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

We describe the internal structure, spatial organization and dynamic formation of coronary artery thrombi from ST-segment elevation myocardial infarction patients. Scanning electron microscopy (SEM) revealed significant differences among four groups of patients (<2 hours; 2-6 hours; 6-12 hours, and >12 hours) related to the time of ischemia. Coronary artery thrombi from patients presenting less than 2 hours after the infarction were almost entirely composed of platelets, with small amounts of fibrin and red blood cells. In contrast, thrombi from late presenters (>12 hours) consisted of mainly platelets at the distal end, where clotting was initiated, with almost no platelets at the proximal end, while the red blood cell content went from low at the initiating end to more than 90% at the proximal end. Furthermore, fibrin was present mainly on the outside of the thrombi and older thrombi contained thicker fibers. The red blood cells in late thrombi were compressed to a close-packed, tessellated array of polyhedral structures, called polyhedrocytes. Moreover, there was redistribution from the originally homogeneous composition to fibrin and platelets to the outside, with polyhedrocytes on the interior. The presence of polyhedrocytes and the redistribution of components are signs of in vivo clot contraction (or retraction). These results suggest why later thrombi are resistant to fibrinolytic agents and other treatment modalities, since the close-packed polyhedrocytes form a nearly impermeable seal. Furthermore, it is of particular clinical significance that these findings suggest specific disparate therapies that will be most effective at different stages of thrombus development.

• MeSH
• čas zasáhnout při rozvinutí nemoci MeSH
• časové faktory MeSH
• erytrocyty patologie   MeSH
• fibrin analýza   MeSH
• fibrinolytika * aplikace a dávkování   škodlivé účinky   MeSH
• hemokoagulace účinky léků   fyziologie   MeSH
• infarkt myokardu s elevacemi ST úseků * etiologie   terapie   MeSH
• koronární trombóza * diagnostické zobrazování   farmakoterapie   metabolismus   patologie   MeSH
• léková rezistence fyziologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mikroskopie elektronová rastrovací metody   MeSH
• trombektomie metody   MeSH
• trombocyty patologie   MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články 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

Myelodysplastic syndromes (MDS) are a heterogeneous group of hematological malignancies with a high risk of transformation to acute myeloid leukemia (AML). MDS are associated with posttranslational modifications of proteins and variations in the protein expression levels. In this work, we present a novel interactomic diagnostic method based on both protein array and surface plasmon resonance biosensor technology, which enables monitoring of protein-protein interactions in a label-free manner. In contrast to conventional methods based on the detection of individual biomarkers, our presented method relies on measuring interactions between arrays of selected proteins and patient plasma. We apply this method to plasma samples obtained from MDS and AML patients, as well as healthy donors, and demonstrate that even a small protein array comprising six selected proteins allows the method to discriminate among different MDS subtypes and healthy donors.

• MeSH
• analýza hlavních komponent MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mapování interakce mezi proteiny * MeSH
• mladý dospělý MeSH
• myelodysplastické syndromy krev   diagnóza   MeSH
• povrchová plasmonová rezonance MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• vazba proteinů MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Myelodysplastic syndromes (MDS) represent a heterogeneous group of pre-leukemic disorders, characterized by ineffective hematopoiesis and the abnormal blood cell development of one or more lineages. Oxidative stress, as an important factor in the carcinogenesis of onco-hematological diseases, is also one of the known factors involved in the pathogenesis of MDS. An increase of reactive oxygen species (ROS) may lead to the oxidation of DNA, lipids, and proteins, thereby causing cell damage. Protein carbonylation caused by ROS is defined as an irreversible post-translational oxidative modification of amino acid side chains, and could play an important role in signaling processes. The detection of protein carbonyl groups is a specific useful marker of oxidative stress. In this study, we examined 32 patients divided into three different subtypes of MDS according to the World Health Organization (WHO) classification criteria as refractory anemia with ringed sideroblasts (RARS), refractory cytopenia with multilineage dysplasia (RCMD), refractory anemia with excess blasts-1,2 (RAEB-1,2). We found significant differences in protein carbonylation between the group of all MDS patients and healthy controls (P=0.0078). Furthermore, carbonylated protein levels were significantly elevated in RARS patients compared to healthy donors (P=0.0013) and to RCMD patients (P=0.0277). We also found a significant difference in the total iron binding capacity (TIBC) between individual subgroups of MDS patients (P=0.0263). Moreover, TIBC was decreased in RARS patients compared to RCMD patients (P=0.0203). TIBC moderately negatively correlated with carbonyl levels (r=-0.5978, P=0.0054) in the MDS patients as a whole. Additionally we observed changes in the carbonylated proteins of RARS patients in comparison with healthy controls and their negative controls. Using tandem mass spectrometry (LC-MS/MS) we identified 27 uniquely carbonylated proteins of RARS patients, which were generated by ROS and could influence the pathophysiology of low-risk MDS. These data indicate that increased protein carbonylation is related with RARS as low-risk MDS subgroup. We suggest that this type of post-translational modification in MDS disease is not "only" a consequence of oxidative stress, but also plays an active role in the pathophysiology and iron metabolism within the RARS subgroup of MDS. Measurement of plasma carbonyl levels and the isolation of carbonylated plasma proteins, followed by their identification, could serve as a potential diagnostic and prognostic tool in MDS.

• MeSH
• dospělí MeSH
• karbonylace proteinů MeSH
• krevní proteiny metabolismus   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• myelodysplastické syndromy diagnóza   metabolismus   MeSH
• oxidační stres MeSH
• prognóza MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• refrakterní anemie MeSH
• senioři MeSH
• tandemová hmotnostní spektrometrie MeSH
• vazba proteinů MeSH
• železo metabolismus   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• 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

Vrozená afibrinogenémie je velmi vzácná vrozená krvácivá choroba způsobená absencí fibrinogenu v krevní plazmě. Pacienti mohou být postiženi závažným spontánním krvácením, včetně intrakraniálního, a to v kterémkoliv věku. Uvádíme kazuistiku chlapce s vrozenou afibrinogenémií, která je způsobena nově zjištěnou mutací v genu pro fibrinogen – homozygotní delecí Aα 6477A. Diagnóza byla stanovena po krvácení při plastice rozštěpu rtu, čelisti a patra po narození. Ve věku 11 měsíců došlo ke spontánnímu intrakraniálnímu krvácení, rozsáhlý hematom byl lokalizován okcipitálně. Akutní neurochirurgický výkon s evakuací hematomu proběhl za substituce plazmatickým derivátem fibrinogenu, stejně jako pooperační období. Poté jsme pacientovi zavedli pravidelnou profylaktickou substituci koncentrátem fibrinogenu 2krát týdně, dosud nebyla zaznamenána žádná další krvácivá epizoda. Spontánní intrakraniální krvácení u pacientů s vrozenou krvácivou chorobou je často závažné. Rozhodujícími faktory pro přežití pacienta jsou včasná diagnostika, substituce koncentrátem a případná neurochirurgická intervence. Profylaktická substituční léčba je po takovéto příhodě doporučeným léčebným přístupem.

Congenital afibrinogenemia is very rare inherited bleeding disorder caused by absence of fibrinogen in plasma. Serious bleeding including intracranial haemorrhage may occur at any age. We present a case report of a boy with afibrinogenemia caused by novel mutation – homozygous deletion Aα 6477A. Diagnosis was set after cleft lip and palate plastic surgery after birth. Spontaneous intracranial bleeding in occipital region occurred spontaneously when the boy was 11 month old. Replacement therapy with plasma derived fibrinogen concentrate successfully covered neurosurgical evacuation of hematoma. Then we commenced the patient on regular prophylaxis with fibrinogen concentrate two times a week. No further bleeding episodes occurred until present time. Spontaneous intracranial bleeding in patients with inherited bleeding disorders is often life threatening. Immediate replacement therapy is crucial, together with exact diagnostics and surgery, when necessary. Prophylactic substitution therapy after a severe bleeding episode is usually recommended.

• MeSH
• afibrinogenemie * diagnóza   genetika   patofyziologie   terapie   MeSH
• fibrinogen aplikace a dávkování   fyziologie   genetika   terapeutické užití   MeSH
• hematologie MeSH
• intrakraniální krvácení * diagnóza   etiologie   genetika   patofyziologie   prevence a kontrola   terapie   MeSH
• kojenec MeSH
• lidé MeSH
• neurochirurgické výkony MeSH
• počítačová rentgenová tomografie MeSH
• výsledek terapie MeSH
• Check Tag
• kojenec MeSH
• lidé MeSH
• mužské pohlaví MeSH
• Publikační typ
• kazuistiky MeSH