Lupeol supplementation improves the developmental competence of bovine embryos in vitro

Lupeol is a triterpene with various pharmacological properties. This study investigated the effect of lupeol on the in vitro development of bovine embryos. Oocytes (270 per group, 1620 in total) obtained from slaughterhouse-derived ovaries were matured and fertilized in vitro and then cultured for 8 days in a humidified atmosphere of 5% CO₂ in air at 38.5 °C. The in vitro maturation medium was supplemented with 0.5, 1.0, 2.0, 3.0, and 4.0 μM lupeol. Treatment with 2.0 μM lupeol significantly (P < 0.05) improved blastocyst development. Hoechst 33342 staining and terminal deoxynucleotidyl transferase dUTP nick-end labeling showed that treatment with 2.0 μM lupeol improved blastocyst quality by increasing the total cell number and reducing the apoptotic cell number. Confocal microscopy confirmed that treatment with 2.0 μM lupeol significantly (P < 0.05) reduced the level of 8-oxoguanine, an indicator of reactive oxygen species. Lupeol treatment also significantly attenuated protein expression of nuclear factor-kappa B subunit 1 (NFKB1), cyclooxygenase (COX) 2, and CASP3. Real-time PCR analysis of nitric oxide synthase 2, NFKB1, COX2, CASP3, and BCL2-associated X protein supported the immunofluorescence data. In conclusion, lupeol is a potent antioxidant that improves bovine embryo development in vitro.