128 EFFECT OF OXYGEN TENSION ON INDIVIDUAL IN VITRO BOVINE EMBRYO DEVELOPMENT IN CUMULUS CELL COCULTURE

Abstract

An in vitro production system, in which a single oocyte can be followed from the moment of retrieval up to the blastocyst stage, would be a valuable tool for studies linking developmental competence and embryo metabolism to immature oocyte quality and follicular environment. Earlier studies revealed that cumulus cell (CC) coculture during IVC enhances individual development, in contrast to group culture. These studies were performed in 5% O2, whereas generally an atmosphere of 20% oxygen is used for coculture with somatic cells. This study aimed to investigate the effect of oxygen tension on individual embryo development in CC coculture. As a control, the effect of oxygen tension on embryo group culture without CC was assessed simultaneously. Therefore, 692 COC from slaughterhouse ovaries (4 replicates) were routinely matured (TCM-199 + 20% serum) and fertilized in groups and then assigned to 4 culture treatments (SOF + 5% serum under oil). Treatments were T1 = 1 zygote in 20 μL + CC in 5% O2; T2 = 1 zygote in 20 μL + CC in 20% O2; T3 = 20 to 25 zygotes in 50 μL in 5% O2; and T4 = 20 to 25 zygotes in 50 μL in 20% O 2. Cleavage, blastocyst, and hatching rates were assessed 2, 8, and 10 days after fertilization, respectively. Possible effects of oxygen tension in individual and group culture were evaluated with binary logistic regression. No interactions between replicate and treatment could be found. Cleavage rates of individual culture showed a tendency (P < 0.1) to be lower in 5% O2 (62.1 v. 71.0% in 20% O2), whereas blastocyst rates were significantly (P < 0.05) higher in 5% O2 (26.6 v. 16.6% in 20% O2). Hatching rates did not differ significantly between the 2 individual treatments (Table 1). Oxygen tension did not have a significant effect on cleavage rates when embryos were cultured in groups, but blastocyst rates were significantly higher in 5% O2 (41.7 v. 27.6% in 20% O2). The group results confirm other studies (Yuan YQ et al. 2003 Theriogenology 59, 1585–1596). In conclusion, higher blastocyst rates can be obtained when an atmosphere of 5% O2 is used for culturing individual zygotes in CC coculture. Because cleavage rates showed a tendency to be higher in 20% O2, an alternating treatment, with 20% O2 until 2 days after fertilization, followed by 5% O2 until the blastocyst stage, should be investigated. Table 1. Cleavage, blastocyst, and hatching rates in 5 and 20% oxygen