Nanotechnology is an emerging field that involves the development, manufacture and measurement of materials and systems in the submicron to nanometer range. Its development is expected to have a large socio-economical impact in practically all fields of industrial activity. However, there is still a lack of information about the potential risks of manufactured nanoparticles for the environment and for human health. In this work, we studied the cytotoxicity, genotoxicity and morphological transforming activity of cobalt nanoparticles (Co-nano) and cobalt ions (Co(2+)) in Balb/3T3 cells. We also evaluated Co-nano dissolution in culture medium and cellular uptake of both Co-nano and Co(2+). Our results indicated dose-dependent cytotoxicity, assessed by colony-forming efficiency test, for both compounds. The toxicity was higher for Co-nano than for Co(2) after 2 and 24 h of exposure, while dose-effect relationships were overlapping after 72 h. Statistically significant results were observed for Co-nano with the micronucleus test and the comet assay, while for Co(2+) positive results were observed only with the latter. In addition, even when Co-nano was genotoxic (at >1 microM), no evident dose-dependent effect was observed. Concerning morphological transformation, we found a statistically significant increase in the formation of type III foci (morphologically transformed colonies) only for Co-nano. Furthermore, we observed a higher cellular uptake of Co-nano compared with Co(2+).

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Chromium compounds, besides being occupational carcinogens, can also induce allergic contact dermatitis (ACD) and other immunomodulatory effects. In this study we investigate cell viability, uptake and intracellular distribution of chromium in human primary dendritic cells (DCs), either immature (iDCs) or driven to differentiate by a specific maturation stimulus (LPS) (mature DCs, mDCs), when exposed for 48 h to concentrations of soluble radiolabelled NatCr04 ranging from 5 to 0.5 /lM. The modulation of the expression of membrane markers (CD80, CD86, MOC class II) correlated with the immunological functions of DCs was also measured. After 48 h of exposure the mean IC so values in 4 donors were 36 and 31 /lM in iDCs and mDC respectively, as detected by propidium iodide incorporation. Cellular uptake of chromium was nearly linear with increasing doses. At 48 h post-exposure chromium was accumulated preferentially in the nuclear and cytosolic fractions (44.1 to 66% and 13.1 to 31% of total cellular chromium, respectively). Although a high inter-individual variability was observed, an increase in the expression of CD86 and, to a lower extent, CD80 and MOC class II membrane markers was found in mDCs of single donors. These results highlight the relevance of searching for the biodistribution of trace metals in primary cells of the immune system. Moreover, they suggest that DCs differentiation markers can help in measuring the immunotoxicity of metalIn the last decades a tremendous amount of research effort has been dedicated to understanding the human health effect of Cr(VI) compounds and the molecular mechanisms by which they occur (I). Nevertheless, several questions related to chromium toxicity and carcinogenicity are still unclear and only partly understood. In particular, concerning the effects of chromium on the immune system, similarly to other trace metals which can have immunostimulatory, immunosuppressive or autoimmunity effects, depending on the dose (2) Cr(VI) compounds at low doses, have in vitro a stimulatory effect on human lymphocytes whereas at higher concentrations they display an inhibitory effect (3-4). Moreover, dosage seems to be non-influent on NK cell functions (5). On the other hand, no effect of chromium was observed on the proliferation of the lymphocytes of workers chronically exposed to welding fumes (6). Furthermore, higher doses of Cr(VI) depress the phagocytic activity of alveolar macrophages, whereas lower doses stimulate this biological

The recent European Commission REACH (Registration, Evaluation and Authorisation of Chemicals) policy outlines a plan for toxicological testing by using alternative non-animal in vitro methods. In this context, there is a need to develop and standardise high-throughput screening (HTS) methods for studying the cytotoxicity induced by chemicals. Electrochemical impedance spectroscopy (EIS) can be considered as a complementary technique to alternative in vitro testing for studying cell adhesion to the substrate, and can give real-time and kinetic information on cell responses to a toxicant. This paper describes the development of a home-made chip based on impedance spectroscopy, and its application in studying the kinetics of BALB/3T3 cell adhesion and the cellular responses to a toxic product as a function of time. Concentrations of sodium arsenite, ranging from 10μM up to 1000μM, were tested in the system, and the results were compared with those obtained with standard protocols used to study basal cytotoxicity induced by chemicals in the BALB/3T3 cell line. The results show that the sensitivity of the developed chip was better than that with the MTT test, with the additional advantages of online monitoring.

The Balb/c3T3 cell transformation assay (CTA) is an available in vitro system to detect the carcinogenic potential of chemicals. Currently, the European Centre for the Validation of Alternative Methods (ECVAM) is validating this test, assessing its reliability and relevance. Its endpoint is the formation of type III foci, which is, when using clone A31-1-1, a very rare event that usually does not occur at all for negative controls. The carcinogenic potential of a compound tested is assessed by comparing the number of foci in treated and untreated cells. The objective of the present work is to optimise the data analysis for this endpoint by applying the most commonly used approach by a t-test and the Fisher's exact test as an alternative approach. For this purpose selected metal compounds classified as carcinogenic (NaAsO2, CdCl2, cisPt), as suspected carcinogenic (C6H5)4AsCl, CH3HgCl), or as compounds without evidence of carcinogenic properties in humans ((NH4)2PtCl6, NaVO3) as well as a non-carcinogenic (AgNO3) were analysed. Our evaluation revealed that the t-test approach, which assumes normality of data, is not appropriate. The results demonstrated that the statistical analysis by Fisher's exact test, better reflecting the data properties, greatly facilitates the interpretation of Balb/c3T3 CTA data regarding carcinogenic potential.