136 Long-term effects of maternal undernutrition on DNA methylation in skeletal muscle of postnatal Wagyu cattle.

Abstract We have previously reported that maternal undernutrition altered the metabolic system of postnatal skeletal muscle in Wagyu (Japanese Black) offspring (Nishino et al., 2025, accepted). The objectives of this study were to investigate the effects of maternal nutritional restriction on DNA methylation in postnatal offspring muscle to elucidate potential epigenetic regulatory mechanisms. Wagyu cows were divided into nutrient-adequate (control, CNT; n=4, 120% of requirements) and nutrient-restricted (NR; n=4; 60% of requirements) groups, and treated from day 35 of gestation until parturition. Diets were delivered on the basis of crude protein requirements, meeting 100% and 80% of dry matter requirements in the CNT and NR groups, respectively. All offspring were provided with the same diet from birth to 300 days of age (d) and biopsy samples of the longissimus thoracis muscle (LM) were collected for analysis. Maternal undernutrition dramatically impacted DNA methylation patterns in LM of 300 d-offspring. The NR offspring LM had 7,173 hypo-differentially methylated regions (DMR) and 6,195 hyper-DMR. Enrichment analysis revealed that genes with hypo-DMR in their promoter regions or upstream of promoters were associated with cell structural organization and neurodevelopment, while genes with hyper-DMR were involved in protein catabolic process and cell development. Previously, we demonstrated that maternal nutritional restriction reduced tricarboxylic acid (TCA) cycle-related metabolites, including nicotinamide adenine dinucleotide (NADH) and succinate (Nishino et al., 2025, accepted). This was accompanied by decreased expression of genes essential for mitochondrial energy production in skeletal muscle, including nicotinamide phosphoribosyltransferase (NAMPT), NADH: ubiquinone oxidoreductase subunit A12 (NDUFA12), and NADH dehydrogenase subunit 5 (ND5), pyruvate dehydrogenase E1 subunit alpha 1 (PDHA1), and dihydrolipoamide dehydrogenase (DLD). However, the present study found that maternal undernutrition did not significantly affect DNA methylation levels of NAMPT, NDUFA12, ND5, PDHA1, and DLD in 300-d offspring LM. These results indicate that maternal nutritional restriction has long-term effects on biological processes that are fundamentally established during the fetal period. However, no clear relationships were found between DNA methylation and gene expression of key metabolic regulators, suggesting the involvement of more complex regulatory networks. Future studies should focus on both the indirect effects of DNA methylation and the potential role of other epigenetic modifications to explore the underlying regulatory mechanisms that govern these adaptive responses.