The inevitable exposure of humans to micro/nanoplastics has become a pressing global environmental issue, with growing concerns regarding their impact on health. While the direct effects of micro/nanoplastics on human health remain largely unknown, increasing attention is being given to their potential role as carriers of environmental pollutants and organic substances. This study investigates the direct toxicity of 500 nm polystyrene nanoplastics (NPs) on human hepatocytes (HepG2) in vitro, both alone and in combination with cadmium (Cd), a hazardous heavy metal and a prevalent environmental pollutant. One-hour exposure to 100 µg/mL of NPs causes a significant increase in ROS production (+25% compared to control) but cell viability remains unaffected even at concentrations much higher than environmental levels. Interestingly, NPs significantly reduce Cd cytotoxicity at LC concentrations (cell viability compared to control: 55.4% for 50 µM Cd, 66.9% for 50 µM Cd + 10 µg/mL NPs, 68.4% for 50 µM Cd + 100 µg/mL NPs). Additionally, NPs do not alter the cellular lipid content after short-term exposure (24 h). However, when Cd and fatty acids are added to the medium, NPs appear to sequester fatty acids, reducing their availability and impairing their uptake by cells in a dose-dependent manner. We confirmed by Dynamic Light Scattering and Scanning Electron Microscopy the interaction between NPs, Cd and free fatty acids. Although polystyrene NPs exhibited minimal cytotoxicity in our experimental model, collectively our findings suggest that predicting the effects of cell exposure to NPs is extremely challenging, due to the potential interaction between NPs, environmental pollutants and specific components of the biological matrix.