Book of abstract of XXIIIrd Symposium of immunology and biology of reproduction
Kubátová, Alena
2017 - anglický This Symposium was mainly focused on diabetes melllitus type 1 (mouse and human model), gene expression during spermatogenesis and spermiogenesis,\nrole of tetraspanins family and other proteins in sperm-egg interaction, role of estrogen receptors (sperm cells) and bisphenol S (oocytes). Klíčová slova: reproduction; sperm-egg interaction; tetraspanins; estrogen receptors; CD molecules; diabetes mellitus; oocytes; sperm surface molecules; bisphenol S; gene expression Plné texty jsou dostupné na jednotlivých ústavech Akademie věd ČR. Book of abstract of XXIIIrd Symposium of immunology and biology of reproduction

This Symposium was mainly focused on diabetes melllitus type 1 (mouse and human model), gene expression during spermatogenesis and spermiogenesis,\nrole of tetraspanins family and other proteins in ...

Kubátová, Alena
Biotechnologický ústav, 2017

Book of abstract of XXIInd Symposium of immunology and biology of reproduction
Pěknicová, Jana; Elzeinová, Fatima; Kubátová, Alena
2016 - anglický Book of abstracts. The Symposium was focused on immunology of reproduction and specific problems in reproduction (mainly in human infertility). Klíčová slova: spermatology; biochemistry; cell interaction; monoclonal antibody; sterility/fertility; gene expression Plné texty jsou dostupné na jednotlivých ústavech Akademie věd ČR. Book of abstract of XXIInd Symposium of immunology and biology of reproduction

Book of abstracts. The Symposium was focused on immunology of reproduction and specific problems in reproduction (mainly in human infertility).

Pěknicová, Jana; Elzeinová, Fatima; Kubátová, Alena
Biotechnologický ústav, 2016

Recombinant proteins and their usage in the studies of male gamete function
Dorosh, Andriy; Žatecká, Eva; Pěknicová, Jana; Dvořáková-Hortová, Kateřina
2016 - anglický Klíčová slova: recombinant protein; spermatogenesis; glycolytic enzyme Plné texty jsou dostupné na jednotlivých ústavech Akademie věd ČR. Recombinant proteins and their usage in the studies of male gamete function

Dorosh, Andriy; Žatecká, Eva; Pěknicová, Jana; Dvořáková-Hortová, Kateřina
Biotechnologický ústav, 2016

Estrogen receptor beta (ERβ) in testicular cells and sperm
Dostálová, Pavla; Žatecká, Eva; Děd, Lukáš; Dorosh, Andriy; Postlerová, Pavla; Jonáková, Věra; Dvořáková-Hortová, Kateřina; Pěknicová, Jana
2016 - anglický Estrogen is a steroid hormone that plays an important role during sperm development in the male and female reproductive tract. Estrogen signalling is a complex process that depends on cell milieu and presence of receptors. Thanks to the steroid nature of estrogens, they can pass through the plasmatic membrane and bind to the intracellular estrogen receptors (ERs). Within the cell, there are several pools of ERs. One of them is localized to the cell nucleus and their activation leads to direct or indirect binding to DNA and ultimately to alternation in gene expression (genomic pathway). Other pools of ERs are associated with plasma membrane or are located in cytosol. Activation of membrane associated ERs leads to rapid non-genomic responses. Nowadays, two classical estrogen receptors are known – ERα and ERβ. Since ERβ is a predominant variant in testes, we focused our study on expression of ERβ variants in murine testes and sperm. We detected two variants of ERβ at mRNA level in both, testes and sperm. These variants differ in 54 nucleotids within the ligand binding domain and this variability results in different affinity to estrogen. We analyzed individual testicular cell types (spermatogonia, spermatocytes, spermatids, Sertoli cells) by RT-qPCR. Our results suggest that both ERβ variants are coexpressed in the same cell type and may therefore interact together. This may have consequences in mediating of estrogen signalling. Moreover, ERβ is expressed more in the later stages of spermatogenesis suggesting the role of ERβ in these stages or alternatively in spermatozoa alone. At the protein level, we detected ERβ in nuclear, membrane and cytosolic fraction prepared from testicular tissue suggesting the involvement of both, genomic and non-genomic, pathways of estrogen signaling in testes. In sperm, anti-ERβ antibodies localized ERβ in acrosome region and tail which is in accordance with the known role of estrogen on capacitation, acrosome reaction and motility. Klíčová slova: estrogen receptor; gene expression; testicular cell types; RT-qPCR; monoclonal antibodies Plné texty jsou dostupné na jednotlivých ústavech Akademie věd ČR. Estrogen receptor beta (ERβ) in testicular cells and sperm

Estrogen is a steroid hormone that plays an important role during sperm development in the male and female reproductive tract. Estrogen signalling is a complex process that depends on cell milieu and ...

Dostálová, Pavla; Žatecká, Eva; Děd, Lukáš; Dorosh, Andriy; Postlerová, Pavla; Jonáková, Věra; Dvořáková-Hortová, Kateřina; Pěknicová, Jana
Biotechnologický ústav, 2016

Epididymal maturation – a crucial step in the post-testicular sperm development
Postlerová, Pavla; Pohlová, Alžběta; Zigo, Michal; Jonáková, Věra
2016 - anglický Mammalian spermatozoa after their development in testis undergo the post-testicular maturation in epididymis where acquire their fertilization ability and competence of movement. The epididymis is tissue with very active fluid-absorbing and fluid-secreting activity. Epididymal fluid contains ions and small molecules, proteins, glycoproteins and enzymes. The surface of spermatozoa is exposed directly to the epididymal fluid, and the sperm plasma membrane is significantly changed. Some testicular proteins are altered, masked, or replaced by new proteins/glycoproteins of epididymal origin. Several proteins produced by epididymis have been described in various mammalian species and shown to be associated with spermatozoa suggesting a role in the sperm maturation and/or sperm-egg binding and fusion. We isolated proteins from fluid, tissue and sperm of boar epididymis, and separated them by chromatographic and electrophoretic methods. We searched for known proteins using panel of antibodies and tested proteins of epididymal fluid for binding abilities. In the epididymis, we found proteins described as proteins of seminal plasma and associated with the sperm surface, such as spermadhesins, beta-microseminoprotein and acrosin inhibitor. These proteins were detected in epididymal sperm, fluid and tissue. We showed that some epididymal proteins may bind the spermatozoa and change the binding sites on the sperm surface. We determined and identified some proteins from boar epididymal fluid with affinity to heparin, hyaluronan and zona pellucida glycoproteins. These phenomena indicate that epididymal fluid proteins bind to the sperm surface during epididymal maturation and might subsequently play role in the sperm capacitation or sperm-zona pellucida binding. Klíčová slova: sperm maturation; glycoproteins; spermadhesins; epididymal fluid; monoclonal antibodies Plné texty jsou dostupné na jednotlivých ústavech Akademie věd ČR. Epididymal maturation – a crucial step in the post-testicular sperm development

Mammalian spermatozoa after their development in testis undergo the post-testicular maturation in epididymis where acquire their fertilization ability and competence of movement. The epididymis is ...

Postlerová, Pavla; Pohlová, Alžběta; Zigo, Michal; Jonáková, Věra
Biotechnologický ústav, 2016

Super-resolution Stimulated Emission Depletion (STED) microscopy imaging of selected integrins in mouse acrosome intact sperm
Frolíková, Michaela; Šebková, Nataša; Děd, Lukáš; Dvořáková-Hortová, Kateřina
2016 - anglický Klíčová slova: actin; integrins family; STED; protein relocalization; acrosome membrane Plné texty jsou dostupné na jednotlivých ústavech Akademie věd ČR. Super-resolution Stimulated Emission Depletion (STED) microscopy imaging of selected integrins in mouse acrosome intact sperm

Frolíková, Michaela; Šebková, Nataša; Děd, Lukáš; Dvořáková-Hortová, Kateřina
Biotechnologický ústav, 2016

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
2016 - anglický 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. Klíčová slova: single cell expresion analysis; RT-qPCR; oocytes; Laser capture microdissection; in situ hybridization Plné texty jsou dostupné na jednotlivých ústavech Akademie věd ČR. Single cell expression analysis of genes with potential mrna gradient in mouse oocytes

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 ...

Dorosh, Andriy; Margaryan, Hasmik; Vodička, Martin; Ergang, Peter; Šídová, Monika; Dvořáková-Hortová, Kateřina
Biotechnologický ústav, 2016

Distribution of CD46 and β1 integrin molecules with respect to different membrane structures of the sperm head
Šebková, Nataša; Frolíková, Michaela; Děd, Lukáš; Dvořáková-Hortová, Kateřina
2016 - anglický CD46 protein plays an important role during fertilization and its role is associated with acrosome stability. CD46 is probably involved in signalling pathways triggering the acrosome reaction (AR). It also associates through membrane integrins with specific MAP kinases involved in the AR. Our aim was to monitor the dynamics of relocation of CD46 and β1 integrin during sperm maturation and its preparation for the fertilization. The dependence of this localization changes on the dynamic of actin cytoskeleton was studied. Our results show the changes in the localization of these proteins associated with the AR and their co-localization was observed using proximity ligation assay. After the AR CD46 and β1 integrin spreads across the sperm head, entering the post-acrosomal compartment, and permeates the borders of different domains. It was shown previously that actin dynamics is necessary for acrosome reaction-associated translocation of Izumo1 protein that is required for sperm-egg fusion. Therefore Latrunculin A was used during sperm incubation. The co-incubation of capacitated sperm with Latrunculin A leads to a decrease of the percentages of sperm, which express relocation pattern after induced AR. 3D models and visualizations of potential membrane processes responsible for the relocation of proteins from the acrosomal area to the other compartments of the sperm head were prepared. Our results deliver new information that proteins CD46 and β1 integrin undergo dynamic relocation towards the sites of sperm-egg fusion during the AR in vitro. The inhibitor of actin dynamics abrogates significantly the AR-associated changes in proteins localization. We speculate that this relocation is of importance for the successful sperm-egg interaction, adhesion and subsequent gamete fusion. Klíčová slova: CD46; β1 integrin; acrosome reaction; latrunculin A; sperm-egg fussion; Proximity ligation assay Plné texty jsou dostupné na jednotlivých ústavech Akademie věd ČR. Distribution of CD46 and β1 integrin molecules with respect to different membrane structures of the sperm head

CD46 protein plays an important role during fertilization and its role is associated with acrosome stability. CD46 is probably involved in signalling pathways triggering the acrosome reaction (AR). It ...

Šebková, Nataša; Frolíková, Michaela; Děd, Lukáš; Dvořáková-Hortová, Kateřina
Biotechnologický ústav, 2016

Visualisation of CD46 and β1 integrins in mouse sperm head by super-resolution Stimulated Emission Depletion (STED) microscopy
Frolíková, Michaela; Šebková, Nataša; Děd, Lukáš; Dvořáková-Hortová, Kateřina
2015 - anglický Klíčová slova: CD46 monoclonal antibody; β1 integrins; STED; Proximity ligation assay Plné texty jsou dostupné na jednotlivých ústavech Akademie věd ČR. Visualisation of CD46 and β1 integrins in mouse sperm head by super-resolution Stimulated Emission Depletion (STED) microscopy

Frolíková, Michaela; Šebková, Nataša; Děd, Lukáš; Dvořáková-Hortová, Kateřina
Biotechnologický ústav, 2015

CD46 and β1integrin interaction in mouse sperm head
Šebková, Nataša; Frolíková, Michaela; Dvořáková-Hortová, Kateřina
2015 - anglický CD46 protein plays an important role during fertilization and its role is associated with acrosome stability. CD46 is probably involved in signalling pathways triggering the acrosome reaction. Integrins interact with many cytoskeletal proteins such as actin, therefore changes in the actin cytoskeleton before and after AR may lead to changes in the association and localization of CD46 and β1integrin. Our aim was to monitor mutual CD46 and β1integrin interaction detected by the proximity ligation assay. It generates a localized signal in a form of spots revealing the exact position of the recognition event. Proteins interaction was study in freshly released sperm and sperm during the acrosome reaction, during which there is a gradual relocation of these proteins towards the equatorial segment and the whole sperm head. Proteins α and β tubulin were used as a positive control, α tubulin and β1 integrin as a negative control. In situ PLA showed a distinct spotted signal indicating the mutual interaction of CD46 and β1integrin. A positive response was demonstrated not only in freshly released sperm but also in sperm during the acrosome reaction. Freshly released sperm were distinctively labelled in the acrosome region and the neck, similarly to the positive control. Sperm during the acrosome reaction showed the signal across the whole sperm head region. No signal or sporadic nonspecific staining was detected in the case of the negative control. In summary, our results deliver new information that proteins CD46 and β1 integrin interact with each other. These results suppose the theory that β1 integrin can mediate a connection between CD46 and sperm cytoskeleton thereby molecules of signalling pathways leading to activation of the acrosome reaction. Klíčová slova: CD46; β1 integrin; acrosome reaction Plné texty jsou dostupné na jednotlivých ústavech Akademie věd ČR. CD46 and β1integrin interaction in mouse sperm head

CD46 protein plays an important role during fertilization and its role is associated with acrosome stability. CD46 is probably involved in signalling pathways triggering the acrosome reaction. ...

Šebková, Nataša; Frolíková, Michaela; Dvořáková-Hortová, Kateřina
Biotechnologický ústav, 2015