Development of a repeated exposure protocol of human bronchial epithelium in vitro to study the long-term effects of atmospheric particles.

Chronic exposure to atmospheric particles is suspected of exacerbating chronic inflammatory respiratory diseases but the underlying mechanisms remain poorly understood. An experimental strategy using human bronchial epithelial cells (NHBE) known to be one of the main target cells of particles in the lung was developed to investigate the long term effects of repeated exposure to particles. Primary cultures of NHBE cells were grown at an air-liquid interface and subjected to repeated treatments to particles. Fate of particles, pro inflammatory response and epithelial differentiation were studied during the 5 weeks following the final treatment. Ultrastructural observations revealed the biopersistence of particles in the bronchial epithelium. The expression of cytochrome P450 1A1, was transiently induced, suggesting that organic compounds could have been metabolized. The release of GM-CSF and IL-6 (biomarkers of pro-inflammatory response), was induced by particle treatments and was maintained up to 5weeks after treatments. The release of amphiregulin and TGFα (Growth Factor) was induced after each treatment. The number of cells expressing the mucin MUC5AC, a differentiation marker, was increased in particle-exposed epithelium. The experimental strategy we developed is suitable for investigating in greater depth the long term effects of particles on bronchial epithelial cells repeatedly exposed to atmospheric particles in vitro.