*6.1. Inflammation*

An in vitro study using various sizes of polystyrene particles found that larger particles (202 nm and 535 nm) produced inflammatory effects on human A549 lung cells. There was higher IL-8 expression by the lung cells treated with the larger particles, in comparison with the same cells exposed to 64 nm particles [112]. Furthermore, unaltered or carboxylated polystyrene nanoparticles brought on a substantial up-regulation of IL-6 and IL-8 genes in human gastric adenocarcinoma, leukemia, and histiocytic lymphoma cells, which suggests that the increase in inflammatory reactions to polystyrene particles is likely to be due to the constitution of the particle, or down to simple particle occurrence rather than because of the particle charge [113,114].

A study on the impact of carboxylated and amino-modified polystyrene particles (120 nm) on the polarization of human macrophages into M1 or M2 phenotypes revealed no change in the expression of M1 markers like CD86, NOS2, TNF α, and IL-1β [115]. However, the introduction of both types of nanoparticles negatively impacted the expression of scavenger receptors CD163 and CD200R, and the release of IL-10 by M2 cells. There was a reduction of *Escherichia coli* phagocytosis by both M1 and M2 macrophages with the introduction of amino-modified particles. On the other hand, phagocytosis by M2 was unaffected by the carboxylated particles. The carboxylated particles also caused increases in the protein mass in M1 and M2, enhanced the release of TGFβ1 by M1, and heightened levels of ATP in M2 [115]. Similarly, in vitro study also shown that unmodified polyethylene particles measuring between 0.3 and 10 μm caused murine macrophages to produce significant levels of cytokines, such as IL-6, IL-1β, and TNF α [116,117].

Furthermore, a number of previous studies indicated that when polyethylene components are used as prostheses, they can fragmentize as a result of wear and tear, and form debris in the joints [118–120]. The polyethylene wear particles trigger the TNF α and IL-1 pro-inflammatory factors, as well as pro-osteoclastic factors, including the receptor activator of NF-κB ligand (RANKL), which causes periprosthetic bone resorption and could eventually result in the patient losing the prosthesis [121]. High levels of plastic particles measuring between 0.2 and 10 μm have been observed in the periprosthetic tissue of ultrahigh molecular weight polyethylene-based implants. In addition, the presence of macrophages in the tissues in the vicinity of the implant area indicates the stimulation of the inflammatory response [121–123]. A study of failed titanium alloy total hip arthroplasty cases found that a majority of wear debris in the interfacial membranes consisted of polyethylene particles with an average diameter of 530 nm [124]. To overcome the observed negative effects, surgeons are now increasingly employing metal-on-metal joint replacements.


**Table 1.** Summary of potential toxic effects of micro- and nanoplastics on human health.

### *6.2. Oxidative Stress and Apoptosis*

A number of in vitro studies have shown that different polystyrene nanoparticles can induce oxidative stress, apoptosis and autophagic cell death in cell context-dependent manner. For instance, amine-modified polystyrene nanoparticles were shown to interact and aggregate with mucin strongly, and induce apoptosis of mucin- and non-mucinsecreting intestinal epithelial cells [126]. Cationic polystyrene nanoparticles were shown to induce reactive oxygen species (ROS) production and endoplasmic reticulum (ER) stress in mouse macrophages and lung epithelial cells via aggregation of misfolded protein, leading to autophagic cell death of RAW 264.7 mouse macrophages and BEAS-2B lung epithelial cells [127,128]. While, unmodified or functionalized polystyrene was shown to induce apoptosis in several human cell types, including primary human alveolar macrophages (MAC), primary human alveolar type 2 (AT2) epithelial cells, human monocytic leukemia cell line (THP-1), human immortalized alveolar epithelial type 1 cells (TT1), human colon carcinoma cells (Caco-2), and human lung cancer cells (Calu-3) [129–132]; and polystyrene nanoparticles were shown to regulate ROS via long non-coding RNAs (e.g., *linc-61, linc-50, linc-9, and linc-2*) in *Caenorhabditis elegans* [142].

Despite the toxic effects observed in the in vitro models, no obvious severe toxicity was observed in liver, duodenum, ileum, jejunum, large intestine, testes, lungs, heart, spleen, and kidneys of mice following oral exposure of a mixture of microplastics [139]. While, other studies have demonstrated that oral exposure (either through oral gavage or drinking water) caused liver inflammation [125], neurotoxic responses [125], reduced body and liver weight [140], reduced mucin excretion in colon [134,140], altered amino acid and bile acid metabolism [134,135], and altered microbiota composition [134,140,141,143]. Interestingly, some of the effects such as altered lipid metabolism was observed even in the offspring of the mice following microplastic exposure [141].
