Polystyrene nanoplastics as an ecotoxicological hazard: cellular and transcriptomic evidences on marine and freshwater in vitro teleost models

The contamination of marine and freshwater environments by nanoplastics is considered a global threat for aquatic biota. Taking into account the most recent concentration range estimates reported globally and recognizing a knowledge gap in polystyrene nanoplastics (PS-NPs) ecotoxicology, the present work investigated the harmful effects of 20 nm and 80 nm PS-NPs, at increasing biological complexity, on the rainbow trout Oncorhynchus mykiss RTG-2 and gilthead seabream Sparus aurata SAF-1 cell lines. Twenty nm PS-NPs exerted a greater cytotoxicity than 80 nm ones and SAF-1 were approximately 4-fold more vulnerable to PS-NPs than RTG-2. The engagement of PS-NPs with plasma membranes was accompanied by discernible uptake patterns and morphological alterations along with a nuclear translocation already within a 30-min exposure. Cells were structurally damaged only by the 20 nm PS-NPs in a time-dependent manner as indicated by distinctive features of the execution phase of the apoptotic cell death mechanism such as cell shrinkage, plasma membrane blebbing, translocation of phosphatidylserine to the outer leaflet of the cell membrane and DNA fragmentation. At last, functional analyses unveiled marked transcriptional impairment at both sublethal and lethal doses of 20 nm PS-NPs, with the latter impacting the "Steroid biosynthesis", "TGF-beta signaling pathway", "ECM-receptor interaction", "Focal adhesion", "Regulation of actin cytoskeleton" and "Protein processing in endoplasmic reticulum" pathways. Overall, a distinct ecotoxicological hazard of PS-NPs at environmentally relevant concentrations was thoroughly characterized on two piscine cell lines. The effects were demonstrated to depend on size, exposure time and model, emphasizing the need for a comparative evaluation of endpoints between freshwater and marine ecosystems.