Autophagy is one of the basic cellular mechanism during preimplantation development of mammalian embryos, and it plays crucial role in several physiological processes. It is induced by interleukin (IL)-1β in mammalian cells. Our present study shows that IL-1β is important for autophagy activation in embryo development. Our in vitro culture system analysis shows effect of IL-1β in medium on the development of mouse embryos and it was found to be concentration dependent. A preimplantation embryo culture using medium containing IL-1β did not improve cleavage and blastocyst development rates of mouse embryos; however, blastocyst quality was significantly improved by increasing total cell number, especially in supplementary 20 ng/mL IL-1β. Furthermore, autophagy activation mainly occurs in 2 to 4 cell embryo and blastocyst, 20 ng/mL IL-1β into culture medium can effectively enhance levels of messenger RNA and protein of autophagy-related-factors in 2 to 4 cell embryos and blastocyst, while these factors reduce in VGX-1027 (IL-1β inhibitor) groups that also reduce the quality of blastocyst. Effects of IL-1β on the development of embryo reduced in 20 ng/mL IL-1β supplemented group when 5 mM 3-methyladenine (3-MA) was also added, which used to inhibit autophagy activation in endogenous PtdIns3Ks signal pathway. Our current results show that exogenous IL-1β can effectively induce autophagy in mouse embryos at stages of 2 to 8 cell and blastocyst, that also help to improve the quality of blastocyst.

• Klíčová slova
• IL-1β, assisted reproduction, autophagy, blastocyst quality, cytokine,
• MeSH
• autofagie * MeSH
• blastocysta účinky léků   patologie   MeSH
• embryo savčí účinky léků   patologie   MeSH
• embryonální vývoj účinky léků   MeSH
• interleukin-1beta farmakologie   MeSH
• kultivace embrya MeSH
• myši MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• interleukin-1beta MeSH

Maternal starvation coincident with preimplantation development has profound consequences for placental-fetal development, with various identified pathologies persisting/manifest in adulthood; the 'Developmental Origin of Health and Disease' (DOHaD) hypothesis/model. Despite evidence describing DOHaD-related incidence, supporting mechanistic and molecular data relating to preimplantation embryos themselves are comparatively meager. We recently identified the classically recognized stress-related p38-mitogen activated kinases (p38-MAPK) as regulating formation of the extraembryonic primitive endoderm (PrE) lineage within mouse blastocyst inner cell mass (ICM). Thus, we wanted to assay if PrE differentiation is sensitive to amino acid availability, in a manner regulated by p38-MAPK. Although blastocysts appropriately mature, without developmental/morphological or cell fate defects, irrespective of amino acid supplementation status, we found the extent of p38-MAPK inhibition induced phenotypes was more severe in the absence of amino acid supplementation. Specifically, both PrE and epiblast (EPI) ICM progenitor populations remained unspecified and there were fewer cells and smaller blastocyst cavities. Such phenotypes could be ameliorated, to resemble those observed in groups supplemented with amino acids, by addition of the anti-oxidant NAC (N-acetyl-cysteine), although PrE differentiation deficits remained. Therefore, p38-MAPK performs a hitherto unrecognized homeostatic early developmental regulatory role (in addition to direct specification of PrE), by buffering blastocyst cell number and ICM cell lineage specification (relating to EPI) in response to amino acid availability, partly by counteracting induced oxidative stress; with clear implications for the DOHaD model.

• Klíčová slova
• cell fate, developmental origin of health and disease (DOHaD), mouse blastocyst, oxidative stress, p38-mitogen activated kinases, primitive endoderm,
• Publikační typ
• časopisecké články MeSH

Formation of the hatching mouse blastocyst marks the end of preimplantation development, whereby previous cell cleavages culminate in the formation of three distinct cell lineages (trophectoderm, primitive endoderm and epiblast). We report that dysregulated expression of Wwc2, a genetic paralog of Kibra/Wwc1 (a known activator of Hippo-signaling, a key pathway during preimplantation development), is specifically associated with cell autonomous deficits in embryo cell number and cell division abnormalities. Division phenotypes are also observed during mouse oocyte meiotic maturation, as Wwc2 dysregulation blocks progression to the stage of meiosis II metaphase (MII) arrest and is associated with spindle defects and failed Aurora-A kinase (AURKA) activation. Oocyte and embryo cell division defects, each occurring in the absence of centrosomes, are fully reversible by expression of recombinant HA-epitope tagged WWC2, restoring activated oocyte AURKA levels. Additionally, clonal embryonic dysregulation implicates Wwc2 in maintaining the pluripotent epiblast lineage. Thus, Wwc2 is a novel regulator of meiotic and early mitotic cell divisions, and mouse blastocyst cell fate.

• Klíčová slova
• blastocyst cell number, cell division, cell lineage decision, cell-fate, oocyte maturation, preimplantation mouse embryo,
• Publikační typ
• časopisecké články MeSH

Individual blastomeres of 2-, 4-, 8- and 16-cell embryos were fused with enucleated blastomeres of 2-cell embryos. The percentage of fused nuclear donor/recipient pairs was high (92 to 100%). Most of the reconstituted embryos cleaved twice within 24 h of in vitro culture and 83% to 95% developed to the morula or blastocyst stage. Increasing the number of blastomeres in reconstituted embryos by the construction of aggregates from blastomeres obtained 24 h after nuclear donor/recipient fusion was effective. All constructed aggregates reached the morula or blastocyst stage within 30 h of in vitro culture. The normal day-13 implantations were recorded after the transfer of reconstituted "half" embryos and aggregates when 8-cell blastomeres were used as nuclear donors.

• MeSH
• blastocysta cytologie   MeSH
• blastomery cytologie   MeSH
• buněčné jádro fyziologie   MeSH
• embryonální a fetální vývoj * MeSH
• fúze buněk MeSH
• kultivační techniky MeSH
• morula cytologie   MeSH
• myši MeSH
• přenos embrya MeSH
• stadium rýhování vajíčka MeSH
• techniky jaderného přenosu * MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

The fusion of sperm and oocytes determines the fertilization competence and subsequent development of embryos, which, in turn, can be affected by various proteins and DNA methylation. However, several factors in this whole regulation process remain unknown, especially in yaks. Here, we report that fibroblast growth factor 10 (FGF10) is an important growth factor that can enhance the maturation rate of yak oocytes and the motility of frozen spermatozoa. Subsequent blastocyst quality was also improved by increasing the total cell number and level of pregnancy-associated protein in blastocysts. These effects were significantly high in the group that received the 5 ng/ml FGF10 treatment, during both in vitro maturation (IVM) and capacitation. Our data show that the effects of FGF10 were dose-dependent at vital steps of embryogenesis in vitro. Furthermore, quantitative polymerase chain reaction, western blot analysis, and immunofluorescence demonstrated that the levels of CD9, CD81, DNMT1, and DNMT3B in both mature cumulus-oocyte complexes and capacitated sperms were regulated by FGF10, which was also highly expressed in the group treated with 5 ng/ml FGF10 during both IVM and capacitation. From our present study, we concluded that FGF10 promotes yak oocyte fertilization competence and subsequent blastocyst quality, and could also regulate CD9, CD81, DNMT1, and DNMT3B to optimize sperm-oocyte interactions and DNA methylation during fertilization.

• Klíčová slova
• DNA methylation, embryonic development, fertilization, fibroblast growth factors (FGFs), sperm-oocyte fusion, yak,
• MeSH
• antigeny CD81 genetika   metabolismus   MeSH
• antigeny CD9 genetika   metabolismus   MeSH
• blastocysta účinky léků   fyziologie   MeSH
• DNA-(cytosin-5-)methyltransferasa genetika   metabolismus   MeSH
• DNA-(cytosin-5)-methyltransferasa 1 genetika   metabolismus   MeSH
• DNA-methyltransferasa 3B MeSH
• embryonální vývoj účinky léků   genetika   fyziologie   MeSH
• fertilizace in vitro veterinární   MeSH
• fertilizace účinky léků   genetika   fyziologie   MeSH
• fibroblastový růstový faktor 10 aplikace a dávkování   fyziologie   MeSH
• IVM techniky veterinární   MeSH
• messenger RNA genetika   metabolismus   MeSH
• oocyty účinky léků   fyziologie   MeSH
• skot embryologie   genetika   fyziologie   MeSH
• těhotenství MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• skot embryologie   genetika   fyziologie   MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antigeny CD81 MeSH
• antigeny CD9 MeSH
• DNA-(cytosin-5-)methyltransferasa MeSH
• DNA-(cytosin-5)-methyltransferasa 1 MeSH
• fibroblastový růstový faktor 10 MeSH
• messenger RNA MeSH

The aim of this study was to compare the effect of purified GPBoS and commonly used FCS on porcine oocyte maturation and subsequent embryonic development after their parthenogenetic activation. COCs were obtained from dissected follicles and cultured for 18, 24, 30, 36, 42 and 48 h in M-199 medium either with GPBoS or FCS. After 24 h with GPBoS, 91% of oocytes reached MI stage while in the medium supplemented with FCS, only 29% of oocytes reached the same stage (P < 0.05). The majority of oocytes from the FCS group (61%) reached MI stage approximately 6 h later. In the time periods between 36 to 48 h both groups of oocytes reached the same stage of maturation. After 48 h of culture the oocytes with extruded polar bodies were activated by a single electric pulse and then cultured with 4 mM 6-DMAP. Activated oocytes were cultured in PZM-3 medium supplemented with 3 mg/ml of BSA. After 7 days, the development and the quality of embryos were evaluated. The results showed that the maturation of oocytes in the presence of GPBoS significantly increased their subsequent developmental ability when compared with FCS supplementation (27% vs. 19% of blastocysts, P < 0.05). However, differential staining revealed that once blastocysts were formed in either group, they had the same total cell number (40 vs. 41) and also the ICM/total cell ratio (0.27 vs. 0.29).

• MeSH
• blastocysta cytologie   účinky léků   metabolismus   MeSH
• buněčná diferenciace účinky léků   fyziologie   MeSH
• buněčné dělení účinky léků   fyziologie   MeSH
• buněčné jádro účinky léků   metabolismus   ultrastruktura   MeSH
• časové faktory MeSH
• krevní proteiny metabolismus   farmakologie   MeSH
• kultivace embrya metody   MeSH
• kultivační média metabolismus   farmakologie   MeSH
• kultivované buňky MeSH
• oocyty cytologie   účinky léků   metabolismus   MeSH
• proliferace buněk účinky léků   MeSH
• Sus scrofa MeSH
• zvětšování buněk účinky léků   MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• krevní proteiny MeSH
• kultivační média MeSH

The aim of this work was to assess the FSH-stimulated expression of epidermal growth factor (EGF)-like peptides in cultured cumulus-oocyte complexes (COCs) and to find out the effect of the peptides on cumulus expansion, oocyte maturation, and acquisition of developmental competence in vitro. FSH promptly stimulated expression of amphiregulin (AREG) and epiregulin (EREG), but not betacellulin (BTC) in the cultured COCs. Expression of AREG and EREG reached maximum at 2 or 4 h after FSH addition respectively. FSH also significantly stimulated expression of expansion-related genes (PTGS2, TNFAIP6, and HAS2) in the COCs at 4 and 8 h of culture, with a significant decrease at 20 h of culture. Both AREG and EREG also increased expression of the expansion-related genes; however, the relative abundance of mRNA for each gene was much lower than in the FSH-stimulated COCs. In contrast to FSH, AREG and EREG neither stimulated expression of CYP11A1 in the COCs nor an increase in progesterone production by cumulus cells. AREG and EREG stimulated maturation of oocytes and expansion of cumulus cells, although the percentage of oocytes that had reached metaphase II was significantly lower when compared to FSH-induced maturation. Nevertheless, significantly more oocytes stimulated with AREG and/or EREG developed to blastocyst stage after parthenogenetic activation when compared to oocytes stimulated with FSH alone or combinations of FSH/LH or pregnant mares serum gonadotrophin/human chorionic gonadotrophin. We conclude that EGF-like peptides do not mimic all effects of FSH on the cultured COCs; nevertheless, they yield oocytes with superior developmental competence.

• MeSH
• buněčná diferenciace účinky léků   genetika   MeSH
• embryonální vývoj účinky léků   genetika   MeSH
• epidermální růstový faktor chemie   farmakologie   MeSH
• folikuly stimulující hormon farmakologie   MeSH
• gonadotropiny farmakologie   MeSH
• kultivace embrya MeSH
• kultivované buňky MeSH
• kumulární buňky účinky léků   metabolismus   fyziologie   MeSH
• oocyty účinky léků   metabolismus   fyziologie   MeSH
• oogeneze účinky léků   genetika   MeSH
• partenogeneze účinky léků   genetika   fyziologie   MeSH
• peptidové fragmenty chemie   farmakologie   MeSH
• prasata * genetika   metabolismus   fyziologie   MeSH
• proliferace buněk účinky léků   MeSH
• stanovení celkové genové exprese MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• epidermální růstový faktor MeSH
• folikuly stimulující hormon MeSH
• gonadotropiny MeSH
• peptidové fragmenty MeSH

Preimplantation mouse embryo development involves temporal-spatial specification and segregation of three blastocyst cell lineages: trophectoderm, primitive endoderm and epiblast. Spatial separation of the outer-trophectoderm lineage from the two other inner-cell-mass (ICM) lineages starts with the 8- to 16-cell transition and concludes at the 32-cell stages. Accordingly, the ICM is derived from primary and secondary contributed cells; with debated relative EPI versus PrE potencies. We report generation of primary but not secondary ICM populations is highly dependent on temporal activation of mammalian target of Rapamycin (mTOR) during 8-cell stage M-phase entry, mediated via regulation of the 7-methylguanosine-cap (m7G-cap)-binding initiation complex (EIF4F) and linked to translation of mRNAs containing 5' UTR terminal oligopyrimidine (TOP-) sequence motifs, as knockdown of identified TOP-like motif transcripts impairs generation of primary ICM founders. However, mTOR inhibition-induced ICM cell number deficits in early blastocysts can be compensated by the late blastocyst stage, after inhibitor withdrawal; compensation likely initiated at the 32-cell stage when supernumerary outer cells exhibit molecular characteristics of inner cells. These data identify a novel mechanism specifically governing initial spatial segregation of mouse embryo blastomeres, that is distinct from those directing subsequent inner cell formation, contributing to germane segregation of late blastocyst lineages.

• Klíčová slova
• EIF4EBP1/4EBP1, TOP-motif, cell fate, inner cell mass/ICM and cell positioning, mTOR/mTORC1, preimplantation mouse embryo,
• MeSH
• blastocysta * MeSH
• buněčná diferenciace fyziologie   MeSH
• buněčný rodokmen MeSH
• embryo savčí * MeSH
• mTORC1 MeSH
• myši MeSH
• savci MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• mTORC1 MeSH

Growth of the remnant embryonic kidney (the mesonephros), as expressed by wet weight, was more rapid in the chick embryos with experimentally induced unilateral renal agenesis compared to controls. The difference was significant between embryonic days 8-12, when the doubled weights of remnant kidneys were increased compared with the weights of paired control kidneys. The excessive growth of the mesonephros ceased on day 14, when the normal physiological regression of the embryonic kidney begins. In the definitive kidney, the metanephros, no significant differences in weights of the control vs. remnant metanephros were found on days 10-14. The characteristics of increased mesonephric growth were evaluated by determination of DNA/protein ratios in homogenates of the kidneys. Significant cellular hypertrophy was found in both the mesonephros and metanephros of the embryos with URA on day 10. Additionally, a non-significant cellular hyperplasia was also revealed in the remnant mesonephros on day 8. This gives evidence that the growth stimuli to the mesonephroi were probably strongest between days 8-10 and that they manifested in the remnant mesonephros only.

• MeSH
• časové faktory MeSH
• hyperplazie MeSH
• hypertrofie MeSH
• kultivace embrya MeSH
• kuřecí embryo MeSH
• ledviny abnormality   embryologie   růst a vývoj   patologie   MeSH
• mezonefros růst a vývoj   patologie   MeSH
• proliferace buněk MeSH
• replikace DNA MeSH
• zvířata MeSH
• Check Tag
• kuřecí embryo MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Data on the frequency of aneuploidy in farm animals are lacking and there is the need for a reliable technique which is capable of detecting all chromosomes simultaneously in a single cell. With the employment of comparative genomic hybridization coupled with the whole genome amplification technique, this study brings new information regarding the aneuploidy of individual chromosomes in pigs. Focus is directed on in vivo porcine blastocysts and late morulas, 4.7% of which were found to carry chromosomal abnormality. Further, ploidy abnormalities were examined using FISH in a sample of porcine embryos. True polyploidy was relatively rare (1.6%), whilst mixoploidy was presented in 46.8% of embryos, however it was restricted to only a small number of cells per embryo. The combined data indicates that aneuploidy is not a prevalent cause of embryo mortality in pigs.

• MeSH
• aneuploidie * MeSH
• blastocysta cytologie   metabolismus   fyziologie   MeSH
• embryo savčí MeSH
• gestační stáří MeSH
• nemoci prasat diagnóza   embryologie   genetika   MeSH
• oocyty cytologie   metabolismus   fyziologie   MeSH
• prasata genetika   MeSH
• srovnávací genomová hybridizace metody   veterinární   MeSH
• těhotenství MeSH
• ztráta embrya diagnóza   genetika   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• práce podpořená grantem MeSH