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Analýza faktorů působících na produkci a kvalitu ejakulátu plemenných kanců
Střechová, Vendula
The aim of this study was an evaluation of quantitative and qualitative characteristics of boar semen and morphology of spermatozoa in relation to selected outer and inner factors. Data from 391 boars of five different breeds from one insemination station in the Czech Republic were collected between 2015 and 2019. Data from a total of 17.810 collected ejaculates were used for evaluation. The analysed factors included breed, birth season of the boar, age of the boar, season of semen collection and frequency of semen collection. In terms of total sperm count, the boars of large white showed below average. A significantly higher concentration of spermatozoa with morphological abnormalities was observed in the semen of the duroc boars. The results documented, that boars born in summer and partially also boars born in winter achieve better values in comparison with boars from spring and autumn litters. While the semen volume and total sperm count increased with the age, with motility of sperm cells it was contrarywise. Semen collected in winter was characteristic with the highest values of volume and total sperm count. On the contrary, the lowest volume and total sperm count was found for the semen collected in summer, and this season was also characterized by the highest frequency of morphologically abnormal spermatozoa. Prolongation of sexual resting led to an increase in sperm concentration and total sperm count in semen. The most significant increase was recorded when the frequency of semen collection prolonged from 4–5 days to 6–7 days. With prolonged sexual resting, the occurrence of spermatozoa with the distal cytoplasmic droplet decreased very significantly.
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Sperm morphology and molecular mechanism associated with sperm fertilizing ability in mice.
Šebková, Nataša ; Komrsková, Kateřina (advisor) ; Jonáková, Věra (referee) ; Linhart, Otomar (referee)
The process of sperm capacitation and acrosome reaction (AR) are highly dynamic processes essential for the fertilization, including cytoskeleton proteins in the sperm head. The study of the distribution of actin, spectrin and α-tubulin in the head of representative mammalian sperm indicates that before and after the AR there were changes in the distribution of the cytoskeleton structures in the sperm head of all representatives, mainly in the apical part of the acrosome, but also in the equatorial and postacrosome part of the sperm head. The particularly well-studied actin cytoskeleton plays an important role during the capacitation and before and after the AR. Environmental estrogens can interfere with the function of endogenous hormones in very low concentrations. They can interact with specific receptors affecting several signalling pathways leading to sperm capacitation and AR. The effect of 17β-estradiol, estrone, estriol and synthetic 17α-ethynylestradiol was evaluated by the ratio of tyrosine phosphorylation and the state of the acrosome during in vitro capacitation in the sperm head. This study has provided the evidence that estrogens significantly stimulate capacitation progress in a concentration-dependent manner. Estrogens decrease number of sperm after the induced AR too. The raising...
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In vitro fertilization in horses
Müller, Filip ; Šušor, Andrej (advisor) ; Šimoník, Ondřej (referee)
Reproductive management in horse breeding increasingly relies on in vitro fertilization (IVF) techniques. This technology entails a complex process, including the acquisition of immature oocytes, their in vitro maturation, fertilization with sperm, and subsequent cultivation of the early embryo before transfer into a synchronized recipient. However, the success of equine IVF is influenced by various factors such as oocyte quality, sperm capacitation, and technical issues unique to this process, compared to other mammalian models. This bachelor's thesis aims to summarize and evaluate current knowledge on IVF of horse eggs, identify key factors influencing success, and describe current procedures for gamete preparation. By incorporating the latest IVF approaches and optimizing them, a higher success rate can be achieved. Key aspects for improving equine IVF include gamete collection and handling, in vitro oocyte maturation, sperm capacitation, and subsequent embryonic development.
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Sperm DNA damage: detection methods used in clinical practice
Tušlová, Magdaléna ; Postlerová, Pavla (advisor) ; Koubek, Pavel (referee)
The current routine semen analysis, known as a spermiogram, is not able to provide a complete picture of male fertility as it does not routinely include an assessment of DNA integrity. This parameter is closely related to difficulties during conception and increased risk of miscarriage. This paper discusses the causes of sperm DNA damage, the detection of this damage by various methods and concludes with a discussion of the possibilities of incorporating these tests into clinical practice to improve the assessment and treatment of male fertility. Mechanisms of sperm DNA fragmentation include impaired spermatogenesis, oxidative stress and impaired apoptosis, which can be influenced by genetic, anatomical and external factors. The main part of this thesis was devoted to a review of methods for the detection of sperm DNA fragmentation, including sperm chromatin structure analysis (SCSA), sperm chromatin dispersion test (SCD), TUNEL method, single-strand DNA break recognition test (ISNT), comet assay and chromatin staining. Although some of these methods are frequently used, challenges remain in their standardization and full integration into clinical practice. However, the possibility of their routine use could bring significant advances in the diagnosis and treatment of male infertility. Among the...
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The effect of oxidative stress on the structure and function of sperm cell cytoplasmic membrane during the fertilization process.
Tojnerová, Kateřina ; Krylov, Vladimír (advisor) ; Šanovec, Ondřej (referee)
Oxidative stress is caused by an imbalance between antioxidants and oxidants, and thus a consequent excess of reactive oxygen species (ROS). It can also be caused by insufficient antioxidant protection, which normally reduces ROS levels and protects the cell from damage. Oxidative stress causes changes in the structure and function of the sperm cytoplasmic membrane during the fertilization process. Here it has a positive effect on physiological functions, where low levels of ROS are necessary for normal sperm development such as sperm capacitation or sperm hyperactivation. However, at higher levels ROS exerts a pathophysiological effect, reducing membrane fluidity due to membrane lipid peroxidation, DNA fragmentation, loss of motility and subsequent inability of the sperm to fertilize can also occur. Keywords: oxidative stress, cytoplasmic membrane, sperm cell, fertilization
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Detection of positive selection in reproductive genes of songbirds
Cakl, Lukáš ; Reifová, Radka (advisor) ; Těšický, Martin (referee)
Reproductive genes are assumed to play an important role in adaptive evolution and speciation, yet little is known about the patterns of molecular evolution in these genes within avian species. This thesis is focused on identifying reproductive genes under positive selection and analyzing their function in songbirds, the largest and most diverse suborder of modern birds. Using existing bioinformatic approaches and published genomic data of 14 songbird species, we have first constructed 12000 groups of orthologous genes and detected significant traces of recurrent positive selection within 385 of them. Using proteomic data, this genome wide set of genes was then filtered to obtain genes expressed within songbird spermatozoa and fluids from the female reproductive tract. Within spermatozoa 22 out of 940 expressed genes were positively selected, while female fluids were found to be more conserved, as only 6 out of 529 expressed genes have shown traces of positive selection. We have then computed the enrichment of gene ontology terms within the positively selected genes. The enriched terms suggest evolutionary pres- sures acting on spermatozoa cytoskeleton, molecular motors and energetic metabolism, highlighting the importance of sperm morphology and swimming speed. Furthermore, the enrichment results...
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Význam vitaminu D a jeho metabolitů v reprodukci živočišných organismů
Červenka, Jiří
Vitamin D is considered to be a group of molecules that have many functions in animal organisms. The best known of these is the key role in calcium and phosphorus homeostasis and regulation of bone mineralization. In recent years, however, many new functions of this vitamin have been discovered, such as the regulation of cell proliferation and differentiation, its influence on the immune system or its function in animal reproduction. This thesis deals with the importance of vitamin D in animal reproduction with an emphasis on livestock. In the literature review, the occurrence of receptors for vitamin D in cells of various types of different tissues of reproductive system, particularly in mammals and birds, and the function of various metabolites mediated by these receptors are discussed in the individual chapters. The thesis includes the design of methodology, which would test the effect of vitamin D on reproduction of the male and female domestic chickens. The hypothesis could verify, whether or not, Rhode Island Red cockerels, that would be fed a diet with a higher vitamin D content would demonstrate better ejaculate parameters and higher fertilization of eggs laid by Rhoade Island White hens inseminated with this semen. Furthemore, a solution is proposed to verify the hypothesis, assuming higher hatchability and lower embryonic mortality in hens fed higher vitamin D feed mixture. The result would be compared with hens and roosters of the same breeds, reared in an environment with the same conditions, fed a vitamin D deficient feed mixture.
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Rab proteins in sperm life
Vojtová, Kristýna ; Páleníková, Veronika (advisor) ; Zelenková, Natálie (referee)
Rab proteins are the largest group of small GTPases. They work as key regulators in the intracellular vesicular transport of cells. This bachelor's thesis contains a literature review of current information on the function of Rab proteins in mammalian spermatozoa. The first part of the work is devoted to their structure, function, cycle, and diseases related to the dysfunction of Rab proteins. The second part is focused on sperm development and maturation linked with specific Rab proteins involved in individual steps during the life of the sperm. Rab proteins are important in cytoskeletal organization and cytokinesis of sperm during spermatogenesis. They participate in the acrosome biogenesis mediated by the Golgi apparatus and the morphological changes of sperm during spermiogenesis. They also play a key role in the acrosome reaction and interaction with the zona pellucida. Due to their necessity in the sperm development and maturation, Rab proteins are proposed as a marker of male fertility, which is summarized in the last part of the thesis. Keywords Rab proteins, GTPases, sperm, spermatogenesis, spermiogenesis, fertilization, infertility
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Proteomic architecture of sperm-egg interactions
Otčenášková, Tereza ; Stopka, Pavel (advisor) ; Petr, Jaroslav (referee) ; Vrbacký, Marek (referee)
Recent advances in proteomic methods provide new insights for biological research including the field of reproductive biology. Determination of the proteomic basis of spermatozoa is pivotal for understanding the complex process of gamete interactions during fertilization such as acrosome reaction. Great differences imposed by postcopulatory sexual selection and phylogeny can be observed regarding the size, shape, and molecular composition of sperm across animal taxa. The first objective of this doctoral thesis is to characterize the protein contents of the acrosome to ascertain its further functional significance in sperm-egg interaction. Also, we aim to investigate the potential relationships between sperm protein composition and sperm morphology diversification, risk of sperm competition, and species phylogenetic background. Wild-caught males from natural populations of species of Mus musculus musculus, Apodemus flavicollis, Microtus arvalis (order Rodentia), Acrocephalus palustris, Chloris chloris, Phylloscopus collybita, Cinclus cinclus, Hirundo rustica, and Taeniopygia guttata from a captive population (order Passeriformes) were subject to the analyses. Nano-liquid chromatography with tandem mass spectrometry was applied as the main methodological approach in this thesis. Our data implicate...
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Ezrin-Radixin-Moesin (ERM) proteins: The link between tetraspanin web and cytoskeleton in gametes
Picková, Jana ; Frolíková, Michaela (advisor) ; Lánská, Eva (referee)
Ezrin, which belongs to the ERM protein family along with radixin and moesin, plays an important role in linking membrane proteins to the actin cytoskeleton. In sperm, ezrin was described so far in bovine and humans, where it probably contributes in actin polymerization during capacitation. During the acrosome reaction, actin plays a significant role in relocation of protein from the inner acrosomal membrane to the equatorial segment, which is an essential process for successful fertilization. One of the proteins that relocates to the equatorial segment, the site of initiation of gamete fusion, is protein CD9. This protein probably stabilizes protein interactions between sperm and oocyte during gamete fusion. In this diploma thesis, we focused on the role of ezrin in linking the CD9 protein to the actin cytoskeleton in mouse sperm. Using indirect immunofluorescence, we detected ezrin, CD9 and actin in acrosome intact mouse sperm. The co-localization of these proteins in the apical acrosomal region suggests their interactions. However, this was not confirmed by our co-immunoprecipitation experiments. Nevertheless, in the apical acrosomal region we also detected EWI-2 protein, which could act as linker between CD9 and ezrin. In this thesis, we also investigated changes in levels of active...
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