(P16-023-23) Mammary Adipocytes Are a Source of Murine Breast Milk Glutamine

Objectives: Twenty percent of the amino acids found in milk, as protein and free amino acids, are glutamine and glutamate. As lactation progresses, free glutamine and glutamate in milk increase by 350% and 40%, respectively. In pigs, up to 50% of milk glutamine is produced within the mammary gland by glutamine synthetase (GS). However, milk glutamine levels may not be optimal because supplementing glutamine to piglets or lactating sows improves growth performance, gut development, and immunity in neonates. Branched-chain amino acids (BCAA) are believed to be precursors for endogenous glutamine synthesis. This work examined the changes of GS and BCAA metabolizing enzymes (BCAA transaminase 2—BCAT2 and branched chain ketoacid dehydrogenase—BCKDHA) in mouse mammary gland during lactation.

Methods: Mouse mammary glands (n=5 unless otherwise noted) were sampled at four time points: 8-week-old virgin/non-lactating (control), post-delivery day 5 (early lactation, PD5), PD15 (peak lactation), and involution (4 days after weaning at PD21, n=4). Samples were subject to RT-qPCR and immunoblotting for GS, BCAT2, and BCKDHA. GS localization within the mammary gland was visualized by immunohistochemistry staining.

Results: GS mRNA expression and protein level were significantly lower at PD5 and PD15 compared with the control. During involution, these levels were restored. Results from immunohistochemistry staining revealed that GS localized in mammary adipocytes with no staining in mammary epithelial cells (MEC). During lactation, the ratio of GS-positive adipocytes to GS-negative MEC decreases, which explains the lower GS abundance seen at PD5 and PD15. Compared with the control, mRNA and protein levels of BCAT2 and BCKDHA significantly decreased during lactation and involution.

Conclusions: Overall, this work demonstrates the importance of mammary adipocytes in milk amino acid synthesis and suggests a unique crosstalk between adipocytes and MEC within the mammary gland regarding milk production and secretion. It also lays a foundation for future studies that will inform clinical practices to improve maternal diets and/or make changes to formula compositions, ultimately improving the health outcomes of both mothers and neonates.

Funding Sources: USDA, NIH