Polystyrene nanoplastics perturb trophoblastic copper homeostasis with reduced ATP7B expression

BACKGROUND: Polystyrene nanoplastics (PS-NPs) have been detected in human placental and chorionic villous tissues and have been increasingly linked to adverse pregnancy outcomes. However, whether PS-NPs disrupt copper homeostasis in trophoblast cells and whether such disturbances may be relevant to missed abortion (MA) remain unclear. This study aimed to characterize PS-NPs-induced alterations in trophoblast copper homeostasis, explore the underlying copper-regulatory mechanisms, and assess whether related molecular alterations were also present in villous tissues from patients with MA. METHODS: HTR-8/SVneo cells were exposed to PS-NPs, and exploratory analyses were conducted in human placental villous samples. Cell viability, proliferation, and apoptosis were assessed via CCK-8, EdU, and TUNEL assays, respectively. Disruptions in copper homeostasis and metabolic remodeling were evaluated by measuring intracellular copper levels, antioxidant defense-related indicators, and tricarboxylic acid (TCA) cycle metabolites, while associated gene expression was quantified by qRT-PCR. Further validation was performed using ATP7B knockdown, WNT3A-mediated modulation of ATP7B mRNA expression and copper chelation with tetrathiomolybdate (TTM). Exploratory ROC analysis was performed to describe the discriminative ability of ATP7B and FDX1 transcript levels within the present cohort. RESULTS: Our findings revealed that exposure to PS-NPs at 200 μg/mL markedly impaired trophoblast function, evidenced by reduced cell viability and proliferation alongside enhanced apoptotic activity. PS-NPs treatment significantly induced intracellular copper accumulation, disrupted redox homeostasis, and disrupted TCA cycle-associated metabolic pathways, while concurrently suppressing ATP7B mRNA expression and upregulating FDX1 and LIAS mRNA. Notably, ATP7B silencing partially recapitulated the PS-NPs-induced phenotype, including copper overload, impaired antioxidant defense, metabolic dysregulation, and trophoblastic injury. WNT3A treatment increased ATP7B mRNA expression, reduced intracellular copper accumulation, and partially restored cell viability, whereas TTM partially alleviated PS-NPs-induced copper dyshomeostasis, redox imbalance, metabolic perturbations, and cellular injury. Exploratory analysis of villous samples showed reduced ATP7B mRNA and increased FDX1 mRNA in the MA group compared with normal early-pregnancy controls. ROC analysis showed that ATP7B, FDX1, and the combined ATP7B-FDX1 model yielded AUC values of 0.859, 0.750, and 0.832, respectively. CONCLUSIONS: PS-NPs disrupt copper homeostasis and impair trophoblast viability, in which reduced ATP7B expression is functionally involved. Reduced ATP7B expression in MA villous tissues further suggests that ATP7B-associated copper dysregulation may be relevant to the pathophysiology of MA.