Defining the mechanism of action of multi-walled carbon nanotubes on immune cells in vitro

Abstract

Engineered nanomaterials (ENM) are a rapidly expanding class of chemicals with applications spanning from cosmetics to food, and from industrial applications to pharmaceuticals. Some ENM are already known to be harmful to humans. Majority of the adverse effects associated with ENM exposure are mediated by the cells of the innate immunity. The respiratory system offers a vulnerable route of exposure for ENM as they are small enough to pass through many of the physical barriers of the body and even translocate to sites beyond the point of entry. In the lungs, ENM tend to induce an inflammatory reaction that can turn chronic and lead to other conditions such as fibrosis. As the production of ENM is constantly increasing and novel materials are reaching the market in an increasing pace, their toxic potential must be well characterised. While the traditional hazard assessment is driven by apical endpoints and heavily focused on expensive and time-consuming animal experiments, toxicogenomics approaches turn the focus onto the mechanisms of toxicity. In this thesis, the transcriptional mechanism of action (tMOA) of multi-walled carbon nanotubes (MWCNT) was characterised in THP-1 monocytes and PMA-differentiated macrophages using DNA microarrays. The exposures were carried out for 24, 48 and 72 hours with three doses (5, 10 and 20 μg/ml), for determining the dose-response relationship with benchmark dose modelling. The results suggest distinct transcriptomic responses in monocytes and macrophages, with macrophages being significantly more sensitive. In macrophages, MWCNT induced the expression of a wide range of immune related genes and pathways, including proinflammatory signalling pathways such as TNF and IL-17 signalling. The proinflammatory response was more pronounced after 24 hours of exposure, while transcriptomic alterations probably mirroring indirect changes in the microenvironment increased between 48 hours and 72 hours, as suggested by the decreased proportion of dose-dependent genes at 72 hours. From this analysis, the genes associated to the NF-kappa B signalling pathway emerged as directly affected by the exposure, suggesting their role as key regulators of the macrophage response to MWCNT. These results contribute to a better understanding of the molecular alterations caused by MWCNT exposure on innate immune cells.