Equine cloning: in vitro and in vivo development of aggregated embryos.
Abstract: The production of cloned equine embryos remains highly inefficient. Embryo aggregation has not yet been tested in the equine, and it might represent an interesting strategy to improve embryo development. This study evaluated the effect of cloned embryo aggregation on in vitro and in vivo equine embryo development. Zona-free reconstructed embryos were individually cultured in microwells (nonaggregated group) or as 2- or 3-embryo aggregates (aggregated groups). For in vitro development, they were cultured until blastocyst stage and then either fixed for Oct-4 immunocytochemical staining or maintained in in vitro culture where blastocyst expansion was measured daily until Day 17 or the day on which they collapsed. For in vivo assays, Day 7-8 blastocysts were transferred to synchronized mares and resultant vesicles, and cloned embryos were measured by ultrasonography. Embryo aggregation improved blastocyst rates on a per well basis, and aggregation did not imply additional oocytes to obtain blastocysts. Embryo aggregation improved embryo quality, nevertheless it did not affect Day 8 and Day 16 blastocyst Oct-4 expression patterns. Equine cloned blastocysts expanded and increased their cell numbers when they were maintained in in vitro culture, describing a particular pattern of embryo growth that was unexpectedly independent of embryo aggregation, as all embryos reached similar size after Day 7. Early pregnancy rates were higher using blastocysts derived from aggregated embryos, and advanced pregnancies as live healthy foals also resulted from aggregated embryos. These results indicate that the strategy of aggregating embryos can improve their development, supporting the establishment of equine cloned pregnancies.
Publication Date: 2012-07-19 PubMed ID: 22553223DOI: 10.1095/biolreprod.112.098855Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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The research article is about a study that investigated the possible benefits of aggregating cloned equine embryos on their in vitro and in vivo development. The results indicated that this strategy may enhance their development and support the establishment of equine cloned pregnancies.
Research Methodology
- The researchers divided zona-free reconstructed embryos into two groups. One group of embryos were individually cultured in microwells (nonaggregated group), while the other group was made up of 2- or 3-embryo aggregates (aggregated groups).
- For in vitro development, they cultured these embryos until they reached the blastocyst stage. At this point, the embryos were either fixed for Oct-4 immunocytochemical staining or kept in in vitro culture. Here, researchers took daily measurements of blastocyst expansion until Day 17 or the day on which the embryos collapsed.
- For in vivo assays, researchers transferred Day 7-8 blastocysts to synchronized mares. They then measured resultant vesicles and cloned embryos by ultrasonography.
Findings and Conclusions
- The results indicated that embryo aggregation led to improved blastocyst rates on a per well basis. It was also found that aggregation did not require additional oocytes to obtain blastocysts.
- Aggregation enhanced embryo quality, but it did not affect the expression patterns of Oct-4 in the Day 8 and Day 16 blastocysts.
- Researchers also found that equine cloned blastocysts that were kept in in vitro culture expanded and increased their cell numbers. This process described an unusual pattern of embryo growth that was independent of embryo aggregation, as all embryos reached comparable size after Day 7.
- The study showed that early pregnancy rates were higher using blastocysts derived from aggregated embryos. Aggregated embryos also resulted in advanced pregnancies and live healthy foals.
- These findings imply that embryo aggregation can be a potential strategy to improve the development of cloned equine embryos and support the establishment of equine cloned pregnancies.
Cite This Article
APA
Gambini A, Jarazo J, Olivera R, Salamone DF.
(2012).
Equine cloning: in vitro and in vivo development of aggregated embryos.
Biol Reprod, 87(1), 15-9.
https://doi.org/10.1095/biolreprod.112.098855 Publication
Researcher Affiliations
- Laboratory of Animal Biotechnology, Agriculture Faculty, University of Buenos Aires, Buenos Aires, Argentina.
MeSH Terms
- Animals
- Animals, Newborn
- Blastocyst / cytology
- Blastocyst / metabolism
- Cell Aggregation
- Cloning, Organism / methods
- Cloning, Organism / veterinary
- Embryo Culture Techniques / veterinary
- Embryo Transfer / veterinary
- Embryonic Development
- Female
- Horses / metabolism
- Male
- Mice
- Octamer Transcription Factor-3 / metabolism
- Parthenogenesis
- Pregnancy
- Pregnancy Outcome / veterinary
Citations
This article has been cited 10 times.- Salamone D, Maserati M. Horse Somatic Cell Nuclear Transfer Using Zona Pellucida-Enclosed and Zona-Free Oocytes. Methods Mol Biol 2023;2647:269-281.
- Hisey EA, Ross PJ, Meyers S. Genetic Manipulation of the Equine Oocyte and Embryo. J Equine Vet Sci 2021 Apr;99:103394.
- Hisey E, Ross PJ, Meyers SA. A Review of OCT4 Functions and Applications to Equine Embryos. J Equine Vet Sci 2021 Mar;98:103364.
- Gambini A, Duque Rodríguez M, Rodríguez MB, Briski O, Flores Bragulat AP, Demergassi N, Losinno L, Salamone DF. Horse ooplasm supports in vitro preimplantation development of zebra ICSI and SCNT embryos without compromising YAP1 and SOX2 expression pattern. PLoS One 2020;15(9):e0238948.
- Xu L, Mesalam A, Lee KL, Song SH, Khan I, Chowdhury MMR, Lv W, Kong IK. Improves the In Vitro Developmental Competence and Reprogramming Efficiency of Cloned Bovine Embryos by Additional Complimentary Cytoplasm. Cell Reprogram 2019 Feb;21(1):51-60.
- Olivera R, Moro LN, Jordan R, Pallarols N, Guglielminetti A, Luzzani C, Miriuka SG, Vichera G. Bone marrow mesenchymal stem cells as nuclear donors improve viability and health of cloned horses. Stem Cells Cloning 2018;11:13-22.
- Olivera R, Moro LN, Jordan R, Luzzani C, Miriuka S, Radrizzani M, Donadeu FX, Vichera G. In Vitro and In Vivo Development of Horse Cloned Embryos Generated with iPSCs, Mesenchymal Stromal Cells and Fetal or Adult Fibroblasts as Nuclear Donors. PLoS One 2016;11(10):e0164049.
- Buemo CP, Gambini A, Moro LN, Hiriart MI, Fernández-Martín R, Collas P, Salamone DF. Embryo Aggregation in Pig Improves Cloning Efficiency and Embryo Quality. PLoS One 2016;11(2):e0146390.
- Gambini A, De Stefano A, Bevacqua RJ, Karlanian F, Salamone DF. The aggregation of four reconstructed zygotes is the limit to improve the developmental competence of cloned equine embryos. PLoS One 2014;9(11):e110998.
- Veraguas-Dávila D, Zapata-Rojas C, Aguilera C, Saéz-Ruiz D, Saravia F, Castro FO, Rodriguez-Alvarez L. Proteomic Analysis of Domestic Cat Blastocysts and Their Secretome Produced in an In Vitro Culture System without the Presence of the Zona Pellucida. Int J Mol Sci 2024 Apr 14;25(8).
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