Ozone induced acute and chronic alterations in the lung in mice: a combined digital imaging and functional analysis.

Ozone is a major environmental pollutant inducing severe pulmonary damages. We have compared the effect of ozone on mice subjected to acute (1 ppm, 1 h) and chronic (1.5 ppm, 1 h, twice weekly for 6 weeks) ozone exposure. Our investigations focused on lesions of the lung tissue, inflammation and hyperreactivity using both conventional methods, and a novel digital imaging histological approach based on machine learning programs. Acute exposure caused disruption in the respiratory epithelium with leakage of protein and recruitment of neutrophils in the bronchoalveolar space, leading to lung inflammation and airway hyperresponsiveness to methacholine. All these parameters were also increased upon chronic exposure. Digital imaging analysis allowed to quantify accurately structural changes occurring in the bronchiolar epithelium. Acute exposure mainly induced a significant decrease of epithelium area and thickness in agreement with its desquamation. Chronic exposure caused significant increase of epithelial, bronchial and lumen areas, unlike acute exposure. Morphological changes of epithelial bronchi were accompanied by a significant increase of peribronchial collagen content under both acute and chronic exposure. However, unlike acute exposure, chronic exposure resulted in a significant increase of the area and diameter of airspaces, but a decrease in their number, all of which reflecting the presence of emphysema. In conclusion, the digital imaging approach provides reliable quantitative analysis of pulmonary structural changes which, in combination with functional approaches, allows a comprehensive analysis of the effects of ozone on lung tissue.