Direct Warming of Vitrified In Vivo Equine Embryos

Area of Science:

• Reproductive Biology
• Embryology
• Cryopreservation

Background:

• Vitrification and warming protocols for in vitro embryos are established.
• In vivo embryos present greater challenges for cryopreservation due to their unique structure.
• Optimizing warming protocols for in vivo embryos is crucial for improving assisted reproductive technologies.

Purpose of the Study:

• To evaluate the efficacy of warming vitrified in vivo embryos using room temperature (RT) isotonic medium.
• To compare pregnancy rates between RT warming and traditional stepwise serial dilution warming.
• To investigate the influence of embryo size on the success of different warming protocols.

Main Methods:

• Vitrification of 117 in vivo embryos using a commercial kit, categorized by size (≤300 μm, >300-400 μm, >400-500 μm).
• Warming protocols included direct transfer to RT isotonic medium or stepwise serial dilution at 42°C.
• Warmed embryos were transferred to recipient mares, with Day 14 pregnancy rates compared between groups.

Main Results:

• Overall pregnancy rates showed no significant difference between RT warming (70.1%) and stepwise warming (82.5%).
• Pregnancy rates were comparable for smaller embryo groups (≤400 μm) regardless of warming method.
• Stepwise warming yielded significantly higher pregnancy rates for the largest embryo group (>400 μm) (p < 0.001).

Conclusions:

• Direct warming in RT isotonic medium is a viable and effective protocol for vitrified in vivo embryos up to 390 μm.
• For clinical application, RT warming is recommended for embryos ≤360 μm due to a noted decline in pregnancy rates in larger embryos.
• Stepwise serial dilution remains superior for warming larger in vivo embryos (>400 μm) to maximize pregnancy outcomes.