112 Growth factor receptors in bison blastocysts and expanded blastocysts

Assisted reproductive technologies can be used in bison for managing genetics and mitigating disease risks for commercial and conservation purposes. In vitro embryo production (IVP) provides an option for exchanging genetics between herds that avoids the stress and biosecurity risks of moving live animals. Systems to produce cattle embryos have been used to generate bison embryos but are roughly half as efficient; IVP rates are 25–30% per oocyte for cattle versus 10% for bison. Thus, cattle IVP systems are not optimised for bison. One approach to addressing this insufficiency is evaluating bison embryos’ specific needs across developmental stages. The goal of this experiment was to identify specific growth factor receptors (GFR) on bison blastocysts (BL) and expanded blastocysts (EXB) as a first step to developing a bison-specific media system. Abattoir-derived oocytes were used to generate embryos using a standard IVP system. Briefly, Grade 1 and 2 oocytes were matured for 23 h then fertilised with frozen-thawed bison sperm at 0.5 × 10⁶ sperm/mL. Sperm and oocytes were co-incubated for 18 h, and presumed zygotes were stripped of their cumulus cells and cultured for 56 h in CDM1 at 38.5°C in a humid tri-gas incubator. After 56 h, embryos were evaluated for developmental progression and all embryos with more than four cells cultured until Day 7 in CDM-2 at 38.5°C in a humid tri-gas incubator. On Day 7, embryos were evaluated for stage and grade. Grade 1 and 2 BL and EXB were snap frozen in groups of 10 (n = 30 for each stage across three replicates) in RIPA buffer. JESS capillary Western blotting was used to analyse protein expression. Protein expression of each GFR was normalised against total protein. GFRs were considered present if they had more than 50 relative units. All antibodies against the targets were first validated using bison follicular cells. Embryos were evaluated for the presence of 16 GFRs previously detected in embryos of other species. Six GFRs were detected in BL and EXB: FGFR1, FGFR2, IGF1R, LIFR, BMPR2, and GCSF-R. FGFR1, LIFR, BMPR3, and GCSF-R were more highly expressed in EXB than BL. IGF1R and FGFR2 were more highly expressed in BL than EXB. The growth factors present are involved in cell division, regulation of cell growth, and maturation. GCSF-R plays a role in implantation and pregnancy maintenance. Ongoing experiments to determine the expression of GFR in earlier stages of development, including in bison oocytes, are in progress. These data will be considered with the data presented here to design a follow-up experiment evaluating the supplementation of bison culture media with growth factors at specific stages of development with the ultimate goal of producing a bison-specific media system. Improving in vitro production of bison embryos has direct applicability to preserving bison genetics and other conservation and commercial applications.