Abstract

Orotic acid (OA) is known to promote cell proliferation and influence triglyceride (TG) and protein metabolism in several animal species and cell types. However, its biological function in ovine mammary epithelial cells (OMECs) remains unclear. In this study, we evaluated the effects of OA on cell proliferation, oxidative stress, and synthesis of lactose, milk fat, and protein, and the explored potential regulatory mechanism using transcriptomic analysis. Treatment with 30 µg/mL of OA significantly promoted OMEC proliferation. OA also improved oxidative stress related indicators, increasing CAT (14.71 ± 3.27 VS 6.90 ± 1.40 U/106cell, P < 0.05) and SOD (7.08 ± 0.86 VS 4.18 ± 1.32 U/106cell, P < 0.05) activates, and reducing MDA (38.55 ± 12.06 VS 80.19 ± 12.34 U/106cell, P < 0.05). TG concentrations increased significantly both in cells (12.58 ± 1.86 VS 2.69 ± 0.23 µg/106 cells, P < 0.05) and culture supernatant (8.54 ± 0.62 VS 2.82 ± 0.12 ng/mL, P < 0.001). In contrast, OA significantly decreased the CSN2 concentration in the supernatant (24.41 ± 0.98 VS 34.62 ± 0.59 ng/mL, P < 0.001). Transcriptomic analysis identified 365 differentially expressed genes (DEGs), including 94 up-regulated and 271 down-regulated genes in the OA treated group. Up-regulated DEGs were enriched in the JAK-STAT signaling pathway and so on, while down-regulated DEGs were enriched in the protein digestion and absorption and so on. DEGs such as RPS6KA2 and FOXO4 were associated with cell proliferation, and ALDOC, SULT2B1, ADRB1, STAT5A were associated with fat metabolism. These findings suggest that OA enhances cells proliferation, improves oxidative stress indicators, and promotes TG synthesis in OMECs. The study provides a useful reference for understanding the regulation mechanism of lactation traits and for dairy sheep breeding.