RESUMO
PURPOSE: Antioxidant and anti-apoptotic effects of melatonin on development of in vitro fertilization (IVF)/vitrified two-cell mouse embryos were evaluated in this study. METHODS: The IVF two-cell embryos were vitrified by cryotop, and were cultured in KSOM medium in different concentrations of melatonin (10(-6), 10(-9), 10(-12) M) and without melatonin. The blastocyst cell number, apoptotic cells and glutathione (GSH) level were evaluated by differential, TUNEL and cell tracker blue staining, respectively. The expression of Bax and Bcl-xl genes was evaluated by qPCR. The expression of melatonin receptors (Mtnr1a and Mtnr1b) in mouse 2-cell embryos and blastocysts was evaluated by RT-PCR. RESULTS: Melatonin increased the rate of cleavage and blastulation at 10(-12) M concentration (p < 0.05). The number of trophectoderm and inner cell mass showed a significant increase (p < 0.05) in 10(-9) M melatonin. The 10(-9) M and 10(-12) M melatonin treatments significantly reduced (p < 0.05) the apoptotic index. The significant increase in the expression of Bcl-xl observed at 10(-9) M concentration however, reduced expression of Bax was not statistically significant. The levels of GSH in 10(-9) and 10(-12) M groups were significantly improved relative to the control group (p < 0.05). The Mtnr1a was expressed in 2-cell embryos and blastocysts in all groups, but the expression of Mntr1b was not detected. CONCLUSION: Melatonin may have a special role against oxidative stress in protection of IVF/vitrified embryos.
