Intracellular Staphylococcus aureus Infection Decreases Milk Protein Synthesis by Preventing Amino Acid Uptake in Bovine Mammary Epithelial Cells

Abstract

Staphylococcus aureus (S. aureus) is one of the main pathogens in cow mastitis, colonizing mammary tissues and being internalized into mammary epithelial cells, causing intracellular infection in the udder. Milk that is produced by cows that suffer from mastitis due to S. aureus is associated with decreased production and changes in protein composition. However, there is limited information on how mastitis-inducing bacteria affect raw milk, particularly with regard to protein content and protein composition. The main purpose of this work was to examine how S. aureus infection affects milk protein synthesis in bovine mammary epithelial cells (BMECs). BMECs were infected with S. aureus, and milk protein and amino acid levels were determined by ELISA after S. aureus invasion. The activity of mTORC1 signaling and the transcription factors NF-κB and STAT5 and the expression of the amino acid transporters SLC1A3 and SLC7A5 were measured by western blot or immunofluorescence and RT-qPCR. S. aureus was internalized by BMECs in vitro, and the internalized bacteria underwent intracellular proliferation. Eight hours after S. aureus invasion, milk proteins were downregulated, and the level of BMECs that absorbed Glu, Asp, and Leu from the culture medium and the exogenous amino acids induced β-casein synthesis declined. Further, the activity of mTORC1 signaling, NF-κB, and STAT5 was impaired, and SLC1A3 and SLC7A5 were downregulated. Eight hours of treatment with 100 nM rapamycin inhibited NF-κB and STAT5 activity, SLC1A3 and SLC7A5 expression, and milk protein synthesis in BMECs. Thus mTORC1 regulates the expression of SLC1A3 and SLC7A5 through NF-κB and STAT5. These findings constitute a model by which S. aureus infection suppresses milk protein synthesis by decreasing amino acids uptake in BMECs.

Crosstalk between integrin/FAK and Crk/Vps25 governs invasion of bovine mammary epithelial cells by S. agalactiae

“…Bacteria and BMECs were stained with Alexa Fluor® 594 Phalloidin. 64 Briefly, BMECs were placed on slides and incubated overnight. S. agalactiae were washed with PBS and stained with CFSE at room temperature for 20 min.…”

“…The samples were examined by TEM (Hitachi HT7700, Hitachi, Ltd., Tokyo, Japan) to detect intracellular bacteria. 64 …”

Bovine Milk Proteome: Milk Fat Globule Membrane Protein Is the Most Sensitive Fraction in Response to High Somatic Cell Count

“…The composition of casein plays a crucial role in determining the nutritional value and production yield of dairy products. With an elevated SCC and increasing severity of udder health, the composition of casein changes due to a decrease in amino acid uptake in bovine mammary epithelial cells and an increase in bacteria hydrolysis, resulting in a decrease in milk quality. Caseins do not directly participate in the udder inflammation process .…”