Browsing by Author "Saruga, Andreia"
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- Biological impact of metal nanomaterials in relation to their physicochemical characteristicsPublication . Louro, Henriqueta; Saruga, Andreia; Santos, Joana; Pinhão, Mariana; Silva, Maria JoaoSeveral metal and metal oxide nanomaterials (NMs), e.g., cerium dioxide NMs(CeO2), barium sulphate NMs(BaSO4) and titanium dioxide NMs(TiO2), display advantageous properties over the bulk materials and have a broad range of innovative applications in food, industry and consumer products. Whether these materials are hazardous and impact on human health or the environment remains an issue that needs to be addressed by reliable studies focused on nano-bio interactions. To contribute to the comprehensive investigation of the toxicological effects of metal NMs, we have assessed the cytotoxic and genotoxic effects of benchmark NMs in human respiratory cells, concomitantly with the analysis of their secondary properties in the cellular moiety. This study shows no effects of BaSO4, while some, but not all, of the other metal-related NMs analyzed have adverse effects. Human respiratory cells were prone to CeO2 cytotoxicity and to DNA damage induction following exposure to anatase TiO2 (NM-100, NM-101 and NM-102), but not rutile TiO2. No clastogenic/aneugenic effects were ascribed to any of the tested NMs. Using correlation analysis, this work also suggests that among these TiO2, the size in the cellular moiety may be the most relevant secondary feature that determines their biological consequences.
- Cyto- and Genotoxicity assessment of manufactured nano cerium dioxide in the A549 cell linePublication . Saruga, Andreia; Louro, Henriqueta; Silva, Maria JoãoIn the past decades the growing application of nanomaterials (NMs) in diverse consumer products has raised various concerns in the field of toxicology. They have been extensively used in a broad range of applications and cover most of the industrial sectors as well as the medicine and the environmental areas. The most common scenarios for human exposure to NMs are occupational, environmental and as consumers and inhalation is the most frequent route of exposure, especially in occupational settings. Cerium dioxide NMs (nano-CeO2) are widely used in a number of applications such as in cosmetics, outdoor paints, wood care products as well as fuel catalysts. For such reason, nano-CeO2 is one of the selected NMs for priority testing within the sponsorship program of the Working Party of Manufactured Nanomaterials of the OECD. In this context, the aim of this study is to assess the safety of nano-CeO2 (NM-212, Joint Research Center Repository) through the characterization of its cytotoxicity and genotoxicity in a human alveolar epithelial cell line. A dispersion of the NM in water plus 0.05% BSA was prepared and sonicated during 16 minutes, according to a standardized protocol. DLS analysis was used to characterize the quality of the NM dispersion in the culture medium. To evaluate the cytotoxicity of nano-CeO2 in the A549 cell line, the colorimetric MTT assay was performed; the capacity of cells to proliferate when exposed to CeO2 was also assessed with the Clonogenic assay. The genotoxicity of this NM was evaluated by the Comet Assay (3 and 24h of exposure) to quantify DNA breaks and the FPG-modified comet assay to assess oxidative DNA damage. The Cytokinesis-Block Micronucleus (CBMN) assay was used to further detect chromosome breaks or loss. The nano-CeO2 particles are spherical, displaying a diameter of 33 nm and 28 m2/g of surface area. The results of the MTT assay did not show any decreased in cells viability following treatment with a dose-range of nano-CeO2 during 24h. Nevertheless, the highest concentrations of this NM were able to significantly reduce the colony forming ability of A549 cells, suggesting that a prolonged exposure may be cytotoxic to these cells. Data from both genotoxicity assays revealed that nano-CeO2 was neither able to induce DNA breaks nor oxidative DNA damage. Likewise, no significant micronucleus induction was observed. Taken together, the present results indicate that this nano-CeO2 is not genotoxic in this alveolar cell line under the tested conditions, although further studies should be performed, e.g., gene mutation in somatic cells and in vivo chromosome damage (rodent micronucleus assay) to ensure its safety to human health.
- Cyto- and Genotoxicity Assessment of Manufactured Nanomaterials in the A549 Cell LinePublication . Saruga, Andreia; Silva, Maria João; Louro, Maria HenriquetaA number of nanomaterials (NMs) have been applied in different fields due to their unique physico-chemical properties. As the use and applications have increased in some industries, serious concerns about their potential impact on the environment and the human health have been raised and have been a challenge for the regulatory authorities. This work aimed at assessing the toxicity of three classes of NMs, namely cerium dioxide, CeO2 (NM-212), titanium dioxide, TiO2 (NM-101 and NM-100) and barium sulphate, BaSO4 (NM-220) since they already have a broad range of applications in industry and consumer products. A standardized protocol for NMs dispersion was followed and the quality of the dispersion in the culture medium was evaluated by the dynamic light scattering technique. Different concentrations (0, 1, 3, 10, 30, 75 and 100 μg/cm2) of each nanomaterial were used to expose A549 cells (human lung carcinoma cells) for cytotoxic evaluation through the MTT and clonogenic assays and genotoxicity assessment through the comet and the cytokinesis-blocked micronucleus (CBMN) assays. The results showed a decrease in cell proliferation after exposure to cerium dioxide nanomaterials for 8 days, at the highest concentrations tested and a slight increase in the level of DNA breaks. Concerning the TiO2 NMs, a statistically significant increase in the level of DNA breaks was found for both NMs; however the CBMN assay did not show any increase in the frequency of chromosomal breaks. BaSO4 was the NM that showed the lowest toxicity in cyto- and genotoxicity assays.Even though the present results contribute to assess the hazard of the tested NMs, the real effects of nanomaterials’ exposure to human health are still unclear and an unequivocal conclusion is difficult to present, given the inconsistent and often conflicting results found in the literature. Thus, the application of some nanomaterials in consumer products should be carefully evaluated until definite conclusions about their safety are available.
- Hazard assessement of metallic nanomaterials in human respiratory cellsPublication . Saruga, Andreia; Louro, Henriqueta; Santos, Joana; Silva, Maria JoãoAs the applications of nanomaterials (NM) have been exponentially increasing, serious concerns about their potential impact on human health have been raised and challenging questions to regulators came up. This work investigated the toxicity of widely used metallic NM, namely cerium dioxide (CeO2, NM-212), titanium dioxide (TiO2, NM-100 and NM-101) and barium sulphate (BaSO4, NM-220). A standardized protocol for NM dispersion was followed and the quality of the dispersion in the culture medium was evaluated by dynamic light scattering. A human alveolar cell line (A549) was exposed to 1-100 μg/cm2 of each NM for cytotoxicity (MTT and clonogenic assays) and genotoxicity (comet and micronucleus assays) assessment. A decrease in cells’ proliferative capacity was detected after exposure to the two highest concentrations of CeO2 for 7 days (p=0.01 and p=0.002, respectively) while the remaining NM were not cytotoxic. Concerning genotoxicity, TiO2 NM significantly increased the level of DNA breaks but those lesions seemed to be efficiently repaired because no chromosome instability was detected by the micronucleus assay. The CeO2 NM induced a two-fold (non-significant) increase of in the level of oxidative DNA. BaSO4 NM was neither cytotoxic nor genotoxic under the tested conditions. Although the present results contribute to the risk assessment of these NM, the real effects from human exposure, e.g., in the workplace, are still unclear. Thus, the implementation of high throughput methodologies to allow cost-efficient strategies and experimental models that better mimic in vivo responses is an urgent need to allow nanosafety studies to keep pace with innovation.
- Hazard assessment of benchmark metallic nanomaterials in alveolar epithelial cellsPublication . Saruga, Andreia; Louro, Henriqueta; Pinhão, Mariana; Silva, Maria JoãoThe fast development of nanotechnology has led to the manufacturing of a wide array of nanomaterials (NMs). Despite the number of studies addressing NMs toxicity, uncertainties about their safety remain, representing a challenge to regulatory authorities. This work intended to assess the toxicity of metallic NMs in alveolar epithelial cells and to relate the effects with their physicochemical properties. Benchmark NMs (JRC) - CeO2 (NM-212), TiO2 (NM-100) and BaSO4 (NM-220) - were dispersed and their properties in the culture medium were evaluated by DLS. A549 cells were exposed to each NM for cytotoxicity (MTT and plating efficiency assays) and genotoxicity (comet and cytokinesis-blocked micronucleus, CBMN, assays) assessment. A homogeneous dispersion that remained stable in culture medium was achieved for all NMs. The CeO2NM was the only one that decreased cells’ proliferative capacity after 8 days exposure. As to the genotoxicity, the TiO2NM significantly increased the level of DNA damage following 3h and 24h exposure, whereas the CeO2NM caused only a slight increase in DNA damage at 3h exposure. None of the NMs tested positive by the CBMN assay. BaSO4NM was neither cytotoxic nor genotoxic. In conclusion, this study contributed to the hazard assessment of different benchmark metallic NMs, disclosing diverse biological effects that will be interpreted considering the inherent physicochemical properties. The identification of key features and pathways that drive NMs’ toxicity is paramount to allow prediction of their adverse effects, avoiding the huge task of testing every new NM.
- Nanotoxicology and nanotechnology interplay towards a safe and responsible innovationPublication . Silva, Maria João; Ventura, Célia; Dias, Kamila; Saruga, Andreia; Pinhão, Mariana; Louro, HenriquetaThe European Union Strategic and Action Plan for Nanosciences and Nanotechnologies emphasizes the need of ensuring the development of safe, integrated, and responsible nanotechnologies and nanotechnology products. Indeed, although the use of manufactured nanomaterials (NM) is widespread and has been considered greatly beneficial for several scientific and technological areas, uncertainties about their safety still remain, representing a challenge to scientists, risk assessors and regulators. Recent studies of the team have been aimed at assessing the potential deleterious effects, e.g., genotoxicity that is intimately associated with carcinogenicity, of several classes of NM, using complementary in vitro and in vivo approaches, nested within the conventional risk assessment paradigm. Overall, the results have suggested that specific physicochemical properties of NM are crucial to define their nano-bio interactions and their toxic potential. Furthermore, our findings reinforced previous reports suggesting that factors inherent to experimental conditions and methodologies may affect the effects observed. These and other nano-specific issues have been hindering the categorization of NM according to their toxicity and, consequently, their risk assessment and management. Nonetheless, considering the wide array of NM produced or under development, a case-by-case approach to the risks of each NM seems an unreasonably extensive task. Thus, the development of high-throughput omics-based tools adapted for the toxicity assessment of NM and, on the other hand, the elucidation of the cellular and molecular mechanisms underlying NM toxicity is of utmost importance to progress faster in the field of nanotoxicology. Such information will allow the development of cost-efficient screening strategies and promote predictive nanotoxicology, driving the synthesis of safer NM. The ultimate goal is fostering a sustainable and safe innovation in the field of nanotechnologies.
- Tackling the uncertainties of emergent nanomaterials for public health: are metal nanoparticles hazardous pollutants?Publication . Louro, Henriqueta; Saruga, Andreia; Pinhão, Mariana; Santos, Joana; Silva, Maria JoãoWhen materials are produced at the nanoscale range, their surface chemistry changes, chemical reactivity increases and other important material properties may be altered, including optical, magnetic and electrical properties. Nanomaterials (NMs) display advantageous characteristics that enable innovative applications, as is the case of several metal and metal oxides such as titanium dioxide nanomaterials (TiO2), cerium dioxide (CeO2) and barium sulphate (BaSO4), that already have a broad range of applications in industry and consumer products. However, the exponential development of nanotechnologies contrasts with the insufficient risk assessment for human health and the environment and reliable studies focused on nano-bio interactions are necessary1. With the aim of contributing to a comprehensive investigation of the hazard of metal oxides NMs in human respiratory cells, we have used cytotoxicity and genotoxicity assays2, concomitantly with the analysis of the NMs’ properties in the cellular moiety. The results show that only CeO2 has cytotoxic effects and TiO2 is genotoxic in the comet assay, but not clastogenic in the micronucleus assay. The remaining NMs are not genotoxic. The present results contribute to evaluate the hazard of the tested NMs, suggesting that some, but not all, metal NMs affect human respiratory cells. Considering the reports describing workers occupationally exposed to TiO23, this finding raises concerns for human health. Further studies are described that were designed to understand if this finding could be generalized to other TiO2 that are frequently used in consumer products. For tackling NMs’ uncertainties, an in-depth investigation of the nano-bio interactions must be foreseen, where correlation analysis with the primary and secondary physicochemical characteristics may enable grouping strategies valuable for risk assessment.