title: Decoding oral impact
title: tobacco and nicotine effects on the microbiome-immune-metabolome axis
titleType: Subtitle
Creators: name: Burns IV, William J
Publisher: name: University of Iowa
Year: 2025

description: BACKGROUND Tobacco smoking is a well-established risk factor for periodontal disease. Within the last two decades, electronic nicotine delivery systems (ENDS) have grown in popularity and are often marketed as a safer alternative to cigarette smoking. While studies exist characterizing the subgingival environment of nicotine users, there is a relative dearth of literature describing the effect of nicotine consumption on the total oral environment, including saliva and the oral mucosal barrier. As the first contact point for smoke or vapor, the changes that occur in this environment may provide insight into risk and possible key biomarkers that could be used to predict disease onset and progression. Therefore, the aim of this study was to characterize the salivary and mucosal environments of tobacco and nicotine users using multi-omics approaches.
description: METHODS 31 participants (11 control, 11 cigarette smokers, and 10 ENDS users) were recruited for participation in this study. Importantly, all participants were in good health and had no active periodontal disease. Saliva, buccal mucosal swabs, and buccal mucosal biopsies were collected and subjected to downstream processing. DNA was isolated from saliva samples and 16S rRNA sequencing were performed with the Illumina MiSeq platform. Analyses were conducted with DADA2 and QIIME2. Taxonomy identification was referenced against the HOMD database. Alpha and beta-diversity were assessed. Results were visualized with PhyloToAST and core species were identified. Immune mediators were analyzed using Millipore Multiplex kit. Salivary metabolites were assessed using gas chromatography/mass spectrometry (GC/MS) analysis at the Metabolomics Core (FOEDRC). The networks were analyzed within the context of KEGG. Biopsies were paraffin embedded, sectioned, and subjected to spatial transcriptomic analysis using Xenium Analyzer.
description: RESULTS Salivary alpha diversity was not different between groups, though beta diversity was significantly different for cigarette smokers compared to control subjects and ENDS users (PERMANOVA, p = 0.001). Smokers and ENDS users displayed altered salivary abundance of key commensal and pathogenic microbial genera, including Limosilactobacillus, Bacteroidates, and Aggregatibacter. Smokers showed increased concentration of the metabolites aminoadipate, histamine, and mannose, while levels of hypotaurine were significantly lower in smokers, followed by ENDS users. Immune profiling revealed significant alterations in inflammatory species in both smokers and ENDS users. Spatial transcriptomics revealed increases in complement-related genes in smokers, while ENDS users showed increases in TGFB2 and CXC motif transcripts. Network analysis indicated that while control networks were primarily driven by anti-inflammatory components, smokers and ENDS users had networks supported by inflammatory species like IL-1, IL-6, and IL-17.
description: CONCLUSION Our pilot study established the baseline salivary microbiome in tobacco and nicotine users with periodontal health. Immune profiling revealed that both pro- and anti-inflammatory mediators are impacted by these habits. Specific metabolites were increased in smokers, while hypotaurine levels were decreased in concordance with nicotine consumption. At the oral mucosal barrier, key transcripts related to immunity and inflammation were differentially expressed. These results yielded promising lines of future inquiry and may eventually establish biomarkers that could be used for risk assessment.