Effect of zona pellucida removal on early development of in vitro produced bovine embryos

Abstract

SUMMARYDuring early embryo development the zona pellucida acts as a barrier against polyspermia and guarantees communication between blastomeres before and during compaction. However, the development of new technologies of embryo production such as "Handmade Cloning" demands removal of this membrane. The aim of this study was to determine the effect of zona pellucida removal on in vitro bovine embryo development. First, the consequences of zona pellucida removal was assessed by comparing the percentage of first cleavage, percentage of blastocysts and cell number among zona-included and zona-free embryos; either through the removal of the zona pellucida immediately after IVF or parthenogenesis. Embryo development was also evaluated when zona pellucida was removed before parthenogenesis. In a second set of experiments, the gene expression levels of BAX, BCL2, CASP3, CDH1, OCT4 and SOX2 were evaluated in zona-free and zona-included IVF-derived embryos. No significant differences were found in the percentage of first cleavage, percentage of blastocyst and cell number on IVF-embryos cultured with or without zona. Parthenogenetic embryos followed the same general pattern even when the zona pellucida was eliminated before activation; however there was a significant increase in the rate of first cleavage when the zona pellucida was removed after activation but this did not impact upon further development. Furthermore, no significant differences in gene expression level were found between zona-free and zona-included IVF-embryos for the studied genes. We concluded that the lack of zona pellucida did not affect the early development when an appropriate system is used for embryo culture to ensure blastomeric contact and normal compaction.

In vitro versus In vivo: Development-, Apoptosis-, and Implantation-Related Gene Expression in Mouse Blastocyst

“…According to obtained results, hatching rate was significantly lower in the IVF-derived embryos than in the in vivo control. Although a correct hatching process occurs normally in a good quality blastocyst, it is shown that in vitro conditions cause zona hardening, and disturb the hatching ability of a produced embryo(39). Furthermore, the success of in vitro hatching is dependent on a sufficiently high number of embryonic cells(40) as well as trophectodermal lysins (41).…”

The transfer of parthenogenetic embryos following artificial insemination in cows can enhance pregnancy recognition via the secretion of interferon tau

“…Indeed, the cell cycle length of the PA embryos appeared to be longer than that of the IVF embryos [23]. PA embryos produced by ethanol activation (a method similar to that of the present study) have few cells [24], and the apoptotic indexes of PA embryos are higher than that of IVF embryos [25]. (2) It is possible that the PA embryos had only a small amount of the enzyme that melts the zona pellucida.…”

The use of a two-step removal protocol and optimized culture conditions improve development and quality of zona free mouse embryos

“…Previous studies have established that bovine zona removal is most effectively achieved using 0.2% proteinase K for 5 min and that this process exhibited low toxicity [19]. In contrast, treatment with 0.5% proteinase K took approximately 6.8 min to completely remove the ZP in mouse embryos suggesting that it is not as efficient in mice.…”