Abstract
Non-esterified fatty acids (NEFAs) such as oleic acid (OA) and linoleic acid (LA) are associated with a higher incidence of infectious diseases such as metritis and mastitis during the bovine peripartum. Fatty acids can induce an increase in the release of ATP, and changes in the expression levels of purinergic receptors in bovine polymorphonuclears (PMN) during peripartum have also been reported. PMN respond to inflammatory processes with production of ROS, release of proteolytic and bactericidal proteins, and formation of neutrophil extracellular traps (NETs). NETs formation is known to require ATP production through glycolysis. Studies have shown that the above-mentioned metabolic changes alter innate immune responses, particularly in PMN. We hypothesized that NEFAs induce the formation of NETs through ATP release by Pannexin 1 and activation of purinergic receptors. In this study, we found that OA and LA induce NET formation and extracellular ATP release. Carbenoxolone, a pannexin-1 (PANX1) inhibitor, reduced OA- and LA-induced ATP release. We also found that P2X1, P2X4, P2X5, P2X7, and PANX1 were expressed at the mRNA level in bovine PMN. Additionally, NEFA-induced NET formation was completely abolished with exposure to NF449, a P2X1 antagonist, and partially inhibited by treatment with etomoxir, an inhibitor of fatty acid oxidation (FAO). Our results suggest that OA and LA induce NET formation and ATP release via PANX1 and activation of P2X1. These new data contribute to explaining the effects of NEFA high concentrations during the transition period of dairy cattle and further understanding of pro-inflammatory effects and outcome of postpartum diseases.
Highlights
In the transition period of dairy cows, which includes 3 weeks pre- and 3 weeks postpartum, there is a high demand for nutrients and a decrease in feed ingestion, producing a transient negative energy balance [1]
These results suggested that the formation of neutrophil extracellular traps (NETs) that was induced by short-term culturing with these fatty acids was independent of FFA1 and NADPH oxidase
We evaluated whether Non-esterified fatty acids (NEFAs) could induce NET release in bovine PMN and found that oleic acid (OA) and linoleic acid (LA) can induce the formation of NETs staining positive for H4 citrulline 3 (H4Cit3)
Summary
In the transition period of dairy cows, which includes 3 weeks pre- and 3 weeks postpartum, there is a high demand for nutrients and a decrease in feed ingestion, producing a transient negative energy balance [1] This period is characterized by hypoglycemia, mobilization of non-esterified fatty acids (NEFAs), and an increase in plasma beta-hydroxybutyrate levels [2, 3]. Cows with high blood levels of NEFAs during peripartum have altered polymorphonuclear (PMN) trafficking and phagocytosis, as well as a reduced ability to kill microorganisms [8] This suggests that NEFAs can contribute to the modulation of the innate immune response, leading to the onset of metabolic and infectious diseases [8, 9]
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