Co-exposure of polystyrene nanoplastics and copper induces development toxicity and intestinal mitochondrial dysfunction in vivo and in vitro

Nanoplastics (NPs) have raised concerns about the combined toxicity to living organisms due to their ability to adsorb heavy metals. There is still uncertainty, however, whether NPs combined with heavy metals exert adverse effects on intestinal microenvironment, especially the intestinal cells and microbiota. Herein, the combined effects of 500 nm spherical-shaped polystyrene nanoplastics (PSNPs) and copper ions (Cu2+) on intestinal cells and gut microbiota were assessed using HCT-116 cells and zebrafish models. The combined exposure of PSNPs (10 mg/L) and Cu2+ (0.5 mg/L) induced more severer hatching interference of zebrafish embryos, deformation, and mortality. In larval stage, PSNPs (10 mg/L) accumulated and carried more Cu2+ in the gastrointestinal tract (GIT) of zebrafish after co-exposure for 5 days. Excessive neutrophil recruitment and oxidative stress in GIT of zebrafish larvae were observed. The mechanism of the combined toxicity was revealed by transmission electron microscopy (TEM) showing the injuries of GIT, transcriptome and 16S rDNA gene sequencing showing the toxicity pathways, including oxidative phosphorylation and respiratory electron transport chain, as well as microbial community analysis showing the induced microbiota dysbiosis. In vitro tests using HCT-116 cells showed that PSNPs (10 mg/L) and Cu2+ (0.5 mg/L) increased cell death while decreasing ATP concentration and mitochondrial membrane potential after 48 h exposure. These findings may provide new insights into the combined toxicity of nanoplastics and heavy metals in the intestinal microenvironment.