A Comparison of Cell Culture Models and Exposure Routes for In Vitro Inhalation Toxicity Testing

Download Poster PDF: Inhalation Toxicity Testing SOT 2020 Poster PDF

Authors: Scott Slattery, Artik Mistry, Larry Bowen, Michael Dzierlenga, and Jessica Hartman

Development of in vitro assays for point-of-contact inhalation toxicity should consider two factors: 1) the physiological relevance of the cell culture model, and 2) the route of exposure. Organotypic culture models may better replicate in vivo adverse outcomes and allow for repeat exposures, whereas simpler cell-line-based models may be more applicable to high-throughput acute testing. Exposure of submerged cultures to compounds in solution is a commonplace approach, compatible with many in vitro assays and amenable to scaling for high-throughput testing.  However, exposure of cultures at the air-liquid interface may better simulate in vivo inhalation exposure to vapors.  

To better understand how these factors affect testing results, we have investigated a four-quadrant matrix of testing approaches that combines simple (BEAS-2B) or organotypic (EpiAirway, MatTek) cell culture models with in-solution or vapor-phase exposures. As an initial test compound, we have chosen methyl iodide for its well-understood and direct mechanism of action in airway epithelia (depletion of glutathione and subsequent cytotoxicity). Sensitivity to methyl iodide-induced cytotoxicity was similar for the two cell culture models, whether exposure was in solution (IC50s: 2.6 mM for BEAS-2B and 3.1 mM for EpiAirway) or at ALI (IC50s: 972 ppm for BEAS-2B and 1191 ppm for EpiAirway). However, we found that the in-solution sensitivities could not predict the vapor-phase sensitivities when converted to equilibrium vapor concentrations using an air-water partition coefficient. The vapor IC50s calculated from solution IC50s were approximately ten times higher than the measured vapor IC50s. This may be explained by the fast evaporation of methyl iodide from exposure medium, with a half-time of ten minutes. Therefore, in-solution exposure was of limited duration, whereas vapor-phase exposure could be maintained for longer durations. The EpiAirway model maintains homeostasis for several weeks, allowing repeated exposures. We observed cumulative effects with repeated exposures to low concentrations of methyl iodide that were not seen after acute exposure.

We conclude that vapor-phase exposure better mimic real-life exposure scenarios for volatile compounds, and that long-lived organotypic cultures allow for detection of effects after repeated exposures.