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Gonadal development during the lifetime of the fastest maturing model vertebrate- turquoise killifish (Nothobrachius furzerí)
LANDOVÁ, Magdaléna
Turquoise killifish had to adapt to the inhospitable conditions in which they live, especially drying temporal water bodies, which means certain death. The life sprint of the representatives of this genus is at its peak within one-month post-hatching, when both sexes have fully developed gonads and can reproduce. This rate comes with a high cost, as the killifish gonads begin to show signs of tissue degradation and germ cell apoptosis as early as three months post-hatching. Germ cell loss increases with age. A description of the development and degradation of the gonads in males and their breeding was elaborated. For the evaluation of aging-specific changes, immunochemical methods were used, focusing on the binding of specific antibodies against target epitopes and their visualization using fluorescence microscopy. Procedures for histological specimens have also been described, both for classical light and fluorescence microscopy.
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Rozdíly ve vývoji gonád u kapra obecného ve věkové kategorii K2-K3 při rozdílných podmínkách chovu
FÁBRIK, Jan
The work contains information about development of gonads in carp aged 2-3 years in specified conditions. A breed of common carp (Cyprinus carpio, L) called the amur mirror carp, which was kept in RAS and ponds, was used for the research. The objective was to confirm the hypothesis that it is possible to reduce the generation interval in females from 4-5 years to 3. The observed values were fish weight, gonadal weight, gonadosomatic index, Fulton's coefficient and degree of oocyte development. Another factor examined was the dependence of GSI on oocyte size.
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Gasotransmise v epigenetických regulacích gametogeneze a embryogeneze
BRICHCÍN, Jiří
As the problems with the reproduction of livestock and humans starting to increase, the need for knowledge of mechanisms involved in regulating the correct process of gametogenesis and embryogenesis also rises. For the experimental part of this work, two components from two different, mostly separately explored fields, i.e. gasotransmitters and epigenetic mechanisms, which are necessary for the correct process of gametes production and early embryonic evolution were chosen. Hydrogen sulfide was chosen from the series of gasotransmitters, and histone deacetylase Sirtuin 1 (SIRT1) was chosen as its possible substrate. Confirmation of the presence of these components was carried out on oocytes and embryos of laboratory mice (Mus musculus).
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Study of changes on spindle and chromosomes vitrified and in vitro maturated human oocytes
Holubová, Kateřina ; Kratochvílová, Irena (advisor) ; Libusová, Lenka (referee)
The aim of this work is the comparison of cryoprotection methods for human oocytes regarding their survival rate and subsequent success in in vitro fertilization. This work showed the effect of oocyte maturation characterized by the state of the meiotic spindle, chromosomes and polar body on the success of in vitro fertilization of standard (fresh) and vitrified oocytes. The acquired knowledge will lead to a better understanding of the relationships between the state of oocytes and specific in vitro fertilization and cryoprotection procedures. The final, application-interesting goal will be to propose the optimization of methods for working with oocytes so that the probability of their success in the IVF cycle is as high as possible.
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Study of changes on spindle and chromosomes vitrified and in vitro maturated human oocytes
Holubová, Kateřina ; Kratochvílová, Irena (advisor) ; Libusová, Lenka (referee)
The aim of this work is the comparison of cryoprotection methods for human oocytes regarding their survival rate and subsequent success in in vitro fertilization. This work showed the effect of oocyte maturation characterized by the state of the meiotic spindle, chromosomes and polar body on the success of in vitro fertilization of standard (fresh) and vitrified oocytes. The acquired knowledge will lead to a better understanding of the relationships between the state of oocytes and specific in vitro fertilization and cryoprotection procedures. The final, application-interesting goal will be to propose the optimization of methods for working with oocytes so that the probability of their success in the IVF cycle is as high as possible.
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Epigenetic and structural characteristics of mammalian oocytes and embryos: extrapolation for human ART
Langerová, Alena ; Fulka, Josef (advisor) ; Pěknicová, Jana (referee) ; Chod, Jiří (referee)
It is now more than 35 years since the first world test-tube baby, Louise Brown, was born (1978) in England and it is estimated that since then more than 4 000 000 of children were produced by in vitro fertilization (IVF) worldwide. The initial success of IVF was less than 20% in best clinics, but now it reaches about 40%. This is a consequence of introduction of new methods, standardization and exploitation of new manipulation and culture media, as well as the incorporation of research results. Nevertheless, the most important still remains the skill and experience of IVF clinics and IVF laboratories staff, especially their ability to critically evaluate the quality of biological material and to decide which cure and treatment are the best one. At least, some biological material (immature and low quality oocytes) can be used for training and also for some experiments aiming to explain some questions, which are not yet fully understood (for example aneuploidies in human oocytes and embryos). In addition, this training can facilitate the introduction of new progressive approaches and may also improve indirectly the quality of infertility treatments. The first part of thesis is focused on the quality evaluation of oocytes collected by aspiration from follicles of stimulated patients. For labeling...
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Kinázová signalizace v meióze I savčích oocytů
Brzáková, Adéla ; Šolc, Petr (advisor) ; Dráber, Pavel (referee)
PLK1 belongs to the extended family of serine/threonine kinases controlling the cell cycle. It is well known for its role in the control of mitosis and contributes also to the regulation of meiotic division. On a basis of Live Cell Imaging (LCI) experiments we can describe the phenotype of the oocytes with PLK1 inhibited by small molecular inhibitor BI2536. PLK1 inhibition leads to delayed nuclear envelope breakdown (NEBD) and chromatin condensation (CC) and also causes desynchronization of NEBD and CC; in contrast to control oocytes, PLK1 inhibited oocytes break down their nuclear envelope with chromatin almost fully condensed. Also duration of these two early nuclear events is prolonged in oocytes with inhibited PLK1. In contrast to somatic cells, PLK1 inhibition in mouse oocytes does not prevent assembly of spindle with two distinct poles but affects the final spindle volume. Similar to somatic cells, mouse oocytes with PLK1 inhibited from the beginning of the meiotic maturation stay arrested in metaphase I but in the case of mouse oocytes, this block is not dependent on Spindle Assembly Checkpoint (SAC) persisting activity. When mouse oocytes are synchronized on metaphase I/anaphase I transition by proteasome inhibition and then PLK1 kinase activity is inhibited, about 2/3 of the oocytes stay arrested...
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Single cell expression analysis of genes with potential mrna gradient in mouse oocytes
Dorosh, Andriy ; Margaryan, Hasmik ; Vodička, Martin ; Ergang, Peter ; Šídová, Monika ; Dvořáková-Hortová, Kateřina
In frogs, there are clearly visible differently pigmented animal and vegetal poles of the egg determined before fertilization and leading to asymmetrical divisions. Mammalian egg does not show any comparable differentiation and it has been generally accepted that even the individual blastomeres in 2-cell and 4-cell embryos are homogenous. However, recent findings suggest that those blastomeres display different gene expression patterns and might already possess some inclinations to specific cell lineages. We therefore raised a question, whether there could be any mRNA or protein gradients in pre-fertilization oocytes similar to a previously described amphibian egg one. In mammalian eggs, there is a membrane region that is poor in microvilli, cortical granules are absent beneath plasma membrane and sperm cells generally do not bind to this location. This microvilli free region also covers the egg nucleus, and cytoskeleton localization differs markedly to the rest of the cortical space, forming actin –myosin II cortical cap/ring and is considered as animal pole. The purpose of this study was to determine gene products that can be detected at single cell level using qPCR and display gradient like distribution in mature oocytes. We checked expression of 12 selected genes in a pool of 10 oocytes and single mature oocytes. Then, we analysed gene expression in fixed intact oocytes and those undergoing laser capture microdissection procedure (LCMD). Eventually, we have determined six candidate genes for the study of intracellular spatial gene expression in mature mammalian oocytes by subcellular qPCR and in situ hybridization.
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