Discrimination between the effects of pulsed electrical stimulation and electrochemically conditioned medium on human osteoblasts.

BACKGROUND: Electrical stimulation is used for enhanced bone fracture healing. Electrochemical processes occur during the electrical stimulation at the electrodes and influence cellular reactions. Our approach aimed to distinguish between electrochemical and electric field effects on osteoblast-like MG-63 cells. We applied 20 Hz biphasic pulses via platinum electrodes for 2 h. The electrical stimulation of the cell culture medium and subsequent application to cells was compared to directly stimulated cells. The electric field distribution was predicted using a digital twin. RESULTS: Cyclic voltammetry and electrochemical impedance spectroscopy revealed partial electrolysis at the electrodes, which was confirmed by increased concentrations of hydrogen peroxide in the medium. While both direct stimulation and AC-conditioned medium decreased cell adhesion and spreading, only the direct stimulation enhanced the intracellular calcium ions and reactive oxygen species. CONCLUSION: The electrochemical by-product hydrogen peroxide is not the main contributor to the cellular effects of electrical stimulation. However, undesired effects like decreased adhesion are mediated through electrochemical products in stimulated medium. Detailed characterisation and monitoring of the stimulation set up and electrochemical reactions are necessary to find safe electrical stimulation protocols.

Gold Nanocolumnar Templates for Effective Chemical Sensing by Surface-Enhanced Raman Scattering.

Herein, we investigate the chemical sensing by surface-enhanced Raman scattering regarding two templates of gold nanocolumns (vertical and tilted) manufactured by glancing angle deposition with magnetron sputtering. We selected this fabrication technique due to its advantages in terms of low-cost production and ease of implementation. These gold nanocolumnar structures allow producing a high density of strongly confined electric field spots within the nanogaps between the neighboring nanocolumns. Thiophenol molecules were used as model analytes since they have the principal property to adsorb well on gold surfaces. Regarding chemical sensing, the vertical (tilted) nanocolumnar templates showed a detection threshold limit of 10 nM (20 nM), an enhancement factor of 9.8 × 108 (4.8 × 108), and a high quality of adsorption with an adsorption constant Kads of 2.0 × 106 M-1 (1.8 × 106 M-1) for thiophenol molecules.

Luminescence enhancement effects on nanostructured perovskite thin films for Er/Yb-doped solar cells.

Recent attempts to improve solar cell performance by increasing their spectral absorption interval incorporate up-converting fluorescent nanocrystals on the structure. These nanocrystals absorb low energy light and emit higher energy photons that can then be captured by the solar cell active layer. However, this process is very inefficient and it needs to be enhanced by different strategies. In this work, we have studied the effect of nanostructuration of perovskite thin films used in the fabrication of hybrid solar cells on their local optical properties. The perovskite surface was engraved with a focused ion beam to form gratings of one-dimensional grooves. We characterized the surfaces with a fluorescence scanning near-field optical microscope, and obtained maps showing a fringe pattern oriented in a direction parallel to the grooves. By scanning structures as a function of the groove depth, ranging from 100 nm to 200 nm, we observed that a 3-fold luminescence enhancement could be obtained for the deeper ones. Near-field luminescence was found to be enhanced between the grooves, not inside them, independent of the groove depth and the incident polarization direction. This indicates that the ideal position of the nanocrystals is between the grooves. In addition, we also studied the influence of the inhomogeneities of the perovskite layer and we observed that roughness tends to locally modify the intensity of the fringes and distort their alignment. All the experimental results are in good agreement with numerical simulations.

Large-Area Nanopillar Arrays by Glancing Angle Deposition with Tailored Magnetic Properties.

Ferromagnetic films down to thicknesses of tens of nanometers and composed by polycrystalline Fe and Fe2O3 nanopillars are grown in large areas by glancing angle deposition with magnetron sputtering (MS-GLAD). The morphological features of these films strongly depend on the growth conditions. Vertical or tilted nanopillars have been fabricated depending on whether the substrate is kept rotating azimuthally during deposition or not, respectively. The magnetic properties of these nanopillars films, such as hysteresis loops squareness, adjustable switching fields, magnetic anisotropy and coercivity, can be tuned with the specific morphology. In particular, the growth performed through a collimator mask mounted onto a not rotating azimuthally substrate produces almost isolated well-defined tilted nanopillars that exhibit a magnetic hardening. The first-order reversal curves diagrams and micromagnetic simulations revealed that a growth-induced uniaxial anisotropy, associated with an anisotropic surface morphology produced by the glancing angle deposition in the direction perpendicular to the atomic flux, plays an important role in the observed magnetic signatures. These results demonstrate the potential of the MS-GLAD method to fabricate nanostructured films in large area with tailored structural and magnetic properties for technological applications.

Composition-Dependent Cytotoxic and Antibacterial Activity of Biopolymer-Capped Ag/Au Bimetallic Nanoparticles against Melanoma and Multidrug-Resistant Pathogens.

Nanostructured silver (Ag) and gold (Au) are widely known to be potent biocidal and cytotoxic agents as well as biocompatible nanomaterials. It has been recently reported that combining both metals in a specific chemical composition causes a significant enhancement in their antibacterial activity against antibiotic-resistant bacterial strains, as well as in their anticancer effects, while preserving cytocompatibility properties. In this work, Ag/Au bimetallic nanoparticles over a complete atomic chemical composition range were prepared at 10 at% through a green, highly reproducible, and simple approach using starch as a unique reducing and capping agent. The noble metal nanosystems were thoroughly characterized by different analytical techniques, including UV-visible and FT-IR spectroscopies, XRD, TEM/EDS, XPS and ICP-MS. Moreover, absorption spectra simulations for representative colloidal Ag/Au-NP samples were conducted using FDTD modelling. The antibacterial properties of the bimetallic nanoparticles were determined against multidrug-resistant Escherichia coli and methicillin-resistant Staphylococcus aureus, showing a clear dose-dependent inhibition even at the lowest concentration tested (5 µg/mL). Cytocompatibility assays showed a medium range of toxicity at low and intermediate concentrations (5 and 10 µg/mL), while triggering an anticancer behavior, even at the lowest concentration tested, in a process involving reactive oxygen species production per the nanoparticle Au:Ag ratio. In this manner, this study provides promising evidence that the presently fabricated Ag/Au-NPs should be further studied for a wide range of antibacterial and anticancer applications.

Effects of nanostructuration on the electrochemical performance of metallic bioelectrodes.

The use of metallic nanostructures in the fabrication of bioelectrodes (e.g., neural implants) is gaining attention nowadays. Nanostructures provide increased surface area that might benefit the performance of bioelectrodes. However, there is a need for comprehensive studies that assess electrochemical performance of nanostructured surfaces in physiological and relevant working conditions. Here, we introduce a versatile scalable fabrication method based on magnetron sputtering to develop analogous metallic nanocolumnar structures (NCs) and thin films (TFs) from Ti, Au, and Pt. We show that NCs contribute significantly to reduce the impedance of metallic surfaces. Charge storage capacity of Pt NCs is remarkably higher than that of Pt TFs and that of the other metals in both morphologies. Circuit simulations of the electrode/electrolyte interface show that the signal delivered in voltage-controlled systems is less filtered when nanocolumns are used. In a current-controlled system, simulation shows that NCs provide safer stimulation conditions compared to TFs. We have assessed the durability of NCs and TFs for potential use in vivo by reactive accelerated aging test, mimicking one-year in vivo implantation. Although each metal/morphology reveals a unique response to aging, NCs show overall more stable electrochemical properties compared to TFs in spite of their porous structure.

Pepper-Mediated Green Synthesis of Selenium and Tellurium Nanoparticles with Antibacterial and Anticancer Potential.

The production of nanoparticles for biomedical applications (namely with antimicrobial and anticancer properties) has been significantly hampered using traditional physicochemical approaches, which often produce nanostructures with poor biocompatibility properties requiring post-synthesis functionalization to implement features that such biomedical applications require. As an alternative, green nanotechnology and the synthesis of environmentally friendly nanomaterials have been gaining attention over the last few decades, using living organisms or biomolecules derived from them, as the main raw materials to produce cost-effective, environmentally friendly, and ready-to-be-used nanomaterials. In this article and building upon previous knowledge, we have designed and implemented the synthesis of selenium and tellurium nanoparticles using extracts from fresh jalapeño and habanero peppers. After characterization, in this study, the nanoparticles were tested for both their antimicrobial and anticancer features against isolates of antibiotic-resistant bacterial strains and skin cancer cell lines, respectively. The nanosystems produced nanoparticles via a fast, eco-friendly, and cost-effective method showing different antimicrobial profiles between elements. While selenium nanoparticles lacked an antimicrobial effect at the concentrations tested, those made of tellurium produced a significant antibacterial effect even at the lowest concentration tested. These effects were correlated when the nanoparticles were tested for their cytocompatibility and anticancer properties. While selenium nanoparticles were biocompatible and had a dose-dependent anticancer effect, tellurium-based nanoparticles lacked such biocompatibility while exerting a powerful anti-cancer effect. Further, this study demonstrated a suitable mechanism of action for killing bacteria and cancer cells involving reactive oxygen species (ROS) generation. In summary, this study introduces a new green nanomedicine synthesis approach to create novel selenium and tellurium nanoparticles with attractive properties for numerous biomedical applications.

Green synthesis and characterization of gold-based anisotropic nanostructures using bimetallic nanoparticles as seeds.

Nanostructured noble metals are of great interest because of their tunable optical and electronic properties. However, the green synthesis of anisotropic nanostructures with a defined geometry by the systematic nanoassembly of particles into specific shape, size, and crystallographic facets still faces major challenges. The present work aimed to establish an environmentally friendly methodology for synthesizing gold-based anisotropic nanostructures using starch-capped bimetallic silver/gold nanoparticles as seeds and hydrogen peroxide as a reducing agent.

Correlative confocal and scanning electron microscopy of cultured cells without using dedicated equipment.

This protocol enables correlative light and electron microscopy (CLEM) imaging of cell surface features without using dedicated equipment. Cells are cultured and fixed on transparent substrates for confocal microscopy imaging. No conductive coating is employed in the scanning electron microscopy workflow, providing a clean cell surface observation, with fiducial markers assisting alignment of optical and topographical images. This protocol describes CLEM imaging for midbody remnants in MDCK cells but can also be applied to different cell types and surface features. For complete details on the use and execution of this protocol, please refer to Casares-Arias et al. (2020).

Green synthesis of starch-capped Cu2O nanocubes and their application in the direct electrochemical detection of glucose.

Glucose determination is an essential procedure in different fields, used in clinical analysis for the prevention and monitoring of diabetes. In this work, modified carbon paste electrodes with Cu2O nanocubes (Cu2O NCs) were developed to test electrochemical glucose detection. The synthesis of the Cu2O NCs was achieved by a green method using starch as the capping agent, obtaining cubic-like morphologies and particle sizes from 227 to 123 nm with increasing amounts of the capping agent, as corroborated by electron microscopy analysis. Their crystalline structure and purity were determined by X-ray diffraction. The capability of starch as a capping agent was verified by Fourier-transform infrared spectroscopy, in which the presence of functional groups of this biopolymer in the Cu2O NCs were identified. The electrochemical response to glucose oxidation was determined by cyclic voltammetry, obtaining a linear response of the electrical current as a function of glucose concentration in the range 100-700 µM, with sensitivities from 85.6 to 238.8 µA mM-1 cm-2, depending on the amount of starch used in the synthesis of the Cu2O NCs.

Aloe Vera-Mediated Te Nanostructures: Highly Potent Antibacterial Agents and Moderated Anticancer Effects.

Cancer and antimicrobial resistance to antibiotics are two of the most worrying healthcare concerns that humanity is facing nowadays. Some of the most promising solutions for these healthcare problems may come from nanomedicine. While the traditional synthesis of nanomaterials is often accompanied by drawbacks such as high cost or the production of toxic by-products, green nanotechnology has been presented as a suitable solution to overcome such challenges. In this work, an approach for the synthesis of tellurium (Te) nanostructures in aqueous media has been developed using aloe vera (AV) extracts as a unique reducing and capping agent. Te-based nanoparticles (AV-TeNPs), with sizes between 20 and 60 nm, were characterized in terms of physicochemical properties and tested for potential biomedical applications. A significant decay in bacterial growth after 24 h was achieved for both Methicillin-resistant Staphylococcus aureus and multidrug-resistant Escherichia coli at a relative low concentration of 5 µg/mL, while there was no cytotoxicity towards human dermal fibroblasts after 3 days of treatment. AV-TeNPs also showed anticancer properties up to 72 h within a range of concentrations between 5 and 100 µg/mL. Consequently, here, we present a novel and green approach to produce Te-based nanostructures with potential biomedical applications, especially for antibacterial and anticancer applications.

Midbody Remnant Inheritance Is Regulated by the ESCRT Subunit CHMP4C.

The inheritance of the midbody remnant (MBR) breaks the symmetry of the two daughter cells, with functional consequences for lumen and primary cilium formation by polarized epithelial cells, and also for development and differentiation. However, despite its importance, neither the relationship between the plasma membrane and the inherited MBR nor the mechanism of MBR inheritance is well known. Here, the analysis by correlative light and ultra-high-resolution scanning electron microscopy reveals a membranous stalk that physically connects the MBR to the apical membrane of epithelial cells. The stalk, which derives from the uncleaved side of the midbody, concentrates the ESCRT machinery. The ESCRT CHMP4C subunit enables MBR inheritance, and its depletion dramatically reduces the percentage of ciliated cells. We demonstrate (1) that MBRs are physically connected to the plasma membrane, (2) how CHMP4C helps maintain the integrity of the connection, and (3) the functional importance of the connection.

TiO2 Nanocolumn Arrays for More Efficient and Stable Perovskite Solar Cells.

Organic-inorganic hybrid perovskite solar cells have attracted much attention due to their high power conversion efficiency (>25%) and low-cost fabrication. Yet, improvements are still needed for more stable and higher-performing solar cells. In this work, a series of TiO2 nanocolumn photonic structures have been intentionally fabricated on half of the compact TiO2-coated fluorine-doped tin oxide substrate by glancing angle deposition with magnetron sputtering, a method particularly suitable for industrial applications due to its high reliability and reduced cost when coating large areas. These vertically aligned nanocolumn arrays were then applied as the electron transport layer into triple-cation lead halide perovskite solar cells based on Cs0.05(FA0.83MA0.17)0.95Pb(I0.83Br0.17)3. By comparison to solar cells built onto the same substrate without nanocolumns, the use of TiO2 nanocolumns can significantly enhance the power conversion efficiency of the perovskite solar cells by 7% and prolong their shelf life. Here, detailed characterizations on the morphology and the spectroscopic aspects of the nanocolumns, their near-field and far-field optical properties, solar cells characteristics, as well as the charge transport properties provide mechanistic insights on how one-dimensional TiO2 nanocolumns affect the performance of perovskite halide solar cells in terms of charge transport, light harvesting, and stability, knowledge necessary for the future design of higher-performing and more stable perovskite solar cells.

Integrated miRNA and mRNA expression profiling identifies novel targets and pathological mechanisms in autoimmune thyroid diseases.

BACKGROUND: The mechanisms underlying autoimmune thyroid disease (AITD) remain elusive. Identification of such mechanisms would reveal novel and/or better therapeutic targets. Here, we use integrated analysis of miRNAs and mRNAs expression profiling to identify potential therapeutic targets involved in the mechanisms underlying AITD. METHODS: miRNA and mRNA from twenty fresh-frozen thyroid tissues (15 from AITD patients and 5 from healthy controls) were subjected to next-generation sequencing. An anti-correlated method revealed potential pathways and disease targets, including proteins involved in the formation of primary cilia. Thus, we examined the distribution and length of primary cilia in thyroid tissues from AITD and controls using immunofluorescence and scanning electron microscopy, and parsed cilia formation in thyroid cell lines in response to inflammatory stimuli in the presence of miRNA mimics. FINDINGS: We found that the expression of miR-21-5p, miR-146b-3p, miR-5571-3p and miR-6503-3p was anti-correlated with Enolase 4 (ENO4), in-turned planar cell polarity protein (INTU), kinesin family member 27 (KIF27), parkin co-regulated (PACRG) and serine/threonine kinase 36 (STK36) genes. Functional classification of these miRNA/mRNAs revealed that their differential expression was associated with cilia organization. We demonstrated that the number and length of primary cilia in thyroid tissues was significantly lower in AITD than in control (frequency of follicular ciliated cells in controls = 67.54% vs a mean of 22.74% and 21.61% in HT and GD respectively p = 0.0001, by one-way ANOVA test). In addition, pro-inflammatory cytokines (IFNγ and TNFα) and specific miRNA mimics for the newly identified target genes affected cilia appearance in thyroid cell lines. INTERPRETATION: Integrated miRNA/gene expression analysis has identified abnormal ciliogenesis as a novel susceptibility pathway that is involved in the pathogenesis of AITD. These results reflect that ciliogenesis plays a relevant role in AITD, and opens research pathways to design therapeutic targets in AITD. FUNDING: Instituto de Salud Carlos III, Comunidad de Madrid, Grupo Español de Tumores Neuroendocrinos y Endocrinos, Ministerio de Economía y Empresa and FEDER.

On the Large Near-Field Enhancement on Nanocolumnar Gold Substrates.

One of the most important and distinctive features of plasmonic nanostructures is their ability to confine large electromagnetic fields on nanometric volumes; i.e., the so-called hot spots. The generation, control and characterization of the hot spots are fundamental for several applications, like surface-enhanced spectroscopies. In this work, we characterize the near-field distribution and enhancement of nanostructured gold thin films fabricated by glancing angle deposition magnetron sputtering. These films are composed of columnar nanostructures with high roughness and high density of inter-columnar gaps, where the electromagnetic radiation can be confined, generating hot spots. As expected, the hot spots are localized in the gaps between adjacent nanocolumns and we use scattering-type scanning near-field optical microscopy to image their distribution over the surface of the samples. The experimental results are compared with finite-difference time-domain simulations, finding an excellent agreement between them. The spectral dependence of the field-enhancement is also studied with the simulations, together with surface-enhanced Raman spectroscopy at different excitation wavelengths in the visible-NIR range, proving a broad-band response of the substrates. These findings may result in interesting applications in the field of surface-enhanced optical spectroscopies or sensing.

Comparison of cytocompatibility and anticancer properties of traditional and green chemistry-synthesized tellurium nanowires.

BACKGROUND: Traditional physicochemical approaches for the synthesis of compounds, drugs, and nanostructures developed as potential solutions for antimicrobial resistance or against cancer treatment are, for the most part, facile and straightforward. Nevertheless, these approaches have several limitations, such as the use of toxic chemicals and production of toxic by-products with limited biocompatibility. Therefore, new methods are needed to address these limitations, and green chemistry offers a suitable and novel answer, with the safe and environmentally friendly design, manufacturing, and use of minimally toxic chemicals. Green chemistry approaches are especially useful for the generation of metallic nanoparticles or nanometric structures that can effectively and efficiently address health care concerns. OBJECTIVE: Here, tellurium (Te) nanowires were synthesized using a novel green chemistry approach, and their structures and cytocompatibility were evaluated. METHOD: An easy and straightforward hydrothermal method was employed, and the Te nanowires were characterized using transmission electron microscopy, scanning electron microscopy, energy-dispersive X-ray spectroscopy, Fourier-transform infrared spectroscopy, X-ray powder diffraction, X-ray photoelectron spectroscopy, and optical microscopy for morphology, size, and chemistry. Cytotoxicity tests were performed with human dermal fibroblasts and human melanoma cells (to assess anticancer properties). The results showed that a treatment with Te nanowires at concentrations between 5 and 100 µg/mL improved the proliferation of healthy cells and decreased cancerous cell growth over a 5-day period. Most importantly, the green chemistry -synthesized Te nanowires outperformed those produced by traditional synthetic chemical methods. CONCLUSION: This study suggests that green chemistry approaches for producing Te nanostructures may not only reduce adverse environmental effects resulting from traditional synthetic chemistry methods, but also be more effective in numerous health care applications.

Mapping plasmon-enhanced upconversion fluorescence of Er/Yb-doped nanocrystals near gold nanodisks.

Fluorescence enhancement effects have many potential applications in the domain of biochemical sensors and optoelectronic devices. Here, the emission properties of up-converting nanocrystals near nanostructures that support surface plasmon resonances have been investigated. Gold nanodisks of various diameters were illuminated in the near-infrared (λ = 975 nm) and a single fluorescent nanocrystal glued at the end of an atomic force microscope tip was scanned around them. By detecting its visible fluorescence around each structure, it is found that the highest fluorescence enhancement occurs in a zone that forms a two-lobe pattern near the nanodisks and which corresponds to the map of the near-field intensity calculated at the excitation wavelength. In agreement with numerical simulations, it is also observed that the maximum fluorescence enhancement takes place when the disk diameter is around 200 nm. Surprisingly, this disk size is small when compared to that yielding the highest far-field scattering resonance, which occurs for disks with a diameter of 300-350 nm at the same excitation wavelength. This shift between the near and far-field resonances should be taken into account in the design of structures in systems that use plasmon enhanced fluorescence effects.

Synergic antibacterial coatings combining titanium nanocolumns and tellurium nanorods.

Nanocolumnar titanium coatings have been fabricated in two sputtering systems with very different characteristics (a laboratory setup and semi-industrial equipment), thus possessing different morphologies (150 nm long columns tilted 20° from the normal and 300 nm long ones tilted 40°, respectively). These coatings exhibit similar antibacterial properties against Gram positive (Staphylococcus aureus) and Gram negative (Escherichia coli) bacteria. When a synergic route is followed and these coatings are functionalized with tellurium (Te) nanorods, the antibacterial properties are enhanced, especially for the long nanocolumns case. The biocompatibility is preserved in all the nanostructured coatings.

Fabrication of black-gold coatings by glancing angle deposition with sputtering.

The fabrication of black-gold coatings using sputtering is reported here. Glancing angle deposition with a rotating substrate is needed to obtain vertical nanostructures. Enhanced light absorption is obtained in the samples prepared in the ballistic regime with high tilt angles. Under these conditions the diameter distribution of the nanostructures is centered at about 60 nm and the standard deviation is large enough to obtain black-metal behavior in the visible range.

Magnetic field modulation of chirooptical effects in magnetoplasmonic structures.

In this work we analyse the magnetic field effects on the chirooptical properties of magnetoplasmonic chiral structures. The structures consist of two-dimensional arrays of Au gammadions in which thin layers of Co have been inserted. Due to the magnetic properties of the Au/Co interface the structures have perpendicular magnetic anisotropy which favours magnetic saturation along the surface normal, allowing magnetic field modulation of the chirooptical response with moderate magnetic fields. These structures have two main resonances. The resonance at 850 nm has a larger chirooptical response than the resonance at 650 nm, which, on the other hand, exhibits a larger magnetic field modulation of its chirooptical response. This dissimilar behaviour is due to the different physical origin of the chirooptical and magneto-optical responses. Whereas the chirooptical effects are due to the geometry of the structures, the magneto-optical response is related to the intensity of the electromagnetic field in the magnetic (Co) layers. We also show that the optical chirality can be modulated by the applied magnetic field, which suggests that magnetoplasmonic chiral structures could be used to develop new strategies for chirooptical sensing.