated. Analyses were performed with cBioPortal, Estimation of Stromal and Immune cells in Malignant Tumors Using Expression Data, Tumor Immune Estimation Resource, the integrated repository portal for tumor-immune system interactions, Tumor Immune Single-cell Hub 2, and the receiver operating characteristic plotter. Six independent breast cancer cohorts were used to study the tumor coagulome and treatment response to chemotherapy. RESULTS: Twenty-two differentially expressed coagulation-related genes were identified in breast tumors. Several coagulome factors were correlated with tumor microenvironment characteristics and were expressed by nonmalignant cells in the tumor microenvironment. PLAT and F8 were independent predictors of better overall survival and progression-free survival, respectively. F12 and PLAU were predictors of worse progression-free survival. The PROCR-THBD-PLAT signature showed a promising predictive value (area under the curve, 0.75; 95% CI, 0.69-0.81; P = 3.6 × 10-17) for combination chemotherapy with fluorouracil, epirubicin, and cyclophosphamide. CONCLUSION: The breast tumor coagulome showed potential in prediction of prognosis and chemotherapy response. Cells within the tumor microenvironment are sources of coagulome factors and may serve as therapeutic targets of coagulation factors.
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Coagulación Sanguínea , Neoplasias de la Mama , Microambiente Tumoral , Humanos , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Femenino , Coagulación Sanguínea/efectos de los fármacos , Persona de Mediana Edad , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/sangre , Resultado del Tratamiento , Noruega , Pronóstico , Regulación Neoplásica de la Expresión Génica , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Anciano , Factores de Coagulación Sanguínea/genética , AdultoRESUMEN
Remarkably, it has been 40 years since the isolation of the 2 genes involved in hemophilia A (HA) and hemophilia B (HB), encoding clotting factor (F) VIII (FVIII) and FIX, respectively. Over the years, these advances led to the development of purified recombinant protein factors that are free of contaminating viruses from human pooled plasma for hemophilia treatments, reducing the morbidity and mortality previously associated with human plasma-derived clotting factors. These discoveries also paved the way for modified factors that have increased plasma half-lives. Importantly, more recent advances have led to the development and Food and Drug Administration approval of a hepatocyte-targeted, adeno-associated viral vector-mediated gene transfer approach for HA and HB. However, major concerns regarding the durability and safety of HA gene therapy remain to be resolved. Compared with FIX, FVIII is a much larger protein that is prone to misfolding and aggregation in the endoplasmic reticulum and is poorly secreted by the mammalian cells. Due to the constraint of the packaging capacity of adeno-associated viral vector, B-domain deleted FVIII rather than the full-length protein is used for HA gene therapy. Like full-length FVIII, B-domain deleted FVIII misfolds and is inefficiently secreted. Its expression in hepatocytes activates the cellular unfolded protein response, which is deleterious for hepatocyte function and survival and has the potential to drive hepatocellular carcinoma. This review is focused on our current understanding of factors limiting FVIII secretion and the potential pathophysiological consequences upon expression in hepatocytes.
Asunto(s)
Hemofilia A , Hemofilia B , Animales , Humanos , Factor VIII/metabolismo , Hemofilia A/genética , Hemofilia A/terapia , Hemofilia A/metabolismo , Factores de Coagulación Sanguínea/genética , Terapia Genética , Hemofilia B/terapia , Hemofilia B/tratamiento farmacológico , Mamíferos/genética , Mamíferos/metabolismoRESUMEN
OBJECTIVE: To screen better promoters and provide more powerful tools for basic research and gene therapy of hemophilia. METHODS: Bioinformatics methods were used to analyze the promoters expressing housekeeping genes with high abundance, so as to select potential candidate promoters. The GFP reporter gene vector was constructed, and the packaging efficiency of the novel promoter was investigated with EF1 α promoter as control, and the transcription and activities of the reporter gene were investigated too. The activity of the candidate promoter was investigated by loading F9 gene. RESULTS: The most potential RPS6 promoter was obtained by screening. There was no difference in lentiviral packaging between EF1 α-LV and RPS6-LV, and their virus titer were consistent. In 293T cells, the transduction efficiency and mean fluorescence intensity of RPS6pro-LV and EF1 αpro-LV were proportional to the lentiviral dose. The transfection efficiency of both promoters in different types of cells was in the following order: 293T>HEL>MSC; Compared with EF1 αpro-LV, RPS6pro-LV could obtain a higher fluorescence intensity in MSC cells, and RPS6pro-LV was more stable in long-term cultured HEL cells infected with two lentiviruses respectively. The results of RT-qPCR, Western blot and FIX activity (FIXâ¶C) detection of K562 cell culture supernatant showed that FIX expression in the EF1 α-F9 and RPS6-F9 groups was higher than that in the unloaded control group, and there was no significant difference in FIX expression between the EF1 α-F9 and RPS6-F9 groups. CONCLUSION: After screening and optimization, a promoter was obtained, which can be widely used for exogenous gene expression. The high stability and viability of the promoter were confirmed by long-term culture and active gene expression, which providing a powerful tool for basic research and clinical gene therapy of hemophilia.
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Vectores Genéticos , Hemofilia A , Humanos , Transducción Genética , Hemofilia A/genética , Transfección , Factores de Coagulación Sanguínea/genética , Lentivirus/genéticaRESUMEN
Rare bleeding disorders (RBDs), including factor (F) I, FII, FV, FVII, combined FV and FVIII (CF5F8), FXI, FXIII and vitamin-K dependent coagulation factors (VKCF) deficiencies, are a heterogeneous group of hemorrhagic disorder with a variable bleeding tendency. RBDs are due to mutation in underlying coagulation factors genes, except for CF5F8 and VKCF deficiencies. FVII deficiency is the most common RBD with >330 variants in the F7 gene, while only 63 variants have been identified in the F2 gene. Most detected variants in the affected genes are missense (>50% of all RBDs), while large deletions are the rarest, having been reported in FVII, FX, FXI and FXIII deficiencies. Most were located in the catalytic and activated domains of FXI, FX, FXIII and prothrombin deficiencies. Understanding the proper molecular basis of RBDs not only can help achieve a timely and cost-effective diagnosis, but also can help to phenotype properties of the disorders.
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Trastornos de la Coagulación Sanguínea Heredados , Trastornos de la Coagulación Sanguínea , Trastornos de las Proteínas de Coagulación , Trastornos Hemorrágicos , Humanos , Trastornos de la Coagulación Sanguínea Heredados/diagnóstico , Trastornos de la Coagulación Sanguínea Heredados/genética , Trastornos de la Coagulación Sanguínea Heredados/terapia , Factores de Coagulación Sanguínea/genética , Hemorragia/etiología , Hemorragia/genética , Vitamina KRESUMEN
Vertebrate blood coagulation is controlled by a cascade containing more than 20 proteins. The cascade proteins are found in the blood in their zymogen forms and when the cascade is triggered by tissue damage, zymogens are activated and in turn activate their downstream proteins by serine protease activity. In this study, we examined proteomes of 21 chordates, of which 18 are vertebrates, to reveal the modular evolution of the blood coagulation cascade. Additionally, two Arthropoda species were used to compare domain arrangements of the proteins belonging to the hemolymph clotting and the blood coagulation cascades. Within the vertebrate coagulation protein set, almost half of the studied proteins are shared with jawless vertebrates. Domain similarity analyses revealed that there are multiple possible evolutionary trajectories for each coagulation protein. During the evolution of higher vertebrate clades, gene and genome duplications led to the formation of other coagulation cascade proteins.
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Factores de Coagulación Sanguínea , Cordados , Animales , Factores de Coagulación Sanguínea/genética , Factores de Coagulación Sanguínea/metabolismo , Vertebrados/genética , Coagulación Sanguínea/genética , Cordados/genética , GenomaRESUMEN
BACKGROUND: Prothrombin, protein C, and factors VII, IX, and X are vitamin K (VK)-dependent coagulation proteins that play an important role in the initiation, amplification, and subsequent attenuation of the coagulation response. Blood coagulation evolved in the common vertebrate ancestor as a specialization of the complement system and immune response, which in turn bear close evolutionary ties with developmental enzyme cascades. There is currently no comprehensive analysis of the evolutionary changes experienced by these coagulation proteins during the radiation of vertebrates and little is known about conservation of residues that are important for zymogen activation and catalysis. OBJECTIVES: To characterize the conservation level of functionally important residues among VK-dependent coagulation proteins from different vertebrate lineages. METHODS: The conservation level of residues important for zymogen activation and catalysis was analyzed in >1600 primary sequences of VK-dependent proteins. RESULTS: Functionally important residues are most conserved in prothrombin and least conserved in protein C. Some of the most profound functional modifications in protein C occurred in the ancestor of bony fish when the basic residue in the activation site was replaced by an aromatic residue. Furthermore, during the radiation of placental mammals from marsupials, protein C acquired a cysteine-rich insert that introduced an additional disulfide in the EGF1 domain and evolved a proprotein convertase cleavage site in the activation peptide linker that also became significantly elongated. CONCLUSIONS: Sequence variabilities at functionally important residues may lead to interspecies differences in the zymogen activation and catalytic properties of orthologous VK-dependent proteins.
Asunto(s)
Protrombina , Vitamina K , Embarazo , Animales , Femenino , Vitamina K/metabolismo , Protrombina/metabolismo , Proteína C , Placenta , Factores de Coagulación Sanguínea/genética , Factores de Coagulación Sanguínea/metabolismo , Análisis de Secuencia , Mamíferos/metabolismoRESUMEN
The coagulation factor 9 gene (FIX) point mutation contributes to most hemophilia B cases, providing ideal gene correction models. Here we identified the frequent mutation G20519A (R226Q) in FIX, which resulted in many severe and moderate hemophilia B patients. This study aimed to investigate the effect of HDR and base editing in correcting FIX mutant. We first constructed HEK293 and liver-derived cell lines Huh7 cells stabling carrying mutated FIX containing G20519A (HEK293-FIXmut and Huh7-FIXmut). Then, CRISPR/Cas9-based homology-directed repair (HDR) and base editing were used for the correction of this mutated point. We used Cas9 nickase (nCas9) mediated HDR and the advanced base editor ABE8e to correct G20519A and then measured the concentration and activity of FIX. Furthermore, we used the star-shaped poly(lysine) gene nanocarriers to deliver the ABE8e correction systems into HEK293-FIXmut and Huh7-FIXmut stem cells to correct mutated FIX. As a result, we found that gRNAs directed inefficient HDR in correcting G20519A. The ABE8e corrected the mutation efficiently in both HEK293-FIXmut and Huh7-FIXmut stem cells. In addition, the star-shaped poly(lysine) carriers delivered non-viral vectors into stem cells efficiently. The nanocarriers-delivered ABE8e system corrected mutated FIX in stem cells, and the stem cells secreted active FIX in high concentration. In conclusion, our study provides a potential alternative for correcting mutated FIX in hemophilia B patients.
Asunto(s)
Edición Génica , Hemofilia A , Hemofilia B , Aminohidrolasas/genética , Factores de Coagulación Sanguínea/genética , Factores de Coagulación Sanguínea/metabolismo , Sistemas CRISPR-Cas/genética , Desoxirribonucleasa I/metabolismo , Edición Génica/métodos , Células HEK293 , Hemofilia A/genética , Hemofilia A/metabolismo , Hemofilia B/genética , Hemofilia B/terapia , Humanos , Mutación , Mutación Missense , Polilisina/química , Células Madre/metabolismoRESUMEN
BACKGROUND: DFNA9 is a form of autosomal progressive sensorineural hearing loss, caused by more than 30 variants in the COCH gene. p.Pro51Ser (p.P51S) variant is characterized by late-onset functional deterioration toward bilateral severe hearing loss and vestibulopathy. Focal sclerosis on computed tomography (CT) and T2-weighted magnetic resonance imaging (MRI) signal loss of semicircular canals are presumably radiologic biomarkers of advanced otovestibular deterioration. OBJECTIVES: The aim of this study was to investigate whether these biomarkers are more frequent in cochlear implant candidates carrying the p.P51S mutation versus noncarriers. Second, the correlation between the hearing and vestibular function and carrier status was studied. Finally, the relationship between the presence of these radiologic features and the degree of hearing and vestibular deterioration was investigated. METHODS: A prospective cohort study was performed on 38 candidates for cochlear implantation in a tertiary referral center. Patients underwent pure tone audiometry, videonystagmography, video head impulse tests and vestibular-evoked myogenic potentials. In addition, three dizziness questionnaires were used. All subjects were administered CT, MRI, and molecular genetic analysis. RESULTS: Sixteen of 38 patients were carriers of the p.P51S COCH mutation. Radiologic lesions were almost exclusively observed in carriers. MRI was more sensitive in showing lesions than CT. Furthermore, p.P51S carriers showed significantly lower function on most vestibular tests, including questionnaires, than noncarriers. Patients with imaging abnormalities showed more pronounced vestibulopathy. CONCLUSION: The present study supplements previous data that endorse the hypothesis that focal sclerosis of semicircular canals are biomarkers of advanced vestibular deterioration, especially in DFNA9.
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Implantación Coclear , Implantes Cocleares , Pérdida Auditiva Sensorineural , Factores de Coagulación Sanguínea/genética , Proteínas de la Matriz Extracelular , Humanos , Mutación , Estudios Prospectivos , EsclerosisRESUMEN
Current hemostatic agents are obtained from pooled plasma from multiple donors requiring costly pathogen screening and processing. Recombinant DNA-based production represents an engineering solution that could improve supply, uniformity, and safety. Current approaches are typically for single gene candidate peptides and often employ non-human cells. We devised an approach where multiple gene products could be produced from a single population of cells. We identified gene specific Synergistic Activation Mediators (SAM) from the CRISPR/Cas9 system for targeted overexpression of coagulation factors II, VII, IX, X, and fibrinogen. The components of the CRISPR-SAM system were expressed in Human Embryonic Kidney Cells (HEK293), and single (singleplex) or multi-gene (multiplex) upregulation was assessed by quantitative RT-PCR (qRT-PCR) and protein expression by ELISA analysis. Factor II, VII, IX, and X singleplex and multiplex activation resulted in 120-4700-fold and 60-680-fold increases in gene expression, respectively. Fibrinogen sub-unit gene activation resulted in a 1700-92,000-fold increases and 80-5500-fold increases in singleplex or multiplex approaches, respectively. ELISA analysis showed a concomitant upregulation of candidate gene products. Our findings demonstrate the capability of CRISPR/Cas9 SAMs for single or multi-agent production in human cells and represent an engineering advance that augments current recombinant peptide production techniques.
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Factores de Coagulación Sanguínea , Sistemas CRISPR-Cas , Factores de Coagulación Sanguínea/biosíntesis , Factores de Coagulación Sanguínea/genética , Fibrinógeno/genética , Edición Génica/métodos , Células HEK293 , Humanos , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/genética , Activación TranscripcionalRESUMEN
Hemostasis is a complex and tightly regulated system that attempts to maintain a homeostatic balance to permit normal blood flow, without bleeding or thrombosis. Hemostasis reflects the subtle balance between procoagulant and anticoagulant factors in the pathways of primary hemostasis, secondary hemostasis, and fibrinolysis. The major components in this interplay include the vascular endothelium, platelets, coagulation factors, and fibrinolytic factors. After vessel wall injury, the subendothelium is exposed to the blood stream, followed by rapid activation of platelets via collagen binding and von Willebrand factor-mediated platelet adhesion to the damaged vessel wall through platelet glycoprotein receptor Ib/IX/V. Activated platelets change their shape, release bioactive molecules from their granules, and expose negatively charged phospholipids on their surface. For a proper function of this process, an adequate number of functional platelets are required. Subsequently, a rapid generation of sufficient amounts of thrombin begins; followed by activation of the coagulation system and its coagulation factors (secondary hemostasis), generating fibrin that consolidates the platelet plug. To maintain equilibrium between coagulation and anticoagulation, the naturally occurring anticoagulants such as protein C, protein S, and antithrombin keep this process in balance. Deficiencies (inherited or acquired) at any level of this fine-tuned system result in pathologic bleedings or increased hypercoagulability states leading to thrombosis. This review will focus on genetic diagnosis of inherited bleeding, thrombotic, and platelet disorders, discussing strengths and limitations of existing diagnostic settings and genetic tools and highlight some important considerations necessary for clinical application.
Asunto(s)
Trastornos de las Plaquetas Sanguíneas , Trombosis , Humanos , Proteína S/metabolismo , Factor de von Willebrand/metabolismo , Trombina/metabolismo , Proteína C , Hemostasis/genética , Trastornos de las Plaquetas Sanguíneas/genética , Trastornos de las Plaquetas Sanguíneas/metabolismo , Trombosis/metabolismo , Plaquetas/metabolismo , Factores de Coagulación Sanguínea/genética , Factores de Coagulación Sanguínea/metabolismo , Hemorragia/genética , Fibrina/metabolismo , Anticoagulantes , Glicoproteínas de Membrana Plaquetaria/metabolismo , Antitrombinas/metabolismo , Fosfolípidos/metabolismo , Colágeno/metabolismoRESUMEN
As per the classical view of the coagulation system, it functions solely in plasma to maintain hemostasis. An experimental approach modeling vascular reconstitution was used to show that vascular endothelial cells (ECs) endogenously synthesize coagulation factors during angiogenesis. Intracellular thrombin generated from this synthesis promotes the mitotic function of vascular endothelial cell growth factor A (VEGF-A). The thrombin concurrently cleaves C5a from EC-synthesized complement component C5 and unmasks the tethered ligand for EC-expressed protease-activated receptor 4 (PAR4). The two ligands jointly trigger EC C5a receptor-1 (C5ar1) and PAR4 signaling, which together promote VEGF receptor 2 growth signaling. C5ar1 is functionally associated with PAR4, enabling C5a or thrombin to elicit Gαi and/or Gαq signaling. EC coagulation factor and EC complement component synthesis concurrently down-regulate with contact inhibition. The connection of these processes with VEGF receptor 2 signaling provides new insights into mechanisms underlying angiogenesis. Knowledge of endogenous coagulation factor/complement component synthesis and joint PAR4/C5ar1 signaling could be applied to other cell types.
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Factores de Coagulación Sanguínea/biosíntesis , Células Endoteliales/metabolismo , Neovascularización Fisiológica , Receptor de Anafilatoxina C5a/metabolismo , Receptores Proteinasa-Activados/metabolismo , Transducción de Señal , Animales , Factores de Coagulación Sanguínea/genética , Femenino , Masculino , Ratones , Ratones Noqueados , Receptor de Anafilatoxina C5a/genética , Receptores Proteinasa-Activados/genéticaRESUMEN
INTRODUCTION: Hemophilia comprises a group of X-linked hemorrhagic disorders that result from a deficiency of coagulation factors. The disorder affects mainly males and leads to chronic pain, joint deformity, reduced mobility, and increased mortality. Current therapies require frequent administration of replacement clotting factors, but the emergence of alloantibodies (inhibitors) diminishes their efficacy. New therapies are being developed to produce the deficient clotting factors and prevent the emergence of inhibitors. AREAS COVERED: This article provides an update on the characteristics and disease pathophysiology of hemophilia A, as well as current treatments, with a special focus on ongoing clinical trials related to gene replacement therapies. EXPERT OPINION: Gene replacement therapies provide safe, durable, and stable transgene expression while avoiding the challenges of clotting factor replacement therapies in patients with hemophilia. Improving the specificity of the viral construct and decreasing the therapeutic dose are critical toward minimizing cellular stress, induction of the unfolded protein response, and the resulting loss of protein production in liver cells. Next-generation gene therapies incorporating chimeric DNA sequences in the transgene can increase clotting factor synthesis and secretion, and advance the efficacy, safety, and durability of gene replacement therapy for hemophilia A as well as other blood clotting disorders.
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Hemofilia A , Hemofilia B , Factores de Coagulación Sanguínea/genética , Factor VIII/uso terapéutico , Terapia Genética/tendencias , Hemofilia A/genética , Hemofilia A/terapia , Hemofilia B/tratamiento farmacológico , Hemofilia B/terapia , Humanos , Isoanticuerpos/genética , Masculino , TransgenesRESUMEN
In a previous study, we reported that human endothelial cells (ECs) express and produce their own coagulation factors (F) that can activate cell surface FX without the additions of external proteins or phospholipids. We now describe experiments that detail the expression and production in ECs and fibroblasts of the clotting proteins necessary for formation of active prothrombinase (FV-FX) complexes to produce thrombin on EC and fibroblast surfaces. EC and fibroblast thrombin generation was identified by measuring: thrombin activity; thrombin-antithrombin complexes; and the prothrombin fragment 1.2 (PF1.2), which is produced by the prothrombinase cleavage of prothrombin (FII) to thrombin. In ECs, the prothrombinase complex uses surface-attached FV and γ-carboxyl-glutamate residues of FX and FII to attach to EC surfaces. FV is also on fibroblast surfaces; however, lower fibroblast expression of the gene for γ-glutamyl carboxylase (GGCX) results in production of vitamin K-dependent coagulation proteins (FII and FX) with reduced surface binding. This is evident by the minimal surface binding of PF1.2, following FII activation, of fibroblasts compared to ECs. We conclude that human ECs and fibroblasts both generate thrombin without exogenous addition of coagulation proteins or phospholipids. The two cell types assemble distinct forms of prothrombinase to generate thrombin.
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Factores de Coagulación Sanguínea/biosíntesis , Factores de Coagulación Sanguínea/genética , Células Endoteliales/metabolismo , Fibroblastos/metabolismo , Trombina/biosíntesis , Antitrombina III/genética , Antitrombina III/metabolismo , Carbohidrato Epimerasas/biosíntesis , Carboxiliasas/genética , Línea Celular , Factor V/genética , Factor V/metabolismo , Factor Xa/metabolismo , Expresión Génica , Células Endoteliales de la Vena Umbilical Humana , Humanos , Cetona Oxidorreductasas/biosíntesis , Modelos Biológicos , Fragmentos de Péptidos/metabolismo , Proteolisis , Protrombina/biosíntesis , Protrombina/genética , Protrombina/metabolismo , Trombomodulina/genética , Trombomodulina/metabolismo , Vitamina K Epóxido Reductasas/genéticaRESUMEN
Although metabolic syndrome (MetS) is linked to an elevated risk of cardiovascular disease (CVD), the cardiac-specific risk mechanism is unknown. Obesity, hypertension, and diabetes (all MetS components) are the most common form of CVD and represent risk factors for worse COVID-19 outcomes compared to their non MetS peers. Here, we use obese Yorkshire pigs as a highly relevant animal model of human MetS, where pigs develop the hallmarks of human MetS and reproducibly mimics the myocardial pathophysiology in patients. Myocardium-specific mass spectroscopy-derived metabolomics, proteomics, and transcriptomics enabled the identity and quality of proteins and metabolites to be investigated in the myocardium to greater depth. Myocardium-specific deregulation of pro-inflammatory markers, propensity for arterial thrombosis, and platelet aggregation was revealed by computational analysis of differentially enriched pathways between MetS and control animals. While key components of the complement pathway and the immune response to viruses are under expressed, key N6-methyladenosin RNA methylation enzymes are largely overexpressed in MetS. Blood tests do not capture the entirety of metabolic changes that the myocardium undergoes, making this analysis of greater value than blood component analysis alone. Our findings create data associations to further characterize the MetS myocardium and disease vulnerability, emphasize the need for a multimodal therapeutic approach, and suggests a mechanism for observed worse outcomes in MetS patients with COVID-19 comorbidity.
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COVID-19/patología , Susceptibilidad a Enfermedades , Síndrome Metabólico/patología , Animales , Factores de Coagulación Sanguínea/genética , Factores de Coagulación Sanguínea/metabolismo , COVID-19/complicaciones , COVID-19/virología , Ciclooxigenasa 2/genética , Ciclooxigenasa 2/metabolismo , Dieta Alta en Grasa/veterinaria , Modelos Animales de Enfermedad , Humanos , Inmunidad Innata/genética , Síndrome Metabólico/complicaciones , Síndrome Metabólico/metabolismo , Metiltransferasas/genética , Metiltransferasas/metabolismo , Miocardio/metabolismo , Estrés Oxidativo/genética , Agregación Plaquetaria , Receptores Purinérgicos P2Y1/genética , Receptores Purinérgicos P2Y1/metabolismo , Sistema Renina-Angiotensina , Factores de Riesgo , SARS-CoV-2/aislamiento & purificación , Porcinos , Activador de Plasminógeno de Tipo Uroquinasa/genética , Activador de Plasminógeno de Tipo Uroquinasa/metabolismoRESUMEN
Depression is a severe disabling psychiatric illness and the pathophysiological mechanisms remain unknown. In previous work, we found the changes in extrinsic coagulation (EC) pathway proteins in depressed patients compared with healthy subjects were significant. In this study, we screened differentially expressed proteins (DEPs) in the EC pathway, and explored the molecular mechanism by constructing a protein-protein interaction (PPI) network. The DEPs of the EC pathwaywere initially screened by isobaric tags for relative and absolute quantification (iTRAQ) in plasma samples obtained from 20 depression patients and 20 healthy controls, and were then identified by Enzyme-linked immunosorbent assays (ELISAs). Ingenuity Pathway Analysis (IPA) software was used to analyse pathway. The differentially expressed genes (DEGs) were identified by analyzing the GSE98793 microarray data from the Gene Expression Omnibus database using the Significance Analysis for Microarrays (SAM, version 4.1) statistical method. Cytoscape version 3.4.0 software was used to construct and visualize PPI networks. The results show that Fibrinogen alpha chain (FGA), Fibrinogen beta chain (FGB), Fibrinogen gamma chain (FGG) and Coagulation factor VII (FVII) were screened in the EC pathway from depression patient samples. FGA, FGB, and FGG were significantly up-regulated, and FVII was down-regulated. Thirteen DEGs related to depression and EC pathways were identified from the microarray database. Among them NF-κB Inhibitor Beta (NFKBIB) and Heat shock protein family B (small) member 1 (HSPB1) were highly correlated with EC pathway. We conclude that EC pathway is associated with depression, which provided clues for the biomarker development and the pathogenesis of depression.
Asunto(s)
Factores de Coagulación Sanguínea , Coagulación Sanguínea/genética , Depresión , Adulto , Biomarcadores/sangre , Factores de Coagulación Sanguínea/análisis , Factores de Coagulación Sanguínea/genética , Factores de Coagulación Sanguínea/metabolismo , Biología Computacional , Depresión/sangre , Depresión/diagnóstico , Depresión/genética , Depresión/metabolismo , Femenino , Humanos , Masculino , Persona de Mediana Edad , TranscriptomaRESUMEN
INTRODUCTION: Venous thromboembolism (VTE), including deep vein thrombosis (DVT) and/or pulmonary embolism (PE), is a common, acute, multifactorial disease with a five-years cumulative incidence of recurrence of approximately 25%. Actually, no single genetic defect can predict the risk of recurrence of VTE. Therefore, individual genetic risk profiling could be useful for the prediction of VTE recurrence. AIM OF THE STUDY: To assess the combined effect of the common prothrombotic genotypes on the risk of recurrence of VTE in recently diagnosed unprovoked VTE patients. PATIENTS AND METHODS: This population based, prospective follow-up study was carried out from January 2015 to December 2020 in (internal medicine, cardiovascular medicine and anesthesia and ICU departments, Tanta University Hospital, Egypt) on 224 recently diagnosed unprovoked VTE patients. Whole blood was collected by standard venipuncture at the time of admission prior to the beginning of anticoagulant therapy. Genomic DNA was extracted and was genotyped for the 5-SNPs Genetic risk score (GRS), previously validated for first venous thrombosis (FVL rs6025, PTM rs1799963, ABO rs8176719, FGG rs2066865 and FXI rs2036914). RESULTS: The main important finding in the present study was that patients having ≥3 risk alleles were associated with higher risk of VTE recurrence compared to those having ≤2 risk alleles (the reference group) (HR 2.5, 95% CI 1.48-4.21) (p = 0.001). Patients with GRS ≥ 3 had a significantly shorter time recurrence free survival (43.07 months) compared to the low risk group of patients with GRS (0-2) (p < 0.001). CONCLUSION: GRS model could be an effective and useful model in risk stratification of VTE patients, and genetic risk profiling of VTE patients could be used for the prediction of recurrence of VTE.