93 Supplementation of tauroursodeoxycholic acid to the in vitro culture medium: effects on the development rate and cryopreservability of bovine embryos

This study focused on the improvement of embryo quality under in vitro conditions using bile acids. In contrast to ex vivo embryos, in vitro-derived bovine embryos demonstrate lower development rates, lower pregnancy rates after transfer to recipients, and inferior viabilities after freeze-warming. Some evidence suggests that tauroursodeoxycholic acid (TUDCA) can reduce endoplasmic stress by supporting the folding of proteins and reducing cell damage caused by misfolding in embryos. This has been shown to improve embryo quality during in vitro production (IVP). Therefore, the present study aimed to analyze potential beneficial effects of TUDCA supplementation of culture medium on early embryonic development and cryogenic viability of bovine blastocysts. Cumulus–oocyte complexes from slaughterhouse ovaries were collected by follicle aspiration. Maturation (TCM-199 + 10% estrous cow serum (ECS), 38.5°C, 5% CO₂, 5% O₂) and IVF (Fert.-TALP, 38.5°C, 5% CO₂, 5% O₂) of oocytes were done via routine procedures using frozen thawed semen for IVF (2 × 10⁶ mL⁻¹). Presumptive zygotes (six replicates, n = 1230) were randomly allocated to two treatment groups represented by either in vitro culture for up to 9 days in SOFaa + 10% ECS (38.5°C, 5% CO₂, 20% O₂) supplemented with 200 µM TUDCA (n = 614) or without TUDCA (n = 616). At Day 7 of in vitro culture, only expanded blastocysts of the two treatment groups (n = 171) were cryopreserved using routine slow-freezing procedures (2 mM ethylene glycol, seeding at −6°C, gradual cooling by −0.5°C s⁻¹). For thawing, straws were placed in a 25°C water bath for 15 s followed by subsequent in vitro culture (SOFaa + 10% ECS) for 86 h to investigate survival, expansion, and hatching rates. For statistical analysis, an analysis of variance was conducted (GraphPad Prism®) with P-values < 0.05 considered to indicate significantly different values. Based on the results, TUDCA supplementation significantly improved blastocyst rate on Day 8 (31.2% vs. 37.5%) and on Day 9 (35.8% vs. 42.2%) of in vitro culture. Moreover, thawed blastocysts of the group supplemented with TUDCA during in vitro culture displayed significantly higher (P < 0.05) survival rates (50.1% vs. 65.3%) compared with control embryos. In agreement, hatching rates of blastocysts cultured in medium supplemented with TUDCA were significantly higher compared with control embryos after warming (19.6% vs. 45.4%). Taken together, the results of our study confirm our hypothesis that supplementation of the bile acid TUDCA to the in vitro culture medium exerts a positive effect on developmental rates of bovine IVP embryos during in vitro culture and on cryopreservability of subsequent blastocysts.