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Silica-coated magnetic nanoparticles impair proteasome activity and increase the formation of cytoplasmic inclusion bodies in vitro.

ABSTRACT: The potential toxicity of nanoparticles, particularly to neurons, is a major concern. In this study, we assessed the cytotoxicity of silica-coated magnetic nanoparticles containing rhodamine B isothiocyanate dye (MNPs@SiO2(RITC)) in HEK293 cells, SH-SY5Y cells, and rat primary cortical and dopaminergic neurons. In cells treated with 1.0??g/?l MNPs@SiO2(RITC), the expression of several genes related to the proteasome pathway was altered, and proteasome activity was significantly reduced, compared with control and with 0.1??g/?l MNPs@SiO2(RITC)-treated cells. Due to the reduction of proteasome activity, formation of cytoplasmic inclusions increased significantly in HEK293 cells over-expressing the ?-synuclein interacting protein synphilin-1 as well as in primary cortical and dopaminergic neurons. Primary neurons, particularly dopaminergic neurons, were more vulnerable to MNPs@SiO2(RITC) than SH-SY5Y cells. Cellular polyamines, which are associated with protein aggregation, were significantly altered in SH-SY5Y cells treated with MNPs@SiO2(RITC). These findings highlight the mechanisms of neurotoxicity incurred by nanoparticles.

SUBMITTER: Phukan G

PROVIDER: S-EPMC4932509 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature

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Silica-coated magnetic nanoparticles impair proteasome activity and increase the formation of cytoplasmic inclusion bodies in vitro.

Phukan Geetika G Shin Tae Hwan TH Shim Jeom Soon JS Paik Man Jeong MJ Lee Jin-Kyu JK Choi Sangdun S Kim Yong Man YM Kang Seong Ho SH Kim Hyung Sik HS Kang Yup Y Lee Soo Hwan SH Mouradian M Maral MM Lee Gwang G

Scientific reports 20160705

The potential toxicity of nanoparticles, particularly to neurons, is a major concern. In this study, we assessed the cytotoxicity of silica-coated magnetic nanoparticles containing rhodamine B isothiocyanate dye (MNPs@SiO2(RITC)) in HEK293 cells, SH-SY5Y cells, and rat primary cortical and dopaminergic neurons. In cells treated with 1.0 μg/μl MNPs@SiO2(RITC), the expression of several genes related to the proteasome pathway was altered, and proteasome activity was significantly reduced, compared ...[more]

PMID: 27378605

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Project description:BackgroundNanoparticles 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 [MNPs@SiO2(RITC)].MethodsWe investigated MNPs@SiO2(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 MNPs@SiO2(RITC) on neuronal cells were assessed by Transwell-based coculture with MNPs@SiO2(RITC)-treated microglia. MNPs@SiO2(RITC)-induced biological changes in the mouse brain in vivo were examined by immunohistochemical analysis.ResultsBV2 murine microglial cells were morphologically activated and the expression of Iba1, an activation marker protein, was increased after MNPs@SiO2(RITC) treatment. Transmission electron microscopy analysis revealed lysosomal accumulation of MNPs@SiO2(RITC) and the formation of vesicle-like structures in MNPs@SiO2(RITC)-treated BV2 cells. The expression of several genes related to metabolism and inflammation were altered in 100 µg/ml MNPs@SiO2(RITC)-treated microglia when compared with that in non-treated (control) and 10 µg/ml MNPs@SiO2(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, MNPs@SiO2(RITC), D-serine, and ubiquitin aggresomes were distributed in the MNPs@SiO2(RITC)-treated mouse brain.ConclusionsMNPs@SiO2(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.

Project description:The molecular imaging of in vivo targets allows non-invasive disease diagnosis. Nanoparticles offer a promising platform for molecular imaging because they can deliver large payloads of imaging reagents to the site of disease. Magnetic resonance imaging (MRI) is often preferred for clinical diagnosis because it uses non-ionizing radiation and offers both high spatial resolution and excellent penetration. We have explored the use of plant viruses as the basis of for MRI contrast reagents, specifically Tobacco mosaic virus (TMV), which can assemble to form either stiff rods or spheres. We loaded TMV particles with paramagnetic Gd ions, increasing the ionic relaxivity compared to free Gd ions. The loaded TMV particles were then coated with silica maintaining high relaxivities. Interestingly, we found that when Gd(DOTA) was loaded into the interior channel of TMV and the exterior was coated with silica, the T1 relaxivities increased by three-fold from 10.9 mM-1 s-1 to 29.7 mM-1s-1 at 60 MHz compared to uncoated Gd-loaded TMV. To test the performance of the contrast agents in a biological setting, we focused on interactions with macrophages because the active or passive targeting of immune cells is a popular strategy to investigate the cellular components involved in disease progression associated with inflammation. In vitro assays and phantom MRI experiments indicate efficient targeting and imaging of macrophages, enhanced contrast-to-noise ratio was observed by shape-engineering (SNP > TMV) and silica-coating (Si-TMV/SNP > TMV/SNP). Because plant viruses are in the food chain, antibodies may be prevalent in the population. Therefore we investigated whether the silica-coating could prevent antibody recognition; indeed our data indicate that mineralization can be used as a stealth coating option to reduce clearance. Therefore, we conclude that the silica-coated protein-based contrast agent may provide an interesting candidate material for further investigation for in vivo delineation of disease through macrophage imaging.

Project description:BackgroundNanoparticles are being increasingly used in biomedical applications owing to their unique physical and chemical properties and small size. However, their biophysical assessment and evaluation of side-effects remain challenging. We addressed this issue by investigating the effects of silica-coated magnetic nanoparticles containing rhodamine B isothiocyanate [MNPs@SiO2(RITC)] on biophysical aspects, such as membrane fluidity and traction force of human embryonic kidney 293 (HEK293) cells. We further extended our understanding on the biophysical effects of nanoparticles on cells using a combination of metabolic profiling and transcriptomic network analysis.ResultsOverdose (1.0 μg/µL) treatment with MNPs@SiO2(RITC) induced lipid peroxidation and decreased membrane fluidity in HEK293 cells. In addition, HEK293 cells were morphologically shrunk, and their aspect ratio was significantly decreased. We found that each traction force (measured in micropillar) was increased, thereby increasing the total traction force in MNPs@SiO2(RITC)-treated HEK293 cells. Due to the reduction in membrane fluidity and elevation of traction force, the velocity of cell movement was also significantly decreased. Moreover, intracellular level of adenosine triphosphate (ATP) was also decreased in a dose-dependent manner upon treatment with MNPs@SiO2(RITC). To understand these biophysical changes in cells, we analysed the transcriptome and metabolic profiles and generated a metabotranscriptomics network, which revealed relationships among peroxidation of lipids, focal adhesion, cell movement, and related genes and metabolites. Furthermore, in silico prediction of the network showed increment in the peroxidation of lipids and suppression of focal adhesion and cell movement.ConclusionTaken together, our results demonstrated that overdose of MNPs@SiO2(RITC) impairs cellular movement, followed by changes in the biophysical properties of cells, thus highlighting the need for biophysical assessment of nanoparticle-induced side-effects.

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Silica-coated magnetic nanoparticles impair proteasome activity and increase the formation of cytoplasmic inclusion bodies in vitro.

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Silica-coated magnetic nanoparticles impair proteasome activity and increase the formation of cytoplasmic inclusion bodies in vitro.

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