In vitro toxicity evaluation of single walled carbon nanotubes on human A549 lung cells

• Published in: Toxicology in vitro 2007-04, Vol.21 (3), p.438-448
• Main Authors: Davoren, Maria, Herzog, Eva, Casey, Alan, Cottineau, Benjamin, Chambers, Gordon, Byrne, Hugh J., Lyng, Fiona M.
• Format: Article
• Language: English
• Subjects: Adenylate Kinase - metabolism; Cell Line; Cell Nucleus - drug effects; Cell Nucleus - ultrastructure; Cell Survival - drug effects; Culture Media, Conditioned - chemistry; Cytoplasm - drug effects; Cytoplasm - ultrastructure; Cytotoxicity; Dose-Response Relationship, Drug; Epithelial Cells - drug effects; Epithelial Cells - metabolism; Epithelial Cells - pathology; Foetal bovine serum; Formazans - metabolism; Humans; Indicators and Reagents - metabolism; Interleukin-8 - metabolism; Lamellar bodies; Lung - drug effects; Lung - metabolism; Lung - pathology; Microscopy, Electron, Transmission; Nanotechnology; Nanotubes, Carbon - chemistry; Nanotubes, Carbon - toxicity; Nanotubes, Carbon - ultrastructure; Necrosis - chemically induced; Neutral Red - metabolism; Oxazines - metabolism; Quartz - metabolism; Quartz - toxicity; Rosaniline Dyes - metabolism; Single walled carbon nanotubes; TEM; Tetrazolium Salts - metabolism; Xanthenes - metabolism
• ISSN: 0887-2333; 1879-3177
• DOI: 10.1016/j.tiv.2006.10.007

This paper describes the in vitro cytotoxicity assessment of single walled carbon nanotubes (SWCNT) on A549 cells, a human lung cell line. Cellular viability was determined using the alamar blue (AB), neutral red (NR) and MTT assays, which evaluated metabolic, lysosomal and mitochondrial activity respectively. In addition, the total protein content of the cells was measured using the coomassie brilliant (CB) blue assay. Supernatants were also assayed for Adenylate Kinase (AK) release and Interleukin 8 (IL-8) which indicated a loss of cell membrane integrity and an inflammation response respectively. To investigate the interactions between serum components in the test medium and the test materials, exposures were conducted both in serum containing (5%) and serum-free medium. Results from the cytotoxicity tests (AB, CB, MTT) revealed the SWCNT to have very low acute toxicity to the A549 cells as all but one of the reported 24 h EC 50 values exceeded the top concentration tested (800 μg/ml). The SWCNT were found to interfere with a number of the dyes used in the cytotoxicity assessment and we are currently conducting a comprehensive spectroscopic study to further investigate these interactions. Of the multiple cytotoxicity assays used, the AB assay was found to be the most sensitive and reproducible. Transmission electron microscopy (TEM) studies confirmed that there was no intracellular localization of SWCNT in A549 cells following 24 h exposure; however, increased numbers of surfactant storing lamellar bodies were observed in exposed cells.