Abstract
BackgroundIn cryopreservation, oocytes are subjected to extreme hyperosmotic conditions, inducing large volume changes that, along with an abrupt temperature drop, interfere with their developmental competence. Our objectives in this work were to find conditions enabling an increase in oocyte cryosurvival and subsequent development.MethodsAbattoir-derived bovine oocytes were cultured without (Control group) or with trans-10,cis-12 conjugated linoleic acid isomer (CLA group). Comparative observations were made for 1) the oocyte developmental competence after exposure to cryoprotectants followed or not by vitrification/warming, 2) the oocyte membrane permeability to water (using the non-permeant cryoprotectant sucrose) and 3) the oocyte membrane permeability to two cryoprotectants (ethylene glycol, EG, and dimethyl sulfoxide, DMSO). Mature oocytes cultured with or without CLA and vitrified/warmed or only exposed to cryoprotectants without vitrification were subjected to in vitro fertilization; embryo culture proceeded until the blastocyst stage. The oocyte membrane permeabilities to water and cryoprotectants were estimated using mature oocytes subjected to hyperosmotic challenges. For water permeability, 200 mM sucrose was used, whereas for the cryoprotectant permeability, a 10 % solution of both EG and DMSO was used. The data were analyzed using the MIXED procedure and Student’s T-test.ResultsCLA supplementation improves the developmental competence of vitrified/warmed and cryoprotectants exposed oocytes (p < 0.01) and reduces their membrane permeability to water (37 %, p < 0.001) and to cryoprotectants (42 %, p < 0.001).Conclusions: By slowing the fluxes of water and of permeant cryoprotectants, CLA contributed to improved oocyte cryosurvival and post-thawed viability. This isomer supplementation to the maturation media should be considered when designing new protocols for oocyte cryopreservation.
Highlights
In cryopreservation, oocytes are subjected to extreme hyperosmotic conditions, inducing large volume changes that, along with an abrupt temperature drop, interfere with their developmental competence
Oocyte maturation and embryo production In experiment 1, supplementation of oocyte culture medium with conjugated linoleic acid (CLA) improved the rate of viable Cumulus oocyte complexes (COC) that survived the vitrification process (CLA vitrified: 80.4 ± 5.6 % vs. Control vitrified: 71.7 ± 5.5 %, p = 0.01) and the cleavage rate (CLA vitrified: 10.4 ± 2.5 % vs. control vitrified: 4.5 ± 2.5 %, p = 0.01) (Fig. 1)
Our results show a CLA effect (p < 0.001) on the water outflow rate induced by an osmotic challenge with sucrose, a non-permeant cryoprotectant currently used in oocyte cryopreservation media [17, 21]
Summary
Oocytes are subjected to extreme hyperosmotic conditions, inducing large volume changes that, along with an abrupt temperature drop, interfere with their developmental competence. Oocyte cryopreservation is increasingly in demand due to the need for preserving gametes of both humans and animals. The cumulative osmotic stress upon the cell due to water and cryoprotectant fluxes triggers abrupt cell volume changes that may compromise cell viability and cryopreservation success [3, 4]. The cryopreservation of female gametes shows a lower success rate. This problem may be partially due to the large oocyte size and shape, with a low surface to volume ratio. Membrane permeability plays an important cryobiological role in cell survival after vitrification [1, 3,4,5]
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