Adsorption of hematite nanoparticles onto Caco-2 cells and the cellular impairments: effectof particle size

Abstract

The increasing applications of engineered nanomaterials nowadays have elevated thepotential of human exposure through various routes including inhalation, skin penetrationand digestion. To date there is scarce information on a quantitative description of theinteractions between nanoparticles (NPs) and cell surfaces and the detrimental effects from theexposure. The purpose of this work was to study in vitro exposure of Caco-2 cells to hematite (α-Fe2O3) NPs and to determine the particle size effects on the adsorption behaviors. Cellularimpairment was also investigated and compared. Hematite NPs were synthesized as part ofthis study with a discrete size distribution and uniform morphology examined by dynamiclight scattering (DLS) and confirmed by transmission electron microscopy (TEM). Caco-2cells were cultured as a model epithelium to mirror human intestinal cells and used toevaluate the impacts of the exposure to NPs by measuring transepithelial electricalresistance (TEER). Cell surface disruption, localization and translocation of NPs throughthe cells were analyzed with immunocytochemical staining and confocal microscopy.Results showed that hematite NPs had mean diameters of 26, 53, 76 and 98 nm and werepositively charged with minor aggregation in the buffer solution. Adsorption of thefour sizes of NPs on cells reached equilibrium within approximately 5 min butadsorption kinetics were found to be size-dependent. The adsorption rates expressed asmg m − 2 min − 1 were greater for large NPs (76 and 98 nm) than those for small NPs(26 and 53 nm). However, adsorption rates, expressed in units ofm − 2 min − 1, were much greater for small NPs than large ones. After the adsorptionequilibrium was reached, the adsorbed mass of NPs on a unit area of cells wascalculated and showed no significant size dependence. Longer exposure time (>3 h) induced adverse cellular effects as indicated by the drop in TEER compared tothe control cells without the exposure to NPs. NPs initially triggered a dynamicreorganization and detachment of microvilli structures on Caco-2 cell surfaces.Following this impact, the drop in TEER occurred more significantly, particularlyfor the exposure to 26 nm NPs, which was consistent with the observations withconfocal microscopy that the junctions were more severely disrupted by 26 nm NPsthan other sizes. In conclusion, this paper demonstrates the interactions at theultrastructural level from initial surface adsorption of NPs upon cells, to thesubsequent microvilli reorganization, membrane penetration and the disruption ofadherens junction and provides the fundamental information on size effects on NPbehavior which is often poorly addressed for in vitro cytotoxicity studies of NPs.