Abstract
In this methodological study, we demonstrated the relevance of 3D imaging performed at various scales for the ex vivo detection and location of cerium oxide nanomaterials (CeO2-NMs) in mouse lung. X-ray micro-computed tomography (micro-CT) with a voxel size from 14 µm to 1 µm (micro-CT) was combined with X-ray nano-computed tomography with a voxel size of 63 nm (nano-CT). An optimized protocol was proposed to facilitate the sample preparation, to minimize the experimental artifacts and to optimize the contrast of soft tissues exposed to metal-based nanomaterials (NMs). 3D imaging of the NMs biodistribution in lung tissues was consolidated by combining a vast variety of techniques in a correlative approach: histological observations, 2D chemical mapping and speciation analysis were performed for an unambiguous detection of NMs. This original methodological approach was developed following a worst-case scenario of exposure, i.e. high dose of exposure with administration via intra-tracheal instillation. Results highlighted both (i) the non-uniform distribution of CeO2-NMs within the entire lung lobe (using large field-of-view micro-CT) and (ii) the detection of CeO2-NMs down to the individual cell scale, e.g. macrophage scale (using nano-CT with a voxel size of 63 nm).
Highlights
IntroductionNanotechnology has undergone rapid development because of the enhanced or modified properties (e.g. magnetic, electronic, optic, surface chemical reactivity, etc.) of nanomaterials (NMs) compared to their bulk counterpart
In the past years, nanotechnology has undergone rapid development because of the enhanced or modified properties of nanomaterials (NMs) compared to their bulk counterpart
As initial CeO2-NMs can be subjected to biotransformation when reaching biological media (e.g.)[42], in-situ Ce speciation analyses were required. These analyses were performed by X-ray Absorption Near Edge Structure (XANES) measurements to obtain information on the site geometry and on the electronic structure of the probed element (i.e. Ce)
Summary
Nanotechnology has undergone rapid development because of the enhanced or modified properties (e.g. magnetic, electronic, optic, surface chemical reactivity, etc.) of nanomaterials (NMs) compared to their bulk counterpart. The challenge in nanotoxicology is to detect and locate small objects as NMs and NMs aggregates in soft organs, tissues and biological cells This required the use of bio-imaging techniques with high spatial resolution and sufficient contrast. The quality of the results and the data interpretation are strongly sample preparation-dependent To get around those difficulties, non-invasive 3D imaging techniques are preferable to visualize internal structure of intact biological samples. Such well-established (e.g. confocal microscopy) and emerging techniques (e.g. optical projection tomography or magnetic resonance microscopy)[8,9] exist but exhibit own limitations. Micro-CT allows the visualization of small pulmonary structures (e.g. bronchiole, alveolar duct) and alveolar architecture in ex vivo fixed lung, with resolution of 1–2 μm[21,23,24,25]
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