Characterization of Bovine Neutrophil Extracellular Traps Induced by Histamine

Background: Histamine plays an central role in many allergic diseases including allergic asthma and allergic rhinitis, and is also involved in bovine laminitis through regulating immune responses. Neutrophil extracellular traps (NETs) formation is a novel effector mechanism of neutrophils to defend against various stimuli. In the present study, we aimed to investigate the role of histamine on bovine NET formation and examined its fundamental molecular mechanisms. ResultsFirstly, the effects of histamine on neutrophil viability was measured by Cell Counting Kit-8 (CCK-8) and Lactate dehydrogenase (LDH) assays. The results showed that histamine had no significant influence on neutrophil viability. Then we characterized histamine-triggered NET formation by confocal microscopy and PicoGreen-derived NETs quantification. Confocal microscopy analyses illustrated NET structures by co-localizing the main components of NETs, and NET quantification revealed that histamine-triggered NETs were released in a dose-dependent manner. In addition, we found reactive oxygen species (ROS) production, phosphorylated extracellular signal-regulated kinase (ERK) and p38 proteins were significantly elevated in histamine-challenged neutrophils. By applying functional inhibitors of nicotinamide adenine dinucleotide phosphate-oxidase (NADPH oxidase), ERK and p38, histamine-triggered NETs were markedly reduced, indicating their importance in histamine-triggered NET formation. ConclusionsOur findings described histamine-triggered NET formation, and revealed its potential molecular mechanisms via NADPH oxidase, ERK and p38 pathways. This is the first study to depict NET formation induced by histamine, which could provide a new insight into histamine-related diseases.