# Glutamine alleviates Staphylococcus aureus-induced mastitis by modulating macrophage polarisation

## TL;DR

This study shows that glutamine and its metabolite αKG can reduce mastitis in mice by promoting anti-inflammatory macrophages and improving gut-mammary health.

## Contribution

The study reveals a novel gut-mammary axis mechanism where glutamine and αKG regulate macrophage polarization to combat mastitis.

## Key findings

- Oral glutamine reduces S. aureus-induced mastitis and protects the blood-milk barrier in mice.
- αKG promotes M2 macrophage polarization by enhancing autophagy and activating the JAK-STAT pathway.
- In vitro experiments confirm αKG's role in driving anti-inflammatory macrophage responses.

## Abstract

Mastitis, a major dairy industry challenge, is often caused by Staphylococcus aureus (S. aureus). Previous research linked gut microbiota disruption—induced by vancomycin in mice—to reduced microbial glutamine and increased mastitis susceptibility. This study hypothesizes that oral glutamine may regulate macrophage function and mitigate S. aureus mastitis. Our results showed that glutamine attenuated S. aureus-induced mastitis, protected the blood-milk barrier function, increased M2-type macrophages in the mammary gland, and increased the concentration of its metabolite α-Ketoglutaric acid (αKG) in both serum and mammary gland. In addition, the addition of αKG in mice significantly increased the transformation of M2-type macrophages and attenuated S. aureus-induced mastitis. Further studies revealed that αKG could initiate autophagy via oxidative phosphorylation, which in turn regulated the expression of Suppressor of cytokine signaling (SOCS3), a negative feedback inhibitor of the JAK/STAT signalling pathway. In vitro experiments verified that αKG enhances autophagy in macrophages, activates the JAK-STAT pathway, thereby promoting M2 polarization and alleviating S. aureus mastitis. This study revealed the key role of glutamine and its metabolite αKG in the mammary gland’s resistance to pathogenic infections, and reveal the mechanism by which they regulate macrophage polarization, providing theoretical guidance for solving the prevention and treatment problems of mastitis. This highlights a potential dietary intervention for mastitis management in dairy cows, bridging gut health and infection resistance.

Mastitis, a major challenge in the dairy industry, is commonly caused by S. aureus. This study demonstrates that oral glutamine supplementation alleviates S. aureus-induced mastitis in mice, protects the blood-milk barrier, and promotes an M2-polarized macrophage phenotype in mammary tissue. Its metabolite, α-ketoglutarate (αKG) enhances oxidative phosphorylation-driven autophagy, which downregulates SOCS3, a negative feedback inhibitor of the JAK-STAT pathway. This activation of JAK-STAT signaling promotes M2 macrophage polarization, thereby mitigating inflammation. In vitro experiments using RAW264.7 cells confirm that αKG enhances autophagy, activates JAK-STAT, and drives M2 polarization. These findings reveal a gut-mammary axis mediated by glutamine metabolism and highlight αKG as a critical immunometabolite that regulates macrophage function, offering a potential dietary strategy for mastitis prevention and therapy in dairy cows.