{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,24]],"date-time":"2026-03-24T14:53:20Z","timestamp":1774364000767,"version":"3.50.1"},"reference-count":61,"publisher":"MDPI AG","issue":"5","license":[{"start":{"date-parts":[[2021,5,6]],"date-time":"2021-05-06T00:00:00Z","timestamp":1620259200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Animals"],"abstract":"<jats:p>In embryos subjected to assisted reproductive techniques, epigenetic modifications may occur that can influence embryonic development and the establishment of pregnancy. In horses, the storage temperature during transport of fresh embryos before transfer is a major concern. The aim of this study was, therefore, to determine the effects of two storage temperatures (5 \u00b0C and 20 \u00b0C) on equine embryos, collected at day seven after ovulation and stored for 24 h, on: (i) morphological development; (ii) expression of candidate genes associated with embryo growth and development, maternal recognition of pregnancy, methylation and apoptosis, and (iii) gene-specific and global DNA methylation. Embryos (n = 80) were collected on day seven or day eight after ovulation and assigned to four groups: day seven control (E7F, fresh); day seven, stored for 24 h at 5 \u00b0C (E5C); day seven, stored for 24 h at 20 \u00b0C (E20C) and day eight control (E8F, fresh 24h time control). The embryos and the storage medium (EquiHold, holding medium, Minitube, Tiefenbach, Germany) from all treatment groups were analyzed for (i) medium temperature, pH, and lipid peroxidation (malondialdehyde; MDA) and (ii) embryo morphology, mRNA expression and DNA methylation (immunohistochemistry and gene-specific DNA methylation). The size of embryos stored at 5 \u00b0C was larger (p &lt; 0.01), whereas embryos stored at 20 \u00b0C were smaller (p &lt; 0.05) after 24 h. There were no changes in pH and MDA accumulation irrespective of the group. The mRNA expression of specific genes related to growth and development (POU5F1, SOX2, NANOG), maternal recognition of pregnancy (CYP19A1, PTGES2), DNA methylation (DNMT1, DNMT3A, DNMT3B) and apoptosis (BAX) in the E5C and E20C were either up or downregulated (p &lt; 0.05) when compared to controls (E7F and E8F). The immune expression of 5mC and 5hmC was similar among treatment groups. Percentage of methylation in the CpG islands was lower in the specific genes ESR1, NANOG and DNMT1 (p &lt; 0.001) in E20C embryos when compared to E8F (advanced embryo stage). Therefore, our study demonstrates for the first time the gene-specific and global DNA methylation status of fresh equine embryos collected on days seven and eight after ovulation. Although our results suggest some beneficial effects of storage at 20 \u00b0C in comparison to 5 \u00b0C, the short-term storage, regardless of temperature, modified gene expression and methylation of genes involved in embryo development and may compromise embryo viability and development after transfer.<\/jats:p>","DOI":"10.3390\/ani11051325","type":"journal-article","created":{"date-parts":[[2021,5,6]],"date-time":"2021-05-06T11:10:27Z","timestamp":1620299427000},"page":"1325","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Epigenetic Changes in Equine Embryos after Short-Term Storage at Different Temperatures"],"prefix":"10.3390","volume":"11","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-5317-2207","authenticated-orcid":false,"given":"Gustavo D. A.","family":"Gastal","sequence":"first","affiliation":[{"name":"Center for Artificial Insemination and Embryo Transfer, Department for Small Animals and Horses, University of Veterinary Medicine Vienna, 1210 Vienna, Austria"},{"name":"Instituto Nacional de Investigaci\u00f3n Agropecuaria INIA, Estaci\u00f3n Experimental La Estanzuela, Ruta 50 km 11, 39173 Colonia, Uruguay"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9599-0830","authenticated-orcid":false,"given":"Dragos","family":"Scarlet","sequence":"additional","affiliation":[{"name":"Division of Obstetrics, Gynecology and Andrology, Department for Small Animals and Horses, University of Veterinary Medicine Vienna, 1210 Vienna, Austria"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Maria","family":"Melchert","sequence":"additional","affiliation":[{"name":"Center for Artificial Insemination and Embryo Transfer, Department for Small Animals and Horses, University of Veterinary Medicine Vienna, 1210 Vienna, Austria"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7485-3661","authenticated-orcid":false,"given":"Reinhard","family":"Ertl","sequence":"additional","affiliation":[{"name":"Vetcore Facility, University of Veterinary Medicine Vienna, 1210 Vienna, Austria"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6077-7362","authenticated-orcid":false,"given":"Christine","family":"Aurich","sequence":"additional","affiliation":[{"name":"Center for Artificial Insemination and Embryo Transfer, Department for Small Animals and Horses, University of Veterinary Medicine Vienna, 1210 Vienna, Austria"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2021,5,6]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"102981","DOI":"10.1016\/j.jevs.2020.102981","article-title":"Current reproductive technologies impacting equine embryo production","volume":"89","author":"Squires","year":"2020","journal-title":"J. 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