Particle-size-associated differences in follicular accessibility, granulosa cell oxidative injury, and oocyte...

Polystyrene particles are emerging environmental contaminants with potential reproductive hazards, but whether particle size influences access to the follicular microenvironment and subsequent oocyte dysfunction remains unclear. Here, fluorescent and non-fluorescent PS particles with nominal diameters of 0.1, 0.5, and 1.0 μm were compared using mouse cumulus-oocyte complexes, ovarian tissue, and KGN granulosa-like cells under equal-mass and theoretically matched particle-number exposure conditions. Smaller PS particles showed greater follicular-associated fluorescence, with particle-associated signals predominantly detected in cumulus/granulosa-cell regions rather than the oocyte compartment. This size-dependent localization pattern was independently reproduced using same-green-fluorophore particles. Trypan blue quenching, phalloidin-guided confocal imaging, and dye-leaching assays further supported particle-associated fluorescence signals that were resistant to extracellular quenching and consistent with intracellular retention. Functionally, PS particle exposure impaired COC expansion, altered mitochondrial spatial organization in metaphase II oocytes, and reduced subsequent embryonic developmental competence. Time-course analysis revealed an early increase in ROS in cumulus cells followed by a later increase in oocytes within intact COCs, indicating a temporally ordered oxidative response during particle exposure. hUCMSC-derived extracellular vesicles partially attenuated PS particle-associated oxidative and apoptosis-related alterations in KGN granulosa-like cells. Because extracellular vesicles exert broad biological effects, antioxidant validation was further performed in intact COCs. N-acetylcysteine reduced ROS-sensitive fluorescence and partially restored first-polar-body extrusion and mitochondrial membrane potential in mature oocytes. Collectively, these findings support a particle-size-associated follicular accessibility-toxicity framework, in which smaller PS particles exhibit greater follicular-associated signals and are accompanied by enhanced oxidative disturbances in follicular somatic cells and impaired oocyte developmental competence. This study highlights oxidative stress as a functionally relevant process underlying PS particle-associated reproductive dysfunction.