Abstract
The present study investigated the vitrification-induced deterioration of mitochondrial functions that may reduce the developmental ability of post-warming bovine embryos. In addition, the effect of supplementation of the culture medium with resveratrol on the mitochondrial functions and post-warming embryonic development was examined. Two days after in vitro fertilization, embryos with 8-12 cells (referred to hereafter as 8-cell embryos) were vitrified and warmed, followed by in vitro incubation for 5 days in a culture medium containing either the vehicle or 0.5 μM resveratrol. Vitrification reduced embryonic development until the blastocyst stage, reduced the ATP content of embryos, and impaired the mitochondrial genome integrity, as determined by real-time polymerase chain reaction. Although the total cell number and mitochondrial DNA copy number (Mt-number) of blastocysts were low in the vitrified embryos, the Mt-number per blastomere was similar among the blastocysts derived from fresh (non-vitrified) and vitrified-warmed embryos. Supplementation of the culture medium with resveratrol enhanced the post-warming embryonic development and reduced the Mt-number and reactive oxygen species level in blastocysts and blastomeres without affecting the ATP content. An increase in the content of cell-free mitochondrial DNA in the spent culture medium was observed following cultivation of embryos with resveratrol. These results suggested that vitrification induces mitochondrial damages and that resveratrol may enhance the development of post-warming embryos and activates the degeneration of damaged mitochondria, as indicated by the increase in the cell-free mitochondrial DNA content in the spent culture medium and the decrease in the Mt-number of blastocysts and blastomeres.