ABSTRACT: The central goal of the following studies was to understand how FGF2, LIF, and IGF1, a cocktail called “FLI,” modulates embryo development and pregnancy outcomes in cattle. All embryos were produced in vitro with or without FLI supplementation at the beginning of culture. For each treatment, embryos were collected at the 4-6 cell, 9-16 cell, morula, or blastocyst stages and RNA was isolated and sequenced at a depth of 50 million reads per sample. In the FLI group, at the 9-16 cell stage, there were 7 upregulated and 6 downregulated differentially expressed genes (DEGs). Of those gene messages, ISG15 and USP18, mediators of interferon signaling were downregulated while message for MGLL, a gene involved in lipid regulation and the PI3K-ATK pathway was upregulated. At the morula stage, 1856 DEGs (580 upregulated in FLI) increased MAPK signaling, TGF-beta signaling, and Hippo signaling which all help regulate cell adhesion, lineage commitment, and growth regulation in the developing embryo. In FLI blastocyst stage embryo, 199 upregulated and 545 downregulated DEGs revealed an increase in processes associated with interferon gamma production and cell differentiation. Many of the downregulated DEGs are involved in MAPK signaling and WNT signaling pathways warranting further research of the fine regulation and interplay of these mechanisms. The addition of FLI at the beginning of culture initiates changes in the early embryo that are reflected during the entire preimplantation period. FLI modulates many of the regulatory pathways in the developing embryo to drive increased cell survival, cell integrity, and overall embryo development.
