Assessment of a new in vitro maturation system for mouse and human cumulus-enclosed oocytes: three-dimensional prematuration culture in the presence of a phosphodiesterase 3-inhibitor

Background: Controlling nuclear maturation during oocyte culture might improve nuclear-cytoplasmic maturation synchrony. In the present study, the quality of mouse and human cumulus-enclosed oocytes (CEOs) was examined after a two-step culture consisting of a three-dimensional prematuration culture (3D-PMC), followed by in vitro maturation (IVM). Methods: Mouse and human CEOs were embedded in an extra-cellular matrix (Collagen gel Type I). The gels containing the CEOs were cultured in medium with a phosphodiesterase 3-inhibitor (PDE3-I; cilostamide 1 µM) for 24 h. Afterwards, CEOs were removed from the gel, washed away from inhibitor and underwent IVM. The optimal concentration of collagen (diluted 1:2 versus undiluted) was first determined in a mouse model. Cytoplasmic maturation after IVM of human and mouse oocytes was assessed in relation to fertilization and embryonic developmental capacity.Results: The diluted form of collagen supported better the structure of the expanding CEOs and meiotic competence of the oocytes. Electron microscopy in combination with Lucifer Yellow dye coupling assay revealed that oocyte-cumulus cell connections could be preserved during 3D-PMC. Percentages of mouse 2-cell embryos upon IVF were higher in the 3D-PMC group compared to in vitro controls and 2D-PMC oocytes, but lower compared to in vivo controls. In the human model, percentages of polar body-extruded oocytes were significantly higher in the 3D-PMC group compared to conventionally matured oocytes. Three-dimensional PMC had also a beneficial effect on embryonic development on day 3 post-ICSI.Conclusions: Applying a 3D-PMC in the presence of a PDE3-I preserve oocyte-cumulus cell connections and influences oocyte developmental capacity.