Effective vitrification and warming of porcine embryos using a pH-stable, chemically defined medium.

Cristina Cuello,Cristina A Martinez,Inmaculada Parrilla,Alicia Nohalez,Jordi Roca,Emilio A Martinez,Maria A Gil

doi:10.1038/srep33915

Abstract

The use of pH-stable media would simplify embryo vitrification and the warming of porcine embryos and might facilitate the application of embryo transfer in practice. In this work, we investigated whether a pH-stable basal medium constituted of Tyrode’s lactate medium, polyvinyl alcohol, and HEPES for buffering was suitable for porcine embryo vitrification warming in place of the conventional gas-equilibrated media. A high percentage (>90%) of embryos survived vitrification and warming in this medium, achieving in vitro survival rates similar to embryos vitrified-warmed using the conventional protocol and their fresh counterparts. The pH-stable medium did not affect the in vivo developmental competence of the vitrified-warmed embryos. A farrowing rate of 71.4% (5/7) with 10.4 ± 3.1 piglets born was obtained for the embryos vitrified and warmed in this medium and transferred to selected recipients. This medium will enable the use of simple, safe and standardized protocols for the vitrification and warming of porcine embryos for optimal embryo survival and quality when applied under field conditions. This study opens new possibilities for the widespread use of embryo transfer in pigs.

Highlights

At present, there is an increasing demand for efficient porcine embryo cryopreservation protocols for industrial and research purposes
The results obtained in this study with the TL-polyvinyl alcohol (PVA) medium buffered with HEPES might be useful for the practical application of these technologies
Several buffers have been used for mammalian embryos, recent results obtained in bovines[19] indicated that HEPES-buffered media were highly efficient for the storage of embryos, leading to a highly stable pH and the preservation of embryo viability

Summary

Introduction

There is an increasing demand for efficient porcine embryo cryopreservation protocols for industrial and research purposes. For the commercial use of this technology, practitioners demand simple, safe and standardized protocols that warrant optimal embryo survival and quality when applied under field conditions In this sense, important progress has been made in recent years with the development of a one-step warming procedure[7] that is useful for direct embryo transfer (ET) or the combined application of vitrification and the non-surgical deep-intrauterine ET procedure[13,14,15] developed by our laboratory in the early 2000s16,17. The development of chemically-defined vitrification-warming media for in vivo[18] and in vitro-derived[12] embryos has improved the reproducibility of results and reduced the risk of disease transmission associated with animal origin compounds. The use of a single medium for the different steps of the technology (embryo collection, vitrification, warming and transfer) would simplify the process and enhance the traceability of the vitrified embryos

Objectives

Methods

Results

Conclusion