Kristin L. Bunker

Single-walled carbon nanotube-induced mitotic disruption

Carbon nanotubes were among the earliest products of nanotechnology and have many potential applications in medicine, electronics, and manufacturing. The low density, small size, and biological persistence of carbon nanotubes create challenges for exposure control and monitoring and make respiratory exposures to workers likely. We have previously shown mitotic spindle aberrations in cultured primary and immortalized human airway epithelial cells exposed to 24, 48 and 96 μg/cm(2) single-walled carbon nanotubes (SWCNT). To investigate mitotic spindle aberrations at concentrations anticipated in exposed workers, primary and immortalized human airway epithelial cells were exposed to SWCNT for 24-72 h at doses equivalent to 20 weeks of exposure at the Permissible Exposure Limit for particulates not otherwise regulated. We have now demonstrated fragmented centrosomes, disrupted mitotic spindles and aneuploid chromosome number at those doses. The data further demonstrated multipolar mitotic spindles comprised 95% of the disrupted mitoses. The increased multipolar mitotic spindles were associated with an increased number of cells in the G2 phase of mitosis, indicating a mitotic checkpoint response. Nanotubes were observed in association with mitotic spindle microtubules, the centrosomes and condensed chromatin in cells exposed to 0.024, 0.24, 2.4 and 24 μg/cm(2) SWCNT. Three-dimensional reconstructions showed carbon nanotubes within the centrosome structure. The lower doses did not cause cytotoxicity or reduction in colony formation after 24h; however, after three days, significant cytotoxicity was observed in the SWCNT-exposed cells. Colony formation assays showed an increased proliferation seven days after exposure. Our results show significant disruption of the mitotic spindle by SWCNT at occupationally relevant doses. The increased proliferation that was observed in carbon nanotube-exposed cells indicates a greater potential to pass the genetic damage to daughter cells. Disruption of the centrosome is common in many solid tumors including lung cancer. The resulting aneuploidy is an early event in the progression of many cancers, suggesting that it may play a role in both tumorigenesis and tumor progression. These results suggest caution should be used in the handling and processing of carbon nanotubes.

What is asbestos and why is it important? Challenges of defining and characterizing asbestos

Asbestos is a term used to describe a group of six fibrous silicate minerals whose unique set of properties has led to widespread use in a variety of commercial products. Asbestos is also commonly associated with potential disease, increasing government regulation, and the upward spiralling costs associated with asbestos abatement and litigation. Yet what exactly is asbestos? The term is in common use and has often been incorrectly applied to many elongated or fibre-shaped mineral particles. However, it has become important to be more precise: which elongated or fibre-shaped mineral particles should be defined as asbestos and which analytical methods should be used to make an accurate identification? This review article is intended to highlight differences among the various mineral particles identified as asbestos and to address controversies that have arisen from the use of the term by a wide range of interested parties. Historical information and summaries of the latest research trends are presented for various academic and professional communities, including geologists, medical doctors and health researchers, regulatory professionals, and legal professionals, in order for them to better understand asbestos-related issues as they consider potential solutions to specific questions.

In Vivo Toxicity Assessment of Occupational Components of the Carbon Nanotube Life Cycle To Provide Context to Potential Health Effects

Pulmonary toxicity studies on carbon nanotubes focus primarily on as-produced materials and rarely are guided by a life cycle perspective or integration with exposure assessment. Understanding toxicity beyond the as-produced, or pure native material, is critical, due to modifications needed to overcome barriers to commercialization of applications. In the first series of studies, the toxicity of as-produced carbon nanotubes and their polymer-coated counterparts was evaluated in reference to exposure assessment, material characterization, and stability of the polymer coating in biological fluids. The second series of studies examined the toxicity of aerosols generated from sanding polymer-coated carbon-nanotube-embedded or neat composites. Postproduction modification by polymer coating did not enhance pulmonary injury, inflammation, and pathology or in vitro genotoxicity of as-produced carbon nanotubes, and for a particular coating, toxicity was significantly attenuated. The aerosols generated from sanding composites embedded with polymer-coated carbon nanotubes contained no evidence of free nanotubes. The percent weight incorporation of polymer-coated carbon nanotubes, 0.15% or 3% by mass, and composite matrix utilized altered the particle size distribution and, in certain circumstances, influenced acute in vivo toxicity. Our study provides perspective that, while the number of workers and consumers increases along the life cycle, toxicity and/or potential for exposure to the as-produced material may greatly diminish.

Factors Affecting the Ambient Physicochemical Properties of Cerium-Containing Particles Generated by Nanoparticle Diesel Fuel Additive Use

Despite the use of cerium oxide nanoparticles (nCe) in some regions as a diesel fuel additive, the physicochemical properties of the resulting exhaust particles in the ambient atmosphere are not well known. The mixing state of ceria with other exhaust particles is one such physicochemical property that has been shown to potentially affect ecosystem/human health. In this study, cerium-containing particles associated with an nCe additive were collected in the laboratory and in Newcastle-upon-Tyne, UK where the local bus fleet uses the Envirox nCe additive. Electron microscopy of laboratory-generated exhaust samples indicated both individual ceria and soot particles (external mixture) and ceria contained within soot agglomerations (internal mixture). Low ambient concentrations prevented quantification of the ceria particle mixing state in the atmosphere; therefore, a multicomponent sectional aerosol dynamic model was used to predict the size, chemical composition, and mixing state of ceria particles as a function of distance from an idealized roadway. Model simulations predicted that most ceria particles remain nonmixed in the ambient atmosphere (300 m downwind from the roadway) due to slow coagulation, with the mixing rate most sensitive to the ceria content of emitted nuclei-mode particles and the particle concentration upwind of the road. Although microscopy analysis showed both external and internal mixtures of ceria and soot in freshly emitted particles, the ambient mass concentration, and size distribution of ceria particles predicted by the model was insensitive to the emitted mixing state. Copyright 2015 American Association for Aerosol Research

Abstract 5464: Genotoxicity of multi-walled carbon nanotubes at occupationally relevant doses

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Carbon nanotubes represent a creative alternative for applications in medicine as therapeutic vectors, imaging and controlled release of active molecules. Although the low density and small size of carbon nanotubes makes respiratory exposures to workers likely during the production or use of commercial products, the potential genotoxicity of multi-walled carbon nanotubes has not been investigated. We have previously shown mitotic spindle aberrations in cultured primary and immortalized human airway epithelial cells exposed to single-walled carbon nanotubes (SWCNT). In order to investigate whether genetic damage was unique to SWCNT, we examined mitotic spindle aberrations following dosing of cells to multi-walled carbon nanotubes (MWCNT) at concentrations anticipated in the workplace. Immortalized and primary respiratory epithelial cells were exposed to 0.024, 0.24, 2.4, and 24μg/cm2 carbon nanotubes. The minimal dose of MWCNT considered for this research was based on the permissible exposure limit for particles with an aerodynamic diameter of less than 5 microns (Occupation Safety and Health Administration). MWCNT induced a dose response of disrupted centrosomes, mitotic spindles and aneuploid chromosome number. The data further showed that monopolar mitotic spindles comprised 95% of the disrupted mitoses. The monopolar phenotype of the disrupted mitotic spindles is in sharp contrast to the multi-polar spindle of SWCNT-induced disruption. Three-dimensional reconstructions showed carbon nanotubes integrated with the microtubules, the DNA and within the centrosome structure. The lower doses did not cause cytotoxicity or apoptosis 24 hours after exposure; however, after 72 hours, significant cytotoxicity was observed in the MWCNT-exposed cells. Disruption of the centrosome is common in many solid tumors including lung cancer. The resulting aneuploidy is a key event in the progression of cancer and is correlated with tumor stage. Our results demonstrate significant disruption of the mitotic spindle by MWCNT at occupationally relevant doses. These results suggest caution should be used in the handling and processing of carbon nanotubes. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5464. doi:1538-7445.AM2012-5464

Application of X-ray Photoelectron Spectroscopy (XPS) for the Surface Characterization of Gunshot Residue (GSR)

Gunshot residue (GSR) is typically found on the hands or clothing of persons who have been in the environment of a discharging firearm, but it may also be found on objects in the vicinity of the discharge. Computer-controlled scanning electron microscopy (CCSEM) is the method preferred by the forensic community for the automated analysis of GSR. GSR samples are therefore typically collected from a crime suspects hands and/or clothing using SEM sample stubs coated with a conductive adhesive. The three major components in modern firearm cartridge primers are lead styphanate (initiator), antimony sulfide (fuel), and barium nitrate (oxidizer). GSR consists of the products of combustion of these primer materials. Large populations of particles in the size range of ~1–10 μm are rapidly screened by energy-dispersive X-ray spectroscopy (EDS) for the presence of combinations of Pb, Sb, and Ba. When combinations of these three elements are detected, the particles are flagged as potential GSR particulate. Flagged particles with significant Pb/Sb/Ba compositions are subsequently relocated for a live identification and a positive confirmation of GSR. Positive particles are classified as being either “characteristic of GSR” (that is, all three key elements are present) or “consistent with GSR” (that is, any two of the key elements are present) based on the particle composition and morphology. Classification is based on the presence of the constituent elements and is not dependant on the element amounts. If a particle does not meet the proper compositional or morphological criteria, it is rejected as GSR. However, CCSEM generally does not provide information regarding the population of particles much less than 1 μm or the surface chemistry of the GSR particles. This article examines the fine fraction of GSR particles with high-resolution electron microscopy and complements the microscopy data with surface chemistry information from X-ray photoelectron spectroscopy (XPS).

Amphiboles between the sheets: observations of interesting morphologies by TEM and FESEM

We observed interesting amphibole morphologies by transmission electron microscopy (TEM) in air samples collected by the United States Environmental Protection Agency (USEPA) in Libby, Montana. Individual amphibole particles were identified and photographed on approximately 200 randomly selected air samples. Transfer of the TEM grids and relocation of these amphiboles in the field emission scanning electron microscope (FESEM) produced high-resolution secondary electron images that allowed a closer examination of these particles. Some of the amphiboles were perfect euhedral single crystals (several micrometers long and 200–500 nanometers wide) showing the classic {110} forms, while other single crystals appeared to be attached to sheet silicates and yet others appeared to have “wings” or “fins,” often less than one micrometer in width, attached to their edges. Energy Dispersive X-Ray Spectroscopy (EDS) and selected area electron diffraction (SAED) from the TEM confirmed that these particles were intergrown amphiboles and sheet silicates. Soil samples collected by the USEPA in Libby, Montana were also analyzed. FESEM investigation revealed that the soil samples contained amphiboles with the same morphologies as observed in the air samples, and were also intergrown with sheet silicates. Thus, these amphiboles occur as single crystals between the layers of vermiculite and hydrobiotite.