In vitro-produced equine embryos: production of foals after transfer, assessment by differential staining and effect of medium calcium concentrations during culture.
Abstract: Viability of equine embryos produced by oocyte maturation, intracytoplasmic sperm injection and embryo culture to the blastocyst stage in vitro was evaluated after transfer of embryos to recipient mares. No pregnancies were produced after transfer of five blastocysts that had been cultured in G media. Transfer of 10 blastocysts cultured in modified DMEM/F-12 medium produced five pregnancies and three live foals; the two lost pregnancies developed only trophoblast (based on transrectal ultrasonography). To evaluate the status of the inner cell mass, equine blastocysts produced in vivo and in vitro were assessed after differential staining. A discrete inner cell mass could not be appreciated in blastocysts of either source after staining; this was attributed to the presence of a network of cells within the trophoblastic vesicle. Because increased medium calcium concentrations have been reported to decrease the incidence of trophoblast-only pregnancy after transfer of equine nuclear transfer embryos, we investigated the effect of increased calcium concentrations during oocyte maturation or during embryo culture. Increasing calcium concentration of culture medium from 2 to 5.6mM during in vitro oocyte maturation did not affect maturation rate (75 and 68%, respectively) or blastocyst development after fertilization (23 and 27%). However, increasing calcium concentration (from 1.3 to 4.9 mM) of medium used for embryo culture significantly decreased blastocyst development (27% versus 13%, respectively) and adversely affected embryo morphology. More work is needed to optimize culture systems for in vitro production of equine embryos.
Publication Date: 2007-06-21 PubMed ID: 17586036DOI: 10.1016/j.theriogenology.2007.04.046Google Scholar: Lookup
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- Comparative Study
- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This study focuses on the viability of horse embryos produced in a lab setting, and assesses their survivability after being transferred to horses. The results varied depending on the culture medium used for embryo creation, with better results seen when using a modified DMEM/F-12 medium. The study also investigated the impact of various calcium concentrations in the culture medium on embryo health.
Equine Embryo Production and Viability
- This research evaluated the viability of horse embryos produced through oocyte maturation, intracytoplasmic sperm injection, and in vitro (lab-based) embryo culture. Specifically, it observed their development until the blastocyst stage, followed by their transfer to recipient mares.
- The embryo viability was influenced by the culture medium used. No pregnancies resulted from embryos cultured in G media, but the transfer of a different set of embryos produced using a modified DMEM/F-12 medium resulted in five pregnancies and three live foals. Two of these pregnancies were lost and had only developed trophoblast (part of the blastocyst stage in early embryonic development).
Inner Cell Mass Assessment
- To gain a clearer understanding of the status of the inner cell mass of horse-sourced embryos, both in vivo (live) and in vitro-produced embryos were examined using differential staining. This staining process, however, did not reveal a distinct inner cell mass. The researchers concluded that this was due to the presence of a network of cells within the trophoblastic vesicle.
Impact of Calcium Concentration
- Earlier studies suggested that higher calcium concentrations in the culture medium could negatively impact the chances of successful, full-term pregnancies after transferring horse embryos. Therefore, this study further investigated the potential impact of varying calcium concentration levels during oocyte maturation and during embryo culture.
- When the scientists increased the calcium concentration in the maturation medium from 2mM to 5.6mM, it was found that neither the maturation rate nor the success of blastocyst development post-fertilization was affected.
- Yet, when the calcium concentration in the medium used for embryo development was increased (from 1.3mM to 4.9mM), there was a significant decrease in successful blastocyst development and the overall morphology of the embryo.
Conclusions and Avenues for Future Research
- The research concluded with the observation that additional work is required to optimize in vitro culture systems for the production of equine embryos.
Cite This Article
APA
Hinrichs K, Choi YH, Walckenaer BE, Varner DD, Hartman DL.
(2007).
In vitro-produced equine embryos: production of foals after transfer, assessment by differential staining and effect of medium calcium concentrations during culture.
Theriogenology, 68(4), 521-529.
https://doi.org/10.1016/j.theriogenology.2007.04.046 Publication
Researcher Affiliations
- Department of Veterinary Physiology and Pharmacology, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX 77843, USA. khinrichs@cvm.tamu.edu
MeSH Terms
- Animals
- Animals, Newborn
- Benzimidazoles / chemistry
- Blastocyst / cytology
- Blastocyst / drug effects
- Blastocyst / physiology
- Calcium / pharmacology
- Cell Culture Techniques
- Culture Media
- Embryo Transfer / veterinary
- Embryonic Development / drug effects
- Embryonic Development / physiology
- Female
- Fluorescent Dyes / chemistry
- Horses / embryology
- Horses / physiology
- Male
- Microscopy, Fluorescence
- Pregnancy
- Sperm Injections, Intracytoplasmic / veterinary
Citations
This article has been cited 7 times.- Siemieniuch-Tartanus M. The early pregnancy in mares - What do we still not know?. Vet Anim Sci 2025 Jun;28:100441.
- Brooks KE, Daughtry BL, Metcalf E, Masterson K, Battaglia D, Gao L, Park B, Chavez SL. Assessing equine embryo developmental competency by time-lapse image analysis. Reprod Fertil Dev 2019 Jan;31(12):1840-1850.
- Roberts MA, London K, Campos-Chillón LF, Altermatt JL. Presumed monozygotic twins develop following transfer of an in vitro-produced equine embryo. J Equine Sci 2015;26(3):89-94.
- Martino NA, Reshkin SJ, Ciani E, Dell'Aquila ME. Calcium-sensing receptor-mediated osteogenic and early-stage neurogenic differentiation in umbilical cord matrix mesenchymal stem cells from a large animal model. PLoS One 2014;9(11):e111533.
- Martino NA, Lange-Consiglio A, Cremonesi F, Valentini L, Caira M, Guaricci AC, Ambruosi B, Sciorsci RL, Lacalandra GM, Reshkin SJ, Dell'Aquila ME. Functional expression of the extracellular calcium sensing receptor (CaSR) in equine umbilical cord matrix size-sieved stem cells. PLoS One 2011 Mar 17;6(3):e17714.
- Smits K, Goossens K, Van Soom A, Govaere J, Hoogewijs M, Vanhaesebrouck E, Galli C, Colleoni S, Vandesompele J, Peelman L. Selection of reference genes for quantitative real-time PCR in equine in vivo and fresh and frozen-thawed in vitro blastocysts. BMC Res Notes 2009 Dec 11;2:246.
- Barfield JP, McCue PM, Squires EL, Seidel GE Jr. Effect of dehydration prior to cryopreservation of large equine embryos. Cryobiology 2009 Aug;59(1):36-41.
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