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.

• 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

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.

• 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
• 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

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

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.

• 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

STUDY QUESTION: Do embryologists from different European countries agree on embryo disposition decisions ('use' or 'discard') about Day 7 (>144 h post-insemination) and/or low-quality blastocysts (LQB;

• MeSH
• aneuploidie MeSH
• blastocysta * fyziologie   MeSH
• lidé MeSH
• přenos embrya metody   MeSH
• retrospektivní studie MeSH
• umělá inteligence * MeSH
• věk matky MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články 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

The goal of this study was to examine the effect of insulin-like growth factor I (IGF-I; added during post-thaw culture (48 h)) on the preimplantation viability and quality of cryopreserved bovine in vivo recovered embryos. The morula stage embryos, non-surgically recovered from superovulated dairy cows of Czech Fleckvieh cattle breed, had previously been cryopreserved by a slow freezing technique and stored in liquid nitrogen since 1989-1990. Following thawing, the embryos were cultured for 48 h either alone (no IGF-I) or in the presence of IGF-I (10 or 100 ng/ml); non-cultured embryos served as a control. Thereafter, the embryos were analyzed for cleavage to the blastocyst stage, apoptosis (TUNEL), embryo cell number and quality of actin cytoskeleton. Following post-thaw culture 41% of embryos developed to advanced blastocysts. IGF-I increased this per cent and, at a higher dose, essentially reduced the per cent of degenerated embryos. In cultured embryos, IGF-I at both doses elevated the cell number compared with non-cultured embryos. However, in comparison with embryos cultured without IGF-I, only the higher IGF-I dose resulted in elevating the embryo cell number. The TUNEL index was significantly lowered by IGF-I treatment. Thawed embryos were mostly of the grade III actin type and fewer (12%) had grade II actin, whilst no grade I actin embryos were noted. The addition of IGF-I resulted in the appearance of grade I actin embryos (8.33 and 6.9% for 10 and 100 ng/ml, respectively). These observations indicate that the addition of IGF-I during post-thaw culture can improve the quality of bovine cryopreserved embryos.

• MeSH
• aktiny metabolismus   MeSH
• apoptóza účinky léků   MeSH
• blastocysta cytologie   účinky léků   metabolismus   MeSH
• embryonální vývoj MeSH
• insulinu podobný růstový faktor I farmakologie   MeSH
• kryoprezervace metody   MeSH
• kultivace embrya metody   MeSH
• kultivační média chemie   farmakologie   MeSH
• morula účinky léků   MeSH
• počet buněk MeSH
• skot MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• skot MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The similarity between humans and pigs, when it comes to tissue morphology, makes Sus scrofa not only a good research model, but also a potential source of cells for tissue engineering. Cell samples obtained from the pig donor, could be influenced in vitro, in order to become a source of tissue material for xenotransplantation, reconstructive and regenerative medicine. Significant amounts of data point to especially major similarities in pig and human reproductive systems. Because of that, particular scientific focus is centered on research concerning porcine COCs, theca and granulosa cells in primary cultures. One of the aspects of the reproductive process, that is still largely undiscovered, is the interaction between preimplantation blastocyst and maternal uterine tissues. In this study, we used molecular analysis techniques, such as RT-qPCR and immunocytochemistry, to analyze the expression and distribution of cytokeratin 18 and panCytokeratins 8, 18 and 19 and vimentin in porcine luminal endometrial epithelial cells, coupled with analysis of their behavior in RTCA. The results have confirmed the presence of epithelial, as well as stromal cell markers in the cells, varying in levels at different stages of culture. They have also given insight into the modes of proliferation and differentiation of studied cells in in vitro culture, as well as providing additional proof for the possible mesenchymal transdifferentiation of epithelial cells.

• MeSH
• biologické markery metabolismus   MeSH
• biologické modely MeSH
• buněčná diferenciace MeSH
• buňky stromatu cytologie   metabolismus   MeSH
• časové faktory MeSH
• endometrium cytologie   MeSH
• epitelové buňky cytologie   metabolismus   MeSH
• kultivované buňky MeSH
• lidé MeSH
• modely u zvířat MeSH
• prasata MeSH
• proliferace buněk * MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

This study examined the impact of cow body condition on the quality of bovine preimplantation embryos. The embryos (n = 107) were flushed from dairy cows and classified according to a five-point scale body condition score (BCS2 n = 17; BCS3 n = 31; BCS4 n = 11) on the 7th day after insemination and then analyzed for development, dead cell index (DCI), cell number and actin cytoskeleton quality. The highest embryo recovery rate (P < 0.05) was recorded in the BCS3 group and the lowest in the BCS4 group. More transferable (morula, blastocyst) embryos were obtained from the BCS4 cows (79%), compared with the BCS2 (64%) or BCS3 (63%) animals. However, cell numbers were higher in the BCS2 and BCS3 groups (P < 0.05) compared with the BCS4 embryos. Conversely, the DCI was lowest in the BCS2 (3.88%; P < 0.05) and highest in the BCS4 (6.56%) embryos. The proportion of embryos with the best actin quality (grade I) was higher in the BCS2 and BCS3 cows compared with the BCS4 group. Almost 25% of all embryos showed fragmented morphology and a higher DCI (5.65%) than normal morulas (1.76%). More fragmented embryos were revealed in the BCS2 (28.6%) and BCS4 (31.25%) groups, and less (19.15%) in the BCS3 group. The cell numbers in such embryos were lower in the BCS4 (22.57) than in the BCS2 (46.25) or BCS3 (42.4) groups. In conclusion, the body condition of dairy cows affects the quality of preimplantation embryos. A BCS over 3.0 resulted in a higher incidence of poor (fragmented) embryos.

• MeSH
• apoptóza MeSH
• blastocysta cytologie   metabolismus   MeSH
• buněčné dělení MeSH
• fertilizace in vitro metody   MeSH
• koncové značení zlomů DNA in situ MeSH
• konfokální mikroskopie MeSH
• mikrofilamenta metabolismus   MeSH
• mlékárenství MeSH
• morula cytologie   metabolismus   MeSH
• počet buněk MeSH
• přenos embrya metody   MeSH
• skot MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• skot MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Cíl studie: Ověřit využití oocytů bez zony pellucidy (zona free oocyty) pro léčbu neplodnostimetodami asistované reprodukce.Typ studie: Pilotní studie.Název a sídlo pracoviště: Centrum asistované reprodukce, Gynekologicko-porodnická klinika 1.LF UK a VFN, Praha.Metodika: Oocyty bez zony pellucidy byly oplozeny metodou injekce jedné spermie do cytoplazmy(ICSI) a kultivovány běžným způsobem do stadia blastocysty, které byly kryokonzervovány.Výsledky: První stadium po oplození oocytů metodou ICSI probíhalo fyziologicky, oocyty bez ZPvyloučily polární tělísko a ve standardní době, tedy za 18 hodin, obsahovaly v cytoplazmě dvěmorfologicky normální prvojádra. V průběhu časného rýhování (první až třetí den kultivace invitro) byla dynamika rýhování normální, ale prostorové uspořádání buněk neodpovídalo jejichtrojrozměrnému uskupení u embryí uzavřených v zoně pellucidě. Ve stádiu kompaktizace a vývo-je blastocysty se morfologický vývoj nelišil od standardního embrya.Závěr: Oocyty bez ZP lze úspěšně oplodnit metodou ICSI. U pacientek s opakovaně nízkým počtemzískaných oocytů a nebo u žen s defektem ZP může ICSI oocytu bez ZP významně zvýšit šanci naotěhotnění.

Objective: The study of zona free oocytes fertilization using intracytoplasmic sperm injection(ICSI) with the aim to increase the pregnancy rate in the assisted reproduction.Design: Pilot study.Setting: Assisted Reproduction Centre, Gynecology Obstetrics Department of the 1st MedicalFaculty and General Faculty Hospital, Charles University, Prague.Methods: Zona free oocytes were fertilized using ICSI and cultured in standard conditions in vitrofor 5 days until the blastocyst stage. Blastocysts were cryopreserved using 8% glycerol.Results: The fertilization process of zona free oocyte was physiological, oocytes expulsed the polarbody and in standard time, i.e. 18 hour post insemination, two distinct, morphologically normalpronuclei were apparent in cytoplasm. During early cleavage, from the first to the third day of invitro culture, the dynamics of cell division was normal, the three-dimensional arrangement ofcells was more flattened than in normal, zona intact embryo. The fourth and fifth day of culture,the morphological appearance of embryo corresponded to normal development.Conclusion: The fertilization of zona free oocytes using ICSI can give morphologically normalzygote and after 5 days culture in vitro these zygotes can develop into blastocyst stage. Thismethod can increase the pregnancy rate in patients with repeatedly low number of oocytes orwith defects of zona pellucida.

• MeSH
• blastocysta MeSH
• buněčná diferenciace MeSH
• dospělí MeSH
• embryo savčí MeSH
• intracytoplazmatické injekce spermie metody   MeSH
• lidé MeSH
• oocyty MeSH
• ženská infertilita terapie   MeSH
• zona pellucida MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• přehledy MeSH