Effect Of Palmitic Acid On The Formation Of Neutrophil Extracellular Traps In Peripartum Dairy Cows

The perinatal period is the most important period of the cow’s entire production cycle.The increase in serum non-esterified fatty acid（NEFA）of cows may be the main cause of subacute inflammation in transition period.The concentration of palmitic acid（PA）changes greatly in the transition period and may be involved in inflammation and immunosuppression.The formation of neutrophil extracellular traps（NETs）is considered as an important immune response pathways of neutrophil.The goal of this study is determine whether the formation of NETs is a mechanism of palmitic acid-induced inflammation and immune response in transition cows.To determine whether PA can induce the formation of NETs,peripheral blood of cows（DIM 7-12）were taken to isolate polymorphonuclear（PMN）,the activity of lactate dehydrogenase and myeloperoxidase（MPO）was detected,the double-stranded DNA was quantified,and the morphology of NETs was observed by fluorescence microscopy（SYTOX Orange）and scanning electron microscope（SEM）,and the expression of of Akt,p-Akt,NE,Nox2,H3 and protein related to PI3K/Akt pathway was detected by Western blot.As a result,it was found that the average viability of the PMN was 97%and the purity of PMN was 96%.Compared with the control group,PA can induce a significant decrease in the activity of lactate dehydrogenase,a significant increase in the release of extracellular DNA,a significant increase in the extracellular double-stranded DNA content,and a significant increase in the number of NTEs structures.Meanwhile,PA can induce a significant increase in intracellular ROS and H₂O₂ levels in neutrophils,but L-Epicatechin significantly reduces this PA-induced ROS and H₂O₂ levels.PA can induce a significant increase in the level of MPO activity and L-Epicatechin can inhibit this PA-induced extracellular MPO levels.PA induced a significant increase in the expression levels of Nox2,Akt and p-Akt proteins,and L-Epicatechin inhibited this PA-induced increase in Nox2,Akt and p-Akt protein expression levels.Conclusion:Palmitic acid can lead to the formation of NETs.The mechanism of formation of PA-induced NETs may be that the PI3K/Akt pathway-associated protein was activated by PA,causing oxidative stress in neutrophils,leading to the formation of NETs.