Development of an in vitro approach to point-of-contact inhalation toxicity testing of volatile compounds, using organotypic culture and air-liquid interface exposure

August 15, 2020

Artik Mistry, Larry E. Bowen, Michael W. Dzierlenga, Jessica K. Hartman, Scott D. Slattery
ScitoVation, LLC, Durham, NC 27713, United States

In this project, an incremental approach to assay development, taking small steps that built understanding of the biology and confidence in the methods were taken to attempted. Focused on general cell health assays (viability,cytotoxicity, TEER), as well as a mechanism-of-action-specific assay (GSH depletion), we found all assays to be sufficiently sensitive over the full range of possible values for both cell culture models (BEAS-2B cell and EpiAirway cultures)


In vitro chemical risk assessment using human cells is emerging as an alternative to in vivo animal testing with reduced costs, fewer animal welfare concerns, and the possibility of greater human health relevance. In vitro inhalation toxicity testing of volatile compounds poses particular challenges. Here we report our efforts to establish a testing protocol in our own lab using the EpiAirway bronchial epithelium cell culture model and the Vitrocell 12/12 system for air-liquid interface (ALI) exposures. For purposes of method development, we used methyl iodide (MeI) as a test compound. We examined viability, cytotoxicity, and epithelial integrity responses. Dose-dependent, reproducible responses were observed with all assays. EpiAirway and BEAS-2B cytotoxicity responses to acute exposure were roughly similar, but EpiAirway was more resistant than BEAS-2B by the viability measurement, suggesting a proliferative response at low MeI concentrations. If wells were sealed to prevent evaporation, in-solution MeI concentration-response could be used to predict the response to MeI vapor within 2-fold by converting from the media- to the air-concentration at equilibrium using the blood:air partition coefficient for MeI. The long-term stability of EpiAirway cultures enabled repeated exposures over a 5-d period, which produced responses at lower concentrations than did acute exposure.

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