# Increased neutrophil extracellular traps formation in the bronchoalveolar lavage fluid of dogs with bronchiectasis

## TL;DR

This study found increased neutrophil extracellular traps in dogs with bronchiectasis, suggesting they may play a role in the disease's progression.

## Contribution

The study is the first to validate and quantify neutrophil extracellular traps in bronchoalveolar lavage fluid of dogs with bronchiectasis and related conditions.

## Key findings

- Neutrophil extracellular traps (NETs) were detectable in bronchoalveolar lavage fluid of dogs with bronchiectasis and bronchial disease.
- MPO–DNA complex levels were significantly higher in dogs with bronchiectasis compared to healthy controls.
- NETs correlated with neutrophil counts and disease duration, especially in bronchiectasis cases.

## Abstract

Chronic bronchial diseases (CBD) associated with bronchiectasis (BE) and/or bronchomalacia (BM) are commonly diagnosed in dogs. Their pathogenesis remains incompletely understood, and recent studies have questioned the clinical relevance of distinguishing BE and/or BM, suggesting that their pathogenic mechanisms may overlap and that differentiating between them may have limited clinical significance. To date, the role of neutrophil extracellular traps (NETs) in BE and/or BM remains undetermined. This study aimed to validate NETs detection techniques in banked canine bronchoalveolar lavage fluid (BALF) using healthy (H) dogs, as negative controls, dogs with bacterial bronchopneumonia (BBP), as positive controls, and to compare NET detectability in dogs with CBD associated with BE and/or BM. A retrospective observational study was conducted using client-owned dogs presented with CBD and BE, BM, or BEBM based on endoscopic findings, or BBP. NETs were measured in BALF by quantification of cell-free DNA (cfDNA), detection of myeloperoxidase (MPO)-DNA complexes via enzyme-linked immunosorbent assay (ELISA) and by confocal laser scanning microscopy of immunofluorescent labeled samples. cfDNA concentrations were significantly higher in BBP dogs than in CBD or H-dogs, with no significant differences among the CBD groups. MPO–DNA complex levels were significantly higher in dogs with CBD and BBP than in H-dogs, with BE dogs showing the greatest increase compared with H and BM dogs, and BEBM dogs also exceeding H-dogs. Across all dogs, MPO–DNA complexes correlated positively with total and absolute neutrophil counts, neutrophil percentage, cfDNA concentrations, and duration of clinical signs. These associations varied between groups, with the strongest and most consistent correlations observed in BE-dogs. This study indicates that NETs are detected and quantifiable in the BALF of dogs with CBD and BE and/or BM and dogs with BBP. Their presence suggests that NET formation may play a role in the pathogenesis of CBD, especially in cases involving BE. Further research is needed to explore the role of NETs as well as the therapeutic potential of NET modulation in respiratory diseases.