Effects of stimulation or inhibition of lipid peroxidation on freezing- thawing of mouse embryos

Abstract

This study was conducted to determine whether the tolerance of embryos to the stress of freezing and thawing can be modified by including in the incubation medium stimulators or inhibitors of membrane lipid peroxidation. Mouse zygotes were cultured in medium M16, supplemented or not with FeSO4, apotransferrin, and/or ascorbate. In each culture supplement, 8-cell embryos were randomly allocated to an untreated (nonfrozen) control group, or treatment by freezing using slow or ultrarapid cooling. In the slow-frozen group, FeSO4 (49.3%, 66 of 134), decreased embryo survival, whereas apotransferrin (82.7%, 110 of 133) and ascorbate (86.4%, 114 of 132), increased significantly the percentage of intact embryos recovered after thawing compared to those cultured in the basic medium M16 (66.9%, 99 of 148). Apotransferrin and ascorbate also increased significantly the percentage of blastocysts on Day 5 (79.7%, 106 of 133, and 89.4%, 118 of 132 vs. 62.2%, 92 of 148, respectively). Ascorbate in addition, increased significantly the percentage of implantations compared to those in the basic medium M16 (84.5%, 60 of 71 vs. 61.1%, 37 of 56). Changes in the ultrarapidly frozen group were not so evident. However, a significant fetal wastage after implantation was observed when embryos were cultured without additives and then ultrarapidly frozen (31.7%, 13 of 41 vs. 7.0%, 3 of 43, in the nonfrozen control group). Fetal weight was similar between culture conditions, but it was significantly lower in slow-frozen and ultrarapidly frozen embryos than in the nonfrozen control group. These data indicate that embryo tolerance to the freezing-thawing stress can be modified by incubating the embryos in the presence of stimulators or inhibitors of membrane lipid peroxidation. This effect is more evident in embryos cryopreserved by slow cooling than by ultrarapid freezing techniques.