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The characterisation of sperm and seminal plasma glycoproteins: Their role within the tetraspanin network and male fertility
Páleníková, Veronika ; Postlerová, Pavla (advisor) ; Ryšlavá, Helena (referee) ; Pěknicová, Jana (referee)
Mgr. Veronika Páleníková only to describe the localization and heterodimerization of α3, α6, αV, β1, and β4 integrins in mentioned integrins. Importantly, we detected the presence of integrin β4 subunit in mouse sperm, and characterised that it forms functional heterodimer with α6 subunit. In tion, we described α6β1 and α3β1integrin pairs, which contribute to the integrity and complexity of sperm membranes. Moreover, we found that α6 integrin subunit interacts with proposed CD9 and CD81 as tetraspanin partners for αV integrin in tetraspanin web. Our goal
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Glycoprotein profile of human ejaculate as a marker of its quality
Pavlová, Hana ; Postlerová, Pavla (advisor) ; Krejčová, Tereza (referee)
Glycoproteins are an indispensable component of the sperm glycocalyx. They arise through the process of co-translational or post-translational modification, so-called glycosylation, during which carbohydrate chains are attached to a peptide chain. Glycoproteins are involved in reproductive processes and their presence is often key to successful fertilization. The degree of glycosylation of glycoproteins is highly variable and is determined by the representation of carbohydrate units in glycans. It has already been found that the degree of glycosylation can be related to the fertility of an individual. The diploma thesis is focused on the comparison of the representation of terminal saccharides, sialic acid and fucose, in the glycoproteins of sperm and seminal plasma of patients with normal and abnormal ejaculate parameters. The total rate of sialylation and fucosylation in the seminal plasma was determined, as well as the rate of sialylation and fucosylation of specifically detected glycoproteins of sperm and seminal plasma. A significant difference was noted only in the detection of total fucosylation of seminal plasma glycoproteins. However, subsequent correlation tests revealed important relationship between the degree of sialylation and fucosylation of glycoproteins and ejaculate parameters,...
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Evaluation of antisperm antibodies (ASA) in diagnostics of idiopathic infertility
Pastyříková, Soňa ; Postlerová, Pavla (advisor) ; Novák, Jan (referee)
Infertility is a problem affecting almost 15% of couples. It can be caused by genetic, hormonal, anatomical or infectious problems, and it can also be caused by an immunological factor. One of the immunological causes of infertility is the presence of anti-sperm antibodies (ASA), which bind to sperm and adversely affect their function. The occurrence of ASA is manifested, for example, by reduced sperm motility, agglutination, and impaired sperm capacitation. Alternatively, ASA reduces the sperm ability to pass through the female urogenital tract, negatively affecting the sperm-oocyte interaction and the development of the embryo. The aim of this bachelor thesis was to compare existing methods for ASA determination. The main parameters were the complexity of the method and the assessment of results according to clinical studies focused on the detection and evaluation of ASA. The exact comparison of ASA detection methods based on the performed studies proved to be complicated, mainly due to the development of individual methods over time, different approaches to the evaluation of results, and low standardization of the methods. However, the most commonly recommended methods are MAR, ELISA and IBT tests, which are standardized and, due to commercially available kits, are considered to be easy to...
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Molecular basis of the sperm selection in the mammalian female reproductive tract
Šrámková, Adéla ; Krylov, Vladimír (advisor) ; Postlerová, Pavla (referee)
Internal fertilization is typical for all mammals. The male transfers sperm cells into the female reproductive tract, where usually one of them reach the oocyte and fertilize it. The female reproductive tract must be adapted to provide a suitable environment for the sperm cells to survive until fertilization. On the other hand, it must also be able to select developmentally competent sperm cells and prevent others to get into the close contact with the oocyte. Physical and molecular mutual communication of sperm cells with the female reproductive tract is ensured by sperm surface molecules and the receptors of the female epithelia lining. It appears that these interactions are required for key transformations, both on the sperm surface and in the cells of the female reproductive tract, where gene expression is affected. As a result of these changes, the most suitable sperm cell with all the required characteristics is allowed to fertilize the matured oocyte.
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Isolation and study of proteins with zona pellucida binding activity
Zigo, Michal ; Postlerová, Pavla (advisor) ; Ryšlavá, Helena (referee) ; Antalíková, Jana (referee)
Binding of sperm to the oocyte is mediated by complementary molecules on the surface of both gametes and involves the interaction of sperm protein receptors with the zona pellucida (ZP) saccharide structures. It has been shown that many proteins receptors are involved in the sperm-ZP interaction, and potential primary sperm receptors for ZP glycoproteins have been investigated in various mammals. The majority of proteins with identified sperm-ZP binding activity belong to the plasma membrane proteins. However, the exact methods for isolation of sperm membrane proteins are still to be standardized. This study is focused on investigating how employment of various isolation protocols leads to acquisition of various protein mixtures. Further in the work, two possible approaches towards identification of potential ZP-binding partners are implemented, in order to disclose novel primary ZP-binding receptor candidates. Sperm proteins of ejaculated and in vitro capacitated boar sperms were isolated by: Triton X-100, Triton X-114, acetic acid, sodium dodecyl sulphate (SDS), N-octyl-β-D- glucopyranoside (OBG), rehydration buffer (RHB), and finally by freezing-thawing extraction and they were characterized by 1-D, 2-D protein profiles, glycoprotein staining and substrate zymographic methods. The results have...
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Study of sperm proteins in different mammalian species
Pohlová, Alžběta ; Postlerová, Pavla (advisor) ; Ryšlavá, Helena (referee)
Reproduction is an essential feature of all animals and a fundamental step to produce new generations. Study of sperm proteins is crucial for understanding of the sperm-egg recognition. We searched out sperm surface proteins involving in the zona pellucida (ZP) binding and studied whether these proteins are preserved throughout mammalian species. Indirect immunofluorescent technique was used to test a panel of monoclonal antibodies prepared against boar sperm surface proteins on spermatozoa of bull and mice. We found a cross-reactivity of some antibodies against boar sperm with bull ejaculated and mouse epididymal spermatozoa. Further, we isolated sperm proteins from different mammalian species, such as pig, bull, dog, cat, mouse and human. Proteins were separated by SDS- electrophoresis and protein/glycoprotein profiles from epididymal, ejaculated and in vitro capacitated sperm were compared. The interaction of sperm with ZP was studied on electrophoretically-separated sperm surface proteins from pig and bull with biotin-labeled ZP glycoproteins. Antibodies, which reacted with boar sperm surface proteins with ZP- binding activity, therefore could be potential egg-binding receptors, were used for monitoring of the sperm protein origin in reproductive fluids and tissues. (In Czech) Keywords: sperm...
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Use of transcriptomics to study mechanism of the action of complex mixtures of organic compounds occurring in the ambient air focusing on polycyclic aromatic hydrocarbons
Líbalová, Helena ; Topinka, Jan (advisor) ; Krásný, Libor (referee) ; Postlerová, Pavla (referee)
Polycyclic aromatic hydrocarbons (PAH) represent a large group of organic compounds occuring as pollutants in ambient air. Besides their genotoxic effect, some of them are known to be complete carcinogens and act via nongenotoxic and tumor promoting mechanism. Although effects of many individual compounds are well-documented, human exposure to polycyclic aromatic hydrocarbons in ambient air occurs through complex mixtures and only few studies describe the behavior of PAH in real complex mixtures. The first part of the thesis is dealing with the global gene expression changes in human embryonic lung fibroblasts (HEL) as a consequence of the effect of complex mixtures containing PAH extracted from the respirable airborne particles PM2.5. These particles were collected in 4 localities in the Czech republic (Ostrava - Bartovice, Ostrava - Poruba, Karviná, Třeboň) differing in the level of the air pollution. Gene expression changes induced by three subtoxic concentrations of organic extracts (EOM - extractable organic matter) from each locality after 24 hour incubation were examined by microarray analysis. Pathway analysis using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database was applied to interpret gene expression data. In each locality we identified several deregulated signaling pathways...
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Secreted proteins by male reproductive tract
Cozlová, Nina ; Postlerová, Pavla (advisor) ; Železná, Blanka (referee) ; Antalíková, Jana (referee)
1 AbstractAbstractAbstractAbstract Proteins secreted in the male reproductive tract play a key role in post-testicular development of sperm and in further steps needed for fertilization. Sperm maturation represents a key step in the reproduction process. Sperm, during the passage through the epididymis undergoes significant changes due to proteolytic and glycolytic activities in the epididymal fluid. Inhibitor of acrosin protects spermatozoa and reproductive epithelium against proteolytic degradation and also protects binding sites for ZP on sperm plasma membrane. In boar reproductive system acrosin inhibitor (AI) was found in seminal plasma and on sperm plasma membrane. Polyclonal antibody recognized AI in extracts of the cauda epididymidis, seminal vesicles, prostate, and Cowper's glands. Using immunofluorescence method has revealed the AI in the epithelium and lumen of these organs but also on the surface of epididymal and ejaculated spermatozoa. We registered the increasing signal of AI from caput to cauda epididymis. Gene expression of AI mRNA was detected in the epididymis, seminal vesicles, prostate, and Cowper's glands and increased gradually throughout the epididymal duct. In present study, we also monitored AI in boar epididymal fluid and spermatozoa along the organ. In the epididymis, AI may...
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