Yongbin Zhang

Cytotoxicity Effects of Graphene and Single-Wall Carbon Nanotubes in Neural Phaeochromocytoma-Derived PC12 Cells

Graphitic nanomaterials such as graphene layers (G) and single-wall carbon nanotubes (SWCNT) are potential candidates in a large number of biomedical applications. However, little is known about the effects of these nanomaterials on biological systems. Here we show that the shape of these materials is directly related to their induced cellular toxicity. Both G and SWCNT induce cytotoxic effects, and these effects are concentration- and shape-dependent. Interestingly, at low concentrations, G induced stronger metabolic activity than SWCNT, a trend that reversed at higher concentrations. Lactate dehydrogenase levels were found to be significantly higher for SWCNT as compared to the G samples. Moreover, reactive oxygen species were generated in a concentration- and time-dependent manner after exposure to G, indicating an oxidative stress mechanism. Furthermore, time-dependent caspase 3 activation after exposure to G (10 microg/mL) shows evidence of apoptosis. Altogether these studies suggest different biological activities of the graphitic nanomaterials, with the shape playing a primary role.

Mechanistic toxicity evaluation of uncoated and PEGylated single-walled carbon nanotubes in neuronal PC12 cells.

We investigated and compared the concentration-dependent cytotoxicity of single-walled carbon nanotubes (SWCNTs) and SWCNTs functionalized with polyethylene glycol (SWCNT-PEGs) in neuronal PC12 cells at the biochemical, cellular, and gene expressional levels. SWCNTs elicited cytotoxicity in a concentration-dependent manner, and SWCNT-PEGs exhibited less cytotoxic potency than uncoated SWCNTs. Reactive oxygen species (ROS) were generated in both a concentration- and surface coating-dependent manner after exposure to these nanomaterials, indicating different oxidative stress mechanisms. More specifically, gene expression analysis showed that the genes involved in oxidoreductases and antioxidant activity, nucleic acid or lipid metabolism, and mitochondria dysfunction were highly represented. Interestingly, alteration of the genes is also surface coating-dependent with a good correlation with the biochemical data. These findings suggest that surface functionalization of SWCNTs decreases ROS-mediated toxicological response in vitro.

Genotoxicity of silver nanoparticles evaluated using the Ames test and in vitro micronucleus assay

Silver nanoparticles (AgNPs) have antimicrobial properties, which have contributed to their widespread use in consumer products. A current issue regarding nanomaterials is the extent to which existing genotoxicity assays are useful for evaluating the risks associated with their use. In this study, the genotoxicity of 5 nm AgNPs was assessed using two standard genotoxicity assays, the Salmonella reverse mutation assay (Ames test) and the in vitro micronucleus assay. Using the preincubation version of the Ames assay, Salmonella strains TA102, TA100, TA1537, TA98, and TA1535 were treated with 0.15-76.8 μg/plate of the AgNPs. Toxicity limited the doses that could be assayed to 2.4-38.4 μg/plate; no increases in mutant frequency over the vehicle control were found for the concentrations that could be assayed. Human lymphoblastoid TK6 cells were treated with 10-30 μg/ml AgNPs, and additional cells were treated with water and 0.73 gy X-rays as vehicle and positive controls. Micronucleus frequency was increased by the AgNP treatment in a dose-dependent manner. At a concentration of 30 μg/ml (with 45.4% relative population doubling), AgNPs induced a significant, 3.17-fold increase with a net increase of 1.60% in micronucleus frequency over the vehicle control, a weak positive response by our criteria. These results demonstrate that the 5 nm AgNP are genotoxic in TK6 cells. Also, the data suggest that the in vitro micronucleus assay may be more appropriate than the Ames test for evaluating the genotoxicity of the AgNPs.

Toxicity and efficacy of carbon nanotubes and graphene: the utility of carbon-based nanoparticles in nanomedicine

Abstract Carbon-based nanomaterials have attracted great interest in biomedical applications such as advanced imaging, tissue regeneration, and drug or gene delivery. The toxicity of the carbon nanotubes and graphene remains a debated issue although many toxicological studies have been reported in the scientific community. In this review, we summarize the biological effects of carbon nanotubes and graphene in terms of in vitro and in vivo toxicity, genotoxicity and toxicokinetics. The dose, shape, surface chemistry, exposure route and purity play important roles in the metabolism of carbon-based nanomaterials resulting in differential toxicity. Careful examination of the physico-chemical properties of carbon-based nanomaterials is considered a basic approach to correlate the toxicological response with the unique properties of the carbon nanomaterials. The reactive oxygen species-mediated toxic mechanism of carbon nanotubes has been extensively discussed and strategies, such as surface modification, have been proposed to reduce the toxicity of these materials. Carbon-based nanomaterials used in photothermal therapy, drug delivery and tissue regeneration are also discussed in this review. The toxicokinetics, toxicity and efficacy of carbon-based nanotubes and graphene still need to be investigated further to pave a way for biomedical applications and a better understanding of their potential applications to humans.

Silver nanoparticle‐induced mutations and oxidative stress in mouse lymphoma cells

Silver nanoparticles (Ag‐NPs) have increasingly been used for coatings on various textiles and certain implants, for the treatment of wounds and burns, as a water disinfectant, and in air‐freshener sprays. The wide use of Ag‐NPs may have potential human health impacts. In this study, the mutagenicity of 5‐nm Ag‐NPs was evaluated in the mouse lymphoma assay system, and modes of action were assessed using standard alkaline and enzyme‐modified Comet assays and gene expression analysis. Treatments of L5178Y/Tk+/‐ mouse lymphoma cells with 5‐nm uncoated Ag‐NPs resulted in a significant yield of mutants at doses between 3 and 6 μg/mL; the upper range was limited by toxicity. Loss of heterozygosity analysis of the Tk mutants revealed that treatments with uncoated Ag‐NPs induced mainly chromosomal alterations spanning less than 34 megabase pairs on chromosome 11. Although no significant induction of DNA damage in Ag‐NP‐treated mouse lymphoma cells was observed in the standard Comet assay, the Ag‐NP treatments induced a dose‐responsive increase in oxidative DNA damage in the enzyme‐modified Comet assay in which oxidative lesion‐specific endonucleases were added. Gene expression analysis using an oxidative stress and antioxidant defense polymerase chain reaction (PCR) array showed that the expressions of 17 of the 59 genes on the arrays were altered in the cells treated with Ag‐NPs. These genes are involved in production of reactive oxygen species, oxidative stress response, antioxidants, oxygen transporters, and DNA repair. These results suggest that 5 nm Ag‐NPs are mutagenic in mouse lymphoma cells due to induction of oxidative stress by the Ag‐NPs. Environ. Mol. Mutagen. 2012.

Neurotoxicity of nanoscale materials.

Nanotechnology has been applied in consumer products and commercial applications, showing a significant impact on almost all industries and all areas of society. Significant evidence indicates that manufactured nanomaterials and combustion-derived nanomaterials elicit toxicity in humans exposed to these nanomaterials. The interaction of the engineered nanomaterials with the nervous system has received much attention in the nanotoxicology field. In this review, the biological effects of metal, metal oxide, and carbon-based nanomaterials on the nervous system are discussed from both in vitro and in vivo studies. The translocation of the nanoparticles through the blood-brain barrier or nose to brain via the olfactory bulb route, oxidative stress, and inflammatory mechanisms of nanomaterials are also reviewed.

Silver Nanoparticles Decrease Body Weight and Locomotor Activity in Adult Male Rats

Silver nanoparticles (Ag-NPs) are widely used in FDA regulated products. The physical-chemical properties of Ag-NPs are characterized using various instruments. The dose-dependent activity and body weight alterations are evaluated after rats were exposed to Ag nanoparticles, suggesting a major human health risk, given the wide application of silver nanomaterials.

Raman spectroscopy as a detection and analysis tool for in vitro specific targeting of pancreatic cancer cells by EGF‐conjugated, single‐walled carbon nanotubes

Single‐walled carbon nanotubes (SWCNTs) were covalently linked to epidermal growth factor (EGF) proteins through an esterification process that was found to be responsible for the docking of SWCNTs on the human pancreatic cancer cells (PANC‐1) surface, thus providing a mechanism for the enhanced delivery and internalization of the nanotubes. Micro Raman spectroscopy and enzyme‐linked immunosorbent assay were used to evaluate the delivery process and kinetics of the SWCNTs. In vitro studies indicated that the delivery kinetics of SWCNT–EGF conjugates, at a concentration of 85 µg ml−1, to the PANC‐1 cell surfaces was significant in the first 30 min of incubation, but reached a plateau with time in accordance with the establishment of equilibrium between the association and the dissociation of EGF with the cell receptors. SWCNT–EGF conjugates could act as strong thermal ablation agents and could induce higher percentages of cellular death compared with the nontargeted SWCNTs alone. Copyright

Carbon nanotubes enhance the internalization of drugs by cancer cells and decrease their chemoresistance to cytostatics

Etoposide is a semisynthetic, chemotherapeutic drug widely recommended to treat an extensive range of human cancers. Our studies indicate that, while etoposide is capable of killing human cancer cells, exposure to single-walled carbon nanotubes (SWCNTs) and etoposide results in enhanced cell death that appears to be synergistic and not merely additive. In this study, we used high pressure liquid chromatography and mass spectrometry to quantify the internal effective dose of etoposide when the human pancreatic cancer cell (PANC-1) was exposed to the combination of these agents. Our results unequivocally indicate that SWCNTs improve etoposide uptake and increase its capacity to kill cancer cells. We suggest that a combination of SWCNTs and etoposide may prove to be a more efficient chemotherapeutic protocol, especially because of the potential to lower toxic drug doses to levels that may be useful in decreasing adverse side effects, as well as in lowering the probability of inducing chemoresistance in exposed cancer cells.

Radio-frequency induced in vitro thermal ablation of cancer cells by EGF functionalized carbon-coated magnetic nanoparticles

Carbon-shelled, iron-based magnetic nanoparticles (C/Fe MNPs) were found to act as strong heat generating agents when exposed to radio-frequency (RF) energy with the ability to thermally destroy cancer cells. In order to efficiently deliver MNPs to cancer cells and to enhance the effectiveness of the RF treatment, human epidermal growth factor (EGF) was bioconjugated with the C/Fe MNPs for their specific delivery to two cancer cell lines, MCF-7 breast cancer cells and Panc-1 pancreatic cancer cells, respectively. These cell lines overexpress the epidermal growth factor receptors (EGFRs) and were used in this study as models. EGF-MNPs have shown higher surface binding efficiency towards the MCF-7 cells based on the comparative ζ-potential measurements. Confocal optical microscopy further confirmed that EGF-bioconjugated MNPs highly accumulated around and inside of these cancer cells. RF treatment was found to destroy 92.8% of MCF-7 breast cancer cells during 10 minutes of treatment when EGF was bound to the nanoparticles, while 37.3% of cells died when MNPs alone were used in identical conditions. Panc-1 cancer cells exhibit a higher resistance than MCF-7 cells when they were exposed to MNPs or RF treatment. Cytotoxicity studies demonstrated that the EGF-C/Fe MNP bioconjugates present lower toxicity compared to the C/Fe MNP. Caspase assay studies demonstrated that the MCF-7 cancer cells underwent an apoptotic process by the caspase 3 deficiency pathway showing no evidence of morphological changes such as membrane blebbing.