Differential cytotoxicity to human cells in vitro of tire wear particles emitted from typical road friction patterns: The dominant role of environmental persistent free radicals

Tire wear particles (TWPs) have been recognized as one of the major sources of microplastics (MPs), however, effects of initial properties and photochemical behavior of TWPs on cytotoxicity to human cells in vitro have not been reported. Therefore, here, three TWPs generated from typical wear of tires and pavements (i.e., rolling friction (R-TWPs) and sliding friction (S-TWPs)) and cryogenically milled tire tread (C-TWPs), respectively, and their photoaging counterparts were used to study the reasons for their differential cytotoxicity to 16HBE cells in vitro. Results showed in addition to changes of surface structure and morphology, different preparation methods could also induce formation of different concentration levels of environmental persistent free radicals (EPFRs) (from 1.24 to 3.06 × 1017 spins/g with g-factors ranging 2.00307-2.00310) on surfaces of TWPs, which contained 7.3%-65.8% of reactive EPFRs (r-EPFRs). Meanwhile, photoaging for 90 d could strengthen formation of EPFRs (from 4.03 to 4.61 × 1017 spins/g) with containing 74.7%-78.1% r-EPFRs on surfaces of TWPs and improve their g-factor indexes (ranging 2.00309-2.00313). At 100 μg mL-1 level, compared to C-TWPs, both R-TWPs and S-TWPs (whether photoaging or not) carried higher intensity EPFRs could significantly inhibit 16HBE cells proliferation activity, cause more cells oxidative stress and induce more cell apoptosis/necrosis and secretion of inflammatory factor (P < 0.05). However, regardless of how TWPs were prepared, photoaged or not, exposure at a concentration of 1 μg mL-1 appeared to be non-acute cytotoxic. Correlation analysis suggested dominant toxicity of TWPs was attributed to the formation of r-EPFRs on their surfaces, which could promote accumulation of excess reactive oxygen species in cells and the massive deposition of intracellular particles. This study provides direct evidence of TWPs cytotoxicity, and underlining the need for a better understanding of the influences of initial properties and photochemical characteristics on risk assessment of TWPs released into the environment.