title: High resolution and dynamic imaging of biopersistence and bioreactivity of extra and intracellular MWNTs exposed to microglial cells.
abstract: Multi-walled carbon nanotubes (MWNTs) are increasingly being developed both as neuro-therapeutic drug delivery systems to the brain and as neural scaffolds to drive tissue regeneration across lesion sites. MWNTs with different degrees of acid oxidation may have different bioreactivities and propensities to aggregate in the extracellular environment, and both individualised and aggregated MWNTs may be expected to be found in the brain. Before practical application, it is vital to understand how both aggregates and individual MWNTs will interact with local phagocytic immune cells, the microglia, and ultimately to determine their biopersistence in the brain. The processing of extra- and intracellular MWNTs (both pristine and when acid oxidised) by microglia was characterised across multiple length scales by correlating a range of dynamic, quantitative and multi-scale techniques, including: UV-vis spectroscopy, light microscopy, focussed ion beam scanning electron microscopy and transmission electron microscopy. Dynamic, live cell imaging revealed the ability of microglia to break apart and internalise micron-sized extracellular agglomerates of acid oxidised MWNTs, but not pristine MWNTs. The total amount of MWNTs internalised by, or strongly bound to, microglia was quantified as a function of time. Neither the significant uptake of oxidised MWNTs, nor the incomplete uptake of pristine MWNTs affected microglial viability, pro-inflammatory cytokine release or nitric oxide production. However, after 24 h exposure to pristine MWNTs, a significant increase in the production of reactive oxygen species was observed. Small aggregates and individualised oxidised MWNTs were present in the cytoplasm and vesicles, including within multilaminar bodies, after 72 h. Some evidence of morphological damage to oxidised MWNT structure was observed including highly disordered graphitic structures, suggesting possible biodegradation. This work demonstrates the utility of dynamic, quantitative and multi-scale techniques in understanding the different cellular processing routes of functionalised nanomaterials. This correlative approach has wide implications for assessing the biopersistence of MWNT aggregates elsewhere in the body, in particular their interaction with macrophages in the lung.
authors: Department of Materials, Imperial College London, Exhibition Road, London, SW7 2AZ, UK. Electronic address: aeg08@ic.ac.uk.
authors: Angela E
authors: AE
authors: Goode
authors: Centre for Neuroinflammation and Neurodegeneration, Department of Medicine, Division of Brain Sciences, Imperial College London, London, W12 0NN, UK.
authors: Daniel A
authors: DA
authors: Gonzalez Carter
authors: Centre for Neuroinflammation and Neurodegeneration, Department of Medicine, Division of Brain Sciences, Imperial College London, London, W12 0NN, UK.
authors: Michael
authors: M
authors: Motskin
authors: Centre for Neuroinflammation and Neurodegeneration, Department of Medicine, Division of Brain Sciences, Imperial College London, London, W12 0NN, UK.
authors: Ilse S
authors: IS
authors: Pienaar
authors: Department of Materials, Imperial College London, Exhibition Road, London, SW7 2AZ, UK.
authors: Shu
authors: S
authors: Chen
authors: Department of Chemistry, Imperial College London, Exhibition Road, London, SW7 2AZ, UK.
authors: Sheng
authors: S
authors: Hu
authors: Department of Materials, Imperial College London, Exhibition Road, London, SW7 2AZ, UK.
authors: Pakatip
authors: P
authors: Ruenraroengsak
authors: Department of Materials, Imperial College London, Exhibition Road, London, SW7 2AZ, UK.
authors: Mary P
authors: MP
authors: Ryan
authors: Department of Chemistry, Imperial College London, Exhibition Road, London, SW7 2AZ, UK.
authors: Milo S P
authors: MS
authors: Shaffer
authors: Centre for Neuroinflammation and Neurodegeneration, Department of Medicine, Division of Brain Sciences, Imperial College London, London, W12 0NN, UK.
authors: David T
authors: DT
authors: Dexter
authors: Department of Materials, Imperial College London, Exhibition Road, London, SW7 2AZ, UK. Electronic address: a.porter@imperial.ac.uk.
authors: Alexandra E
authors: AE
authors: Porter
mesh_terms: D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D016207:Cytokines / Q000096:biosynthesis; D004705:Endocytosis / Q000187:drug effects; D005110:Extracellular Space / Q000737:chemistry*; D021621:Imaging, Three-Dimensional / Q000379:methods*; D042541:Intracellular Space / Q000737:chemistry*; D051379:Mice; D017628:Microglia / Q000166:cytology* / Q000187:drug effects / Q000648:ultrastructure; D037742:Nanotubes, Carbon / Q000633:toxicity* / Q000648:ultrastructure; D010084:Oxidation-Reduction; D013056:Spectrophotometry, Ultraviolet
chemical_list: D016207:Cytokines; D037742:Nanotubes, Carbon
keywords: Aggregation; Biocompatibility; Brain; Carbon nanotube (CNT); Microglia