Recent progress in bovine in vitro‐derived embryo cryotolerance: Impact of in vitro culture systems, advances in cryopreservation and future considerations

Abstract

Contents Cryopreservation of in vitro‐derived bovine embryos is a crucial step for the widespread reproduction and conservation of valuable high‐merit animals. Given the current popularity of bovine in vitro embryo production (IVP), there is a demand for a highly efficient ultra‐low temperature storage method in order to maximize donor ovum pickup (OPU) turn‐over, recipient availability/utilization and domestic/overseas commercial trading opportunities. However, IVP bovine embryos are still very sensitive to chilling and cryopreservation, and despite recent progress, a convenient (simple and robust) protocol has not yet been developed. At the moment, there are two methods for bovine IVP embryo cryopreservation: slow programmable freezing and vitrification. Both of the aforementioned techniques have pros and cons. While controlled‐rate slow cooling can easily be adapted for direct transfer (DT), ice crystal formation remains an issue. On the other hand, vitrification solved this problem but the possibility of successful DT commercial incorporation remains to be determined. Moreover, simplification of the vitrification protocol (including warming) through the use of an in‐straw dilution without the use of a microscope is a prerequisite for its use under farm conditions. This review summarizes the bovine IVP embryo cryopreservation achievements, strengths and limitations of both freezing systems and prospective improvements to enhance cryosurvival, as well as perspectives on future directions of this assisted reproductive technology.

In Vitro Production of Embryos from Prepubertal Holstein Cattle and Mediterranean Water Buffalo: Problems, Progress and Potential

“…In addition to yielding similar rates of embryos and pregnancies following transfer, another goal is for prepubertal-derived embryos to have cryotolerance similar to that of adult-derived embryos. It is well documented that in vivo produced embryos are more cryotolerant than their in vitro produced counterparts [ 135 , 136 , 137 ]. As such, embryo quality plays a major role in post-thaw survivability, with the cytoplasmic lipid content, i.e., the number and size of lipid droplets, shown to affect cryotolerance significantly, with more lipids being detrimental [ 135 ].…”

The incompletely fulfilled promise of embryo transfer in cattle—why aren’t pregnancy rates greater and what can we do about it?

“…It is a reasonable expectation that the most competent embryos for pregnancy establishment will be derived from culture systems that either closely mimic conditions in vivo for oocyte maturation, fertilization, and embryonic development or utilize alternative solutions to place embryos in the same physiological state as would occur in vivo. There have been important changes in culture systems since the first calf from an IVP embryo was produced in 1981 ( Brackett et al, 1982 ), including modifications of procedures for oocyte maturation, fertilization, and embryo culture (see Parrish, 2014 ; Sirard, 2018 ; and Ferré et al, 2020 for some historical perspective). Even today, however, culture systems differ in many ways from the situation in vivo, including the substratum embryos reside on (plastic instead of epithelium), availability of energy substrates, amino acids, micronutrients and cell-signaling molecules, and fluid dynamics around the embryo.…”

PGE2 Supplementation of Oocyte Culture Media Improves the Developmental and Cryotolerance Performance of Bovine Blastocysts Derived From a Serum-Free in vitro Production System, Mirroring the Inner Cell Mass Transcriptome

“…Studies performed in cattle have shown that embryos produced in vitro are more vulnerable to cooling than their counterparts generated in vivo ( Nedambale et al, 2004 ; Yu et al, 2010 ). In addition to the absence of a natural environment for oocytes and embryos, the composition of the culture media used throughout the successive steps of in vitro production (IVP); namely in vitro maturation (IVM), in vitro fertilization (IVF), and in vitro development (IVD), may modulate embryo survival after thawing ( Agarwal et al, 2014 ; Swain et al, 2016 ; Ferre et al, 2020 ). In cattle, it has been well known for decades that adding serum to culture media negatively affects embryo cryotolerance ( Yamashita et al, 1999 ).…”