How to use an in vitro approach to characterize the toxicity of airborne compounds

As part of the development of new approach methodologies (NAMs), numerous in vitro methods are being developed to characterize the potential toxicity of inhalable xenobiotics (gases, volatile organic compounds, polycyclic aromatic hydrocarbons, particulate matter, nanoparticles). However, the materials and methods employed are extremely diverse, and no single method is currently in use. Method standardization and validation would raise trust in the results and enable them to be compared. This four-part review lists and compares biological models and exposure methodologies before describing measurable biomarkers of exposure or effect. The first section emphasizes the importance of developing alternative methods to reduce, if not replace, animal testing (3R principle). The biological models presented are mostly to cultures of epithelial cells from the respiratory system, as the lungs are the first organ to come into contact with air pollutants. Monocultures or cocultures of primary cells or cell lines, as well as 3D organotypic cultures such as organoids, spheroids and reconstituted tissues, but also the organ(s) model on a chip are examples. The exposure methods for these biological models applicable to airborne compounds are submerged, intermittent, continuous either static or dynamic. Finally, within the restrictions of these models (i.e. relative tiny quantities, adhering cells), the mechanisms of toxicity and the phenotypic markers most commonly examined in models exposed at the air-liquid interface (ALI) are outlined.