Effects of antifreeze glycoprotein 8 (AFGP8) supplementation during vitrification on the in vitro developmental capacity of expanded bovine blastocysts.

Cryopreservation is an effective method for the long-term storage of valuable germplasm in the field of reproductive research. The present study examined the developmental capacity of post-thaw bovine blastocysts during vitrification after supplementation with antifreeze glycoprotein 8 (AFGP8). Survival and re-expansion rates in culture during the 12h after thawing were significantly higher in the AFGP8-treated than untreated group. In addition, blastocysts from the AFGP8-treated group exhibited lower rates of apoptosis. Real-time reverse transcription-polymerase chain reaction analysis showed that the expression of the Bcl-2 gene, coding for an anti-apoptotic protein, was increased significantly, whereas the expression of the pro-apoptotic gene Bax was decreased significantly in the AFGP8-treated group. The cellular proliferation rate and mitochondrial membrane potential were significantly higher in post-thaw re-expanded blastocysts from the AFGP8-treated compared with untreated group. In addition, outgrowth potential in post-thaw blastocysts in re-expanded blastocysts after vitrification was significantly increased in the AFGP8-treated compared with untreated group. Together, these results are the first to demonstrate that the addition of AFGP8 during vitrification can help protect bovine blastocysts against chill-induced injury.