USE OF COMBINED OXIDATIVE SUBSTRATES IN PORCINE OOCYTE IN VITRO MATURATION

FLORENCIA IRIARTE; CAROLINA LETO; STEPHANIA MADRID GAVIRIA; TOMÁS GADZE; ELIZABETH BREININGER; PABLO CETICA; SERGIO MORADO

Jornada; VI Jornadas Internacionales INITRA; 2021

The aim of this work was to study the implication of endogenous lipids, amino acids (Aa) or their combination as oxidative substrates for porcine oocyte nuclear and cytoplasmic maturation in vitro. Immature cumulus-oocyte complexes (COCs) were obtained by aspiration of ovarian antral follicles from slaughtered gilts and they were randomly distributed into 6 groups according to their maturation medium: NCSU-37 without pyruvate or glucose, NCSU-37 + glucose, NCSU-37 + L-carnitine (fatty acid β-oxidation activator), NCSU-37 + Aa, NCSU-37 + L-carnitine + Aa, NCSU-37 + L-carnitine + Aa + glucose. All the groups were matured for 44h at 39ºC, 5% CO2 and 100% humidity. In each group, oocyte endogenous lipids consumption was determined using Nile Red assay and the meiotic maturation percentage was evaluated by epifluorescence microscopy using Hoechst 33342 solution. To analyze cytoplasmic maturation, boar semen was centrifuged and resuspended in modified Tris Buffer medium (mTBM) to reach a 1x106 motile sperm/ml final concentration. Sperm and COCs were co-incubated for 3h in mTBM + bovine serum albumin and then putative zygotes were cultured in NCSU-23 supplemented with essential and non-essential Aa under mineral oil at 39°C, 5% CO2 and 100% humidity. Blastocyst percentages were determined at day 7 after in vitro fertilization. Meiotic maturation and blastocyst percentages were compared using a Chi-square analysis for non-parametric data. Endogenous lipids levels were expressed as mean ± standard error mean and interactions were analyzed by two-way ANOVA, using post-hoc general contrasts for comparison among treatments. The addition of L-carnitine, alone or in combination with glucose or Aa, to the maturation media increased oocyte endogenous lipid consumption respect to oocytes matured in the presence of glucose or Aa as unique oxidative substrates (p