Evaluation of Equine Oocytes from Preserved Ovaries Using Intracytoplasmic Sperm Injection

抄録

This study was conducted to examine if oocytes obtained from one day preserved equine ovaries could be used for IVM and IVF research. Ovaries were transported at 20°C over a long distance from a slaughterhouse to a laboratory taking about 24 h. Oocytes with compact cumulus cell layers were collected from ovaries and cultured for 42 h in IVMD101 medium at 38.5°C in 5% CO₂ in air. The proportions of metaphase-II stage oocytes increased significantly between 24 and 30 h of culture and reached a plateau at 30 h. After 30 h of culture, the denuded oocytes with a first polar body were subjected to intracytoplasmic sperm injection (ICSI) using frozen-thawed stallion spermatozoa. The ICSI oocytes were activated by 10 min treatment with ionophore A23187 and 6 h treatment with cycloheximide, and cultured for up to 10 days in CR1aa medium. The proportion of ICSI oocytes fertilized normally (defined as those with 2 pronuclei and 2 polar bodies) was 44%. In vitro culture of ICSI oocytes resulted in a cleavage rate of 52% and a blastocyst development rate of 9%. These results indicate that equine oocytes derived from one day preserved ovaries can reach metaphase-II stage during 30 h of IVM, and that IVM oocytes can develop into blastocysts after ICSI.