Respiratory toxicity and immunotoxicity evaluations of microparticle and nanoparticle C60 fullerene aggregates in mice and rats following nose-only inhalation for 13 weeks

Abstract

C60 fullerene (C60), or buckminsterfullerene, is a spherical arrangement of 60 carbon atoms, having a diameter of approximately 1 nm, and is produced naturally as a by-product of combustion. Due to its small size, C60 has attracted much attention for use in a variety of applications; however, insufficient information is available regarding its toxicological effects. The effects on respiratory toxicity and immunotoxicity of C60 aggregates (50 nm [nano-C60] and 1 μm [micro-C60] diameter) were examined in B6C3F1/N mice and Wistar Han rats after nose-only inhalation for 13 weeks. Exposure concentrations were selected to allow for data evaluations using both mass-based and particle surface area-based exposure metrics. Nano-C60 exposure levels selected were 0.5 and 2 mg/m³ (0.033 and 0.112 m²/m³), while micro-C60 exposures were 2, 15 and 30 mg/m³ (0.011, 0.084 and 0.167 m²/m³). There were no systemic effects on innate, cell-mediated, or humoral immune function. Pulmonary inflammatory responses (histiocytic infiltration, macrophage pigmentation, chronic inflammation) were concentration-dependent and corresponded to increases in monocyte chemoattractant protein (MCP)-1 (rats) and macrophage inflammatory protein (MIP)-1α (mice) in bronchoalveolar lavage (BAL) fluid. Lung overload may have contributed to the pulmonary inflammatory responses observed following nano-C60 exposure at 2 mg/m³ and micro-C60 exposure at 30 mg/m³. Phenotype shifts in cells recovered from the BAL were also observed in all C60-exposed rats, regardless of the level of exposure. Overall, more severe pulmonary effects were observed for nano-C60 than for micro-C60 for mass-based exposure comparisons. However, for surface-area-based exposures, more severe pulmonary effects were observed for micro-C60 than for nano-C60, highlighting the importance of dosimetry when evaluating toxicity between nano- and microparticles.

Keywords: Buckminsterfullerene; immunotoxicity; inhalation; nanoparticles; pulmonary inflammation.

Figure 1. MCP-1 and IL-1 levels in the BAL fluid of female Wistar Han rats following C60 fullerene inhalation for 13 weeks

Combined control group results are shown. Values of control animals assigned to the micro-C60 and nano-C60 studies were not significantly different. Individual study control values (pg/ml) for MCP-1 were: 65 ± 22 (micro-C60 study) and 93 ± 24 (nano-C60 study). Individual study control values (pg/ml) for IL-1 were: 4 ± 4 (micro-C60 study) and 18 ± 9 (nano-C60 study). Values represent the mean ± SE (N = 8 animals per test group; N = 16 for combined control group). Significant differences from individual study control group (not shown): *p < 0.05 and **p < 0.01. Significant differences from combined control group (shown): ^†p < 0.05 and ^††p < 0.01. Pairwise significant differences between C60 exposure groups: ^##p < 0.01.

Figure 2. MIP-1α levels in the BAL fluid of female B6C3F1/N mice following C60 fullerene inhalation for 13 weeks

Combined control group results are shown. Values of control animals assigned to the micro-C60 and nano-C60 studies were not significantly different. Individual study control values (pg/ml) were: 0.74 ± 0.7 (micro-C60 study) and 6.8 ± 3.3 (nano-C60 study). Values represent the mean ± SE (N = 8 animals per test group; N = 16 for combined control group). Significant differences from individual study control group (not shown): **p < 0.01. Significant differences from combined control group (shown): ^†p < 0.05 and ^††p < 0.01. Pairwise significant differences between C60 exposure groups: ^#p < 0.05.