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Molecular aspects of mammalian oocyte and embryo quality
Chyská, Anna ; Vaškovičová, Michaela (advisor) ; Jansová, Denisa (referee)
According to statistics in the Czech Republic, up to 20 % of couples experience infertility, which is still rising. One of the infertility treatment options is to undergo one of the assisted reproduction methods. In vitro fertilization (IVF) is among the most commonly used methods. In order to increase the success rate of fertilization, it is preferable to have a quality oocyte with specific characteristics. Such an oocyte is then a prerequisite for a good quality embryo, a correct course of pregnancy, and proper fetal development. Meiosis, a key step in oocyte formation, is regulated by complex signalling pathways. If these pathways are dysfunctional, the chances of the oocyte developing into a competent embryo are reduced. For example, errors in segregation during meiosis lead to aneuploidy, which is a major cause of miscarriages and birth defects. Therefore, understanding the signaling pathways can help identify the origin of the errors that result in aneuploidy and thus contribute to improving women's reproductive health. Key words: aneuploidy, spindle, in vitro fertilization, spindle assembly checkpoint, oocyte quality, chromosome segregation
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Smell tests in Alzheimer's disease diagnostics and their cognitive correlates
Vaškovicová, Michaela ; Lukavský, Jiří (advisor) ; Krámská, Lenka (referee)
This thesis is focused on the use of olfactory tests in neuropsychological diagnostics of Alzheimer's disease and on the cognitive correlates of these tests. The literature review briefly summarizes current knowledge of the diagnostics of Alzheimer's disease and its dynamically developing criteria. Attention is also paid to the olfactory impairment that occurs during the course of this disease, as well as to specific psychophysical olfactory tests. The research part summarizes the results of research within the grant GACR 17-05292S, 01. 01. 2017 - 31. 12. 2019 - New blood biomarkers for early diagnosis, prognosis and course of Alzheimer's disease held at the Psychiatric Clinic of the First Faculty of Medicine and General Teaching Hospital. The exploratory analysis examines possible cognitive correlates of the olfactory Sniffin' Sticks test. Practical experience with olfactory test is evaluated in the discussion. Keywords Alzheimer's disease, olfactory deficit, smell tests, Sniffin'-Sticks, neuropsychological diagnostics
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Recognition of expressed double-stranded RNAs in mammalian cells
Vaškovičová, Michaela ; Svoboda, Petr (advisor) ; Petr, Jaroslav (referee)
Long double-stranded RNA (dsRNA) is a unique structure formed during viral replication or transcription of repetitive elements. Mammalian cells evolved several mechanisms how to respond to dsRNA. dsRNA can be engaged in one of three pathways: interferon response, RNA editing, and RNA interference (RNAi). RNAi is evolutionary conserved effect of dsRNA, which results in sequence-specific messenger RNA degradation. However, in mammals, RNAi is functional only in mouse oocytes, which express truncated version of Dicer (DicerO ). In somatic cells, dsRNA triggers sequence-independent interferon pathway. The main aim of this Master's thesis was to examine how specific double-stranded RNA-binding proteins (DRBPs) influence distribution of long dsRNA into RNAi and sequence-independent pathways. We used a luciferase-based reporter RNAi assay to monitor sequence-specific and sequence-independent effects of dsRNA co-expressed with selected DRBPs. Our results suggest that none of the tested DRBPs is sufficient to stimulate RNAi in somatic cells. Interestingly, the overexpression of either TARBP2 or PACT suppressed RNAi in cells expressing DicerO . Moreover, microRNA pathway, which employs the same protein factors as RNAi, is not inhibited by TARBP2 or PACT. Therefore, we propose that DRBPs overexpression...
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Smell tests in Alzheimer's disease diagnostics and their cognitive correlates
Vaškovicová, Michaela ; Lukavský, Jiří (advisor) ; Krámská, Lenka (referee)
This thesis is focused on the use of olfactory tests in neuropsychological diagnostics of Alzheimer's disease and on the cognitive correlates of these tests. The literature review briefly summarizes current knowledge of the diagnostics of Alzheimer's disease and its dynamically developing criteria. Attention is also paid to the olfactory impairment that occurs during the course of this disease, as well as to specific psychophysical olfactory tests. The research part summarizes the results of research within the grant GACR 17-05292S, 01. 01. 2017 - 31. 12. 2019 - New blood biomarkers for early diagnosis, prognosis and course of Alzheimer's disease held at the Psychiatric Clinic of the First Faculty of Medicine and General Teaching Hospital. The exploratory analysis examines possible cognitive correlates of the olfactory Sniffin' Sticks test. Practical experience with olfactory test is evaluated in the discussion. Keywords Alzheimer's disease, olfactory deficit, smell tests, Sniffin'-Sticks, neuropsychological diagnostics
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Olfactory deficiency in neuropsychological diagnostics of Alzheimer's disease
Vaškovicová, Michaela ; Kulišťák, Petr (advisor) ; Dvořáková, Zuzana (referee)
This bachelor thesis is focused on the application of olfactory deficiency in neuropsychological diagnostics of Alzheimer's disease. Firstly, in the theoretical part, the work shortly summarizes current knowledge regarding neuropsychological diagnostics of Alzheimer's disease, with an emphasis on the new diagnostics criteria and usage of various kinds of biomarkers. Furthermore there is thoroughly covered the topic of olfactory deficiency as an early biomarker of Alzheimer's disease, while focusing on olfactometry employing psychophysical subjective tests. The work also stresses out the possibility of cognitive conditionality of some of the smell tests, most prominently the odor identification test. The issue of cognitive demands of smell tests is also further entertained in the research part, where a study of possible cognitive correlates of the Odor Identification test is proposed. For this purpose was chosen the Odor Identification test from the extended set of Sniffin' Sticks and the neuropsychological test battery Uniform Data Set. The main presumed candidates for cognitive correlates of the smell test are tests of language functions, especially the Boston Naming Test measuring naming ability. The research proposed here is based on controlling influence of cognitive deficits on the Odor...
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Double strand DNA breaks response in Huntington´s disease transgenic minipigs
Vaškovičová, Michaela ; Šmatlíková, Petra ; Herbert, A. ; Motlík, Jan ; Šolc, Petr
Huntington’s disease (HD) is progressive neurodegenerative disorder caused by presence of CAG expansion in the huntingtin gene, which gives rise to mutated form of huntingtin protein (mHtt). There is a strong evidence that DNA damage response is compromised by presence of mHtt in cells and increase of double strand DNA breaks (DSBs) is an early event in HD pathology. It was shown, that level of γH2AX is significantly higher in R6/2 mice compared to wild-type animals. Moreover, level of γH2AX is higher also in striatal neurons and fibroblasts of human HD patients. Furthermore, protein p53, key player in DNA damage response, is hyperactivated in cells expressing mHtt and inhibition of p53 or ATM ameliorates phenotypes of HD animal models. However, exact mechanism of mHtt action is not clear and therefore further investigation of mHtt effects on DSBs response is very important for the understanding of HD pathology.
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Oxidative stress in primary porcine fibroblasts expressing mutated huntingtin
Šmatlíková, Petra ; Askeland, G. ; Vaškovičová, Michaela ; Klíma, Jiří ; Motlík, Jan ; Eide, L. ; Ellederová, Zdeňka
Molecular events, such as protein aggregation, mitochondrial dysfunction, and transcriptional dysregulation have been linked to Huntington’s disease (HD) pathogenesis. Oxidative stress has been considered as one of the key players in disease progression. Though, it is still not clear whether oxidative stress causes HD, or if it is a consequence of other primary events.
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The use of RNA affinity tags in RNA biology
Vaškovičová, Michaela ; Svoboda, Petr (advisor) ; Drbal, Karel (referee)
RNA molecules play essential role in many important processes in living organisms. Messenger RNAs transmit genetic information from the DNA to the cellular sites of protein synthesis. RNAs, which do not encode for proteins, have numerous specialized functions in regulation of gene expression, splicing, transport, localization, translation, and stability of RNAs. For example, ribosomal and transfer RNAs are required for the translation of proteins; small nuclear RNAs are involved in splicing, and small nucleolar RNAs modify other RNAs. MicroRNAs and small interfering RNAs are essential regulators of gene expression. Other non-coding RNAs regulate chromatin and transcription. RNAs are typically present in ribonucleoprotein (RNP) complexes containing various proteins, which are essential for functions of the complexes. Understanding RNA-protein interactions and composition of RNP complexes is critical for delineating their functions. The common strategy in the study of RNP complexes is tagging the RNA component, in order to isolate, purify and analyze the tagged RNAs with their binding partners. Tagged RNP complexes can be subsequently subjected to further studies. In the first part of my work, I describe four basic strategies of tagging the RNA, which can be used for study of RNAs and RNP complexes....
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Recognition of expressed double-stranded RNAs in mammalian cells
Vaškovičová, Michaela ; Svoboda, Petr (advisor) ; Petr, Jaroslav (referee)
Long double-stranded RNA (dsRNA) is a unique structure formed during viral replication or transcription of repetitive elements. Mammalian cells evolved several mechanisms how to respond to dsRNA. dsRNA can be engaged in one of three pathways: interferon response, RNA editing, and RNA interference (RNAi). RNAi is evolutionary conserved effect of dsRNA, which results in sequence-specific messenger RNA degradation. However, in mammals, RNAi is functional only in mouse oocytes, which express truncated version of Dicer (DicerO ). In somatic cells, dsRNA triggers sequence-independent interferon pathway. The main aim of this Master's thesis was to examine how specific double-stranded RNA-binding proteins (DRBPs) influence distribution of long dsRNA into RNAi and sequence-independent pathways. We used a luciferase-based reporter RNAi assay to monitor sequence-specific and sequence-independent effects of dsRNA co-expressed with selected DRBPs. Our results suggest that none of the tested DRBPs is sufficient to stimulate RNAi in somatic cells. Interestingly, the overexpression of either TARBP2 or PACT suppressed RNAi in cells expressing DicerO . Moreover, microRNA pathway, which employs the same protein factors as RNAi, is not inhibited by TARBP2 or PACT. Therefore, we propose that DRBPs overexpression...
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