Durability of carbon nanotubes and their potential to cause inflammation

It has been suggested that carbon nanotubes (CNTs) might conform to the fibre pathogenicity paradigm that explains the pathogenicity of asbestos and other fibres on a continuum based on length, aspect ratio and biopersistence. Multiwalled carbon nanotubes have shown that they may satisfy the first two aspects of the fibre paradigm but biopersistence has not been investigated to a substantial degree for carbon nanotubes in general. Whilst biopersistence is complex and requires animal studies, durability, the chemical mimicking of the process of fibre dissolution using in vitro treatment, is a rapid alternative that is easier to determine and is closely related to biopersistence but does not require large numbers of animals. The University of Edinburgh, the Institute of Occupational Medicine (Edinburgh), and the Commonwealth Scientific & Industrial Research Organisation (Australia) have collaborated to determine the durability of carbon nanotubes in simulated biological fluid and subsequent fibre pathogenicity, compared with well-characterised fibre controls. This collaborative project was financially supported under Safe Work Australia’s Nanotechnology Work Health and Safety Program.The study had two main components. First, known masses of four types of carbon nanotubes (one type of single-walled CNT (CNTSW) and three types of multi-walled CNTs (CNTSPIN, CNTTANG2 and CNTLONG1)), two types of asbestos fibres (one type of chrysotile asbestos (LFC) and one type of amosite asbestos (LFA)), and one type of glass wool fibre (X607) were incubated in simulated biological fluid (Gambles solution) for up to 24 weeks, with samples removed from incubation, filtered, dried, and weighed at defined time-points (0wk, 3wk, 6wk, 10wk and 24wk). Two of the four types of carbon nanotubes, CNTSPIN and CNTSW, showed no loss of mass, and no change in morphology or average fibre length when viewed by electron microscopy. A third, CNTTANG2, showed a possible loss of mass (~25%) after 24wk incubation, but no change in morphology. However, the fourth type of carbon nanotube, CNTLONG1, was recovered at only 70% of original mass at all time-points from 3wk onwards. Electron microscopy confirmed that the average fibre length had decreased slightly and the proportion of long fibres had also decreased, indicating that this type of nanotube had undergone some kind of modification during the incubation in Gambles solution that had resulted in fibre dissolution and/or breakage.The second part of the study was designed to investigate the impact of incubation in Gambles solution on carbon nanotube pathogenicity in mice in comparison to the asbestos and glass wool fibres. Two types of CNT samples included in the durability assessment (CNTSPIN and CNTTANG2) were excluded from in vivo assessment leaving one type of multi-walled CNT (CNTLONG1) and one type of single-walled CNT (CNTSW). The CNTLONG1 sample had previously been shown to induce an asbestos-like response when injected into the peritoneal cavity of mice, a model for the biological response of exposure at the mesothelium following inhalation of high aspect ratio fibres. The incubation in Gambles solution and subsequent loss of mass and fibre shortening for CNTLONG1 was associated with decreased pathogenicity compared to the strong inflammatory and granuloma response induced in mice by injection of CNTLONG1 fibres that had been incubated for 0wk. A similar loss of pathogenicity with incubation was also observed for the less durable of the asbestos fibres included here (LFC), whereas the second, more durable asbestos (LFA) showed no significant loss of mass, no change in morphology and no loss of pathogenicity. The CNTSW sample was shown by electron microscopy to form very tightly agglomerated particle-like bundles and did not elicit an inflammatory response in mice regardless of incubation. These results, therefore, support the view that carbon nanotubes can be durable but may also be subject to bio-modification in a sample-specific manner. They also suggest that if they are of sufficient length and aspect ratio, pristine carbon nanotubes can induce asbestos-like responses in mice, but that this may be mitigated if the nanotubes are of a less durable nature.