Ultrastructural changes of sheep cumulus–oocyte complexes following different methods of vitrification

Abstract

To determine the ultrastructural changes of sheep cumulus-oocyte complexes (COCs) following different methods of vitrification, good quality isolated COCs (GV stage) were randomly divided into the non-vitrified control, conventional straw, cryotop and solid surface vitrification groups. In both conventional and cryotop methods, vitrified COCs were respectively loaded by conventional straws and cryotops, and then plunged directly into liquid nitrogen (LN2); whereas in the solid surface group, vitrified COCs were first loaded by cryotops and then cooled before plunging into LN2. Post-warming survivability and ultrastructural changes of healthy COCs in the cryotop group especially in comparison with the conventional group revealed better viability rate and good preservation of the ooplasm organization. However in all vitrification groups except the cryotop group, mitochondria were clumped. Solely in the conventional straw group, the mitochondria showed different densities and were extremely distended. Moreover in the latter group, plenty of large irregular connected vesicles in the ooplasm were observed and in some parts their membrane ruptured. Also, in the conventional and solid surface vitrification groups, cumulus cells projections became retracted from the zona pellucida in some parts. In conclusion, the cryotop vitrification method as compared with other methods seems to have a good post-warming survivability and shows less deleterious effects on the ultrastructure of healthy vitrified-warmed sheep COCs.

In vitromaturation is slowed in prepubertal lamb oocytes: ultrastructural evidences

“…Concerning the morphology of other organelles, our data are generally in agreement with previous studies on ovine oocyte ultrastructure. In fact, the morphology of mitochondria, lipid droplets, Golgi complexes and endoplasmic reticulum, before and after maturation, were comparable with previous reports on prepubertal and adult sheep oocytes matured in vitro [ 16 , 17 , 22 ] or in vivo [ 27 – 29 ]. As well, we observed that the organelle redistribution during maturation was coincident with what described by others, both in vitro and in vivo [ 16 , 28 ].…”

Vitrification of human immature oocytes before and after in vitro maturation: a review

“…Deleterious effects of vitrification, such as retraction of cumulus cell projections, were observed in vitrified sheep COCs [79], similar to immature vitrified COCs from primed rats, were a reduction of F-actin in ooplasm and oocyte cortex was associated to irreversible but irregular retraction of transzonal projections [35]. The mechanical removal of cumulus cells from immature bovine oocytes did not affect their maturation competence but reduced blastocyst rates, when compared with intact COCs [80].…”

Cytoskeleton structure, pattern of mitochondrial activity and ultrastructure of frozen or vitrified sheep embryos

“…Higher degrees of mitochondrial swelling were observed in previous studies on cryopreserved grade I and II sheep embryos [2,5]. Mitochondrial swelling is also commonly described in cryopreserved oocytes [14,16,23]. Using in vitro produced embryos and similar procedures of slow freezing and vitrification Bettencourt et al [3] achieved satisfactory pregnancy rates of 68.4% and 54.6% on day 45, respectively.…”