Cytological and transcriptomic responses of WI-38 human lung fibroblasts to plant-exposed metal nanoparticle residues.

Metal-based nanoparticles are increasingly encountered in human exposure scenarios, yet the relationship between residue-level exposure and cellular hazard remains unclear. In this study, iron (Fe), copper (Cu), and silver (Ag) nanoparticles were applied to chilli pepper plants to estimate plant-associated metal nanoparticle residues in the edible fruits. Subsequently, exposure-relevant concentration ranges were defined and examined in normal human WI-38 fibroblasts using integrated cytological and transcriptomic approaches. Metal accumulation was higher in vegetative tissues than fruits, with Fe ≤ 10.6 mg/kg, Cu ≤ 2.4 mg/kg, and Ag ≤0.1 mg/kg. In vitro cytotoxicity analysis following 24 h exposure revealed material-specific dose-response relationships, with IC50 values of 15.2 ppm for Fe nanoparticles, 0.32 ppm for Cu nanoparticles, and 6.7 ppm for Ag nanoparticles. Exposure at IC50 concentrations induced measurable cytological alterations, including increased total apoptosis (up to 11.9%) and accumulation of cells in the G0/G1 phase. RNA sequencing identified a distinct transcriptional response to combined nanoparticle exposure, with enrichment of pathways related to inflammatory signaling (TNF, NF-κB, IL-17), cytokine-cytokine receptor interaction, metal and oxidative stress handling (mineral absorption, ferroptosis), and stress-associated remodeling pathways. Guided by these enriched pathways, targeted RT-qPCR validation demonstrated nanoparticle-specific gene regulation, including strong induction of TNF (7.06-fold) and the membrane stress-associated gene PROM2 (8.63-fold) following Cu nanoparticle exposure, while Ag nanoparticles selectively upregulated PROM2 (4.98-fold). These results highlight the disconnect between low residue levels and cellular hazard, emphasizing cumulative exposure, sensitive endpoints, and the value of integrating residue analysis with human cell-based mechanistic assessment for risk evaluation.