Development of bioassays for gaseous chemicals using air-liquid interface culture of human lung epithelial cells

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To establish a series of toxicitiy evaluation systems for chemicals or environmental samples in gas phase, we employed air-liquid interface culture (ALIC) of human bronchial (air way) and alveolus epithelial cells as a basic culture method, and investigated their applications to volatile organic compounds and suspended paniculate matter (SPM).First, using a human bronchial epithelial cell line, Calu-3, we developed a simple closed system for direct gas exposure, and tested the toxicity expression of Calu-3 to several organic compounds during 48 hours of loading. The toxicity in ALIC exposure was higher than that in conventional exposure in the liquid phase. The reason was largely explained by numerical estimation that chemical concentration on the cell surface in the liquid culture is lower than that in ALIC culture, in the cause of the diffusion process of molecules in the surface liquid layer. These results indicate that basic concept of the combination of ALIC of lung cells and a si … More mple closed system is promising as a testing cytotoxicrly to gaseous compounds.Second, in vivo mimicking exposure of actual SPM samples to human lung alveolar epithelia was achieved using an air-liquid interface culture of human lung alveolar A549 cells, resulting in successful observation of the toxicity in terms of the cell survivability after 48 hours exposure. In addition, an integrated use of the ALIC-based A549 cell assay for general toxicity and the EROD-based Hep G2 cell assay for PAHs was recommended to evaluate the primary toxicity of SPM. We thereby understood the permeation kinetics of SPM toxicity across the alveolar epithelia in terms of the EROD capacity of Hep G2 cells, and were able to obtain basic information on SPM toxicity in humans.Third, we developed a numerical simulation model that describes elution of chemicals initially adsorbed onto SPM samples, intake and accumulation of these chemicals into alveolus cell layer, and their final permeation to the systemic blood circulation. By focusing on the EROD activity of SPM samples, we quantified the comprehensive toxicity of the relevant samples and used these toxicitis in determining parameters involved in these numerical simulations. In addition, we extended the numerical models in actual situations that reflect low-dose and long-term exposure of SPM to humans across the very thin layer of the alveolar tissue. Less