Cationic Polystyrene Nanosphere Toxicity Depends on Cell-Specific Endocytic and Mitochondrial Injury Pathways

The exponential increase in the number of new nanomaterials that are being produced increases the likelihood of adverse biological effects in humans and the environment. In this study we compared the effects of cationic nanoparticles in five different cell lines that represent portal-of-entry or systemic cellular targets for engineered nanoparticles. Although 60 nm NH₂-labeled polystyrene (PS) nanospheres were highly toxic in macrophage (RAW 264.7) and epithelial (BEAS-2B) cells, human microvascular endothelial (HMEC), hepatoma (HEPA-1), and pheochromocytoma (PC-12) cells were relatively resistant to particle injury. While the death pathway in RAW 264.7 cells involves caspase activation, the cytotoxic response in BEAS-2B cells is more necrotic in nature. Using fluorescent-labeled NH₂-PS, we followed the routes of particle uptake. Confocal microscopy showed that the cationic particles entered a LAMP-1 positive lysosomal compartment in RAW 264.7 cells from where the particles could escape by lysosomal rupture. A proton pump inhibitor interfered in this pathway. Subsequent deposition of the particles in the cytosol induced an increase in mitochondrial Ca²⁺ uptake and cell death that could be suppressed by cyclosporin A (CsA). In contrast, NH₂-PS toxicity in BEAS-2B cells did not involve the LAMP-1 endosomal compartment, stimulation of proton pump activity, or an increase in mitochondrial Ca²⁺. Particles were taken up by caveolae, and their toxicity could be disrupted by cholesterol extraction from the surface membrane. Although the particles induced mitochondrial damage and ATP depletion, CsA did not affect cytotoxicity. Cationic particles were taken up into HEPA-1, HMEC, and PC-12 cells, but this did not lead to lysosomal permeabilization, increased Ca²⁺ flux, or mitochondrial damage. Taken together, the results of this study demonstrate the importance of cell-specific uptake mechanisms and pathways that could lead to sensitivity or resistance to cationic particle toxicity.