Effects of oocyte activation and sperm preparation on the development of porcine embryos derived from in vitro-matured oocytes and intracytoplasmic sperm injection.
The objective was to determine the effects of various methods of oocyte activation and sperm pretreatment on development of porcine embryos derived from in vitro-matured oocytes and intracytoplasmic sperm injection (ICSI). The second polar body was extruded in the majority (>78.4%) of in vitro-matured (IVM) oocytes 4h after electrical pulse activation. In embryos generated by ICSI and sham-ICSI, a combination of an electrical pulse, with various chemical activators 4 h later, improved (P < 0.05) blastocyst formation rate compared to activation only with a pulse. Treatment with 6-dimethylaminopurine (DMAP) after electrical activation significantly increased the oocyte activation rate. The effects of exposure of sperm to repeated freeze-thaw cycles (without cryoprotectant) on oocyte activation and the effects of sperm pre-incubated with dithiothreitol (DTT) or Triton X-100 on early embryo development were also examined. Blastocyst formation rates after ICSI did not differ between motile sperm and those rendered immotile by one-time freezing and thawing without cryoprotectant. However, sperm rendered immotile by three cycles of freezing/thawing without cryoprotectant had a significantly lower blastocyst formation rate. Although oocytes injected with sperm pre-incubated with Triton X-100 had a higher normal fertilization rate than those pre-incubated with DTT or one-time frozen/thawed sperm, rates of blastocyst formation and cell numbers were similar among the three groups. In conclusion, various methods of oocyte activation and sperm preparation significantly affected the developmental capacity of early porcine embryos derived from IVM and ICSI.
Tian
Jian-Hui
JH
Key Laboratory of Animal Genetics and Breeding of The Ministry of Agriculture, College of Animal Science and Technology, China Agricultural University, Beijing 100094, PR China. tianjh@cau.edu.cn
Wu
Zhong-Hong
ZH
Liu
Lin
L
Cai
Yuan
Y
Zeng
Shen-Ming
SM
Zhu
Shi-En
SE
Liu
Guo-Shi
GS
Li
Ying
Y
Wu
Chang-Xin
CX
D000818:Animals; D002970:Cleavage Stage, Ovum / Q000187:drug effects; D003451:Cryoprotective Agents / Q000494:pharmacology*; D003470:Culture Media / Q000737:chemistry; D004229:Dithiothreitol / Q000494:pharmacology; D004560:Electricity; D046149:Embryo Culture Techniques / Q000379:methods / Q000662:veterinary*; D005260:Female; D008297:Male; D009865:Oocytes / Q000187:drug effects / Q000502:physiology*; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary; D020554:Sperm Injections, Intracytoplasmic / Q000379:methods / Q000662:veterinary*; D013081:Sperm Motility; D013552:Swine / Q000196:embryology*
D003451:Cryoprotective Agents; D003470:Culture Media; D004229:Dithiothreitol