title: Characterization of in vitro genotoxic, cytotoxic and transcriptomic responses following exposures to amorphous silica of different sizes.
abstract: The objectives of the present study were to investigate the underlying mechanisms of genetic and cellular toxicity induced by silica nanoparticles (SiNPs) and determine if such toxicity is influenced by particle size. Commercially available amorphous SiNPs (12 nm, 5-10 nm, and 10-15 nm) and micrometer sized (SiP2 μm) silica were characterised for size, chemical composition, and aggregation state. Mouse lung epithelial (FE1) cells derived from Muta™Mouse were exposed to various concentrations (12.5, 25, 50, 100 μg/ml) of SiNPs and SiP2 μm. Cellular viability, clonogenic potential, oxidative stress, micronucleus formation, and mutant frequency were measured at different post-exposure time points. Cellular internalization of particles was assessed using nanoscale hyperspectral microscopy. Biological pathway and functional perturbations were assessed using DNA microarrays. Detailed characterization of particles confirmed their size, purity, and uniform dispersion in the exposure medium. Decreased cellular viability was observed acutely at 24h at concentrations higher than 25 μg/ml for all particle types, with SiNPs being the most sensitive; loss of viability was surface area dependent at the lowest concentration tested. However, only SiNP12 showed poor long-term survival. A size-dependent increase in micronucleus formation was also observed for SiNPs. In contrast to the viability results, SiP2 μm exhibited the highest potential to induce oxidative stress compared to the SiNPs at all tested concentrations. Gene ontology and biological pathway analysis revealed significant changes in the expression of genes implicated in lysosomal functions in SiNP12-treated cells, which appear closely associated with higher SiNP12 internalization and lysosomal rearrangements in the cytoplasm of these cells. These results suggest that SiNPs induce cellular and genetic toxicity in a size-dependent manner and that the observed toxicity may be the results of higher particle internalization of smaller SiNP and subsequent lysosomal overload.
authors.affiliation: Environmental Health Science and Research Bureau, Environmental and Radiation Health Sciences Directorate, Health Canada, Ottawa, Canada.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Decan
authors.affiliation: Environmental Health Science and Research Bureau, Environmental and Radiation Health Sciences Directorate, Health Canada, Ottawa, Canada.
authors.forename: Dongmei
authors.initials: D
authors.lastname: Wu
authors.affiliation: Environmental Health Science and Research Bureau, Environmental and Radiation Health Sciences Directorate, Health Canada, Ottawa, Canada.
authors.forename: Andrew
authors.initials: A
authors.lastname: Williams
authors.affiliation: New Substances Assessment and Control Bureau, Safe Environments Directorate, Health Canada, Ottawa, Canada.
authors.forename: Stéphane
authors.initials: S
authors.lastname: Bernatchez
authors.affiliation: Centre for Biologics Evaluation, Biologics and Genetic Therapies Directorate, Health Canada, Ottawa, Canada.
authors.forename: Michael
authors.initials: M
authors.lastname: Johnston
authors.affiliation: New Substances Assessment and Control Bureau, Safe Environments Directorate, Health Canada, Ottawa, Canada.
authors.forename: Myriam
authors.initials: M
authors.lastname: Hill
authors.affiliation: Environmental Health Science and Research Bureau, Environmental and Radiation Health Sciences Directorate, Health Canada, Ottawa, Canada. Electronic address: sabina.halappanavar@hc-sc.gc.ca.
authors.forename: Sabina
authors.initials: S
authors.lastname: Halappanavar
mesh_terms: D026921:Amino Acid Transport Systems, Neutral / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D055598:Chemical Phenomena; D004249:DNA Damage; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000235:genetics / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects; D005978:Glutathione / Q000378:metabolism; D008168:Lung / Q000166:cytology / Q000187:drug effects; D051907:Lysosomal Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D015162:Micronucleus Tests; D053758:Nanoparticles / Q000737:chemistry / Q000633:toxicity*; D018384:Oxidative Stress / Q000187:drug effects; D010316:Particle Size; D017382:Reactive Oxygen Species / Q000378:metabolism; D012822:Silicon Dioxide / Q000737:chemistry / Q000633:toxicity*; D059467:Transcriptome; D050051:Transient Receptor Potential Channels / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation; D025262:Vacuolar Proton-Translocating ATPases / Q000235:genetics / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D052118:Lysosomal-Associated Membrane Protein 1
chemical_list: D026921:Amino Acid Transport Systems, Neutral; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; D005978:Glutathione; D051907:Lysosomal Membrane Proteins; D017382:Reactive Oxygen Species; D012822:Silicon Dioxide; D050051:Transient Receptor Potential Channels; D025262:Vacuolar Proton-Translocating ATPases; D020691:rab GTP-Binding Proteins; C111445:CTNS protein, human; C052255:Lamp1 protein, mouse; D052118:Lysosomal-Associated Membrane Protein 1; C416674:MCOLN1 protein, human; C100151:dipeptidyl peptidase II; C000594810:ATP6V1G1 protein, human; C512960:Rab27B protein, human; C506979:ATP6V0B protein, human
keywords: Gene expression; Hyperspectral microscopy; Micronucleus assay; Nanoparticles; Silica nanoparticles