Periodic table screening for enhanced positive contrast in MRI and in vivo uptake in glioblastoma.

The quest for nanomaterial-based imaging probes that can provide positive contrast in MRI is fueled by the necessity of developing novel diagnostic applications with potential for clinical translation that current gold standard probes cannot provide. Although interest in nanomaterials for positive contrast has increased in recent years, their study is less developed than that of traditional negative contrast probes in MRI. In our search for new magnetic materials with enhanced features as positive contrast probes for MRI, we decided to explore the chemical space to comprehensively analyze the effects of different metals on the performance of iron oxide nanomaterials already able to provide positive contrast in MRI. To this end, we synthesized 30 different iron oxide-based nanomaterials. Thorough characterization was performed, including multivariate analysis, to study the effect of different variables on their relaxometric properties. Based on these results, we identified the best combination of metals for in vivo imaging and tested them in different experiments. First, we tested its performance on magnetic resonance angiography using a concentration ten times lower than that clinically approved for Gd. Finally, we studied the capability of these nanomaterials to cross the affected blood-brain barrier in a glioblastoma model. The results showed that the selected nanomaterials provided excellent positive contrast at large magnetic field and were able to accumulate at the tumor site, highlighting the affected tissue.

Cellular and Molecular Processes Are Differently Influenced in Primary Neural Cells by Slight Changes in the Physicochemical Properties of Multicore Magnetic Nanoparticles.

Herein, we use two exemplary superparamagnetic iron oxide multicore nanoparticles (SPIONs) to illustrate the significant influence of slightly different physicochemical properties on the cellular and molecular processes that define SPION interplay with primary neural cells. Particularly, we have designed two different SPION structures, NFA (i.e., a denser multicore structure accompanied by a slightly less negative surface charge and a higher magnetic response) and NFD (i.e., a larger surface area and more negatively charged), and identified specific biological responses dependent on SPION type, concentration, exposure time, and magnetic actuation. Interestingly, NFA SPIONs display a higher cell uptake, likely driven by their less negative surface and smaller protein corona, more significantly impacting cell viability and complexity. The tight contact of both SPIONs with neural cell membranes results in the significant augmentation of phosphatidylcholine, phosphatidylserine, and sphingomyelin and the reduction of free fatty acids and triacylglycerides for both SPIONs. Nonetheless, NFD induces greater effects on lipids, especially under magnetic actuation, likely indicating a preferential membranal location and/or a tighter interaction with membrane lipids than NFA, in agreement with their lower cell uptake. From a functional perspective, these lipid changes correlate with an increase in plasma membrane fluidity, again larger for more negatively charged nanoparticles (NFD). Finally, the mRNA expression of iron-related genes such as Ireb-2 and Fth-1 remains unaltered, while TfR-1 is only detected in SPION-treated cells. Taken together, these results demonstrate the substantial impact that minor physicochemical differences of nanomaterials may exert in the specific targeting of cellular and molecular processes. A denser multicore structure generated by autoclave-based production is accompanied by a slight difference in surface charge and magnetic properties that become decisive for the biological impact of these SPIONs. Their capacity to markedly modify the lipidic cell content makes them attractive as lipid-targetable nanomedicines.

Different coatings on magnetic nanoparticles dictate their degradation kinetics in vivo for 15 months after intravenous administration in mice.

BACKGROUND: The surface coating of iron oxide magnetic nanoparticle (MNPs) drives their intracellular trafficking and degradation in endolysosomes, as well as dictating other cellular outcomes. As such, we assessed whether MNP coatings might influence their biodistribution, their accumulation in certain organs and their turnover therein, processes that must be understood in vivo to optimize the design of nanoformulations for specific therapeutic/diagnostic needs. RESULTS: In this study, three different MNP coatings were analyzed, each conferring the identical 12 nm iron oxide cores with different physicochemical characteristics: 3-aminopropyl-triethoxysilane (APS), dextran (DEX), and dimercaptosuccinic acid (DMSA). When the biodistribution of these MNPs was analyzed in C57BL/6 mice, they all mainly accumulated in the spleen and liver one week after administration. The coating influenced the proportion of the MNPs in each organ, with more APS-MNPs accumulating in the spleen and more DMSA-MNPs accumulating in the liver, remaining there until they were fully degraded. The changes in the physicochemical properties of the MNPs (core size and magnetic properties) was also assessed during their intracellular degradation when internalized by two murine macrophage cell lines. The decrease in the size of the MNPs iron core was influenced by their coating and the organ in which they accumulated. Finally, MNP degradation was analyzed in the liver and spleen of C57BL/6 mice from 7 days to 15 months after the last intravenous MNP administration. CONCLUSIONS: The MNPs degraded at different rates depending on the organ and their coating, the former representing the feature that was fundamental in determining the time they persisted. In the liver, the rate of degradation was similar for all three coatings, and it was faster than in the spleen. This information regarding the influence of coatings on the in vivo degradation of MNPs will help to choose the best coating for each biomedical application depending on the specific clinical requirements.

Iron oxide and iron oxyhydroxide nanoparticles impair SARS-CoV-2 infection of cultured cells.

BACKGROUND: Coronaviruses usually cause mild respiratory disease in humans but as seen recently, some human coronaviruses can cause more severe diseases, such as the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the global spread of which has resulted in the ongoing coronavirus pandemic. RESULTS: In this study we analyzed the potential of using iron oxide nanoparticles (IONPs) coated with biocompatible molecules like dimercaptosuccinic acid (DMSA), 3-aminopropyl triethoxysilane (APS) or carboxydextran (FeraSpin™ R), as well as iron oxyhydroxide nanoparticles (IOHNPs) coated with sucrose (Venofer®), or iron salts (ferric ammonium citrate -FAC), to treat and/or prevent SARS-CoV-2 infection. At non-cytotoxic doses, IONPs and IOHNPs impaired virus replication and transcription, and the production of infectious viruses in vitro, either when the cells were treated prior to or after infection, although with different efficiencies. Moreover, our data suggest that SARS-CoV-2 infection affects the expression of genes involved in cellular iron metabolism. Furthermore, the treatment of cells with IONPs and IOHNPs affects oxidative stress and iron metabolism to different extents, likely influencing virus replication and production. Interestingly, some of the nanoparticles used in this work have already been approved for their use in humans as anti-anemic treatments, such as the IOHNP Venofer®, and as contrast agents for magnetic resonance imaging in small animals like mice, such as the FeraSpin™ R IONP. CONCLUSIONS: Therefore, our results suggest that IONPs and IOHNPs may be repurposed to be used as prophylactic or therapeutic treatments in order to combat SARS-CoV-2 infection.

The surface coating of iron oxide nanoparticles drives their intracellular trafficking and degradation in endolysosomes differently depending on the cell type.

Magnetic nanoparticles (MNPs) are potential theranostic tools that are biodegraded through different endocytic pathways. However, little is known about the endolysosomal network through which MNPs transit and the influence of the surface coating in this process. Here, we studied the intracellular transit of two MNPs with identical iron oxide core size but with two distinct coatings: 3-aminopropyl-trietoxysilane (APS) and dimercaptosuccinic acid (DMSA). Using endolysosomal markers and a high throughput analysis of the associated proteome, we tracked the MNPs intracellularly in two different mouse cell lines, RAW264.7 (macrophages) and Pan02 (tumor cells). We did not detect differences in the MNP trafficking kinetics nor in the MNP-containing endolysosome phenotype in Pan02 cells. Nonetheless, DMSA-MNPs transited at slower rate than APS-MNPs in macrophages as measured by MNP accumulation in Rab7+ endolysosomes. Macrophage DMSA-MNP-containing endolysosomes had a higher percentage of lytic enzymes and catalytic proteins than their APS-MNP counterparts, concomitantly with a V-type ATPase enrichment, suggesting an acidic nature. Consequently, more autophagic vesicles are induced by DMSA-MNPs in macrophages, enhancing the expression of iron metabolism-related genes and proteins. Therefore, unlike Pan02 cells, the MNP coating appears to influence the intracellular trafficking rate and the endolysosome nature in macrophages. These results highlight how the MNP coating can determine the nanoparticle intracellular fate and biodegradation in a cell-type bias.

Nanoparticles for Magnetic Heating: When Two (or More) Is Better Than One.

The increasing use of magnetic nanoparticles as heating agents in biomedicine is driven by their proven utility in hyperthermia therapeutic treatments and heat-triggered drug delivery methods. The growing demand of efficient and versatile nanoheaters has prompted the creation of novel types of magnetic nanoparticle systems exploiting the magnetic interaction (exchange or dipolar in nature) between two or more constituent magnetic elements (magnetic phases, primary nanoparticles) to enhance and tune the heating power. This process occurred in parallel with the progress in the methods for the chemical synthesis of nanostructures and in the comprehension of magnetic phenomena at the nanoscale. Therefore, complex magnetic architectures have been realized that we classify as: (a) core/shell nanoparticles; (b) multicore nanoparticles; (c) linear aggregates; (d) hybrid systems; (e) mixed nanoparticle systems. After a general introduction to the magnetic heating phenomenology, we illustrate the different classes of nanoparticle systems and the strategic novelty they represent. We review some of the research works that have significantly contributed to clarify the relationship between the compositional and structural properties, as determined by the synthetic process, the magnetic properties and the heating mechanism.

Mixing iron oxide nanoparticles with different shape and size for tunable magneto-heating performance.

Tuning the magnetic properties of nanoparticles is a strategic goal to use them in the most effective way to perform specific functions in the nanomedicine field. We report a systematic study carried out on a set of samples obtained by mixing together iron oxide nanoparticles with different shape: elongated with aspect ratio ∼5.2 and mean volume of the order of 103 nm3 (excluding the silica coating) and spherical with mean volume one order of magnitude larger. These structural features of the nanoparticles together with their aggregation state determine the magnetic anisotropy and the magnetic relaxation processes. In particular, the spherical nanoparticles turn out to be more stable against superparamagnetic relaxation. Mixing the nanoparticles in different proportions allows to modulate the magnetic response of the samples. The two populations of nanoparticles magnetically influence each other through a mean field mechanism, which depends crucially on temperature and rules the hysteretic magnetic properties and their thermal evolution. This magnetic phenomenology has a direct impact on the ability of the mixed samples to generate heat under an alternating magnetic field, a key function in view of nanomedicine applications. Under proper testing conditions, the heating efficiency of the mixed samples is larger compared to that obtained as the sum of those of the parent nanoparticles. This occurs thanks to the mean field produced by the magnetically blocked spherical nanoparticles that stabilizes the thermally fluctuating moments of the elongated ones, which therefore contribute more effectively to the heat production.

Aggregation effects on the magnetic properties of iron oxide colloids.

Magnetic nanoparticles (MNPs), and in particular iron oxide nanoparticles (mainly magnetite and maghemite), are being widely used in the form of aqueous colloids for biomedical applications. In such colloids, nanoparticles tend to form assemblies, either aggregates, if the union is permanent, or agglomerates, if it is reversible. These clustering processes have a strong impact on the MNPs' properties that are often not well understood. In this review, the causes and consequences of MNPs aggregation/agglomeration are reviewed and discussed. Special attention has been paid to the impact of the MNPs aggregation/agglomeration on their magnetic properties and heating properties, when exposed to an alternating magnetic field in the frame of magnetic hyperthermia. In addition, a model system with MNPs of two different sizes coated with three different molecules oleic acid, meso-2, 3-dimercaptosuccinic acid and poly(maleic anhydride-alt-1-octadecene) has been characterized and the results used to support the ideas reviewed.

Improving the reliability of the iron concentration quantification for iron oxide nanoparticle suspensions: a two-institutions study.

Most iron oxide nanoparticles applications, and in special biomedical applications, require the accurate determination of iron content as the determination of particle properties from measurements in dispersions is strongly dependent on it. Inductively coupled plasma (ICP) and spectrophotometry are two typical worldwide used analytical methods for iron concentration determination. In both techniques, precise determination of iron is not straightforward and nanoparticle digestion and dilution procedures are needed prior to analysis. The sample preparation protocol has been shown to be as important as the analytical method when accuracy is aimed as many puzzling reported results in magnetic, colloidal, and structural properties are simply attributable to inadequate dissolution procedures. Therefore, a standard sample preparation protocol is needed to ensure the adequate and complete iron oxide nanoparticle dissolution and to harmonize this procedure. In this work, an interlaboratory evaluation of an optimized iron oxide nanoparticle digestion/dilution protocol was carried out. The presented protocol is simple, inexpensive, and does not involve any special device (as microwave, ultrasound, or other high-priced digestion devices). Then, iron concentration was measured by ICP-OES (performed in ICMM/CSIC-Spain) and spectrophotometry (NanoPET-Germany) and the obtained concentration values were analyzed to determine the most probable error causes. Uncertainty values as low as 1.5% were achieved after the optimized method was applied. Moreover, this article provides a list of recommendations to significantly reduce uncertainty in both sample preparation and analysis procedures. Graphical abstract ᅟ.

One-Step Fast Synthesis of Nanoparticles for MRI: Coating Chemistry as the Key Variable Determining Positive or Negative Contrast.

Iron oxide nanomaterial is a typical example of a magnetic resonance imaging probe for negative contrast. It has also been shown how this nanomaterial can be synthesized for positive contrast by modification of the composition and size of the core. However, the role of the organic coating in the relaxometric properties is largely unexplored. Here, maghemite nanoparticles with either excellent positive or very good negative contrast performance are obtained by modifying coating thickness while the core is kept unchanged. Different nanoparticles with tailored features as contrast agent according to the coating layer thickness have been obtained in a single-step microwave-driven synthesis by heating at different temperatures. A comprehensive analysis is conducted of how the composition and structure of the coating affects the final magnetic, relaxometric, and imaging performance. These results show how the organic coating plays a fundamental role in the intrinsic relaxometric parameters of iron oxide-based contrast media.

Time-course assessment of the aggregation and metabolization of magnetic nanoparticles.

To successfully develop biomedical applications for magnetic nanoparticles, it is imperative that these nanoreagents maintain their magnetic properties in vivo and that their by-products are safely metabolized. When placed in biological milieu or internalized into cells, nanoparticle aggregation degree can increase which could affect magnetic properties and metabolization. To evaluate these aggregation effects, we synthesized citric acid-coated iron oxide nanoparticles whose magnetic susceptibility can be modified by aggregation in agar dilutions and dextran-layered counterparts that maintain their magnetic properties unchanged. Macrophage models were used for in vitro uptake and metabolization studies, as these cells control iron homeostasis in the organism. Electron microscopy and magnetic susceptibility studies revealed a cellular mechanism of nanoparticle degradation, in which a small fraction of the particles is rapidly degraded while the remaining ones maintain their size. Both nanoparticle types produced similar iron metabolic profiles but these profiles differed in each macrophage model. Thus, nanoparticles induced iron responses that depended on macrophage programming. In vivo studies showed that nanoparticles susceptible to changes in magnetic properties through aggregation effects had different behavior in lungs, liver and spleen. Liver ferritin levels increased in these animals showing that nanoparticles are degraded and their by-products incorporated into normal metabolic routes. These data show that nanoparticle iron metabolization depends on cell type and highlight the necessity to assess nanoparticle aggregation in complex biological systems to develop effective in vivo biomedical applications. STATEMENT OF SIGNIFICANCE: Magnetic iron oxide nanoparticles have great potential for biomedical applications. It is however imperative that these nanoreagents preserve their magnetic properties once inoculated, and that their degradation products can be eliminated. When placed in a biological milieu nanoparticles can aggregate and this can affect their magnetic properties and their degradation. In this work, we showed that iron oxide nanoparticles trigger the iron metabolism in macrophages, the main cell type involved in iron homeostasis in the organism. We also show that aggregation can affect nanoparticle magnetic properties when inoculated in animal models. This work confirms iron oxide nanoparticle biocompatibility and highlights the necessity to assess in vivo nanoparticle aggregation to successfully develop biomedical applications.

Formation Mechanism of Maghemite Nanoflowers Synthesized by a Polyol-Mediated Process.

Magnetic nanoparticles are being developed as structural and functional materials for use in diverse areas, including biomedical applications. Here, we report the synthesis of maghemite (γ-Fe2O3) nanoparticles with distinct morphologies: single-core and multicore, including hollow spheres and nanoflowers, prepared by the polyol process. We have used sodium acetate to control the nucleation and assembly process to obtain the different particle morphologies. Moreover, from samples obtained at different time steps during the synthesis, we have elucidated the formation mechanism of the nanoflowers: the initial phases of the reaction present a lepidocrocite (γ-FeOOH) structure, which suffers a fast dehydroxylation, transforming to an intermediate "undescribed" phase, possibly a partly dehydroxylated lepidocrocite, which after some incubation time evolves to maghemite nanoflowers. Once the nanoflowers have been formed, a crystallization process takes place, where the γ-Fe2O3 crystallites within the nanoflowers grow in size (from ∼11 to 23 nm), but the particle size of the flower remains essentially unchanged (∼60 nm). Samples with different morphologies were coated with citric acid and their heating capacity in an alternating magnetic field was evaluated. We observe that nanoflowers with large cores (23 nm, controlled by annealing) densely packed (tuned by low NaAc concentration) offer 5 times enhanced heating capacity compared to that of the nanoflowers with smaller core sizes (15 nm), 4 times enhanced heating effect compared to that of the hollow spheres, and 1.5 times enhanced heating effect compared to that of single-core nanoparticles (36 nm) used in this work.

In-situ particles reorientation during magnetic hyperthermia application: Shape matters twice.

Promising advances in nanomedicine such as magnetic hyperthermia rely on a precise control of the nanoparticle performance in the cellular environment. This constitutes a huge research challenge due to difficulties for achieving a remote control within the human body. Here we report on the significant double role of the shape of ellipsoidal magnetic nanoparticles (nanorods) subjected to an external AC magnetic field: first, the heat release is increased due to the additional shape anisotropy; second, the rods dynamically reorientate in the orthogonal direction to the AC field direction. Importantly, the heating performance and the directional orientation occur in synergy and can be easily controlled by changing the AC field treatment duration, thus opening the pathway to combined hyperthermic/mechanical nanoactuators for biomedicine. Preliminary studies demonstrate the high accumulation of nanorods into HeLa cells whereas viability analysis supports their low toxicity and the absence of apoptotic or necrotic cell death after 24 or 48 h of incubation.

Fast synthesis and bioconjugation of (68) Ga core-doped extremely small iron oxide nanoparticles for PET/MR imaging.

Combination of complementary imaging techniques, like hybrid PET/MRI, allows protocols to be developed that exploit the best features of both. In order to get the best of these combinations the use of dual probes is highly desirable. On this sense the combination of biocompatible iron oxide nanoparticles and 68Ga isotope is a powerful development for the new generation of hybrid systems and multimodality approaches. Our objective was the synthesis and application of a chelator-free 68Ga-iron oxide nanotracer with improved stability, radiolabeling yield and in vivo performance in dual PET/MRI. We carried out the core doping of iron oxide nanoparticles, without the use of any chelator, by a microwave-driven protocol. The synthesis allowed the production of extremely small (2.5 nm) 68Ga core-doped iron oxide nanoparticles. The microwave approach allowed an extremely fast synthesis with a 90% radiolabeling yield and T1 contrast in MRI. With the same microwave approach the nano-radiotracer was functionalized in a fast and efficient way. We finally evaluated these dual targeting nanoparticles in an angiogenesis murine model by PET/MR imaging. Copyright © 2016 John Wiley & Sons, Ltd.

Efficient and safe internalization of magnetic iron oxide nanoparticles: two fundamental requirements for biomedical applications.

We have performed a series of in vitro tests proposed for the reliable assessment of safety associated with nanoparticles-cell interaction. A thorough analysis of toxicity of three different coating iron oxide nanoparticles on HeLa cells has been carried out including, methyl thiazol tetrazolium bromide (MTT) and Trypan blue exclusion tests, cell morphology observation by optical and Scanning Electron Microscopy (SEM), study of cytoskeletal components, analysis of cell cycle and the presence of reactive oxygen species (ROS). We have quantified magnetic nanoparticle internalization, determined possible indirect cell damages and related it to the nanoparticle coating. The results confirm a very low toxicity of the analyzed iron oxide nanoparticles into HeLa cells by multiple assays and pave the way for a more successful cancer diagnostic and treatment without secondary effects. FROM THE CLINICAL EDITOR: In this paper, three different iron oxide nanoparticles are studied and compared from the standpoint of safety and toxicity in HeLa cells, demonstrating low toxicity for each preparation, and paving the way to potential future clinical applications.

Deferiprone and idebenone rescue frataxin depletion phenotypes in a Drosophila model of Friedreich's ataxia.

Friedreich's ataxia (FRDA), the most common inherited ataxia, is a neurodegenerative disease caused by a reduction in the levels of the mitochondrial protein frataxin, the function of which remains a controversial matter. Several therapeutic approaches are being developed to increase frataxin expression and reduce the intramitochondrial iron aggregates and oxidative damage found in this disease. In this study, we tested separately the response of a Drosophila RNAi model of FRDA (Llorens et al., 2007) to treatment with the iron chelator deferiprone (DFP) and the antioxidant idebenone (IDE), which are both in clinical trials. The FRDA flies have a shortened life span and impaired motor coordination, and these phenotypes are more pronounced in oxidative stress conditions. In addition, under hyperoxia, the activity of the mitochondrial enzyme aconitase is strongly reduced in the FRDA flies. This study reports that DFP and IDE improve the life span and motor ability of frataxin-depleted flies. We show that DFP eliminates the excess of labile iron in the mitochondria and thus prevents the toxicity induced by iron accumulation. IDE treatment rescues aconitase activity in hyperoxic conditions. These results validate the use of our Drosophila model of FRDA to screen for therapeutic molecules to treat this disease.

Large scale production of biocompatible magnetite nanocrystals with high saturation magnetization values through green aqueous synthesis.

In this work, a straightforward aqueous synthesis for mass production (up to 20 g) of uniform and crystalline magnetite nanoparticles with core sizes between 20 and 30 nm, which are the optimum nanoparticle core sizes for hyperthermia applications, is proposed. Magnetic and heating properties have been analyzed showing very high saturation magnetization and magnetic heating values. To stabilize the naked magnetite nanocrystals at physiological pH and increase their circulation time in blood, they have been covalently coated with carboxymethyl dextran, a biocompatible polymer. The influence of this superficial modification on the magnetic and heating properties has been studied showing that these biocompatible magnetic nanocrystals maintain high saturation magnetization values, good colloidal stability and hyperthermia properties in the presence of the polymeric external layer. These particles, suitably functionalized, could be used to selectively kill cancer cells under a moderate alternating magnetic field (44 mT and 70 kHz).

Liver and brain imaging through dimercaptosuccinic acid-coated iron oxide nanoparticles.

BACKGROUND & AIM: Uptake, cytotoxicity and interaction of improved superparamagnetic iron oxide nanoparticles were studied in cells, tissues and organs after single and multiple exposures. MATERIAL & METHOD: We prepared dimercaptosuccinic acid-coated iron oxide nanoparticles by thermal decomposition in organic medium, resulting in aqueous suspensions with a small hydrodynamic size (< 100 nm), high saturation magnetization and susceptibility, high nuclear magnetic resonance contrast and low cytotoxicity. RESULTS: In vitro and in vivo behavior showed that these nanoparticles are efficient carriers for drug delivery to the liver and brain that can be combined with MRI detection.

Uniform nanosized goethite particles obtained by aerial oxidation in the FeSO4-Na2CO3 system.

Pure goethite particles in the nanometer size range (from approximately 200 to approximately 80 nm) with an elongated shape (axial ratio from approximately 5 to approximately 8) useful as iron precursors for magnetic recording have been prepared by oxidation of the suspensions resulting from the addition of sodium carbonate to Fe(II) sulfate aqueous solutions under a restrictive set of experimental conditions (Fe(II) concentration, carbonate/Fe(II) mole ratio, temperature, and air flow rate). In all cases, the goethite particles were formed by a dissolution-recrystallization mechanism through an intermediate green-rust phase. The particle size was determined by the carbonate/Fe(II) ratio (which controls the formation pH), the FeSO(4) concentration, and the air flow rate. The smallest particles (length 80 nm) were obtained for a high carbonate/Fe(II) mole ratio (>/=3), a low Fe(II) concentration (0.075 mol dm(-3)), and an air flow rate of 2 dm(3) min(-1). The goethite particles were also characterized by the electron diffraction and high-resolution TEM finding that they were monocrystalline, having the crystalline c axis parallel to the longest particle dimension.