Unravelling the toxicity of carbon nanomaterials - From cellular interactions to mechanistic understanding.

The application of carbon nanomaterials in diverse fields has substantially increased their demand for commercial usage. Within the earliest decade, the development of functional materials has further increased the significance of this element. Despite the advancements recorded, the potential harmful impacts of embracing carbon nanomaterials for biological applications must be balanced against their advantages. Interestingly, many studies have neglected the intriguing and dynamic cellular interaction of carbon nanomaterials and the mechanistic understanding of their property-driven behaviour, even though common toxicity profiles have been reported. Reiterating the toxicity issue, several researchers conclude that these materials have minimal toxicity and may be safe for contact with biological systems at certain dosages. Here, we aim to provide a report on the significance of some of the properties that influence their toxicity. After that, a description of the implication of nanotoxicology in humans and living systems, revealing piece by piece their exposure routes and possible risks, will be provided. Then, an extensive discussion of the mechanistic puzzle modulating the interface between various human cellular systems and carbon nanomaterials such as carbon nanotubes, carbon dots, graphene, fullerenes, and nanodiamonds will follow. Finally, this review also sheds light on the organization that handles the risk associated with nanomaterials.

Immunotoxicity of engineered nanomaterials and their role in asthma.

The toxicity of engineered nanomaterials (ENMs) in vivo and in vitro has formed the basis of most studies. However, the toxicity of ENMs, particularly on the immune system, i.e. immunotoxicity, and their role in manipulating it, are less known. This review addresses the initiation or exacerbation as well as the attenuation of allergic asthma by a variety of ENMs and how they may be used in drug delivery to enhance the treatment of asthma. This review also highlights a few research gaps in the study of the immunotoxicity of ENMs, for example, the potential drawbacks of assays used in immunotoxicity assays; the potential role of hormesis during dosing of ENMs; and the variables that result in discrepancies among different studies, such as the physicochemical properties of ENMs, differences in asthmatic animal models, and different routes of administration.

Exposure Assessment of Silver and Gold Nanoparticles Generated During the Synthesis Process in a South African Research Laboratory.

During the synthesis of engineered nanomaterials (ENMs), various occupational exposures occur, leading to health consequences. To date, there is paucity of studies focused on modeling the deposition of nanoparticles emitted from ENMs synthesis processes. This study aimed to characterise and assess exposure to gold (AuNPs) and silver nanoparticles (AgNPs) during a synthesis process in a research laboratory in South Africa. AuNPs and AgNPs synthesis processes were monitored for an hour in a laboratory using a Scanning Mobility Particle Sizer. The monitoring was conducted at a height of 1.2-1.5 m (m) and 1.5 m away from the hood, assuming a 30 cm (cm) breathing circumference zone. Each synthesis process was monitored thrice to generate reliable point estimates, which were used to assess exposure over 8 hours. A time-weighted average concentration was calculated and compared to the derived 8-h occupational exposure limit (OEL) for AgNPs (0.19 µg/m3) and the proposed provisional nano reference value for AuNPs (20,000 particles/cm3). The Multiple-Path Particle Dosimetry model was used to calculate the deposition and retention of both AuNPs and AgNPs. NPs emitted during the synthesis process were dominant in the nuclei (79% for AuNPs and 54% for AgNPs), followed by the Aitken (12% for AuNPs and 29% for AgNPs), with fewer particles in the accumulation mode (9.2% for AuNPs and 17% for AgNPs). AuNPs and AgNPs generated during the synthesis process were determined at 1617.3 ± 102 cm3 (0.046 µg/m3) and 2,687 cm3 ± 620 (0.077 µg/m3), respectively. For the three exposure scenarios, none exceeded the occupational exposure limit for both AuNPs (provisional) and AgNPs (OEL). Workers in the synthesis laboratory are exposed to a concentration below the recommended occupational exposure limit for silver and the proposed provisional nano reference value for gold. Although, the concentrations to which laboratory workers are exposed to are below safe levels, the assessment of the lung deposition patterns indicate a high particle lung retention which raise concerns about long term safety of workers.

Importance of Surface Topography in Both Biological Activity and Catalysis of Nanomaterials: Can Catalysis by Design Guide Safe by Design?

It is acknowledged that the physicochemical properties of nanomaterials (NMs) have an impact on their toxicity and, eventually, their pathogenicity. These properties may include the NMs' surface chemical composition, size, shape, surface charge, surface area, and surface coating with ligands (which can carry different functional groups as well as proteins). Nanotopography, defined as the specific surface features at the nanoscopic scale, is not widely acknowledged as an important physicochemical property. It is known that the size and shape of NMs determine their nanotopography which, in turn, determines their surface area and their active sites. Nanotopography may also influence the extent of dissolution of NMs and their ability to adsorb atoms and molecules such as proteins. Consequently, the surface atoms (due to their nanotopography) can influence the orientation of proteins as well as their denaturation. However, although it is of great importance, the role of surface topography (nanotopography) in nanotoxicity is not much considered. Many of the issues that relate to nanotopography have much in common with the fundamental principles underlying classic catalysis. Although these were developed over many decades, there have been recent important and remarkable improvements in the development and study of catalysts. These have been brought about by new techniques that have allowed for study at the nanoscopic scale. Furthermore, the issue of quantum confinement by nanosized particles is now seen as an important issue in studying nanoparticles (NPs). In catalysis, the manipulation of a surface to create active surface sites that enhance interactions with external molecules and atoms has much in common with the interaction of NP surfaces with proteins, viruses, and bacteria with the same active surface sites of NMs. By reviewing the role that surface nanotopography plays in defining many of the NMs' surface properties, it reveals the need for its consideration as an important physicochemical property in descriptive and predictive toxicology. Through the manipulation of surface topography, and by using principles developed in catalysis, it may also be possible to make safe-by-design NMs with a reduction of the surface properties which contribute to their toxicity.

Intracellular and extracellular targets as mechanisms of cancer therapy by nanomaterials in relation to their physicochemical properties.

Cancer nanomedicine has evolved in recent years and is only expected to increase due to the ease with which nanomaterials (NMs) may be manipulated to the advantage of the cancer patient. The success of nanomedicine is dependent on the cell death mechanism, which in turn is dependent on the organelle initially targeted. The success of cancer nanomedicine is also dependent on other cellular mechanisms such as the induction of autophagy dysfunction, manipulation of the tumor microenvironment (TME) and secretome or induction of host immune responses. Current cancer phototherapies for example, photothermal- or photodynamic therapies as well as radio enhancement also form a major part of cancer nanomedicine. In general, cancer nanomedicine may be grouped into those NMs exhibiting inherent anti-cancer properties that is, self-therapeutic NMs (Group 1), NMs leading to localization of phototherapies or radio-enhancement (Group 2), and NMs as nanocarriers in the absence or presence of external radiation (Group 3). The recent advances of these three groups, together with their advantages and disadvantages as well as their cellular mechanisms and ultimate outcomes are summarized in this review. By exploiting these different intracellular mechanisms involved in initiating cell death pathways, it is possible to synthesize NMs that may have the desirable characteristics to maximize their efficacy in cancer therapy. Therefore, a summary of these important physicochemical characteristics is also presented that need to be considered for optimal cancer cell targeting and initiation of mechanisms that will lead to cancerous cell death. This article is categorized under: Therapeutic Approaches and Drug Discovery > Nanomedicine for Oncologic Disease Toxicology and Regulatory Issues in Nanomedicine > Toxicology of Nanomaterials Toxicology and Regulatory Issues in Nanomedicine > Regulatory and Policy Issues in Nanomedicine.

The toxicity of respirable South African mine tailings dust in relation to their physicochemical properties.

INTRODUCTION: Decades of mining in South Africa has given rise to hundreds of tailings storage facilities (TSFs) and several tonnes of waste. These TSFs have contributed to air pollution due to the lack of proper rehabilitation measures. Currently, it is not known whether tailings emissions could be the cause of respiratory-related ill effects. In addition, the physicochemical properties that may govern their toxicity have not yet been identified. AIM: The aim of this research was to determine the toxicity of tailings dust and identify the physicochemical properties likely to govern toxicity. METHODS: Dust samples were collected from five TSFs in the Gauteng and North West Provinces of South Africa and sieved to enrich the airborne particle fraction more likely to be inhaled. Thereafter, their physicochemical characteristics were assessed i.e. size distribution, specific surface area, shape, surface elemental composition, mineral composition, total elemental composition and surface activity. In addition, the toxicity and cellular internalization of the particles were assessed using the BEAS-2B epithelial and U937 monocytic-macrophage cell lines.Results: The results showed that all tailings dusts showed toxicity, particularly in the BEAS-2B cell line. This toxicity could have been governed by either their elemental composition, e.g. high transition elements e.g. Fe, Cu, Cr and V in the dusts from TSF 4, or a combination of other physicochemical properties, e.g. higher quartz content, lower size and higher surface area in the dusts from TSF 1. CONCLUSION: These results provide mechanistic evidence to support future epidemiological studies attempting to link tailings dust exposure to adverse health effects.

Interference: A Much-Neglected Aspect in High-Throughput Screening of Nanoparticles.

Despite several studies addressing nanoparticle (NP) interference with conventional toxicity assay systems, it appears that researchers still rely heavily on these assays, particularly for high-throughput screening (HTS) applications in order to generate "big" data for predictive toxicity approaches. Moreover, researchers often overlook investigating the different types of interference mechanisms as the type is evidently dependent on the type of assay system implemented. The approaches implemented in the literature appear to be not adequate as it often addresses only one type of interference mechanism with the exclusion of others. For example, interference of NPs that have entered cells would require intracellular assessment of their interference with fluorescent dyes, which has so far been neglected. The present study investigated the mechanisms of interference of gold NPs and silver NPs in assay systems implemented in HTS including optical interference as well as adsorption or catalysis. The conventional assays selected cover all optical read-out systems, that is, absorbance (XTT toxicity assay), fluorescence (CytoTox-ONE Homogeneous membrane integrity assay), and luminescence (CellTiter Glo luminescent assay). Furthermore, this study demonstrated NP quenching of fluorescent dyes also used in HTS (2',7'-dichlorofluorescein, propidium iodide, and 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethyl-benzamidazolocarbocyanin iodide). To conclude, NP interference is, as such, not a novel concept, however, ignoring this aspect in HTS may jeopardize attempts in predictive toxicology. It should be mandatory to report the assessment of all mechanisms of interference within HTS, as well as to confirm results with label-free methodologies to ensure reliable big data generation for predictive toxicology.

Exceedance of environmental exposure limits to crystalline silica in communities surrounding gold mine tailings storage facilities in South Africa.

BACKGROUND: Unlike occupational silica exposures, the association between non-occupational silica exposures and adverse health effects is not well researched, despite its occurrence in communities close to dust-generating sources such as tailings storage facilities (TSFs). Recent studies have shown that communities surrounding TSFs in South Africa often complain about the onset of dust-related health effects. Even though international interim non-occupational crystalline silica limits have been established, South Africa is yet to enforce its own limits for residential areas close to TSF sites. OBJECTIVE: The objective of the study was to assess the need to enforce non-occupational crystalline silica limits for South Africa. METHODS: The methods involved (1) Quantifying the silica polymorph content in bulk dust collected from TSFs in South Africa; (2) Assessing the possibility of the dust to reach surrounding communities through ambient and personal filter sampling and; (3) Conducting risk characterisation for both cancer and non-cancer endpoints. RESULTS: All bulk dust samples consisted mostly of crystalline silica (73.14-87.09%). Large percentages of nanoparticles were identified in all bulk samples (66.8-70.7%) indicating the possibility of the dust to lodge deep within the lungs. The crystalline silica levels obtained from ambient PM10 sampling and personal PM4 sampling all exceeded the international crystalline silica interim limits and reached maximum levels of 90 and 50.9µg·m-3, respectively. For three TSFs, sampling sites close to the TSFs showed higher PM4 silica levels compared to sites further from TSFs. Risk characterisation revealed the possibility of cancer and non-cancer health effects when chronically exposed to silica levels recorded during the study. CONCLUSION: The results indicate unacceptable crystalline silica exposures in surrounding communities and the need for enforcement of an ambient silica limit for South Africa.

In Search of a Converging Cellular Mechanism in Nanotoxicology and Nanomedicine in the Treatment of Cancer.

Multiple applications of nanomaterials have raised concern with regard to their toxicity. With increasing research into nanomaterial safety, mechanisms involved in the toxic effects of nanomaterials have begun to emerge. The importance of nanomaterial-induced lysosomal membrane permeabilization through overloading or direct damage of the lysosomal compartment, resulting in the blockade of autophagosome-lysosome fusion and autophagy dysfunction, as well as inflammasome activation were cited as emerging mechanisms of nanomaterial toxicity. It has recently been proposed that these very mechanisms leading to nanomaterial toxicity may be utilized in nanotherapeutics. This review discusses these nanomaterial-induced mechanisms in detail and how it has been exploited in cancer research. This review also addresses certain considerations that need to be kept in mind when using nanomaterials in therapeutics.

Systematic Review of Screening and Surveillance Programs to Protect Workers from Nanomaterials.

BACKGROUND: Screening and surveillance approaches for workers exposed to nanomaterials could aid in early detection of health effects, provide data for epidemiological studies and inform action to decrease exposure. The aim of this review is to identify such screening and surveillance approaches, in order to extract available data regarding (i) the studies that have successfully been implemented in present day, (ii) identification of the most common and/or toxic nano-related health hazards for workers and (iii) possible exposure surveillance markers. This review contributes to the current understanding of the risk associated with nanomaterials by determining the knowledge gap and making recommendations based on current findings. METHODS: A systematic review was conducted. PubMed and Embase were searched to identify articles reporting on any surveillance-related study that described both exposure to nanomaterials and the health indicators that were measured. Four reviewers worked in pairs to independently assess the eligibility of studies and risk of bias before extraction of data. Studies were categorised according to the type of study and the medical surveillance performed, which included the type of nanomaterial, any exposure details provided, as well as health indicators and biomarkers tested. RESULTS: Initially 92 studies were identified, from which 84 full texts were assessed for eligibility. Seven studies met all the inclusion criteria, i.e. those performed in Taiwan, Korea, Czech Republic and the US. Of these, six compared health indicators between exposed and unexposed workers and one study described a surveillance program. All studies were at a high risk of bias. Workers were exposed to a mix of nanomaterials in three studies, carbon-based nanomaterials in two studies, nano-silver in one study and nano-titanium oxide in the other study. Two studies did not find a difference in biomarkers between exposed and unexposed workers. In addition, differences in early effects on pulmonary function or neurobehavioral tests were not observed. One study found an increased prevalence of allergic dermatitis and "sneezing" in the exposed group. CONCLUSIONS: This review of recently published data on surveillance studies proves that there is a gap in the current knowledge, where most of the surveillance-related studies reported do not follow a set format that provides the required information on ENM characterisation, the type of exposure and the measured indicators/biomarkers. Hence, there is very low quality evidence that screening and surveillance might detect adverse health effects associated with workplace exposure. This systematic review is relevant because it proves that, although surveillance programs have been initiated and preliminary results are being published, the current studies are actually not answering the important questions or solving the overall problem regarding what the potential health hazards are among workers either handling or potentially exposed to ENMs. The recommendations, thus proposed, are based on an obvious need for (i) exposure registries, where longitudinal follow-up studies should inform surveillance, (ii) known exposure measurements or summary indices for ENMs as a reference (iii) validation of candidate biomarkers and (iv) studies that compare the effects of these surveillance approaches to usual care, e.g. those commonly followed for bulk-size hazardous materials.

Gold nanoparticle interference study during the isolation, quantification, purity and integrity analysis of RNA.

Investigations have been conducted regarding the interference of nanoparticles (NPs) with different toxicological assay systems, but there is a lack of validation when conducting routine tests for nucleic acid isolation, quantification, integrity, and purity analyses. The interference of citrate-capped gold nanoparticles (AuNPs) was investigated herein. The AuNPs were added to either BEAS-2B bronchial human cells for 24 h, the isolated pure RNA, or added during the isolation procedure, and the resultant interaction was assessed. Total RNA that was isolated from untreated BEAS-2B cells was spiked with various concentrations (v/v%) of AuNPs and quantified. A decrease in the absorbance spectrum (220-340 nm) was observed in a concentration-dependent manner. The 260 and 280 nm absorbance ratios that traditionally infer RNA purity were also altered. Electrophoresis was performed to determine RNA integrity, but could not differentiate between AuNP-exposed samples. However, the spiked post-isolation samples did produce differences in spectra (190-220 nm), where shifts were observed at a shorter wavelength. These shifts could be due to alterations to chromophores found in nucleic acids. The co-isolation samples, spiked with 100 µL AuNP during the isolation procedure, displayed a peak shift to a longer wavelength and were similar to the results obtained from a 24 h AuNP treatment, under non-cytotoxic test conditions. Moreover, hyperspectral imaging using CytoViva dark field microscopy did not detect AuNP spectral signatures in the RNA isolated from treated cells. However, despite the lack of AuNPs in the final RNA product, structural changes in RNA could still be observed between 190-220 nm. Consequently, full spectral analyses should replace the traditional ratios based on readings at 230, 260, and 280 nm. These are critical points of analyses, validation, and optimization for RNA-based techniques used to assess AuNPs effects.

Label-free in vitro toxicity and uptake assessment of citrate stabilised gold nanoparticles in three cell lines.

BACKGROUND: Reliable in vitro toxicity testing is needed prior to the commencement of in vivo testing necessary for hazard identification and risk assessment of nanoparticles. In this study, the cytotoxicity and uptake of 14 nm and 20 nm citrate stabilised gold nanoparticles (AuNPs) in the bronchial epithelial cell line BEAS-2B, the Chinese hamster ovary cell line CHO, and the human embryonic kidney cell line HEK 293 were investigated. METHODS: Cytotoxicity of the AuNPs was assessed via traditional XTT-, LDH-, and ATP-based assays, followed by cell impedance studies. Dark-field imaging and hyperspectral imaging were used to confirm the uptake of AuNPs into the cells. RESULTS: Interference of the AuNPs with the XTT- and ATP-based assays was overcome through the use of cell impedance technology. AuNPs were shown to be relatively non-toxic using this methodology; nevertheless CHO cells were the most sensitive cell type with 20 nm AuNPs having the highest toxicity. Uptake of both 14 nm and 20 nm AuNPs was observed in all cell lines in a time- and cell type-dependent manner. CONCLUSIONS: Using the cell impedance and dark-field hyperspectral imaging technologies, it was possible to study the toxicity of AuNPs in different cell lines and show that these cells could internalize AuNPs with their subsequent intracellular aggregation. It was also possible to show that this toxicity would not correlate with the level of uptake but it would correlate with cell-type and the size of the AuNPs. Therefore, these two label-free methodologies used in this study are suitable for in vitro studies on the effects of AuNPs, and could present themselves as appropriate and valuable methodologies for future nanoparticle toxicity and uptake studies.

Vitamin D receptor gene methylation is associated with ethnicity, tuberculosis, and TaqI polymorphism.

The Vitamin D receptor (VDR) gene encodes a transcription factor which, on activation by vitamin D, modulates diverse biologic processes, including calcium homeostasis and immune function. Genetic variation involving VDR shows striking differences in allele frequency between populations and has been associated with disease susceptibility, including tuberculosis and autoimmunity, although results have often been conflicting. We hypothesized that methylation of VDR may be population specific and that the combination of differential methylation and genetic variation may characterize tuberculosis (TB) predisposition. We use bisulfite conversion and/or pyrosequencing to analyze the methylation status of 17 CpGs of VDR and to genotype 7 SNPs in the 3' CpG Island (CpG island [CGI] 1060), including the commonly studied SNPs ApaI (rs7975232) and TaqI (rs731236). We show that, for lymphoblastoid cell lines from two ethnically diverse populations (Yoruba from HapMap, n = 30 and Caucasians, n = 30) together with TB cases (n = 32) and controls (n = 29) from the Venda population of South Africa, there are methylation variable positions in the 3' end that significantly distinguish ethnicity (9/17 CpGs) and TB status (3/17 CpGs). Moreover, methylation status shows complex association with TaqI genotype highlighting the need to consider both genetic and epigenetic variants in genetic studies of VDR association with disease.