In Vitro Maturation with Leukemia Inhibitory Factor Prior to the Vitrification of Bovine Oocytes Improves Their Embryo Developmental Potential and Gene Expression in Oocytes and Embryos.

Meritxell Vendrell-Flotats,Manel Lopez-Bejar,Jonathan Lamarre,Marc Yeste,Iris Martínez-Rodero,Teresa Mogas,Tania García-Martínez

doi:10.3390/ijms21197067

Abstract

Oocyte cryopreservation has a significant impact on subsequent embryonic development. Herein, we investigated whether supplementing in vitro maturation medium with Leukemia Inhibitory Factor (LIF) prior to vitrification affects embryo development and gene expression at different embryo developmental stages. A panel of genes including maternal effect, epigenetics, apoptosis and heat stress was relatively quantified. The results show reduced cleavage rates after vitrification, regardless of the LIF treatment. Although not statistically different from control-vitrified oocytes, oocyte apoptosis and the blastocyst yield of LIF-vitrified oocytes were similar to their non-vitrified counterparts. Vitrification increased oocyte ZAR1, NPM2 and DPPA3 gene expression while its expression decreased in LIF-vitrified oocytes to similar or close levels to those of non-vitrified oocytes. With a few gene-specific exceptions, vitrification significantly increased the expression of DNMT3A, HDAC1, KAT2A, BAX and BCL2L1 in oocytes and most stages of embryo development, while comparable expression patterns for these genes were observed between LIF-vitrified and non-vitrified groups. Vitrification increased HSPA1A expression in oocytes and HSP90AA1 in 2-cell embryos. Our data suggest that vitrification triggers stage-specific changes in gene expression throughout embryonic development. However, the inclusion of LIF in the IVM medium prior to vitrification stimulates blastocyst development and several other developmental parameters and induces oocytes and embryos to demonstrate gene expression patterns similar to those derived from non-vitrified oocytes.

Highlights

Over the past several decades, the cryopreservation of bovine oocytes has become an important part of assisted reproductive technologies
A key factor determining blastocyst yield, can be directly affected by in vitro maturation [23]. This is significant in the context of cryopreservation, where a major obstacle to the use of vitrified oocytes is the impact of freezing on oocyte quality
Similar to the results reported by Mo et al [22], a significantly higher number of inner cell mass (ICM) cells and a clear trend towards higher hatching ability was consistently observed in blastocysts derived from oocytes matured in vitro with Leukemia Inhibitory Factor (LIF)

Summary

Introduction

Over the past several decades, the cryopreservation of bovine oocytes has become an important part of assisted reproductive technologies. The basic mechanisms behind the disturbance of oocyte processes through vitrification are likely to be complex and involve the disruption of cellular structures, such as the cellular skeleton and meiotic spindles; the premature release of cortical granules [5,6] leading to zona hardening and impairment of normal fertilization [7]; compromised mitochondrial integrity [8]; changes in oocyte gene expression [9,10,11], and the initiation of oocyte apoptosis [11] While these injuries threaten the viability and subsequent embryo development of oocytes, few reports have examined the relationship between gene expression and aberrant early embryonic development or on growth retardation of embryos derived from vitrified bovine oocytes

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