Analysis of the In Vitro Toxicity of Nanocelluloses in Human Lung Cells as Compared to Multi-Walled Carbon Nanotubes

Cellulose micro/nanomaterials (CMNM), comprising cellulose microfibrils (CMF), nanofibrils (CNF), and nanocrystals (CNC), are being recognized as promising bio-nanomaterials due to their natural and renewable source, attractive properties, and potential for applications with industrial and economical value. Thus, it is crucial to investigate their potential toxicity before starting their production at a larger scale. The present study aimed at evaluating the cell internalization and in vitro cytotoxicity and genotoxicity of CMNM as compared to two multi-walled carbon nanotubes (MWCNT), NM-401 and NM-402, in A549 cells. The exposure to all studied NM, with the exception of CNC, resulted in evident cellular uptake, as analyzed by transmission electron microscopy. However, none of the CMNM induced cytotoxic effects, in contrast to the cytotoxicity observed for the MWCNT. Furthermore, no genotoxicity was observed for CNF, CNC, and NM-402 (cytokinesis-block micronucleus assay), while CMF and NM-401 were able to significantly raise micronucleus frequency. Only NM-402 was able to induce ROS formation, although it did not induce micronuclei. Thus, it is unlikely that the observed CMF and NM-401 genotoxicity is mediated by oxidative DNA damage. More studies targeting other genotoxicity endpoints and cellular and molecular events are underway to allow for a more comprehensive safety assessment of these nanocelluloses.This research received funding from the Portuguese Foundation for Science and Technology (FCT/MCTES), through national funds (PIDDAC) under the project ToxApp4NanoCELFI (PTDC/SAUPUB/32587/2017). The following financial grants are also acknowledged: Projects UIDB/00009/2020 and UIDP/00009/2020 (ToxOmics); Projects UIDB/04138/2020 and UIDP/04138/2020 (iMed.Ulisboa) and L. Gonçalves Principal Researcher grant (CEECIND/03143/2017); FEDER and POR Lisboa2020 through the COMPETE 2020 POCI and PORL; POCI-01-0145-FEDER-007688 (UIDB/50025/2020-2023); NanoCell2SEC (PTDC/CTM-REF/30529/2017); CIEPQPF (UIDB/00102/2020); European Topology Interdisciplinary Action (EUTOPIA CA17139); Inpactus Project—Innovative Products and Technologies from Eucalyptus (Project Nº 21874, funded by Portugal 2020 through ERDF in the frame of COMPETE 2020 Nº246/AXIS II/2017). Nádia Vital holds an FCT/MCTES PhD Scholarship grant (2020.07168.BD)