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Study of placental specific microRNA expression in pacients with spontaneous preterm birth and preterm prelabor rupture of membranes
Vintrová, Iva ; Hromadníková, Ilona (advisor) ; Daňková, Pavlína (referee)
MicroRNAs (miRNAs) are small non-coding RNAs with a length of 18 to 25 nucleotides playing a pivotal role in post-transcriptional regulation of gene expression. There are miRNAs whose expression is limited to a certain tissue type and diseases which are characterized by a unique miRNA expression profile. I assumed spontaneous preterm birth (PTB) and preterm prelabor rupture of membranes (PPROM) would be characterized by a unique miRNA expression profile. I observed the gene expression of 15 placental specific miRNAs (miR-512-5p, miR-515-5p, miR-516b-5p, miR-517-5p, miR-518b, miR-518f-5p, miR-519a-5p, miR-519d-3p, miR-519e- 5p, miR-520a-5p, miR-520h, miR-524-5p, miR-525-5p, miR-526a and miR-526b-5p) in placental tissue of pacients with PTB, PPROM and women with term in labor pregnancies (FG). PTB group consisted of 24 pacients, PPROM group of 75 pacients and FG group of 20 pacients. Quantitative real-time PCR was used to quantify gene expression. In the group of PTB pregnancies I identified 3 significantly upregulated miRNAs (miR-516b-5p, miR-519d-3p and miR-524-5p) and 4 miRNAs (miR-518b, miR-519a-5p, miR-520h and miR-526a) with a trend to upregulation compared to controls (FG). In the group of PPROM pregnancies I identified 3 miRNAs (miR-519d-3p, miR-520h and miR-256b-5p) with a trend to...
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Using of quantitative DNA method as a screening tool for effecient genotyping of samples in forensic DNA laboratory.
Koljenšič, Ivana ; Stenzl, Vlastimil (advisor) ; Daňková, Pavlína (referee)
Quantification of human DNA in forensic samples is an important step during STR profiling because the STR genotyping is sensitive to the quantity of DNA used in the PCR reaction. This study focuses on the importance of quantification in the entire process of genetic analysis. Two real time PCR platforms (Roche LightCycler480 System and ABI 7900 RT PCR) were used to compare two commercial kits in terms of DNA quantification. It was found out that accuracy of absolute quantification values in commercial quantification kits is strongly dependent on the construction of calibration curve. Especially low template DNA samples were used to assess whether QuantifilerTM or Plexor® HY System can determinate a minimum quantification value (cut off value) below which STR profiles would consistently fail to be detected. The usage of Plexor® HY System enabled to determine the cut off quantification value more exactly probably due to different molecular background and chemistry used in this kit. Reliability and other issues connected with cut off value are discussed. In order to better understand the relationship between the quantity of DNA and the number of detectable loci series the dilution experiment with standard DNA007 was done. Quantitative and qualitative consequences of input DNA amount in evaluation of...
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Human gut microbiome: Origin, ontogenetic development, diversity and its use in anthropology
Dvořáková, Barbora ; Daňková, Pavlína (advisor) ; Schierová, Dagmar (referee)
Human gut microbiome is a broad term encompassing all microorganismal life inhabiting the human gut. The bacteria living in the human gut represents the largest group of the human microbiome, make up the significant percentage of the human cellular composition and their genomes comprise a big part of the human genome. Gut microbiome has a significant role in human health and changes throughout the human life in reaction e.g. to change of diet and medical drug usage. This work pursues the acquisition and development of gut microbiome, the factors influencing its formation and diversity, and its use in anthropology.
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Cytogenetic methods in genotoxicology
Bártů, Linda ; Daňková, Pavlína (advisor) ; Langová, Martina (referee)
We are constantly exposed to a variety of factors which may be a cause of DNA mutations. The influence of mutagens of physical, chemical and biological origin is studied by genotoxicology. Ionic radiation is among the most common physical mutagens, benzene, vinylchloride or some drugs represent the chemical mutagens, while some viruses and may act as biological mutagens. The repair mechanisms of double strand breaks can be divided into those that require HRR-homologous sequences and those that may use of microhomologies consisting of a short DNA sequence (NHEJ). Both mechanisms can lead to aberrations of chromosomes, if they are not precise. Acquired chromosomal aberrations include translocation, common in cancer cells; deletion; or the production of acentric fragments, dicentrics and rings. Chromatid aberrations includes chromatid breaks and chromatide exchanges. There are various methods for detecting/examining such mutations and these can be categorised according to the phases of the cell cycle. The basic method is clasic Giemsa stain which reveals the most of aberrations except translocations and inversions and numeric abnormalities in metaphasic cells. Another way of testing mutagenicity is determining the rate of sister chromatide exchange; or the so called micronucleus test used to measure...
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Mechanisms of action of bisphenol A with emphasis on metabolism and fertility
Houška, Jakub ; Daňková, Pavlína (advisor) ; Novák, Jan (referee)
Bisphenol A is an endocrine disruptor, a chemical which is found in environment and also in water and food consumed by people and which disrupts endocrine system of humans and other organisms. Being endocrine disruptor it has a wide scale of negative effects on human health. I have attempted reviewing the molecular mechanism of its action with special respect to obesity and reproduction in this paper including interactions of bisphenol with specific receptors, its impact on enzyme synthesis and also on epigenetic mechanisms as DNA methylation or changes in miRNA expression. Possible ways of elimination of bisphenol A effects are examined in the end of this work. Keywords bisphenol A, molecular mechanisms, obesity, fertility, receptor, epigenetic mechanisms, elimination of effects
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Endometriosis, its aetiopathogenesis and influence on female's fertility
Selková, Katarína ; Daňková, Pavlína (advisor) ; Crha, Igor (referee)
Endometriosis is an estrogen-dependent disease of women of reproductive age. It is often considered as one of the main causes of female infertility. Its development and progression are influenced by several exogenous and endogenous factors. It is a heterogeneous disease and one of the most important tasks is to identify the candidate genes. That will allow us to better understand why and under what circumstances endometriosis develops. Because not all women with endometriosis are infertile, I focused in this thesis at selected candidate genes, their alleles and epigenetic dysregulation that could link endometriosis to infertility. From the available literature WNT4, HOXA gene cluster, ESR1, PGR, FN1, VETZ, GREB1 appear to play a major role in the pathogenesis of endometriosis. These genes also play an important role in various biological pathways and their dysregulation leads to low implantation / complete implantation failure, insufficient oogenesis, embryogenesis, and infertility in patients with endometriosis. Identification of genes whose dysregulation induces the development of endometriosis in the population is important, given the increasing incidence of endometriosis and infertility rates in recent years. With further research on pathogenesis of endometriosis it would be possible to make...
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Role of immune and adipose cells in the development of adipose tissue inflammation induced by stress associated with obesity
Kračmerová, Jana ; Rossmeislová, Lenka (advisor) ; Cahová, Monika (referee) ; Daňková, Pavlína (referee)
1 SUMMARY Obesity and overfeeding are associated not only with increased circulating levels of nutrients and metabolites, but also with increased risk of the development of additional disorders, such as cardiovascular diseases, cancer or insulin resistance. Plausible link between obesity and its comorbidities is inflammatory state, observed on the whole body level as well as in AT. As possible initiators of this inflammation, hypertrophied adipocytes were suggested. Adipocytes per se secrete a spectrum of heterogeneous molecules including cytokines. Under the stress conditions, adipocytes and subsequently AT resident immune cells switch to pro-inflammatory state and via secretory signaling attract additional immune cells. Furthermore, hypertrophic adipocytes release higher levels of metabolites that may also contribute to pro- inflammatory polarization of immune cells, mainly macrophages. General aim of this thesis was to investigate connection between impaired levels of nutrients and pro-inflammatory statue and activation of immune cells in healthy (obese and lean) subjects. In the Part one of this thesis, we analyzed acute reaction of immune cells in circulation and AT on artificially elevated levels of nutrients, imitating its increased values typical for metabolic syndrome. HFM ingestion led to...
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The significance of extracellular DNA in osteoclastogenesis from peripheral blood precursors - in vitro study
Jelínková, Ivana ; Daňková, Pavlína (advisor) ; Korabečná, Marie (referee)
Introduction: Extracellular DNA (ecDNA) is a common component of blood plasma. Increased levels of ecDNA in plasma can be found in some autoimmune diseases like systemic lupus erythematosus (SLE), rheumatoid arthritis or celiac disease which are associated with inflammatory processes. These diseases are also associated with an increased risk of osteoporosis. Bone is a dynamic structure undergoing constant modelling caused by osteoblasts, osteocytes and osteoclasts. Shifting their equilibrium can lead to pathological conditions such as osteoporosis. In this thesis we focused on elucidating whether ecDNA, an inflammatory agent with proven immunoregulatory effects can alter differentiation potential of monocytes and alternatively lead to osteoclastogenesis via TLR9. Material and methods: We obtained monocytes from peripheral blood of healthy donors and cultivated them with four types of ODNs control (CO), stimulatory (ST), inhibitory (INH, telomeric (TLM) with phosphodiester (-pO) or phosphorothioate (-pS) backbone for two weeks to establish their effect on differentiation potential of monocytes into osteoclasts. Osteoclastogenesis was evaluated by number of yielded osteoclasts observed on a light microscope. To establish the effect of ODNs on osteoclast activity samples were analysed by qPCR for...
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Role of inflammation in pathologic bone resorption in axial spondyloarthritis
Šebová, Eva ; Daňková, Pavlína (advisor) ; Tencerová, Michaela (referee)
Introduction: Axial spondyloarthritis (ax-SpA) is an inflammatory rheumatic disease. It is a unique model of bone remodeling disorders because, although one of the main diagnostic parameters is the rate of bone formation, inflammation present in patients' bodies increases the risk of pathological bone resorption, which can lead to osteoporosis. The processes of pathological resorption in ax-SpA have not been fully investigated, both in the disease as such and in the individual forms of the disease, i.e. non-radiographic (nr-axSpA), radiographic axial spondyloarthritis (r-axSpA) and ankylosing spondylitis (AS). This work deals with the influence of inflammatory serum of patients on the process of osteoclast differentiation from peripheral precursors of patients and healthy donors. Material and methods: Monocytes separated from the peripheral blood of either axSpA patients or healthy donors were stimulated for 14 days in vitro with serum from patients and in parallel with serum of age and sex of the corresponding healthy donors. Osteoclasts were evaluated as multinucleated, TRAP positive cells. Their numbers were statistically processed. Results: The inflammatory serum environment of patients with axSpA stimulated the osteoclastogenesis of axSpA monocytes significantly more (P <0,05) than the...
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Marfan syndrome, the main genetic causes
Šinknerová, Eliška ; Rothová, Olga (advisor) ; Daňková, Pavlína (referee)
This thesis is a review with the topic of Marfan syndrome (MFS) that hasn't been sufficiently described in Czech scientific literature yet. The first chapters pursue general informations and history of research of MFS, which was first desribed in 1896 and following research continued. During 1990s the main interest was to describe manifestations of Marfan syndrome and to find main genetic causes. Research continues until today and it is focused on searching specific mutations causing this syndrome. The thesis furthermore describes clinical manifestations classical MFS and neonatal MFS, e. g. manifestations in many body systems as sketal, cardiovascular, pulmonar and other systems. Thesis is focused on process which enables to diagnose MFS. The main part of the thesis is a desctiption of molecular nature and genetic aspects of this syndrome. The thesis describes in detail gene FBN1 and different types of mutationon which are responsible for formation of MFS. Attention is paid to genotype/phenotype correlation. This thesis also includes suggestions for work with this topic (MFS) in teaching at high schools and grammar schools.
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