Back to Search Start Over

Efficient one-step direct transfer to recipients of thawed bovine embryos cultured in vitro and frozen in chemically defined medium

Efficient one-step direct transfer to recipients of thawed bovine embryos cultured in vitro and frozen in chemically defined medium

Authors :
José Manuel Prendes
Enrique J. Gómez
Javier Prendes
Isabel Gimeno
Antonio Murillo
Marta Muñoz
Alejandro Vázquez
Susana Carrocera
David C. Martin
Juan José Pérez-Jánez
Source :
Theriogenology. 146:39-47
Publication Year :
2020
Publisher :
Elsevier BV, 2020.

Abstract

Direct transfer (DT) of cryopreserved embryos to recipients facilitates on-farm application. We analyzed a new freezing/thawing (F/T) procedure for in vitro produced (IVP) embryos, integrating: 1) an ethylene-glycol based system; 2) a culture step without protein; and 3) a synthetic protein substitute (CRYO3) in cryopreservation medium. IVP embryos from abattoir ovaries were cultured in groups in BSA-containing synthetic oviduct fluid with or without 0.1% fetal calf serum (FCS) until Day-6. Morulae and early blastocysts were subsequently cultured without protein from Day-6 onwards. Day 7 and Day 8 expanded blastocysts (EXB) were subjected to F/T or vitrification/warming (V/W). Thawed and warmed EXB were cultured in vitro, and development rates, cell counts and dead cells were analyzed in surviving embryos. V/W improved survival over F/T (live and hatching rates at 2 h, 24 h and 48 h) (P 0.0001), and FCS before Day 6 did not affect in vitro survival. After F/T, embryos had lower cell counts in the ICM, TE and total cells than after V/W. Day-7 embryos after F/T showed % apoptotic, % pycnotic and % total dead cells higher (p 0.05) than their Day-8 counterparts, probably because F/T reduced the numbers of ICM cells within Day-8 embryos. Thereafter, Day-7 blastocysts were transferred to heifers in an experimental herd. There were no differences in birth rates with frozen (-FCS [n = 40]: 45%; +FCS [n = 14]: 28%), vitrified (-FCS [n = 47]: 53%; +FCS [n = 11]: 36%) and fresh (-FCS [n = 30]: 47%; +FCS [n = 17]: 53%) embryos. However, frozen embryos produced with FCS showed 5/9 miscarriages after Day-40. Calves born from frozen (n = 22), vitrified (n = 29) and fresh (n = 22) transfers did not differ in birth weight, gestation length and daily gain weight (P 0.10). Subsequently, transfer of frozen embryos (n = 29) derived from oocytes collected from live, hormonally stimulated cows in experimental herd, led to pregnancy rates of 57% (heifers) and 40% (dry cows). with EXB on Day-62 Finally, embryos produced with BSA were transferred to cows in an on-field trial (frozen [n = 80]; fresh [n = 58]), with no differences in pregnancy rates (days 30-40). Pregnancy and birth rates could not be predicted from in vitro approaches. The new F/T system yields pregnancy and birth rates comparable to vitrified and fresh embryos without birth overweight. The absence of products of animal origin, defined chemical composition, and direct transfer entail sanitary, manufacturing and application advantages.

Details

ISSN :
0093691X
Volume :
146
Database :
OpenAIRE
Journal :
Theriogenology
Accession number :
edsair.doi.dedup.....cd382628d2821a85575eaa5c74efaade
Full Text :
https://doi.org/10.1016/j.theriogenology.2020.01.056