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Molecular-cytogenetic diagnostics of marker chromosomes
Tesner, Pavel ; Kočárek, Eduard (advisor) ; Zemanová, Zuzana (referee) ; Šubrt, Ivan (referee)
Supernumerary marker chromosomes (sSMCs) are a relatively rare cytogenetic phenomenon. Their laboratory examination is often difficult, and the clinical interpretation is even more challenging. The main reason is that most sSMC carriers have no clinical manifestations. The chromosome origin and exact range of the aberration are very important, as well as the fact that sSMCs are often found in mosaics that can strongly influence both the phenotype and the interpretation of result. Prenatal sSMC finding is one of the most challenging situations in both clinical and laboratory genetics. This work deals with the investigation process of sSMC carriers using molecular cytogenetic techniques, especially fluorescence in situ hybridization (FISH). We investigated a total of 67 families collected both prospectively and retrospectively, and we found 70 unique sSMCs in a total of 74 individuals. Six cases were familial and in three cases two sSMCs were found in one individual. According to the initial karyotype finding, the cases were divided into two groups, sSMCs supernumerary to a normal karyotype (group A) and sSMCT s supernumerary to the Turner karyotype (group B). The chromosomal origin was successfully determined in 88,6 % sSMCs. In group A the most common findings were sSMCs derived from chromosome 15,...
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The significance of thrombophilic and immunologic factors in human reproduction
Šubrt, Ivan ; Ulčová-Gallová, Zdeňka (advisor) ; Kužel, David (referee) ; Šantavý, Jiří (referee)
The aim of presented study was to compare frequencies of eight antiphospholipid antibodies (aPL) in serum and assorted genetic thrombophilic factors and their mutual relation in patients with recurrent pregnancy loss (RPL) and controls. Enzyme-linked immunosorbent assay was used for detection of aPL against phosphatidyl-L-serine, phosphatidylethanolamine, phosphatidylinositol, phosphatidyl- DL-glycerol, phosphatidic acid, annexin V, cardiolipin, and beta2-GPI. Thrombophilic mutations factor V Leiden (F5 G1691A), F II G20210A, and MTHFR C677T and A1298C variants were determined using a melting curve analysis of the PCR amplification product detected by the fluorescence resonance energy transfer (FRET). PAI1 (-675)4G/5G, PROZ intron F G79A, PROZ A(-13)G and PROZ R255H variants were determined using standard PCR-RFLP method. Genotypes distribution and allelic frequencies were calculated. Correlation between aPL and thrombophilic factors was tested by chi-square and Fisher exact test. Our results showed significantly increased prevalence of aPL against phosphatidylinositol (17 - 19.6 % dependent on number of spontaneous miscarriages) and against phosphatidyl-L-serine (18-25 %). aPL in IgG prevailed. In 96 % of studied group we found at least one risk factor (either aPL positivity or thrombophilic factor). Both...
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Clonal evolution of leukemic cells and its role in the progression of leukemia and preleukemia
Svobodová, Karla ; Zemanová, Zuzana (advisor) ; Urbánková, Helena (referee) ; Šubrt, Ivan (referee)
Clonal evolution is a multistep process characterized by progression of the disease, adverse prognosis and shortening of overall survival. The aim of the dissertation was a detailed characterization of identified changes in patients with myelodysplastic syndromes (MDS) and clonal evolution and evaluation of their prognostic impact. We performed detail cytogenomic analyses in 36/469 (8%) patients with confirmed linear clonal evolution. We described 57 primary abnormalities (32% MDS-specific) at the time of diagnosis, the most frequent was deletion of long arm of chromosome 5. We proved 156 secondary aberrations (21% MDS-specific) during the course of the clonal evolution, the most frequent were trisomies/tetrasomies of chromosome 8. We identified acquired uniparental disomies (aUPD) in 19% of patients. In MDS-specific aUPDs 4q, 11q and 17p, we proved homozygous mutations of TET2, c-CBL and TP53 genes. We found a statistically significant difference in overall survival between the groups of patients divided according to their diagnostic cytogenomic findings. In patients with clonal evolution before treatment 54% of aberrations were gains of whole chromosomes, by contrast 44% of abnormalities identified in patients with clonal evolution after treatment were monosomies or deletions. The study of clonal...
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Variants of human chromosome 9 - from norm to pathology Epidemiology and significance for medical genetics.
Šípek, Antonín ; Panczak, Aleš (advisor) ; Šubrt, Ivan (referee) ; Vallová, Vladimíra (referee)
Heterochromatin variants of human chromosome 9 belong to the most common variabilities of human karyotype. The variability involves the large block of constitutive heterochromatin in the pericentric region of chromosome 9, which is composed of various types of repetitive DNA sequences. Those variants can be studied from population epidemiologic, molecular cytogenetic and clinical genetic point of view. We have performed a broad epidemiologic study of the incidence of pericentric inversion of chromosome 9 (inv(9)) and other variants of chromosome 9 in 6 different laboratory cohorts, which included the evaluation of more than 26.000 of cytogenetic reports, the study we published is currently the largest in the world. We expressed the overall incidence of inv(9) to be 1.6% and the total incidence of variants of chromosome 9 to be 3.3-3.9%. Inv(9) was more common in females, however the difference was not statistically significant. Molecular cytogenetic part of the project was based on our own diagnostic approach, which involved the combination of three different commercial FISH probes. Combination of those probes allowed us to differentiate particular subvariants of chromosome 9, which cannot be analyzed only by using G- or C-banding. Using our method, we tested 49 carriers of chromosome 9...
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Molecular-cytogenetic diagnostics of marker chromosomes
Tesner, Pavel ; Kočárek, Eduard (advisor) ; Zemanová, Zuzana (referee) ; Šubrt, Ivan (referee)
Supernumerary marker chromosomes (sSMCs) are a relatively rare cytogenetic phenomenon. Their laboratory examination is often difficult, and the clinical interpretation is even more challenging. The main reason is that most sSMC carriers have no clinical manifestations. The chromosome origin and exact range of the aberration are very important, as well as the fact that sSMCs are often found in mosaics that can strongly influence both the phenotype and the interpretation of result. Prenatal sSMC finding is one of the most challenging situations in both clinical and laboratory genetics. This work deals with the investigation process of sSMC carriers using molecular cytogenetic techniques, especially fluorescence in situ hybridization (FISH). We investigated a total of 67 families collected both prospectively and retrospectively, and we found 70 unique sSMCs in a total of 74 individuals. Six cases were familial and in three cases two sSMCs were found in one individual. According to the initial karyotype finding, the cases were divided into two groups, sSMCs supernumerary to a normal karyotype (group A) and sSMCT s supernumerary to the Turner karyotype (group B). The chromosomal origin was successfully determined in 88,6 % sSMCs. In group A the most common findings were sSMCs derived from chromosome 15,...
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The significance of thrombophilic and immunologic factors in human reproduction
Šubrt, Ivan ; Ulčová-Gallová, Zdeňka (advisor) ; Kužel, David (referee) ; Šantavý, Jiří (referee)
The aim of presented study was to compare frequencies of eight antiphospholipid antibodies (aPL) in serum and assorted genetic thrombophilic factors and their mutual relation in patients with recurrent pregnancy loss (RPL) and controls. Enzyme-linked immunosorbent assay was used for detection of aPL against phosphatidyl-L-serine, phosphatidylethanolamine, phosphatidylinositol, phosphatidyl- DL-glycerol, phosphatidic acid, annexin V, cardiolipin, and beta2-GPI. Thrombophilic mutations factor V Leiden (F5 G1691A), F II G20210A, and MTHFR C677T and A1298C variants were determined using a melting curve analysis of the PCR amplification product detected by the fluorescence resonance energy transfer (FRET). PAI1 (-675)4G/5G, PROZ intron F G79A, PROZ A(-13)G and PROZ R255H variants were determined using standard PCR-RFLP method. Genotypes distribution and allelic frequencies were calculated. Correlation between aPL and thrombophilic factors was tested by chi-square and Fisher exact test. Our results showed significantly increased prevalence of aPL against phosphatidylinositol (17 - 19.6 % dependent on number of spontaneous miscarriages) and against phosphatidyl-L-serine (18-25 %). aPL in IgG prevailed. In 96 % of studied group we found at least one risk factor (either aPL positivity or thrombophilic factor). Both...
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