Comparison of Ethylene Glycol and Propylene Glycol for the Vitrification of Immature Porcine Oocytes

Abstract

Our aim was to optimize a cryoprotectant treatment for vitrification of immature porcine cumulus-oocyte complexes (COCs). Immature COCs were vitrified either in 35% ethylene glycol (EG), 35% propylene glycol (PG) or a combination of 17.5% EG and 17.5% PG. After warming, the COCs were in vitro matured (IVM), and surviving oocytes were in vitro fertilized (IVF) and cultured. The mean survival rate of vitrified oocytes in 35% PG (73.9%) was higher (P<0.05) than that in 35% EG (27.8%). Oocyte maturation rates did not differ among vitrified and non-vitrified control groups. Blastocyst formation in the vitrified EG group (10.8%) was higher (P<0.05) than that in the vitrified PG group (2.0%) but was lower than that in the control group (25.0%). Treatment of oocytes with 35% of each cryoprotectant without vitrification revealed a higher toxicity of PG on subsequent blastocyst development compared with EG. The combination of EG and PG resulted in 42.6% survival after vitrification. The maturation and fertilization rates of the surviving oocytes were similar in the vitrified, control and toxicity control (TC; treated with EG+PG combination without cooling) groups. Blastocyst development in the vitrified group was lower (P<0.05) than that in the control and TC groups, which in turn had similar development rates (10.7%, 18.1% and 23.3%, respectively). In conclusion, 35% PG enabled a higher oocyte survival rate after vitrification compared with 35% EG. However, PG was greatly toxic to oocytes. The combination of 17.5% EG and 17.5% PG yielded reasonable survival rates without toxic effects on embryo development.

Effects of two combinations of cryoprotectants on the in vitro developmental capacity of vitrified immature porcine oocytes

“…Our results are similar to those previously reported for EG-based media [1,22]. However, Somfai et al [41,46] recently reported viability of approximately 28% using EG as the only permeant CPA and 42.6% viability with a combination of EG þ PG [46]. Although the differences in viability between the studies could be due to technical aspects of the vitrification procedure or the quality of the oocytes used, the dissimilarities in the final CPA concentration, which was 32% in our study and 35% in Somfai et al [46], could play a key role.…”

“…Although the reasons for these differences are unclear, the results suggest that the higher permeability of PG better protects oocytes compared with DMSO in terms of fertilization efficiency. As in previous studies [10,13,46], our results indicate that regardless of CPA combination, vitrified oocytes exhibit a reduced ability to develop to the blastocyst stage after IVF compared with controls.…”

“…In our study, those oocytes exposed to CPAs and classified as viable after IVM displayed similar fertilization [46] at similar concentrations. Although some authors have reported that DMSO is more toxic than PG to bovine oocytes [39] or EG to porcine embryos [1], we did not observe such differences between the treatment groups.…”

“…However, Somfai et al [41,46] recently reported viability of approximately 28% using EG as the only permeant CPA and 42.6% viability with a combination of EG þ PG [46]. Although the differences in viability between the studies could be due to technical aspects of the vitrification procedure or the quality of the oocytes used, the dissimilarities in the final CPA concentration, which was 32% in our study and 35% in Somfai et al [46], could play a key role. Supporting this speculation, it has been proved that slight variations in CPA concentration may significantly affect the vitrification efficiency of porcine embryos [26].…”

“…Although the CPA toxicity mechanisms are unclear, CPAs are known to cause changes in the organization of the microtubules in the oocytes of mice [42,43], rabbits [42], cattle [44], and humans [45]. In pigs, the exposure of immature oocytes to high concentrations of CPAs can trigger cytoskeletal disruption [12], and the excessive osmotic stress induced by CPAs can cause membrane damage in the oocytes [46]. All of these effects could be responsible for the increased number of degenerated oocytes found after IVM.…”