A review of pulmonary neutrophilia and insights into the key role of neutrophils in particle-induced pathogenesis in the lung from animal studies of lunar dusts and other poorly soluble dust particles.

The mechanisms of particle-induced pathogenesis in the lung remain poorly understood. Neutrophilic inflammation and oxidative stress in the lung are hallmarks of toxicity. Some investigators have postulated that oxidative stress from particle surface reactive oxygen species (psROS) on the dust produces the toxicopathology in the lungs of dust-exposed animals. This postulate was tested concurrently with the studies to elucidate the toxicity of lunar dust (LD), which is believed to contain psROS due to high-speed micrometeoroid bombardment that fractured and pulverized lunar surface regolith. Results from studies of rats intratracheally instilled (ITI) with three LDs (prepared from an Apollo-14 lunar regolith), which differed 14-fold in levels of psROS, and two toxicity reference dusts (TiO2 and quartz) indicated that psROS had no significant contribution to the dusts' toxicity in the lung. Reported here are results of further investigations by the LD toxicity study team on the toxicological role of oxidants in alveolar neutrophils that were harvested from rats in the 5-dust ITI study and from rats that were exposed to airborne LD for 4 weeks. The oxidants per neutrophils and all neutrophils increased with dose, exposure time and dust's cytotoxicity. The results suggest that alveolar neutrophils play a critical role in particle-induced injury and toxicity in the lung of dust-exposed animals. Based on these results, we propose an adverse outcome pathway (AOP) for particle-associated lung disease that centers on the crucial role of alveolar neutrophil-derived oxidant species. A critical review of the toxicology literature on particle exposure and lung disease further supports a neutrophil-centric mechanism in the pathogenesis of lung disease and may explain previously reported animal species differences in responses to poorly soluble particles. Key findings from the toxicology literature indicate that (1) after exposures to the same dust at the same amount, rats have more alveolar neutrophils than hamsters; hamsters clear more particles from their lungs, consequently contributing to fewer neutrophils and less severe lung lesions; (2) rats exposed to nano-sized TiO2 have more neutrophils and more severe lesions in their lungs than rats exposed to the same mass-concentration of micron-sized TiO2; nano-sized dust has a greater number of particles and a larger total particle-cell contact surface area than the same mass of micron-sized dust, which triggers more alveolar epithelial cells (AECs) to synthesize and release more cytokines that recruit a greater number of neutrophils leading to more severe lesions. Thus, we postulate that, during chronic dust exposure, particle-inflicted AECs persistently release cytokines, which recruit neutrophils and activate them to produce oxidants resulting in a prolonged continuous source of endogenous oxidative stress that leads to lung toxicity. This neutrophil-driven lung pathogenesis explains why dust exposure induces more severe lesions in rats than hamsters; why, on a mass-dose basis, nano-sized dusts are more toxic than the micron-sized dusts; why lung lesions progress with time; and why dose-response curves of particle toxicity exhibit a hockey stick like shape with a threshold. The neutrophil centric AOP for particle-induced lung disease has implications for risk assessment of human exposures to dust particles and environmental particulate matter.

A tiered approach to investigate the inhalation toxicity of cobalt substances. Tier 4: Effects from a 28-day inhalation toxicity study with tricobalt tetraoxide in rats.

Lung cancer following inhalation in rodents is a major concern regarding exposure to cobalt substances. However, little information is available on adverse effects and toxicity following long-term inhalation exposure to poorly soluble cobalt substances with low bioavailability. Thus, the present study focused on pulmonary effects of the poorly soluble tricobalt tetraoxide (5, 20, 80 mg/m³) in a 28-day inhalation exposure study. Lung weights increased with increasing exposures. Bronchoalveolar lavage fluid analysis and histopathology revealed lung tissue inflammation at the mid-dose with increasing severity in the high-dose group and post-exposure persistency. Markers for cellular damage and cell proliferation were statistically significantly increased. No increase in 8-OH-dG lesions was observed, indicating an absence of oxidative DNA lesions. The primary effect of inhaled Co3O4 particles is inflammation of the respiratory tract strongly resembling responses of inhaled "inert dust" substances, with a NOAEC of 5 mg/m³ under the conditions of this test.

Risk Governance of Emerging Technologies Demonstrated in Terms of its Applicability to Nanomaterials.

Nanotechnologies have reached maturity and market penetration that require nano-specific changes in legislation and harmonization among legislation domains, such as the amendments to REACH for nanomaterials (NMs) which came into force in 2020. Thus, an assessment of the components and regulatory boundaries of NMs risk governance is timely, alongside related methods and tools, as part of the global efforts to optimise nanosafety and integrate it into product design processes, via Safe(r)-by-Design (SbD) concepts. This paper provides an overview of the state-of-the-art regarding risk governance of NMs and lays out the theoretical basis for the development and implementation of an effective, trustworthy and transparent risk governance framework for NMs. The proposed framework enables continuous integration of the evolving state of the science, leverages best practice from contiguous disciplines and facilitates responsive re-thinking of nanosafety governance to meet future needs. To achieve and operationalise such framework, a science-based Risk Governance Council (RGC) for NMs is being developed. The framework will provide a toolkit for independent NMs' risk governance and integrates needs and views of stakeholders. An extension of this framework to relevant advanced materials and emerging technologies is also envisaged, in view of future foundations of risk research in Europe and globally.

Comment on Saber et al. (2019), "Commentary: the chronic inhalation study in rats for assessing lung cancer risk may be better than its reputation".

In their Commentary Saber et al. (Part Fibre Toxicol 16: 44, 2019) argue that chronic inhalation studies in rats can be used for assessing the lung cancer risk of insoluble nanomaterials. The authors make several significant errors in their interpretation and representation of the underlying science. In this Letter to the Editor we discuss these inaccuracies to correct the scientific record. When the science is recounted accurately it does not support Saber et al's statements and conclusions.

Toxicity testing of poorly soluble particles, lung overload and lung cancer.

In 2013, an ECETOC Task Force evaluated scientific understanding of the 'lung overload' hypothesis. As there is no evidence that humans develop lung tumours following exposure to poorly soluble particles (PSPs), emphasis was given to the observed higher sensitivity and specificity of rat lung responses and potential impacts of this on human risk assessment. Key arguments and outcomes are summarised here, together with discussion of additional findings published since 2013. Inhalation exposure to PSPs in all species is associated with localised pulmonary toxicity initiated by a persistent pro-inflammatory response to particle deposition. Events in the rat indicate a plausible adverse outcome pathway for lung tumour development following exposure to PSPs under overload conditions. A different particle lung translocation pattern compared to rats make humans less sensitive to developing comparable lung overload conditions and appears to also preclude tumour formation, even under severe and prolonged exposure conditions. Evidence continues to suggest that the rat lung model is unreliable as a predictor for human lung cancer risk. However, it is a sensitive model for detecting various thresholded inflammatory markers, with utility for non-neoplastic risk assessment purposes. It is noteworthy that preventing inflammatory rat lung responses will also inhibit development of neoplastic outcomes.

Case studies putting the decision-making framework for the grouping and testing of nanomaterials (DF4nanoGrouping) into practice.

Case studies covering carbonaceous nanomaterials, metal oxide and metal sulphate nanomaterials, amorphous silica and organic pigments were performed to assess the Decision-making framework for the grouping and testing of nanomaterials (DF4nanoGrouping). The usefulness of the DF4nanoGrouping for nanomaterial hazard assessment was confirmed. In two tiers that rely exclusively on non-animal test methods followed by a third tier, if necessary, in which data from rat short-term inhalation studies are evaluated, nanomaterials are assigned to one of four main groups (MGs). The DF4nanoGrouping proved efficient in sorting out nanomaterials that could undergo hazard assessment without further testing. These are soluble nanomaterials (MG1) whose further hazard assessment should rely on read-across to the dissolved materials, high aspect-ratio nanomaterials (MG2) which could be assessed according to their potential fibre toxicity and passive nanomaterials (MG3) that only elicit effects under pulmonary overload conditions. Thereby, the DF4nanoGrouping allows identifying active nanomaterials (MG4) that merit in-depth investigations, and it provides a solid rationale for their sub-grouping to specify the further information needs. Finally, the evaluated case study materials may be used as source nanomaterials in future read-across applications. Overall, the DF4nanoGrouping is a hazard assessment strategy that strictly uses animals as a last resort.

Applied Nanotoxicology.

Nanomaterials, including nanoparticles and nanoobjects, are being incorporated into everyday products at an increasing rate. These products include consumer products of interest to toxicologists such as pharmaceuticals, cosmetics, food, food packaging, household products, and so on. The manufacturing of products containing or utilizing nanomaterials in their composition may also present potential toxicologic concerns in the workplace. The molecular complexity and composition of these nanomaterials are ever increasing, and the means and methods being applied to characterize and perform useful toxicologic assessments are rapidly advancing. This article includes presentations by experienced toxicologists in the nanotoxicology community who are focused on the applied aspect of the discipline toward supporting state of the art toxicologic assessments for food products and packaging, pharmaceuticals and medical devices, inhaled nanoparticle and gastrointestinal exposures, and addressing occupational safety and health issues and concerns. This symposium overview article summarizes 5 talks that were presented at the 35th Annual meeting of the American College of Toxicology on the subject of "Applied Nanotechnology."

A decision-making framework for the grouping and testing of nanomaterials (DF4nanoGrouping).

The European Centre for Ecotoxicology and Toxicology of Chemicals (ECETOC) 'Nano Task Force' proposes a Decision-making framework for the grouping and testing of nanomaterials (DF4nanoGrouping) that consists of 3 tiers to assign nanomaterials to 4 main groups, to perform sub-grouping within the main groups and to determine and refine specific information needs. The DF4nanoGrouping covers all relevant aspects of a nanomaterial's life cycle and biological pathways, i.e. intrinsic material and system-dependent properties, biopersistence, uptake and biodistribution, cellular and apical toxic effects. Use (including manufacture), release and route of exposure are applied as 'qualifiers' within the DF4nanoGrouping to determine if, e.g. nanomaterials cannot be released from a product matrix, which may justify the waiving of testing. The four main groups encompass (1) soluble nanomaterials, (2) biopersistent high aspect ratio nanomaterials, (3) passive nanomaterials, and (4) active nanomaterials. The DF4nanoGrouping aims to group nanomaterials by their specific mode-of-action that results in an apical toxic effect. This is eventually directed by a nanomaterial's intrinsic properties. However, since the exact correlation of intrinsic material properties and apical toxic effect is not yet established, the DF4nanoGrouping uses the 'functionality' of nanomaterials for grouping rather than relying on intrinsic material properties alone. Such functionalities include system-dependent material properties (such as dissolution rate in biologically relevant media), bio-physical interactions, in vitro effects and release and exposure. The DF4nanoGrouping is a hazard and risk assessment tool that applies modern toxicology and contributes to the sustainable development of nanotechnological products. It ensures that no studies are performed that do not provide crucial data and therefore saves animals and resources.

How meaningful are risk determinations in the absence of a complete dataset? Making the case for publishing standardized test guideline and 'no effect' studies for evaluating the safety of nanoparticulates versus spurious 'high effect' results from single investigative studies.

A recent review article critically assessed the effectiveness of published research articles in nanotoxicology to meaningfully address health and safety issues for workers and consumers. The main conclusions were that, based on a number of flaws in study designs, the potential risk from exposures to nanomaterials is highly exaggerated, and that no 'nano-specific' adverse effects, different from exposures to bulk particles, have been convincingly demonstrated. In this brief editorial we focus on a related tangential issue which potentially compromises the integrity of basic risk science. We note that some single investigation studies report specious toxicity findings, which make the conclusions more alarming and attractive and publication worthy. In contrast, the standardized, carefully conducted, 'guideline study results' are often ignored because they can frequently report no adverse effects; and as a consequence are not considered as novel findings for publication purposes, and therefore they are never considered as newsworthy in the popular press. Yet it is the Organization for Economic Cooperation and Development (OECD) type test guideline studies that are the most reliable for conducting risk assessments. To contrast these styles and approaches, we present the results of a single study which reports high toxicological effects in rats following low-dose, short-term oral exposures to nanoscale titanium dioxide particles concomitant with selective investigative analyses. Alternatively, the findings of OECD test guideline 408, standardized guideline oral toxicity studies conducted for 90 days at much higher doses (1000 mg kg-1) in male and female rats demonstrated no adverse effects following a very thorough and complete clinical chemical, as well as histopathological evaluation of all of the relevant organs in the body. This discrepancy in study findings is not reconciled by the fact that several biokinetic studies in rats and humans demonstrate little or no uptake of nanoscale or pigment-grade TiO2 particles following oral exposures. We conclude that to develop a competent risk assessment profile, results derived from standardized, guideline-type studies, and even 'no effect' study findings provide critically useful input for assessing safe levels of exposure; and should, in principle, be readily acceptable for publication in peer-reviewed toxicology journals. This is a necessary prerequisite for developing a complete dataset for risk assessment determinations.

A critical appraisal of existing concepts for the grouping of nanomaterials.

The grouping of substances serves to streamline testing for regulatory purposes. General grouping approaches for chemicals have been implemented in, e.g., the EU chemicals regulation. While specific regulatory frameworks for the grouping of nanomaterials are unavailable, this topic is addressed in different publications, and preliminary guidance is provided in the context of substance-related legislation or the occupational setting. The European Centre for Ecotoxicology and Toxicology of Chemicals Task Force on the Grouping of Nanomaterials reviewed available concepts for the grouping of nanomaterials for human health risk assessment. In their broad conceptual design, the evaluated approaches are consistent or complement each other. All go beyond the determination of mere structure-activity relationships and are founded on different aspects of the nanomaterial life cycle. These include the NM's material properties and biophysical interactions, specific types of use and exposure, uptake and kinetics, and possible early and apical biological effects. None of the evaluated grouping concepts fully take into account all of these aspects. Subsequent work of the Task Force will aim at combining the available concepts into a comprehensive 'multiple perspective' framework for the grouping of nanomaterials that will address all of the mentioned aspects of their life cycles.

Safety of titanium dioxide (E171) as a food additive for humans.

Titanium dioxide (TiO2), also known as E171, is commonly used as a white colorant in food, pharmaceuticals, cosmetics, and toothpaste. However, in May 2021, the European Food Safety Authority (EFSA) expert panel, in evaluating the safety of titanium dioxide (E171) as a food additive, concluded that a concern for genotoxicity could not be ruled out. This occurred several years after EFSA had previously considered titanium dioxide to be safe as a food additive. EFSA based this new interpretation on the results of genotoxicity tests of TiO2 nanomaterials. EFSA noted that available data are insufficient to define threshold doses/concentrations of TiO2 particles below which genotoxicity will not occur in tissues containing these particles. Here, it is argued that EFSA made a manifest error regarding the safety of titanium dioxide (E171) particles as a food additive for humans. First, the notion of particle size distribution of TiO2 particles is explained. Second, the changing opinions from the various EFSA evaluations in 2016, 2018, 2019 vs. 2021 are discussed. Third, the low toxicity of TiO2 particles is described in rats exposed by oral gavage and feeding studies in rats and mice. Fourth, the importance of low absorption rates from the gastrointestinal tract vs. circulation in rats and humans but not in mice is identified. Fifth, other international health scientists have weighed in on the EFSA (EFSA J, 2021, 19 (5), 6585) decision and generally disagreed with EFSA's opinion on the safety of E171 TiO2. A common theme voiced by the United Kingdom, Canada, Australia, and New Zealand agencies is that it is inappropriate to compare nanoparticle toxicity studies of dispersed/sonicated nanoparticles with the content of E171 TiO2 in foods because the test materials used in key studies considered by EFSA (EFSA J, 2021, 19 (5), 6585) are not representative of E171 TiO2 particles. Finally, a group of experts recently considered the genotoxicity of TiO2 and could not find support for a direct DNA damaging mechanism of TiO2 (nano and other forms). For these reasons, it is suggested that EFSA made a manifest error on the safety of E171 as a food additive.

Embracing a weight-of-evidence approach for establishing NOAELs for nanoparticle inhalation toxicity studies.

The goal of this article is to evaluate a recently published subchronic inhalation study with carbon nanofibers in rats and discuss the importance of a weight-of-evidence (WOE) framework for determining no adverse effect levels (NOAELs). In this Organization for Economic Cooperation and Development (OECD) 413 guideline inhalation study with VGCF-H carbon nanofibers (CNFs), rats were exposed to 0, 0.54, 2.5 or 25 mg/m(3) CNF for 13 weeks. The standard toxicology experimental design was supplemented with bronchoalveolar lavage (BAL) and respiratory cell proliferation (CP) endpoints. BAL fluid (BALF) recovery of inflammatory cells and mediators (i.e., BALF- lactate dehydrogenase [LDH], microprotein [MTP], and alkaline phosphatase [ALKP] levels) were increased only at 25 mg/m(3), 1 day after exposure. No differences versus control values in were measured at 0.54 or 2.5 mg/m(3) exposure concentrations for any BAL fluid endpoints. Approximately 90% (2.5 and 25 mg/m(3)) of the BAL-recovered macrophages contained CNF. CP indices at 25 mg/m(3) were increased in the airways, lung parenchyma, and subpleural regions, but no increases in CP versus controls were measured at 0.54 or 2.5 mg/m(3). Based upon histopathology criteria, the NOAEL was set at 0.54 mg/m(3), because at 2.5 mg/m(3), "minimal cellular inflammation" of the airways/lung parenchyma was noted by the study pathologist; while the 25 mg/m(3) exposure concentration produced slight inflammation and occasional interstitial thickening. In contrast, none of the more sensitive pulmonary biomarkers such as BAL fluid inflammation/cytotoxicity biomarkers or CP turnover results at 2.5 mg/m(3) were different from air-exposed controls. Given the absence of convergence of the histopathological observations versus more quantitative measures at 2.5 mg/m(3), it is recommended that more comprehensive guidance measures be implemented for setting adverse effect levels in (nano)particulate, subchronic inhalation studies including a WOE approach for establishing no adverse effect levels; and a suggestion that some findings should be viewed as normal physiological adaptations (e.g., normal macrophage phagocytic responses-minimal inflammation) to long-term particulate inhalation exposures.

Occupational Exposure to Poorly Soluble Low Toxicity Particles and Cardiac Disease: A Look at Carbon Black and Titanium Dioxide.

Environmental particulate exposure and the potential risk to people with various types of cardiac diseases, most notably cardiovascular disease, have aroused scientific and regulatory interest worldwide. Epidemiological studies have shown associations between exposure to airborne environmental particulate matter (PM) and mortality from cardiovascular disease (CVD). The associations reported, however, are complex and may not involve a direct role for PM, since air pollutants are diverse and highly correlated. This study examines the potential role of occupational exposure to two types of particles, namely, manufactured carbon black (CB) and titanium dioxide (TiO2), on the risk of cardiovascular disease. To address the risk of cardiovascular disease from exposure to carbon black and titanium dioxide, as reflective of poorly soluble low toxicity particles, we reviewed the published cohort mortality studies of occupational exposure to carbon black and titanium dioxide. Mortality studies of carbon black have been conducted in the United States, Germany, and the United Kingdom. Five mortality studies related to workers involved in the manufacture of titanium dioxide in the United States and Europe have also been conducted. In addition, a meta-analysis of the three-carbon black mortality studies was performed. In the random-effects meta-analysis, full cohort meta-SMRs were 1.01 (95% confidence interval (CI): 0.79-1.29) for heart disease; 1.02 (95% CI: 0.80-1.30) for ischemic heart disease; and 1.08 (95% CI: 0.74-1.59) for acute myocardial infarction (AMI) mortality. A small but imprecise increased AMI mortality risk was suggested for cumulative exposure by a meta-HR = 1.10 per 100 mg/m3-years (95% CI: 0.92-1.31) but not for lugged exposures, that is, for recent exposures. Results of five cohort mortality studies of titanium dioxide workers in the United States and Europe showed no excess in all heart disease or cardiovascular disease. In the most recent study in the United States, an internal analysis, that is, within the cohort itself, with no lag time, showed that the exposure group 15-35 mg/m3-years yielded a significantly increased risk for heart disease; however, there was no evidence of increasing risk with increasing exposure for any of the exposure categories. In contrast to environmental studies, the results of cohort mortality studies do not demonstrate that airborne occupational exposure to carbon black and titanium dioxide particulates increases cardiovascular disease mortality. The lack of a relationship between carbon black and titanium dioxide and CVD mortality suggests that the associations reported in air pollution studies may not be driven by the particulate component.

Debunking some misconceptions about nanotoxicology.

Nanotechnology is currently undergoing an impressive expansion in material science research and development of systems that have novel properties due to their small size. Most of the research efforts have been focused on applications, while the implications efforts (i.e., environmental health and safety) have lagged behind. As a consequence, the success of nanotechnology will require assurances that the products being developed are safe from an environmental, health, and safety standpoint. These concerns have led to a debate among governmental agencies and advocacy groups on whether implementation of special regulations should be required for commercialization of products containing nanomaterials. Therefore the assessments of nanomaterial-related health risks must be accurate and verifiable. A mechanism for conducting well-designed toxicology studies includes rigorous attention to nanoparticle physicochemical characterization, as well as consideration of potential routes of exposure, justification of nanoparticle doses, and inclusion of benchmark controls. Unfortunately, some results obtained from earlier studies have fostered general perceptions and fears about nanoparticle health hazards-based mainly upon simple metrics such as particle size, surface area, and particle dose. In addition, there are currently held views that results of screening in silico or in vitro cell culture assays can serve as adequate screening substitutes for identifying health hazards. Some of these "misconceptions" should be challenged or confirmed by the implementation of thorough and accurately detailed nanotoxicology studies. In this article, the author briefly discusses some of the generalized "misconceptions" regarding nanomaterial toxicity and presents alternative views on these issues.

Pulmonary bioassay studies with brake lining components - Nonfibrous potassium octatitanate - Terracess JS particles in rats.

Nonfibrous potassium octatitanate particles are commercially utilized in applications such as brake pads or brake linings. The aim of this study was to assess lung toxicity in rats exposed to Terracess JS particle-types, one form of nonfibrous octatitanate particulates, and compare the effects to vehicle controls and to Min-U-Sil α-quartz particles as a positive benchmark control particle. Groups of male rats were intratracheally instilled with doses of either 1 or 5 mg/kg of Terracess JS particles or α-quartz particles in phosphate-buffered saline. Phosphate-buffered saline (PBS) solution instilled rats served as vehicle controls. Following exposures, the lungs of PBS and particle-exposed rats were evaluated for bronchoalveolar lavage (BAL) fluid inflammatory biomarkers at post-instillation time points of 1 week, 1 month, and 3 months. In addition, lung tissue morphologies from PBS or 5 mg/kg particle-exposed (Terracess JS or α-quartz) rats were evaluated at postexposure time points of 1 month and 3 months. The BAL fluid results demonstrated that pulmonary instillation exposures in rats to quartz particles produced sustained pulmonary inflammation and significant cytotoxic effects measured at 1 week, 1 month and 3 months postexposure. In contrast, exposures to Terracess JS particle-types produced no significant lung inflammatory or cell injury effects when compared to PBS vehicle control exposed rats. With regard to histopathology of lung tissue, pulmonary exposures to quartz particles in rats produced a progressive, dose-dependent lung inflammatory response characterized by neutrophils and foamy lipid-containing alveolar macrophage accumulation, as well as evidence of early lung tissue thickening consistent with the development of pulmonary fibrosis at the 3-month postexposure time period. In contrast, histopathological analyses of lung tissues revealed that pulmonary exposures to Terracess JS particulates resulted in no significant adverse effects when compared to PBS-exposed controls, as evidenced by the normal lung architecture observed in the exposed animals at post-instillation exposure time periods ranging from 1 month to 3 months. The results described herein demonstrate the benign nature of the pulmonary instillation response in rats following particle exposures to 1 or 5 mg/kg (approximately 1.25 mg) of Terracess JS particle-types in these pulmonary bioassay studies, using appropriate benchmark control particles for comparative evaluations. Thus, based on these results, it is concluded that inhaled Terracess JS particles are expected to have a low-risk potential for producing adverse pulmonary health effects in exposed workers.

Assessing health risks of inhaled nanomaterials: development of pulmonary bioassay hazard studies.

This brief discussion provides an overview of current concepts and perceptions regarding the pulmonary toxicity of ultrafine or nanoparticles. These aspects include, but are not limited to comparisons of fine particle vs. ultrafine particle effects and the unique response of pulmonary effects in rats vs. other rodent species, particularly at particle overload concentrations. In the final section, two studies are described which demonstrate that particle size is not the most significant particulate factor in producing exposure-related pulmonary effects.

Changing the dose metric for inhalation toxicity studies: short-term study in rats with engineered aerosolized amorphous silica nanoparticles.

Inhalation toxicity and exposure assessment studies for nonfibrous particulates have traditionally been conducted using particle mass measurements as the preferred dose metric (i.e., mg or microg/m(3)). However, currently there is a debate regarding the appropriate dose metric for nanoparticle exposure assessment studies in the workplace. The objectives of this study were to characterize aerosol exposures and toxicity in rats of freshly generated amorphous silica (AS) nanoparticles using particle number dose metrics (3.7 x 10(7) or 1.8 x 10(8) particles/cm(3)) for 1- or 3-day exposures. In addition, the role of particle size (d(50) = 37 or 83 nm) on pulmonary toxicity and genotoxicity endpoints was assessed at several postexposure time points. A nanoparticle reactor capable of producing, de novo synthesized, aerosolized amorphous silica nanoparticles for inhalation toxicity studies was developed for this study. SiO(2) aerosol nanoparticle synthesis occurred via thermal decomposition of tetraethylorthosilicate (TEOS). The reactor was designed to produce aerosolized nanoparticles at two different particle size ranges, namely d(50) = approximately 30 nm and d(50) = approximately 80 nm; at particle concentrations ranging from 10(7) to 10(8) particles/cm(3). AS particle aerosol concentrations were consistently generated by the reactor. One- or 3-day aerosol exposures produced no significant pulmonary inflammatory, genotoxic, or adverse lung histopathological effects in rats exposed to very high particle numbers corresponding to a range of mass concentrations (1.8 or 86 mg/m(3)). Although the present study was a short-term effort, the methodology described herein can be utilized for longer-term inhalation toxicity studies in rats such as 28-day or 90-day studies. The expansion of the concept to subchronic studies is practical, due, in part, to the consistency of the nanoparticle generation method.

Health effects related to nanoparticle exposures: environmental, health and safety considerations for assessing hazards and risks.

The field of nanotechnology currently is undergoing a dramatic expansion in material science research and development. Most of the research efforts have been focused on applications; the implications (i.e., health and environmental effects) research has lagged behind. The success of nanotechnology will require assurances that the products being developed are safe from an environmental, health, and safety (EHS) standpoint. In this regard, it has been previously reported in pulmonary toxicity studies that lung exposures to ultrafine or nanoparticles (defined herein as particle size <100 nm in one dimension) produce enhanced adverse inflammatory responses when compared to larger particles of similar composition. Surface properties (particularly particle surface area) and free radical generation, resulting from the interactions of particles with cells may play important roles in nanoparticle toxicity. This brief review identifies some of the key factors for studying EHS risks and hazard effects related to nanoparticle exposures. Health and environmental risk evaluations are products of hazard and exposure assessments. The key factors for discussion herein include the importance of particle characterization studies; development of a nanomaterial risk framework; as well as corresponding hypothesis-driven, mechanistically-oriented investigations, concomitant with base set hazard studies which clearly demonstrate that particle size is only a single (and perhaps minor) factor in influencing the safety of nanomaterials.

A role for nanoparticle surface reactivity in facilitating pulmonary toxicity and development of a base set of hazard assays as a component of nanoparticle risk management.

Results of some lung toxicology studies in rats indicate that pulmonary exposures to ultrafine or nanoparticles produce enhanced inflammatory responses compared to fine-sized particles. Apart from particle size and corresponding surface area considerations, several additional factors may influence the lung toxicity of nanoparticles. These include surface reactivity or surface treatments/coatings of particles, and aggregation potential of aerosolized particles. Conclusions from three pulmonary bioassay hazard/safety studies are summarized herein and demonstrate that particle surface characteristics such as chemical reactivity often correlate better with pulmonary toxicity than particle size or surface area considerations. In the first study, fine-sized quartz particle exposures in rats (500 nm) produced similar effects (inflammation, cytotoxicity, cell proliferation, and/or histopathology) compared to smaller 12-nm nanoscale quartz particles. In another study, no measurable differences in lung toxicity indices were quantified when comparing exposure effects in rats to (1) fine-sized TiO(2) particles (300 nm, 6 m(2)/g [surface area]); (2) TiO(2) nanodots (6-10 nm, 169 m(2)/g); or (3) TiO(2) nanorods (27 m(2)/g). In a third study, exposures to ultrafine TiO(2) particles of similar sizes and different surface areas produced differential degrees of toxicity--based in large part upon surface reactivity endpoints--rather than particle size or surface area indices. Finally, in a related issue for nanotechnology implications, a concept for developing a risk assessment system for the development of new nanomaterials is presented. Embodied in a Nanorisk framework process, implementation of a base set of toxicity tests for evaluating the health and environmental hazards related to nanoparticle exposures is discussed.