Abstract
OBJECTIVE: It was found that the levels of antineutrophil cytoplasmic antibodies (ANCA) are elevated and linked to disease severity of bronchiolitis obliterans (BO) in children. This study aims to explore the mechanism of ANCA in the process of BO. METHODS: Plasma from BO patients (n = 40) and healthy controls (n = 11) was analyzed for ANCA and neutrophil extracellular traps (NETs) components. Plasma IgG from ANCA-positive BO children and normal controls were used to stimulate neutrophils, measuring reactive oxygen species (ROS) and NETs production. Small airway epithelial cells (SAECs) were exposed to NETs, assessing viability by CCK8 and cytokine release by ELISA. The IgG treated neutrophils were co-cultured with SAECs, and cytokines were measured by ELISA. RESULTS: The levels of ANCA and NETs components including dsDNA, neutrophil elastase (NE) and myeloperoxidase (MPO) in the plasma of BO children were significantly higher than those of healthy controls. ANCA-positive IgG induced neutrophils produce ROS and NETs. The cell viability of SAECs was significantly reduced upon treatment with NETs in a concentration-dependent manner. The levels of IL-8, IL-17, TNF-α, and TGF-β secreted by SAECs treated with NETs were increased significantly, and the degree of increase was positively correlated with the concentration of NETs. The co-culture of neutrophils stimulated by ANCA IgG with SAECs significantly increased the expression of cytokines including IL-8, IL-17, TNF-α, and TGF-β. CONCLUSIONS: NETs induced by ANCA may exacerbate airway inflammation in children with BO.