Elsevier

Carbon

Volume 44, Issue 6, May 2006, Pages 1100-1105
Carbon

Effects of fullerenes and single-wall carbon nanotubes on murine and human macrophages

https://doi.org/10.1016/j.carbon.2005.11.009Get rights and content

Abstract

The discovery in 1985 of C-fullerenes, a novel carbon allotrope with a polygonal structure made up solely by 60 carbon atoms, and in 1991 of C-nanotubes, thin carbon filaments (1–3 μm in length and 0.001 μm in diameter) with extraordinary mechanical properties, opened a wide field of activity in carbon research. While toxicity and biocompatibility of C-fullerenes have been widely investigated, literature data concerning the biological properties and biotoxicity of C-nanotubes are poor and contradictory. Here we test the ability of highly purified C-Single-Walled-Nanotubes (SWNTs) and C-fullerenes to elicit an inflammatory response by murine and human macrophage cells in vitro. In order to determine the potential of these C-derivatives as biological inducers of inflammatory reactions we evaluate the ability of C-single-walled nanotubes and C-fullerenes to induce the release of NO by murine macrophages cells, to stimulate the phagocytic activity of human macrophage cells and to be cytotoxic against these cells. We show that SWNTs-C-nanotubes, when highly purified, as well as C-fullerenes, do not stimulate the release of NO by murine macrophage cells in culture, their uptake by human macrophage cells is very low, and they possess a very low toxicity against human macrophage cells.

Introduction

The discovery in 1985 of C-fullerenes, a novel carbon allotrope with a polygonal structure made up solely by 60 carbon atoms, and in 1991 of C-nanotubes, thin carbon filaments (1–3 μm in length and 1–3 nm in diameter) with extraordinary mechanical properties, opened a wide field of activity in carbon research. During the last few years practical application of fullerenes as biological as well as pharmacological agents was investigated. Various fullerene-based compounds were tested for biological activity, including antiviral, antioxidant and chemiotactic activities [1]. C-nanotubes , because of their aspect, their high modulus and strength have been considered as a promising filler in polymer as well as ceramics and metal-matrix composites as a substitute for carbon fibers [2], [3], [4]. Furthermore, the selective adhesion and enhanced function of osteoblasts as well as the increase in nerve cell functions and the decreased astrocyte adhesion on nanometer diameter carbon fibers and on composites containing carbon nanofibers has been recently demonstrated [5], [6], [7]. Thus, because of these properties, carbon nanostructures are more and more considered a promising material for orthopedic and dental artificial devices and neural probes.

In the last few years there has been an ever growing interest in the development, production and use of carbon nanoparticles in a wide range of industrial and scientific fields due to the unique surface properties of these nanostructures. As a result, human exposure to them will occur more and more frequently. Because of their extremely light weight carbon nanostructures could become airborne and be easily inhaled by human lungs, mostly in laboratory environments. Recently it has been demonstrated that, after inhalation exposure to ultrafine carbon nanoparticles, a translocation of the inhaled particles to the central nervous system could occur via the blood brain barrier and via the olfactory nerve from deposits on the nasal olfactory mucosa [8]. But, at present, while toxicity and biocompatibility of C-fullerenes have been widely investigated [9], [10], [11], [12], [13], [14], few literature data exist concerning the biological properties and biotoxicity of C-nanotubes [15], [16], [17], [18].

We aimed to assess the ability of pure Single-Walled-Carbon-Nanotubes (SWNTs) and C-fullerenes to elicite an inflammatory response by murine and human macrophage cells in vitro and to be cytotoxic against these cells. Moreover, in order to determine if the behaviour of these C-nanoparticles towards the biological environment could be due, at least in part, to graphite, one of the main component of C-nanomaterial preparations, we compared their effects with those induced on the same cells by graphite particles. Macrophages, also known as antigen presenting cells, are the primary and most important cells of the immune system that react against all foreign body materials. When these cells get in contact with material particles or pathogens, they become activated and, after having phagocytosed particles, secrete a lot of chemical mediators of inflammation, very aggressive against either the host tissues and the foreign molecules or particles. Nitric oxide (NO) is one of these molecules and it is regarded as a key marker of activation–inflammation of these cells [19], [20]. We evaluated the release of NO by murine macrophages stimulated with C-fullerenes, pure C-nanotubes and graphite particles. In addition we assessed the phagocytic activity and cell viability of human macrophages challenged with the same C-nanoparticles.

Section snippets

Carbon nanoparticles

The following C-nanoparticle preparations have been studied:

  • Highly pure C-SWNTs produced by the Chemical Vapour Deposition technique (CVD) (Colomer JF, University of Namur, Belgique). They were purified by a thermal and acidic treatment. The analysis by the X-ray Diffraction (XRD) technique (resolution=0.08 A−1) showed the presence of SWNTs (zone 0.23 A−1), the absence of nickel and cobalt (zone 3.1 A−1) and a very low amount of amorphous carbon (zone 1.7 A−1) (Fig. 1). Furthermore they were

Results

We observed that the release of nitric oxide by murine macrophages was the same both when the cells were stimulated by LPS and when the cells, previously stimulated by the positive control LPS, were challenged with fullerenes and pure SWNTs (Fig. 3a and b). So the macrophage stimulation induced by these C-nanoparticles can be considered not specific. On the contrary, when different concentrations of Graphite particles (15, 30 and 60 μg/ml) were added to the cell cultures previously stimulated

Discussion

Our data, showing that C60-fullerenes, very highly purified carbon nanostructures, do not stimulate the release of NO by murine macrophage cells in culture, their uptake by human macrophage cells is very low, and that they do not induce apoptosis and cell death compared to graphite particles, suggest that these C-nanostructures are not able to elicite a reactive-inflammatory response by mammalian cells in vitro and are not cytotoxic. These findings are in agreement with previously reported data

Conclusions

Based on these preliminary results, we think that a more detailed physico-chemical characterization of these C-nanomaterials must be performed, before testing their biocompatibility and before any conclusion on their toxicity could be drawn.

We believe that the toxicity of C-nanomaterials need to be understood in the framework of the material characterization. Determining toxicity of C-nanoparticles is quite complicated. The surface coating, the presence of metal catalysts and/or graphite, the

Acknowledgements

J.F. Colomer, Namur, is kindly acknowledged for having supplied highly purified SWNTs. The financial support from the GDR 1752 “Nanotubes” (CNRS) is kindly acknowledged.

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