Toxicity assessment and bioaccumulation in zebrafish embryos exposed to carbon nanotubes suspended in Pluronic® F-108

Carbon nanotubes (CNTs) are often suspended in Pluronic® surfactants by sonication, which may confound toxicity studies because sonication of surfactants can create degradation products that are toxic to mammalian cells. Here, we present a toxicity assessment of Pluronic® F-108 with and without suspended CNTs using embryonic zebrafish as an in vivo model. Pluronic® sonolytic degradation products were toxic to zebrafish embryos just as they were to mammalian cells. When the toxic Pluronic® fragments were removed, there was little effect of pristine multi-walled CNTs (pMWNTs), carboxylated MWNTs (cMWNTs) or pristine single-walled carbon nanotubes (pSWNTs) on embryo viability and development, even at high concentrations. A gel electrophoretic method coupled with Raman imaging was developed to measure the bioaccumulation of CNTs by zebrafish embryos, and dose-dependent uptake of CNTs was observed. These data indicate that embryos accumulate pMWNTs, cMWNTs and pSWNTs yet there is very little embryo toxicity.     

Pulmonary toxicity of single-wall carbon nanotubes in mice 7 and 90 days after intratracheal instillation.

Nanomaterials are part of an industrial revolution to develop lightweight but strong materials for a variety of purposes. Single-wall carbon nanotubes are an important member of this class of materials. They structurally resemble rolled-up graphite sheets, usually with one end capped; individually they are about 1 nm in diameter and several microns long, but they often pack tightly together to form rods or ropes of microscopic sizes. Carbon nanotubes possess unique electrical, mechanical, and thermal properties and have many potential applications in the electronics, computer, and aerospace industries. Unprocessed nanotubes are very light and could become airborne and potentially reach the lungs. Because the toxicity of nanotubes in the lung is not known, their pulmonary toxicity was investigated. The three products studied were made by different methods and contained different types and amounts of residual catalytic metals. Mice were intratracheally instilled with 0, 0.1, or 0.5 mg of carbon nanotubes, a carbon black negative control, or a quartz positive control and euthanized 7 d or 90 d after the single treatment for histopathological study of the lungs. All nanotube products induced dose-dependent epithelioid granulomas and, in some cases, interstitial inflammation in the animals of the 7-d groups. These lesions persisted and were more pronounced in the 90-d groups; the lungs of some animals also revealed peribronchial inflammation and necrosis that had extended into the alveolar septa. The lungs of mice treated with carbon black were normal, whereas those treated with high-dose quartz revealed mild to moderate inflammation. These results show that, for the test conditions described here and on an equal-weight basis, if carbon nanotubes reach the lungs, they are much more toxic than carbon black and can be more toxic than quartz, which is considered a serious occupational health hazard in chronic inhalation exposures.     

Cytotoxicity of single-wall carbon nanotubes on human fibroblasts.

We present a toxicological assessment of five carbon nanomaterials on human fibroblast cells in vitro. We correlate the physico-chemical characteristics of these nanomaterials to their toxic effect per se, i.e. excluding catalytic transition metals. Cell survival and attachment assays were evaluated with different concentrations of refined: (i) single-wall carbon nanotubes (SWCNTs), (ii) active carbon, (iii) carbon black, (iv) multi-wall carbon nanotubes, and (v) carbon graphite. The refined nanomaterial that introduced the strongest toxic effect was subsequently compared to its unrefined version. We therefore covered a wide range of variables, such as: physical dimensions, surface areas, dosages, aspect ratios and surface chemistry. Our results are twofold. Firstly, we found that surface area is the variable that best predicts the potential toxicity of these refined carbon nanomaterials, in which SWCNTs induced the strongest cellular apoptosis/necrosis. Secondly, we found that refined SWCNTs are more toxic than its unrefined counterpart. For comparable small surface areas, dispersed carbon nanomaterials due to a change in surface chemistry, are seen to pose morphological changes and cell detachment, and thereupon apoptosis/necrosis. Finally, we propose a mechanism of action that elucidates the higher toxicity of dispersed, hydrophobic nanomaterials of small surface area.     

Adipocytes differentiation in the presence of Pluronic F127–coated carbon nanotubes

Carbon nanotubes are considered important nanodevices in biomedicine, and several applications in drug and gene delivery in different organs and tissues are presently under investigation. Among them very little attention has been dedicated to white adipose tissue (WAT), despite its wide distribution and endocrine role, which could potentially be used to release therapeutic agents. An important premise to use nanotubes in WAT is to determine their potential for toxicity on adipocytes. Here we show that Pluronic F127 (PF127)–coated multiwalled carbon nanotubes (MWCNTs) can be tolerated by NIH-3T3-L1 pre-adipocytes without affecting their growth. Moreover, the differentiation process of NIH-3T3-L1 pre-adipocytes to adipocytes is not compromised by the presence of 5 μg/mL MWCNTs in the medium. These results suggest that reasonable concentrations of PF127-MWCNTs are not toxic for adipocytes and do not interfere with their differentiation process.From the Clinical EditorThis study established that Pluronic F127 (PF127)–coated multiwalled carbon nanotubes (MWCNTs) are tolerated by NIH-3T3-L1 pre-adipocytes. The differentation of these pre-adipocytes is not compromised by the presence of 5 μg/mL MWCNTs, suggesting that these nanotubes are not toxic for adipocytes.     

Copper oxide nanoparticles are highly toxic: a comparison between metal oxide nanoparticles and carbon nanotubes

Since the manufacture and use of nanoparticles are increasing, humans are more likely to be exposed occupationally or via consumer products and the environment. However, so far toxicity data for most manufactured nanoparticles are limited. The aim of this study was to investigate and compare different nanoparticles and nanotubes regarding cytotoxicity and ability to cause DNA damage and oxidative stress. The study was focused on different metal oxide particles (CuO, TiO2, ZnO, CuZnFe2O4, Fe3O4, Fe2O3), and the toxicity was compared to that of carbon nanoparticles and multiwalled carbon nanotubes (MWCNT). The human lung epithelial cell line A549 was exposed to the particles, and cytotoxicity was analyzed using trypan blue staining. DNA damage and oxidative lesions were determined using the comet assay, and intracellular production of reactive oxygen species (ROS) was measured using the oxidation-sensitive fluoroprobe 2',7'-dichlorofluorescin diacetate (DCFH-DA). The results showed that there was a high variation among different nanoparticles concerning their ability to cause toxic effects. CuO nanoparticles were most potent regarding cytotoxicity and DNA damage. The toxicity was likely not explained by Cu ions released to the cell medium. These particles also caused oxidative lesions and were the only particles that induced an almost significant increase (p = 0.058) in intracellular ROS. ZnO showed effects on cell viability as well as DNA damage, whereas the TiO2 particles (a mix of rutile and anatase) only caused DNA damage. For iron oxide particles (Fe3O4, Fe2O3), no or low toxicity was observed, but CuZnFe2O4 particles were rather potent in inducing DNA lesions. Finally, the carbon nanotubes showed cytotoxic effects and caused DNA damage in the lowest dose tested. The effects were not explained by soluble metal impurities. In conclusion, this study highlights the in vitro toxicity of CuO nanoparticles.     

In vitro investigation of oxide nanoparticle and carbon nanotube toxicity and intracellular accumulation in A549 human pneumocytes

If released in the environment, nanomaterials might be inhaled by populations and cause damage to the deepest regions of the respiratory tract, i.e., the alveolar compartment. To model this situation, we studied the response of A549 human pneumocytes after exposure to aluminium oxide or titanium oxide nanoparticles, and to multi-walled carbon nanotubes. The influence of size, crystalline structure and chemical composition was investigated. After a detailed identification of nanomaterial physico-chemical characteristics, cells were exposed in vitro and viability and intracellular accumulation were assessed. In our conditions, carbon nanotubes were more toxic than metal oxide nanoparticles. Our results confirmed that both nanotubes and nanoparticles are able to rapidly enter into cells, and distribute in the cytoplasm and intracellular vesicles. Among nanoparticles, we demonstrate significant difference in biological response as a function of size, crystalline phase and chemical composition. Their toxicity was globally lower than nanotubes toxicity. Among nanotubes, the length did not influence cytotoxicity, neither the presence of metal catalyst impurities.     

Salinity‐dependent toxicity of water‐dispersible,single‐walled carbon nanotubes to Japanese medaka embryos

To investigate the effects of salinity on the behavior and toxicity of functionalized single‐walled carbon nanotubes (SWCNTs), which are chemical modified nanotube to increase dispersibility, medaka embryos were exposed to non‐functionalized single‐walled carbon nanotubes (N‐SWCNTs), water‐dispersible, cationic, plastic‐polymer‐coated, single‐walled carbon nanotubes (W‐SWCNTs), or hydrophobic polyethylene glycol‐functionalized, single‐walled carbon nanotubes (PEG‐SWCNTs) at different salinities, from freshwater to seawater. As reference nanomaterials, we tested dispersible chitin nanofiber (CNF), chitosan‐chitin nanofiber (CCNF) and chitin nanocrystal (CNC, i.e. shortened CNF). Under freshwater conditions, with exposure to 10 mg l⁻¹ W‐SWCNTs, the yolk sacks of 57.8% of embryos shrank, and the remaining embryos had a reduced heart rate, eye diameter and hatching rate. Larvae had severe defects of the spinal cord, membranous fin and tail formation. These toxic effects increased with increasing salinity. Survival rates declined with increasing salinity and reached 0.0% in seawater. In scanning electron microscope images, W‐SWCNTs, CNF, CCNF and CNC were adsorbed densely over the egg chorion surface; however, because of chitin's biologically harmless properties, only W‐SWCNTs had toxic effects on the medaka eggs. No toxicity was observed from N‐SWCNT and PEG‐SWCNT exposure. We demonstrated that water dispersibility, surface chemistry, biomedical properties and salinity were important factors in assessing the aquatic toxicity of nanomaterials. Copyright © 2016 John Wiley & Sons, Ltd.     

Influence of carboxylic acid functionalization on the cytotoxic effects induced by single wall carbon nanotubes on human endothelial cells (HUVEC)

A vast variety of nanomaterials have been developed in the recent years, being carbon nanotubes (CNTs) the ones that have attracted more attention, due to its unique properties which make them suitable for numerous applications. Consequently, it is predicted that tons of CNTs will be produced worldwide every year, being its exposure of toxicological concern. Nanomaterials, once into the body, can translocate from the uptake sites to the blood circulation or the lymphatic system, resulting in distribution throughout the body. Thus, the vascular endothelium can be in contact with them and can suffer from their toxic effects. In this regard, the aim of this work was to investigate the cytotoxicity of single-walled carbon nanotubes (SWCNTs) on human endothelial cells evaluating the influence of acid carboxylic functionalization and also the exposure time (24 and 48 h). Biomarkers assessed were neutral red uptake, protein content, a tetrazolium salt metabolization and cell viability by means of the Trypan blue exclusion test. Cells were exposed to concentrations between 0 and 800 μg/mL SWCNTs for 24 and 48 h. Results have shown that both SWCNTs and carboxylic acid functionalized single-walled carbon nanotubes (COOH-SWCNTs) induce toxic effects in HUVEC cells in a concentration- and time-dependent way. Moreover, the carboxylic acid functionalization results in a higher toxicity compared to the SWCNTs.     

Mitigation of the impact of single-walled carbon nanotubes on a freshwater green algae: Pseudokirchneriella subcapitata

This study investigates the biological response of Pseudokirchneriella subcapitata to single-walled carbon nanotubes (SWNTs) suspended in gum Arabic (GA), using typical 96-hour algal bioassays and long-term growth studies. Changes in algal biomass and cell morphology associated with specific SWNT-treatments were monitored and the mechanisms of observed biological responses investigated through a combination of biochemical and spectroscopic methods. Results from short-term bioassays showed a growth inhibition in culture media containing >0.5 mg SWNT/L and a final GA concentration of 0.023% (v/v). Interestingly, the observed toxicity disappears when GA concentrations are brought to levels ≥ 0.046%. Long-term experiments based on toxic combination of SWNTs and GA showed that P. subcapitata would easily recover from an initial growth inhibition effect. Overall, these findings point to the possibility of GA to mitigate the toxicity of SWNTs, making it an ideal surfactant if SWNT suspension in GA does not alter the performance sought from these nanotubes.     

Comparative toxicity of three differently shaped carbon nanomaterials on Daphnia magna: does a shape effect exist?

The acute toxicity of three differently shaped carbon nanomaterials (CNMs) was studied on Daphnia magna, comparing the induced effects and looking for the toxic mechanisms. We used carbon nano-powder (CNP), with almost spherical primary particle morphology, multi-walled carbon nanotubes (CNTs), tubes of multi-graphitic sheets, and cubic-shaped carbon nanoparticles (CNCs), for which no ecotoxicological data are available so far. Daphnids were exposed to six suspensions (1, 2, 5, 10, 20 and 50?mg L^?1) of each CNM, and then microscopically analyzed. Ultrastructural analyses evidenced cellular uptake of nanoparticle in CNP and CNT exposed groups, but not in samples exposed to CNCs. Despite this difference, very similar effects were observed in tissues exposed to the three used CNMs: empty spaces between cells, cell detachment from the basal lamina, many lamellar bodies and autophagy vacuoles. These pathological figures were qualitatively similar among the three groups, but they differed in frequency and severity. CNCs caused the most severe effects, such as partial or complete dissolution of the brush border and thinning of the digestive epithelium. Being the cubic shape not allowed to be internalized into cells, but more effective than others in determining physical damages, we can conclude that shape is an important factor for driving nanoparticle uptake by cells and for determining the acute toxicological endpoints. Shape also plays a key role in determining the kind and the severity of pathologies, which are linked to the physical interactions of CNMs with the exposed tissues.     

Single-walled carbon nanotubes and graphene oxides induce autophagosome accumulation and lysosome impairment in primarily cultured murine peritoneal macrophages

The wide application of carbon nanomaterials in various fields urges in-depth understanding of the toxic effects and underlying mechanisms of these materials on biological systems. Cell autophagy was recently recognized as an important lysosome-based pathway of cell death, and autophagosome accumulation has been found to be associated with the exposure of various nanoparticles, but the underlying mechanisms are still uncertain due to the fact that autophagosome accumulation can result from autophagy induction and/or autophagy blockade. In this study, we first evaluated the toxicity of acid-functionalized single-walled carbon nanotubes and graphene oxides, and found that both carbon nanomaterials induced adverse effects in murine peritoneal macrophages, and GOs were more potent than AF-SWCNTs. Both carbon nanomaterials induced autophagosome accumulation and the conversion of LC3-I to LC3-II. However, degradation of the autophagic substrate p62 protein was also inhibited by both nanomaterials. Further analyses on lysosomes revealed that both carbon nanomaterials accumulated in macrophage lysosomes, leading to lysosome membrane destabilization, which indicates reduced autophagic degradation. The effects of AF-SWCNTs and GOs on cell autophagy revealed by this study may shed light on the potential toxic mechanism and suggest caution on their utilization.     

Multi-walled carbon nanotube-induced inhalation toxicity: Recognizing nano bis-demethoxy curcumin analog as an ameliorating candidate

Human beings and ecosystems are being possibly exposed to CNTs, as there is a rise in global production rate of carbon nanotubes (CNTs). This may affect the health of humans and increases the environmental risk. We have already reported the pulmonary toxicity due to the inhalation of MWCNTs. We claim that a compound with anti-inflammatory and antioxidant activity may ameliorate the CNT-induced toxic effect. With this view, we have investigated the ameliorative effect of intravenously-administered nano bis-demethoxy curcumin analog (NBDMCA) against MWCNTs-induced inhalation toxicity by examining the lung histopathology for inflammatory cell dynamics, pulmonary remodeling and estimating the inflammatory biomarkers in the broncho-alveolar lavage fluid. We observed that NBDMCA could ameliorate the injury as evidenced by the decline in the levels of markers of inflammation, cell damage, and the histopathological changes induced by MWCNTs. We conclude that NBDMCA may be used to reduce the risk of MWCNTs-induced inhalation toxicity.     

Toxicity and clearance of intratracheally administered multiwalled carbon nanotubes from murine lung

Carbon nanotubes (CNT) are known to have widespread industrial applications; however, several reports indicated that these compounds may be associated with adverse effects in humans. In this study, multiwalled carbon nanotubes were administered to murine lungs intratracheally to determine whether acute and chronic pulmonary toxicity occurred. In particular, pristine multiwalled carbon nanotubes (PMWCNT) and acid-treated multiwalled carbon nanotubes (TMWCNT) were used in this study. In broncheoalveolar lavage fluid (BALF) cell analysis, PMWCNT induced more severe acute inflammatory cell recruitment than TMWCNT. Histopathologically, both PMWCNT and TMWCNT induced multifocal inflammatory granulomas in a dose-dependent manner. The observed granulomas were reversible, with TMWCNT-induced granulomas diminishing faster than PMWCNT-induced granulomas. Although the area of granuloma reduced with time, hyperplasia and dysplastic characteristics such as mitotic figures, anisokaryosis, and anisocytosis were still observed. These findings demonstrate that MWCNT induces granulomatous inflammation, and the duration and pattern of inflammation seem to vary depending upon the types of MWCNT to which mice are exposed. Therefore, toxicity studies on various types of CNT are needed as the responsiveness to these compounds differs.     

Influence of acid functionalization on the cardiopulmonary toxicity of carbon nanotubes and carbon black particles in mice

Engineered carbon nanotubes are being developed for a wide range of industrial and medical applications. Because of their unique properties, nanotubes can impose potentially toxic effects, particularly if they have been modified to express functionally reactive chemical groups on their surface. The present study was designed to evaluate whether acid functionalization (AF) enhanced the cardiopulmonary toxicity of single-walled carbon nanotubes (SWCNT) as well as control carbon black particles. Mice were exposed by oropharyngeal aspiration to 10 or 40 μg of saline-suspended single-walled carbon nanotubes (SWCNTs), acid-functionalized SWCNTs (AF-SWCNTs), ultrafine carbon black (UFCB), AF-UFCB, or 2 μg LPS. 24 hours later, pulmonary inflammatory responses and cardiac effects were assessed by bronchoalveolar lavage and isolated cardiac perfusion respectively, and compared to saline or LPS-instilled animals. Additional mice were assessed for histological changes in lung and heart. Instillation of 40 μg of AF-SWCNTs, UFCB and AF-UFCB increased percentage of pulmonary neutrophils. No significant effects were observed at the lower particle concentration. Sporadic clumps of particles from each treatment group were observed in the small airways and interstitial areas of the lungs according to particle dose. Patches of cellular infiltration and edema in both the small airways and in the interstitium were also observed in the high dose group. Isolated perfused hearts from mice exposed to 40 μg of AF-SWCNTs had significantly lower cardiac functional recovery, greater infarct size, and higher coronary flow rate than other particle-exposed animals and controls, and also exhibited signs of focal cardiac myofiber degeneration. No particles were detected in heart tissue under light microscopy. This study indicates that while acid functionalization increases the pulmonary toxicity of both UFCB and SWCNTs, this treatment caused cardiac effects only with the AF-carbon nanotubes. Further experiments are needed to understand the physico-chemical processes involved in this phenomenon.     

Significant toxic role for single-walled carbon nanotubes during normal embryogenesis

In order to investigate the effect of SWCNTs in the embryo, we examined the outcome of SWCNTs in avian embryo at an early stage of development. We found that SWCNTs-treatment inhibits the angiogenesis of the chorioallantoic membrane (CAM) and in the chicken embryo. Moreover, we showed that SWCNTs can harm the normal development of the embryo since all SWCNTs-exposed embryos are smaller in comparison with their matched controls. We also found that the majority of SWCNTs-exposed embryos die before 12 days of incubation. Macroscopic examination did not reveal any anomalies in these embryos. However, RT-PCR analysis of eleven genes, which are important regulators of cell proliferation, apoptosis, survival and angiogenesis, shows that these genes are deregulated in brain and liver tissues from SWCNTs-treated embryos in comparison with their matched controls. This study suggests that SWCNTs could have a very toxic effect on the normal development of the embryo.From the Clinical EditorIn this study, a significant toxicity of single-walled carbon nanotubes was observed during normal embryogenesis: the nanotubes inhibited the angiogenesis of the chorioallantoic membrane (CAM) in chicken embryos. All exposed embryos died before 12 days of incubation, suggesting a severe effect.     

Short‐term splenic impact of single‐strand DNA functionalized multi‐walled carbon nanotubes intraperitoneally injected in rats

In recent years, a great deal of studies have focused on the possible toxicity of carbon nanotubes (CNT), as a result of their potential applications in the field of nanotechnologies. The investigation of spleen toxicity is part of the carbon nanotubes‐induced toxicity assessment. In this study, we investigated the possible toxic effects of CNT on the rat spleen, after intraperitoneally (i.p.) administration of a single dose [1.5 ml; 2 mg multi‐walled (MW) CNT per body weight (bw)] of multi‐walled carbon nanotubes (exterior diameter 15–25 nm, interior diameter 10–15 nm, surface 88 m² g^–1) functionalized 1:1 with single‐strand DNA (ss‐DNA‐MWCNT, 270 mg l^–1). CNT functionalization with DNA determines a stable dispersion in the body fluids. For the detection of carbon nanotubes in the spleen, Raman spectroscopy, histopathologic examination, confocal microscopy and transmission electron microscopy (TEM) were performed at different time points (1, 6, 24, 48 and 144 h) after MWCNT administration. The dynamics of oxidative stress parameters (malondialdehyde, protein carbonyls and reduced glutathione), along with nitrosative stress parameters (nitric oxide, inducible NO synthase), the pro‐inflammatory cytokines [interleukin‐(IL)‐1β] and the number of cells expressing caspase 3 and proliferating cell nuclear antigen (PCNA) were assessed. Our results indicate that, after i.p. administration, MWCNT translocate progressively in the spleen, with a peak of concentration after 48 h, and determine lymphoid hyperplasia and an increase in the number of cells which undergo apoptosis, in parallel with the enhancement of the mitosis in the white pulp and with transient alterations of oxidative stress and inflammation that need further investigations for a longer period of monitoring. Copyright © 2013 John Wiley & Sons, Ltd.     

Exposure to Carbon Nanotube Material: Assessment of Nanotube Cytotoxicity using Human Keratinocyte Cells

Carbon nanotubes are new members of carbon allotropes similar to fullerenes and graphite. Because of their unique electrical, mechanical, and thermal properties, carbon nanotubes are important for novel applications in the electronics, aerospace, and computer industries. Exposure to graphite and carbon materials has been associated with increased incidence of skin diseases, such as carbon fiber dermatitis, hyperkeratosis, and naevi. We investigated adverse effects of single-wall carbon nanotubes (SWCNT) using a cell culture of immortalized human epidermal keratinocytes (HaCaT). After 18 h of exposure of HaCaT to SWCNT, oxidative stress and cellular toxicity were indicated by formation of free radicals, accumulation of peroxidative products, antioxidant depletion, and loss of cell viability. Exposure to SWCNT also resulted in ultrastructural and morphological changes in cultured skin cells. These data indicate that dermal exposure to unrefined SWCNT may lead to dermal toxicity due to accelerated oxidative stress in the skin of exposed workers.     

Exposure to carbon nanotube material: assessment of nanotube cytotoxicity using human keratinocyte cells

Carbon nanotubes are new members of carbon allotropes similar to fullerenes and graphite. Because of their unique electrical, mechanical, and thermal properties, carbon nanotubes are important for novel applications in the electronics, aerospace, and computer industries. Exposure to graphite and carbon materials has been associated with increased incidence of skin diseases, such as carbon fiber dermatitis, hyperkeratosis, and naevi. We investigated adverse effects of single-wall carbon nanotubes (SWCNT) using a cell culture of immortalized human epidermal keratinocytes (HaCaT). After 18 h of exposure of HaCaT to SWCNT, oxidative stress and cellular toxicity were indicated by formation of free radicals, accumulation of peroxidative products, antioxidant depletion, and loss of cell viability. Exposure to SWCNT also resulted in ultrastructural and morphological changes in cultured skin cells. These data indicate that dermal exposure to unrefined SWCNT may lead to dermal toxicity due to accelerated oxidative stress in the skin of exposed workers.     

In vitro toxicity of acid-functionalized single-walled carbon nanotubes: effects on murine macrophages and gene expression profiling

Single-walled carbon nanotubes (SWCNTs) are widely used in industrial and medical sectors, and the increasing exposure of SWCNTs necessitates the studies of their potential environmental and health effects. Considerable efforts have been made to improve the dispersion of SWCNTs by chemical modifications. However, the toxicological effects of such modifications on SWCNTs are mostly unknown. This study was designed to determine the influences of acid functionalization on SWCNT toxicity and to understand the molecular toxic mechanisms. RAW264.7 cells were exposed to 0-50 μg/mL of as-synthesized SWCNTs or acid-functionalized SWCNTs (AF-SWCNTs) for 24 hours and then their toxicities were compared via viability analysis. After that the global gene expression profiles of cells exposed to AF-SWCNTs were obtained and analyzed. The results showed that AF-SWCNTs penetrated cell membrane and aggregated in cell cytoplasm and nuclear areas, resulting in enhanced toxicity. In addition, AF-SWCNTs altered the expression of genes related to ribosome, mitochondria, inflammatory response, cell cycle/apoptosis, and proteasome pathway. The gene expression study excluded the interference of metallic impurities and suggested similar toxic mechanism to that of ultra-fine particulate matters.