Pediatric Buccal Epithelial Cell Proportions Decrease Reliably With Age: Considerations for PedBE in Pediatric Research

Children’s cheek swabs, cellularly heterogeneous samples consisting primarily of buccal epithelial cells (BECs), are widely used in epigenetic association studies of pediatric DNA methylation and biomarker creation, e.g., the Pediatric Buccal Epigenetic (PedBE) clock. Although BEC proportion is suggested to decrease with age, this was not investigated during childhood. Here, we analyzed cheek swabs from 3799 typically developing children 2-months to 20-years-old. Estimated BEC proportion declined, while estimated neutrophil proportion increased, with chronological and predicted epigenetic age, and variability in both cell types increased with age. Although estimated BEC proportion was weakly related to epigenetic age acceleration (EAA), this was attenuated to non-significance after correcting for BECs during initial EAA estimation. Notably, correcting for BEC proportion attenuated the association of PedBE EAA with obsessive-compulsive disorder and strengthened the association with diurnal cortisol slope. Thus, the change in children’s oral cell proportions with age is a crucial consideration for cell type-sensitive research.