A critical review of the biological mechanisms underlying the in vivo and in vitro toxicity of carbon nanotubes: The contribution of physico-chemical characteristics

Abstract

This critical review of the available human health safety data, relating to carbon nanotubes (CNTs), was conducted in order to assess the risks associated with CNT exposure. Determining the toxicity related to CNT exploitation is of great relevance and importance due to the increased potential for human exposure to CNTs within occupational, environmental and consumer settings. When this information is combined with knowledge on the likely exposure levels of humans to CNTs, it will enable risk assessments to be conducted to assess the risks posed to human health. CNTs are a diverse group of materials and vary with regards to their wall number (single and multi-walled CNTs are evident), length, composition, and surface chemistry. The attributes of CNTs that were identified as being most likely to drive the observed toxicity have been considered, and include CNT length, metal content, tendency to aggregate/agglomerate and surface chemistry. Of particular importance, is the contribution of the fibre paradigm to CNT toxicity, whereby the length of CNTs appears to be critical to their toxic potential. Mechanistic processes that are critical to CNT toxicity will also be discussed, with the findings insinuating that CNTs can exert an oxidative response that stimulates inflammatory, genotoxic and cytotoxic consequences. Consequently, it may transpire that a common mechanism is responsible for driving CNT toxicity, despite the fact that CNTs are a diverse population of materials. The similarity of the structure of CNTs to that of asbestos has prompted concern surrounding the exposure of humans, and so the applicability of the fibre paradigm to CNTs will be evaluated. It is also necessary to determine the systemic availability of CNTs following exposure, to determine where potential targets of toxicity are, and to thereby direct in vitro investigations within the most appropriate target cells. CNTs are therefore a group of materials whose useful exploitable properties prompts their increased production and utilization within diverse applications, so that ensuring their safety is of vital importance.     

Biodistribution and toxicity of nanodiamonds in mice after intratracheal instillation

Nanodiamonds (NDs) are receiving increasing attention in materials science and nanotechnology-based industries for a large variety of applications, including protein immobilization, biosensors, therapeutic molecule delivery, and bioimaging. However, limited information is known about their biokinetic behavior and toxicity in vivo. In this article, we investigated the biodistribution of NDs using radiotracer techniques and evaluated its acute toxicity in Kun Ming mice after intratracheal instillation. The biodistribution showed that, besides having the highest retention in the lung, NDs were distributed mainly in the spleen, liver, bone and heart. An analysis of histological morphology and biochemical parameters indicated that NDs could induce dose-dependent toxicity to the lung, liver, kidney and blood. This work provided fundamental data for understanding the biodistribution of NDs and will provide guidance for further study of their toxicity.     

Carbon nanotubes part II: a remarkable carrier for drug and gene delivery

Introduction: Carbon nanotubes (CNT) have recently been studied as novel and versatile drug and gene delivery vehicles. When CNT are suitably functionalized, they can interact with various cell types and are taken up by endocytosis.

Areas covered: Anti-cancer drugs cisplatin and doxorubicin have been delivered by CNT, as well as methotrexate, taxol and gemcitabine. The delivery of the antifungal compound amphotericin B and the oral administration of erythropoietin have both been assisted using CNT. Frequently, targeting moieties such as folic acid, epidermal growth factor or various antibodies are attached to the CNT-drug nanovehicle. Different kinds of functionalization (e.g., polycations) have been used to allow CNT to act as gene delivery vectors. Plasmid DNA, small interfering RNA and micro-RNA have all been delivered by CNT vehicles. Significant concerns are raised about the nanotoxicology of the CNT and their potentially damaging effects on the environment.

Expert opinion: CNT-mediated drug delivery has been studied for over a decade, and both in vitro and in vivo studies have been reported. The future success of CNTs as vectors in vivo and in clinical application will depend on achievement of efficacious therapy with minimal adverse effects and avoidance of possible toxic and environmentally damaging effects.     

Pharmacokinetics and in vivo fate of drug loaded chitosan nanoparticles

Chitosan is a natural polysaccharide which is generally biodegradable, biocompatible and mucoadhesive, thus, attracting considerable interest of scientific researchers. The application of chitosan as nanocarriers for drug delivery thrived. And some of their pharmacokinetics and biodistribution profiles were studied, which are crucial to develop a promising drug delivery system. In this article, we will first give an introduction for the chitosan as drug delivery system, especially as nanoparticles. Then, we focus on pharmacokinetics studies of various chitosan nanoparticles both in vitro and in vivo. In a following part, we refer to researches on biodistribution properties of chitosan nanoparticles. Here we crucially discuss the in vivo fate of chitosan nanoparticles. And finally, toxicity issue is discussed and conclusions are drawn.     

Application of carbon nanotubes in neurology: clinical perspectives and toxicological risks

Nanomedicine is an emerging field that proposes the application of precisely engineered nanomaterials for the prevention, diagnosis and therapy of certain diseases, including neurological pathologies. Carbon nanotubes (CNT) are a new class of nanomaterials, which have been shown to be promising in different areas of nanomedicine. In this review, the application of CNT interfacing with the central nervous system (CNS) will be described, and representative examples of neuroprosthetic devices, such as neuronal implants and electrodes will be discussed. Furthermore, the possible application of CNT-based materials as regenerative matrices of neuronal tissue and as delivery systems for the therapy of CNS will be presented.     

Determinants of carbon nanotube toxicity

In the last few years questions have been raised regarding the potential toxicity of carbon nanotubes (CNTs) to humans and environment. It is believed that the physico-chemical characteristics of these materials are key determinants of CNT interaction with living organisms, and hence determine their toxicity. As for other nanomaterials, the most important of these characteristics are the length, diameter, surface area, tendency to agglomerate, bio-durability, presence and nature of catalyst residues as well as chemical functionalization of the CNT. This review highlights the recent advancements in the understanding of the CNT properties which are essential in determining CNT toxicity. Hence the focus is on CNT dimensions, surface properties, bio-durability and corona formation as these fields have evolved greatly in recent years.     

Effect of BSA on carbon nanotube dispersion for in vivo and in vitro studies

Carbon nanotubes (CNT) are one of the most promising nanomaterials because of their intrinsic properties. So, it becomes urgent to assess their toxicity. However, CNT are insoluble in aqueous media required for toxicological studies. Thus, we propose a simple method to disperse CNT for toxicological studies using a biomolecule: The albumin. To evaluate this method, several nanotubes were suspended in saline solution (NaCl 0.9%) without or with albumin at a concentration of 0.5 mg/ml or equal as CNT concentration. These suspensions were visually compared to suspensions obtained with classical dispersing methods using Tween 80 or serum. Homogeneity of the suspensions with or without BSA and CNT structure were analyzed by TEM, agglomerates quantification and total carbon dosage. The effect of coupled albumin-CNT was then tested on A549 and U937 cells in vitro and on rats in vivo. Total carbon dosage, agglomerates quantification and TEM revealed that, in the presence of albumin, the tested nanotubes were better dispersed without any modification of their structure. The CNT suspension was tested in vitro and in vivo in rats. Albumin solution alone induced no modification of the biological responses studied (i.e., cell viability in vitro and inflammatory response and histopathology in vivo) compared to the saline. CNT in NaCl or BSA altered cellular viability in vitro in a similar way but results obtained with CNT suspension in the presence of albumin showed a better reproducibility that can be explained by the better homogeneity of the suspensions. CNT in BSA but not in NaCl significantly increased the cell number in BAL and also the number of apparent CNT-containing cells. Taken together, these results highlight the potential importance of CNT dispersion (and thus of the vehicle) for the toxicological studies.     

Respiratory toxicity of multi-wall carbon nanotubes

Carbon nanotubes focus the attention of many scientists because of their huge potential of industrial applications, but there is a paucity of information on the toxicological properties of this material. The aim of this experimental study was to characterize the biological reactivity of purified multi-wall carbon nanotubes in the rat lung and in vitro. Multi-wall carbon nanotubes (CNT) or ground CNT were administered intratracheally (0.5, 2 or 5 mg) to Sprague-Dawley rats and we estimated lung persistence, inflammation and fibrosis biochemically and histologically. CNT and ground CNT were still present in the lung after 60 days (80% and 40% of the lowest dose) and both induced inflammatory and fibrotic reactions. At 2 months, pulmonary lesions induced by CNT were characterized by the formation of collagen-rich granulomas protruding in the bronchial lumen, in association with alveolitis in the surrounding tissues. These lesions were caused by the accumulation of large CNT agglomerates in the airways. Ground CNT were better dispersed in the lung parenchyma and also induced inflammatory and fibrotic responses. Both CNT and ground CNT stimulated the production of TNF-alpha in the lung of treated animals. In vitro, ground CNT induced the overproduction of TNF-alpha by macrophages. These results suggest that carbon nanotubes are potentially toxic to humans and that strict industrial hygiene measures should to be taken to limit exposure during their manipulation.     

Two faces of carbon nanotube: toxicities and pharmaceutical applications

In the field of nanotechnology, carbon nanotube (CNT) is gaining importance for the delivery of therapeutic agents and diagnosis of diseases. CNT is emerging as an efficient nanocarrier system with cylindrical nanostructure. Due to its nanoscale dimensions, CNTs have a high cell-penetration quality that allows its use in site-specific targeting. Another aspect of the utilization of CNT lies in its hollow structure through which an active moiety can be delivered in a controlled manner via CNTs' nano channels. Despite these positive aspects of CNT, scientists are still working to improve its biocompatibility and solubility and eliminating toxicity in vivo, which are creating problems with the use of CNTs. Therefore, functionalization becomes an important aspect to be studied because it decreases the toxicity of CNTs and make them nonimmunogenic. In this review, different functionalization techniques of CNTs and their biomedical applications-in particular for cancer therapy to date-are reviewed in detail to present the potential of this nanovector.     

Advances in mechanisms and signaling pathways of carbon nanotube toxicity

Abstract

Carbon nanotubes (CNT) have been developed into new materials with a variety of industrial and commercial applications. In contrast, the physicochemical properties of CNT at the nanoscale render them the potency to generate toxic effects. Indeed, the potential health impacts of CNT have drawn a great deal of attention in recent years, owing to their identified toxicological and pathological consequences including cytotoxicity, inflammation, fibrosis, genotoxicity, tumorigenesis, and immunotoxicity. Understanding the mechanisms by which CNT induce toxicity and pathology is thus urgently needed for accurate risk assessment of CNT exposure in humans, and for safe and responsible development and commercialization of nanotechnology. Here, we summarize and discuss recent advances in this area with a focus on the molecular interactions between CNT and mammalian systems, and the signaling pathways important for the development of CNT toxicity such as the NF-κB, NLRP3 inflammasome, TGF-β1, MAPK, and p53 signaling cascades. With the current mechanistic evidence summarized in this review, we expect to provide new insights into CNT toxicology at the molecular level and offer new clues to the prevention of health effects resulting from CNT exposure. Moreover, we disclose questions and issues that remain in this rapidly advancing field of nanotoxicology, which would facilitate ascertaining future research directions.     

Carbon nanotubes size classification,characterization and nasal airway deposition

Abstract

Workers and researchers in the carbon nanotubes (CNT)-related industries and laboratories might be exposed to CNT aerosols while generating and handling CNT materials. From the viewpoint of occupational health, it is essential to study the deposition of CNT aerosol in the human respiratory tract to investigate the potential adverse health effects. In this study, a human nasal airway replica and two types of CNT materials were employed to conduct CNT nasal airway deposition studies. The two CNT materials were aerosolized by a nebulizer-based wet generation method, with size classified by three designated classification diameters (51, 101 and 215?nm), and then characterized individually in terms of their morphology and aerodynamic diameter. The nasal deposition experiments were carried out by delivering the size classified CNTs into the nasal airway replica in three different inspiratory flow rates. From the characterization study, it showed that the morphology of the size classified CNTs could be in a variety of complex shapes with their physical dimension much larger than their classification diameter. In addition, it was found that the aerodynamic diameters of the classified CNTs were slightly smaller than their classification diameter. The nasal deposition data acquired in this study showed that the deposition efficiency of CNTs in the nasal airway were generally less than 0.1, which implies that the majority of the CNTs inhaled into the nose could easily penetrate through the entire nasal airway and transit further down to the lower airways, possibly causing adverse health effects.     

Molecular nanomedicine towards cancer: ¹¹¹In-labeled nanoparticles

Nanomedicine is the medical application of materials, devices, or systems in the nanometer scale and is currently undergoing explosive development. Molecular imaging of cancer using nanosized materials comprises an important part in diagnosis, therapy, and drug discovery in medical nanosciences. Radiopharmaceuticals are a key tool of molecular imaging in the field of nuclear medicine. The in vivo administration of radiolabeled nanoparticles (NPs) can provide an accurate biodistribution profile of the nanoformulations, as well as visualization of their route in vivo. Surface modifications of NPs with antibodies, peptides, or other small molecules that bind to tumor cell receptors have resulted in the development of sensitive and specific targeted imaging and diagnostic modalities for in vitro and in vivo applications. Radiometals are the most favorable of all radionuclides for labeling applications and they have the most suitable properties for single-photon emission computed tomography imaging. Indium-111((111)In), specifically, is a readily available gamma-emitting radiometal, which is widely used in clinical practice for diagnosis and/or therapy. Herein, we will overview the latest evolvement on (111)In-labeled nanoparticles for biodistribution assessment and/or imaging evaluation of nanocarriers, as well as therapy in cancer.     

Biodistribution of [125I]-labeled therapeutic proteins: Application in protein drug development beyond oncology

The majority of biodistribution studies of therapeutic proteins published to date focus on tumor-targeting agents. In this report we present a number of case studies that demonstrate the utility of biodistribution studies during preclinical development of biotherapeutics for non oncology indications, as well as provide a practical perspective on the methodology applied to these studies. For the commonly used classes of biologics (such as human monoclonal antibodies), biodistribution profiles may be compared to those of other therapeutics of the same class and compounds with unexpected off-target mediated uptake may be identified. Temporal biodistribution profiles may be used to address kinetics and reversibility of target- and/or off-target-mediated accumulation. In cases when circulating biotherapeutic is rapidly eliminated from circulation due to the formation of anti-product antibodies, tissue data may provide useful insight on test article exposure at the site of therapeutic action (or at the site of toxicity). Comparison of temporal biodistribution profiles between the genetically engineered and wild-type mouse strains or between the disease models and healthy animals may provide useful insight on sites and kinetics of target-mediated elimination. Finally, biodistribution studies will be a useful tool to study in vivo disposition for a variety of existing and upcoming novel classes of protein compounds. © 2009 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 99:1028–1045, 2010     

Pulmonary toxicity of carbon nanotubes and asbestos — Similarities and differences

Carbon nanotubes are a valuable industrial product but there is potential for human pulmonary exposure during production and their fibrous shape raises the possibility that they may have effects like asbestos, which caused a worldwide pandemic of disease in the20th century that continues into present. CNT may exist as fibres or as more compact particles and the asbestos-type hazard only pertains to the fibrous forms of CNT. Exposure to asbestos causes asbestosis, bronchogenic carcinoma, mesothelioma, pleural fibrosis and pleural plaques indicating that both the lungs and the pleura are targets. The fibre pathogenicity paradigm was developed in the 1970s–80s and has a robust structure/toxicity relationship that enables the prediction of the pathogenicity of fibres depending on their length, thickness and biopersistence. Fibres that are sufficiently long and biopersistent and that deposit in the lungs can cause oxidative stress and inflammation. They may also translocate to the pleura where they can be retained depending on their length, and where they cause inflammation and oxidative stress in the pleural tissues. These pathobiological processes culminate in pathologic change — fibroplasia and neoplasia in the lungs and the pleura. There may also be direct genotoxic effects of fibres on epithelial cells and mesothelium, contributing to neoplasia. CNT show some of the properties of asbestos and other types of fibre in producing these types of effects and more research is needed. In terms of the molecular pathways involved in the interaction of long biopersistent fibres with target tissue the events leading to mesothelioma have been a particular area of interest. A variety of kinase pathways important in proliferation are activated by asbestos leading to pre-malignant states and investigations are under way to determine whether fibrous CNT also affects these molecular pathways. Current research suggests that fibrous CNT can elicit effects similar to asbestos but more research is needed to determine whether they, or other nanofibres, can cause fibrosis and cancer in the long term.     

Hypouricemic effects of novel concentrative nucleoside transporter 2 inhibitors through suppressing intestinal absorption of purine nucleosides

We have developed concentrative nucleoside transporter 2 (CNT2) inhibitors as a novel pharmacological approach for improving hyperuricemia by inhibiting intestinal absorption of purines. Dietary purine nucleosides are absorbed in the small intestines by CNTs expressed in the apical membrane. In humans, the absorbed purine nucleosides are rapidly degraded to their final end product, uric acid, by xanthine oxidase. Based on the expression profile of human CNTs in digestive tract tissues, we established a working hypothesis that mainly CNT2 contributes to the intestinal absorption of purine nucleosides. In order to confirm this possibility, we developed CNT2 inhibitors and found that (2R,3R,4S,5R)-2-(6-amino-8-{[3'-(3-aminopropoxy)-biphenyl-4-ylmethyl]-amino}-9H-purin-9-yl)-5-hydroxymethyl-tetrahydrofuran-3,4-diol (KGO-2142) and 1-[3-(5-{[1-((2R,3R,4S,5R)-3,4-dihydroxy-5-hydroxymethyl-tetrahydrofuran-2-yl)-1H-benzimidazol-2-ylamino]-methyl}-2-ethoxyphenoxy)-propyl]-piperidine-4-carboxylic acid amide (KGO-2173) were inhibitory. These CNT2 inhibitors had potent inhibitory activity against inosine uptake via human CNT2, but they did not potently interfere with nucleoside uptake via human CNT1, CNT3 or equilibrative nucleoside transporters (ENTs) in vitro. After oral administration of KGO-2173 along with [(14)C]-inosine, KGO-2173 significantly decreased the urinary excretion of radioactivity at 6 and 24h in rats. Since dietary purine nucleosides are not utilized in the body and are excreted into the urine rapidly, this decrease in radioactivity in the urine represented the inhibitory activity of KGO-2173 toward the absorption of [(14)C]-inosine in the small intestines. KGO-2142 almost completely inhibited dietary RNA-induced hyperuricemia and the increase in urinary excretion of uric acid in cebus monkeys. These novel CNT2 inhibitors, KGO-2142 and KGO-2173, could be useful therapeutic options for the treatment of hyperuricemia.     

Polymeric radiotracers in nuclear imaging

Water-soluble polymers have been used in the last two decades to modify the pharmacokinetics and physicochemical properties of targeted therapeutic agents. Non-invasive imaging techniques such as nuclear imaging can be used to assess the drug delivery efficiency of novel formulations in a cost-effective fashion and thereby facilitate their development process. Polymeric radiopharmaceuticals have also been investigated on their own right as potential nuclear imaging agents. Clinical applications of polymeric radiopharmaceuticals include blood-pool imaging and targeted molecular imaging. In the latter case, water-soluble polymers are often used to modify the pharmacokinetics and biodistribution pattern of ligands that target receptors or antigens at disease sites. As advances are continue to be made in the emerging field of molecular imaging, nuclear imaging will play an increasingly important role in the development of polymeric drug delivery systems. Similarly, polymer technology will also be integrated into the development of molecularly targeted radiopharmaceuticals. Here, we review various aspects of polymeric radiotracers and their applications in nuclear imaging.     

A stochastic model of carbon nanotube deposition in the airways and alveoli of the human respiratory tract

Context: In the past two decades, possible exposure of workers to nanoparticles has excited the attention of occupational medicine, resulting in the conception of related risk assessments. Although most nanoparticles have been categorized as hazardous substances in the meantime, their behavior in the human respiratory tract still bears some enigmas, which require clarification.

Objectives: The study pursues the goal to provide detailed theoretical lung deposition data of carbon nanotubes (CNT) with various diameters and lengths. Besides a quantification of total and regional deposition, also airway generation-specific deposition has been subjected to the modeling process.

Methods: Theoretical approach of CNT deposition in the human lungs has been conducted by assuming a stochastic structure of the bronchial network, within which particle transport takes place along randomly selected paths. Fluid-dynamic particle characteristics have been simulated by application of a rigid fiber model, which considers diverse forces and torques acting on the particles during their translocation within the inhaled air. Particle deposition in the entire lungs has been approximated by using the aerodynamic/thermodynamic diameter concept and related empirical deposition formulae.

Results: Theoretical deposition data reflect a significant dependence of CNT deposition on (a) the effective size of the particles and (b) the conditions, under which they are taken up into the respiratory tract. Extremely small CNT (～1?nm) are primarily filtered in the extrathoracic airways, intermediately sized CNT (～10?nm) exhibit a preference to deposit in the alveoli, and large CNT (～100?nm) are marked by minimum deposition.

Conclusion: Pulmonary deposition of CNT is subject to a partly remarkable variation. According to the model of this study, particles of intermediate size seem to bear highest potential to act as hazardous substances.     

Evaluation of domain of unknown function 1220 (DUF1220) for detection of human genome by quantitative polymerase chain reaction: Potential use in assessing the biodistribution of transplanted therapeutic human cells

The biodistribution profile of cell-based therapy products in animal models is important for evaluation of their safety and efficacy. Because of its quantitative nature and sensitivity, the quantitative polymerase chain reaction (qPCR) is a useful method for detecting and quantifying xenogeneic cell-derived DNA in animal models, thereby allowing a biodistribution profile to be established. Although the restriction endonuclease family from Arthrobacter luteus (Alu) of repetitive elements in human genome sequences has been used to assess the biodistribution of human cells, high background signals are detected. In the present study, we evaluate the potential of domain of unknown function 1220 (DUF1220), which is a human lineage-specific, multiple-copy gene similar to Alu sequences, for such analysis. Using qPCR analysis for DUF1220, human genome could be detected against a mouse genome background at a level comparable to that of Alu sequences with no detectable background signals. Moreover, using this approach, the human genome could be distinguished from the cynomolgus monkey genome. Further investigation of the quantitative aspects of this DUF1220-based qPCR assay might prove its usefulness for biodistribution studies of human cells transplanted into animals in the future.