Effects of Emissions from Heated Tobacco Products and Reference Cigarettes on Gene Expression and Mitochondrial Function in Human Lung Epithelial BEAS-2B Cells †

Heated tobacco products (HTPs) are marketed as lower-risk alternatives to conventional cigarettes; however, their toxicological impacts remain insufficiently characterized. This study evaluated the effects of HTP emissions on gene expression and mitochondrial function in comparison with conventional cigarettes. Whole cigarette smoke condensates (WCSCs), comprising both gas and particulate phases, were prepared from three commercially available HTPs and from 3R4F reference cigarettes. Human lung epithelial BEAS-2B cells were exposed to WCSCs at 3 μg nicotine/mL for 24 h, followed by transcriptome profiling using RNA sequencing. Principal component analysis demonstrated that HTP-WCSCs induced weaker gene expression changes than 3R4F-WCSC, with only modest variation among HTPs. Gene set enrichment analysis revealed that both HTP- and 3R4F-WCSCs significantly downregulated oxidative phosphorylation (OXPHOS)-related pathways, indicating potential mitochondrial impairment. Functional assays confirmed that both exposures elevated mitochondrial reactive oxygen species (ROS), while mitochondrial morphology, ATP production, membrane potential, and cytosolic ROS were largely unaffected. Collectively, these results show that although HTP emissions elicit weaker transcriptomic perturbations than conventional cigarette emissions, both converge on mitochondrial targets by suppressing OXPHOS gene expression and increasing mitochondrial ROS. Mitochondrial dysfunction may therefore represent a common mechanism underlying tobacco product toxicity.