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Mutation Screening in Familial Cardiovascular Diseases
Čapek, Pavel ; Brdička, Radim (advisor) ; Gregor, Pavel (referee) ; Baxová, Alice (referee)
Introduction: Hypertrophic cardiomyopathy is a congenital cardiac disease with autosomal dominant pattern of inheritance and incomplete penetrance. With the knowledge of the responsible genes, the ability to detect the underlying genetic change and with the study of functional analysis of defected protein, we might be able to determine whether specific genotypes lead to different phenotypes. Aims of Study: To comprehensively analyze the mechanism of genesis of hypertrophic cardiomyopathy in Czech patients afflicted with this disorder from molecular genetic point of view (MYH7, TNNT2 gene) to functional analysis of the 3D molecular model of defected β-myosin heavy chain protein in silico. Beside these aims of the study, the reduction of production of inflammatory aggregates in the cardiovascular system was studied in patients with type 2 diabetes mellitus. The reason of this study was to look into possibilities of therapeutical effect on selected cardiovascular risks in patients with hypertrophic cardiomyopathy simultaneously suffering from type 2 diabetes mellitus. Both of these groups of patients have substantially increased risk of cardiovascular diseases due to development of premature atherosclerosis. Material and Methods: A total of 170 probands were enrolled in this study of MYH7 gene. DNA...
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The importance of Neolithic expansion in Central Europe - an assessment of phylogenetic age of mtDNA haplogroups in the Czech population.
Priehodová, Edita ; Černý, Viktor (advisor) ; Brdička, Radim (referee)
Agriculture, with different Neolithic cultures, starts in the Near East more than 10,000 years ago. This new way of life has very different archaeological manifestations that previous Mesolithic. After its Near Eastern emergence, the farming practices rapidly penetrated into southeastern Europe and the first signs of Neolithic in Central Europe are already 7,000 years old. It is being considered that the cultural innovations influenced demographic growth of the populations that have taken part in the Neolithic spread. In such situation, new mutations would have to fix and could form new specific haplogroups for Europe with ancestral ties to the Near East. Phylogeographic studies such as founder analysis of European and Near Eastern mtDNA sequences found that the European Neolithic component was enriched mainly by haplogroups J and T1, and that the genetic contribution of farming economy in European gene pool is about 10 - 20%. However, studies like these have not been yet realized in particular parts of Europe. The aim of this thesis is to disentangle the internal variability of Central European haplogroups J and T1 thought to be involved in the Neolithic demic diffusion. We classified these haplogroups from the HVS-I mtDNA sequences of 281 samples of the recent population of the Czech Republic. We...
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P-selectin - a novel protein in hemostasis
Hrachovinová, Ingrid ; Brdička, Radim (advisor) ; Kvasnička, Jan (referee) ; Penka, Miroslav (referee)
Originally, P-selectin played an important role in the vascular response to inflammation. In the first half of the nineties there were published papers demonstrated that P-selectin also played a significant role in blood coagulation and thrombosis. In these experiments anti-P-selectin antibodies blocked fibrin formation and suppressed the developing of thrombus. Shortly thereafter, P-selectin was shown to upregulate tissue factor (TF) generation on monocytes. The active role of P-selectin in hemostasis was supported also with our findings that overexpression of soluble P-selectin (sP-sel) can induce a procoagulant state in plasma/blood. I have focused in my postgraduate study on pathophysiology of sP-sel , its role in formation of microparicles (MPs) bearing TF. In the detail study of sP-sel we found, that procoagulant state in plasma is due to procoagulant MPs, part of them contained TF. We confirmed that recently characterized "blood-borne" TF could be induced by sP-sel. Moreover, because the production of microparticles was suppressed by inhibiting antibody against PSGL-1 (receptor of P-selectin on leukocytes), we proposed the origin of MPs from leukocytes. Recruitment of TF-bearing MPs from monocytes I demonstrated later with FACS screening of presence of specific CD-markers. I realized that...
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Genetic factors in aetiology and phatogenesisof low gamma-glutamyltransferase cholestasis and hereditary jaundice
Cebecauerová, Dita ; Jirsa, Milan (advisor) ; Brdička, Radim (referee) ; Macek, Milan (referee)
(English) Recent progress in understanding the molecular mechanism of hepatobiliary disorders enabled the improvement of diagnostic accuracy and promoted the study of the regulation of gene expression and its potential modifying factors. Current achievement in the field of genetically determined cholestatic disorders is well illustrated in this thesis, focused on low gamma-glutamyltransferase (γGT) cholestasis and hereditary jaundice. The study describes several distinct defects of hepatocyte transport system, characterises underlying mutations and their phenotypic consequences and, finally, extends these studies for detailed characterisation of ATP8B1 gene regulatory regions. Chapters related to low γGT cholestasis - characterise rare type of mutation associated with benign course of PFIC type I (formerly BRIC1) and explain the putative mechanisms of mutation origin. - provide extensive study of severe forms of ABCB11 deficiency (PFIC2) including genotype-phenotype correlations in 109 affected families, evaluation of the specific ABCB11 genotypes' impact on BSEP immunostaining and risk of hepatobiliary malignancy. - identify and characterise yet unknown regulatory regions of ATP8B1, a gene mutated in Progressive Familial Intrahepatic Cholestasis type I. The studies demonstrate the complex structure...
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Fetal hemoglobin in myelodysplastic syndrome patients.
Staňková, Nora ; Beličková, Monika (advisor) ; Brdička, Radim (referee)
5 Abstract Aims Determination of gene expression of HBG1 gamma globin in myelodysplastic syndrome (MDS) patients in CD34+ pluripotent hæmatopoietic cells and connection of HBG1 gene expression with various subtypes of MDS. Furthermore, detection of single nucleotide polymorphisms rs 4671393 and rs 11886868 in these patients and in healthy Czech population donors and to determine whether a connection exists between the occurrence of the above polymorphisms and HBG1 gene over-expression, as demonstrated in some hæmatological disorders. Samples The source of genetic material to identify gene expression were 80 HBG1 RNA samples isolated from the pluripotent hematopoietic CD34+ cells of MDS patients. As a sample of healthy controls, 6 samples of commercially purchased CD 34+ cells from the Lonza com- pany were used. The source of genetic material for the detection of polymorphisms were 140 DNA samples isolated from purified granulocytes of MDS patients and 49 samples of DNA isolated from peripheral blood granulocytes from healthy Czech population donors. Methods Real-Time PCR was used to determine HBG1 gene expression and detection of single nu- cleotide polymorphisms. Taqman Gene Expression Assay was used to determine the level of expression and the results were evaluated using the comparative ΔΔCT method....
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The mitochondrial genome in the ontogenesis
Töröková, Petra ; Brdička, Radim (advisor) ; Černý, Viktor (referee)
The main goal of this study is the comparison of sequences of the HVRII region of the mitochondrial genome in the cord blood sample and the saliva sample of the same individual, taken at average ten years from his/her birth. It is known that during ontogenesis the human genome changes. All the more the mitochondrial genome which shows a higher mutation rate, and moreover it is not taken care of it by repair mechanisms. In older individuals, there was found a distinctive amount of mitochondrial variations cumulated in different tissues in the process of the ontogenesis. This study is focused on the detection of these changes already in younger individuals. The tissue-specific variability which is created during ontogenesis might have an adverse influence on all sorts of the mtDNA based studies. The samples were taken in two regions (Teplice / Prachatice) that differ in the pollution of environment. With regard to that, the samples with discovered changes were compared from the standpoint of the region, which they had come from, with the aim to prove the influence of environment on the mutagenesis of the mitochondrial DNA. Samples were also compared from the point of view of sex. Furthermore the variability of the collection of Czech population was evaluated and the estimation of the genetic...
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Gene flow and its consequences for microevolution in Taraxacum (Asteraceae)
Marván, Richard ; Vančata, Václav (advisor) ; Brdička, Radim (referee) ; Lhota, Stanislav (referee)
Microsatellites(SIR) are very abundantand spread over the entire genome of all living organisms,so markers can be developedfor any genetic objective.Some SfRs exhibit very high levels of allelic variation.SIRs are generallyeasy to obtain and rapidlyassayed using polymerase chain reaction (PCR), requiringonly minute amountsof biologicalmaterial.Their distinctattributes made them attractiveand the popular tool of choice in a numberof applicationswithinthe area of biology' The core of this Íheslsconslsts of five chaptersrepresentingsťudieswhere we investigated autosomal or Y_linked SIRs Íocarry out our tasks. our objectives Were (i) to investigateallele frequencies for a set of autosomal SIRS, to estimatestandard population and forensic genetic parameters,and to perŤorman inter_populationcomparisonof allele frequencydata;(ii)to design a novel PCR-based Y chromosomeSIR multiplexassay for the analysisof highlydiverseloci, and to evaluateťhissysťemfor forensicand populationgenetic purposes;and (iii)to carryout the firststudy in a Pan paniscusgroup wherea combinationof behaviouraldata on male dominancerank and matingsuccess withgeneticdataonpaternityallowsthetestingof thepatternof relationshipsbetween thesevariables,and to evaluatepossib/econsequencesof the establishedgenetic relationshipsfor...
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Molecular Genetic Applications in Archeological and Anthropological Context
Urbanová, Markéta ; Černý, Viktor (advisor) ; Brdička, Radim (referee) ; Brůžek, Jaroslav (referee)
6 Coxcr-usroN The aim of our studywas to establisha laboratoryprotocolfor isolationand analysisof aDNA. By studying three different skeletal collections answersabout sex, kinship and presenceof pathogenicbacteriawere searched. During the course of this thesisnew protocols for aDNA study were developedand continuously optimized'We wereableto evaluateinformationaboutsexof 37 subadultindividuals(.l8'7%)from Mikulčice settlementcemetery(9thcentury)andto confirmpÍesenceof Mycobacteriumlepraeon 2 skeletonsfromŽatec cemetery(l3th century).Subadult skeletonsexaminationbrought interestingresults of extremelyhigher presenceof maleindividuals. Unfortunately,notall aimswerefulťrlled.By establishedprotocolfor aDNA kinshipanalysisit wasnot possibleto retrievesufficientinťormationfor explanationof familial relationshipof individualsfrom thesame doubleburialsite. All acquiredexperienceand establishedprotocols are very valuable and in fururewe supposetheir full utilizationandimorovementin thefield of kinshioanalvsisof aDNA molecule. 28
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Body composition changes in obese children during the weight reducing programme. The study of selected candidate genes for obesity and their associations with anthropometric and biochemical parameters.
Zamrazilová, Hana ; Bláha, Pavel (advisor) ; Štich, Vladimír (referee) ; Brdička, Radim (referee)
L. Abstrakt Background: Theprevalenceof obesityhasoverthe pasttwentyyearsgrownto suchanextentthat it is now commonlyrefenedto as a pandemic.From the point of view of healthrisks and complicationsconnectedwith obesity,the fact that this increasingtrend is prominenteven in childrenseemsto beveryserious.Thestudieshaveprovedclearrelationsbetweenearlyobesityand obesitypersistinguntil adulthood,the amountof visceralfat, cardiovascularrisks,diabetestype2, hypertensionandhyperlipidemia. Aims: To detect weight changesand body compositionchangesin children during the comprehensive5-6 weekin-patientweightreducingprogramme.To comparethe amountof body fat determinedby anthropomehicmethodsandDEXA method.To evaluateselectedbiochemical and hormonal parametersof lipid and glucosemetabolismsin obesechildren. To detectthe dependenceof anthropometricand biochemicalparameterson polymorphismsprolgl9pro, Ser343SerandGln2?3Argof the LEPRgene,andpolymorphismLeul62Valof theppARalfagene. Results: The study showed significant difftrences in body weight decreasesexpressedin percentagefromtheinitialbodyweightin thegroupof obesegirls(p & 0.001).Girlsthelowestage category(7-9 years)hadsignificantlygreaterweightlossescomparedto theirweightat thestartof theirstay,opposedto girlsin higheragecategories(11-17years,p & 0.05).At thebeginningof...
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Detection of minimal residual disease in patients with early breast cancer
Janků, Filip ; Matouš, Bohuslav (advisor) ; Brdička, Radim (referee) ; Melichar, Bohuslav (referee)
Breast cancer is one of the most serious health problems in our society. In the Czech Republic there are nearly 6000 women newly diagnosed annually. Despite the increasing incidence the mortality is leveling off or even decreasing in many countries (152,153). It is probably attributed to earlier diagnosis and the introduction of screening mammograms in many developed countries (154), and new findings in molecular biology of tumors. Several molecular factors are already routinely used in routine clinical practice as prognostic (estrogen and progesterone receptors, HER-2/neu, p53, Ki-67, vascular endothelial growth factor-VEGF), and predictive factors (estrogen and progesterone receptors, HER-2/neu) or therapeutic targets of anticancer treatment (estrogen and progesterone receptors, EGFR, HER- 2/neu, HER3, HER4, VEGF, mTOR) (7,45,155-157). The detection of minimal residual disease in early breast cancer is another attempt to implement modern diagnostic technologies in order to improve treatment outcomes. The aim of the study was to investigate diagnosis and prognostic implications of minimal residual disease in axillary lymph nodes, and bone marrow of patients with early breast cancer. The most promising material was bone marrow. From the clinical point of view it is necessary to validate both...
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