P52 Investigating the impact of carboxylated polystyrene nanoplastics in liver using hepatic cell lines and human precision-cut liver slices

Background and Aims Large plastic products degrade during use and after disposal into small particles described as microplastics (<5 mm, MP) and nanoplastics (<1µm, NP). These MP/NPs accumulate in the environment and can enter the human body via ingestion, inhalation or dermal contact. Several studies have reported MP/NP in human tissue, raising concerns about potential adverse health outcomes. Existing research on NP toxicity is variable and lacks sufficient understanding of the liver-associated pathophysiology. To understand the impact of NP on the liver, we determined the uptake and effects of commercially available carboxylate-modified 20 nm polystyrene nanoparticles (PS NPs) in three hepatic models: human carcinoma hepatocytes (HepG2), immortalised human hepatocytes (IHH), and human precision-cut liver slices (hPCLS). Methods We visualised and quantified PS NPs uptake in all three hepatic models using confocal microscopy and flow cytometry. We analysed post-exposure cytotoxicity by assessing cell viability via viability staining, and cell injury by measuring intracellular enzymes (LDH, AST and ALT) levels in the media. Furthermore, we examined the subtle physiological effects of PS NPs in HepG2 cells by estimating relative reactive oxygen species (ROS) production with the DCFHDA assay and comparing mitochondrial respiration using the Agilent Seahorse XFe96 analyser. Results Exposure experiments show model and dose-dependent uptake of PS NPs after 24 and 48 hours of exposure in all three hepatic models. 63.0 ± 31.5% IHH cells showed PS NPs uptake at a concentration of 0.1 µg/mL, which is considerably lower than the estimated plastic concentration in human blood (1.8 -4.7 µg/mL). Cell viability of IHH cells decreased from 95.2 ± 2% in the control group to 9.7 ± 7.1% at 100 µg/mL after 48 hours of exposure. Early signs of cell injury were observed with elevated AST levels in the media of IHH and hPCLS at high PS NP concentrations. HepG2 cells showed no changes in cell viability, redox state, or mitochondrial respiratory parameters after exposure. Conclusion The study demonstrates that PS NPs can readily accumulate in hepatocytes, even at a concentration as low as 0.1 µg/mL in IHH cells, and cause cell damage and reduce cell viability at a high dose of 100 µg/mL. The effect of PS NPs is model-dependent, highlighting the need for physiologically relevant toxicity models to study the underlying mechanisms of NP cytotoxicity. These findings emphasise the urgent need to investigate the health effects of NP pollution on human liver health in complex toxicity models.