192 Effect of l-carnitine and resveratrol supplementation to the maturation medium on in vitro bovine embryo production

l-Carnitine (LC) and resveratrol (RES) are two additives with antioxidant properties that have proven effective in stimulating energy metabolism and reducing oxidative stress in cells. Supplementation with these additives in the IVM medium of bovine oocytes could provide benefits in nuclear maturation, fertilization, and embryonic development. In this context, this study evaluated the effect of supplementation with LC (0.3 mg mL⁻¹), RES (1 µM), and their combination (LC+RES = 0.15 mg mL⁻¹ + 0.5 µM) in the IVM medium of oocytes on the nuclear maturation rate (MII) and embryonic development rate (cleavage and blastocyst) after IVF. A total of 1209 cumulus–oocyte complexes (COCs) were retrieved from bovine ovaries and matured in 60-µL drops (25 oocytes/drop) of IVM medium (TCM-199, 10% FBS, 0.2 mM sodium pyruvate, 2 µg mL⁻¹ estradiol-17β, 5 µg mL⁻¹ LH, 25 µg mL⁻¹ FSH, 2 mM l-glutamine, 100 µM cysteamine, and 50 µg mL⁻¹ gentamicin) supplemented with LC (n = 304), RES (n = 300), LC+RES (n = 302), and non-supplemented (control, n = 303). After 24 h of incubation (6% CO₂ and maximum humidity), the oocytes were in vitro fertilized with frozen-thawed bovine semen and in vitro cultured for 7 days. After maturation, 100 presumptively matured oocytes from each treatment were fixed with formaldehyde (2%), stained with Hoechst, and evaluated for meiotic maturation (MII) using epifluorescence microscopy (UV-2E/C filter, excitation 330–380 nm, emission 420 nm). The remaining oocytes were in vitro cultured, and the cleavage and blastocyst rates were evaluated at 3 and 7 days post-IVF, respectively. A one-way ANOVA and Fisher’s LSD tests were used to evaluate the effects of the additive treatments on embryonic development. The results showed no significant differences in the nuclear maturation rate (P > 0.05; LC = 86.8%; RES = 81.4%; LC+RES = 79.5%; and control = 75.3%). The cleavage rate was higher (P < 0.05) in oocytes matured with LC compared with the control (78.9 ± 2.86% vs. 68.5 ± 3.27%, respectively). Similarly, the blastocyst rate was significantly higher (P < 0.05) in oocytes matured with LC compared with those matured with LC+RES and the control (39.7 ± 3.43% vs. 27.2 ± 3.14% and 30.0 ± 3.23%, respectively). Of the total blastocysts produced in this experiment (263 embryos from 809 presumed zygotes [32.5%]), the LC group produced a higher percentage (P < 0.05) of compact blastocysts compared with the control; and all LC, RES, and LC+RES groups produced higher percentages (P < 0.05) of expanded blastocysts compared with the control group. In conclusion, LC supplemented in the maturation medium of bovine oocytes resulted in better embryonic development, while RES, alone or combined with LC, had no significant effect on the in vitro production of bovine embryos