Investigations of acute effects of polystyrene and polyvinyl chloride micro- and nanoplastics in an advanced in vitro triple culture model of the healthy and inflamed intestine.

The continuous degradation of plastic waste in the environment leads to the generation of micro- and nanoplastic fragments and particles. Due to the ubiquitous presence of plastic particles in natural habitats as well as in food, beverages and tap water, oral exposure of the human population with plastic particles occurs worldwide. We investigated acute toxicological effects of polystyrene (PS) and polyvinyl chloride (PVC) micro- and nanoparticles in an advanced in vitro triple culture model (Caco-2/HT29-MTX-E12/THP-1) mimicking the healthy and inflamed human intestine to study the effect of inflammatory processes on plastic particle toxicity. We monitored barrier integrity, cytotoxicity, cell layer integrity, DNA damage, the release of pro-inflammatory cytokines (IL-1ß, IL-6, IL-8 and TNF-α) and mucus distribution after 24 h of particle exposure. In addition, we investigated cytotoxicity, DNA damage and IL-1ß release in monocultures of the three cell lines. Amine-modified polystyrene nanoparticles (PS-NH2) served as a positive control for particle-induced toxicity. No acute effects in the investigated endpoints were observed in the model of the healthy intestine after PS or PVC exposure. However, during active inflammatory processes, exposure to PVC particles was found to augment the release of IL-1ß and to cause a loss of epithelial cells. Our results suggest that prevalent intestinal inflammation might be an important factor to consider when assessing the hazard of ingested micro- and nanoplastic particles.

Assessing the NLRP3 Inflammasome Activating Potential of a Large Panel of Micro- and Nanoplastics in THP-1 Cells.

Due to the ubiquity of environmental micro- and nanoplastics (MNPs), inhalation and ingestion by humans is very likely, but human health effects remain largely unknown. The NLRP3 inflammasome is a key player of the innate immune system and is involved in responses towards foreign particulate matter and the development of chronic intestinal and respiratory inflammatory diseases. We established NLRP3-proficient and -deficient THP-1 cells as an alternative in vitro screening tool to assess the potential of MNPs to activate the NLRP3 inflammasome. By investigating cytokine release (IL-1ß and IL-8) and cytotoxicity after treatment with engineered nanomaterials, this in vitro approach was compared to earlier published ex vivo murine bone marrow-derived macrophages and in vivo data. This approach showed a strong correlation with previously published data, verifying that THP-1 cells are a suitable model to investigate NLRP3 inflammasome activation. We then investigated the proinflammatory potential of eight MNPs of different size, shape, and chemical composition. Only amine-modified polystyrene (PS-NH2) acted as a direct NLRP3 activator. However, polyethylene terephthalate (PET), polyacrylonitrile (PAN), and nylon (PA6) induced a significant increase in IL-8 release in NLRP3-/- cells. Our results suggest that most MNPs are not direct activators of the NLRP3 inflammasome, but specific MNP types might still possess pro-inflammatory potential via other pathways.

An inverted in vitro triple culture model of the healthy and inflamed intestine: Adverse effects of polyethylene particles.

As environmental pollution with plastic waste is increasing, numerous reports show the contamination of natural habitats, food and drinking water with plastic particles in the micro- and nanometer range. Since oral exposure to these particles is virtually unavoidable, health concerns towards the general population have been expressed and risk assessment regarding ingested plastic particles is of great interest. To study the intestinal effects of polymeric particles with a density of <1 g/cm³ in vitro, we spatially inverted a triple culture transwell model of the healthy and inflamed intestine (Caco-2/HT29-MTX-E12/THP-1), which allows contact between buoyant particles and cells. We validated the inverted model against the original model using the enterotoxic, non-steroidal anti-inflammatory drug diclofenac and subsequently assessed the cytotoxic and pro-inflammatory effects of polyethylene (PE) microparticles. The results show that the inverted model exhibits the same distinct features as the original model in terms of barrier development and inflammatory parameters. Treatment with 2 mM diclofenac causes severe cytotoxicity, DNA damage and complete barrier disruption in both models. PE particles induced cytotoxicity and pro-inflammatory effects in the inverted model, which would have remained undetected in conventional in vitro approaches, as no effect was observed in non-inverted control cultures.

Evaluation of cytotoxic effects and oxidative stress with hydroxyapatite dispersions of different physicochemical properties in rat NR8383 cells and primary macrophages.

Enhanced cytotoxicity and oxidative stress through reactive oxygen species (ROS) formation are discussed as relevant parameters regarding potential hazardous properties of nanomaterials. In this study, the biocompatibility of five hydroxyapatite materials of different size and morphology, i.e., nano/needle-shaped (HA-NN), nano/rod-like (HA-NR), nano/plate-like (HA-NP), fine/dull needle-shaped (HA-FN), and a hydroxyapatite-protein-composite (HPC), was investigated in rat NR8383 and primary alveolar macrophages. Lipopolysaccharide (LPS) and DQ12 quartz served as positive controls. In the water-soluble tetrazolium salt 1 (WST-1) and lactate dehydrogenase (LDH) assays with NR8383 cells, no cytotoxicity was observed for HPC and the pure hydroxyapatite samples up to 3000 microg/ml, while HA-FN showed a significant effect at the highest dose in the LDH assay. In primary cells, no cytotoxicity was observed with all samples up to 300 microg/ml. ROS generation measured by electron paramagnetic resonance (EPR) technique was significantly enhanced with HA-NN and HPC in NR8383 cells. No effect was detected in primary cells, which are considered more relevant to physiological conditions. All hydroxyapatites elicited TNF-alpha release from the NR8383 cells, but with significantly lower potency than DQ12 quartz and LPS. In conclusion, combined findings in both cell types support a good biocompatibility of the pure hydroxyapatite samples as well as of the hydroxyapatite-protein-composite.