194 Methionine and guanidinoacetic acid supplementation influence bovine oocyte quality during in vitro maturation

During oocyte maturation and early embryonic development, methylation can influence the quality of the oocyte and subsequent embryo. If methylation is reduced, then abnormal oocyte maturation and reduced embryo quality can occur. Methionine (Met) is an important one-carbon metabolite that can increase methylation, while guanidinoacetic acid (GAA) sequesters methyl groups, leading to global methylation deficiency. We hypothesized that supplementation of GAA will lead to methyl deficiency and incorporation of Met will improve methylation status in the matured oocyte. Follicles (2–7mm) were aspirated from slaughterhouse ovaries to obtain cumulus–oocyte complexes (n = 800 per treatment) and matured for 23 h in maturation medium containing Control, Met (21 µM), GAA (20 mM), or Met + GAA. After maturation, a subset of oocytes were randomly selected for transcript abundance analysis of markers associated with methylation and oocyte quality, 5-methylcytosine analysis, and chromatin and microtubule analysis. The remaining oocytes underwent IVF with sex-sorted male semen, followed by in vitro embryo culture. Cleavage and blastocyst rates, microtubule distribution, and chromatin configuration were collected and analyzed using the GLIMMIX procedure of SAS, while 5-methylcytosine and transcript abundance was analyzed using the MIXED procedure of SAS, with the main effect of supplementation as fixed effects and replication as random effect. Compared with the control group, 5-methylcytosine was reduced when Met and GAA were supplemented alone and increased when they were supplemented together (P < 0.05). Transcript abundance of BMP15, CEPT1, and COX2 were significantly reduced (P < 0.05) and a tendency for reduction was observed in CHPT1 and CPEB4 in the GAA group compared with the control group. A significant interaction (P < 0.05) between Met and GAA was observed for DNMT3A and DNMT3B, with a reduction in the Met and GAA supplementation alone and an increase in Met + GAA supplementation. Microtubule distribution abnormalities were increased (P < 0.05) with GAA supplementation compared with the control group, while no differences were observed for the combined supplementation. No differences were observed in Met, GAA, or the Met + GAA for cleavage or blastocyst rate. Taken together, Met and GAA appears to have a negative impact on global methylation and oocyte quality. The negative impact observed with Met or GAA alone is diminished in the oocytes when Met is combined with GAA. Using this model, we can begin to understand the implications of supplementing one-carbon metabolites during IVM and start to understand long-term implications on early embryonic development.

USDA is an equal opportunity employer and provider.