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Статті в журналах з теми "Bovine embryonic"
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Toralová, Tereza, Veronika Benešová, Kateřina Vodičková Kepková, Petr Vodička, Andrej Šušor, and Jiří Kaňka. "Bovine preimplantation embryos with silenced nucleophosmin mRNA are able to develop until the blastocyst stage." REPRODUCTION 144, no. 3 (September 2012): 349–59. http://dx.doi.org/10.1530/rep-12-0033.
Повний текст джерелаАнотація:
This study was conducted to investigate the effect of silencing nucleophosmin in the development of in vitro-produced bovine embryos. Nucleophosmin is an abundant multifunctional nucleolar phosphoprotein that participates, for example, in ribosome biogenesis or centrosome duplication control. We showed that although the transcription of embryonic nucleophosmin started already at late eight-cell stage, maternal protein was stored throughout the whole preimplantation development and was sufficient for the progression to the blastocyst stage. At the beginning of embryogenesis, translation occurs on maternally derived ribosomes, the functionally active nucleoli emerge during the fourth cell cycle in bovines. We found that nucleophosmin localisation reflected the nucleolar formation during bovine preimplantation development. The protein was detectable from the beginning of embryonic development. Before embryonic genome activation, it was dispersed throughout the nucleoplasm. The typical nucleolar localisation emerged with the formation of active nucleoli. At the blastocyst stage, nucleophosmin tended to localise especially to the trophectoderm. To see for how long is maternal nucleophosmin preserved, we silenced the nucleophosmin mRNA using RNA interference approach. Although a large portion of nucleophosmin was degraded in embryos with silenced nucleophosmin mRNA, an amount sufficient for normal development was preserved and we detected only a temporal delay in nucleophosmin relocalisation to nucleoli. Moreover, we observed no defects in nuclear shape or cytoskeleton previously found in somatic cells and only a non-significant decrease in embryonic developmental competence. Thus, our results show that the preserved amount of maternal nucleophosmin is sufficient for preimplantation development of bovine embryo.
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Fu, Yao, Jia-Jun Xu, Xu-Lei Sun, Hao Jiang, Dong-Xu Han, Chang Liu, Yan Gao, Bao Yuan, and Jia-Bao Zhang. "Function of JARID2 in bovines during early embryonic development." PeerJ 5 (December 21, 2017): e4189. http://dx.doi.org/10.7717/peerj.4189.
Повний текст джерелаАнотація:
Histone lysine modifications are important epigenetic modifications in early embryonic development. JARID2, which is a member of the jumonji demethylase protein family, is a regulator of early embryonic development and can regulate mouse development and embryonic stem cell (ESC) differentiation by modifying histone lysines. JARID2 can affect early embryonic development by regulating the methylation level of H3K27me3, which is closely related to normal early embryonic development. To investigate the expression pattern of JARID2 and the effect of JARID2-induced H3K27 methylation in bovine oocytes and early embryonic stages, JARID2 mRNA expression and localization were detected in bovine oocytes and early embryos via qRT-PCR and immunofluorescence in the present study. The results showed that JARID2 is highly expressed in the germinal vesicle (GV), MII, 2-cell, 4-cell, 8-cell, 16-cell and blastocyst stages, but the relative expression level of JARID2 in bovine GV oocytes is significantly lower than that at other oocyte/embryonic stages (p < 0.05), and JARID2 is expressed primarily in the nucleus. We next detected the mRNA expression levels of embryonic development-related genes (OCT4, SOX2 and c-myc) after JARID2 knockdown through JARID2-2830-siRNA microinjection to investigate the molecularpathwayunderlying the regulation of H3K27me3 by JARID2 during early embryonic development. The results showed that the relative expression levels of these genes in 2-cell embryos weresignificantly higher than those in the blastocyst stage, and expression levels were significantly increased after JARID2 knockdown. In summary, the present study identified the expression pattern of JARID2 in bovine oocytes and at each early embryonic stage, and the results suggest that JARID2 plays a key role in early embryonic development by regulating the expression of OCT4, SOX2 and c-myc via modification of H3K27me3 expression. This work provides new data for improvements in the efficiency ofin vitroembryo culture as well as a theoretical basis for further studying the regulatory mechanisms involved in early embryonic development.
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Perera, Chalani Dilshani, Muhammad Idrees, Abdul Majid Khan, Zaheer Haider, Safeer Ullah, Ji-Su Kang, Seo-Hyun Lee, Seon-Min Kang та Il-Keun Kong. "PDGFRβ Activation Induced the Bovine Embryonic Genome Activation via Enhanced NFYA Nuclear Localization". International Journal of Molecular Sciences 24, № 23 (1 грудня 2023): 17047. http://dx.doi.org/10.3390/ijms242317047.
Повний текст джерелаАнотація:
Embryonic genome activation (EGA) is a critical step during embryonic development. Several transcription factors have been identified that play major roles in initiating EGA; however, this gradual and complex mechanism still needs to be explored. In this study, we investigated the role of nuclear transcription factor Y subunit A (NFYA) in bovine EGA and bovine embryonic development and its relationship with the platelet-derived growth factor receptor-β (PDGFRβ) by using a potent selective activator (PDGF-BB) and inhibitor (CP-673451) of PDGF receptors. Activation and inhibition of PDGFRβ using PDGF-BB and CP-673451 revealed that NFYA expression is significantly (p < 0.05) affected by the PDGFRβ. In addition, PDGFRβ mRNA expression was significantly increased (p < 0.05) in the activator group and significantly decreased (p < 0.05) in the inhibitor group when compared with PDGFRα. Downregulation of NFYA following PDGFRβ inhibition was associated with the expression of critical EGA-related genes, bovine embryo development rate, and implantation potential. Moreover, ROS and mitochondrial apoptosis levels and expression of pluripotency-related markers necessary for inner cell mass development were also significantly (p < 0.05) affected by the downregulation of NFYA while interrupting trophoblast cell (CDX2) differentiation. In conclusion, the PDGFRβ-NFYA axis is critical for bovine embryonic genome activation and embryonic development.
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Degrelle, Séverine A., Kim-Anh Lê Cao, Yvan Heyman, Robin E. Everts, Evelyne Campion, Christophe Richard, Céline Ducroix-Crépy, et al. "A small set of extra-embryonic genes defines a new landmark for bovine embryo staging." REPRODUCTION 141, no. 1 (January 2011): 79–89. http://dx.doi.org/10.1530/rep-10-0174.
Повний текст джерелаАнотація:
Axis specification in mouse is determined by a sequence of reciprocal interactions between embryonic and extra-embryonic tissues so that a few extra-embryonic genes appear as ‘patterning’ the embryo. Considering these interactions as essential, but lacking in most mammals the genetically driven approaches used in mouse and the corresponding patterning mutants, we examined whether a molecular signature originating from extra-embryonic tissues could relate to the developmental stage of the embryo proper and predict it. To this end, we have profiled bovine extra-embryonic tissues at peri-implantation stages, when gastrulation and early neurulation occur, and analysed the subsequent expression profiles through the use of predictive methods as previously reported for tumour classification. A set of six genes (CALM1, CPA3, CITED1, DLD, HNRNPDL, and TGFB3), half of which had not been previously associated with any extra-embryonic feature, appeared significantly discriminative and mainly dependent on embryonic tissues for its faithful expression. The predictive value of this set of genes for gastrulation and early neurulation stages, as assessed on naive samples, was remarkably high (93%). In silico connected to the bovine orthologues of the mouse patterning genes, this gene set is proposed as a new trait for embryo staging. As such, this will allow saving the bovine embryo proper for molecular or cellular studies. To us, it offers as well new perspectives for developmental phenotyping and modelling of embryonic/extra-embryonic co-differentiation.
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Stice, S. L. "Pluripotent bovine embryonic cell lines direct embryonic development following nuclear transfer." Biology of Reproduction 54, no. 1 (January 1, 1996): 100–110. http://dx.doi.org/10.1095/biolreprod54.1.100.
Повний текст джерела
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Asl, M. Pashai, K. Khodadadi, M. K. Holland, N. M. Richings, and P. J. Verma. "430. Investigation of pluripotency in derived embryonic stem cell (ESC) lines." Reproduction, Fertility and Development 20, no. 9 (2008): 110. http://dx.doi.org/10.1071/srb08abs430.
Повний текст джерелаАнотація:
To produce autologous ESCs for a bovine model of cell therapy, we activated oocytes by calcium ionophore (CI) and 6 dimethylaminopurine (6 DMAP) and isolated ESCs from the resulting parthenotes. Parthenote ESC lines (pbESC) would also provide a valuable tool for epigenetic studies on ESCs. Five pbESC like-cell lines were expanded for 12 passages over 120 days and differentiated to form embryoid bodies by suspension culture. The pbESC lines demonstrated typical ESC morphology and expressed ESC markers including alkaline phosphate and stage-specific embryonic antigen, SSEA1 and SSAE4 asssessed by histochemical and immuno-fluorescence staining, respectively. In addition, gene expression of Oct4, Rex1, SSEA1 and ALP was confirmed using RT–PCR. These cells had a normal karyotype. The cells formed EBs and showed expression of the markers of three embryonic germ layers. In summary, we show that ESCs can be derived from bovine parthenogenetic blastocysts and that these cells express pluripotent markers and have ability to form EBs and differentiate into cells indicative of the three embryonic germ layers. Additional work will focus on imprinted gene expression and will provide further evidence of the parthenogenetic origin of the pbESC lines.
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Saadeldin, Islam M., Ahmed Abdelfattah-Hassan, and Ayman Abdel-Aziz Swelum. "Feeder Cell Type Affects the Growth of In Vitro Cultured Bovine Trophoblast Cells." BioMed Research International 2017 (2017): 1–6. http://dx.doi.org/10.1155/2017/1061589.
Повний текст джерелаАнотація:
Trophectoderm cells are the foremost embryonic cells to differentiate with prospective stem-cell properties. In the current study, we aimed at improving the current approach for trophoblast culture by using granulosa cells as feeders. Porcine granulosa cells (PGCs) compared to the conventional mouse embryonic fibroblasts (MEFs) were used to grow trophectoderm cells from hatched bovine blastocysts. Isolated trophectoderm cells were monitored and displayed characteristic epithelial/cuboidal morphology. The isolated trophectoderm cells expressed mRNA of homeobox protein (CDX2), cytokeratin-8 (KRT8), and interferon tau (IFNT). The expression level was higher on PGCs compared to MEFs throughout the study. In addition, primary trophectoderm cell colonies grew faster on PGCs, with a doubling time of approximately 48 hrs, compared to MEFs. PGCs feeders produced a fair amount of 17β-estradiol and progesterone. We speculated that the supplementation of sex steroids and still-unknown factors during the trophoblasts coculture on PGCs have helped to have better trophectoderm cell’s growth than on MEFs. This is the first time to use PGCs as feeders to culture trophectoderm cells and it proved superior to MEFs. We propose PGCs as alternative feeders for long-term culture of bovine trophectoderm cells. This model will potentially benefit studies on the early trophoblast and embryonic development in bovines.
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Wu, Shanshan, Xiaoyu Zhao, Meiling Wu, Lei Yang, Xuefei Liu, Danyi Li, Han Xu, et al. "Low Expression of Mitofusin 1 Gene Leads to Mitochondrial Dysfunction and Embryonic Genome Activation Failure in Ovine-Bovine Inter-Species Cloned Embryos." International Journal of Molecular Sciences 23, no. 17 (September 4, 2022): 10145. http://dx.doi.org/10.3390/ijms231710145.
Повний текст джерелаАнотація:
Inter-species somatic cell nuclear transfer (iSCNT) is significant in the study of biological problems such as embryonic genome activation and the mitochondrial function of embryos. Here, we used iSCNT as a model to determine whether abnormal embryo genome activation was caused by mitochondrial dysfunction. First, we found the ovine-bovine iSCNT embryos were developmentally blocked at the 8-cell stage. The reactive oxygen species level, mitochondrial membrane potential, and ATP level in ovine-bovine cloned embryos were significantly different from both bovine-bovine and IVF 8-cell stage embryos. RNA sequencing and q-PCR analysis revealed that mitochondrial transport, mitochondrial translational initiation, mitochondrial large ribosomal subunit, and mitochondrial outer membrane genes were abnormally expressed in the ovine-bovine embryos, and the mitochondrial outer membrane and mitochondrial ribosome large subunit genes, mitochondrial fusion gene 1, and ATPase Na+/K+ transporting subunit beta 3 gene were expressed at lower levels in the ovine-bovine cloned embryos. Furthermore, we found that overexpression and knockdown of Mfn1 significantly affected mitochondrial fusion and subsequent biological functions such as production of ATP, mitochondrial membrane potential, reactive oxygen species and gene expressions in cloned embryos. These findings enhance our understanding of the mechanism by which the Mfn1 gene regulates embryonic development and embryonic genome activation events.
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Speckhart, Savannah L., Mary A. Oliver, and Alan D. Ealy. "Developmental Hurdles That Can Compromise Pregnancy during the First Month of Gestation in Cattle." Animals 13, no. 11 (May 25, 2023): 1760. http://dx.doi.org/10.3390/ani13111760.
Повний текст джерелаАнотація:
Several key developmental events are associated with early embryonic pregnancy losses in beef and dairy cows. These developmental problems are observed at a greater frequency in pregnancies generated from in-vitro-produced bovine embryos. This review describes critical problems that arise during oocyte maturation, fertilization, early embryonic development, compaction and blastulation, embryonic cell lineage specification, elongation, gastrulation, and placentation. Additionally, discussed are potential remediation strategies, but unfortunately, corrective actions are not available for several of the problems being discussed. Further research is needed to produce bovine embryos that have a greater likelihood of surviving to term.
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Yang, Cai-Xia, Zichuan Liu, Renaud Fleurot, Pierre Adenot, Véronique Duranthon, Xavier Vignon, Qi Zhou, Jean-Paul Renard, and Nathalie Beaujean. "Heterochromatin reprogramming in rabbit embryos after fertilization, intra-, and inter-species SCNT correlates with preimplantation development." REPRODUCTION 145, no. 2 (February 2013): 149–59. http://dx.doi.org/10.1530/rep-11-0421.
Повний текст джерелаАнотація:
To investigate the embryonic genome organization upon fertilization and somatic cell nuclear transfer (SCNT), we tracked HP1β and CENP, two well-characterized protein markers of pericentric and centromeric compartments respectively, in four types of embryos produced by rabbit in vivo fertilization, rabbit parthenogenesis, rabbit-to-rabbit, and bovine-to-rabbit SCNT. In the interphase nuclei of rabbit cultured fibroblasts, centromeres and associated pericentric heterochromatin are usually isolated. Clustering into higher-order chromatin structures, such as the chromocenters seen in mouse and bovine somatic cells, could not be observed in rabbit fibroblasts. After fertilization, centromeres and associated pericentric heterochromatin are quite dispersed in rabbit embryos. The somatic-like organization is progressively established and completed only by the 8/16-cell stage, a stage that corresponds to major embryonic genome activation in this species. In SCNT embryos, pericentric heterochromatin distribution typical for rabbit and bovine somatic cells was incompletely reverted into the 1-cell embryonic form with remnants of heterochromatin clusters in 100% of bovine-to-rabbit embryos. Subsequently, the donor cell nuclear organization was rapidly re-established by the 4-cell stage. Remarkably, the incomplete remodeling of bovine-to-rabbit 1-cell embryos was associated with delayed transcriptional activation compared with rabbit-to-rabbit embryos. Together, the results confirm that pericentric heterochromatin spatio-temporal reorganization is an important step of embryonic genome reprogramming. It also appears that genome reorganization in SCNT embryos is mainly dependent on the nuclear characteristics of the donor cells, not on the recipient cytoplasm.
Дисертації з теми "Bovine embryonic"
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Pant, Disha. "Towards the derivation of bovine embryonic stem cells." College Park, Md.: University of Maryland, 2008. http://hdl.handle.net/1903/7854.
Повний текст джерелаАнотація:
Thesis (Ph. D.) -- University of Maryland, College Park, 2008.Thesis research directed by: Dept. of Animal and Avian Sciences. Title from t.p. of PDF. Includes bibliographical references. Published by UMI Dissertation Services, Ann Arbor, Mich. Also available in paper.
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Silva, Bruna Mazeti [UNESP]. "Efeitos do agente modificador epigenético (Tricostatina A) no desenvolvimento embrionário pré-implantacional de embriões fêmeas de bovinos." Universidade Estadual Paulista (UNESP), 2011. http://hdl.handle.net/11449/92589.
Повний текст джерелаАнотація:
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A acetilação das histonas é um dos principais mecanismos de reprogramação epigenética do genoma dos gametas a fim de estabelecer um estado totipotente para o desenvolvimento normal. O objetivo deste trabalho foi avaliar o efeito da utilização da Tricostatina A (TSA) sobre os níveis de acetilação das histonas e o desenvolvimento embrionário, avaliando-se a contagem do número de células da massa celular interna (MCI), trofectoderma (TE), e células totais, e os níveis da reação de imunocitoquímica para H3K9ac. Para isso, três experimentos foram delineados. No primeiro experimento, verificou-se que o grupo tratado com 5nM de TSA por 144h durante o cultivo in vitro (CIV) aumentou (p<0,01) o nível de H3K9ac em relação ao grupo controle. A taxa de clivagem e o desenvolvimento embrionário foram avaliados em um segundo experimento, e o grupo tratado com 5nM de TSA não apresentou diferença na produção de embriões comparado ao grupo controle. No experimento 3, o tratamento com a TSA não apresentou efeitos significativos em relação ao número de células da MCI, porém houve redução (p<0,01) tanto do número de células da TE quanto de células totais comparado ao grupo controle. Portanto, conclui-se que o tratamento com TSA não altera o desenvolvimento embrionário pré-implantacional de embriões fêmeas de bovinos, mostrando efeito negativo sobre o desenvolvimento dos embriões tratados
Histone acetylation is one of the major mechanisms of epigenetic reprogramming of gamete genome to establish a totipotent state for normal development. The aim of this work was to evaluate the use of Trichostatin A (TSA) on the levels of histone acetylation and embryonic development, the assessment of counting the cell number of inner cell mass (ICM), trophectoderm (TE), and total cells, and the levels of H3K9ac for immunocytochemical reaction. Therefore, three experiments were designed. In the first experiment, it was verified that the group treated with TSA for 5nM for 144h during in vitro culture (IVC) increased (p <0.01) the level of H3K9ac in the control group. Cleavage rate and embryo development were evaluated in a second experiment, and the group treated with 5nM TSA showed no difference in embryo production compared to control. In Experiment 3, treatment with TSA had no significant effect on the cell number of ICM, but decreased (p <0.01) both the cell number of TE and total cells as compared to control. Therefore, it is concluded that treatment with TSA does not alter the preimplantation embryonic development female bovine embryos, showing a negative effect on the development of embryos treated
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Silva, Bruna Mazeti. "Efeitos do agente modificador epigenético (Tricostatina A) no desenvolvimento embrionário pré-implantacional de embriões fêmeas de bovinos /." Jaboticabal : [s.n.], 2011. http://hdl.handle.net/11449/92589.
Повний текст джерелаАнотація:
Resumo: A acetilação das histonas é um dos principais mecanismos de reprogramação epigenética do genoma dos gametas a fim de estabelecer um estado totipotente para o desenvolvimento normal. O objetivo deste trabalho foi avaliar o efeito da utilização da Tricostatina A (TSA) sobre os níveis de acetilação das histonas e o desenvolvimento embrionário, avaliando-se a contagem do número de células da massa celular interna (MCI), trofectoderma (TE), e células totais, e os níveis da reação de imunocitoquímica para H3K9ac. Para isso, três experimentos foram delineados. No primeiro experimento, verificou-se que o grupo tratado com 5nM de TSA por 144h durante o cultivo in vitro (CIV) aumentou (p<0,01) o nível de H3K9ac em relação ao grupo controle. A taxa de clivagem e o desenvolvimento embrionário foram avaliados em um segundo experimento, e o grupo tratado com 5nM de TSA não apresentou diferença na produção de embriões comparado ao grupo controle. No experimento 3, o tratamento com a TSA não apresentou efeitos significativos em relação ao número de células da MCI, porém houve redução (p<0,01) tanto do número de células da TE quanto de células totais comparado ao grupo controle. Portanto, conclui-se que o tratamento com TSA não altera o desenvolvimento embrionário pré-implantacional de embriões fêmeas de bovinos, mostrando efeito negativo sobre o desenvolvimento dos embriões tratadosAbstract: Histone acetylation is one of the major mechanisms of epigenetic reprogramming of gamete genome to establish a totipotent state for normal development. The aim of this work was to evaluate the use of Trichostatin A (TSA) on the levels of histone acetylation and embryonic development, the assessment of counting the cell number of inner cell mass (ICM), trophectoderm (TE), and total cells, and the levels of H3K9ac for immunocytochemical reaction. Therefore, three experiments were designed. In the first experiment, it was verified that the group treated with TSA for 5nM for 144h during in vitro culture (IVC) increased (p <0.01) the level of H3K9ac in the control group. Cleavage rate and embryo development were evaluated in a second experiment, and the group treated with 5nM TSA showed no difference in embryo production compared to control. In Experiment 3, treatment with TSA had no significant effect on the cell number of ICM, but decreased (p <0.01) both the cell number of TE and total cells as compared to control. Therefore, it is concluded that treatment with TSA does not alter the preimplantation embryonic development female bovine embryos, showing a negative effect on the development of embryos treated
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Torres, Ana Catarina Belejo Mora. "Embryo-maternal interactions leading to embryonic development and survival in the bovine : role of progesterone and prostaglandins." Doctoral thesis, Universidade Técnica de Lisboa. Faculdade de Medicina Veterinária, 2012. http://hdl.handle.net/10400.5/5230.
Повний текст джерелаАнотація:
Tese de Doutoramento em Ciências Veterinárias. Especialidade de ClínicaThe objectives of this thesis were to evaluate steroidogenic and prostanoid embryo-maternal interactions leading to embryonic development and survival in cattle, and to evaluate therapeutic strategies at embryo transfer (ET) designed to enhance embryo survival. In vitro experiments (three experimental chapters) - bovine early (Day 7) embryos i) had transcription of genes coding for enzymes progesterone (P4) and of prostaglandins (PGs) synthesis pathways (StAR, P450scc,3β-HSD, PTGS2, PGFS, PGES); ii) produced these mediators (P4, PGF2α, PGE2) into culture medium; iii) had a significant increase in transcription levels of the above genes (except StAR) associated to first embryonic cellular differentiation; iv) derived from pre-pubertal oocyte donors had transcription levels of the above genes similar to those of embryos derived from post-pubertal cyclic heifers (except for 3β-HSD, which tended to be higher in embryos from cyclic heifers). Additionally, v) in a developed luteal cells (LC) co-culture model, LC induced an embryotrophic effect, significantly increasing blastocyst yield and quality; however, this embryotrophic effect was not associated with an increase in embryonic gene transcription or production of P4, PGF2α, PGE2; vi) Embryos co-cultured with LC did not exert a luteotrophic effect upon the cells; and vii) oil overlaying of culture wells exerted an embryotrophic effect, but absorbed P4 from culture medium. In vivo experiments (two experimental chapters) - novel in-vivo models considering poor developmental competence embryos (demi-embryos) and either sub-normal fertility recipients (lactating high-yielding dairy cow) or high fertility recipients (virgin dairy heifers) were used to evaluate the effect of hCG and carprofen treatment at embryo transfer on embryo survival and on plasma P4 and PSPB concentrations of recipients. Conclusions were that: i) treatment with hCG induced formation of secondary CL, increased plasma P4 concentrations, survival rate of demi-embryos and pregnancy rate of recipients (only in cows). Embryos were rescued beyond maternal recognition of pregnancy (MPR), but later embryonic survival, growth until implantation and placental PSPB secretion until Day 63 of pregnancy (only tested in cows) were not affected; ii) embryonic survival following MRP is not under direct dependency of maternal P4 concentrations; iii) treatment with carprofen had no significant effect on plasma P4 concentrations and embryonic survival, but decreased the luteotrophic effect of hCG.
RESUMO - Interações embrio-maternas relevantes para o desenvolvimento e sobrevivência embrionários em bovinos – papel da progesterona e das prostaglandinas - Os objetivos desta tese foram: avaliar interações embrio-maternas esteroidogénicas e prostanoides no desenvolvimento e sobrevivência embrionárias; testar estratégias terapêuticas na transferência embrionária (TE) com vista ao aumento da sobrevivência embrionária. Experiências in vitro (três capítulos experimentais) – embriões bovinos (Dia 7): i) revelaram transcrição de genes codificantes das enzimas das vias sintéticas da progesterona (P4) e PGs (PTGS2, PGFS, PGES, StAR, P450scc,3β-HSD); ii) produziram estes mediadores (P4, PGF2α, PGE2) para o meio de cultura; iii) apresentaram um aumento significativo dos níveis de transcrição destes genes (à exceção da StAR) associado à primeira diferenciação celular embrionária; iv) derivados de dadoras de oócitos pré-púberes revelaram níveis de transcrição dos genes mencionados similares aos de embriões de dadoras cíclicas (à exceção dos níveis de transcrição para a 3β-HSD, tendencialmente mais elevados em embriões provenientes de fêmeas cíclicas). Adicionalmente, v) num modelo de co-cultura de células lúteas desenvolvido, estas exerceram um efeito embriotrófico, aumentando significativamente a taxa de desenvolvimento e qualidade embrionárias; porém, este efeito não foi associado a aumento na transcrição génica ou produção de P4, PGF2α, PGE2; vi) Embriões em co-cultura com células lúteas não exerceram um efeito luteotrófico nas células; e vii) o uso de óleo mineral na cobertura dos poços de cultura exerceu um efeito embriotrófico, mas absorveu P4 do meio. Experiências in vivo (dois capítulos experimentais) – novos modelos in vivo - embriões de baixa competência de desenvolvimento (hemi-embriões) e recetoras sub-férteis (vacas leiteiras de alta produção) ou com fertilidade alta (novilhas leiteiras virgens) - foram usados na avaliação do efeito na sobrevivência embrionária e nas concentrações plasmáticas de P4 e PSPB das recetoras, de tratamentos, na TE, com hCG ou carprofen. Concluiu-se que: i) o tratamento com hCG induziu a formação de CLs secundários, aumentou as concentrações plasmáticas de P4, a taxa de sobrevivência dos hemi-embriões e as taxas de gestação das recetoras (em vacas). Os embriões foram resgatados para além do reconhecimento materno da gestação (RMG), mas a sobrevivência embrionária posterior, o crescimento até à implantação e a secreção placentária de PSPB até ao Dia 63 de gestação (testados em vacas) não foram afetados; ii) a sobrevivência embrionária após o RMG não está diretamente dependente das concentrações de P4 maternas; iii) o tratamento com o carprofeno não afetou significativamente as concentrações de P4 ou a sobrevivência embrionária, mas diminuiu o efeito luteotrófico da hCG.
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Abd-Elmaksoud, Ahmed. "Morphological, Glycohistochemical, and Immunohistochemical Studies on the Embryonic and Adult Bovine Testis." Diss., lmu, 2005. http://nbn-resolving.de/urn:nbn:de:bvb:19-39786.
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Hempstock, Joanne. "The role of recombinant trophoblast interferons in embryonic mortality in ruminants." Thesis, University of Reading, 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.307800.
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Walters, Anneke H. "In vitro assessment of fertilization and embryo development with Bovine spermatozoa after scrotal insulation." Diss., Virginia Tech, 2004. http://hdl.handle.net/10919/29721.
Повний текст джерелаАнотація:
Fertilization and cleavage of bovine embryos depend not only on maternal involvement, but also on the paternal contributions that involve more than just providing the haploid male genome. Therefore, the overall objective of this project was to determine the impact of morphologically abnormal spermatozoa on fertilization, subsequent embryonic development, and embryo quality at the cellular level. Four experiments used morphologically abnormal semen samples collected and cryopreserved from four Holstein bulls before (Pre) and after a scrotal insulation (PI) period of 48 h. Zygotes were cultured for 8 d when a developmental score was assigned to each embryo; subpopulations were subjected to either the TUNEL or caspase assays to determine apoptosis. In the final experiment pronuclear decondensation for presumptive zygotes was evaluated by differential interference contrast microscopy at 3 h time intervals from 6 to 18 h post in vitro insemination (hpi). Morphological evaluation of semen samples revealed a decrease (P < 0.01) in the percentages of normal spermatozoa in the PI samples in comparison with the Pre samples for Bulls I and Bull III (74 to 22.3% and 67.7 to 0.5 %, respectively) and the scrotal insulation effects persisted from the time of cleavage through blastocyst formation for Bulls I and III and corresponded with a similar decrease in blastocyst development for PI samples in experiment 1 regardless of which semen separation method was used. Likewise, the overall pronuclear decondensation rate for the PI zygotes of Bull I and III showed no increase over time and remained predominantly at PN1 stage (1.5 ± 0.17; 1.8 ± 0.22, respectively). In contrast, the development for Bull II and Bull IV were unaffected. The embryo quality assessment revealed that the caspase intensity increased significantly for both Bull I (217 ± 147) and Bull III (229 ± 98) for the PI embryo groups compared to those of Bull II (98 ± 115) and Bull IV (90 ± 111). In conclusion, the tested separation methods used seemed inadequate in their ability to provide potentially competent sperm for IVF. The decrease in embryonic development appears to be multifaceted and related to the changes in head shape morphology and we suggest the failure in normal pronuclear formation is associated with an absence of normal decondensation of the penetrating spermatozoon. The inability to consistently measure apoptosis in early stage embryos complicates the assessment of differences in embryo quality. These observations support the hypothesis of uncompensable seminal traits in IVF with abnormal spermatozoa and provide compelling evidence that the effect of morphologically abnormal spermatozoa occurred prior to cleavage, thus is manifested during the early stages of fertilization.Ph. D.
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Jousan, Frank Dean. "Insulin-like growth factor-I and apoptosis as determinants of preimplantation bovine embryonic development." [Gainesville, Fla.] : University of Florida, 2006. http://purl.fcla.edu/fcla/etd/UFE0015320.
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Villafranca, Locher Maria Cristina. "Fusion of bovine fibroblasts to mouse embryonic stem cells: a model to study nuclear reprogramming." Diss., Virginia Tech, 2018. http://hdl.handle.net/10919/82864.
Повний текст джерелаАнотація:
The cells from the inner cell mass (ICM) of an early embryo have the potential to differentiate into all the different cell types present in an adult organism. Cells from the ICM can be isolated and cultured in vitro, becoming embryonic stem cells (ESCs). ESCs have several properties that make them unique: they are unspecialized, can self-renew indefinitely in culture, and given the appropriate cues can differentiate into cells from all three germ layers (ecto-, meso-, and endoderm), including the germline, both in vivo and in vitro. Induced pluripotent stem cells (iPSCs) can be generated from adult, terminally differentiated somatic cells by transient exogenous expression of four transcription factors (Oct4, Sox2, Klf4, and cMyc; OSKM) present normally in ESCs. It has been shown that iPSCs are equivalent to ESCs in terms of morphology, gene expression, epigenetic signatures, in vitro proliferation capacity, and in vitro and in vivo differentiation potential. However, unlike ESCs, iPSCs can be obtained from a specific individual without the need for embryos. This makes them a promising source of pluripotent cells for regenerative medicine, tissue engineering, drug discovery, and disease modelling; additionally, in livestock species such as the bovine, they also have applications in genetic selection, production of transgenic animals for agricultural and biomedical purposes, and species conservancy. Nevertheless, ESC and iPSC lines that meet all pluripotency criteria have, to date, only been successfully produced in mice, rats, humans, and non-human primates.
In the first part of this dissertation, we attempted reprogramming of three types of bovine somatic cells: fetal fibroblasts (bFFs), adult fibroblasts (bAFs), and bone marrow-derived mesenchymal stem cells (bMSCs), using six different culture conditions adapted from recent work in mice and humans. Using basic mouse reprogramming conditions, we did not succeed in inducing formation of ESC-like colonies in bovine somatic cells. The combination of 2i/LIF plus ALK5 inhibitor II and ascorbic acid, induced formation of colony-like structures with flat morphology, that occasionally produced trophoblast-like structures. These trophoblast-like vesicles did not appear when an inhibitor of Rho-associated, coiled-coil containing protein kinase 1 (ROCK) was included in the medium. We screened for expression of exogenous OSKM vector with RT-PCR and found upregulation of OSKM vector 24h after Dox was added to the medium; however, expression was sharply decreased on day 2 after Dox induction, and was not detectable after day 3. In a separate experiment, we induced reprogramming of bFF and bAFs using medium supplemented with 50% of medium conditioned by co-culture with the bovine trophoblast CT1 line. These cells expressed both OCT4 and the OSKM vector 24h after Dox induction. However, similar to our previous observations, both markers decreased expression until no signal was detected after day 3. In summary, we were unable to produce fully reprogrammed bovine iPSCs using mouse and human protocols, and the exact cause of our lack of success is unclear. It is possible that a different method of transgene expression could play a role in reprogramming. However, these ideas would be driven by a rather empirical reasoning, extrapolating findings from other species, and not contributing in our understanding of the particular differences of pluripotecy in ungulates. Our inability to produce bovine iPSCs, combined with the only partial reprogramming observed by others, justifies the need for in depth study of bovine pluripotency mechanisms, before meaningful attempts to reprogram bovine somatic cells to plutipotency are made. Therefore, we focused on getting a better understanding of bovine nuclear reprogramming. This would allow us to rationally target the specific requirements of potential bovine pluripotent cells.
Cell fusion is a process that involves fusion of the membrane of two or more cells to form a multinucleated cell. Fusion of a somatic cell to an ESC is known to induce expression of pluripotency markers in the somatic nucleus. In the second part of this dissertation, we hypothesized that fusion of bFFs to mouse ESCs (mESCs) would induce expression of pluripotency markers in the bFF nucleus. We first optimized a cell fusion protocol based on the use of polyethylene glycol (PEG), and obtained up to 11.02% of multinucleated cells in bFFs. Next, we established a method to specifically select for multinucleated cells originated from the fusion of mESCs with bFFs (heterokaryons), using indirect immunofluorescence. With this in place, flow cytometry was used to select 200 heterokaryons which were further analyzed using RNA-seq. We found changes in bovine gene expression patterns between bFFs and heterokaryons obtained 24h after fusion. Focusing on the bovine transcriptome, heterokaryons presented upregulation of early pluripotency markers OCT4 and KLF4, as well as hypoxia response genes, contrasted with downregulation of cell cycle inhibitors such as SST. The cytokine IL6, known to increase survival of early embryos in vitro, was upregulated in heterokaryons, although its role and mechanism of action is still unclear. This indicates that the heterokaryon cell fusion model recapitulates several of the events of early reprogramming, and can therefore be used for further study of pluripotency in the bovine. The cell fusion model presented here can be used as a tool to characterize early changes in bovine somatic nuclear reprogramming, and to study the effect of different reprogramming conditions on the bovine transcriptome.Ph. D.
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Larsen, Davin M. "Evaluation of the TGF-ß Inhibitor RepSox on the Expression of Pluripotency Pathways in Murine and Bovine Cells." DigitalCommons@USU, 2013. http://digitalcommons.usu.edu/etd/1509.
Повний текст джерелаАнотація:
Embryonic stem cells are pluripotent cells isolated from morula stage embryos or the inner cell mass of blastocyst stage embryos. They are capable of differentiating into tissues of all three primary germ layers. In recent years pluripotent cell lines have been created from somatic cell types using various methods, the primary method being viral transduction of exogenous Oct4, Sox2, Klf4, and c-Myc or Oct4, Sox2, Nanog, and Lin28 transgene constructs. The resulting cell lines are termed induced pluripotency stem cells, and are similar to embryonic stem cells in many ways. However, these cell lines are not acceptable for clinical applications due to the use of both modified viral vectors and insertion of exogenous transgenes in their production. Recently the small molecule RepSox, a TGF-ß pathway inhibitor, was used to replace Sox2 during cellular reprogramming of murine embryonic fibroblasts. We evaluated the effects of RepSox on expression of pathways related to pluripotency in murine embryonic fibroblast, murine embryonic stem, and bovine embryonic fibroblast cells. Each cell type was treated with RepSox for 72 hours and subjected to standard qPCR for gene expression analysis. PCR arrays specific to stem cell pathways were used to initially evaluate the effects of RepSox on candidate genes. A subset of genes was then selected for further analysis based on these initial results. We report that RepSox inhibition of the TGF-ß pathway in murine embryonic fibroblasts results in significant upregulation of components of the Wnt, Notch, and Hedgehog signaling pathways, all of which have been linked to stem cell maintenance. In addition, we observed significant upregulation of genes associated with embryonic, mesenchymal, stem cell, and neural cell lineages, indicating that RepSox may be useful in direct reprogramming of murine cells to other somatic cell types. RepSox treatment of murine embryonic stem cells did not result in consistent upregulation of Wnt, Notch, or Hedgehog pathway components, but did result in upregulation of Sox2 and Klf4 expression. Lastly, RepSox treatment of bovine embryonic fibroblasts did not result in the same effects as seen in murine fibroblasts, indicating a need for further analysis to determine the effects of RepSox on bovine cells.
Книги з теми "Bovine embryonic"
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Shamsuddin, Mohammed. In vitro fertilization in the bovine: Oocyte maturation, fertilization and early embryonic development. Uppsala: Sveriges Lantbruksuniversitet, 1993.
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Albihn, Ann. Maternal influence on the early embryonic development in the bovine: With special emphasis on repeat breeder heifers. Uppsala: Sveriges Lantbruksuniversitet, 1991.
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Elsden, R. P. Manual for embryo transfer. Hastings, Neb: Society for Theriogenology, 1987.
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Rief, Stephanie. Etablierung eines Kokultursystems von bovinen Präimplantationsembryonen mit Eileiterepithelzellen vom Rind sowie dessen Einfluss auf Entwicklung, Metabolismus und Genexpression der Embryonen. München: Hieronymus, 2001.
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Berg, Debra Ann. Identification of the plasminogen activator produced by microdissected day 12 to 14 bovine embryonic tissues. 1990.
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Cannon, Matthew Joel. Evaluation of alterations in the bovine zona pellucida induced by plasmin or embryonic plasminogen activator. 1995.
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Hozab, Adel Abdulla. The effects of hormones and inducers of intracellular messengers on bovine embryo development in vitro: Plasminogen activator production and changes in embryonic size. 1990.
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Seidel, George E. Jr. Embryo Transfer in Dairy Cattle. 2nd ed. W D Hoard & Sons Co, 1997.
Знайти повний текст джерелаЧастини книг з теми "Bovine embryonic"
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Hansen, Peter J. "Early Embryonic Loss Due to Heat Stress." In Bovine Reproduction, 580–88. Hoboken, NJ, USA: John Wiley & Sons, Inc., 2014. http://dx.doi.org/10.1002/9781118833971.ch64.
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Hernández, Aureliano. "Embryonic Survival and Mortality." In Bovine Maternal Support and Embryo Survival, 113–25. Cham: Springer Nature Switzerland, 2024. http://dx.doi.org/10.1007/978-3-031-62391-2_11.
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Mannaerts, B. M. J. L. "Cytological Parameters for Rating Bovine Embryo Quality." In Embryonic Mortality in Farm Animals, 216–22. Dordrecht: Springer Netherlands, 1986. http://dx.doi.org/10.1007/978-94-009-5038-2_18.
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Cao, Shanbo, Fang Wang, and Lin Liu. "Isolation and Culture of Bovine Embryonic Stem Cells." In Epiblast Stem Cells, 111–23. Totowa, NJ: Humana Press, 2013. http://dx.doi.org/10.1007/978-1-62703-628-3_9.
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Hernández, Aureliano. "First Stages of Embryonic Development, Histogenesis of the Placenta, and Pregnancy Maintenance." In Bovine Maternal Support and Embryo Survival, 63–112. Cham: Springer Nature Switzerland, 2024. http://dx.doi.org/10.1007/978-3-031-62391-2_10.
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Watson, Andrew J., Aileen Hogan, Ann Hahnel, and Gilbert A. Schultz. "Activation of the Embryonic Genome: Comparisons Between Mouse and Bovine Development." In Preimplantation Embryo Development, 115–30. New York, NY: Springer New York, 1993. http://dx.doi.org/10.1007/978-1-4613-9317-7_9.
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Soto, Delia A., Micaela Navarro, and Pablo J. Ross. "Derivation of Bovine Primed Embryonic Stem Cells from Somatic Cell Nuclear Transfer Embryos." In Methods in Molecular Biology, 305–15. New York, NY: Springer US, 2023. http://dx.doi.org/10.1007/978-1-0716-3064-8_17.
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Roach, Marsha, Li Wang, Xiangzhong Yang, and X. Cindy Tian. "Bovine Embryonic Stem Cells." In Methods in Enzymology, 21–37. Elsevier, 2006. http://dx.doi.org/10.1016/s0076-6879(06)18002-7.
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Valadão, Loide, Helena Moreira da Silva, and Fernando Moreira da Silva. "Bovine Embryonic Development to Implantation." In Embryology - Theory and Practice. IntechOpen, 2019. http://dx.doi.org/10.5772/intechopen.80655.
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Gokhale, Golden, and Guru Dutt Sharma. "Adverse Impact of Heat Stress on Bovine Development: Causes and Strategies for Mitigation." In Bovine Science [Working Title]. IntechOpen, 2021. http://dx.doi.org/10.5772/intechopen.99307.
Повний текст джерелаАнотація:
Heat stress induces the richness and reproductive domesticated animal’s performance by settling the physiology conceptive steps, through hormonal irregularity, diminished oocyte quality and feeble semen quality, and diminished undeveloped organism advancement and endurance. It depends on principally milk production, nutrition, disease management, sexual activities, and heat stress tolerance capacity in livestock farming. The decreases infertility caused by elevated blood heat influences sex gland regulation, oestrus regulation, and gametocyte disturbance and also affects embryonic development. Heat stress reduces the degree of dominance of the seminal vesicles and this may be observed as reduced steroidogenic capability of its theca and granulose cells as fall in blood oestrogen concentrations. Plasma progestin levels are also diminished counting on whether or not the heat stress is acute and on the metabolic state of the animal. The endocrine changes the cyst activities and alters the ovulatory mechanism leading to a decrease in gametocyte and embryo quality. Summer infertility may be countered through oestrus behaviour can be mitigated by with the help of implementation of ovulation phase treatments to limited period of embryonic transfer and also advanced reproductive technologies involving hormonal treatments, systematic artificial insemination and which may enhance the possibility of establishing pregnancy in domestic animals.
Тези доповідей конференцій з теми "Bovine embryonic"
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Daleffi, Natalia Soriani, Marcella Pecora Milazzotto, and Fernanda Nascimento Almeida. "Modeling and Implementation of a Web Database for RNA-Seq of Bovine Embryonic Cells." In Brazilian e-Science Workshop. Sociedade Brasileira de Computação - SBC, 2023. http://dx.doi.org/10.5753/bresci.2023.234243.
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This paper aims to develop a dedicated RNA-Seq database for bovine embryos, generated to gain insights into reproductive metabolism. The data is categorized into three groups, each obtained from distinct experiments. The primary objective is to streamline data analysis through a platform, named TranscriptomicsSeqDB, which standardizes and organizes RNA-Seq information from the Laboratory of Embryonic Metabolism and Epigenetics at UFABC, São Paulo, Brazil. Apart from data storage and management, TranscriptomicsSeqDB provides a user-friendly search interface with predefined queries to facilitate gene-specific indicator analysis.
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Crouse, M. S., J. S. Caton, K. J. Claycombe-Larson, C. R. Dahlen, L. P. Reynolds, P. P. Borowicz, and A. K. Ward. "Effects of glucose and one-carbon metabolites on mitochondrial respiration in bovine embryonic cells." In 6th EAAP International Symposium on Energy and Protein Metabolism and Nutrition. The Netherlands: Wageningen Academic Publishers, 2019. http://dx.doi.org/10.3920/978-90-8686-891-9_53.
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Koteneva, S. V., A. V. Nefedchenko, T. I. Glotova, and A. G. Glotov. "Development of a multiplex real-time PCR assay for the detection of three species of bovine pestiviruses." In БИОТЕХНОЛОГИЯ: НАУЧНЫЕ ИССЛЕДОВАНИЯ И СВЯЗЬ С ПРОИЗВОДСТВОМ, 120–25. Всероссийский научно-исследовательский и технологический институт биологической промышленности, 2024. https://doi.org/10.47804/978-5-89904-038-2-2024-120-125.
Повний текст джерелаАнотація:
The article presents the results of development of multiplex polymerase chain reaction in real time for detection and differentiation of three species of bovine pestiviruses (Pestivirus A, B and H). Synthetic oligonucleotide primers and probes complementary to the highly conserved 5/-UTR region of the genome, specific for all species of the genus Pestivirus and for each species separately were selected. The main parameters of the reaction were worked out. The analytical sensitivity of the developed real-time PCR was 1,6×102 GE for Pestivirus spp., 7,2×10 GE for Pestivirus A, 9,0×10 GE for Pestivirus B and 1.5×103 GE for Pestivirus H. PCR has high specificity and does not detect RNA of other viruses. Using the developed method, 18 samples of embryonic serum, 12 types of continuous cell culture lines, 10 series of live vaccines for immunization of cattle, 3050 samples of biomaterial from sick animals were studied. As a result of the studies, the Pestivirus H genome was detected in 7 samples of embryonic serum, 18 samples of internal organs of calves and 1 live vaccine. Pestivirus A and Pestivirus B RNA were detected in 173 and 5 samples of biomaterial from animals, 2 and 1 samples of continuous cell lines, respectively. Multiplex real-time PCR can be used for express diagnostics of cattle diseases caused by pestiviruses and control of biological products.
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Knocker, Louisa, and Zheng Rong Yang. "SLC- and NDUF-genes expression dynamics in pre-implantation embryonic development between bovine and mouse — A bioinformatics study." In 2014 7th International Conference on Biomedical Engineering and Informatics (BMEI). IEEE, 2014. http://dx.doi.org/10.1109/bmei.2014.7002857.
Повний текст джерелаЗвіти організацій з теми "Bovine embryonic"
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Hansen, Peter J., and Zvi Roth. Use of Oocyte and Embryo Survival Factors to Enhance Fertility of Heat-stressed Dairy Cattle. United States Department of Agriculture, August 2011. http://dx.doi.org/10.32747/2011.7697105.bard.
Повний текст джерелаАнотація:
The overall goal was to identify survival factors that can improve pregnancy success following insemination or embryo transfer in lactating dairy cows exposed to heat stress. First, we demonstrated that oocytes are actually damaged by elevated temperature in the summer. Then we tested two thermoprotective molecules for their effect on oocyte damage caused by heat shock. One molecule, ceramide was not thermoprptective. Another, insulin-like growth factor-1 (IGF) reduced the effects of heat shock on oocyte apoptosis and oocyte cleavage when added during maturation. We also used lactating cows exposed to heat stress to determine whether bovine somatotropin (bST), which increases IGF1 levels in vivo, would improve fertility in summer. Cows treated with bST received a single injection at 3 days before insemination. Controls received no additional treatment. Treatment with bST did not significantly increase the proportion of inseminated cows diagnosed pregnant although it was numerically greater for the bST group (24.2% vs 17.8%, 124–132 cows per group). There was a tendency (p =0.10) for a smaller percent of control cows to have high plasma progesterone concentrations (≥ 1 ng/ml) at Day 7 after insemination than for bST-treated cows (72.6 vs 81.1%). When only cows that were successfully synchronized were considered, the magnitude of the absolute difference in the percentage of inseminated cows that were diagnosed pregnant between bST and control cows was reduced (24.8 vs 22.4% pregnant for bST and control). Results failed to indicate a beneficial effect of bST treatment on fertility of lactating dairy cows. In another experiment, we found a tendency for addition of IGF1 to embryo culture medium to improve embryonic survival after embryo transfer when the experiment was done during heat stress but not when the experiment was done in the absence of heat stress. Another molecule tested, granulocyte-macrophage colony-stimulating factor (GM-CSF; also called colony-stimulating factor-2), improved embryonic survival in the absence of heat stress. We also examined whether heat shock affects the sperm cell. There was no effect of heat shock on sperm apoptosis (programmed cell death) or on sperm fertilizing ability. Therefore, effects of heat shock on sperm function after ejaculation if minimal. However, there were seasonal changes in sperm characteristics that indicates that some of the decrease in dairy cow fertility during the summer in Israel is due to using semen of inferior quality. Semen was collected from five representative bulls throughout the summer (August and September) and winter (December and January). There were seasonal differences in ion concentration in seminal plasma and in the mRNA for various ion channels known to be involved in acrosome reactions. Furthermore, the proportion of sperm cells with damaged acrosomes was higher in post-thaw semen collected in the summer than in its counterpart collected in winter (54.2 ± 3.5% vs. 51.4 ± 1.9%, respectively; P < 0.08Further examination is required to determine whether such alterations are involved in the low summer fertility of dairy cows.
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Hansen, Peter J., and Amir Arav. Embryo transfer as a tool for improving fertility of heat-stressed dairy cattle. United States Department of Agriculture, September 2007. http://dx.doi.org/10.32747/2007.7587730.bard.
Повний текст джерелаАнотація:
The overall objective of the current proposal is to develop procedures to improve the pregnancy rate achieved following transfer of fresh or cryopreserved embryos produced in the laboratory into heat-stress recipients. The overall hypothesis is that pregnancy rate in heat-stressed lactating cows can be improved by use of embryo transfer and that additional gains in pregnancy rate can be achieved through development of procedures to cryopreserve embryos, select embryos most likely to establish and maintain pregnancy after transfer, and to enhance embryo competence for post-transfer survival through manipulation of culture conditions. The original specific objectives were to 1) optimize procedures for cryopreservation (Israel/US), 2) develop procedures for identifying embryos with the greatest potential for development and survival using the remote monitoring system called EmbryoGuard (Israel), 3) perform field trials to test the efficacy of cryopreservation and the EmbryoGuard selection system for improving pregnancy rates in heat-stressed, lactating cows (US/Israel), 4) test whether selection of fresh or frozen-thawed blastocysts based on measurement of group II caspase activity is an effective means of increasing survival after cryopreservation and post-transfer pregnancy rate (US), and 5) identify genes in blastocysts induced by insulin-like growth factor-1 (IGF-1) (US). In addition to these objectives, additional work was carried out to determine additional cellular determinants of embryonic resistance to heat shock. There were several major achievements. Results of one experiment indicated that survival of embryos to freezing could be improved by treating embryos with cytochalasin B to disrupt the cytoskeleton. An additional improvement in the efficacy of embryo transfer for achieving pregnancy in heat-stressed cows follows from the finding that IGF-1 can improve post-transfer survival of in vitro produced embryos in the summer but not winter. Expression of several genes in the blastocyst was regulated by IGF-1 including IGF binding protein-3, desmocollin II, Na/K ATPase, Bax, heat shock protein 70 and IGF-1 receptor. These genes are likely candidates 1) for developing assays for selection of embryos for transfer and 2) as marker genes for improving culture conditions for embryo production. The fact that IGF-1 improved survival of embryos in heat-stressed recipients only is consistent with the hypothesis that IGF-1 confers cellular thermotolerance to bovine embryos. Other experiments confirmed this action of IGF-1. One action of IGF-1, the ability to block heat-shock induced apoptosis, was shown to be mediated through activation of the phosphatidylinositol 3-kinase pathway. Other cellular determinants of resistance of embryos to elevated temperature were identified including redox status of the embryo and the ceramide signaling pathway. Developmental changes in embryonic apoptosis responses in response to heat shock were described and found to include alterations in the capacity of the embryo to undergo caspase-9 and caspase-3 activation as well as events downstream from caspase-3 activation. With the exception of IGF-1, other possible treatments to improve pregnancy rate to embryo transfer were not effective including selection of embryos for caspase activity, treatment of recipients with GnRH.and bilateral transfer of twin embryos. In conclusion, accomplishments achieved during the grant period have resulted in methods for improving post-transfer survival of in vitro produced embryos transferred into heat-stressed cows and have lead to additional avenues for research to increase embryo resistance to elevated temperature and improve survival to cryopreservation. In addition, embryo transfer of vitrified IVF embryos increased significantly the pregnancy rate in repeated breeder cows.