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

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

• Publikační typ
• abstrakt z konference MeSH

• Publikační typ
• abstrakt z konference MeSH

Fibrinogen—a 340-kDa glycoprotein—plays a crucial role in blood coagulation, platelet aggregation, wound healing, and other physiological processes. A mutation in fibrinogen may lead to congenital dysfibrinogenemia,a rare disease characterized by the functional deficiency of fibrinogen. About 580 cases of abnormal fibrinogens have been reported worldwide; thereof 335 cases in the fibrinogen Aa chain[1]. To our knowledge, only five cases of abnormal fibrinogens with two mutations [2–6] and one case of two different mutations in the same family [7] have been described earlier. A 52-year-old female was examined for bleeding. Routine hemostasis screening resulted in a diagnosis of dysfibrinogenemia. Functional testing revealed prolonged fibrin polymerization, prolonged lysis of the clot, abnormal fibrin morphology,and fibrinopeptides release. Genetic analysis showed two heterozygous nonsense mutations—previously described mutation AaGly13Glu and a novel mutation Aa Ser314Cys. The mutation Aa Gly13-Glu was found in her brother and niece, but there was no evidence in either of the mutation Aa Ser314Cys. While mutation Aa Gly13Glu is responsible for abnormal fibrinopeptide release and prolonged thrombin time, the novel mutation Aa Ser314Cys seems to affect fibrin morphology and fibrinolysis.

• MeSH
• afibrinogenemie krev   komplikace   genetika   MeSH
• bodová mutace MeSH
• dítě MeSH
• dospělí MeSH
• elektroforéza sérových bílkovin MeSH
• fibrin ultrastruktura   MeSH
• fibrinogeny abnormální genetika   izolace a purifikace   MeSH
• fibrinopeptid A metabolismus   MeSH
• hemoragické poruchy etiologie   MeSH
• heterozygot MeSH
• lidé středního věku MeSH
• lidé MeSH
• mikroskopie elektronová rastrovací MeSH
• nesmyslný kodon MeSH
• posttranslační úpravy proteinů MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH

May-Hegglin anomaly (MHA), Sebastian (SBS), Fechtner (FTNS) and Epstein (EPS) syndromes are rare autosomal dominant disorders with giant platelets and thrombocytopenia. Other manifestations of these disorders are combinations of the presence of granulocyte inclusions and deafness, cataracts and renal failure. Currently, MHA, SBS, FTNS and EPS are considered to be distinct clinical manifestation of a single illness caused by mutations of the MYH9 gene encoding the heavy chain of non-muscle myosin IIA (NMMHC-IIA). As the MYH9 gene has a high number of exons, it takes much time and material to use this method for the detection of MYH9 mutations. Recently, a new method has been introduced for scanning DNA mutations without the need for direct sequencing: high-resolution melting analysis (HRMA). Mutation detection with HRMA relies on the intercalation of the specific dye (LC Green plus) in double-strand DNA and fluorescence monitoring of PCR product melting profiles. In our study, we optimized the conditions and used HRMA for rapid screening of mutations in all MYH9 exons in seven affected individuals from four unrelated families with suspected MYH9 disorders. Samples identified by HRMA as positive for the mutation were analysed by direct sequencing. HRMA saved us over 85% of redundant sequencing.

• MeSH
• lidé MeSH
• missense mutace MeSH
• molekulární motory genetika   chemie   MeSH
• mutační analýza DNA metody   MeSH
• těžké řetězce myosinu genetika   chemie   MeSH
• tranzitní teplota MeSH
• trombocytopenie genetika   krev   MeSH
• trombocyty patologie   MeSH
• velikost buňky MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• práce podpořená grantem MeSH

The role of oxidative stress in the initiation and progression of myelodysplastic syndromes (MDS) as a consequence of iron overload remains unclear. In this study we have simultaneously quantified plasma low-molecular-weight aminothiols, malondialdehyde, nitrite, and nitrate and have studied their correlation with serum iron/ferritin levels, patient treatment (chelation therapy), and clinical outcomes. We found significantly elevated plasma levels of total, oxidized, and reduced forms of cysteine (P < 0.001), homocysteine (P < 0.001), and cysteinylglycine (P < 0.006) and significantly depressed levels of total and oxidized forms of glutathione (P < 0.03) and nitrite (P < 0.001) in MDS patients compared to healthy donors. Moreover, total (P < 0.032) and oxidized cysteinylglycine (P = 0.029) and nitrite (P = 0.021) differed significantly between the analyzed MDS subgroups with different clinical classifications. Malondialdehyde levels in plasma correlated moderately with both serum ferritin levels (r = 0.78, P = 0.001) and serum free iron levels (r = 0.60, P = 0.001) and were significantly higher in patients with iron overload. The other analyzed compounds lacked correlation with iron overload (represented by serum iron/ferritin levels). For the first time our results have revealed significant differences in the concentrations of plasma aminothiols in MDS patients, when compared to healthy donors. We found no correlation of these parameters with iron overload and suggest the role of oxidative stress in the development of MDS disease.

• MeSH
• dipeptidy krev   MeSH
• dusičnany krev   MeSH
• dusitany krev   MeSH
• ferritiny krev   MeSH
• lidé MeSH
• malondialdehyd krev   MeSH
• myelodysplastické syndromy krev   komplikace   patologie   MeSH
• oxidace-redukce MeSH
• oxidační stres MeSH
• přetížení železem krev   komplikace   patologie   MeSH
• studie případů a kontrol MeSH
• sulfhydrylové sloučeniny krev   MeSH
• výsledek terapie MeSH
• železo krev   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Fibrinogen is one of the plasma proteins most susceptible to oxidative modification. It has been suggested that modification of fibrinogen may cause thrombotic/bleeding complications associated with many pathophysiological states of organism. We exposed fibrinogen molecules to three different modification reagents-malondialdehyde, sodium hypochlorite, and peroxynitrite-that are presented to various degrees in different stages of oxidative stress. We studied the changes in fibrin network formation and platelet interactions with modified fibrinogens under flow conditions. The fastest modification of fibrinogen was caused by hypochlorite. Fibers from fibrinogen modified with either reagent were thinner in comparison with control fibers. We found that platelet dynamic adhesion was significantly lower on fibrinogen modified with malondialdehyde and significantly higher on fibrinogen modified either with hypochlorite or peroxynitrite reflecting different prothrombotic/antithrombotic properties of oxidatively modified fibrinogens. It seems that, in the complex reactions ongoing in living organisms at conditions of oxidation stress, hypochlorite modifies proteins (e.g., fibrinogen) faster and more preferentially than malondialdehyde. It suggests that the prothrombotic effects of prior fibrinogen modifications may outweigh the antithrombotic effect of malondialdehyde-modified fibrinogen in real living systems.

• MeSH
• adhezivita trombocytů účinky léků   fyziologie   MeSH
• chlornan sodný farmakologie   MeSH
• fibrinogen chemie   metabolismus   farmakologie   MeSH
• indikátory a reagencie chemie   farmakologie   MeSH
• kultivované buňky MeSH
• kyselina peroxydusitá chemie   MeSH
• lidé MeSH
• malondialdehyd chemie   farmakologie   MeSH
• oxidační stres účinky léků   fyziologie   MeSH
• trombocyty účinky léků   metabolismus   MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The aim of this study was to investigate the structure and function of fibrinogen obtained from a patient with normal coagulation times and idiopathic thrombophilia. This was done by SDS-PAGE and DNA sequence analyses, scanning electron microscopy, fibrinopeptide release, fibrin polymerisation initiated by thrombin and reptilase, fibrinolysis, and platelet aggregometry. A novel heterozygous point mutation in the fibrinogen Aα chain, Phe98 to Ile, was found and designated as fibrinogen Vizovice. The mutation, which is located in the RGDF sequence (Aα 95-98) of the fibrinogen coiled-coil region, significantly affected fibrin clot morphology. Namely, the clot formed by fibrinogen Vizovice contained thinner and curled fibrin fibers with reduced length. Lysis of the clots prepared from Vizovice plasma and isolated fibrinogen were found to be impaired. The lysis rate of Vizovice clots was almost four times slower than the lysis rate of control clots. In the presence of platelets agonists the mutant fibrinogen caused increased platelet aggregation. The data obtained show that natural mutation of Phe98 to Ile in the fibrinogen Aα chain influences lateral aggregation of fibrin protofibrils, fibrinolysis, and platelet aggregation. They also suggest that delayed fibrinolysis, together with the abnormal fibrin network morphology and increased platelet aggregation, may be the direct cause of thrombotic complications in the patient associated with pregnancy loss.

• MeSH
• agregace trombocytů MeSH
• batroxobin metabolismus   MeSH
• bodová mutace MeSH
• časové faktory MeSH
• dospělí MeSH
• fibrin metabolismus   MeSH
• fibrinogen genetika   metabolismus   MeSH
• fibrinolýza MeSH
• hemokoagulace MeSH
• heterozygot MeSH
• konformace proteinů MeSH
• lidé MeSH
• samovolný potrat krev   MeSH
• těhotenství MeSH
• trombin metabolismus   MeSH
• trombofilie krev   genetika   MeSH
• vyšetření krevní srážlivosti MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH
• práce podpořená grantem MeSH

• MeSH
• afibrinogenemie krev   genetika   MeSH
• fibrinogen genetika   metabolismus   MeSH
• fibrinogeny abnormální genetika   metabolismus   MeSH
• homozygot MeSH
• lidé MeSH
• předškolní dítě MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• Publikační typ
• dopisy MeSH
• kazuistiky MeSH
• práce podpořená grantem MeSH