Functional consequences for primary human alveolar macrophages following treatment with long, but not short, multiwalled carbon nanotubes

Abstract

Purpose

Multiwalled carbon nanotubes (MWCNTs) are a potential human health hazard, primarily via inhalation. In the lung, alveolar macrophages (AMs) provide the first line of immune cellular defense against inhaled materials. We hypothesized that, 1 and 5 days after treating AMs with short (0.6 μm in length; MWCNT-0.6 μm) and long (20 μm in length; MWCNT-20 μm) MWCNTs for 24 hours, AMs would exhibit increased markers of adverse bioreactivity (cytokine release and reactive oxygen species generation) while also having a modified functional ability (phagocytosis and migration).

Methods

Primary human AMs were treated with short and long MWCNTs for 24 hours, 1 and 5 days after which toxicity end points, including cell death, reactive oxygen species generation, and inflammatory mediator release, were measured. AM functional end points involving phagocytic ability and migratory capacity were also measured.

Results

AM viability was significantly decreased at 1 and 5 days after treatment with MWCNT-20 μm, while superoxide levels and inflammatory mediator release were significantly increased. At the same time, there was reduced phagocytosis and migratory capacity alongside increased expression of MARCO; this coincided with frustrated phagocytosis observed by scanning electron microscopy. In contrast, the adverse bioreactivity of the shorter MWCNT-0.6 μm with AMs (and any resulting reduction in AM functional ability) was substantially less marked or absent altogether.

Conclusion

This study shows that after 24-hour treatment with long, but not short, MWCNTs, AM function is severely affected up to 5 days after the initial exposure. This has potentially significant pathophysiological consequences for individuals who may be intentionally (via therapeutic applications) or unintentionally exposed to these nanomaterials.

Keywords:

• nanotechnology
• MWCNTs
• alveolar macrophages
• cytokines
• phagocytosis
• bioreactivity

Supplementary material

Figure S1 AMs treated with MWCNT-20 μm; viability, phagocytosis, and AM migration expressed as a percent of respective NT controls (100%).

Notes: t-Test comparison shows that the percent viability of treated AMs (yellow bars) was proportionally significantly higher than the percent phagocytic activity (blue bars) and percent migratory activity (red bars), indicating that the ability to phagocytose bacteria and to migrate were markedly inhibited by the long MWCNTs. Significant differences between percent viability and phagocytic activity or migratory activity are indicated, where *P<0.05.

Abbreviations: NT, nontreated; MWCNTs, multiwalled carbon nanotubes; AMs, alveolar macrophages.

Acknowledgments

Financial support for SS from Unilever, UK, is gratefully acknowledged. PR is supported by the Leverhulme Trust, UK. We thank Dr Katerina Kouravelou at Nanothinx, Greece, for her generous advice throughout the study. We also thank Dr Andrew V Rogers (Royal Brompton Hospital, London) for assistance and expert advice on cell imaging using scanning electron microscopy. This research was supported by the Medical Research Council and Public Health England, Center for Environment and Health. The project was also supported by the NIHR Respiratory Disease Biomedical Research Unit at the Royal Brompton and Harefield NHS Foundation Trust and Imperial College London. The views expressed in this publication are those of the authors(s) and not necessarily those of the NHS, the National Institute for Health Research, or the Department of Health.

Disclosure

The authors report no conflicts of interest in this work.