Bioinspired zinc-doped carbon dots protect against polystyrene microplastic-induced spermatogenic dysfunction through Nrf2 activation and cuproptosis suppression

Abstract

Polystyrene microplastics (PS-MPs) are increasingly recognized for their detrimental effects on reproductive health, particularly in the male reproductive system. While mitochondrial oxidative stress is recognized as a driver of MPs-induced reproductive injury, it remains unclear whether this process involves cuproptosis, a newly identified copper-dependent form of regulated cell death, and elucidating this potential link is critical for alleviating such damage. Here, we report that PS-MPs exposure induces testicular injury and spermatogenic dysfunction in mice, associated with redox imbalance, copper transport defects, and upregulation of the cuproptosis regulators Ferredoxin 1 (FDX1) and Lipoic Acid Synthetase (LIAS). In turn, zinc-doped trifolirhizin-derived carbon dots (ZnTFZCDs) were rationally designed via a one-step hydrothermal strategy, embedding the antioxidant properties of a natural flavonoid and the reproductive-protective role of zinc into a biocompatible carbon nanoplatform. ZnTFZCDs are well-dispersed carbon dots rich in oxygen/nitrogen-containing groups with Zn coordination, and demonstrate favorable biosafety without overt cytotoxicity, hemolysis, or organ damage. In vitro, ZnTFZCDs potently scavenge intracellular reactive oxygen species (ROS), maintain antioxidant enzyme activities, preserve mitochondrial membrane potential, suppress lipid peroxidation, and rescue Glutathione peroxidase 4 (GPX4) expression in MPs-exposed RAW264.7 and GC-2 cells. Notably, ZnTFZCDs modulate copper homeostasis by upregulating the copper exporter ATP7B, downregulating the importer SLC31A1, and reducing FDX1/LIAS expression, thereby inhibiting cuproptosis-related signaling. In vivo, ZnTFZCDs administration ameliorates testicular histopathology, improves sperm count and morphology, reactivates the Nrf2/HO-1 antioxidant axis, and suppresses cuproptosis-associated alterations in testicular tissues, confirming the translational potential of the in vitro findings. Collectively, ZnTFZCDs represent a bio-inspired nanoplatform that attenuates PS-MPs-induced male reproductive injury through coordinated modulation of redox and copper homeostasis, offering a new therapeutic strategy for environmental pollutant-related reproductive disorders by targeting the previously overlooked cuproptosis pathway.