88 Effect of culture medium on the development of mouse and cattle embryos

Potassium simplex optimized medium (KSOMaa) was developed by John Biggers and his colleagues during the 1990s, and it is the standard for mouse embryo culture. Variants of KSOMaa have shown to be highly successful for the culture of human embryos to the blastocyst stage. The current experiments were designed to evaluate CaseBioscience CaseMONO, a new variant of KSOMaa. Differences were tested by ANOVA or Fisher’s Exact Test, as appropriate. Mouse embryo culture and transfer experiments were performed by Embryotools S.L. In 61 standard one-cell mouse embryo assays, 1830 zygotes were cultured in CaseMONO medium and compared with 660 cultured in Sigma EmbryoMax (control), an early variant of KSOMaa. Development to expanded blastocyst by 96 h of culture was not different between the groups (96.7 ± 3.6% vs. 97.0 ± 2.5%; P = 0.7197), but the proportion of good-quality blastocysts was significantly greater in CaseMONO (96.3 ± 3.7% vs. 87.4 ± 4.8%, P < 0.0001). In a second experiment, mouse zygotes were cultured in CaseMONO or EmbryoMax for 96 h. Ninety blastocysts from each group were transferred to synchronized CD1 recipients, and the recipients were killed 10 days after transfer. The implantation rate (50.0% vs. 53.3%), fetal rate (32.2% vs. 31.1%), fetal loss rate (35.5% vs. 41.7%), fetal weight (56.8 ± 26.9 mg vs. 49.6 ± 23.0 mg), and fetal crown-rump length (7.95 ± 1.53 mm vs. 7.47 ± 1.68 mm) did not differ between the CaseMONO and control groups. Placental weight for the CaseMONO group was significantly greater than controls (64.7 ± 22.0 mg vs. 50.9 ± 19.0 mg, P = 0.0171), likely related to the significantly greater cell numbers in blastocysts derived from culture in CaseMONO compared with controls (210.9 ± 30.2 vs. 171.3 ± 33.5, P < 0.0001), as determined in a third experiment. The ability of CaseMONO to support the development of cattle embryos was evaluated by the Animal Reproduction and Biotechnology Laboratory, Colorado State University. Cattle oocytes were recovered from abattoir ovaries by aspiration (on Day −1), matured in vitro for 24 h (until Day 0), and then co-incubated with bull sperm for an additional 18 h (until Day 1). The presumptive zygotes were then cultured in either CaseMONO or the laboratory standard CDM-1 for 56 h (until Day 3). The cleaved embryos were transferred from CaseMONO to fresh CaseMONO or from CDM-1 to CDM-2 for culture through to Day 7–8 and then evaluated for the development to the blastocyst stage. In four independent replicates, 414 presumptive zygotes were cultured in CaseMONO and 230 in CDM-1/CDM-2. Blastocyst development was 28.2 ± 7.8% in CaseMONO versus 32.9 ± 10.2% in CDM-1/CDM-2 (P = 0.486). The results suggest that CaseMONO may support development of mouse embryos better than EmbryoMax, including proportion of good-quality blastocysts, blastocyst cell numbers, and placental weight. Differences between CaseMONO and EmbryoMax include Gly-Gln versus Gln, l-lactate versus d- and l-lactate, and gentamicin versus penicillin-streptomycin. The development of cattle embryos to the blastocyst stage in CaseMONO was not different from that in CDM-1/-2, suggesting that it may be useful for cattle embryo culture.