Delipidating in vitro-produced bovine zygotes: Effect on further development and consequences for freezability

To study the effect of partial removal of intracytoplasmatic lipids from bo vine zygotes on their in vitro and in vivo survival, presumptive zygotes we re delipidated by micromanipulation and cocultured with Vero cells in B-2 10% FCS. Blastocyst rates of delipidated (n=960), sham (centrifuged but no t delipidated, n=830) and control embryos (n=950) were 42.1, 42.3 aid 39.9% respectively (P > 0.05). Day 7 blastocysts derived from delipidated zygote s had a mean of 123.9 +/- 45.6 nuclei compared to 137.5 +/- 32.9 for contro l blastocysts (P > 0.05). The full-term development of delipidated blastocy sts after single transfer to recipients was similar to that of control IVF blastocysts (41.2% vs 45.4% respectively). To assess the effect of delipida tion on the embryo tolerance to freezing/thawing, delipidated (n=73), contr ol (n=67) and sham (n=50) Day 7 blastocysts were frozen in 1.36 M glycerol + 0.25 M sucrose in PBS. After thawing, embryos were cocultured for 72 h wi th Vero cells in B-2 + 10% FCS. Survival rates at 24 h were not significant ly different between groups. However, in the delipidated group, the surviva l rate after 48 h in culture was significantly higher than in the control g roup (56.2 vs 39.8, P < 0.02), resulting in a higher hatching rate after 3 days in culture (45.2 vs 22.4, P < 0.02). Pregnancy rates for delipidated a nd control frozen/thawed embryos were respectively 10.5 and 22.2% (P > 0.05 ). Electron microscopic observations showed much fewer lipid droplets (and smaller) in delipated blastocysts than in controls. Taken together, our dat a show that delipidation of one cell stage bovine embryos is compatible wit h their normal development to term and has a beneficial effect on their tol erance to freezing and thawing at the blastocyst stage. This procedure, how ever, alters the developmental potential of such blastocysts, suggesting th at maternally inherited lipid stores interfere with metabolic recovery afte r thawing. (C) 2001 by Elsevier Science Inc.