A Brief Incubation of Cumulus-Enclosed Mouse Eggs in a Calcium-Free Medium Containing a High Concentration of Calcium-Chelator Markedly Improves Preimplantation Development.

Valeria Merico,Silvia Garagna,Maurizio Zuccotti

doi:10.3390/ijerph17103505

Abstract

The presence of cumulus cells (CCs) surrounding ovulated eggs is beneficial to in vitro fertilization and preimplantation development outcomes in several mammalian species. In the mouse, this contribution has a negligible effect on the fertilization rate; however, it is not yet clear whether it has positive effects on preimplantation development. Here, we compared the rates of in vitro fertilization and preimplantation development of ovulated B6C3F1 CC-enclosed vs. CC-free eggs, the latter obtained either after a 5 min treatment in M2 medium containing hyaluronidase or after 5–25 min in M2 medium supplemented with 34.2 mM EDTA (M2-EDTA). We found that, although the maintenance of CCs around ovulated eggs does not increment their developmental rate to blastocyst, the quality of the latter is significantly enhanced. Most importantly, for the first time, we describe a further quantitative and qualitative improvement, on preimplantation development, when CC-enclosed eggs are isolated from the oviducts in M2-EDTA and left in this medium for a total of 5 min prior to sperm insemination. Altogether, our results establish an important advancement in mouse IVF procedures that would be now interesting to test on other mammalian species.

Highlights

In most eutherian mammals, the metaphase II oocyte is ovulated, surrounded by layers of cumulus cells (CCs) embedded in an extracellular matrix of hyaluronic acid
The presence of CCs surrounding ovulated eggs has a negligible effect on the in vitro fertilization rate [23], but it still remains unclear whether they are required at the time of fertilization for improved preimplantation developmental competence or if the procedure with the enzyme hyaluronidase, routinely used for the removal of CCs, may result in a drawback of their early developmental potential
The results of our investigation indicate that the maintenance of CCs around the ovulated egg is beneficial to the outcome of mouse in vitro preimplantation development

Summary

Introduction

The metaphase II oocyte (egg) is ovulated, surrounded by layers of cumulus cells (CCs) embedded in an extracellular matrix of hyaluronic acid. The presence of CCs surrounding the egg is not strictly required for successful in vitro fertilization (IVF), suggesting that they are dispensable, a number of studies have underlined the beneficial effect of their presence on both rates of fertilization and preimplantation development in pigs [13,14,15], buffalos [16], and cattle [17,18,19,20,21,22]. The presence of CCs surrounding ovulated eggs has a negligible effect on the in vitro fertilization rate [23], but it still remains unclear whether they are required at the time of fertilization for improved preimplantation developmental competence or if the procedure with the enzyme hyaluronidase, routinely used for the removal of CCs, may result in a drawback of their early developmental potential

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