Advanced in vitro exposure systems.

In Vitro Cytotoxicity Assessment of 3D Human Airway Tissue Following Exposure to Whole Aerosol/Smoke Generated from Electronic and Combustible Cigarettes

27. Mar. 2022

Poster SOT 2022, 61st Annual Meeting and ToxExpo, March 27-31, San Diego, USA

Thomas Shutsky1, Brian M. Keyser1, Kristen Jordan1, Michael Hollings2, and Emma Rothwell2
1Scientific & Regulatory Affairs, RAI Services Company, Winston-Salem, NC; 
2Labcorp Early Development Laboratories Ltd., Harrogate, UK

In this study, EpiAirway™ tissues were exposed to whole aerosol/smoke generated from six different Vuse Alto electronic nicotine delivery system test articles with various nicotine concentrations and a combustible cigarette. A Vitrocell® VC10® robot was used to generate whole aerosol/smoke with either a modified ISO or Health Canada Intense smoking regimen. Whole smoke/aerosol was puffed with clean air with a series of different air flows to achieve the delivered dose range. 

 

Abstract

The MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay measures the metabolic activity of the cell and is therefore an indicator of cell survival/viability. Characterization of the capacity of the 3D culture systems comprised of human airway tissue (e.g., EpiAirway™) to evaluate the cytotoxicological exposures to the respiratory tract provides a useful tool to distinguish the impact of known and potential respiratory irritants.
In this study, EpiAirway™ tissues were exposed to whole aerosol/smoke generated from six different Vuse Alto electronic nicotine delivery system (ENDS) test articles with various nicotine concentrations and a combustible cigarette (CC). A Vitrocell® VC10® robot was used to generate whole aerosol/smoke with either a modified ISO 20768:2018 (ENDS) or Health Canada Intense (HCI) smoking regimen. Whole smoke/aerosol was puffed with clean air with a series of different air flows (L/minute) to achieve the delivered dose range. The exposure conditions were 4.0 - 0 L/min (undiluted) for the ENDS and 10 - 1 L/min for the combustible cigarette. Liquid traps containing PBS within the exposure module allowed aerosol/smoke dosimetry via nicotine quantification.
The whole smoke generated from the CC was found to be cytotoxic in the EpiAirway™ tissue model, as measured by the MTT assay with a mean calculated IC50 value of 6.5 µg nicotine /mL. Whole aerosol from the ENDS products did not induce any observed cytotoxicity, with tissue viability not falling below 83% at the highest delivered dose (undiluted aerosol). In the absence of ENDS aerosol cytotoxicity, the highest doses delivered based on nicotine concentration (356.75 – 982 µg nicotine/mL depending on ENDS nicotine levels), were orders of magnitude greater than the mean calculated CC IC 50 value (6.5 µg nicotine /mL). These data demonstrate that the MTT assay using the EpiAirway™ model can be used to differentiate cytotoxic responses to whole aerosol/smoke exposure from different tobacco product categories in a relevant human in vitro respiratory test system.

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