研究発表を行った学会；The 15th Transgenic Technology Meeting (TT2019)
2019年 4月 7日〜10日（神戸）
タイトル；N-acetyl cysteine improves the fertilizability of vitrified-warmed mouse oocytes by recovering the thiol levels of zona pellucida post oxidation.
（熊本大学 生命資源研究・支援センター 資源開発分野）
Oocyte vitrification is useful for preserving female genetic resources. However, decline in fertilizability of vitrified-warmed oocytes has prevented the efficiently production of genetic engineered mice. In this study, we addressed the problem of decreasing in fertilizability of vitrified-warmed oocytes by assessing the oxidation of zona pellucida and examining the effect of N-acetyl cysteine (NAC) on the fertilizability of vitrified-warmed oocytes.
Materials and Methods
Male mice aged 12 weeks and female mice aged 4 weeks were used as sperm and oocyte donors, respectively. Superovulation was induced in the female mice by injecting a mixture of 0.1 mL inhibin antiserum and 3.75 IU equine chorionic gonadotropin, followed by an injection of 7.5 IU human chorionic gonadotropin. Oocytes from the superovulated females were collected and vitrified using simple vitrification method. After warming vitrified oocytes, the fertilization rate was evaluated by IVF using fertilization medium with or without NAC. The birth rate of NAC-treated oocytes was then examined by embryo transfer. The protein thiol level of zona pellucida was measured using Alexa Fluor 488 C5 maleimide before and after vitrification.
Results and Discussion
We observed that the protein thiol level in zona pellucida decreased by vitrification, especially after treatment of 1M DMSO. However, NAC recovered the protein thiol level and increased the fertilization rate of vitrified-warmed oocytes. Two-cell embryos produced by IVF normally developed into pups. These results suggested that NAC improves the fertilizability of vitrified-warmed oocytes by rescuing the oxidized status of zona pellucida.