PSIV-18 Combination of 1.8 Mm Choline and Follistatin Supplementation May Reduce Transcript Abundance of CEPT1 and CPBE4 During In-Vitro Bovine Oocyte Maturation

Abstract

Abstract One-carbon metabolite supplementation during in vitro embryonic development improves blastocyst quality; however, our understanding of the effects of combining these metabolites during in vitro maturation (IVM) is limited. Supplementation with follistatin improved oocyte quality following IVM. Choline supplementation was beneficial during early embryonic development; however, the role of choline during oocyte maturation is unknown. The objective of this study was to investigate the combination of choline and follistatin during IVM on oocyte quality. We hypothesized that supplementation of choline with follistatin would synergistically improve oocyte quality. Follicles (2 to 7mm) were aspirated from slaughterhouse ovaries to obtain cumulus oocyte complexes for IVM that were treated with choline (0, 1.3, or 1.8 mM) and follistatin (0 or 10 ng/mL) in a 3 x 2 design for 23 hr and visually graded for maturation. Cumulus cells and oocytes were used for qPCR transcript abundance analysis (cumulus cells: CHPT1, CX43, EFGR1, FST and STAR; oocytes: BMP15, GDF9, CEPT1, CPBE4, MTHFR, NANOS, CX43 and COX2). Oocyte maturation was analyzed using the GLIMMIX procedure of SAS and transcript abundance was analyzed using the GLM procedure of SAS with the main effect of choline, follistatin and the interaction as fixed effects. There were no differences in the visual characteristics of the oocytes during maturation (P > 0.1). No differences in transcript abundance were observed with choline or follistatin in the cumulus cells (P > 0.1). Addition of 1.3 mM choline during IVM may increase CPEB4 (P = 0.1) in oocytes compared with controls. Transcript abundance of CEPT1 tended to be reduced in oocytes supplemented with 1.8 mM choline and follistatin compared with control oocytes (P = 0.07). Incorporation of follistatin with 1.8 mM choline supplementation during IVM tended (P = 0.08) to reduce CPEB4 in oocytes compared with controls. Other markers of oocyte quality (BMP15, GDF9, NANOS, CX43 and COX2) were not different between choline or follistatin. In contrast to the hypothesis, the combination of choline and follistatin appeared to reduce overall in vitro oocyte quality. While no statistical differences were observed for the other markers, it appears that either 1.3 mM choline or follistatin supplementation may provide a beneficial impact on oocyte quality. Choline 1.3 mM supplementation may improve cell cycle progression from MI to MII phase through the up-regulation of CBEP4. This further supports the conclusion that choline is beneficial for in vitro oocyte development. Although follistatin appears to antagonize the actions of choline during oocyte maturation, they target different mechanisms (TGFβ signaling and DNA methylation, respectively) and both provide long-term benefits during in-vitro embryonic development. Further research is, therefore, warranted to investigate the antagonizing relationship between choline and follistatin during IVM and long-term impacts on blastocyst development. USDA is an equal opportunity provider and employer.