Transcriptomic investigation of NP toxicity on HepaRG spheroids

Metal nanoparticles (NPs) are commonly used nanomaterials, however concerns have been raised about their toxicity. Although a few studies have reported the toxicity of NPs on cells, they have generally been restricted to a limited variety of NPs, inappropriate cell lines, or culture methods. Thus, the adverse effects remain inadequately understood, necessitating further analysis. This study focuses on assessing the impacts of diverse NPs of varying materials and sizes on HepaRG spheroids to determine the genes that respond to acute NP toxicity. HepaRG cells, the most appropriate alternative to primary hepatocytes, were cultured in 3D spheroids to better mimic the cellular microenvironment of the liver. To elucidate the toxicity mechanisms of NPs on HepaRG spheroids, transcriptome analysis was conducted by using RNA sequencing (RNA-seq). Among all NPs, lowest and highest numbers of differentially expressed genes (DEGs) were found for 40 nm AuNP (118 genes) and InP/ ZnS (1904 genes), respectively. Remarkably, processes such as drug metabolism, sensitivity to metal ions, oxidative stress, endothelial-mesenchymal transition (EMT) and apoptosis consistently exhibited significant enrichment across all NPs of different materials. Pathways related to stress responses of the cells such as the MAPK, p53 and mTOR pathways are found to be dysregulated upon exposure to various NPs. The genes that are common and unique between DEGs of different NPs were identified. These results provide novel insights into the toxicological mechanisms of NPs on HepaRG spheroids.