Work

Effects of a metabolizable protein supply on metabolic function of CD4+ T lymphocytes in mid-lactation dairy cows

Year: 2026

Type: article

Abstract: The objective of the present experiment was to determine the effects of MP supply on metabolic function of nonactivated (NA) and activated (AC) CD4 + T lymphocytes in mid-lactation dairy cows.Thirty-two multiparous Holstein cows with a mean ± SD lactation of 3.28 ± 0.99, milk yield 51.6 ± 6.1 kg/d, and milk protein 3.38% ± 0.47% were enrolled at 86 ± 18 DIM in a barn equipped with 16 bin feeders.Cows and bins (16 cows/8 bins per treatment) were randomly assigned to treatments.All cows were fed the same herd diet (MP = 3,006 g/d) during a 2-wk adaptation period before enrollment and a 1-wk covariate period following enrollment to allow acclimation to the bins and collect baseline measurements.Following the covariate period, cows received 1 of 2 treatments for 12 wk: diets formulated to be either adequate in MP supply (MPA; MP = 3,111 g/d) or MP inadequate (MPI; MP = 2,983 g/d).The difference in MP supply was achieved by including porcine blood meal in the MPA, whereas it was omitted in the MPI treatment.Blood was collected during wk 11 at 173 ± 18 DIM to isolate peripheral blood mononuclear cells followed by magnetic isolation of CD4 + T lymphocytes using bovine-specific monoclonal antibodies.Isolated CD4 + T lymphocytes from each cow were split into 2 tubes and randomly assigned to incubate in an assay medium (250 µL) or with a combination of phorbol 12-myristate 13-acetate (100 µL, 20 ng/mL) and ionomycin (10 µL, 1 µg/ mL) for 2 h at 37 °C to evaluate metabolic function under metabolically NA and AC states, respectively, using an extracellular flux analyzer.Mitochondrial and glycolytic functional kinetics were recorded for NA and AC CD4 + T lymphocytes based on real-time measurement of oxygen consumption rate (OCR), extracellular acidification rate (ECAR), and proton efflux rate (PER) under basal condi-tions and in response to complex V inhibitor (oligomycin A), a protonophore uncoupler (BAM 15), and complex I and complex III inhibitors (rotenone and antimycin A).Cows fed the MPI diet exhibited lower OCR leading to a reduced maximal respiration compared with MPA-fed cows.Feeding MPI diet decreased maximal respiration in both metabolically NA and AC CD4 + T lymphocytes.Feeding MPI diet decreased sparing respiratory capacity in both metabolically NA and AC CD4 + T lymphocytes.During the T-cell persistence assay, an interaction between diet and time within the assay was observed for ECAR and PER, indicating that CD4 + T cells from cows fed MPI showed a progressive decrease in aerobic glycolysis over the 1.50 to 73.0 min measurement period, whereas CD4 + T cells from cows fed MPA maintained higher glycolytic activity.Mitochondrial, glycolytic, and total ATP production rates were unaffected by MP supply; however, glycolytic ATP production rates were greater in AC than NA CD4 + T lymphocytes.These results indicate that suboptimal supply of MP reduced the metabolic fitness and persistence of CD4 + T lymphocytes, which might compromise immune responses during nutrient imbalance, such as early lactation or health challenges.

Source: Journal of Dairy Science

DOI: 10.3168/jds.2025-27476