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Principles of ovarian stimulation and oocyte collection in mouse and human.
Klinovská, Karolína ; Hortová, Kateřina (advisor) ; Svoboda, Petr (referee)
This thesis deals with methods of controlled ovarian stimulation and oocyte retrieval on physiological basis. Mouse has been major model organism for wide range of key researches and being able to stimulate its ovarian functions correctly is necessary. That is why mouse stimulation is main topic of this thesis. It summarizes available data and stimulation protocols in relation to factors which can affect the result as a condition, strain, quality of hormones used in administration and others. Potential affections on oocyte and embryo quality are mention according to specific types of administration protocols. Human infertility is slowly becoming common and one of the techniques used to treat it in assisted reproduction is also ovarian stimulation. In the end of this thesis is a chapter dealing with this commonly used method. It is focused on stimulation for oocyte retrieval, as well as on stimulation of recipient mother in embryotransfer. Differences based on hormones, success in In vitro fertilization programme (IVF) and application frequency are mentioned.
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Analysis of gene expression during spermatogenesis in mice
Suchan, Jan ; Stopka, Pavel (advisor) ; Hortová, Kateřina (referee)
Spermatogenesis is very unique and complex process regulating and be amenable to reproductive behaviour equally. These processes are affected by outer factors such as social stimuli, represented by interaction between male and female. Mice belong to mammals with high sensitivity to chemical signals. Hence pheromonal communication takes important part in their life. Pheromones have effect on expression activity in testicular tissue throught hormonal pathways. A target of these regulations are for example proteins participating in immunity or competition actions. Phenotype plasticity might proceed on the level of DNA as well as on the level of gene expression. That's why we take interest if factors like social contacts are able to significantly change expression of genes involved in reproduction pathways. We are also interested in possibility that expression changes of target genes take part in speciation events. In this study we demonstrate the effect of short-term social contact to gene expression of biochemical pathways involved in spermatogenesis and by other pathways linked together with reproductive bahaviour. This involvement might be direct or indirect. We found genes, whose level of expression in testicular tissue is different between species Mus m. musculus and Mus m. domesticus. Here I provide...
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The Influence of eco-toxicological factors on spermatogenesis, morphology and function of mammalian sperm
Vašinová, Jitka ; Hortová, Kateřina (advisor) ; Pěknicová, Jana (referee)
An increasing infertility due to pathological changes on sperm has become a serious issue. A rising concentration of fluorides in the environment could be a potentially negative factor affecting the male fertility. Eco-toxicological effect of fluorides can be enhanced in the present of aluminium ions by forming fluoro-metallic complexes, analogues of phosphate groups that interfere with the activity of G-proteins and P-type ATPases, which are part of several signalling pathways during sperm maturation. The aim of this thesis was monitoring the effect of fluorides on mouse sperm capacitation in vitro, analysis of the effect of fluorides on male reproductive organ and on the level of mouse spermatogenesis. Fluorides were administered to mice through table water at the concentrations of 0, 1, 10, 100 ppm and at the same time we monitored their synergic action with aluminium at 10 ppm dose. The level of tyrosine phosphorylation in the sperm head was the main marker for successful capacitation in vitro. The results show that an oversized load of ions, to which mice were exposed, affected negatively spermatogenesis and ability of sperm to capacitate in vitro. Besides that, a multigenerational study in mice demonstrated that even low concentrations of aluminium fluorides can have a negative effect on...
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Role paternálního H4K12ac při utváření pronukleí a v časné embryogenezi u myší.
Dudková, Barbora ; Hortová, Kateřina (advisor) ; Stopka, Tomáš (referee)
During the process of spermatogenesis, histones are replaced by protamines, basic proteins enabling transmission of DNA to the oocyte during fertilization. In mouse sperm, there is only 1% of remaining histones whose N-terminal tails contain post-translationally modified residues. In this study, I was interested in contribution of paternal histone H4 acetylated on lysine K12 residues (H4K12ac) that is present in mature sperm head in remaining nucleosomes. Physiologically, H4K12ac has an important role in transcription factor accumulation and in regulation of gene expression. The presence and abundance of H4K12ac modification in various pronuclei stages of 1-cell embryo and parthenotes were assessed by imunnoflourescent detection with utilization of anti-H4K12ac antibody. Altogether, the paternal pronucleus exhibits a strong acetylation signal on H4K12 since its formation, while in the maternal one, there is a slow continual increase of H4K12ac getting on the same level as paternal pronucleus till the pronuclei fusion. Simultaneously DNA methylation status in both pronuclei was detected. In paternal pronucleus there is a continual decrease in the DNA methylation detectable as a decrease of 5mC and an increase of 5hmC signal. Meanwhile, the maternal pronucleus stays widely methylated. DNA...
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Dynamics of acrosome reaction during intra-specific sperm competition in rodents.
Veselá, Kateřina ; Hortová, Kateřina (advisor) ; Pěknicová, Jana (referee)
Dynamics of acrosome reaction during intra-specific sperm competition in rodents Sperm acrosome integrity is disturbed in promiscuous species field mice (Apodemus) and more than half of the spermatozoa undergoing spontaneous acrosome reaction (AR) before binding to the zona pellucida. In Muridae it is documented a generally high rate of spontaneous AR, and the percentage increases in promiscuous species up to 60 % in 60 min capacitation in vitro. The acrosome integrity positively corellates with presence of CD46 protein which absence in wood mouse is fenotypicaly same as in CD46 knock-out mouse leading to accelerated spontaneous AR. It is necessary to clarify whether for mouse sperm it is essential the primary binding of intact sperm to zona pellucida of the egg or whether it is preferred secondary sperm binding after spontaneous AR. In this context, the question is whether there is a relocalization of the key fusion protein IZUMO in sperm during spontaneous AR. IZUMO relocalization was monitored by immunofluorescence at specific times of capacitation in vitro during spontaneous and induced AR. IZUMO relocalization as closely connected to actin cytoskeleton, and β1 integrins. Dynamics and localization of β1 integrin during spontaneous and induced AR was also detected by immunofluorescence. Our results...
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Effect of cryopreservation on mouse sperm.
Veselá, Kateřina ; Hortová, Kateřina (advisor) ; Pěknicová, Jana (referee)
Cryopreservation or freezing of sperm in the reproductive biology is still actual topic. Today is the only method used for sperm storage, whether for the purposes of assisted reproduction, or for scientific purposes. However, this method has a negative impact on such stored cells and is therefore still a subject of many studies. Among the main causes of sperm damage in rodents there are inappropriately selected cryoprotective agents, poor or no elimination of oxidative stress generated during cryopreservation, as well as a poorly chosen speed and temperature of freezing. Correctly chosen procedures and the composition of the media in which sperm are stored, can significantly affect the quality of sperm. This bachelor theses focuses on the influence of cryopreservation on mouse spermatozoa. The main impact of this method includes the effect of freezing on sperm DNA, plasma membrane, acrosome, and sperm motility.
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