Linoleic acid incorporation and competence of bovine oocytes matured in vitro

CARRO, M.M.; BUSCHIAZZO, J.; ORESTI, G.M.; RIOS, G.; ALBERIO, R.

VANCOUVER

Congreso; 17th International Congress on Animal Reproduction (ICAR); 2012

International Congress on Animal Reproduction

Linoleic acid (LA) is a long chain unsaturated fatty acid which increases the survival of cryopreserved bovine oocytes and embryos when added in culture media at low concentrations (0.3?9 uM). The objectives of this work were to determine the effect of different concentrations of LA on bovine oocyte maturation. We determined how LA influenced nuclear maturation, the levels of fatty acid incorporation and the lipid classes to which the LA is incorporated in bovine oocytes. Cumulus oocyte complexes (COC) aspirated from ovaries recovered after slaughter were in vitro matured in a chemically defined maturation medium supplemented with LA at 9, 43 and 100 uM. Nuclear maturation was determined by staining denuded oocytes with bisbenzamide Hoechst 33 342. To evaluate LA incorporation, oocytes were incubated in the presence of radiolabelled LA (0.8 lCi) and randomly separated into three groups according to incubation intervals: T0 = 0 h (n = 188), T1 = 1 h (n = 173) and T22 = 22 h (n = 252). After incubation, lipids were extracted, separated by thin layer chromatography (TLC) and the incorporation of [14C]LA into each lipid class was measured. The results show that low and medium LA concentrations (9 and 43 uM) did not affect the nuclear maturation, whereas the highest concentration (100 uM) inhibited germinal vesicle breakdown. The incorporation of radiolabelled LA to oocytes was significantly higher (p < 0.05) in T22 group, with an average incorporation of T0 = 1.18 ± 0.69, T1 = 12.18 ± 3.67, T22 = 83.39 ± 19.60 pmol/100 oocytes. The radioactivity associated to each lipid band isolated by TLC showed that the LA is mainly esterified in phospholipids and triglycerides (p < 0.05). The incorporation of an unsaturated fatty acid such as LA to oocyte phospholipids mainly phosphatidylcholine, could affect membrane physical properties such as membrane fluidity, and would explain previous reports on the effects of LA for embryo/oocyte cryopreservation. However, high levels of LA may negatively alter the physical properties of membranes and may explain their inhibitory effect on germinal vesicle breakdown, altering signaling events of the oocyte. Furthermore, triglycerides are reservoirs of energy used during oocyte maturation and early embryonic development, therefore the esterification of LA in this lipid class could impact oocyte and embryo lipid metabolism. This study was supported by grants from the National Agency of Scientific and Technical Research and National Institute of Agricultural Technology