| dc.description.abstract | Cryoprotective agents (CPAs) are crucial for vitrification of e.g., oocytes and embryos, aiding in assisted reproductive technologies (ARTs). CPAs protect the cell when exposed to freezing temperatures of liquid nitrogen (LN2), forming a vitreous glass-like form. However, vitrification requires optimal exposure times and concentration of CPAs to avoid damages such as ice crystallisation, osmotic stress, DNA damage, damage to the zona pellucida (ZP) and oxidative stress. The aim of the present study was to assess the effect of CPAs on bovine oocytes and embryos and utilise these in a vitrification process while evaluating morphology by Hoechst staining. In order to evaluate the effectiveness of the CPAs on oocytes and embryos, parameters of developmental competence were used: cleavage and blastocyst rate (%). Findings showed that EG 7.5% + PG 7.5%, GLY 10%, PG 10% and EG 7.5% + DMSO 7.5% with 73.08%, 77.59%, 81.67% and 80.00% cleavage rate, respectively, performed the best amongst all selected CPAs. Measure of blastocyst rate showed that EG 7.5% + GLY 7.5% and GLY 10% with 40.4% and 39.7%, respectively, performed the best. Parthenogenesis was observed for EG 10% with 7.14% cleavage rate. However, the cleaved oocyte did not develop further. Compact morulae were vitrified using four different CPA (EG 7.5% + PG 7.5%, GLY 10%, PG 10% and EG 7.5% + DMSO 7.5%) concentrations. The post-thaw results showed that these morulae did not develop further and died. This may have been due to several factors such as osmotic stress, damage to the ZP, DNA damage or the low concentration of CPAs. Staining by Hoechst 33342 showed membrane damage and fragmentation of blastomere cells. For future studies to ensure survival and developmental competence after vitrification, higher concentration of CPAs and extra post-thaw procedure steps should be utilised. Furthermore, a TUNEL-assay would aid in providing data on the blastomere state (living/dead) following vitrification. | |
