Nanoplastics and thermal stress induce synergistic toxicity in marine medaka (Oryzias melastigma) through epithelial barrier damage, immuno-metabolic dysregulation, and cancer-associated transcriptional signatures.

Marine organisms are increasingly exposed to nanoplastics (NPs) and rising temperatures due to climate change, yet their combined effects remain poorly understood. Here, we investigated the interactive toxicity of polystyrene nanoplastics (PS-NPs, 80 nm) and elevated temperature (32 °C vs 25 °C) in marine medaka larvae (Oryzias melastigma) using integrated bioaccumulation analysis, histopathology, and transcriptomic profiling. Our results revealed that the digestive tract served as the primary tissue for NPs bioaccumulation, with elevated temperature significantly accelerating particle bioaccumulation (1.20-6.27-fold increase) and exacerbating shedding of intestinal villi. Transcriptomic analysis identified temperature as the dominant driver of transcriptional remodeling, while co-exposure induced unique synergistic effects not observed under individual stressors. Specifically, combined exposure amplified immuno-inflammatory responses through complement cascade and phagosome pathways, with NPs perceived as pseudo-pathogenic threats triggering respiratory burst and apoptosis. Concurrently, antagonistic interactions suppressed critical protective responses including heat shock protein hsp90α1 and immune regulators (e.g., il2rg and ptx3). Most concerning, co-exposure uniquely enriched multiple cancer-related pathways absent from individual stress conditions, with synergistic upregulation of the proto-oncogene myc-2. These findings demonstrated that elevated temperature fundamentally altered NPs toxicity from localized intestinal stress to broad immuno-inflammatory dysregulation, creating a transcriptional state associated with cancer-related pathways. Additionally, a phenotypic interaction analysis on catalase (CAT), superoxide dismutase (SOD), malondialdehyde (MDA), reduced glutathione (GSH) and intestinal villus morphometry (length and width) showed that the standardized Cohen's d interaction indices followed a consistent trend matching transcriptome-level predictions of synergistic antioxidant induction and barrier impairment. Our results highlighted that current single-stressor risk assessments underestimate NPs threat under climate change scenarios, emphasizing the urgent need for multi-stressor frameworks in environmental protection.