The degradation of maternally provided molecules is a very important process during early embryogenesis. However, the vast majority of studies deals with mRNA degradation and protein degradation is only a very little explored process yet. The aim of this article was to summarize current knowledge about the protein degradation during embryogenesis of mammals. In addition to resuming of known data concerning mammalian embryogenesis, we tried to fill the gaps in knowledge by comparison with facts known about protein degradation in early embryos of non-mammalian species. Maternal protein degradation seems to be driven by very strict rules in terms of specificity and timing. The degradation of some maternal proteins is certainly necessary for the normal course of embryonic genome activation (EGA) and several concrete proteins that need to be degraded before major EGA have been already found. Nevertheless, the most important period seems to take place even before preimplantation development-during oocyte maturation. The defects arisen during this period seems to be later irreparable.
- MeSH
- embryo nesavčí metabolismus fyziologie MeSH
- embryo savčí metabolismus fyziologie MeSH
- embryonální vývoj fyziologie MeSH
- genom fyziologie MeSH
- lidé MeSH
- oocyty metabolismus fyziologie MeSH
- proteiny metabolismus MeSH
- vývojová regulace genové exprese fyziologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
Tetraspanins are multifunctional molecules located in specific microdomains on the plasma membrane. Thanks to their ability to form networks with other proteins they can participate in many cellular functions. Tetraspanins are part of the interactive network in gametes; however, their precise role in fertilization is not yet clear. The aim of this study was to compare the localization of CD9 and CD81 tetraspanins during oocyte maturation and early development of the embryos in bovine and porcine model. CD9 was detected on the oocyte plasma membrane and vesicles in the perivitelline space of bovine oocytes and embryos. We suggest that CD9 could be a component involved in transzonal projections. Based on the results of in vitro fertilization assay, CD9 and CD81 seem to be part of a more complex fusion network on the plasma membrane of bovine oocytes. On the other hand, both tetraspanins showed a clustered expression pattern on the plasma membrane and inner margin of zona pellucida (ZP) in porcine oocytes and embryos. We found a new species-specific pattern of CD9 and CD81 distribution in ZP which could reflect their specialized role in processes associated with cell adhesion and intercellular communication upon fertilization.
- MeSH
- antigeny CD81 metabolismus MeSH
- antigeny CD9 metabolismus MeSH
- buněčné linie MeSH
- embryo savčí cytologie metabolismus MeSH
- fertilizace in vitro účinky léků MeSH
- metafáze účinky léků MeSH
- myši inbrední BALB C MeSH
- oocyty cytologie metabolismus MeSH
- partenogeneze účinky léků MeSH
- prasata MeSH
- protilátky farmakologie MeSH
- skot MeSH
- stadium rýhování vajíčka cytologie účinky léků MeSH
- zvířata MeSH
- Check Tag
- skot MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND/AIM: Spontaneous regression (SR) of tumours is a rare phenomenon not yet fully understood. The aim of this study was to investigate immune cells infiltrating progressive and SR tumours in a Lewis rat sarcoma model. MATERIALS AND METHODS: Rats were subcutaneously inoculated with rat sarcoma R5-28 (clone C4) cells. Developing tumours were obtained on day 42 and cryosections were immunohistochemically processed for detection of immune cells. RESULTS: A high density of granulocytes was found in the necrotic areas of both progressive and SR tumours. CD4+ cells and CD8+ cells were rare and sparsely dispersed in the tumour tissue without clear difference between the two types of tumours. On the contrary, CD161+ cells were abundant and evenly distributed in SR tumours, but these cells were very rare in progressive tumours. CONCLUSION: Based on the differences in number and distribution of the immune cell subpopulations, we believe that natural killer (CD161+) cells play a major role in the destruction of cancer cells during SR of tumours in this Lewis rat model.
- MeSH
- buňky NK patologie MeSH
- CD4-pozitivní T-lymfocyty patologie MeSH
- CD8-pozitivní T-lymfocyty patologie MeSH
- imunohistochemie MeSH
- krysa rodu rattus MeSH
- lektinové receptory NK-buněk - podrodina B genetika MeSH
- lidé MeSH
- modely nemocí na zvířatech MeSH
- potkani inbrední LEW MeSH
- regulace genové exprese u nádorů MeSH
- sarkom genetika patologie MeSH
- spontánní regrese nádoru genetika patologie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Porcine oocytes that have matured in in vitro conditions undergo the process of aging during prolonged cultivation, which is manifested by spontaneous parthenogenetic activation, lysis or fragmentation of aged oocytes. This study focused on the role of hydrogen sulfide (H2S) in the process of porcine oocyte aging. H2S is a gaseous signaling molecule and is produced endogenously by the enzymes cystathionine-β-synthase (CBS), cystathionine-γ-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (MPST). We demonstrated that H2S-producing enzymes are active in porcine oocytes and that a statistically significant decline in endogenous H2S production occurs during the first day of aging. Inhibition of these enzymes accelerates signs of aging in oocytes and significantly increases the ratio of fragmented oocytes. The presence of exogenous H2S from a donor (Na2S.9H2O) significantly suppressed the manifestations of aging, reversed the effects of inhibitors and resulted in the complete suppression of oocyte fragmentation. Cultivation of aging oocytes in the presence of H2S donor positively affected their subsequent embryonic development following parthenogenetic activation. Although no unambiguous effects of exogenous H2S on MPF and MAPK activities were detected and the intracellular mechanism underlying H2S activity remains unclear, our study clearly demonstrates the role of H2S in the regulation of porcine oocyte aging.
- MeSH
- cystathionin-beta-synthasa metabolismus MeSH
- cystathionin-gama-lyasa metabolismus MeSH
- embryo savčí účinky léků MeSH
- inhibitory enzymů farmakologie MeSH
- kultivace embrya MeSH
- kultivované buňky MeSH
- oocyty účinky léků fyziologie MeSH
- partenogeneze účinky léků MeSH
- prasata MeSH
- stárnutí buněk účinky léků MeSH
- sulfan metabolismus farmakologie MeSH
- sulfurtransferasy metabolismus MeSH
- zvířata MeSH
- Check Tag
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The processes of oocyte growth, acquisition of meiotic competence and meiotic maturation are regulated by a large number of molecules. One of them could be calcineurin consisting of catalytic subunit A (Aα, Aβ, Aγ isoforms) and regulatory subunit B (B1, B2 isoforms). Calcineurin is involved in the meiotic maturation of oocytes in invertebrates or in lower vertebrates. In the mammalian oocytes, the possible role of calcineurin in the regulation of oocyte meiosis has not been clarified to date. In this study, to investigate the role of calcineurin during porcine oocyte growth, acquisition of meiotic competence and meiotic maturation, we analysed the expression and localisation of calcineurin subunits and the mRNA expression of calcineurin isoforms. Calcineurin was expressed in growing porcine oocytes, in fully grown oocytes and during their in vitro meiotic maturation. We found both subunits of calcineurin. Calcineurin A and calcineurin B were localised mainly in the cortex in all porcine oocytes. The changes in the intracellular localisation of separate calcineurin subunits during meiotic maturation were determined. We detected mRNA for calcineurin isoforms Aβ, Aγ, B2 in oocytes and mRNA for calcineurin isoforms Aβ, Aγ, B1, and B2 in cumular cells. To our knowledge, this is the first confirmation of calcineurin presence in porcine oocytes.
- MeSH
- IVM techniky veterinární MeSH
- kalcineurin genetika metabolismus MeSH
- meióza fyziologie MeSH
- messenger RNA genetika metabolismus MeSH
- oocyty fyziologie MeSH
- prasata * MeSH
- regulace genové exprese fyziologie MeSH
- transport proteinů fyziologie MeSH
- zvířata MeSH
- Check Tag
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The nitric oxide donor (+)-S-nitroso-N-acetylpenicillamine (SNAP) is capable of inducing parthenogenetic activation in pig oocytes matured in vitro. However, quite a long exposure to the nitric oxide donor, exceeding 10 h, is necessary for successful oocyte activation. Repeated short-term treatment with 2 mm SNAP significantly increased the activation rates despite the fact that the overall exposure time to the nitric oxide donor did not exceed 4 h. With regard to the activation rate, 12 repeated treatments lasting 10 min each were found to be the most efficient regimen (63.3%). The continuous exposure to the nitric oxide donor for the same overall time induced parthenogenetic activation in 12.5% oocytes (2-h continuous treatment with 2 mm SNAP). The development of parthenogenetic embryos increased after repeated short-term treatment with SNAP. After continuous treatment with 2 mm SNAP for 10 h, only 6.7% of the oocytes cleaved, and none developed beyond the 4-cell stage. Thirty-minute treatment repeated four times with 2 mm SNAP induced cleavage in 37.5% of the oocytes, 18.3% developed to the morula stage, and 6.7% reached the blastocyst stage. Based on the results, it is concluded that pulsatile treatment can significantly improve parthenogenetic activation rate when compared with the continuous treatment using nitric oxide donors.
- MeSH
- donory oxidu dusnatého aplikace a dávkování MeSH
- kultivace embrya veterinární MeSH
- kultivované buňky MeSH
- oocyty účinky léků fyziologie MeSH
- partenogeneze účinky léků fyziologie MeSH
- prasata MeSH
- S-nitroso-N-acetylpenicilamin aplikace a dávkování MeSH
- stadium rýhování vajíčka MeSH
- zvířata MeSH
- Check Tag
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
In this paper we assessed: (i) the change in nitric oxide synthase (NOS) isoforms' expression and intracellular localization and in NOS mRNA in porcine oocytes during meiotic maturation; (ii) the effect of NOS inhibition by N(omega)-nitro-l-arginine methyl ester (l-NAME) and aminoguanidine (AG) on meiotic maturation of cumulus-oocyte complexes (COC) as well as denuded oocytes (DO); and (iii) nitric oxide (NO) formation in COC. All three NOS isoforms (eNOS, iNOS and nNOS) and NOS mRNA (eNOS mRNA, iNOS mRNA and nNOS mRNA) were found in both porcine oocytes and their cumulus cells except for nNOS mRNA, which was not detected in the cumulus cells. NOS isoforms differed in their intracellular localization in the oocyte: while iNOS protein was dispersed in the oocyte cytoplasm, nNOS was localized in the oocyte cytoplasm and in germinal vesicles (GV) and eNOS was present in dots in the cytoplasm, GV and was associated with meiotic spindles. l-NAME inhibitor significantly suppressed metaphase (M)I to MII transition (5.0 mM experimental group: 34.9% MI, control group: 9.5% MI) and at the highest concentration (10.0 mM) also affected GV breakdown (GVBD); in contrast also AG inhibited primarily GVBD. The majority of the oocytes (10.0 mM experimental group: 60.8%, control group: 1.2%) was not able to resume meiosis. AG significantly inhibited GVBD in DO, but l-NAME had no significant effect on the GVBD of these cells. During meiotic maturation, NO is formed in COC and the NO formed by cumulus cells is necessary for the process of GVBD.
- MeSH
- konfokální mikroskopie MeSH
- kumulární buňky enzymologie MeSH
- meióza MeSH
- messenger RNA metabolismus MeSH
- NG-nitroargininmethylester farmakologie MeSH
- oocyty účinky léků enzymologie MeSH
- protein - isoformy antagonisté a inhibitory genetika metabolismus MeSH
- synthasa oxidu dusnatého antagonisté a inhibitory genetika metabolismus MeSH
- zvířata MeSH
- Check Tag
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- index tělesné hmotnosti MeSH
- lidé středního věku MeSH
- lidé MeSH
- morbidní obezita chirurgie ošetřování terapie MeSH
- ošetřovatelská péče metody MeSH
- pacienti hospitalizovaní MeSH
- změny tělesné hmotnosti MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- kazuistiky MeSH
The effect of protein kinase C (PKC) inhibitors on porcine oocyte activation by calcium ionophore A23187 was studied. Calcium ionophore applied in a 50 microM concentration for 10 min induced activation in 74% of oocytes matured in vitro. When the ionophore-treated oocytes were exposed to the effect of bisindolylmaleimide I, which inhibits calcium-dependent PKC isotypes (PKC-alpha, -beta(I), -beta(II), -gamma,) and calcium-independent PKC isotypes (PKC-delta, -epsilon), the portion of activated oocytes decreased (at a concentration of 100 nM, 2% of the oocytes were activated). Go6976, the inhibitor of calcium-dependent PKC isotypes PKC-alpha, -beta(I) did not prevent the action of the oocytes treated with calcium ionophore in concentrations from 1 to 100 microM. The inhibitor of PKC-beta(I) and beta(II) isotypes, hispidin, in a concentration of 2 microM-2 mM, was not effective either. The inhibitor of PKC-delta isotype, rottlerin, suppressed activation of the oocytes by calcium ionophore (no oocyte was activated at 10 microM concentration). The PKC-delta isotype in matured porcine oocytes, studied by Western blot analysis, appeared as non-truncated PKC-delta of 77.5 kDa molecular weight, on the one hand, and as truncated PKC-delta, which was present in the form of a doublet of approximately 62.5 and 68 kDa molecular weight, on the other hand. On the basis of these results, it can be supposed that PKC participates in the regulation of processes associated with oocyte activation. Calcium-dependent PKC-alpha, -beta isotypes do not seem to play any significant role in calcium activation. The activation seems to depend on the activity of the calcium-independent PKC-delta isoform.
- MeSH
- acetofenony farmakologie MeSH
- benzopyrany farmakologie MeSH
- calcimycin farmakologie MeSH
- indoly farmakologie MeSH
- inhibitory proteinkinas farmakologie MeSH
- ionofory farmakologie MeSH
- izoenzymy antagonisté a inhibitory fyziologie klasifikace MeSH
- karbazoly farmakologie MeSH
- maleimidy farmakologie MeSH
- oocyty fyziologie účinky léků MeSH
- prasata fyziologie MeSH
- proteinkinasa C antagonisté a inhibitory fyziologie klasifikace MeSH
- pyrony farmakologie MeSH
- vápník fyziologie MeSH
- western blotting MeSH
- zvířata MeSH
- Check Tag
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- kazuistiky MeSH
