Exclusive heat-not-burn cigarette smoking alters the profile of circulating microRNAs.
Background. Traditional combustion cigarette (TCC) smoking is an established risk factor for several types of cancer and cardiovascular diseases. Circulating microRNAs (miRNAs) represent key molecules mediating pathogenetic mechanisms, and potential biomarkers for personalized risk assessment. Several transcriptomics studies have showed that TCC smoking globally changes the profile of circulating miRNAs. The use of heat-not-burn cigarettes (HNBCs) as alternative smoking devices is rising exponentially worldwide, with still unknown long-term effects on health. Comprehensive data on the circulating miRNA profile in chronic HNBC smokers are still lacking. Purpose. We aimed at defining the profile of circulating miRNAs in serum samples of chronic exclusive HNBC smokers, and identifying potentially pathogenetic circulating miRNA signatures. Results. Differential expression analysis revealed 101 miRNAs significantly modulated between groups (adjusted p<0.05), with a prevalence of circulating miRNA downregulation in smokers (only 20 miRNAs were upregulated in both groups of smokers). Head-to-head comparison revealed that 65 miRNAs were exclusively modulated in TCC, 26 miRNAs were commonly modulated in both smoker groups, and 10 miRNAs were exclusively modulated in HNBC samples, all versus NS. KEGG pathway analysis performed on target genes of the commonly modulated miRNAs returned cancer-associated and cardiovascular disease pathways. The application of stringent abundance and fold-change criteria to this list nailed down our functional bioinformatic analyses to a network where miR-25-3p and miR-221-3p are main hubs for cardiovascular- and cancer- related molecular pathways, respectively. Conclusions. Our results define for the first time a global miRNA profile in the serum of exclusive chronic HNBC smokers. Despite the globally lower extent of the differences observed, our data suggest a significant detrimental impact of HNBCs on circulating miRNAs. Overall design: Serum samples were obtained from subjects previously enrolled in the SUR-VAPES CHRONIC trial, including three groups of 20 healthy young subjects, stratified as chronic HNBC smokers (mean exclusive use of HNBC 1.5±0.5 years), TCC smokers, and non-smokers (NS). Four sub-groups of five subjects each were randomly created for each group (tested for non-statistical differences in available anthropometric and clinical parameters). Three sub-groups were used to create three pooled samples per group for small RNA sequencing, and the fourth sub-group constituted the validation set. Data were analyzed using DEseq2.