Development of a standardized in vitro approach to evaluate microphysical,chemical,and toxicological properties of combustion-derived fine and ultrafine particles

Ultrafine particles represent a growing concern in the public health community but their precise role in many illnesses is still unknown. This lack of knowledge is related to the experimental difficulty in linking their biological effects to their multiple properties, which are important determinants of toxicity. Our aim is to propose an interdisciplinary approach to study fine(FP) and ultrafine(UFP) particles, generated in a controlled manner using a mini CAST(Combustion Aerosol Standard) soot generator used with two different operating conditions(CAST1 and CAST3). The chemical characterization was performed by an untargeted analysis using ultra-high resolution mass spectrometry. In conjunction with this approach, subsequent analysis by gas chromatography–mass spectrometry(GC–MS) was performed to identify polycyclic aromatic hydrocarbons(PAH). CAST1 enabled the generation of FP with a predominance of small PAH molecules, and CAST3 enabled the generation of UFP, which presented higher numbers of carbon atoms corresponding to larger PAH molecules. Healthy normal human bronchial epithelial(NHBE) cells differentiated at the air-liquid interface(ALI) were directly exposed to these freshly emitted FP and UFP. Expression of MUC5AC, FOXJ1, OCLN and ZOI as well as microscopic observation confirmed the ciliated pseudostratified epithelial phenotype. Study of the mass deposition efficiency revealed a difference between the two operating conditions, probably due to the morphological differences between the two categories of particles. We demonstrated that only NHBE cells exposed to CAST3 particles induced upregulation in the gene expression of IL-8 and NQO1. This approach offers new perspectives to study FP and UFP with stable and controlled properties.