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Transcriptional regulation of miR-17-92 microRNA cluster during macrophage differentiation.
Rybářová, Jana ; Stopka, Tomáš (advisor) ; Pospíšek, Martin (referee)
miR-17-92 cluster (Oncomir1) encodes seven microRNAs (miRNA, miR) regulating many biological processes including proliferation, differentiation or apoptosis. Overexpression of microRNAs encoded by miR-17-92 cluster is found in a number of tumors including acute and chronic myeloid leukemias (Dixon-McIver et al., 2008; Li et al., 2008; Venturini et al., 2007). Myeloid progenitors express miR-17-92 cluster at a high level, while macrophage differentiation associates with its downregulation. Our laboratory found, that miR-17-92 cluster is repressed by transcription factor Early growth response 2 (Egr2) upon differentiation of primary myeloid PUER progenitors, induced with transcription factor PU.1. Aim of this thesis is to further test the abovementioned data by preparing a reporter vectors set, carrying various fragments of miR-17-92 putative promoter, which enables us to study regulation of transcription of miR-17-92 cluster. This task complicated by presence of increased GC content of the miR-17-92 promoter was successfully accomplished resulting in amplification of eight fragments containing the various parts of miR-17-92 promoter including region -3.3 to 0 kb relative to the start of miR-17-5p sequence, that were inserted into pGL3 reporter vector. Transfection of pGL3 reporter vector carrying...
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Epigenetické regulace u autoimunitních onemocnění se zaměřením na revmatoidní artritidu
Horková, Veronika ; Daňková, Pavlína (advisor) ; Hušáková, Markéta (referee)
Exact cause of rheumatoid arthritis, as well as other autoimmune diseases has not been identified yet. In last twenty years, epigenetics showed a new face of immune system. DNA methylation, modification of histones - proteins around which DNA is wrapped, or interference of small RNA sequences - microRNAs, these all are heritable changes outside the DNA sequence that provide another component involved in autoimmunity. Presented epigenetic mechanisms alter gene expression and thus facilitate production of pro- inflammatory factors leading to autoimmune reactions. Moreover, genes regulating apoptosis are also frequently targeted by epigenetic modifications. Not only these mechanisms provide another level of immune defense, they also explain higher female susceptibility to autoimmune diseases and the influence of environment on pathogenesis of these diseases.
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Diagnostic and prognostic impact of miR-122 and miR-142 in colorectal cancer
Kalkušová, Kateřina ; Červená, Klára (advisor) ; Král, Jan (referee)
Colorectal carcinoma is one of the most frequent cancers and it is also one of the most common causes of cancer related death. It is often diagnosed at a late stage which is associated with a worse prognosis. Therefore, there is an effort to identify new biomarkers that allow earlier diagnosis. These biomarkers could predict prognosis of disease and patient's response to treatment. Short noncoding RNAs - microRNA - appears to be promising candidates, these molecules are involved in many cellular processes through RNA interference. The aim of this thesis is to summarize knowledge about application of miR-122 and miR-142 as diagnostic and prognostic biomarkers for colorectal cancer. Keywords: microRNA, biomarkers, colorectal cancer, diagnostics, prognosis
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Vývoj chemických regulátorů drah mikroRNA a RNAi
Bruštíková, Kateřina ; Svoboda, Petr (advisor) ; Bařinka, Cyril (referee) ; Pospíšek, Martin (referee)
MicroRNAs are noncoding RNAs inducing sequence-specific posttranscriptional inhibition of gene expression and represent the major class of small endogenous RNAs in mammalian cells. Over 2,500 of human microRNAs potentially regulating more than 60% of human protein-coding genes have been identified. MicroRNAs participate in the majority of cellular processes, and their expression changes in various diseases, including cancer. Currently, there is no efficient small chemical compound available for the modulation of microRNA pathway activity. At the same time, small chemical compounds represent excellent tools for research of processes involving RNA silencing pathways, for biotechnological applications, and would have a considerable therapeutic potential. The presented work represents a part of a broader project, whose ultimate goal is: (i) to find a set of small molecules allowing for stimulation or inhibition of RNA silencing and (ii) to identify crosstalks between RNA silencing and other cellular pathways. This thesis summarizes results from the first two phases of the project, the development of high-throughput screening assays and the high- throughput screening (HTS) of available libraries of small compounds. To monitor the microRNA pathway activity, we developed and optimized one biochemical...
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Extracellular microRNAs in hematological malignancies and their use for diagnosis and treatment monitoring
Šulcová, Dominika ; Pospíšil, Vít (advisor) ; Čermák, Vladimír (referee)
MicroRNAs are short non-coding RNAs that negatively regulate gene expression at post-transcriptional level by interfering with mRNA translation and stability. Recently, microRNAs were surprisingly found to be present in various body fluids including blood plasma and serum, cerebrospinal fluid, saliva, milk or urine. These extracellular microRNAs are resistant to RNases and stable in high temperature or pH. Extreme stability of extracellular microRNAs is caused by their association with protective protein complexes (mostly with Argonaute proteins). MicroRNAs are frequently deregulated in cancer and specific tumor- related microRNAs can be also detected in body fluids, indicating that extracellular microRNAs can be used as tumor specific markers. This Bachelor thesis reviews basic principles of microRNA function and biogenesis with focus on extracellular microRNAs and their role in intercellular communication, and it highlights the role of extracellular microRNAs in hematological malignancies and their possible use in diagnosis and treatment.
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Extracellular microRNAs and their role in pathologies especially in the field of gynecology and obstetrics.
Štěrbová, Monika ; Hromadníková, Ilona (advisor) ; Balušíková, Kamila (referee)
microRNAs (miRNAs) represent a relatively newly discovered group of RNA molecules and they serve to regulate gene expression. In spite of processes of differentiation, proliferation and apoptosis, miRNAs influence the whole biological systems, such as embryogenesis, oncogenesis, and immunity. There have been a number of experiments in recent years concerning diagnoses and predictions of complications during pregnancy, and tumour growth. Extracellular miRNA molecules participating in circulation of patients are used in the non-invasive diagnostics. RNA molecules usually get into the extracellular fluid during the apoptosis process. I chose four diseases, which extracellular miRNA have diagnostic potential - preeclampsia, intrauterine growth retardation, gestational diabetes mellitus and breast cancer - for my work. An aberrant expression of different levels of various extracellular miRNAs has been reported in these diseases but the clinical use of microRNAs in the diagnosis and prediction of those still requires further research and optimization. Keywords: breast cancer, extracellular nucleic acids, fetal growth retardation, gestational diabetes mellitus, microRNA, PCR, preeclampsia
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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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The role of microRNA in hypoxic pulmonary hypertension
Tichý, Václav ; Hampl, Václav (advisor) ; Kolář, David (referee)
Pulmonary Arterial hypertension (PAH) is a devastating progressive disease that significantly decreases quality of life and has the average survival rate of only few years. One of the significant initiators of PAH is chronic hypoxia. After more than six decades of research that was initiated in 1946 by von Euler and Liljestrand, a new group of potential regulators of this pathology was discovered, that became heavily studied in the last five years. They are highly conserved molecules belonging to non-coding RNA. These 19-23 nucleotides long microRNA (miRNA) act as negative regulators of expression on various proteins. Many of them regulate traditional signalling pathways of hypoxic PAH (HIF-1, BMPR2) and miRNA is in turn regulated by other signalizations. Together, that creates an interconnected network of direct and indirect interactions and feedback loops, that we need to study in order to understand hypoxic PAH. This thesis summarizes findings about important miRNA molecules from the last few years and elucidates part of these regulatory mechanisms on several miRNA molecules (miR-17-92, miR-21, miR-210, miR-204 a miR143/145).
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Study of the regulatory properties of oncogenic microRNAs under normal and pathologically altered conditions in order to detect new tumors.
Dusílková, Nina Borisovna ; Stopka, Tomáš (advisor) ; Machová Poláková, Kateřina (referee) ; Rohoň, Peter (referee)
Oncogenic microRNAs (miRNAs) are small RNA molecules that inhibit post-translational regulatory mechanisms at the epigenetic level. miRNAs are often deregulated in malignancies and due to their stability are detectable in non-cellular fractions of peripheral blood. In our laboratory, we have performed several studies that have investigated and utilized miRNAs as biomarkers for various hematological tumors (e.g., chronic lymphocytic leukemia, Hodgkin`s lymphoma) and solid tumors (e.g., breast cancer). The aim of these studies was to find the association of miRNAs with pathophysiological and clinical aspects of each disease. Here, we confirmed the importance of particular miRNA or its complex during disease monitoring. Combining clinical, molecular biological and statistical analyses, we were able to find miRNA sets that fulfilled not only a diagnostic role but also a prognostic role beyond expectations. The main focus of this thesis is on the investigation of microRNAs in the diagnosis of a hematological malignancy - primary cutaneous T-cell lymphoma (CTCL). Tumor specificity of some miRNAs has been demonstrated. Their aberrant expression in tissue samples of CTCL patients obtained from skin biopsies, correctly distinguished malignant disease from control samples of benign skin lesions. Here, we...
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Postpartum expression of cardiovascular disease-associated microRNAs - comparison of expression levels between plasma, plasma exosomes and whole peripheral venous blood
Ševčíková, Adéla ; Hromadníková, Ilona (advisor) ; Korabečná, Marie (referee)
MicroRNA (miRNA) are short non-coding RNA molecules that regulate gene expression at the post-transcriptional level. Many miRNAs are involved in the pathogenesis of cardiovascular diseases, which is associated with altered gene expression. This work compares miRNA gene expression profiles among various biological sources - whole peripheral venous blood (whole PB), plasma and plasma exosomes. For all tested groups combined, the expression levels of miRNA were maximal in whole PB and lowered in plasma and plasma exosomes, and the expression levels of miRNA were higher in plasma than in plasma exosomes, except miR-126-3p, which had a higher level detected in plasma exosomes compared to plasma. This work also compares expression levels of cardiovascular miRNA between women with anamnesis of gestational diabetes mellitus (GDM) and physiological gravidity 3-11 years postpartum in whole PB, plasma and plasma exosomes. In whole PB, 12 of 29 tested miRNAs were up-regulated in women with prior exposure to GDM. MiR-181a-5p was up-regulated in plasma exosomes and miR-499a-5p in plasma in women with prior exposure to GDM. The changes in whole peripheral venous blood seem to reflect the complex systemic response to the changes that occurred during the onset of GDM. Women with aberrant epigenetic profiles may...
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