Evaluation of cytotoxic, genotoxic and inflammatory responses of nanoparticles from photocopiers in three human cell lines

Part Fibre Toxicol. 2013 Aug 22;10:42. doi: 10.1186/1743-8977-10-42.

Abstract

Background: Photocopiers emit nanoparticles with complex chemical composition. Short-term exposures to modest nanoparticle concentrations triggered upper airway inflammation and oxidative stress in healthy human volunteers in a recent study. To further understand the toxicological properties of copier-emitted nanoparticles, we studied in-vitro their ability to induce cytotoxicity, pro-inflammatory cytokine release, DNA damage, and apoptosis in relevant human cell lines.

Methods: Three cell types were used: THP-1, primary human nasal- and small airway epithelial cells. Following collection in a large volume photocopy center, nanoparticles were extracted, dispersed and characterized in the cell culture medium. Cells were doped at 30, 100 and 300 μg/mL administered doses for up to 24 hrs. Estimated dose delivered to cells, was ~10% and 22% of the administered dose at 6 and 24 hrs, respectively. Gene expression analysis of key biomarkers was performed using real time quantitative PCR (RT-qPCR) in THP-1 cells at 5 μg nanoparticles/mL for 6-hr exposure for confirmation purposes.

Results: Multiple cytokines, GM-CSF, IL-1β, IL-6, IL-8, IFNγ, MCP-1, TNF-α and VEGF, were significantly elevated in THP-1 cells in a dose-dependent manner. Gene expression analysis confirmed up-regulation of the TNF-α gene in THP-1 cells, consistent with cytokine findings. In both primary epithelial cells, cytokines IL-8, VEGF, EGF, IL-1α, TNF-α, IL-6 and GM-CSF were significantly elevated. Apoptosis was induced in all cell lines in a dose-dependent manner, consistent with the significant up-regulation of key apoptosis-regulating genes P53 and Casp8 in THP-1 cells. No significant DNA damage was found at any concentration with the comet assay. Up-regulation of key DNA damage and repair genes, Ku70 and Rad51, were also observed in THP-1 cells, albeit not statistically significant. Significant up-regulation of the key gene HO1 for oxidative stress, implicates oxidative stress induced by nanoparticles.

Conclusions: Copier-emitted nanoparticles induced the release of pro-inflammatory cytokines, apoptosis and modest cytotoxicity but no DNA damage in all three-human cell lines. Taken together with gene expression data in THP-1 cells, we conclude that these nanoparticles are directly responsible for inflammation observed in human volunteers. Further toxicological evaluations of these nanoparticles, including across different toner formulations, are warranted.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Air Pollutants / chemistry
  • Air Pollutants / toxicity*
  • Apoptosis / drug effects*
  • Cell Culture Techniques
  • Cell Line
  • Cell Survival / drug effects
  • Comet Assay
  • Copying Processes*
  • Cytokines / immunology*
  • DNA Damage*
  • Epithelial Cells / drug effects
  • Epithelial Cells / immunology
  • Epithelial Cells / pathology
  • Flow Cytometry
  • Humans
  • Macrophages / drug effects
  • Macrophages / immunology
  • Macrophages / pathology
  • Microscopy, Electron, Transmission
  • Nanoparticles / chemistry
  • Nanoparticles / toxicity*
  • Oxidative Stress / drug effects
  • Particle Size
  • Respiratory Mucosa / drug effects
  • Respiratory Mucosa / immunology
  • Respiratory Mucosa / pathology
  • Solubility
  • Surface Properties

Substances

  • Air Pollutants
  • Cytokines