Modifications in human oral fibroblast ultrastructure, collagen production and lysosomal compartment in response to E-cigarette fluids

Background: Many smokers have recently turned to electronic cigarettes (e-cigarettes) because they have been marketed as a cheaper, safer smokeless alternative to traditional cigarettes and a possible smoking cessation tool. Although the safety of these electronic devices is still not fully known, there is evidence of their cytotoxicity on cells belonging to the oral cavity. In a previous study by the authors, the increase of reactive oxygen production and Bax expression, followed by the occurrence of apoptosis, was demonstrated in human gingival fibroblasts (HGFs). The aim of this paper is to further investigate the effects of the e-cigarette liquids (with and without nicotine) on the same experimental model. Methods: HGFs were treated with e-cigarette fluids containing nicotine (final concentration 1 mg/mL) and the equivalent volume of a fluid without nicotine, for periods ≤48 hours. Lactate dehydrogenase assay (LDH), electronic microscopy analysis, collagen I production, flow cytometry lysosome compartment evaluation, and western blotting light chain 3 (microtubule-associated protein 1A/1B-LC3) expression were performed. Results: Fluids containing nicotine exerted cytotoxicity as demonstrated by increased levels of LDH, in parallel to the presence of numerous vacuoles in the cytoplasm, a decrease in collagen I production, and augmented LC3 II expression. Autophagic vesicles and more procollagen I molecules were present in the cytoplasm of fibroblasts exposed to nicotine-free fluids. In the same samples, time-dependent activation of the lysosomal compartment with no changes in LC3 expression was detected. Conclusion: E-cigarette fluids (with and without nicotine) trigger molecular and morphologic responses in oral fibroblasts, raising concerns about their role in the pathogenesis of oral diseases.