Glucose metabolism by bovine neutrophils characterized by mass spectrometry and [13C6]glucose

Abstract Text:

At the present time, the metabolic priority for and subsequent pathway fate of glucose by bovine blood neutrophils (BBN) is unknown.  During early lactation, glucose availability is low.  Consequently, a resulting energy-compromised state may partly explain the lack of robust immune response observed at this time.  The objective of this study was to characterize the metabolic utilization of glucose by BBN from dairy cows in early and mid-lactation by incubation of BBN with [¹³C₆] glucose followed by gas chromatography–mass spectrometry (GCMS).  Jugular blood (~150 mL) was collected from four healthy multiparous dairy cows in early (n = 2; < 21 DIM) and mid-lactation (n = 2; > 90 DIM). BBN were isolated and adjusted to a final concentration of 6 × 10⁶ cells/mL using RPMI/5% fetal bovine serum containing 4 mM of a 50:50 mix of unlabeled and ¹³C-labeled glucose.  Cells were incubated for 2 h at 37^◦C, 95% humidity, and 5 % CO₂.  Non-essential amino acids in metabolic equilibrium with Krebs cycle intermediates and lactate in BBN were monitored by GCMS.  Data were analyzed by the student’s t-test for unpaired comparison.  Significance was declared at P ≤ 0.05.  No stage of lactation effect was observed.  Although 43% of the pyruvate pool derived from glucose, a small proportion of this pool entered the Krebs cycle. Our results suggest that the pyruvate carboxylase flux was minimal and only 5% of the acetyl-CoA pool was derived from the pyruvate pool.  In conclusion, although BBN exhibit high rates of glycolysis, glucose is not the main substrate oxidized for energy in the Krebs cycle and perhaps other extracellular substrates (e.g. fatty acids, glutamine, aspartate) are preferred.  Future studies are needed to examine the metabolic utilization of other substrates and their influence on the function of BBN. 

Keywords:  cow, glucose, neutrophil