Aberrant glycosylation of glycoproteins has been linked with various pathologies. Therefore, understanding the relationship between aberrant glycosylation patterns and the onset and progression of the disease is an important research goal that may provide insights into cancer diagnosis and new therapy development. In this study, we use a surface plasmon resonance imaging biosensor and a lectin array to investigate aberrant glycosylation patterns associated with oncohematological disease-myelodysplastic syndromes (MDS). In particular, we detected the interaction between the lectins and glycoproteins present in the blood plasma of patients (three MDS subgroups with different risks of progression to acute myeloid leukemia (AML) and AML patients) and healthy controls. The interaction with lectins from Aleuria aurantia (AAL) and Erythrina cristagalli was more pronounced for plasma samples of the MDS and AML patients, and there was a significant difference between the sensor response to the interaction of AAL with blood plasma from low and medium-risk MDS patients and healthy controls. Our data also suggest that progression from MDS to AML is accompanied by sialylation of glycoproteins and increased levels of truncated O-glycans and that the number of lectins that allow discriminating different stages of disease increases as the disease progresses.

• MeSH
• akutní myeloidní leukemie * MeSH
• biosenzitivní techniky * MeSH
• glykoproteiny metabolismus   MeSH
• glykosylace MeSH
• krevní plazma metabolismus   MeSH
• lektiny MeSH
• lidé MeSH
• myelodysplastické syndromy * terapie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

A single-center study was conducted on 120 patients with inherited disorders of primary hemostasis followed at our hematological center. These patients presented a variety of bleeding symptoms; however, they had no definitive diagnosis. Establishing a diagnosis has consequences for the investigation of probands in families and for treatment management; therefore, we aimed to improve the diagnosis rate in these patients by implementing advanced diagnostic methods. According to the accepted international guidelines at the time of study, we investigated platelet morphology, platelet function assay, light-transmission aggregometry, and flow cytometry. Using only these methods, we were unable to make a definitive diagnosis for most of our patients. However, next-generation sequencing (NGS), which was applied in 31 patients, allowed us to establish definitive diagnoses in six cases (variants in ANKRD26, ITGA2B, and F8) and helped us to identify suspected variants (NBEAL2, F2, BLOC1S6, AP3D1, GP1BB, ANO6, CD36, and ITGB3) and new suspected variants (GFI1B, FGA, GP1BA, and ITGA2B) in 11 patients. The role of NGS in patients with suspicious bleeding symptoms is growing and it changes the diagnostic algorithm. The greatest disadvantage of NGS, aside from the cost, is the occurrence of gene variants of uncertain significance.

• MeSH
• krevní proteiny genetika   MeSH
• krvácení MeSH
• lidé MeSH
• trombocytopatie * diagnóza   genetika   MeSH
• vyšetření funkce trombocytů MeSH
• vysoce účinné nukleotidové sekvenování MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Česká republika MeSH

Here, we present the first case of fibrinogen variant FGG c.8G>A. We investigated the behaviour of this mutated fibrinogen in blood coagulation using fibrin polymerization, fibrinolysis, fibrinopeptides release measurement, mass spectrometry (MS), and scanning electron microscopy (SEM). The case was identified by routine coagulation testing of a 34-year-old man diagnosed with thrombosis. Initial genetic analysis revealed a heterozygous mutation in exon 1 of the FGG gene encoding gamma chain signal peptide. Fibrin polymerization by thrombin and reptilase showed the normal formation of the fibrin clot. However, maximal absorbance within polymerization was lower and fibrinolysis had a longer degradation phase than healthy control. SEM revealed a significant difference in clot structure of the patient, and interestingly, MS detected several posttranslational oxidations of fibrinogen. The data suggest that the mutation FGG c.8G>A with the combination of the effect of posttranslational modifications causes a novel case of hypofibrinogenemia associated with thrombosis.

• MeSH
• afibrinogenemie * komplikace   genetika   MeSH
• dospělí MeSH
• fibrin metabolismus   MeSH
• fibrinogen genetika   metabolismus   MeSH
• fibrinogeny abnormální * genetika   metabolismus   MeSH
• hemostatika * MeSH
• lidé MeSH
• oxidační stres MeSH
• posttranslační úpravy proteinů MeSH
• trombóza * komplikace   genetika   MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• mužské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH

Congenital fibrinogen disorders are caused by mutations in genes coding for fibrinogen and may lead to various clinical phenotypes. Here, we present a functional and structural analysis of 4 novel variants located in the FGB gene coding for fibrinogen Bβ chain-heterozygous missense BβY416C and BβA68S, homozygous nonsense BβY345*, and heterozygous nonsense BβW403* mutations. The cases were identified by coagulation screening tests and further investigated by various methods. Fibrin polymerization had abnormal development with decreased maximal absorbance in all patients. Plasmin-induced fibrin degradation revealed different lytic phases of BβY416C and BβW403* than those of the control. Fibrinopeptide cleavage measured by reverse phase high pressure liquid chromatography of BβA68S showed impaired release of fibrinopeptide B. Morphological properties, studied through scanning electron microscopy, differed significantly in the fiber thickness of BβY416C, BβA68S, and BβW403*, and in the fiber density of BβY416C and BβW403*. Finally, homology modeling of BβA68S showed that mutation caused negligible alternations in the protein structure. In conclusion, all mutations altered the correct fibrinogen function or structure that led to congenital fibrinogen disorders.

• MeSH
• afibrinogenemie krev   diagnóza   genetika   MeSH
• fenotyp * MeSH
• fibrinogen chemie   genetika   metabolismus   MeSH
• genetická predispozice k nemoci * MeSH
• genetické asociační studie MeSH
• hemokoagulace MeSH
• konformace proteinů MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• molekulární modely MeSH
• mutace * MeSH
• mutační analýza DNA MeSH
• novorozenec MeSH
• senioři MeSH
• vyšetření krevní srážlivosti MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• novorozenec MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH

Congenital hypofibrinogenemia is a rare bleeding disorder characterized by a proportional decrease of functional and antigenic fibrinogen levels. Hypofibrinogenemia can be considered the phenotypic expression of heterozygous loss of function mutations occurring within one of the three fibrinogen genes (FGA, FGB, and FGG). Clinical manifestations are highly variable; most patients are usually asymptomatic, but may appear with mild to severe bleeding or thrombotic complications. We have sequenced all exons of the FGA, FGB, and FGG genes using the DNA isolated from the peripheral blood in two unrelated probands with mild hypofibrinogenemia. Coagulation screening, global hemostasis, and functional analysis tests were performed. Molecular modeling was used to predict the defect of synthesis and structural changes of the identified mutation. DNA sequencing revealed a novel heterozygous variant c.1421G>A in exon 8 of the FGB gene encoding a Bβ chain (p.Trp474Ter) in both patients. Clinical data from patients showed bleeding episodes. Protein modelling confirmed changes in the secondary structure of the molecule, with the loss of three β sheet arrangements. As expected by the low fibrinogen levels, turbidity analyses showed a reduced fibrin polymerisation and imaging difference in thickness fibrin fibers. We have to emphasize that our patients have a quantitative fibrinogen disorder; therefore, the reduced function is due to the reduced concentration of fibrinogen, since the Bβ chains carrying the mutation predicted to be retained inside the cell. The study of fibrinogen molecules using protein modelling may help us to understand causality and effect of novel genetic mutations.

• Publikační typ
• časopisecké články MeSH

Fibrinogen je klíčový glykoprotein krevní koagulace, během níž je přeměňován na fibrin. Vrozená dysfibrinogenemie je choroba charakterizovaná funkční poruchou molekuly vedoucí k abnormální tvorbě fibrinu. Vrozená hypofibrinogenemie je vzácné onemocnění charakterizované sníženou hladinou funkčního i celkového fibrinogenu v plazmě. Vyšetřili jsme 36 nepříbuzných rodin z celé České republiky s diagnózou suspektní dysfibrinogenemie či afibrinogenemie. Čtyři pacienti měli trombotické komplikace, osm pacientů se manifestovalo krvácivě a ostatní byli asymptomatičtí. Genetické vyšetření odhalilo vrozenou mutaci u třinácti nepříbuzných rodin, u nichž způsobuje vrozenou dysfibrinogenemii. Pomocí genetických, proteomických a funkčních vyšetření bylo odhaleno osm případů dysfibrinogenemie v Aα řetězci, konkrétně se jedná o mutace Aα Arg16Cys; Aα Arg16His; Aα Gly13Glu; Aα Phe98Ile; Aα Asn106Asp; a kombinovanou mutaci Aα Gly13Glu a Aα Ser314Cys. Jeden případ byl nalezen v Bβ řetězci – Bβ Arg237Ser a čtyři případy byly odhaleny v γ řetězci – γ Ser313Gly, γ Tyr262Cys, γ Tyr363Asn a γ Arg275His. V jednom případě byla identifikována molekulární příčina vrozené afibrinogenemie. V jednom případě byla zjištěna příčina získané dysfibrinogenemie v souvislosti s produkcí autoprotilátek proti fibrinogenu u pacienta s mnohočetným myelomem.

Fibrinogen is a key glycoprotein of blood coagulation. During haemocoagulation fibrinogen is converted to fibrin. Congenital dysfibrinogenemia is a disease wherein an inherited abnormality in the fibrinogen molecule results in defective fibrin clot formation. Hereditary hypofibrinogenemia is a disease wherein an inherited abnormality in the fibrinogen molecule results in low fibrinogen level in plasma. 36 unrelated families in the Czech Republic suspected with either dysfibrinogenemia or afibrinogenemia were examined. Four patients presented with thrombosis, eight patients presented with bleeding tendencies and others were asymptomatic. Heterozygous point mutations Aα Arg16Cys (Fibrinogen Nový Jičín, Ostrava I), Aα Arg16His (Fibrinogen Praha II, Ostrava II), Aα Asn106Asp (Fibrinogen Plzeň), γ Tyr363Asn (Fibrinogen Praha III), γ Tyr262Cys (Fibrinogen Liberec) and γ Arg275His (Fibrinogen Brno) were found to be the direct causes of dysfibrinogenemias in the carriers. Molecular genetics experiments revealed inherited mutations in 13 unrelated families causing hereditary dysfibrinogenemia. In one case acquired dysfibrinogenemia secondary to multiple myeloma was found.

• Klíčová slova
• dysfibrinogenemie,
• MeSH
• afibrinogenemie * diagnóza   epidemiologie   genetika   krev   MeSH
• fibrinogen analýza   chemie   MeSH
• fibrinogeny abnormální analýza   genetika   MeSH
• fibrinolýza MeSH
• genetické testování MeSH
• lidé MeSH
• rodina MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• práce podpořená grantem MeSH