Effects of prolonged stimulation with heated tobacco products (Ploom TECH⁺) on gingival epithelial cells

Abstract

Objective and Background: Heated tobacco products have recently become commercially available. These products, as well as combustible cigarettes, produce aerosols; the risk of various diseases associated with heated tobacco products may be the same or higher than that with combustible cigarettes. In this study, we examined the effect of Ploom TECH + extract on gingival epithelial cells.Methods: Tobacco leaves from Ploom TECH + tobacco capsules and water were mixed and heated; the supernatant subsequently collected was the heated tobacco product (HTP; control: HTP not added). Normal human gingival epithelial progenitors were cultured alternately with or without HTP for a total of 1 month. Subsequently, RNA, DNA, and proteins were isolated from these samples and comprehensively analyzed using RNA sequencing (RNA-seq), reduced representation bisulfite sequencing (RRBS), and western blotting, respectively.Results: RNA-seq revealed that 284 genes showed a twofold increase and 145 genes showed a twofold decrease in gene expression. A heat map showed genetic differences between the control and HTP groups. A principal component analysis plot showed a clear genetic distribution between the control and HTP. Gene Ontology (GO) analysis showed that genes related to seven GO terms, including cornification and keratinization, were induced by long-term HTP stimulation. By contrast, GO pathways with a significant decrease in component expression were not detected. RRBS revealed that CpG island methylation increased more than twofold in 158 genes and decreased to less than twofold in 171 genes. Methylation of these CpG islands was not correlated with changes in gene expression levels. HTP treatment increased S100A7 expression. Conclusion:Long-term HTP stimulation affected epithelial differentiation and keratinization of gingival epithelial cells. Thus, habitual use of Ploom TECH + may be a risk factor for tobacco-related oral mucosal diseases.

Electronic Cigarettes, Heated Tobacco Products, and Oral Health: A Systematic Review and Meta-Analysis

“…Similarly, two studies on HTP confirmed gene modifications when oral cells were exposed to HTP [108,109]. When compared to ECS and CS, HTP seems to modify oral cell function [111].…”

“…Regarding studies on oral cancer cells [108], conclusive hypotheses could not be drawn, since some papers suggested that ENDS and HTP might be possible alternatives to cigarette smoking. In contrast, other studies concluded that its effects are similar to those of conventional smoking [109][110][111].…”

“…Of the 41 included studies (Table 3), 8 evaluated the effect of ENDS on commensal bacteria growth or Streptococcus mutans adhesion to enamel [75,76,82,83,88,92,94,102]; 3 evaluated bacterial composition/diversity [87,95,97], 9 considered metabolic or morphological changes of gingival fibroblasts or gingival endothelial cells after ENDS exposure [71,87,88,[95][96][97]99,100,105]; 6 studied the effect of ENDS on oral cancer cells or gene deregulation in oral epithelial cells [84,86,90,103,104]; and 11 evaluated the effects of vaping on heterogeneous outcomes: C. albicans growth, microbial community changes, gene pathways of normal human oral keratinocytes, and whitening effect on extracted teeth [44,71,73,[78][79][80]85,98,103,104,107]. Four studies [108][109][110][111] evaluated the effect of HTP on human cells for biological and genetical alterations.…”