Animal oocyte and embryo cryopreservation

Abstract

Cryopreservation of oocytes and embryos is a crucial step for the widespread and conservation of animal genetic resources. However, oocytes and early embryos are very sensitive to chilling and cryopreservation and although new advances have been achieved in the past few years the perfect protocol has not yet been established. All oocytes and embryos suffer considerable morphological and functional damage during cryopreservation but the extent of the injury as well as differences in survival and developmental rates may be highly variable depending on the species, developmental stage and origin (for example, in vitro produced or in vivo derived, micromanipulated or not). Currently, there are two methods for gamete and embryos cryopreservation: slow freezing and vitrification. We have experienced both techniques but vitrification has become a viable and promising alternative to traditional approaches especially when dealing with in vitro produced or micromanipulated embryos and oocytes. Recently new strategies based on emerging studies in the field of lipid research have been used to reduce intracellular lipid content in bovine in vitro produced embryos and therefore increase their tolerance to micromanipulation and cryopreservation. The addition of a conjugated isomer of linoleic acid, the trans-10, cis-12 octadecadienoic acid to embryo culture medium more than twice improved embryo post-thawing viability after micromanipulation and vitrification. Vitrification was also used for the cryopreservation of embryos belonging to the Portuguese Animal Germplasm Bank project presently running at our facilities.

In vitro maturation, apoptotic gene expression and incidence of numerical chromosomal abnormalities following cryotop vitrification of sheep cumulus-oocyte complexes

“…Presence of lipid droplets and vacuoles in the ooplasm of sheep oocytes is an important obstacle in the way of oocyte cryopreservation; lipid droplets make oocytes more sensitive to cooling, causing irreversible injuries to membrane structures at low temperatures [35][36][37]. Also the number and structure of lipid droplets in different species can cause different cryosensitivity injuries for oocytes [38].…”

Obtaining farm animal embryos <i>in vitro</i>

“…In this case, the short time of vitrification and elimination of costly programmable freezing equipment is of great importance. There are many modifications of freezing and vitrification procedures (Vajta and Kuwayama, 2006;Ledda et al, 2007;Pereira and Marques, 2008;Prentiz and Anzar, 2011;Saragusty and Arav, 2011).…”

“…Oocytes can be cryopreserved at the GV stage and in metaphase II, but compared with embryos, oocytes are much more sensitive. As a result of cryoconservation they develop morphological and functional defects (Martins et al, 2005;Contreras et al, 2008;Pereira and Marques, 2008), therefore, the cryopreservation efficiency is low. The specificity of oocyte cryopreservation is due to the granulosa cells surrounding them; because of their role in oocyte maturation they can not be removed.…”

“…The specificity of oocyte cryopreservation is due to the granulosa cells surrounding them; because of their role in oocyte maturation they can not be removed. Further problems with oocyte cryopreservation are related to the morphology of the metaphase II oocyte cell membrane and cortical granules and spindle apparatus (Pereira and Marques, 2008).…”

“…Cattle and sheep embryos are the most resistant, the least -pig and horse embryos (Pereira and Marques, 2008). In the 1980s and 1990s, progeny was obtained after freezing and thawing of farm animal embryos.…”