| dc.description.abstract | The use of cryopreservation of in vitro produced (IVP) bovine embryos is limited because IVP embryos have a lower cryo-tolerance than in vivo embryos. In general the cryo-tolerance of cells is influenced by the lipid phase behavior: when the lipid gel phase is formed at a lower lipid phase transition temperature this will allow better cryo-survival. The lipid phase transition temperature can be influenced by adaptation of the fatty acids linked to the membrane phospholipids. The lipid phase transition temperature is lower when the fatty acids have a higher unsaturation level and a shorter chain length and when the double bonds have a more central position. This study aimed to increase the unsaturation level of phospholipids of IVP embryos by the addition of unsaturated fatty acids to in vitro maturation (IVM) and in vitro culture (IVC) media with the final goal to improve membrane properties and hence cryo-tolerance of IVP embryos. The study determined the effects of the addition of unsaturated fatty acids in IVM or IVC media influenced in vitro developmental competence, phospholipid acyl composition and cryo-survival properties of oocytes and blastocysts. Oleic acid (OA) and linoleic acid (cis-9,cis-12-octadecadienoic acid; LA) were complexed to lipid free bovine serum albumin (BSA) and added to lipid free IVM or IVC media. In experiment 1, cumulus oocyte complexes (COCs) were exposed to either no fatty acids (control), 700 µM OA, 700 µM LA or a combination of 350 µM LA and 350 µM OA during the entire IVM period before routine embryo culture. In experiment 2, presumptive zygotes were exposed to either control BSA (final concentration of fatty acids 350 µM), ≥96% lipid free BSA, or BSA complexed with either 350 µM OA, 350 µM LA or a combination of 175 µM LA and 175 µM OA during the entire IVC period. The presence of LA, OA or a combination of LA and OA during IVM did not affect blastocysts rates. In contrast, the presence of LA during IVC, whether or not in combination with OA, resulted in significantly lower blastocyst rates. The largest drop in blastocyst rates was observed when IVC was performed with lipid free BSA. Lipid analysis was performed on oocytes and day 8 blastocysts of all experimental conditions. Interestingly, the overall effect of the addition of the unsaturated fatty acids LA, OA or a combination of LA and OA was a decrease in the number of double bonds in fatty acid complexed to phospholipids while no clear pattern in the changes in fatty acid chain length were observed in blastocysts. Therefore, the presence of LA, OA or a combination of LA and OA during IVM or IVC probably did not lower the lipid phase transition temperature of IVP blastocysts. Indeed, the cryo-survival rates after slow freezing of blastocysts exposed to OA or a combination of LA and OA during IVC were not affected. In oocytes, the results of this study indicate that maturation in presence of LA might improve the cryo-tolerance of oocytes due to a higher number of double bonds and a shorter chain length in phosphatidylcholine (PC). | |
