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Objective: Vitrification is a convenient, effective method for freezing and storing embryos. Under certain situations, such as an unsuitable endometrial environment, extra embryos can be re-vitrified for future use. There is inadequate data on the effects of re-vitrification on embryos, so we have evaluated the effects of re-vitrification on the development rate and expression of apoptotic and implantation genes. Methods: Female NMRI mice, ages six-eight weeks were super-ovulated with 7.5 IU PMSG and 7.5 IU hCG. Females were mated with males from the same strain and inspected for the presence of vaginal plugs the following morning. Females with the presence of vaginal plugs were considered to be pregnant and killed 62 h post hCG injection. Eight-cell embryos were flushed from their oviducts and subsequently divided into three experimental groups: fresh, vitrified-warmed 8-cell embryos, and re-vitrified-warmed blastocyst embryos. RNA was extracted and we used real-time PCR to evaluate expressions of Bax, Bcl-2, and ErbB4. Data was analyzed by the chi-square and ANOVA tests. Results: A significant difference existed in blastocyst formation rate, degeneration rate, and expressions of Bax, Bcl-2, and ErbB4 in re-vitrified embryos compared to fresh embryos. Conclusion: The vitrification and warming process did not affect the developmental rate and expressions of Bax, Bcl-2, and ErbB4 in the eight-cell stage embryos. However, we observed a change in development rate and expression rates of Bax, Bcl-2, and ErbB4 after re-vitrification in the early blastocyst stage.

Keywords: Apoptosis, Implantation, Re-vitrification, Blastocyst

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