Assessment of the zona pellucida microdissection on its thickness in mammalian embryos

The zona pellucida (ZP) is a dynamically changing object that plays an important role during the preimplantation stage of embryogenesis. The ZP thickness may affect the implantation success and pregnancy rate, it is considered as a prognostic factor in a number of studies. The study was aimed to assess the dynamic changes in the mouse embryonic ZP thickness after laser assisted hatching (LAH) that involved breaching the ZP integrity at the blastocyst stage. Femtosecond laser pulses were used to perform the zona microsurgery. The zona thickness was measured both at the stage of blastocyst microsurgery (~Е3.5, i.e. 3.5 days of embryogenesis) and at the hatching stage (~Е5). Significant differences in the ZP thickness were revealed in the control group of embryos: from 6.21 µm (Е3.5) to 5.4 µm (Е5). The changes in thickness from 6.6 µm (Е3.5) to 6.2 µm (Е5) observed in the group subjected to LAH were non-significant. Tracing the ZP thickness of a particular embryo from the blastocyst stage to the hatching stage made it possible to estimate the thinning coefficients in the experimental and control groups. The findings that indicate lower tensile strength of the zona in case of LAH can provide the basis for further research on the ZP properties in case of using the embryo cryopreservation protocols.