The effects of beta mercaptoethanol (β-ME) on in vitro development under oxidative stress and cystine uptake of bovine embryos were investigated. Bovine 1-cell embryos obtained by in vitro fertilization were cultured in TCM-199 or synthetic oviductal fluid (SOF) in 20% O2 supplemented with β-ME. Addition of β-ME significantly (P 0.01) promoted embryo development when cultured in both TCM-199 and SOF under high levels of O2, to almost the same rates when they were cultured in 5% O2. To investigate whether the growth-promoting effect of β-ME was related to cystine uptake, which is an important amino acid for intracellular glutathione (GSH) synthesis, 1-cell, 8-cell, morula, and blastocyst stage embryos were incubated in cystine, cysteine-free TCM-199 containing radioisotope-labeled cystine supplemented with or without β-ME. It was found that cystine uptake was consistently low in each embryo stage incubated without β-ME.

It is well known that in vitro development of bovine embryos is highly affected by culture conditions. Therefore, much effort has been made to develop effective culture methods that support embryo development in vitro. The detrimental effect of oxidative stress caused by a high oxygen concentration (95% air; approximately 20% O2 concentration) under culture conditions on the development of mammalian preimplantation embryos was recently studied. It was found that embryo development is promoted under a low O2 concentration as opposed to a high O2 concentration. However, reduced embryo development in a high O2 concentration was restored by addition of free radical scavengers. Therefore, to protect embryos against oxidative stress seems to be one of the keys to improving development. We have demonstrated that development of bovine embryos was promoted by beta mercaptoethanol (β-ME), a low molecular weight thiol that is used as a reducing agent in culture medium. Since that research was conducted, the beneficial effect of β-ME has been widely investigated on maturation of bovine oocytes and development of embryos in vitro cultured in a 20% O2 concentration. However, the effect of β-ME on in vitro development under conditions of oxidative stress such as a high O2 concentration, has not been elucidated precisely in comparison with development under conditions of low O2 concentration.

Effect of BSO on [14C]cystine uptake of bovine embryos. Ten 8-cell stage embryos were incubated for 30 min in cystine, cysteine-free TCM-199 plus [14C]cystine containing 100 μM beta mercaptoethanol with or without 5 mM BSO. After incubation with or without BSO, the radioactivity of incorporated [35S]cystine was measured with a scintillation counter. Data show means ± SEM. n, The number of replications; a vs. b, P 0.0001; b vs. c, P 0.0001; a vs. c, P 0.01.