Global DNA methylation and Reactive Oxygen Species assessment after in vitro exposure to novel cellulose nanomaterials on intestinal cells

Vital, Nádia; Ventura, Célia; Kranendonk, M.; Silva, Maria João; Louro, Henriqueta

http://hdl.handle.net/10400.18/8990

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Abstract(s)

Innovative cellulose nanomaterials (CNMs) find applications in multiple food-related products. However, it is recognized that nanomaterials (NMs) may cause adverse effects frequently by biological mechanisms that include production of intrinsic reactive oxygen species (ROS), or epigenetic alterations, by changes in gene expression through alterations in DNA methylation. To contribute to the safety assessment of two types of micro/nanofibrillated celluloses (CMF/CNFs) derived from industrial Eucalyptus globulus kraft, intracellular ROS levels and global DNA methylation patterns were evaluated using human intestinal cell models exposed to these CNMs. A harmonized protocol for in vitro simulation of human digestion was applied to incorporate the effect of potential changes in the physicochemical properties of CNMs due to the digestion process. After exposure of intestinal cells to digested and undigested CNMs samples (concentrations from 3.1 to 200 μg/mL, 3h and 24h), ROS levels were determined using the 2,7-dichlorofluorescein diacetate probe and fluorescence detection. The global DNA methylation was assessed after 24h exposure, using the methylated DNA Quantification Kit(colorimetric, Abcam) to quantify the levels of methylated cytosines (5-methylcytosine) in CpG dinucleotides. No increased ROS were observed after exposure to CNMs with and without digestion. Conversely, the preliminary results showed significant increased methylation of DNA after exposure to the CMF, without digestion, as compared to the negative control. No effect could be observed for digested sample, possibly due to the high background of the digestion product per se. Our preliminary findings seem to suggest the occurrence of biological effects of CNMs in intestinal cells that require further investigation, as may lead to a toxic impact on gastrointestinal cells, of concern for human health.

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Thanks to P.Alvito, R.Assunção and M.Silva(Food Safety Department, INSA) for support with the in vitro digestion.
Abstract disponível em: https://omp.uv.es/index.php/PUV/catalog/book/537