Influence of smoking on methylation and hydroxymethylation levels in global DNA and specific sites of KRT14, KRT19, MIR-9-3 and MIR-137 genes of oral mucosa.

Objective: The aim of this study was to investigate the influence of smoking on the methylation and hydroxymethylation status of global DNA and specific sites of KRT14, KRT19, miR-9-3 and miR-137 genes in the healthy oral mucosa.

Methods: Samples of oral epithelial cells were collected using mouthwash from a population of former and current smokers and those who had never smoked. Genomic DNA was extracted, and global DNA methylation and hydroxymethylation was performed using an ELISA-based technique; DNA methylation at specific sites was performed using Methylation-Specific PCR (MSP) (KRT14, miR-9-3 and miR-137) or Methylation-Sensitive Restriction Enzymes (MSRE) (KRT19).