title: Differential bioreactivity of neutral, cationic and anionic polystyrene nanoparticles with cells from the human alveolar compartment: robust response of alveolar type 1 epithelial cells.
abstract: BACKGROUND
  Engineered nanoparticles (NP) are being developed for inhaled drug delivery. This route is non-invasive and the major target; alveolar epithelium provides a large surface area for drug administration and absorption, without first pass metabolism. Understanding the interaction between NPs and target cells is crucial for safe and effective NP-based drug delivery. We explored the differential effect of neutral, cationic and anionic polystyrene latex NPs on the target cells of the human alveolus, using primary human alveolar macrophages (MAC) and primary human alveolar type 2 (AT2) epithelial cells and a unique human alveolar epithelial type I-like cell (TT1). We hypothesized that the bioreactivity of the NPs would relate to their surface chemistry, charge and size as well as the functional role of their interacting cells in vivo.
  
  
  METHODS
  Amine- (ANP) and carboxyl- surface modified (CNP) and unmodified (UNP) polystyrene NPs, 50 and 100 nm in diameter, were studied. Cells were exposed to 1-100 μg/ml (1.25-125 μg/cm(2); 0 μg/ml control) NP for 4 and 24 h at 37 °C with or without the antioxidant, N-acetyl cysteine (NAC). Cells were assessed for cell viability, reactive oxygen species (ROS), oxidised glutathione (GSSG/GSH ratio), mitochondrial integrity, cell morphology and particle uptake (using electron microscopy and laser scanning confocal microscopy).
  
  
  RESULTS
  ANP-induced cell death occurred in all cell types, inducing increased oxidative stress, mitochondrial disruption and release of cytochrome C, indicating apoptotic cell death. UNP and CNP exhibited little cytotoxicity or mitochondrial damage, although they induced ROS in AT2 and MACs. Addition of NAC reduced epithelial cell ROS, but not MAC ROS, for up to 4 h. TT1 and MAC cells internalised all NP formats, whereas only a small fraction of AT2 cells internalized ANP (not UNP or CNP). TT1 cells were the most resistant to the effects of UNP and CNP.
  
  
  CONCLUSIONS
  ANP induced marked oxidative damage and cell death via apoptosis in all cell types, while UNP and CNP exhibited low cytotoxicity via oxidative stress. MAC and TT1 cell models show strong particle-internalization compared to the AT2 cell model, reflecting their cell function in vivo. The 50 nm NPs induced a higher bioreactivity in epithelial cells, whereas the 100 nm NPs show a stronger effect on phagocytic cells.
authors.affiliation: Lung Cell Biology, Section of Airways Disease, National Heart & Lung Institute, Imperial College London, Dovehouse Street, London, SW3 6LY, UK. p.ruenraroengsak@imperial.ac.uk.
authors.forename: Pakatip
authors.initials: P
authors.lastname: Ruenraroengsak
authors.affiliation: Lung Cell Biology, Section of Airways Disease, National Heart & Lung Institute, Imperial College London, Dovehouse Street, London, SW3 6LY, UK. t.tetley@imperial.ac.uk.
authors.forename: Teresa D
authors.initials: TD
authors.lastname: Tetley
mesh_terms: D056809:Alveolar Epithelial Cells / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D000838:Anions; D000975:Antioxidants / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D001692:Biological Transport; D002412:Cations; D002460:Cell Line; D048430:Cell Shape; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004337:Drug Carriers; D019803:Glutathione Disulfide / Q000378:metabolism; D006801:Humans; D016676:Macrophages, Alveolar / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D053758:Nanoparticles; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000187:drug effects; D011137:Polystyrenes / Q000737:chemistry / Q000378:metabolism / Q000633:toxicity*; D061251:Primary Cell Culture; D011650:Pulmonary Alveoli / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D017382:Reactive Oxygen Species / Q000378:metabolism; D013499:Surface Properties; D013997:Time Factors
chemical_list: D000838:Anions; D000975:Antioxidants; D002412:Cations; D004337:Drug Carriers; D011137:Polystyrenes; D017382:Reactive Oxygen Species; D019803:Glutathione Disulfide