Browsing by Author "Bettencourt, Ana Francisca"
Now showing 1 - 3 of 3
Results Per Page
Sort Options
- Analysis Of The Cytotoxicity And Genotoxicity of Digested Titanium Dioxide Nanomaterials (TiO2) In Intestinal CellsPublication . Louro, Henriqueta; Vieira, Adriana; Gramacho, Ana Catarina; Rolo, Dora; Vital, Nádia; Martins, Carla; Assunção, Ricardo; Alvito, Paula; Gonçalves, Lídia; Bettencourt, Ana Francisca; Silva, Maria JoãoTitanium dioxide nanomaterials (TiO2) have been frequently applied as food additives, in pharmaceuticals and in personal care products, such as toothpastes. Despite some regulators like EFSA concluded that the absorption of orally administered TiO2 is low, and that the use of TiO2 as a food additive does not raise a genotoxic concern, the presence of TiO2 in human organs was recently reported. This exposure may lead to adverse outcomes and has been poorly investigated. Furthermore, many of the biological effects of TiO2 described in the literature often overlook adequate physicochemical properties and their modification due to NMs interaction with the surrounding physiological matrices happening, e.g, during digestion. This work aimed to investigate in intestinal cells, the cyto- and genotoxic effects of TiO2 after the simulation of the human digestive process using the standardized INFOGEST in vitro digestion method, to better understand their potential negative impacts on the gastrointestinal tract. The TiO2 were characterized before and after digestion using DLS, zeta potential and TEM-EDS. The digestion product was used for cytotoxicity (MTT) and genotoxicity (comet, micronucleus) assays in two types of intestinal cells (Caco-2 and mucus secreting HT29-MTX cells). The results of the cytotoxicity and genotoxicity assays are discussed in view of the TiO2 secondary characteristics, to further understand the potential adverse intestinal outcomes in light of the transformation they suffer during digestion.
- Cellular effects of Titanium Dioxide Nanoparticles in the intestinePublication . Rolo, Dora; Pereira, Joana F.S.; Matos, Paulo; Gonçalves, Lídia; Bettencourt, Ana Francisca; Jordan, Peter; Silva, Maria João; Louro, HenriquetaIntroduction: The increased use of titanium dioxide nanoparticles (TiO2-NPs) as a food additive demands a thorough assessment of their potential risk for human health. Via oral exposure they may lead to adverse local or systemic outcomes, and few studies have focused on the cellular internalization mechanisms (endocytosis) of TiO2-NPs. The objective was to analyze the mechanisms by which TiO2-NPs (NM-102, NM-103 and NM-105, JRC repository) translocate by the intestinal epithelium layer, using monolayers of human intestinal cell lines (Caco-2 and HT29-MTX), as well as polarized Caco-2 cells, and co-cultures of both cells. Results: We evaluated cell differentiation by transepithelial resistance measurements and the translocation of TiO2-NPs tagged with alizarin through the intestinal barrier by confocal microscopy and we confirmed the internalization of the TiO2-NPs in both cell line models. Co-localization studies suggested that the smallest TiO2-NPs were internalized into EEA1-positive early-endosomes and accumulate in late endosomes (Rab7), with only a small fraction following the degradative pathway to the lysosome (LAMP1). This suggested that at least part of the TiO2-NPs could be redirected to the secretory pathway. Consistently, we detected fluorescence passing from the apical (AP) to the basolateral (BL) chamber, depending on the characteristics of cell model and TiO2-NPs tested. Conclusions: Small TiO2-NPs were endocytosed by Caco-2 cells, with an increase in particle diameter suggesting intracellular aggregation, whereas larger agglomerates deposited mainly extracellularly. Following endocytosis, TiO2 NPs were trafficked through different intracellular compartments including early and late endosomes/endo-lysosomes, with part being subjected to AP to BL transport.
- Titanium Dioxide Nanoparticles molecular effects: Internalization in the Human Intestinal EpitheliumPublication . Rolo, Dora; Pereira, Joana FS; Matos, Paulo; Gonçalves, Lídia; Bettencourt, Ana Francisca; Jordan, Peter; Silva, Maria João; Louro, HenriquetaThe use of titanium dioxide nanoparticles (TiO2-NPs) as food additive and in food packaging demands a thorough assessment of their potential risk for human health, specifically with regard to gastrointestinal tract. Using intestinal cells, we analyzed the mechanisms by which digested TiO2-NPs (NM-102, NM-103 and NM-105, JRC repository) translocate through the intestinal epithelium layer, as compared to non-digested particles. Human digestion of TiO2-NPs was simulated using the INFOGEST in vitro harmonized digestion method. Caco-2 cells were grown as polarized cell monolayer for exposure and differentiation was evaluated by TEER. The translocation of TiO2-NPs, tagged with alizarin red, through the cell barrier was analysed by confocal microscopy, using colocalization with antibodies against specific endosomal compartments. The internalization of the TiO2-NPs was confirmed for the three TiO2-NPs tested, both before and after digestion simulation. The smallest TiO2-NPs were internalized into EEA1-positive early- endosomes and accumulated in late endosomes (Rab7), with only a small fraction following the degradative pathway to the lysosome (LAMP1). The data suggested that at least part of the TiO2-NPs could be redirected to the secretory pathway. Consistently, fluorescence passing from the apical to the basolateral chamber was observed, without disruption of the intestinal barrier function. The changes in cell function or signal transduction pathways are being studied and possible consequences to human gastrointestinal tract arediscussed. We thank the support from P. Alvito, C. Martins and R. Assunção (INSA,Lisbon, Portugal).