Biological effects of inhaled hydraulic fracturing sand dust. II. Particle characterization and pulmonary effects 30 d following intratracheal instillation.

Hydraulic fracturing ("fracking") is used in unconventional gas drilling to allow for the free flow of natural gas from rock. Sand in fracking fluid is pumped into the well bore under high pressure to enter and stabilize fissures in the rock. In the process of manipulating the sand on site, respirable dust (fracking sand dust, FSD) is generated. Inhalation of FSD is a potential hazard to workers inasmuch as respirable crystalline silica causes silicosis, and levels of FSD at drilling work sites have exceeded occupational exposure limits set by OSHA. In the absence of any information about its potential toxicity, a comprehensive rat animal model was designed to investigate the bioactivities of several FSDs in comparison to MIN-U-SIL® 5, a respirable α-quartz reference dust used in previous animal models of silicosis, in several organ systems (Fedan, J.S., Toxicol Appl Pharmacol. 00, 000-000, 2020). The present report, part of the larger investigation, describes: 1) a comparison of the physico-chemical properties of nine FSDs, collected at drilling sites, and MIN-U-SIL® 5, a reference silica dust, and 2) a comparison of the pulmonary inflammatory responses to intratracheal instillation of the nine FSDs and MIN-U-SIL® 5. Our findings indicate that, in many respects, the physico-chemical characteristics, and the biological effects of the FSDs and MIN-U-SIL® 5 after intratracheal instillation, have distinct differences.

Three-dimensional printing with nano-enabled filaments releases polymer particles containing carbon nanotubes into air.

Fused deposition modeling (FDM™) 3-dimensional printing uses polymer filament to build objects. Some polymer filaments are formulated with additives, though it is unknown if they are released during printing. Three commercially available filaments that contained carbon nanotubes (CNTs) were printed with a desktop FDM™ 3-D printer in a chamber while monitoring total particle number concentration and size distribution. Airborne particles were collected on filters and analyzed using electron microscopy. Carbonyl compounds were identified by mass spectrometry. The elemental carbon content of the bulk CNT-containing filaments was 1.5 to 5.2 wt%. CNT-containing filaments released up to 1010 ultrafine (d < 100 nm) particles/g printed and 106 to 108 respirable (d ~0.5 to 2 µm) particles/g printed. From microscopy, 1% of the emitted respirable polymer particles contained visible CNTs. Carbonyl emissions were observed above the limit of detection (LOD) but were below the limit of quantitation (LOQ). Modeling indicated that, for all filaments, the average proportional lung deposition of CNT-containing polymer particles was 6.5%, 5.7%, and 7.2% for the head airways, tracheobronchiolar, and pulmonary regions, respectively. If CNT-containing polymer particles are hazardous, it would be prudent to control emissions during use of these filaments.

Differentiating gold nanorod samples using particle size and shape distributions from transmission electron microscope images.

Size and shape distributions of gold nanorod samples are critical to their physico-chemical properties, especially their longitudinal surface plasmon resonance. This interlaboratory comparison study developed methods for measuring and evaluating size and shape distributions for gold nanorod samples using transmission electron microscopy (TEM) images. The objective was to determine whether two different samples, which had different performance attributes in their application, were different with respect to their size and/or shape descriptor distributions. Touching particles in the captured images were identified using a ruggedness shape descriptor. Nanorods could be distinguished from nanocubes using an elongational shape descriptor. A non-parametric statistical test showed that cumulative distributions of an elongational shape descriptor, that is, the aspect ratio, were statistically different between the two samples for all laboratories. While the scale parameters of size and shape distributions were similar for both samples, the width parameters of size and shape distributions were statistically different. This protocol fulfills an important need for a standardized approach to measure gold nanorod size and shape distributions for applications in which quantitative measurements and comparisons are important. Furthermore, the validated protocol workflow can be automated, thus providing consistent and rapid measurements of nanorod size and shape distributions for researchers, regulatory agencies, and industry.

Surgical smoke control with local exhaust ventilation: Experimental study.

This experimental study aimed to evaluate airborne particulates and volatile organic compounds (VOCs) from surgical smoke when a local exhaust ventilation (LEV) system is in place. Surgical smoke was generated from human tissue in an unoccupied operating room using an electrocautery surgical device for 15 min with 3 different test settings: (1) without LEV control; (2) control with a wall irrigation suction unit with an in-line ultra-low penetration air filter; and (3) control with a smoke evacuation system. Flow rate of LEVs was approximately 35 L/min and suction was maintained within 5 cm of electrocautery interaction site. A total of 6 experiments were conducted. Particle number and mass concentrations were measured using direct reading instruments including a condensation particle counter (CPC), a light-scattering laser photometer (DustTrak DRX), a scanning mobility particle sizer (SMPS), an aerodynamic particle sizer (APS), and a viable particle counter. Selected VOCs were collected using evacuated canisters using grab, personal and area sampling techniques. The largest average particle and VOCs concentrations were found in the absence of LEV control followed by LEV controls. Average ratios of LEV controls to without LEV control ranged 0.24-0.33 (CPC), 0.28-0.39 (SMPS), 0.14-0.31 (DustTrak DRX), and 0.26-0.55 (APS). Ethanol and isopropyl alcohol were dominant in the canister samples. Acetaldehyde, acetone, acetonitrile, benzene, hexane, styrene, and toluene were detected but at lower concentrations (<500 µg/m3) and concentrations of the VOCs were much less than the National Institute for Occupational Safety and Health recommended exposure limit values. Utilization of the LEVs for surgical smoke control can significantly reduce but not completely eliminate airborne particles and VOCs.

Accumulation of Ubiquitin and Sequestosome-1 Implicate Protein Damage in Diacetyl-Induced Cytotoxicity.

Inhaled diacetyl vapors are associated with flavorings-related lung disease, a potentially fatal airway disease. The reactive α-dicarbonyl group in diacetyl causes protein damage in vitro. Dicarbonyl/l-xylulose reductase (DCXR) metabolizes diacetyl into acetoin, which lacks this α-dicarbonyl group. To investigate the hypothesis that flavorings-related lung disease is caused by in vivo protein damage, we correlated diacetyl-induced airway damage in mice with immunofluorescence for markers of protein turnover and autophagy. Western immunoblots identified shifts in ubiquitin pools. Diacetyl inhalation caused dose-dependent increases in bronchial epithelial cells with puncta of both total ubiquitin and K63-ubiquitin, central mediators of protein turnover. This response was greater in Dcxr-knockout mice than in wild-type controls inhaling 200 ppm diacetyl, further implicating the α-dicarbonyl group in protein damage. Western immunoblots demonstrated decreased free ubiquitin in airway-enriched fractions. Transmission electron microscopy and colocalization of ubiquitin-positive puncta with lysosomal-associated membrane proteins 1 and 2 and with the multifunctional scaffolding protein sequestosome-1 (SQSTM1/p62) confirmed autophagy. Surprisingly, immunoreactive SQSTM1 also accumulated in the olfactory bulb of the brain. Olfactory bulb SQSTM1 often congregated in activated microglial cells that also contained olfactory marker protein, indicating neuronophagia within the olfactory bulb. This suggests the possibility that SQSTM1 or damaged proteins may be transported from the nose to the brain. Together, these findings strongly implicate widespread protein damage in the etiology of flavorings-related lung disease.

Role of epithelial-mesenchymal transition (EMT) and fibroblast function in cerium oxide nanoparticles-induced lung fibrosis.

The emission of cerium oxide nanoparticles (CeO2) from diesel engines, using cerium compounds as a catalyst to lower the diesel exhaust particles, is a health concern. We have previously shown that CeO2 induced pulmonary inflammation and lung fibrosis. The objective of the present study was to investigate the modification of fibroblast function and the role of epithelial-mesenchymal transition (EMT) in CeO2-induced fibrosis. Male Sprague-Dawley rats were exposed to CeO2 (0.15 to 7mg/kg) by a single intratracheal instillation and sacrificed at various times post-exposure. The results show that at 28days after CeO2 (3.5mg/kg) exposure, lung fibrosis was evidenced by increased soluble collagen in bronchoalveolar lavage fluid, elevated hydroxyproline content in lung tissues, and enhanced sirius red staining for collagen in the lung tissue. Lung fibroblasts and alveolar type II (ATII) cells isolated from CeO2-exposed rats at 28days post-exposure demonstrated decreasing proliferation rate when compare to the controls. CeO2 exposure was cytotoxic and altered cell function as demonstrated by fibroblast apoptosis and aggregation, and ATII cell hypertrophy and hyperplasia with increased surfactant. The presence of stress fibers, expressed as α-smooth muscle actin (SMA), in CeO2-exposed fibroblasts and ATII cells was significantly increased compared to the control. Immunohistofluorescence analysis demonstrated co-localization of TGF-ß or α-SMA with prosurfactant protein C (SPC)-stained ATII cells. These results demonstrate that CeO2 exposure affects fibroblast function and induces EMT in ATII cells that play a role in lung fibrosis. These findings suggest potential adverse health effects in response to CeO2 nanoparticle exposure.

Comparison of the toxicity of sintered and unsintered indium-tin oxide particles in murine macrophage and epidermal cells.

Indium-tin oxide (ITO) is used to produce flat panel displays and several other technology products. Composed of 90% indium oxide (In2O3) and 10% tin oxide (SnO2) by weight, ITO is synthesized under conditions of high heat via a process known as sintering. Indium lung disease, a recently recognized occupational illness, is characterized by pulmonary alveolar proteinosis, fibrosis, and emphysema. Murine macrophage (RAW 264.7) and epidermal (JB6) cells stably transfected with AP-1 to study tumor promoting potential, were used to differentiate between the toxicological profiles of sintered ITO (SITO) and unsintered mixture (UITO). We hypothesized that sintering would play a key role in free radical generation and cytotoxicity. Exposure of cells to both UITO and SITO caused a time and dose dependent decrease of the viability of cells. Intracellular ROS generation was inversely related to the dose of both UITO and SITO, a direct reflection of the decreased number of viable RAW 264.7 and JB6/AP-1 cells observed at higher concentrations. Electron spin resonance showed significantly increased hydroxyl radical (OH) generation in cells exposed to UITO compared to SITO. This is different from LDH release, which showed that SITO caused significantly increased damage to the cell membrane compared to UITO. Lastly, the JB6/AP-1 cell line did not show activation of the AP-1 pathway. Our results highlight both the differences in the mechanisms of cytotoxicity and the consistent adverse effects associated with UITO and SITO exposure.

Pulmonary toxicity and global gene expression changes in response to sub-chronic inhalation exposure to crystalline silica in rats.

Exposure to crystalline silica results in serious adverse health effects, most notably, silicosis. An understanding of the mechanism(s) underlying silica-induced pulmonary toxicity is critical for the intervention and/or prevention of its adverse health effects. Rats were exposed by inhalation to crystalline silica at a concentration of 15 mg/m3, 6 hr/day, 5 days/week for 3, 6 or 12 weeks. Pulmonary toxicity and global gene expression profiles were determined in lungs at the end of each exposure period. Crystalline silica was visible in lungs of rats especially in the 12-week group. Pulmonary toxicity, as evidenced by an increase in lactate dehydrogenase (LDH) activity and albumin content and accumulation of macrophages and neutrophils in the bronchoalveolar lavage (BAL), was seen in animals depending upon silica exposure duration. The most severe histological changes, noted in the 12-week exposure group, consisted of chronic active inflammation, type II pneumocyte hyperplasia, and fibrosis. Microarray analysis of lung gene expression profiles detected significant differential expression of 38, 77, and 99 genes in rats exposed to silica for 3-, 6-, or 12-weeks, respectively, compared to time-matched controls. Among the significantly differentially expressed genes (SDEG), 32 genes were common in all exposure groups. Bioinformatics analysis of the SDEG identified enrichment of functions, networks and canonical pathways related to inflammation, cancer, oxidative stress, fibrosis, and tissue remodeling in response to silica exposure. Collectively, these results provided insights into the molecular mechanisms underlying pulmonary toxicity following sub-chronic inhalation exposure to crystalline silica in rats.

Pulmonary toxicity following acute coexposures to diesel particulate matter and α-quartz crystalline silica in the Sprague-Dawley rat.

The effects of acute pulmonary coexposures to silica and diesel particulate matter (DPM), which may occur in various mining operations, were investigated in vivo. Rats were exposed by intratracheal instillation (IT) to silica (50 or 233 µg), DPM (7.89 or 50 µg) or silica and DPM combined in phosphate-buffered saline (PBS) or to PBS alone (control). At one day, one week, one month, two months and three months postexposure bronchoalveolar lavage and histopathology were performed to assess lung injury, inflammation and immune response. While higher doses of silica caused inflammation and injury at all time points, DPM exposure alone did not. DPM (50 µg) combined with silica (233 µg) increased inflammation at one week and one-month postexposure and caused an increase in the incidence of fibrosis at one month compared with exposure to silica alone. To assess susceptibility to lung infection following coexposure, rats were exposed by IT to 233 µg silica, 50 µg DPM, a combination of the two or PBS control one week before intratracheal inoculation with 5 × 105 Listeria monocytogenes. At 1, 3, 5, 7 and 14 days following infection, pulmonary immune response and bacterial clearance from the lung were evaluated. Coexposure to DPM and silica did not alter bacterial clearance from the lung compared to control. Although DPM and silica coexposure did not alter pulmonary susceptibility to infection in this model, the study showed that noninflammatory doses of DPM had the capacity to increase silica-induced lung injury, inflammation and onset/incidence of fibrosis.

Characterization of chemical contaminants generated by a desktop fused deposition modeling 3-dimensional Printer.

Printing devices are known to emit chemicals into the indoor atmosphere. Understanding factors that influence release of chemical contaminants from printers is necessary to develop effective exposure assessment and control strategies. In this study, a desktop fused deposition modeling (FDM) 3-dimensional (3-D) printer using acrylonitrile butadiene styrene (ABS) or polylactic acid (PLA) filaments and two monochrome laser printers were evaluated in a 0.5 m3 chamber. During printing, chamber air was monitored for vapors using a real-time photoionization detector (results expressed as isobutylene equivalents) to measure total volatile organic compound (TVOC) concentrations, evacuated canisters to identify specific VOCs by off-line gas chromatography-mass spectrometry (GC-MS) analysis, and liquid bubblers to identify carbonyl compounds by GC-MS. Airborne particles were collected on filters for off-line analysis using scanning electron microscopy with an energy dispersive x-ray detector to identify elemental constituents. For 3-D printing, TVOC emission rates were influenced by a printer malfunction, filament type, and to a lesser extent, by filament color; however, rates were not influenced by the number of printer nozzles used or the manufacturer's provided cover. TVOC emission rates were significantly lower for the 3-D printer (49-3552 µg h-1) compared to the laser printers (5782-7735 µg h-1). A total of 14 VOCs were identified during 3-D printing that were not present during laser printing. 3-D printed objects continued to off-gas styrene, indicating potential for continued exposure after the print job is completed. Carbonyl reaction products were likely formed from emissions of the 3-D printer, including 4-oxopentanal. Ultrafine particles generated by the 3-D printer using ABS and a laser printer contained chromium. Consideration of the factors that influenced the release of chemical contaminants (including known and suspected asthmagens such as styrene and 4-oxopentanal) from a FDM 3-D printer should be made when designing exposure assessment and control strategies.

In Vitro Toxicity Evaluation of Lignin-(Un)coated Cellulose Based Nanomaterials on Human A549 and THP-1 Cells.

A significant amount of research toward commercial development of cellulose based nanomaterials (CNM) is now in progress with some potential applications. Using human A549 and THP-1 cells, we evaluated the biological responses of various CNMs, made out of similar material but with functional and morphological variations. While A549 cells displayed minimal or no cytotoxic responses following exposure to CNMs, THP-1 cells were more susceptible to cytotoxicity, cellular damage and inflammatory responses. Further analysis of these biological responses evaluated using hierarchical clustering approaches was effective in discriminating (dis)-similarities of various CNMs studied and identified potential inflammatory factors contributing to cytotoxicity. No correlation between cytotoxicity and surface properties of CNMs was found. This study clearly highlights that, in addition to the source and characteristics of CNMs, cell type-specific differences in the recognition/uptake of CNMs along with their inherent capability to respond to external stimuli are crucial for assessing the toxicity of CNMs.

Differential pulmonary effects of CoO and La2O3 metal oxide nanoparticle responses during aerosolized inhalation in mice.

BACKGROUND: Although classified as metal oxides, cobalt monoxide (CoO) and lanthanum oxide (La2O3) nanoparticles, as representative transition and rare earth oxides, exhibit distinct material properties that may result in different hazardous potential in the lung. The current study was undertaken to compare the pulmonary effects of aerosolized whole body inhalation of these nanoparticles in mice. RESULTS: Mice were exposed to filtered air (control) and 10 or 30 mg/m(3) of each particle type for 4 days and then examined at 1 h, 1, 7 and 56 days post-exposure. The whole lung burden 1 h after the 4 day inhalation of CoO nanoparticles was 25 % of that for La2O3 nanoparticles. At 56 days post exposure, < 1 % of CoO nanoparticles remained in the lungs; however, 22-50 % of the La2O3 nanoparticles lung burden 1 h post exposure was retained at 56 days post exposure for low and high exposures. Significant accumulation of La2O3 nanoparticles in the tracheobronchial lymph nodes was noted at 56 days post exposure. When exposed to phagolysosomal simulated fluid, La nanoparticles formed urchin-shaped LaPO4 structures, suggesting that retention of this rare earth oxide nanoparticle may be due to complexation of cellular phosphates within lysosomes. CoO nanoparticles caused greater lactate dehydrogenase release in the bronchoalveolar fluid (BALF) compared to La2O3 nanoparticles at 1 day post exposure, while BAL cell differentials indicate that La2O3 nanoparticles generated more inflammatory cell infiltration at all doses and exposure points. Histopathological analysis showed acute inflammatory changes at 1 day after inhalation of either CoO or La2O3 nanoparticles. Only the 30 mg/m(3) La2O3 nanoparticles exposure caused chronic inflammatory changes and minimal fibrosis at day 56 post exposure. This is in agreement with activation of the NRLP3 inflammasome after in vitro exposure of differentiated THP-1 macrophages to La2O3 but not after CoO nanoparticles exposure. CONCLUSION: Taken together, the inhalation studies confirmed the trend of our previous sub-acute aspiration study, which reported that CoO nanoparticles induced more acute pulmonary toxicity, while La2O3 nanoparticles caused chronic inflammatory changes and minimal fibrosis.

Evaluation of pulmonary and systemic toxicity following lung exposure to graphite nanoplates: a member of the graphene-based nanomaterial family.

BACKGROUND: Graphene, a monolayer of carbon, is an engineered nanomaterial (ENM) with physical and chemical properties that may offer application advantages over other carbonaceous ENMs, such as carbon nanotubes (CNT). The goal of this study was to comparatively assess pulmonary and systemic toxicity of graphite nanoplates, a member of the graphene-based nanomaterial family, with respect to nanoplate size. METHODS: Three sizes of graphite nanoplates [20 µm lateral (Gr20), 5 µm lateral (Gr5), and <2 µm lateral (Gr1)] ranging from 8-25 nm in thickness were characterized for difference in surface area, structure,, zeta potential, and agglomeration in dispersion medium, the vehicle for in vivo studies. Mice were exposed by pharyngeal aspiration to these 3 sizes of graphite nanoplates at doses of 4 or 40 µg/mouse, or to carbon black (CB) as a carbonaceous control material. At 4 h, 1 day, 7 days, 1 month, and 2 months post-exposure, bronchoalveolar lavage was performed to collect fluid and cells for analysis of lung injury and inflammation. Particle clearance, histopathology and gene expression in lung tissue were evaluated. In addition, protein levels and gene expression were measured in blood, heart, aorta and liver to assess systemic responses. RESULTS: All Gr samples were found to be similarly composed of two graphite structures and agglomerated to varying degrees in DM in proportion to the lateral dimension. Surface area for Gr1 was approximately 7-fold greater than Gr5 and Gr20, but was less reactive reactive per m(2). At the low dose, none of the Gr materials induced toxicity. At the high dose, Gr20 and Gr5 exposure increased indices of lung inflammation and injury in lavage fluid and tissue gene expression to a greater degree and duration than Gr1 and CB. Gr5 and Gr20 showed no or minimal lung epithelial hypertrophy and hyperplasia, and no development of fibrosis by 2 months post-exposure. In addition, the aorta and liver inflammatory and acute phase genes were transiently elevated in Gr5 and Gr20, relative to Gr1. CONCLUSIONS: Pulmonary and systemic toxicity of graphite nanoplates may be dependent on lateral size and/or surface reactivity, with the graphite nanoplates > 5 µm laterally inducing greater toxicity which peaked at the early time points post-exposure relative to the 1-2 µm graphite nanoplate.

Emission of particulate matter from a desktop three-dimensional (3D) printer.

Desktop three-dimensional (3D) printers are becoming commonplace in business offices, public libraries, university labs and classrooms, and even private homes; however, these settings are generally not designed for exposure control. Prior experience with a variety of office equipment devices such as laser printers that emit ultrafine particles (UFP) suggests the need to characterize 3D printer emissions to enable reliable risk assessment. The aim of this study was to examine factors that influence particulate emissions from 3D printers and characterize their physical properties to inform risk assessment. Emissions were evaluated in a 0.5-m(3) chamber and in a small room (32.7 m(3)) using real-time instrumentation to measure particle number, size distribution, mass, and surface area. Factors evaluated included filament composition and color, as well as the manufacturer-provided printer emissions control technologies while printing an object. Filament type significantly influenced emissions, with acrylonitrile butadiene styrene (ABS) emitting larger particles than polylactic acid (PLA), which may have been the result of agglomeration. Geometric mean particle sizes and total particle (TP) number and mass emissions differed significantly among colors of a given filament type. Use of a cover on the printer reduced TP emissions by a factor of 2. Lung deposition calculations indicated a threefold higher PLA particle deposition in alveoli compared to ABS. Desktop 3D printers emit high levels of UFP, which are released into indoor environments where adequate ventilation may not be present to control emissions. Emissions in nonindustrial settings need to be reduced through the use of a hierarchy of controls, beginning with device design, followed by engineering controls (ventilation) and administrative controls such as choice of filament composition and color.

Treated and untreated rock dust: Quartz content and physical characterization.

Rock dusting is used to prevent secondary explosions in coal mines, but inhalation of rock dusts can be hazardous if the crystalline silica (e.g., quartz) content in the respirable fraction is high. The objective of this study is to assess the quartz content and physical characteristics of four selected rock dusts, consisting of limestone or marble in both treated (such as treatment with stearic acid or stearates) and untreated forms. Four selected rock dusts (an untreated and treated limestone and an untreated and treated marble) were aerosolized in an aerosol chamber. Respirable size-selective sampling was conducted along with particle size-segregated sampling using a Micro-Orifice Uniform Deposit Impactor. Fourier Transform Infrared spectroscopy and scanning electron microscopy with energy-dispersive X-ray (SEM-EDX) analyses were used to determine quartz mass and particle morphology, respectively. Quartz percentage in the respirable dust fraction of untreated and treated forms of the limestone dust was significantly higher than in bulk samples, but since the bulk percentage was low the enrichment factor would not have resulted in any major change to conclusions regarding the contribution of respirable rock dust to the overall airborne quartz concentration. The quartz percentage in the marble dust (untreated and treated) was very low and the respirable fractions showed no enrichment. The spectra from SEM-EDX analysis for all materials were predominantly from calcium carbonate, clay, and gypsum particles. No free quartz particles were observed. The four rock dusts used in this study are representative of those presented for use in rock dusting, but the conclusions may not be applicable to all available materials.

Performance of a scanning mobility particle sizer in measuring diverse types of airborne nanoparticles: Multi-walled carbon nanotubes, welding fumes, and titanium dioxide spray.

Direct-reading instruments have been widely used for characterizing airborne nanoparticles in inhalation toxicology and industrial hygiene studies for exposure/risk assessments. Instruments using electrical mobility sizing followed by optical counting, e.g., scanning or sequential mobility particle spectrometers (SMPS), have been considered as the "gold standard" for characterizing nanoparticles. An SMPS has the advantage of rapid response and has been widely used, but there is little information on its performance in assessing the full spectrum of nanoparticles encountered in the workplace. In this study, an SMPS was evaluated for its effectiveness in producing "monodisperse" aerosol and its adequacy in characterizing overall particle size distribution using three test aerosols, each mimicking a unique class of real-life nanoparticles: singlets of nearly spherical titanium dioxide (TiO2), agglomerates of fiber-like multi-walled carbon nanotube (MWCNT), and aggregates that constitutes welding fume (WF). These aerosols were analyzed by SMPS, cascade impactor, and by counting and sizing of discrete particles by scanning and transmission electron microscopy. The effectiveness of the SMPS to produce classified particles (fixed voltage mode) was assessed by examination of the resulting geometric standard deviation (GSD) from the impactor measurement. Results indicated that SMPS performed reasonably well for TiO2 (GSD = 1.3), but not for MWCNT and WF as evidenced by the large GSD values of 1.8 and 1.5, respectively. For overall characterization, results from SMPS (scanning voltage mode) exhibited particle-dependent discrepancies in the size distribution and total number concentration compared to those from microscopic analysis. Further investigation showed that use of a single-stage impactor at the SMPS inlet could distort the size distribution and underestimate the concentration as shown by the SMPS, whereas the presence of vapor molecules or atom clusters in some test aerosols might cause artifacts by counting "phantom particles." Overall, the information obtained from this study will help understand the limitations of the SMPS in measuring nanoparticles so that one can adequately interpret the results for risk assessments and exposure prevention in an occupational or ambient environment.

Effects of amorphous silica coating on cerium oxide nanoparticles induced pulmonary responses.

Recently cerium compounds have been used in a variety of consumer products, including diesel fuel additives, to increase fuel combustion efficiency and decrease diesel soot emissions. However, cerium oxide (CeO2) nanoparticles have been detected in the exhaust, which raises a health concern. Previous studies have shown that exposure of rats to nanoscale CeO2 by intratracheal instillation (IT) induces sustained pulmonary inflammation and fibrosis. In the present study, male Sprague-Dawley rats were exposed to CeO2 or CeO2 coated with a nano layer of amorphous SiO2 (aSiO2/CeO2) by a single IT and sacrificed at various times post-exposure to assess potential protective effects of the aSiO2 coating. The first acellular bronchoalveolar lavage (BAL) fluid and BAL cells were collected and analyzed from all exposed animals. At the low dose (0.15mg/kg), CeO2 but not aSiO2/CeO2 exposure induced inflammation. However, at the higher doses, both particles induced a dose-related inflammation, cytotoxicity, inflammatory cytokines, matrix metalloproteinase (MMP)-9, and tissue inhibitor of MMP at 1day post-exposure. Morphological analysis of lung showed an increased inflammation, surfactant and collagen fibers after CeO2 (high dose at 3.5mg/kg) treatment at 28days post-exposure. aSiO2 coating significantly reduced CeO2-induced inflammatory responses in the airspace and appeared to attenuate phospholipidosis and fibrosis. Energy dispersive X-ray spectroscopy analysis showed Ce and phosphorous (P) in all particle-exposed lungs, whereas Si was only detected in aSiO2/CeO2-exposed lungs up to 3days after exposure, suggesting that aSiO2 dissolved off the CeO2 core, and some of the CeO2 was transformed to CePO4 with time. These results demonstrate that aSiO2 coating reduce CeO2-induced inflammation, phospholipidosis and fibrosis.

Diacetyl and 2,3-pentanedione exposure of human cultured airway epithelial cells: Ion transport effects and metabolism of butter flavoring agents.

Inhalation of butter flavoring by workers in the microwave popcorn industry may result in "popcorn workers' lung." In previous in vivo studies rats exposed for 6 h to vapor from the flavoring agents, diacetyl and 2,3-pentanedione, acquired flavoring concentration-dependent damage of the upper airway epithelium and airway hyporeactivity to inhaled methacholine. Because ion transport is essential for lung fluid balance,we hypothesized that alterations in ion transport may be an early manifestation of butter flavoring-induced toxicity.We developed a system to expose cultured human bronchial/tracheal epithelial cells (NHBEs) to flavoring vapors. NHBEs were exposed for 6 h to diacetyl or 2,3-pentanedione vapors (25 or ≥ 60 ppm) and the effects on short circuit current and transepithelial resistance (Rt) were measured. Immediately after exposure to 25 ppm both flavorings reduced Na+ transport,without affecting Cl- transport or Na+,K+-pump activity. Rt was unaffected. Na+ transport recovered 18 h after exposure. Concentrations (100-360 ppm) of diacetyl and 2,3-pentanedione reported earlier to give rise in vivo to epithelial damage, and 60 ppm, caused death of NHBEs 0 h post-exposure. Analysis of the basolateral medium indicated that NHBEs metabolize diacetyl and 2,3-pentanedione to acetoin and 2-hydroxy-3-pentanone, respectively. The results indicate that ion transport is inhibited transiently in airway epithelial cells by lower concentrations of the flavorings than those that result in morphological changes of the cells in vivo or in vitro.

Detection and quantification of 2H and 3R phases in commercial graphene-based materials.

Graphene-based material (GBM) samples acquired from commercial sources are investigated using X-ray diffraction (XRD). Of the 18 GBM samples investigated here, seven samples show XRD patterns with features characteristic of the graphite structure. The XRD patterns of the seven samples are analyzed showing the presence of both the ABA (2H) structure and the ABCA (3R) structure. After de-convoluting the (101) lines of the 2H and 3R structures, the areas under the peaks are used to determine the relative concentrations of the 2H and 3R phases present, typically yielding the ratio 60/40 for 2H/3R. The presence of the 3R structure is important since the 3R structure is a semiconductor with tunable band gap and it is less stable than the 2H structure. The number of layers determined from the analysis of the XRD data varies between 65 and 109 for different samples yielding thickness of the graphite sheets varying between 22 nm and 37 nm. Scanning electron microscopy and transmission electron microscopy of three representative samples confirms the sheet-like morphology and stacking of the graphene layers in the samples. Relevance of these results in connection with their potential applications and toxicology is briefly discussed.

The group A streptococcal collagen-like protein-1, Scl1, mediates biofilm formation by targeting the extra domain A-containing variant of cellular fibronectin expressed in wounded tissue.

Wounds are known to serve as portals of entry for group A Streptococcus (GAS). Subsequent tissue colonization is mediated by interactions between GAS surface proteins and host extracellular matrix components. We recently reported that the streptococcal collagen-like protein-1, Scl1, selectively binds the cellular form of fibronectin (cFn) and also contributes to GAS biofilm formation on abiotic surfaces. One structural feature of cFn, which is predominantly expressed in response to tissue injury, is the presence of a spliced variant containing extra domain A (EDA/EIIIA). We now report that GAS biofilm formation is mediated by the Scl1 interaction with EDA-containing cFn. Recombinant Scl1 proteins that bound cFn also bound recombinant EDA within the C-C' loop region recognized by the α(9)ß(1) integrin. The extracellular 2-D matrix derived from human dermal fibroblasts supports GAS adherence and biofilm formation. Altogether, this work identifies and characterizes a novel molecular mechanism by which GAS utilizes Scl1 to specifically target an extracellular matrix component that is predominantly expressed at the site of injury in order to secure host tissue colonization.