Mucus obstruction is a key feature of many inflammatory airway diseases. Neutrophil extracellular traps (NETs) are released upon neutrophil stimulation and consist of extracellular chromatin networks studded with cytotoxic proteins. When released in the airways, these NETs can become part of the airway mucus. We hypothesized that the extracellular DNA and/or oxidative stress (e.g., by the release of reactive oxygen species and myeloperoxidase during NETs formation in the airways) would increase mucus viscoelasticity. We collected human airway mucus from endotracheal tubes of healthy patients admitted for elective surgery and coincubated these samples with NETs from phorbol 12-myristate 13-acetate-stimulated neutrophils. Unstimulated neutrophils served as controls, and blocking experiments were performed with dornase alfa for extracellular DNA and the free radical scavenger dimethylthiourea for oxidation. Compared with controls, the coincubation of mucus with NETs resulted in 1) significantly increased mucus viscoelasticity (macrorheology) and 2) significantly decreased mesh pore size of the mucus and decreased movement of muco-inert nanoparticles through the mucus (microrheology), but 3) NETs did not cause visible changes in the microstructure of the mucus by scanning EM. Incubation with either dornase alfa or dimethylthiourea attenuated the observed changes in macrorheology and microrheology. This suggests that the release of NETs may contribute to airway mucus obstruction by increasing mucus viscoelasticity and that this effect is not solely due to the release of DNA but may in part be due to oxidative stress.

粘液阻塞是许多炎性气道疾病的关键特征。中性粒细胞胞外陷阱 (NETs) 在中性粒细胞刺激后释放，由散布有细胞毒性蛋白的胞外染色质网络组成。当在气道中释放时，这些 NET 可以成为气道粘液的一部分。我们假设细胞外 DNA 和/或氧化应激（例如，气道中 NETs 形成过程中活性氧和髓过氧化物酶的释放）会增加粘液的粘弹性。我们从接受择期手术的健康患者的气管插管中收集人气道粘液，并将这些样本与来自佛波醇 12-肉豆蔻酸酯 13-乙酸酯刺激的中性粒细胞的 NET 共培养。未受刺激的中性粒细胞作为对照，用dornase alfa 进行细胞外DNA 和自由基清除剂二甲基硫脲氧化进行封闭实验。与对照组相比，黏液与 NETs 的共培养导致1 ) 显着增加粘液粘弹性 (宏观流变学) 和2 ) 显着减小粘液的网孔尺寸和减少粘液惰性纳米粒子通过粘液的运动 (微流变学)，但3 ) NETs 不会引起粘液微观结构的可见变化通过扫描 EM。与 dornase alfa 或二甲基硫脲一起孵育减弱了观察到的宏观流变学和微观流变学的变化。这表明 NETs 的释放可能通过增加粘液粘弹性导致气道粘液阻塞，这种影响不仅是由于 DNA 的释放，而且可能部分是由于氧化应激。