Untitled

Funder

Organizational Unit

Publications

The use of comet assay to assess global DNA methylation in human biomonitoring studies

Publication . Costa, Carla; Alves, Ana Catarina; Costa, Solange; Soares, Amadeu M.V.M.; Monteiro, Marta S.; Loureiro, Susana; Teixeira, João Paulo

The Comet assay is a valuable tool for the detection of DNA damage in genotoxicity and human biomonitoring studies. Throughout the years, this biomarker has undergone several adaptations in their protocol in order to increase its sensitivity and the possible outcomes. By including an additional step of DNA digestion with lesion-specific endonucleases, the comet assay can provide information regarding the type of DNA damage detected in cells. The use of these enzymes has also allowed the development of a methylation-sensitive modified version of the comet assay. This version enables the routine measurement of global methylation, as well as CpG island DNA methylation in a variety of cells while simultaneously determining the genetic integrity of examined cells (Wentzel, 2012). Briefly, it makes use of isochizomeric restriction enzymes HpaII and MspI (that display differential sensitivity to DNA methylation) to characterize methylation outside CpG islands and restriction enzyme NotI to determine DNA methylation in CpG islands. The technique has been recently adapted to a medium-throughput version (Lewies, 2014) that allows the simultaneous analysis of a larger number of samples and overcomes some technical problems. Nevertheless, this technique has not yet been carried out in human biomonitoring studies. In this context, the aim of this work was to make use of this version of the comet assay to characterize global DNA methylation in approximately 50 human samples. Samples were analysed by the methylation-sensitive modified version of the comet assay (medium-throughput) and by ELISA based assay. Data obtained with both methods were compared and reproducibility of the methylation-sensitive modified version of the comet assay determined. Results obtained contribute to knowledge on the feasibility of this version of the comet assay and its possible usage in human biomonitoring studies as an epigenetic biomarker.

2015-09Conference object

Open access

Protective effect of Crataegus monogyna Jacq. ethanolic extracts in oxidant-induced DNA damage evaluated through comet assay with human peripheral lymphocytes

Publication . Barreira, Joao; Costa, Carla; Teixeira, João Paulo; Ferreira, Isabel; Oliveira, Beatriz

Much attention of preventive medicine research is focused on natural antioxidants. This interest refers not only to isolation and identification of new biologically active molecules for the pharmaceutical industry, but also because of the emergent public interest in using crude plant extracts, such as infusions for self-medication (Krishnaiah et al., 2011). The use of antioxidants, such as the well-known polyphenolic compounds, to prevent genetic damage induced by physical or chemical agents is of considerable interest. This bioactivity might be related to their anticlastogenic effect, due to the presence of specific functional groups. Other antioxidant compounds, such as vitamins C and D, were reported for their DNA-damage decreasing effect, suggesting that reactive oxygen species may be involved in this activity (Benavente-García et al., 2004). Evaluating the antioxidant activity of natural matrices represents one of our primary research challenges (Ferreira et al., 2009). Among hundreds of studied species, Crataegus monogyna Jacq. stood out as being one of the most promising plants due to its high bioactivity. Besides the antioxidant activity, C. monogyna was also studied for the human tumour cells growth inhibitory capacity of its phenolic extracts; furthermore, individual phenolic compounds were fully characterized by high performance liquid chromatography-photo diode array detection-electrospray ionization tandem mass spectrometry (HPLC-DAD-ESI/ MS), revealing high levels of flavonols, flavones, hydroxycinnamic acid derivatives and anthocyanins (Rodrigues et al., 2012). However, there is a high limitation in examining if the detected bioactivity is actually transferred from in vitro to in vivo systems (Carocho and Ferreira, 2013).

2015-09Conference object

Open access

Risk-Benefit Assessment in foods: a tool for a better food and health policy in Europe

Publication . Assunção, R.; Martins, C.; Brazão, R.; Fernandes, P.; Carvalho-Oliveira, I.; Torres, D.; Lopes, Carla; Carvalho, C.; Vasconcelos, F.; Monteiro, S.; Nabais, P.; Carmona, P.; Persson, M.; Thomsen, S.; Jakobsen, L.; Pires, S.; Poulsen, M.; Membré, JM; Boué, G; Alvito, Paula

RiskBenefit4EU – Partnering to strengthen the risk–benefit assessment within EU using a holistic approach, is a recent European pilot project funded by EFSA and coordinated by Portugal (PT), integrating a multidisciplinary team from health and food institutes, national food safety authorities, R&D institutions and academia from PT, Denmark (DK) and France (FR). The main objectives of RiskBenefit4EU concerns the development of a set of Risk–Benefit Assessment (RBA) tools to assess and integrate food risks and benefits in the areas of microbiological, nutritional and chemical components through the development of a harmonised framework. This pilot project will validate the RBA framework created using a Portuguese case study on cereal-based foods. The research idea for food safety in risk assessment is to create an international network on RBA to promote and disseminate the outputs and knowledge acquired under RiskBenefir4EU, at European level. This network aims to promote knowledge and capacity building on RBA (acquired under RiskBenefit4EU) among European early stage researchers and to apply the harmonised framework on their countries. Health risks associated with consumption of cereal-based foods, an important source of nutrients with beneficial health effects, could increase soon due to climate changes in Europe (dry conditions and increased ambient temperatures could promote an increase in toxins production; occurrence of emergent compounds) so the dissemination and use of the RBA harmonized tools related with ingestion of cereal-based foods and derivatives could contribute to support future food and health policy in Europe.

2018-01-17Conference object

Open access

Building capacity in risk-benefit assessment of foods: lessons learned from the RB4EU project

Publication . Assunção, Ricardo; Alvito, Paula; Brazão, Roberto; Carmona, Paulo; Fernandes, Paulo; Jakobsen, Lea S.; Lopes, Carla; Martins, Carla; Membré, Jeanne-Marie; Monteiro, Sarogini; Nabais, Pedro; Thomsen, Sofie T.; Torres, Duarte; Viegas, Silvia; Pires, Sara; Boué, Geraldine

Background: Human diet may present both risks and benefits to consumers’ health. Risk-benefit assessment of foods (RBA) intends to estimate the overall health impact associated with exposure (or lack of exposure) to a particular food or food component. Scope and approach: “RiskBenefit4EU – Partnering to strengthen the risk-benefit assessment within EU using a holistic approach” (RB4EU) is a project funded by the European Food Safety Authority (EFSA) that integrates a multidisciplinary team from Portugal, Denmark and France. This project aims to strengthen the EU capacity to assess and integrate food risks and benefits regarding toxicology, microbiology and nutrition. One of the specific objectives of RB4EU was to build capacity in RBA among the recipient partners from Portugal. In order to achieve this objective, a capacity building strategy including theoretical and hands-on training and the development of a case-study were established. This paper aims to present the strategy used in the RB4EU project to build capacity within RBA, including the main training approaches and the lessons learned. Key findings and conclusions: The capacity-building program included three main activities: theoretical training, focusing on RBA concepts; hands-on training, applying the acquired concepts to a concrete case-study, using the methods and tools displayed; and scientific missions, intending to provide advanced training in specific domains of RBA. The developed strategy can be used in the future to build capacity within RBA.

2019-07Journal article

Restricted access

Caraterização in vitro dos efeitos tóxicos da patulina na integridade do epitélio intestinal e potenciais efeitos protetores da cisteína

Publication . Assunção, Ricardo; Martins, Carla; Ferreira, Mariana; Alvito, Paula

A mucosa intestinal é a primeira barreira biológica encontrada pelas micotoxinas presentes nos alimentos, sendo a patulina, uma micotoxina produzida por fungos do género Penicillium spp., uma preocupação particular atendendo a que a exposição humana a esta micotoxina pode conduzir a efeitos imunológicos, neurológicos e gastrointestinais. Considerando estes efeitos para a saúde, o presente estudo tem como objetivos a avaliação do efeito tóxico da exposição intestinal a patulina, bem como a determinação do potencial efeito protetor da coadministração de patulina e cisteína na membrana intestinal, utilizando para o efeito células Caco-2. A integridade da membrana intestinal foi determinada pela medição da resistência elétrica transepitelial (TEER). Os resultados evidenciaram um decréscimo acentuado nos valores de TEER após 24 horas de exposição celular a 95 μM de patulina. Para as concentrações mais reduzidas verificou-se uma redução máxima inferior a 25% após 24 horas de exposição. A coadministração de patulina (95 μM) e cisteína (40 μM) revelou um decréscimo nos valores de TEER. O tratamento com cisteína em concentrações superiores ( 400 μM) revelou efeito protetor da membrana intestinal, tendo em conta os valores de TEER. Estes resultados contribuem para uma avaliação do risco mais precisa associada à exposição a contaminantes alimentares.

2016-07Journal article

Open access