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ENGENUR - Definition of new ENdpoints to assess and discriminate GENotoxic effects resulting from environmental exposures
Coordinator - Sónia Mendo
Programme - Fundação para a Ciência e a Tecnologia- FEDER- COMPETE PTDC/AAC-AMB/114057/2009
Execution dates - 2011-01-01 - 2014-01-01 (36 Months)
Funding Entity - FCT
Funding for CESAM - 108.693 €
Total Funding - 172.945 €
Proponent Institution - Universidade de Aveiro
Participating Institutions
Centro de Histocompatibilidade do Centro
Instituto Tecnológico Nuclear

Until the beginning of the 21st century, 60 uranium and radium (226Ra) mines were exploited in Portugal. The mining residues, often containing high concentrations of radionuclides, were stockpiled at the surface generally near the uranium mines (Carvalho et al., 2007). Other uranium deposits do exist and several mining companies recently applied for mining licences. Therefore, uranium mining industry may resume activities in the near future. Amongst mining activities, uranium mining is one of high environmental concern, as it constitutes a source of chemical contamination of groundwater resources and adjacent soils and, moreover, radioactivity in mining waste may expose biota to detrimental doses of ionizing radiation (Antunes et al., 2008a; Antunes et al., 2007b; Carvalho et al., 2007 a, b; Pereira et al., 2008; Pereira et al., 2004a). Resident populations in uranium mining areas may be exposed to radiation, metal/radionuclides rich-wastes, as well as, to contaminated water and food products. When the risk assessment of radiological contaminated sites is carried out, information about indicator species may be used to assess adverse effects to species themselves, to other components of the ecosystem, and to humans (Burger et al., 2000). They are also important to support decisions about cleanup actions and, to evaluate the success of past remediation measures (Burger et al., 2000). The bioaccumulation of radioactive substances is responsible for cellular and genetic damages that can seriously affect human health, as well as other important groups of organisms. Regarding uranium radiotoxic and chemotoxic properties, genotoxic responses seem to be a relevant endpoint, as uranium acts on the formation of oxidative DNA damages (Barillet et al., 2005). Few data are available concerning the correlation among genotoxic effects on humans with those recorded on other animal species, highlighting the importance of this study. Increasing our understanding about these correlations, we will be able to use bioindicator species, to characterize human exposures and to predict, in due time, genotoxic effects. Moreover, we will be able to integrate evaluations of risks to humans and to ecosystems,what is crucial to propose mitigation measures, designed to reduce human health risks and simultaneously, to restore the ecosystem. With our experience in the environmental risk assessment of contaminated areas (Antunes et al., 2008a; Antunes et al., 2008b; Antunes et al., 2007a; Antunes et al., 2007b; Marques et al., 2009; Pereira et al., 2008; Pereira et al., 2006; Pereira et al., 2004a; Pereira et al., 2004b), we have concluded that in general, standard ecotoxicological sub-lethal assays or the in situ assays, developed by our team (Antunes et al., 2008a; b;) are not adjusted for sites contaminated with radioactive wastes, as they do not include the evaluation of endpoints related with genotoxic effects. Hence, their application may contribute to underestimate the risks, and to lose information about genotoxic responses of exposed organisms. Furthermore, this team has already validated two vertebrate species, as bioindicators of metals exposure and genotoxicity in two mining areas (Pereira et al., 2006, Marques et al., 2009), nevertheless we think that we still need, for different species, the validation of methods/techniques which have been developed to assess genotoxic effects in humans, in order to help us to perceive mechanisms of action of genotoxic agents in those species and to increase our knowledge about similarities and dissimilarities with humans. Bearing in mind our poor knowledge about all of these aspects, this proposal aims to focus four main aspects: i) development and the validation of new/existing techniques/methods to assess genotoxic responses on different wildlife and domestic species, ii) development of new endpoints to be assessed in standard ecotoxicological assays, usually applied to assess risks to terrestrial communities. But with such improvement will become useful, as well, as an alarm for genotoxic effects induced by environmental exposures. iii) validation of new bio-indicator wildlife and domestic species for exposures to metals, radioactive elements and radiation, which within a risk assessment scenario, can help in predicting exposure and effects on humans, and iv) comprehension of mechanisms of genotoxic effects yield by metals/radionuclides and radiation, under environmental exposure, to analyze similarities with those already know for humans. In this proposal some simple and cost effective techniques like comet assay and flow cytometry will be used, since they can be very useful for uranium/radionuclides genotoxicity evaluation. Expression analyses of genes involved in the carcinogenesis process will be performed by Real-time PCR. These genes are of great importance, as they provide answers about uranium capabilities to generate cell death and cancer. Mutation analyses will be carried out after amplification and amplicon sequencing (MWG, Germany) of all the relevant exons of each gene. Suppression Subtractive Hybridization (SSH) will be used to study the different gene expression levels along early-developmental stages of two wildlife vertebrate species. This proposal is presented by a team with vast expertise in the risk assessment of mining contaminated areas ( Fernando Gonçalves, Ruth Pereira, Sérgio Marques, Sara Antunes) from UA-Aveiro; in radiological protection and safety from ITN (Fernando Carvalho) and in molecular and cell biology from UA-Aveiro and CHC (Sónia Mendo and Artur Paiva respectively).





Sónia Mendo
Principal Investigator

CESAM Funding: