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Cell cycle regulation and genome integrity protection in the early mammalian embryos
Knoblochová, Lucie ; Drutovič, David (advisor) ; Carr, Antony M. (referee) ; Fulková, Helena (referee)
(English version) Infertility is a major health problem, as it affects one in every six people worldwide (Njagi et al., 2023). One of the major reasons for infertility are aneuploidies, additions or losses of an entire or partial chromosome during cell division. Aneuploidies thus negatively influence cellular processes and potentially lead to developmental problems or embryo loss. It has been thought for a long time that aneuploidies arise mostly during oocyte development, and these mechanisms have been well studied. However, recent evidence has shown that aneuploidies arise also de novo after fertilisation and during the early embryonic development; but the molecular mechanisms of these abnormalities still remains elusive. Aneuploidies often originate during cell cycle division from unrepaired DNA damage in mitosis. DNA damage is sensed by DNA damage response (DDR) signalling pathways, which slow down or arrest cell cycle progression until it is resolved. An essential DDR factor during typical cell cycle progression is checkpoint kinase 1 (CHK1). However, the role of DDR factors in the early embryos, and especially CHK1, have not been well studied. Early embryonic development is regulated by maternal factors stored in the oocyte until the transcription of the embryonic genome begins. To study...
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Mathematical modeling of early embryogenesis processes
Dunajová, Michaela ; Šejnoha, Jiří (advisor) ; Drutovič, David (referee)
1 Abstract: Early embryogenesis is a fundamental phenomenon of developmental biology, which involves the establishment of the first structures within embryos. The work focuses on mathematical spatial models of mammalian blastulation, which were created based on spatial approach. We discuss and compare six mo- dels, which simulate the differentiation of cell lines and the formation of blas- tocyst structures. Models are based on local rules, using considerable cellular abstraction. Models focus on the processes of differentiation of the blastomere into the trophoectoderm and the inner cell mass, followed by the establishment of the structure of the epiblast and the primitive endoderm.
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The role of cumulus cells during the maturation of mammalian oocytes
Meniuková, Kateřina ; Krylov, Vladimír (advisor) ; Drutovič, David (referee)
Cumulus cells in the ovarian follicles emerge by differentiation from somatic granulosa cells. They are located around the oocytes in stratified clusters and their innermost layer is called the corona radiata. Cytoplasmic microvilli of cumulus cells pass through zona pellucida to the proximity of the oolema, where formation of gap junctions enables signalling and metabolic codependency. Oocytes produce factors affecting processes in cumulus cells, including their metabolism and viability. By modulating the abundance of cyclic nucleotides in oocytes, cumulus cells allow the maintenance of meiotic arrest, providing time for oocytes to finish their growth and gain competence for maturation. Because oocytes have a very limited ability to process glucose, metabolic cooperation with cumulus cells enables them to gain the energy needed for processes associated with growth and maturation. Several metabolic pathways of glucose processing create energy substrates for the formation of ATP, which they then transport to the oocytes. They also regulate the amount of lipid droplets in oocytes and protect them from cellular toxicity. An increase in the concentration of luteinizing hormone in the follicle just before ovulation induces the transmission of the signal for cumulus expansion. Interruption of the...
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NAD+-dependent histone deacetylase SIRT1 in the process of oogenesis, fertilization and early embryonic development
Valentová, Iveta ; Nevoral, Jan (advisor) ; Drutovič, David (referee)
SIRT1 is a histone deacetylase from the sirtuin family that affects epigenetic and non- epigenetic targets. We can assume that the known SIRT1 substrates are involved in the regulation of gametogenesis and early embryonic development. Our hypotheses say SIRT1 is present in oocytes and early embryos and it plays a physiological role in oocyte maturation, fertilization and early embryonic development. A mouse model of a conditional knock-out line producing Sirt1-deficient oocytes was developed to verify our hypotheses. Oocytes and embryos were analyzed for SIRT1, its selected substrates and other markers by immunocytochemistry. We found out that the presence of SIRT1 contributes to oocyte quality through modulation of the chromatin histone code and stabilization of the spindle. Furthermore, the purely maternal origin of SIRT1 presents in both zygote pronuclei. Last but not least we discovered a significant effect of SIRT1 on early embryonic development, probably mainly due to its role in the activation of the embryonic genome. The results confirm our hypothesis that SIRT1 is present in oocytes and embryos mainly around chromatin. The results show that SIRT1 is a maternal factor determining oocyte quality and it is necessary for the embryonic genome activation.
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Aurora kinase signaling during mammalian oocyte meiosis
Komrsková, Anna ; Drutovič, David (advisor) ; Petr, Jaroslav (referee)
Mammalian oocytes, especially human oocytes, are prone to defective chromosome segregation and are prone to aneuploidies, most commonly in the first meiotic division. Due to these defects, aneuploid embryos are formed after fertilization, resulting in numerous birth defects or infertility. The spindle formation in the first meiotic division in mammalian ooctes is a critical moment for preserving genome integrity. In mammalian oocytes, the spindle assembly is regulated via acentriolar microtubule-organizing centers and chromatine itself, through the activity of Aurora kinases and Ran-GTP gradient. Apart from the Aurora kinase family, other kinases regulate spindle formation, like Polo-like kinase 1 (PLK1). This thesis focused on potential cooperation between Aurora kinases and PLK1 or Ran-GTP. Our results show that the levels of phosphorylated Aurora kinase A (AURKA), but not levels of phosphorylated Aurora kinase B or Aurora kinase C, decrease after pharmacological inhibiton of PLK1. Levels of phosphorylated Aurora kinases do not change after pharmacological inhibition of Ran-GTP signaling. Using live-cell microscopy we showed that overexpression of PLK1 can rescue the phenotype observed in Aurka depleted oocytes. Next, we found that at least one of the signal pathways at chromosomes is necessary...
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Formation of pole assymetry during the Xenopus laevis oocytes growing
Pácalová, Eliška ; Vávra, Jiří (advisor) ; Drutovič, David (referee)
A frog Xenopus laevis is an important model organism used in developmental and cell biology and physiology. Oocytes of this species are frequently used for study of mechanisms of oogenesis and membrane proteins. Through oogenesis an animal-vegetal axis is established by asymmetric distribution of determinant molecules which are important in embryogenesis and establishment for another body axes. The main aim of this thesis is to overview transport mechanisms for establishment of asymmetric molecule (especially mRNA) and organelles asymmetry to animal and vegetal poles and their regulation. Key words: Xenopus laevis, oocyte, animal pole, vegetal pole, cytoskeleton, intracellular transport, mitochondrial cloud, METRO, early pathway, late pathway
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The role of protein kinases in spindle formation and chromosome segregation during mammalian oocyte meiosis
Komrsková, Anna ; Drutovič, David (advisor) ; Libusová, Lenka (referee)
Correctly assembled spindle during the first meiotic division is crucial for proper chromosome segregation. Protein kinases from Aurora kinase and Polo-like kinase families are an essential part of mechanisms that affect spindle assembly and take part in chromosome segregation. These kinases play a very specific role in these processes both individually and in cooperation. A defective spindle assembly is often followed by incorrect chromosome segregation, that can lead to aneuploidy. Some aneuploidies cause infertility, others lead to birth defects like Down's syndrome. Human oocyte meiosis is more sensitive to errors in chromosome segregation than mitosis, meiosis in human sperm or meiosis in female gametes in other organisms. This tendency makes the research of Aurora kinases and Polo-like kinases very appealing and eventually rewarding in the context of human reproduction. Keywords Meiosis, oocytes, spindle, microtubule-organising centers, protein kinases, Aurora kinases, Polo-like kinases
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