Search

Human diseases associated with exposure to asbestos fibers include pleural fibrosis and plaques, pulmonary fibrosis (asbestosis), lung cancer, and diffuse malignant mesothelioma. The critical determinants of fiber bioactivity and toxicity include not only fiber dimensions, but also shape, surface reactivity, crystallinity, chemical composition, and presence of transition metals. Depending on their size and dimensions, inhaled fibers can penetrate the respiratory tract to the distal airways and into the alveolar spaces. Fibers can be cleared by several mechanisms, including the mucociliary escalator, engulfment, and removal by macrophages, or through splitting and chemical modification. Biopersistence of long asbestos fibers can lead to inflammation, granuloma formation, fibrosis, and cancer. Exposure to synthetic carbon nanomaterials, including carbon nanofibers and carbon nanotubes (CNTs), is considered a potential health hazard because of their physical similarities with asbestos fibers. Respiratory exposure to CNTs can produce an inflammatory response, diffuse interstitial fibrosis, and formation of fibrotic granulomas similar to that observed in asbestos-exposed animals and humans. Given the known cytotoxic and carcinogenic properties of asbestos fibers, toxicity of fibrous nanomaterials is a topic of intense study. The mechanisms of nanomaterial toxicity remain to be fully elucidated, but recent evidence suggests points of similarity with asbestos fibers, including a role for generation of reactive oxygen species, oxidative stress, and genotoxicity. Considering the rapid increase in production and use of fibrous nanomaterials, it is imperative to gain a thorough understanding of their biologic activity to avoid the human health catastrophe that has resulted from widespread use of asbestos fibers.

Background The most common causes of granulomatous inflammation are persistent pathogens and poorly-degradable irritating materials. A characteristic pathological reaction to intratracheal instillation, pharyngeal aspiration, or inhalation of carbon nanotubes is formation of epithelioid granulomas accompanied by interstitial fibrosis in the lungs. In the mesothelium, a similar response is induced by high aspect ratio nanomaterials, including asbestos fibers, following intraperitoneal injection. This asbestos-like behaviour of some engineered nanomaterials is a concern for their potential adverse health effects in the lungs and mesothelium. We hypothesize that high aspect ratio nanomaterials will induce epithelioid granulomas in nonadherent macrophages in 3D cultures. Results Carbon black particles (Printex 90) and crocidolite asbestos fibers were used as well-characterized reference materials and compared with three commercial samples of multiwalled carbon nanotubes (MWCNTs). Doses were identified in 2D and 3D cultures in order to minimize acute toxicity and to reflect realistic occupational exposures in humans and in previous inhalation studies in rodents. Under serum-free conditions, exposure of nonadherent primary murine bone marrow-derived macrophages to 0.5 μg/ml (0.38 μg/cm2) of crocidolite asbestos fibers or MWCNTs, but not carbon black, induced macrophage differentiation into epithelioid cells and formation of stable aggregates with the characteristic morphology of granulomas. Formation of multinucleated giant cells was also induced by asbestos fibers or MWCNTs in this 3D in vitro model. After 7-14 days, macrophages exposed to high aspect ratio nanomaterials co-expressed proinflammatory (M1) as well as profibrotic (M2) phenotypic markers. Conclusions Induction of epithelioid granulomas appears to correlate with high aspect ratio and complex 3D structure of carbon nanotubes, not with their iron content or surface area. This model offers a time- and cost-effective platform to evaluate the potential of engineered high aspect ratio nanomaterials, including carbon nanotubes, nanofibers, nanorods and metallic nanowires, to induce granulomas following inhalation.

Malignant mesothelioma (MM) is an aggressive form of cancer that arises in the lining of the peritoneal, pleural or pericardial cavities. It has a long latency period, and the development of this rare cancer is typically associated with asbestos exposure. Mesothelioma may also be a concern for carbon nanotubes (CNTs), which are under development for biomedical applications[1-3] but have been recently shown to produce asbestos-like pathogenic behavior in mice, suggesting their potential to induce mesothelioma upon chronic exposure.[4-6] The diffuse nature of the tumors makes their surgical removal difficult, and they are resistant to conventional chemotherapy.[7,8] As an alternative strategy, non-conventional approaches involving new chemical agents, and targeting methods minimizing side effects are needed. Recently it has been shown that selenium, in the form of selenite and selenocysteine, can selectively inhibit growth and induce apoptotic cell death in MM cells compared to normal mesothelial cells,[9] i.e. exhibits differential toxicity. Selenium is believed to play a role in cancer prevention,[10,11] and selenium deficiency has been linked with increased cancer incidence.[11-13] The fundamental mechanisms of selenium chemo-prevention are not fully understood. At doses marginally higher than the essential dose (Recommended Daily Allowance = 55 μg/day, No Adverse Effect Level = 800 μg/day), selenium can cause toxicity.[14] The combination of the administered dose and the chemical form co-determine selenium’s activity as toxic or carcinostatic.[10,11] For example, the selenoenzyme glutathione peroxidase can protect cells from oxidative damage[15,16] and the seleno-enzymes, glutathione S-transferases, can repair damaged DNA and prevent mutation.[16] Other forms of selenium, however, can produce reactive oxygen species[10] leading to oxidative stress and cell death. In recent years, elemental nano-selenium (nSe) has gained special attention due to its therapeutic properties.[17,18] As a major advantage over other selenium forms, elemental (zero-valent) nanoparticles are a high-Se-density formulation with the potential for local delivery of high doses into cancer cells. To achieve this in practice requires a nSe formulation that is rapidly internalized by the target cell and is sufficiently stable in the extracellular fluid to reach the target cell intact. Here we synthesize two competing formulations of nano-selenium and evaluate their relative effectiveness for high-dose intracellular delivery. We show that a novel selenium-carbon composite nanostructure (CNPnSe) is highly effective in inducing death in malignant mesothelial cells in vitro and shows promise for the development of targeted therapy for malignant mesothelioma.[12]

The nucleoside reverse transcriptase inhibitor AZT induces multiple manifestations of genotoxicity, among which aneuploidy, observed as micronuclei containing whole chromosomes, has been reported (Borojerdi et al., Mut Res, 2009). We have seen a loss of β-tubulin polymerization in about 23% of normal human mammary epithelial cells (NHMECs) exposed for 24 hr to 200 μM AZT, and demonstrated down-regulation of hsa-miR-770-5p, a microRNA known to target STMN1, in the same AZT-exposed cells. STMN1 prevents microtubule polymerization, resulting in destabilization of the mitotic spindle and chromosome misalignment. We therefore hypothesized that, if AZT exposure downregulates hsa-miR-770-5p, there should be increased expression of STMN1, which would result in loss of microtubule polymerization leading to aneuploidy. In this study we exposed cells of the human breast line MCF-10A to 100 μM AZT continuously for 1 to 3 passages (P). By Western Blot we showed that total STMN1 was 3.5-fold increased at P1, in AZT-exposed cells, compared to the unexposed controls. Using the same STMN1 antiserum, by immunohistochemistry (IHC), we observed that some cells stained heavily for STMN1, while others had virtually no staining, and that the overall STMN1 fluorescence was increased in AZT-exposed cells, compared to unexposed controls. When tubulin polymerization was evaluated by IHC, using β-tubulin antiserum, 10.1% of AZT-exposed cells showed a loss of tubulin polymerization, while only 0.7% of unexposed cells showed the same effect (p=0.004). The data suggest that AZT induced aneuploidy may be mediated by inhibition of hsa-miR-770-5p, upregulation of STMN1, impaired polymerization of β-tubulin, aberrant mitotic spindle formation and consequent chromosome misalignment resulting in aneuploidy. If confirmed, this will constitute a novel mechanism of AZT-induced genotoxicity that may or may not be related to AZT-DNA incorporation. Ongoing experiments will explore these relationships further by localizing STMN1 and β-tubulin in specific cells, and examining the same cultures for spindle aberrations at mitosis. Citation Format: Andrea V. Rivera, Vanesa C. Sanchez, Miriam C. Poirier, Ofelia A. Olivero. Zidovudine (AZT)-induced aneuploidy, mediated by Stathmin 1 (STMN1), involves a loss of polymerized β-tubulin. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3603. doi:10.1158/1538-7445.AM2013-3603