Quadrupling efficiency in production of genetically modified pigs through improved oocyte maturation

Abstract

<p id="d2910463e278">The efficiency of production of genetically altered piglets for agricultural and biomedical purposes is very low, in large part because of the poor quality of the in vitro-matured oocytes used to initiate the process. Here, we describe a chemically defined culture medium that enhances porcine oocyte maturation and subsequent embryo development to the blastocyst stage, and increases the number of offspring born after embryo transfer, thereby effectively quadrupling the efficiency of piglet production. This research also provides some insights into the physiological events that lead to oocyte competence and how the efficacy of oocyte in vitro maturation might be improved in other species. </p><p class="first" id="d2910463e281">Assisted reproductive technologies in all mammals are critically dependent on the quality of the oocytes used to produce embryos. For reasons not fully clear, oocytes matured in vitro tend to be much less competent to become fertilized, advance to the blastocyst stage, and give rise to live young than their in vivo-produced counterparts, particularly if they are derived from immature females. Here we show that a chemically defined maturation medium supplemented with three cytokines (FGF2, LIF, and IGF1) in combination, so-called “FLI medium,” improves nuclear maturation of oocytes in cumulus–oocyte complexes derived from immature pig ovaries and provides a twofold increase in the efficiency of blastocyst production after in vitro fertilization. Transfer of such blastocysts to recipient females doubles mean litter size to about nine piglets per litter. Maturation of oocytes in FLI medium, therefore, effectively provides a fourfold increase in piglets born per oocyte collected. As they progress in culture, the FLI-matured cumulus–oocyte complexes display distinctly different kinetics of MAPK activation in the cumulus cells, much increased cumulus cell expansion, and an accelerated severance of cytoplasmic projections between the cumulus cells outside the zona pellucida and the oocyte within. These events likely underpin the improvement in oocyte quality achieved by using the FLI medium. </p>