Assessment of the effect of adding L-carnitine and/or resveratrol to maturation medium before vitrification on in vitro-matured calf oocytes

Abstract

Cryopreservation may lead bovine oocytes to undergo morphological changes and functional damage due to the high-lipid content in the cytoplasm and the formation of reactive oxygen species. Against this background, the present study aimed to improve the cryotolerance and developmental competence of prepubertal calf oocytes by adding L-carnitine (LC) and/or resveratrol (R) to the IVM medium, as the former is involved in lipid metabolism and both are able to scavenge reactive oxygen species. With this purpose, different quality and functional oocyte parameters, such as spindle and chromosome configuration, DNA integrity, caspase activity, and the profile of genes involved in lipid metabolism and oxidative stress were evaluated in IVM bovine oocytes before or after vitrification/warming. Oocytes were matured in the absence (control) or presence of LC (3.03 mM) and/or R (1 μM) and then vitrified/warmed before IVF and embryo culture. All treatment groups (control, LC, R, and LC + R) of nonvitrified IVM oocytes showed similar rates (P > 0.05) of a normal spindle and chromosome configuration to oocytes vitrified/warmed after maturation in the presence of LC + R. When oocytes in all treatment groups were compared before and after vitrification, no significant differences were detected in DNA fragmentation as measured using the TUNEL method. However, the proportion of early apoptotic oocytes increased after vitrification/warming, except when previously matured with R. Vitrified/warmed oocytes matured in the presence of LC did not differ with nonvitrified oocytes in terms of the expression of ACACA, SLC2A1, PLIN2, HSPA1A, GPX1, and SOD1 genes. Similarly, expression of ACACA, SLC2A1, PLIN2, HSPA1A, and SOD1 genes in vitrified/warmed oocytes was similar to that of their fresh counterparts when matured in the presence of R. Finally, while the addition of LC and/or R to IVM medium had no effect on both cleavage and blastocyst rates either in fresh or vitrified oocytes. To conclude, the results of the present study report that the addition of LC and/or R to the IVM medium used for prepubertal bovine oocytes did not increase the embryo development potential of both fresh and vitrified oocytes. However, LC + R supplementation before vitrification decreased spindle damage, R addition–modulated apoptosis, and LC or R addition before vitrification positively affected the gene expression of vitrified/warmed oocytes.