Evaluation of cytotoxic and genotoxic effects of functionalized nanocellulose in a co-culture system approaching the lung environment

Pinto, Fátima; Ventura, Célia; Lourenço, Ana Filipa; Ferreira, Paulo J.T.; Louro, Henriqueta; Silva, Maria João

http://hdl.handle.net/10400.18/7851

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Introduction: Nanocelluloses, obtained from different sources and by various methods and holding different functionalization are innovative environmentally friendly materials in both pure and composite forms that hold great promise for industrial or advanced biomedical applications. Based on previous knowledge on other nanofibers e.g., carbon nanotubes, revealing toxicity at cellular and organismal levels, there is a need to evaluate the toxic potential of new nanocelluloses, particularly cellulose nanofibrils (CNF), before entering the market. In this work, three different types of nanocelluloses produced from Eucalyptus globulus bleached kraft pulp but differing in important physicochemical properties such as the carboxyl content (CCOOH), degree of polymerization (DP), morphology, specific surface area and size of fibrils, were investigated regarding cyto- and genotoxic effects in human alveolar cells and in a co-culture of human alveolar cells (A549 cells) and THP-1 differentiated macrophages. Results: A preliminary evaluation of the cytotoxicity in A549 by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay revealed that all nanocellulose samples showed no toxicity over the examined concentration range during a 24h exposure period. The genotoxicity of the nanocellulose samples was assessed through the cytokinesis-blocked micronucleus assay in A549 co-cultured with THP-1 cells and exposed to 1.5, 3, 6, 12.5, and 25 μg/cm2 of each nanocellulose for 24h. Preliminary results showed no alterations in the frequency of micronucleated binuceated cells for all tested concentrations in one sample of CNF produced with a catalytic oxidation with TEMPO (2,2,6,6-tetramethylpiperidine-1-oxyl) radical (T1300). Moreover, the cytokinesis-block proliferation index of exposed A549 cells did not show differences as compared with non-exposed cells, in line with the MTT results, suggesting that the tested nanocelluloses do not affect A549 cells proliferation. Conclusions: Preliminary results suggest that T1300 sample do not present cytotoxicity or genotoxicity in the examined concentration range and exposure time, as compared to controls. The other two samples are still being investigated.

Summary: Three nanocellulose samples were investigated for their in vitro toxicity. No major cytotoxic or genotoxic effects were detected in alveolar cells co-cultured with THP-1 macrophages.