Co-exposure to polystyrene nanoplastics and glyphosate exacerbates NETs-mediated pyroptosis by activating the NLRP3 inflammasome in mouse liver

Nanoplastics (NPs), highly prevalent due to large-scale plastic production, and glyphosate (Gly), the most utilized herbicides worldwide, are ubiquitous environmental contaminants. Growing concerns highlight that NPs can act as vectors for various pollutants like Gly, but their combined toxic effects in mammals and the underlying mechanisms remain poorly understood. In this study, the hepatotoxicity and potential mechanisms under the exposure of polystyrene nanoplastics (PSNPs) and/or Gly in vivo and in vitro were investigated. Mice were treated with PSNPs (25 mg/kg/day) and/or Gly (50 mg/kg/day) by oral gavage for 5 weeks. Results showed that exposure to PSNPs or Gly caused liver injury in mice, with co-exposure resulting exacerbated hepatotoxicity, evidenced by increased neutrophil infiltration and ultrastructural damages, elevated oxidative stress (LPO, H 2 O 2 , T-AOC and CAT), increased neutrophil chemokines (CCL2, CXCL12) and marker of neutrophil extracellular traps (NETs) formation (MPO), and upregulated pyroptosis-related factors (TLR4, NF-κB, NLRP3, ASC, Caspase-1, GSDMD, IL-1β, IL-18). In addition, a co-culture system of peripheral blood neutrophils (PBNs) and AML12 cells was established, revealing that co-exposure amplified ROS production, NETs formation (SYTOX Green staining) and pyroptosis. Notably, inhibition of the NLRP3 inflammasome significantly reduced NETs production, and degradation of NETs substantially decreased pyroptosis, demonstrating feedback between NETs and the NLRP3 inflammasome that drives inflammation induced by PSNPs and Gly. These results highlight that co-exposure exacerbated NETs-mediated pyroptosis through NLRP3 inflammasome activation. Our study equips new reference for understanding the mechanistic insights and health implications of the combined toxicity of PSNPs and Gly. • Co-exposure to PSNPs and Gly exacerbates liver injury in mice. • Co-exposure to PSNPs and Gly aggravates NETs-mediated pyroptosis. • Feedback between NETs and NLRP3 inflammasome drives inflammation and pyroptosis.