Effects of Ibuprofen and Fusion Protein Inhibitor (FPI) on neutrophil extracellular traps (NETs) in the lung tissue of calves experimentally infected with Bovine Respiratory Syncytial Virus (BRSV)

Abstract

Abstract Neutrophil extracellular traps is a mode of neutrophil death targeting extracellular pathogens. They are composed of a complex of processed chromatin bound to granular and cytoplasmic proteins expelled from the cell onto pathogens. Although NETs play a role in trapping and killing extracellular pathogens they may be injurious and are considered to play a role in a wide array of inflammatory diseases. The exact contribution of NETs to clearance of pathogens versus inflammatory injury is poorly understood. Bovine respiratory syncytial virus (BRSV), like RSV, causes severe respiratory disease. In this study, we quantified NETs in lung tissue of calves experimentally infected with BRSV after treating with the anti-inflammatory drug Ibuprofen and/or an antiviral drug (FPI). Twenty four calves were infected with BRSV, placed into 6 treatment groups and treated with ibuprofen, FPI or both. Calves were euthanized on day 10 post infection and the lungs evaluated for pathological changes that were correlated with the IHC data. Cryosections of lung tissue were stained with primary antibodies [Rabbit anti citrinated histones(citH3), Rabbit anti neutrophil elastase (NE) and counter stained with secondary antibodies conjugated to Alexa 594 or 488. Slides were examined using an EVOS fluorescence microscope with at least 10 fields from each lung section at ×400 magnification evaluated. NETs were analyzed and quantitated using Materials Image Processing and Automated Reconstruction (MIPAR™) with the total NETs score calculated as the average of NETs in the 10 fields. Results show that a combination of Ibuprofen and FPI treatment from day 3 following experimental infection had a significant reduction in clinical score and average NETs in lung tissue.