Fig. 1: Evolving Landscape of Preclinical Tools Used for Toxicological Assessment of Electronic Nicotine Delivery Systems.

a Electronic nicotine delivery systems (ENDS) are proliferating swiftly, with a wide array of e-liquid ingredients and device functionalities, presenting considerable challenges in assessing their potential pulmonary toxicity. Traditional methodologies predominantly include two-dimensional (2D) cell culture models that employ either cell lines or undifferentiated primary human lung cells (b), as well as static three-dimensional (3D) in vitro models such as air-liquid interface (ALI) cultures using primary human airway epithelial cells in transwell inserts (TWIs) (c), microtome-assisted precision cut lung slices (PCLS) (d), laboratory animals, notably rodents (e), and, rarely, lung spheroids and organoids (f). Complementary techniques involve in silico modeling (g) and physicochemical analytical methods such as quartz crystal microbalance (QCM), gas chromatography-mass spectrometry (GC-MS), and nuclear magnetic resonance (NMR) spectroscopy (h, i). Emerging bioengineered technologies, such as Lung-on-a-Chip living devices (j) and the bioinspired non-living robotic platform Human Vaping Mimetic Real-Time Particle Analyzer (HUMITIPAA) (k), offer new opportunities to overcome certain limitations of existing tools, enhance human and clinical relevance, and expedite toxicity evaluation. Images in (j, k) were reproduced from ref. ⁸⁷. and ref. ¹³⁸, respectively. a–g, i Were fully or in part created using BioRender.com, and (j, k) were reproduced with permission from ref. ⁸⁷ and ref. ¹³⁸.