Neutrophil extracellular traps induce airway epithelial-mesenchymal transition in a neutrophil-dominant asthmatic murine model through Hedgehog/Gli1 pathway.

Abstract

BACKGROUND: Neutrophilic asthma is one of the main types of severe asthma. Our previous studies demonstrated that neutrophil extracellular traps (NETs) contribute to its pathological process. However, the underlying mechanisms remains unclear. This study aimed to investigate the potential mechanisms of NETs in neutrophilic asthma. METHODS: Clinical samples were collected from patients with neutrophilic asthma and healthy controls. A neutrophil-dominant asthmatic murine model was established using ovalbumin (OVA), Freund's complete adjuvant (CFA) and lipopolysaccharide (LPS). Airway inflammation and remodeling were assessed by pathological staining. The expression of EMT markers and Hedgehog (Hh)/Gli1 pathway markers were measured by Western blot, qPCR, and immunofluorescence. RESULTS: We found that the expression of dsDNA, one of the skeleton components of NETs, was significantly higher in the peripheral plasma of patients with neutrophilic asthma than that of healthy controls, and neutrophils in neutrophilic asthma patients were more likely to induce the production of NETs. We further demonstrated that NETs induced EMT in airway epithelial cells. Both in vivo and in vitro, we confirmed that reducing NETs formation or enhancing NETs degradation reversed EMT process, attenuated airway hyperresponsiveness (AHR) and alleviated airway inflammation in neutrophil-dominant asthmatic mouse model. We also found that the Hh/Gli1 pathway was activated during this process, and inhibition of the Hh/Gli1 pathway also reversed the EMT process of airway epithelium. Similarly, AHR and airway inflammation in neutrophil-dominant asthmatic mice were reduced. CONCLUSIONS: We confirmed that NETs promote EMT in airway epithelium via activation the Hh/Gli1 signaling pathway, thus playing an important role in the pathogenesis of neutrophilic asthma. Targeting NETs or the Hh/Gli1 pathway may provide a promising therapeutic strategy for the treatment of severe neutrophilic asthma.