Oxygen tension and medium supplements for in vitro maturation of bovine oocytes cultured individually in a chemically defined medium.
The effects of adding cysteamine, EGF, and glucose as an energy substrate under low oxygen tension during in vitro maturation (IVM) were examined to find ways of improving the individual in vitro production (IVP) system in individually cultured bovine oocytes. The basic medium was mSOFaa containing 1 mg/ml polyvinyl alcohol. Immature oocytes were individually cultured in an IVM medium with 10 ng/ml EGF, 100 microM cysteamine, or EGF plus cysteamine under 20% or 5% O(2). Cleavage and blastocyst rates were significantly higher (P<0.05) in IVM culture was under 20% O(2) than in culture under 5% O(2). Under 5% O(2), neither EGF nor cysteamine improved embryonic development. The proportion of matured oocytes was significantly higher (P<0.05) in the presence of 1.5 mM glucose under 20% O(2) (68.6%), and 5.5 mM (66.7%) and 10 mM (65.5%) glucose under 5% O(2). The presence of 5.5 mM glucose significantly (P<0.05) increased the maturation rate compared with the absence of glucose, irrespective of addition of EGF and cysteamine. The addition of cysteamine alone in the maturation medium significantly (P<0.05) increased the intracellular GSH concentration in the oocytes. Also, under 5% O(2) cysteamine and/or EGF significantly (P<0.05) improved the proportions of penetrated oocytes, cleavage and blastocyst formation, which were similar levels to those of oocytes matured under 20% O(2). After vitrification, the re-expanding and hatching rates of blastocysts derived from the individual IVP system containing cysteamine under 5% O(2) were significantly (P<0.05) higher than those of blastocysts derived from the individual IVP system without cysteamine under 5% O(2) and the group IVP system under 20% O(2). The present study showed that a high glucose level (5.5 or 10 mM) was optimal in IVM culture under low (5%) oxygen tension. The addition of EGF and/or cysteamine to the maturation medium had no positive effect on nuclear maturation, but improved fertilizability, developmental competence and cryoresistance following vitrification, probably due to increased GSH synthesis during the IVM process.
Oyamada
Takahiro
T
Laboratory of Animal Reproduction, Obihiro University of Agriculture and Veterinary Medicine, Obihiro 080-8555, Japan.
Fukui
Yutaka
Y
D000818:Animals; D001757:Blastomeres; D002417:Cattle; D018929:Cell Culture Techniques / Q000379:methods*; D002455:Cell Division; D003470:Culture Media / Q000494:pharmacology; D003543:Cysteamine / Q000494:pharmacology; D004734:Energy Metabolism; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005260:Female; D005307:Fertilization in Vitro; D005947:Glucose / Q000494:pharmacology; D009865:Oocytes / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D010100:Oxygen / Q000494:pharmacology*
D003470:Culture Media; D003543:Cysteamine; D004815:Epidermal Growth Factor; D005947:Glucose; D010100:Oxygen