Rob Bedford¹, Emma Rothwell¹, Sophie Martin¹, Cian O’Hanlon¹, Andrew McCune² and Michael Hollings^1
1Genetic and Molecular Toxicology and ²Immunology and Immunotoxicology, Covance Laboratories Ltd., Harrogate, UK

- Exposure to 3R4F resulted in increased levels of IL-4, MDC, GM-CSF, IL-12/IL23p40, IL-10 and IFN-γ in the recovery media. Approximately three-fold increases in MDC, GM-CSF, IL-12/IL23p40 and IFNγ were observed whilst two-fold increases were observed for IL-4 and IL-10. In comparison, no marked effect was observed in the module media.

- In contrast to the response observed from 3R4F exposure, fewer changes in cytokine production were observed following THP and E-cigarette exposure. IFNγ demonstrated a two-fold increase in levels measured in the recovery media at doses 1 and 2 for THP. IL-12/IL23p40 also demonstrated a 1.5-fold increase in recovery media following exposure to THP. In addition, IFN-γ and IL-8 were increased following exposure to E-cigarette at dose 2. IL-1β also demonstrated a 1.5-fold increase in the recovery and module media following exposure to E-cigarette.

- A number of cytokines were reduced following exposure to THP and E-cigarette. For example, GM-CSF, MIP1α, VEGF and MCP-1.

- These results demonstrate the difference in cytokine profiles of MucilAir tissues following exposure to different nicotine-containing products.

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DOI: https://doi.org/10.1016/j.mrgentox.2020.503230 

Authors
Thorne D¹, Hollings M², Kilford J², Clements J², Payne R², Ballantyne M², Dalrymple, A¹, Dillon, D¹, Meredith, C¹, Gaҫa, M^{1  
 
1} British American Tobacco, R&D, Southampton, Hampshire SO15 8TL, UK. 
² Covance Laboratories Ltd, Otely Road, Harrogate, HG1 PY, UK

Highlights

• An experimental concept of exposing L5178Y cells at the air-agar-interface
• Cells were lightly seeded on an agar surface to achieve exposure
• Flowing air, plating and recovery efficiencies from the agar-surface were investigated
• Positive controls were responsive, suggesting a clear cellular interaction with the agar-matrix 
• Cigarette smoke aerosol showed a dose-response relationship with increasing mutant frequencies

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Robert Leverette¹, Brian Keyser¹, Michael Hollings² and Adam Seymour^2
1RAI Services Company, Winston−Salem, NC 27101 USA
²Covance Laboratories Ltd., Harrogate, North Yorkshire HG3 1PY, UK

• Freshly generated whole smoke / aerosol from three different tobacco product types (3R4F, THP and ENDS) was consistently delivered within the AMES 48 and 6/48 exposure modules.
• Biological endpoints: 3R4F whole smoke induced increased revertant counts (Ames) and cytotoxicity (NRU) in the AMES 48 and 6/48 modules, respectively.
• Revertant counts (Ames) and cytotoxicity (IC50) values were comparable to those from the standard exposure modules when run under similar exposure conditions.
• Overall, the AMES 48 and 6/48 modules are deemed “Fit for Use" for the in vitrO evaluation of different tobacco product types (combustible, THP, ENDS).
 

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Adam Seymour, Michael Hollings, Joanne Larner and Julie Clements;

Covance Laboratories Ltd, Harrogate, UK

QCMs were used to assess the deposition of cigarette aerosol from a Vitrocell® VC10® in Vitrocell® Ames, 6/4, 12/4 and 24/4 exposure modules. The power to resolve difference between 8 and 1 SLPM is statistically significant in all module types assessed. Uniformity within each module was confirmed (P >0.98) in all module types, at all airflows. Repeatability was assessed and found to be consistent (no significant difference) in all module types at airflows of 10, 8 and 4 SLPM, however, at 1 SLPM a significant difference was noted in 9 out of 12 experiments – likely an artifact of high deposition.
These results show that exposures utilizing the Vitrocell® VC10® and associated modules are robust; but indicate the necessity of dosimetry measures to aid in the quantification of delivered dose.
 

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¹Leverette R, ¹Keyser B, ²Seymour A, ²Hollings M and ¹Bombick B
¹Scientific & Regulatory Affairs, RAI Services Company Winston-Salem, NC 27102
²Covance Laboratories Ltd Otley Road, Harrogate, North Yorkshire HG3 1PY, UK

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