3-D Nasal Cultures: Systems Toxicological Assessment of a Candidate Modified-Risk Tobacco Product

July 7, 2016

doi: 10.14573/altex.1605041

Anita R. Iskandar, Carole Mathis, Florian Martin, Patrice Leroy, Alain Sewer, Shoaib Majeed, Diana Kuehn, Keyur Trivedi, Davide Grandolfo, Maciej Cabanski, Emmanuel Guedj, Celine Merg, Stefan Frentzel, Nikolai V. Ivanov, Manuel C. Peitsch and Julia Hoeng

Philip Morris International R&D, Philip Morris Products S.A., Neuchâtel, Switzerland

Human nasal epithelium is the first barrier against inhaled toxicants. The air-liquid interface human nasal culture MucilAir™ offers a more physiologically relevant and robust systems for studying the effects of exposure. This cells were exposed to 3R4F cigarettes and THS2.2 tobacco sticks
in the the Vitrocell® 24/48 exposure system. Results have been evaluated by measuring the cytotoxicity, histology processing, immunostaining, cilia beating frequency (CBF), cytochrome P450 (CYP) activity, luminex-based measurement of secreted analytes, RNA/miRNA purification and mRNA microarray.

In vitro toxicology approaches have evolved, from a focus on molecular changes within a cell to understanding of toxicity-related mechanisms in systems that can mimic the in vivo environment. The recent development of three dimensional (3-D) organotypic nasal epithelial culture models offers a physiologically robust system for studying the effects of exposure through inhalation. Exposure to cigarette smoke (CS) is associated with nasal inflammation; thus the nasal epithelium is relevant for evaluating the pathophysiological impact of CS exposure. The present study investigated further the application of in vitro human 3-D nasal epithelial culture models for toxicological assessment of inhalation exposure. Aligned with 3Rs strategy, this study aimed to explore the relevance of a human 3-D nasal culture model to assess the toxicological impact of aerosols generated from a candidate modified risk tobacco product (cMRTP), the Tobacco Heating System (THS)2.2, as compared with smoke generated from reference cigarette 3R4F. A series of experimental repetitions where multiple concentrations of THS2.2 aerosol and 3R4F smoke were applied, were conducted to obtain reproducible measurements to understand the cellular/molecular changes that occur following exposure. In agreement with the Vision and Strategy of the Toxicity Testing in the 21st Century, this study implemented a systems toxicology approach and found that for all tested concentrations, the impact of 3R4F smoke was substantially greater than that of THS2.2 aerosol in terms of cytotoxicity levels, alterations in the tissue morphology, secretion of pro-inflammatory mediators, impaired ciliary function, and increased perturbed transcriptomes and miRNA expression profiles.

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