Short communication: amino acid supplementation and stage of lactation alter apparent utilization of nutrients by blood neutrophils from lactating dairy cows in vitro

Authors: GARCIA, M - University Of Maryland; Elsasser, Theodore; JUENGST, L - University Of Maryland; QU, Y - University Of Maryland; BEQUETTE, B - University Of Maryland; MOYES, K - University Of Maryland

Submitted to: Journal of Dairy Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/22/2016
Publication Date: 3/9/2016
Citation: Garcia, M., Elsasser, T.H., Juengst, L., Qu, Y., Bequette, B., Moyes, K.M. 2016. Short communication: amino acid supplementation and stage of lactation alter apparent utilization of nutrients by blood neutrophils from lactating dairy cows in vitro. Journal of Dairy Science. 99(5):3777-3783.

Interpretive Summary: Cells of the immune system play a major role in maintaining the health of dairy cows by going into the mammary gland when infection threatens and destroying minor populations of bacteria. A significant feature of how this cell-based immune system response occurs is the fact that these immune cells can function very well on alternative nutrients that provide energy to these cells for their function all the while not really compromising the nutrition needs of other tissues in the animal's body. One of these cell-specific nutrients is glutamine. While the impact of glutamine is well documented what is less well appreciated is how other more common nutrients impact the function of these cells. In order to better clarify how changes in nutrient status affects immune cells in cows at different stages of the milk production cycle, investigators at the University of Maryland and USDA-ARS Beltsville, MD harvested immune cells from the circulation of cows at several different stages of lactation. Culturing these cells outside the animal, researchers changed the balance of nutrients in the culture medium and investigated how the administration of a common bacterial toxin, endotoxin, affected the function of these cells. Across the board, cells treated with endotoxin released significant amounts of immune hormones characteristic of the inflammation observed in bacterial disease. An interesting finding was that these immune cells were highly resistant to being diverted from their main functions, moving towards the infection in response to the challenge (called chemotaxis) and surrounding and "eating" invading bacteria (called phagocytosis). The main metabolic feature that was associated with increasing the amount of amino acids in the media was the observation that as amino acid concentrations increased the use of the sugar glucose decreased apparently sparing it for use by other cells. The data suggest that nutritional intervention can play a major role in modifying the whole body response to infection by assisting in conserving energy during times of stress. The concept of timely nutritional support during or in prevention of disease may be of benefit as a collateral strategy to decrease the amount of antibiotics used to fight disease in dairy cows permitting the animal's own immune system to function more efficiently.

Technical Abstract: Glutamine is the preferred AA used by polymorphonuclear leukocytes (PMN) during the inflammatory response. However, the effect of other AA on bovine PMN response during inflammation and how this is altered by stage of lactation has not been fully elucidated. The objective of this study was to determine the effect of additional AA supplementation (pool of AA excluding Gln) on AA profiles, gene expression, and inflammatory function of PMN from dairy cows in early and mid lactation in vitro. We used 18 Holstein cows for this study. Polymorphonuclear leukocytes were isolated. Working solutions of AA (0 or 4 mM) and LPS (0 or 50µg/mL) were added to cell populations suspended in RPMI and incubated for 2h at 37°C. We used a subset of samples for gene and protein expression. Concentrations of AA in medium were determined using gas chromatography-mass spectrometry with norleucine as an internal standard. Apparent AA and glucose utilization were calculated by subtracting the concentration after from that of before incubation. Data were analyzed as a randomized block design. Challenge with LPS increased the expression of proinflammatory genes and AA supplementation decreased both the expression of some proinflammatory genes and the media concentrations of tumor necrosis factor-a. Neither stage of lactation, LPS challenge, nor AA supplementation altered the chemotactic or phagocytic abilities of PMN in vitro. Polymorphonuclear leukocytes supplemented with AA had greater concentrations and apparent utilization of most of the supplemented AA, whereas the unsupplemented group had greater apparent utilization of glucose. Alanine was not provided in the media but was present in spent media, and Ile, Gly, and Pro were greater in spent media than in media before incubation indicating synthesis of these AA. Regarding expression of genes involved in nutrient metabolism, the expression of G6PD, coding for the enzyme glucose 6-phosphate dehydrogenase, was increased and that of PDHA1, coding for the enzyme pyruvate dehydrogenase a 1, tended to increase with AA supplementation. Due to the lower concentration of tumor necrosis factor-a in media coupled with a downregulation of several proinflammatory genes, we concluded that AA, rather than Gln, alter the inflammatory response of bovine blood PMN. Independent from Gln, blood PMN from cows in early lactation may use certain AA as their primary carbon source for energy than cows in later lactation. Evaluating cows during the early postpartum period will provide additional information on the effect of stage of lactation and nutrient supplementation on PMN function.