In vitro maturation of porcine oocytes using a defined medium and developmental capacity after intracytoplasmic sperm injection.
To establish a defined in vitro maturation culture system for porcine oocytes, we examined the effects of adding cysteine (Cys) and epidermal growth factor (EGF) to the maturation medium. Furthermore, to evaluate cytoplasmic maturation, we investigated GSH concentrations and embryo development after intracytoplasmic sperm injection (ICSI). The basic media for IVM were modified TCM199 containing 10% newborn calf serum (NBCS) or 0.1% polyvinyl alcohol (PVA), supplemented with amino acids. Adding EGF (10 ng/ml) or EGF + Cys (0.57 mM) to the defined medium (0.1% PVA + amino acids) increased (P < 0.05) the rate of nuclear maturation relative to the defined medium (without these additives). After ICSI, oocytes matured in a medium supplemented with NBCS, Cys and EGF had a higher (P < 0.05) rate of pronuclear formation rate than oocytes matured in the defined IVM medium. Although there was no significant difference in cleavage rates between NBCS- and PVA-containing media supplemented with both Cys and EGF, the rate of blastocyst development was lower (P < 0.05) in the defined medium than in the NBCS-containing medium. Intracellular GSH concentrations of oocytes matured in the NBCS- and PVA-containing media supplemented with both Cys and EGF were higher (P < 0.05) than in oocytes matured in PVA alone or in oocytes before maturation. Adding Cys and EGF to a defined medium for porcine IVM improved rates of nuclear maturation and cleaved oocytes following ICSI, probably due to increased GSH concentrations. Also, embryos derived from oocytes matured in the defined medium (with the addition of Cys and EGF) developed into blastocysts after ICSI.
Kishida
R
R
Laboratory of Animal Reproduction, Obihiro University of Agriculture and Veterinary Medicine, Obihiro 080-8555, Japan.
Lee
E S
ES
Fukui
Y
Y
D000596:Amino Acids / Q000008:administration & dosage; D000818:Animals; D001755:Blastocyst / Q000502:physiology; D003470:Culture Media; D003545:Cysteine / Q000008:administration & dosage; D047108:Embryonic Development; D004815:Epidermal Growth Factor / Q000008:administration & dosage; D005260:Female; D005978:Glutathione / Q000032:analysis; D008297:Male; D009865:Oocytes / Q000502:physiology*; D011142:Polyvinyl Alcohol / Q000008:administration & dosage; D020554:Sperm Injections, Intracytoplasmic / Q000662:veterinary*; D013552:Swine; D046509:Tissue Culture Techniques
D000596:Amino Acids; D003470:Culture Media; D004815:Epidermal Growth Factor; D011142:Polyvinyl Alcohol; D005978:Glutathione; D003545:Cysteine