RESUMO
Due to their exciting properties, engineered nanoparticles have obtained substantial attention over the last two decades. As many types of nanoparticles are already used for technical and biomedical applications, the chances that cells in the brain will encounter nanoparticles have strongly increased. To test for potential consequences of an exposure of brain cells to engineered nanoparticles, cell culture models for different types of neural cells are frequently used. In this review article we will discuss experimental strategies and important controls that should be used to investigate the physicochemical properties of nanoparticles for the cell incubation conditions applied as well as for studies on the biocompatibility and the cellular uptake of nanoparticles in neural cells. The main focus of this article will be the interaction of cultured neural cells with iron oxide nanoparticles, but similar considerations are important for studying the consequences of an exposure of other types of cultured cells with other types of nanoparticles. Our article aims to improve the understanding of the special technical challenges of working with nanoparticles on cultured neural cells, to identify potential artifacts and to prevent misinterpretation of data on the potential adverse or beneficial consequences of a treatment of cultured cells with nanoparticles.
Assuntos
Compostos Férricos/toxicidade , Nanopartículas/toxicidade , Neurônios/efeitos dos fármacos , Animais , Materiais Biocompatíveis/química , Materiais Biocompatíveis/metabolismo , Materiais Biocompatíveis/toxicidade , Linhagem Celular Tumoral , Compostos Férricos/química , Compostos Férricos/metabolismo , Humanos , Nanopartículas/química , Nanopartículas/metabolismo , Neurônios/metabolismo , Tamanho da PartículaRESUMO
Iron oxide nanoparticles (IONPs) are used for various biomedical and therapeutic approaches. To investigate the uptake and the intracellular trafficking of IONPs in neural cells we have performed nanoparticle pulse-chase experiments to visualize the internalization and the fate of fluorescent IONPs in C6 glioma cells and astrocyte cultures. Already a short exposure to IONPs for 10 min at 4 °C (nanoparticle pulse) allowed binding of substantial amounts of nanoparticles to the cells, while internalization of IONPs into the cell was prevented. The uptake of bound IONPs and the intracellular trafficking was started by increasing the temperature to 37 °C (chase period). While hardly any cellular fluorescence nor any iron staining was detectable directly after the nanoparticle pulse, dotted cellular fluorescence and iron patterns appeared already within a few minutes after start of the chase incubation and became intensified in the perinuclear region during further incubation for up to 90 min. Longer chase incubations resulted in separation of the fluorescent coat from the core of the internalized IONPs. Disruption of actin filaments in C6 cells strongly impaired the internalization of IONPs, whereas destabilization of microtubules traped IONP-containing vesicles to the plasma membrane. In conclusion, nanoparticle pulse-chase experiments allowed to synchronize the cellular uptake of fluorescent IONPs and to identify for C6 cells an actin-dependent early and a microtubule-dependent later process in the intracellular trafficking of fluorescent IONPs.
Assuntos
Citoesqueleto/metabolismo , Glioma/metabolismo , Nanopartículas/metabolismo , Neurônios/metabolismo , Astrócitos/metabolismo , Movimento Celular/fisiologia , Sobrevivência Celular , Células Cultivadas , Citoplasma/metabolismo , Humanos , Microtúbulos/metabolismoRESUMO
Iron oxide nanoparticles (IONPs) are used for diverse medical approaches, although the potential health risks, for example adverse effects on brain functions, are not fully clarified. Several in vitro studies demonstrated that the different types of brain cells are able to accumulate IONPs and reported a toxic potential for IONPs, at least for microglia. However, little information is available for the in vivo effects of direct application of IONPs into the brain over time. Therefore, we examined the cellular responses and the distribution of iron in the rat brain at different time points after local infusion of IONPs into selected brain areas. Dispersed IONPs or an equivalent amount of low molecular weight iron complex ferric ammonium citrate or vehicle were infused into the medial prefrontal cortex (mPFC), the caudate putamen (CPu), or the dorsal hippocampus (dHip). Rats were sacrificed 1 day, 1 week, or 4 weeks post-infusion and brain sections were histologically examined for treatment effects on astrocytes, microglia, and neurons. Glial scar formation was observed in the mPFC and CPu 1 week post-infusion independent of the substance and probably resulted from the infusion procedure. Compared to vehicle, IONPs did not cause any obvious additional adverse effects and no additional tissue damage, while the infusion of ferric ammonium citrate enhanced neurodegeneration in the mPFC. Results of iron staining indicate that IONPs were mainly accumulated in microglia. Our results demonstrate that local infusions of IONPs in selected brain areas do not cause any additional adverse effects or neurodegeneration compared to vehicle.
Assuntos
Corpo Estriado/efeitos dos fármacos , Hipocampo/efeitos dos fármacos , Nanopartículas Magnéticas de Óxido de Ferro/administração & dosagem , Córtex Pré-Frontal/efeitos dos fármacos , Animais , Astrócitos/efeitos dos fármacos , Injeções Intraventriculares , Masculino , Microglia/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Ratos , Ratos WistarRESUMO
Background: Hepatitis C virus (HCV) is a hepatotropic, blood-borne virus, but in up to one-third of infections of the transmission route remained unidentified. Viral genome copies of HCV have been identified in several body fluids, however, non-parental transmission upon exposure to contaminated body fluids seems to be rare. Several body fluids, e.g., tears and saliva, are renowned for their antimicrobial and antiviral properties, nevertheless, HCV stability has never been systematically analyzed in those fluids. Methods: We used state of the art infectious HCV cell culture techniques to investigate the stability of HCV in different body fluids to estimate the potential risk of transmission via patient body fluid material. In addition, we mimicked a potential contamination of HCV in tear fluid and analyzed which impact commercially available contact lens solutions might have in such a scenario. Results: We could demonstrate that HCV remains infectious over several days in body fluids like tears, saliva, semen, and cerebrospinal fluid. Only hydrogen-peroxide contact lens solutions were able to efficiently inactivate HCV in a suspension test. Conclusion: These results indicate that HCV, once it is present in various body fluids of infected patients, remains infective and could potentially contribute to transmission upon direct contact.