doi:10.1016/j.biopha.2025.118950engRozman, IzaGallo-Cordova, AlvaroPuerto Morales, María delMorales Ovalle, Marco A.Goya, Gerardo F.Kološa, KatjaHocevar, DomenŽegura, BojanaŠtern, AljaSafety of ferrite nanoparticles for biomedical applications: Cyto- and genotoxic effects of MxFe3-xO4 (M = Fe, Zn, Mn) in an Advanced 3D human hepatic in vitro modelART-2026-147999Given the growing interest in nanosized spinel-type ferrite nanoparticles for biomedical applications and the limited information on their safety, this study aimed to assess their cellular and genotoxic effects in an in vitro 3D human hepatic cell model (HepG2 spheroids). Ferrite nanoparticles – γFe2O3 (FeNPs; 14 ± 4 nm), Zn0.7Fe2.3O4 (ZnNPs; 14 ± 5 nm), and Mn0.4Fe2.6O4 (MnNPs; 7 ± 2 nm) – were synthesised through a microwave-assisted polyol route, functionalized with citric acid, and characterised using Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES), Transmission Electron Microscopy (TEM), X-ray Diffraction (XRD), and Fourier Transform Infrared Spectroscopy (FTIR). Nanoparticle uptake was analysed using TEM, cytotoxicity was measured with CellTiter-Glo®, and oxidative stress induction was assessed using the 2′,7′-Dichlorodihydrofluorescein diacetate (DCFH-DA) and malondialdehyde (MDA) assay. Genotoxic effects were evaluated using the comet, γH2AX and p-H3 assays. Cellular stress responses were assessed using toxicogenomic analysis. Significant cytotoxicity of the tested nanoparticles (0.1–250 µg/mL) was observed; however, TEM analysis revealed limited penetration to the outermost cell layers of spheroids. Notably, only FeNPs induced ROS generation, while MDA levels remained unchanged in all tested samples. Low DNA damage was detected at 24 h, but a significant increase was observed at 96 h (5–50 µg/mL). No increase in γH2AX or p-H3 was found. No substantial alterations in DNA damage or oxidative stress-response gene expression were detected. Altogether, our findings suggest that the effects of ferrite nanoparticles are time- and composition-dependent, underlining the importance of further mechanistic and chronic exposure evaluations in 3D cell models.2026http://zaguan.unizar.es/record/16861210.1016/j.biopha.2025.118950http://zaguan.unizar.es/record/168612oai:zaguan.unizar.es:168612info:eu-repo/grantAgreement/EC/HORIZON EUROPE/101079113/EU/TWINNING FOR EXCELLENCE TO STRATEGICALLY ADVANCE RESEARCH IN CARCINOGENESIS AND CANCER/CutCancerinfo:eu-repo/grantAgreement/EC/H2020/101007629 /EU/Nanomaterials for Enzymatic Control of Oxidative Stress Toxicity and Free Radical Generation/NESTORThis project has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement No H2020 101007629 -NESTORBIOMEDICINE & PHARMACOTHERAPY 195 (2026), 118950 [15 pp.]byhttps://creativecommons.org/licenses/by/4.0/deed.esinfo:eu-repo/semantics/openAccess