Endogenous 15(S)-hydroxyeicosatetraenoic acid mediates amplified estrogenic responses under nanoplastic-homosalate coexposure.

Micro- and nanoplastics (MNPs) can enhance the toxicity of co-occurring chemicals via a proposed "Trojan horse" effect, yet the underlying mechanisms remain unclear. Here, we investigated the estrogenic effects of coexposure of ultraviolet filter homosalate (HMS) and polystyrene nanosphere (PNS) using ovariectomized mice (HMS: 0.1 and 1 mg/kg; PNS: 2.5 mg/kg) and human cell models (HMS: 0.01 - 1 μM; PNS: 1 mg/L). In mice, HMS-PNS coexposure significantly increased uterine weight, promoted mammary gland proliferation, and upregulated estrogen receptor 1 and its downstream targets amphiregulin and progesterone receptor. Integrated metabolomic and transcriptomic analyses identified endogenous 15(S)-hydroxyeicosatetraenoic acid (15(S)-HETE) as a key mediator of these effects in mammary glands. In MCF-7 cells, HMS-PNS coexposure elevated 15(S)-HETE levels, promoting cell proliferation via the estrogen receptor alpha-arachidonate 15-lipoxygenase (ERα-ALOX15) axis. At a concentration of 100 nM 15(S)-HETE, pharmacological inhibition of phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) abrogated cell proliferation and serum and glucocorticoid-regulated kinase 1 (SGK1) activation. Moreover, immunoprecipitation and docking analyses suggested a direct interaction between 15(S)-HETE and SGK1. Knockdown of ALOX15, or PI3K/AKT inhibition, suppressed HMS-PNS-induced cell proliferation. Taken together, these results demonstrated that HMS-PNS coexposure amplifies estrogenic responses through ERα-ALOX15-dependent 15(S)-HETE production and PI3K/AKT/SGK1 signaling. Our findings uncover a mechanistic pathway beyond the canonical "Trojan horse" effect, providing new insight into how MNPs modulate endocrine-disrupting activity of co-occurring contaminants and informing future risk assessment of combined environmental exposures.