This study investigated the pro-inflammatory and pro-oxidative effects of popular electronic cigarette aerosols (ECAs) compared with conventional cigarette smoke (CS) in the cultured human alveolar epithelial cell line (A549). Using cytotoxicity assays and four ECAs, substantial differences in biological impact were observed. CS exposure led to significant declines in cell viability and pronounced morphological changes, consistent with the presence of toxic combustion byproducts. Most ECAs caused negligible cytotoxicity except for the tobacco-flavoured variant, which demonstrated marked toxicity. DNA damage and altered cell cycle profiles were minor. Oxidative stress analysis revealed stable superoxide dismutase activity but notable glutathione depletion, especially with watermelon- and strawberry-flavoured ECAs, and unaltered mitochondrial transmembrane potential, indicating the importance of individual flavour additives in cellular antioxidant defence. Inflammatory markers, such as TNF-α, NF-κB, and IL-6, were differentially elevated across the CS and ECA groups, with IL-6 consistently increased, underscoring its role in regulating epithelial cells. Advanced double fluorescence analysis revealed increased cellular heterogeneity and inflammation, which was distinct for all ECA flavours. Overall, the findings demonstrate considerable heterogeneity in biological effects among ECA flavourings and propose a simple ECA biomonitoring model. The results emphasise the necessity for individualised toxicity assessments, especially regarding subclinical inflammation and potential long-term health outcomes.

This study investigated the proinflammatory and prooxidative effects of popular electronic cigarette (EC) aerosols compared with conventional cigarette smoke (CS) in cultured human alveolar epithelial cell line (A549). Using cytotoxicity assays and four EC extracts, substantial differences in biological impact were observed. CS exposure led to significant declines in cell viability and pronounced morphological changes, consistent with the presence of toxic combustion byproducts. Most EC extracts caused negligible cytotoxicity except for the tobacco-flavoured variant, which demonstrated marked toxicity. DNA damage and altered cell cycle profiles were minor. Oxidative stress analysis revealed stable superoxide dismutase activity but notable glutathione depletion, especially with watermelon and strawberry-flavoured ECs, indicating the importance of individual flavour additives in cellular antioxidant defence. Inflammatory markers such as TNF-α, NF-κB, and IL-6 were differentially elevated across CS and EC groups, with IL-6 consistently increased, underscoring its role in epithelial cell regulation. Advanced double fluorescence analysis highlighted increased cellular heterogeneity and inflammation, distinct for all EC flavours. Overall, the findings demonstrate considerable heterogeneity in biological effects among EC flavourings and propose a simple biomonitoring model. The results emphasise the necessity for individualised toxicity assessments, especially regarding potential long-term health outcomes.

Electronic cigarettes (ECs) are promoted as a safer alternative to traditional cigarettes, yet their impact on immune cells remains incompletely understood. This study investigates the activation of human primary adherent and non-adherent monocytes exposed ex vivo to aerosols from four flavored ECs (classic tobacco, menthol, watermelon, and strawberry) compared to cigarette smoke (CS) and nicotine alone. EC aerosols (ECEs) induced modest cytotoxicity, oxidative stress, and superoxide dismutase activity compared to CS, with high cell response heterogeneity indicating subpopulation-specific effects. Adherent monocytes showed elevated integrin expression (CD11a, CD11b), ICAM-1 (CD54), TNFα, and oxidative stress versus non-adherent cells, amplified by ECE. Dual fluorescence flow cytometry (green DCF for ROS and red for anti-TNFα Ab) revealed shifts toward pro-inflammatory/oxidative quadrants, particularly upper-right high-intensity relatively small subsets with macrophage M1-like CD68 expression in adherent cells. ECEs reduced phagocytosis in adherent monocytes, mimicking CS effects, probably driven by non-nicotine components. Strawberry flavor (ECE 4) elicited the strongest TNFα induction. These findings demonstrate EC-induced subclinical inflammation via selective monocyte activation, potentially contributing to chronic cardiopulmonary risks despite significantly lower overall toxicity than CS.