Effect of the exposure to methyl-β-cyclodextrin prior to chilling or vitrification on the viability of bovine immature oocytes

Abstract

The present study aimed to evaluate the effect of methyl-β-cyclodextrin (MβCD) as a cholesterol loader to change oocyte plasma membrane and increase its tolerance toward cryopreservation. The first and second experiments were conducted to investigate if MβCD could improve nuclear and cytoplasmic maturation after oocyte exposure to cold stress for 10 or 30 min, respectively. No differences (P>0.05) in either experiment in the metaphase II (MII) rate of oocytes exposed to MβCD and cold stress; but these oocytes presented lower maturation rates than control groups. In the second experiment, a lower percentage of oocytes showed degenerated chromatin (P<0.05) after exposure to 2mg/mL of MβCD compared to the group exposed to 0mg/mL. However, no differences among treatments were observed in cytoplasmic maturation. Groups exposed to cold stress demonstrated a lower (P<0.05) capacity for embryonic development compared to the control groups. In the third experiment immature oocytes were exposed to MβCD and then, vitrified (cryotop). After warming, we observed that the ability to reach MII and chromatin degeneration were altered (P<0.05) by MβCD. The blastocysts rate (P<0.05) on D7 was higher in the 2 mg/mL MβCD group, but an identical finding was not observed on D8 (P>0.05). Chromatin degeneration was higher in the vitrification groups. We conclude that MβCD improved nuclear maturation by reducing oocyte degeneration after cold stress or vitrification; however, more studies are required to clarify the usefulness of MβCD use in oocyte cryopreservation.

Free cholesterol and cholesterol esters in bovine oocytes: Implications in survival and membrane raft organization after cryopreservation

“…In the first study [14], the authors showed that cholesterol diffused through cumulus cells and reached the oocyte but failed to reflect these effects in the production of embryos. The same methodology was applied later in immature bovine oocytes [15]. In this case, both the presence of serum and the compact structure of the cumulus of immature oocytes could have limited the availability of MβCD/cholesterol complexes, affecting the results collected.…”

“…Modulation of plasma membrane cholesterol to increase post-cryopreservation survival is currently a booming topic for the male gamete of different species (for review, see [13]) but has been little addressed in oocytes. Previously, Horvath and Seidel [14] and, more recently, Spricigo et al [15] reported the use of this method to add cholesterol to bovine oocytes prior to vitrification. Horvarth and Seidel [14] claimed that cholesterol could enter mature oocytes through the cumulus cells and the zona pellucida , improving its viability after vitrification in a chemically defined system.…”

“…Horvarth and Seidel [14] claimed that cholesterol could enter mature oocytes through the cumulus cells and the zona pellucida , improving its viability after vitrification in a chemically defined system. Spricigo et al [15] improved nuclear maturation after treating immature bovine oocytes with cholesterol loaded MβCD prior to vitrification. However, these effects were not reflected in the production of embryos in vitro [14,15].…”

“…Spricigo et al [15] improved nuclear maturation after treating immature bovine oocytes with cholesterol loaded MβCD prior to vitrification. However, these effects were not reflected in the production of embryos in vitro [14,15]. Whereas Horvath and Seidel [14] analyzed the presence and absence of serum in their assays, Spricigo et al [15] used fetal calf serum in order to load MβCD with cholesterol from serum, and also during the recovery of cumulus -oocyte complexes (COCs), in vitro maturation and vitrification.…”

“…However, these effects were not reflected in the production of embryos in vitro [14,15]. Whereas Horvath and Seidel [14] analyzed the presence and absence of serum in their assays, Spricigo et al [15] used fetal calf serum in order to load MβCD with cholesterol from serum, and also during the recovery of cumulus -oocyte complexes (COCs), in vitro maturation and vitrification. More recently, rabbit oocytes with increased cholesterol content by exposure to MβCD/cholesterol complexes were found to fail to improve their developmental competence after either vitrification or slow freezing [16].…”