Availability and effects of n-TiO₂ and PCB77 in fish in vitro models of the intestinal barrier and liver under single- and/or co-exposure scenarios

Titanium dioxide nanoparticles (n-TiO₂) and polychlorinated biphenyls (PCBs) can be present in the food of fish, leading to intestinal exposure uptake, and accumulation in inner organs. This study examined combination effects of n-TiO₂ and PCB77 in vitro models of the fish intestinal epithelium and liver, i.e., RTgut-GC cell cultures grown in ThinCerts™ and RTL-W1 cell cultures grown in standard tissue culture plates. Mass spectrometry and microscopy techniques were used to obtain information on nanoparticle translocation across the intestinal barrier model. In addition, the substances’ effect on intestinal barrier permeability, cell viability, expression of dioxin – and antioxidant response element -controlled genes, and induction of cytochrome P450 1a (Cyp1a)-dependent ethoxyresorufin-O-deethylase (EROD) activity were assessed. TiO₂ nanoparticles were taken up by RTgut-GC cells and detected in the bottom compartment of the intestinal epithelial barrier model. It was not possible to conclude definitively if n-TiO₂ translocation occurred via transcytosis or paracellular migration but observations of nanoparticles in the lateral space between adjacent epithelial cells were rare. PCB77 (1 and 10 µM, 24 h) did not affect barrier permeability, i.e., n-TiO2 translocation is probably not facilitated in case of co-exposure. Furthermore, previous and simultaneous exposure to n-TiO₂ (1 and 10 mg/L, 24 h) did not have any influence on PCB77-induced Cyp1a mRNA and enzyme activity levels in RTL-W1 cells. Furthermore, there were no significant differences in expression of antioxidant response element-controlled genes comparing control, single substance, and mixture treatments, not even following long-term exposure (0.01–1 mg/L n-TiO₂ + 1 nM PCB77, 4 weeks). While an underestimation of the effects of n-TiO₂ and PCB77 cannot be fully excluded as concentration losses due to sorption to cell culture plastics were not measured, the results suggest that the test substances probably have a low potential to exhibit combination effects on the assessed endpoints when co-existing in fish tissues.