Genome-wide methylation changes upon Caco-2 cells exposure to undigested and digested titanium dioxide nanoparticles

Titanium dioxide nanoparticles (TiO₂NPs) are relevant nanomaterials (NMs) for biomedicine and industry, which raise concerns about its effects on human health, particularly through ingestion. Several studies found that exposure to NMs can lead to DNA methylation changes. DNA methylation regulates gene expression, playing a vital role in development and disease, with aberrant methylation linked to cancer and other health conditions. We aimed at identifying DNA methylation changes in intestinal cells exposed to three TiO₂NPs (NM-102, NM-103, NM-105), either digested or undigested. Their cellular effects were investigated by functional pathway and gene ontology (GO) analysis. 48, 41, 55 differentially methylated genes (DMG) were identified after exposure to undigested NM-102, NM-103, NM-105; 71, 65, 55 DMG in the digested counterparts. Undigested TiO₂NPs affected many G-proteins/adenylate cyclase-related pathways (PKA, glucagon, GPER1, CREB1, ADORA2B); the digested had lower impact. Cancer-related pathways were shared. Enriched molecular functions were mainly transcription-related; different biological processes were enriched if TiO₂NPs were digested or not. TiO₂NPs exposure causes DNA methylation changes that have a functional impact on intestinal cells, which differs with its physicochemical properties and digestion. NM-105 caused hypermethylation, unlike the other TiO₂NPs. This study highlights DNA methylation relevance in assessing NMs’ toxicity. Titanium dioxide nanoparticles (TiO₂NP) are widely present in our daily lives, including in food as a white pigment to make it better-looking and appealing. Although there are many toxicological studies on TiO₂NP, few have focused on its effects on DNA methylation, which is a mechanism for regulating gene expression, and consequently, cellular functions. In this study, human intestinal cells were exposed to three different types of TiO₂NP, before and after simulated digestion. Our results indicate that the physical-chemical characteristics of TiO₂NP influence its effects and demonstrate the impact of digestion, relevant in the context of oral exposure. Moreover, they highlight the biological impact of TiO₂NP on intestinal cells through DNA methylation changes and, consequently, the relevance of studying these changes when assessing the adverse effects of nanomaterials on human health.