This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research article
Gwang Lee
Ajou University School of Medicine and Graduate School of Medicine
Corresponding Author
ORCiD: https://orcid.org/0000-0002-1299-9478
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background
Nanoparticles have been studied for brain imaging, diagnosis, and drug delivery owing to their versatile properties due to their small sizes. However, there are growing concerns that nanoparticles may exert toxic effects in the brain. In this study, we assessed direct nanotoxicity on microglia, the resident macrophages of the central nervous system, and indirect toxicity on neuronal cells exerted by silica-coated magnetic nanoparticles containing rhodamine B isothiocyanate dye [[email protected]2(RITC)].
Methods
We investigated [email protected]2(RITC)-induced biological changes in BV2 murine microglial cells via RNA-sequencing-based transcriptome analysis and gas chromatography-mass spectrometry-based intracellular and extracellular amino acid profiling. Morphological changes were analyzed by transmission electron microscopy. Indirect effects of [email protected]2(RITC) on neuronal cells were assessed by Transwell-based coculture with [email protected]2(RITC)-treated microglia. [email protected]2(RITC)-induced biological changes in the mouse brain in vivo were examined by immunohistochemical analysis.
Results
BV2 murine microglial cells were morphologically activated and the expression of Iba1, an activation marker protein, was increased after [email protected]2(RITC) treatment. transmission electron microscopy analysis revealed lysosomal accumulation of [email protected]2(RITC) and the formation of vesicle-like structures in [email protected]2(RITC)-treated BV2 cells. The expression of several genes related to metabolism and inflammation were altered in 0.1 µg/µl [email protected]2(RITC)-treated microglia when compared with that in non-treated (control) and 0.01 µg/µl [email protected]2(RITC)-treated microglia. Combined transcriptome and amino acid profiling analyses revealed that the transport of serine family amino acids, including glycine, cysteine, and serine, was enhanced. However, only serine was increased in the growth medium of activated microglia; especially, excitotoxic d-serine secretion from primary rat microglia was the most strongly enhanced. Activated primary microglia reduced intracellular ATP levels and proteasome activity in cocultured neuronal cells, especially in primary cortical neurons, via d-serine secretion. Moreover, ubiquitinated proteins accumulated and inclusion bodies were increased in primary dopaminergic and cortical neurons cocultured with activated primary microglia. In vivo, [email protected]2(RITC), d-serine, and ubiquitin aggresomes were distributed in the [email protected]2(RITC)-treated mouse brain.
Conclusions
[email protected]2(RITC)-induced activation of microglia triggers excitotoxicity in neurons via d-serine secretion, highlighting the importance of neurotoxicity mechanisms incurred by nanoparticle-induced microglial activation.
Keywords
silica-coated magnetic nanoparticles, nanotoxicity, microglia, excitotoxicity, inclusion body
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This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research article
Gwang Lee
Ajou University School of Medicine and Graduate School of Medicine
Corresponding Author
ORCiD: https://orcid.org/0000-0002-1299-9478
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Published
View the published article at Particle and Fibre Toxicology.
Version 1
Posted 17 Feb, 2021
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Physiology
License:
This work is licensed under a CC BY 4.0 License. Read Full License
View the published article at Particle and Fibre Toxicology.
Background
Nanoparticles have been studied for brain imaging, diagnosis, and drug delivery owing to their versatile properties due to their small sizes. However, there are growing concerns that nanoparticles may exert toxic effects in the brain. In this study, we assessed direct nanotoxicity on microglia, the resident macrophages of the central nervous system, and indirect toxicity on neuronal cells exerted by silica-coated magnetic nanoparticles containing rhodamine B isothiocyanate dye [[email protected]2(RITC)].
Methods
We investigated [email protected]2(RITC)-induced biological changes in BV2 murine microglial cells via RNA-sequencing-based transcriptome analysis and gas chromatography-mass spectrometry-based intracellular and extracellular amino acid profiling. Morphological changes were analyzed by transmission electron microscopy. Indirect effects of [email protected]2(RITC) on neuronal cells were assessed by Transwell-based coculture with [email protected]2(RITC)-treated microglia. [email protected]2(RITC)-induced biological changes in the mouse brain in vivo were examined by immunohistochemical analysis.
Results
BV2 murine microglial cells were morphologically activated and the expression of Iba1, an activation marker protein, was increased after [email protected]2(RITC) treatment. transmission electron microscopy analysis revealed lysosomal accumulation of [email protected]2(RITC) and the formation of vesicle-like structures in [email protected]2(RITC)-treated BV2 cells. The expression of several genes related to metabolism and inflammation were altered in 0.1 µg/µl [email protected]2(RITC)-treated microglia when compared with that in non-treated (control) and 0.01 µg/µl [email protected]2(RITC)-treated microglia. Combined transcriptome and amino acid profiling analyses revealed that the transport of serine family amino acids, including glycine, cysteine, and serine, was enhanced. However, only serine was increased in the growth medium of activated microglia; especially, excitotoxic d-serine secretion from primary rat microglia was the most strongly enhanced. Activated primary microglia reduced intracellular ATP levels and proteasome activity in cocultured neuronal cells, especially in primary cortical neurons, via d-serine secretion. Moreover, ubiquitinated proteins accumulated and inclusion bodies were increased in primary dopaminergic and cortical neurons cocultured with activated primary microglia. In vivo, [email protected]2(RITC), d-serine, and ubiquitin aggresomes were distributed in the [email protected]2(RITC)-treated mouse brain.
Conclusions
[email protected]2(RITC)-induced activation of microglia triggers excitotoxicity in neurons via d-serine secretion, highlighting the importance of neurotoxicity mechanisms incurred by nanoparticle-induced microglial activation.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
This is a list of supplementary files associated with this preprint. Click to download.
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