Effects of L-leucine and sodium acetate on milk protein synthesis under heat stress conditions in bovine mammary epithelial cells in vitro

It is widely known that heat stress (HS) has negative effects on dairy cows, such as a reduction in milk production and milk protein. However, there has been no research yet on the effects of HS at the bovine mammary epithelial cells (MAC-T) level and the function of L-leucine (LEU) and sodium acetate (ACE) in reducing HS. In this study, we evaluated the negative effects of HS at various temperatures on MAC-T and verified whether LEU and ACE are effective at reducing HS and increasing protein synthesis. An experiment was conducted by dividing MAC-T into three groups: 39 °C, 41 °C, and 43 °C. In the case of LEU and ACE supplementation experiments, the cells were supplemented with 0, 0.45, 0.9, 1.8, and 3.6 mM of LEU and ACE to reach the differentiation medium. It was observed that under HS at 41 °C, HSP70, BAX, and eIF4EBP1 gene expression were increased, whereas Bcl-2, eIF4E, and PRKAA1 gene expression were decreased. When 1.8 mM of LEU was added under HS at 41 °C, it suppressed apoptosis by reducing the gene expression of HSP70 and controlling the gene expression of apoptosis-related genes such as BAX and Bcl-2. Additionally, mTOR, P-mTOR, and β-casein proteins were increased. In the case of 0.9 mM of ACE, it was found to decrease the gene expression of HSP70 and BAX and increase the amount of β-casein protein synthesis. Simultaneous supplementation of LEU and ACE has been shown to reduce HS, inhibit apoptosis, and increase β-casein protein expression. In summary, HS at 41 °C began to have a negative effect on MAC-T, while LEU and ACE reduced HS and inhibited apoptosis, alleviating cell damage and effectively increasing β-casein protein synthesis. The results suggest that LEU and ACE have the potential to reduce HS and promote protein synthesis under HS conditions in MAC-T.