Lung injury and oxidative stress induced by inhaled chlorine in mice is associated with proinflammatory activation of macrophages and altered bioenergetics.

Chlorine (Cl) gas is a highly toxic and oxidizing irritant that causes life-threatening lung injuries. Herein, we investigated the impact of Cl-induced injury and oxidative stress on lung macrophage phenotype and function. Spontaneously breathing male C57BL/6J mice were exposed to air or Cl (300 ppm, 25 min) in a whole-body exposure chamber. Bronchoalveolar lavage (BAL) fluid and cells, and lung tissue were collected 24 h later and analyzed for markers of injury, oxidative stress and macrophage activation. Exposure of mice to Cl resulted in increases in numbers of BAL cells and levels of IgM, total protein, and fibrinogen, indicating alveolar epithelial barrier dysfunction and inflammation. BAL levels of inflammatory proteins including surfactant protein (SP)-D, soluble receptor for glycation end product (sRAGE) and matrix metalloproteinase (MMP)-9 were also increased. Cl inhalation resulted in upregulation of phospho-histone H2A.X, a marker of double-strand DNA breaks in the bronchiolar epithelium and alveolar cells; oxidative stress proteins, heme oxygenase (HO)-1 and catalase were also upregulated. Flow cytometric analysis of BAL cells revealed increases in proinflammatory macrophages following Cl exposure, whereas numbers of resident and antiinflammatory macrophages were not altered. This was associated with increases in numbers of macrophages expressing cyclooxygenase (COX)-2 and inducible nitric oxide synthase (iNOS), markers of proinflammatory activation, with no effect on mannose receptor (MR) or Ym-1 expression, markers of antiinflammatory activation. Metabolic analysis of lung cells showed increases in glycolytic activity following Cl exposure in line with proinflammatory macrophage activation. Mechanistic understanding of Cl-induced injury will be useful in the identification of efficacious countermeasures for mitigating morbidity and mortality of this highly toxic gas.