1. Academic Validation
  2. β-Hydroxybutyrate impairs the release of bovine neutrophil extracellular traps through inhibiting phosphoinositide 3-kinase-mediated nicotinamide adenine dinucleotide phosphate oxidase reactive oxygen species production

β-Hydroxybutyrate impairs the release of bovine neutrophil extracellular traps through inhibiting phosphoinositide 3-kinase-mediated nicotinamide adenine dinucleotide phosphate oxidase reactive oxygen species production

  • J Dairy Sci. 2022 Apr;105(4):3405-3415. doi: 10.3168/jds.2021-21174.
Siyuan Liu 1 Xiaobing Li 2 Xiaohan Zhou 1 Juan J Loor 3 Qianming Jiang 3 Xiancheng Feng 1 Yuchen Yang 1 Lin Lei 1 Xiliang Du 1 Xinwei Li 1 Wang Zhe 1 Yuxiang Song 4 Guowen Liu 5
Affiliations

Affiliations

  • 1 State Key Laboratory for Zoonotic Diseases, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun, Jilin Province, 130062, China.
  • 2 College of Veterinary Medicine, Yunnan Agricultural University, Kunming 650201, China.
  • 3 Mammalian NutriPhysioGenomics, Department of Animal Sciences and Division of Nutritional Sciences, University of Illinois, Urbana 61801.
  • 4 State Key Laboratory for Zoonotic Diseases, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun, Jilin Province, 130062, China. Electronic address: [email protected].
  • 5 State Key Laboratory for Zoonotic Diseases, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun, Jilin Province, 130062, China. Electronic address: [email protected].
Abstract

Ketosis in dairy cows often occurs in the peripartal period and is accompanied by immune dysfunction. High concentrations of β-hydroxybutyrate (BHB) in peripheral blood during ketosis inhibits the release of neutrophil extracellular traps (NET) and contributes to immune dysfunction. However, the mechanisms whereby BHB affects NET release remains unclear. In this study, 5 healthy peripartal dairy cows (within 3 wk postpartum) with serum BHB concentrations <0.6 mM and glucose concentrations >3.5 mM were used as blood donors. Blood samples were collected before feeding, and the isolated polymorphonuclear neutrophils were incubated with 3 mM BHB for different times. Inhibition of Cit-H3 (citrullinated histone 3) protein abundance, a marker of NET activation, in response to BHB was used to determine an optimal incubation time for in vitro experiments. Four hours was selected as the optimal duration of BHB treatment. Phorbol-12-myristate-13-acetate (PMA) was used to induce the release of NET in vitro. The BHB treatment with or without PMA treatment decreased protein abundance of Cit-H3 and PAD4 (arginine deiminase 4) and increased neutrophil Elastase. Immunofluorescence and scanning electron microscope analyses revealed that BHB treatment inhibited PMA-induced NET release. The BHB treatment also decreased double strain DNA content in the supernatant, further confirming the inhibitory effect of BHB on NET release. Furthermore, BHB treatment decreased the level of intracellular Reactive Oxygen Species (ROS), phosphorylation level of p47, and protein abundance of Rac2, suggesting that BHB-induced NET inhibition may have been caused by decreased NADPH oxidase-derived ROS. The phosphorylation level of phosphoinositide 3-kinase (PI3K), an important upstream regulator of NADPH Oxidase, was attenuated by BHB treatment. To confirm the involvement of PI3K signaling pathway in BHB-induced NET inhibition, 740Y-P, a potent activator of PI3K signaling pathway, was used. Data indicated that 740Y-P relieved the inhibitory effects of BHB on ROS production and NADPH Oxidase activation. Importantly, as revealed by immunofluorescence and scanning electron microscopy analyses, 740Y-P also dampened the inhibitory effect of BHB on NET release and the protein abundance of Cit-H3 and PAD4. Overall, the present study revealed that high concentration of BHB impairs NET release through inhibiting PI3K-mediated NADPH Oxidase ROS production. These findings help partly explain the immune dysfunction in cows experiencing negative energy balance or ketosis in early lactation.

Keywords

ketosis; neutrophil extracellular trap; nicotinamide adenine dinucleotide phosphate; phosphoinositide 3-kinase; reactive oxygen species.

Figures
Products
  • Cat. No.
    Product Name
    Description
    Target
    Research Area
  • HY-P0175
    99.67%, PI3K Activator