Publications

Alzheimer's disease is characterized by a combination of several neuropathological hallmarks, such as extracellular aggregates of beta amyloid (Aβ). Numerous alternatives have been studied for inhibiting Aβ aggregation but, at this time, there are no effective treatments available. Here, we developed the tri-component nanohybrid system AuNPs@POM@PEG based on gold nanoparticles (AuNPs) covered with polyoxometalates (POMs) and polyethylene glycol (PEG). In this work, AuNPs@POM@PEG demonstrated the inhibition of the formation of amyloid fibrils, showing a 75% decrease in Aβ aggregation in vitro. As it is a potential candidate for the treatment of Alzheimer's disease, we evaluated the cytotoxicity of AuNPs@POM@PEG and its ability to cross the blood-brain barrier (BBB). We achieved a stable nanosystem that is non-cytotoxic below 2.5 nM to human neurovascular cells. The brain permeability of AuNPs@POM@PEG was analyzed in an in vitro microphysiological model of the BBB (BBB-on-a-chip), containing 3D human neurovascular cell co-cultures and microfluidics. The results show that AuNPs@POM@PEG was able to cross the brain endothelial barrier in the chip and demonstrated that POM does not affect the barrier integrity, giving the green light to further studies into this system as a nanotherapeutic.

JTD Keywords: beta-amyloid, blood-brain barrier organ-on-a-chip, cellular uptake, citrate, cytotoxicity, electrocatalytic reduction, gold nanoparticles, hypothesis, nanorods, polyoxometalates, size, stability, surface, Alzheimers-disease, Blood–brain barrier organ-on-a-chip, Gold nanoparticles, Nanovehicle, Polyoxometalates, Β-amyloid

This work presents a novel nanoparticle-based thermosensor implant able to reveal the precise temperature variations along the polymer filaments, as it contracts and expands due to changes in the macroscale local temperature. The multimodal device is able to trace the position and the temperature of a polypropylene mesh, employed in abdominal hernia repair, by combining plasmon resonance and Raman spectroscopy with hydrogel responsive system. The novelty relies on the attachment of the biocompatible nanoparticles, based on gold stabilized by a chitosan-shell, already charged with the Raman reporter (RaR) molecules, to the robust prosthesis, without the need of chemical linkers. The SERS enhanced effect observed is potentiated by the presence of a quite thick layer of the copolymer (poly(N-isopropylacrylamide)-co-poly(acrylamide)) hydrogel. At temperatures above the LCST of PNIPAAm-co-PAAm, the water molecules are expulsed and the hydrogel layer contracts, leaving the RaR molecules more accessible to the Raman source. In vitro studies with fibroblast cells reveal that the functionalized surgical mesh is biocompatible and no toxic substances are leached in the medium. The mesh sensor opens new frontiers to semi-invasive diagnosis and infection prevention in hernia repair by using SERS spectroscopy. It also offers new possibilities to the functionalization of other healthcare products.© 2023 Wiley-VCH GmbH.

JTD Keywords: adhesion, blends, chitosan, gold nanoparticles, poly(n-isopropylacrylamide), polypropylene mesh, polypropylene meshes, repair, scattering, silver, surgical implants, thermosensitive hydrogels, toxicity, Chitosan, Gold nanoparticles, Polypropylene meshes, Surgical implants, Thermosensitive hydrogels

The lack of predictive models that mimic the blood-brain barrier (BBB) hinders the development of effective drugs for neurodegenerative diseases. Animal models behave differently from humans, are expensive and have ethical constraints. Organ-on-a-chip (OoC) platforms offer several advantages to resembling physiological and pathological conditions in a versatile, reproducible, and animal-free manner. In addition, OoC give us the possibility to incorporate sensors to determine cell culture features such as trans-endothelial electrical resistance (TEER). Here, we developed a BBB-on-a-chip (BBB-oC) platform with a TEER measurement system in close distance to the barrier used for the first time for the evaluation of the permeability performance of targeted gold nanorods for theranostics of Alzheimer's disease. GNR-PEG-Ang2/D1 is a therapeutic nanosystem previously developed by us consisting of gold nanorods (GNR) functionalized with polyethylene glycol (PEG), angiopep-2 peptide (Ang2) to overcome the BBB and the D1 peptide as beta amyloid fibrillation inhibitor, finally obtaining GNR-PEG-Ang2/D1 which showed to be useful for disaggregation of the amyloid in in vitro and in vivo models. In this work, we evaluated its cytotoxicity, permeability, and some indications of its impact on the brain endothelium by employing an animal-free device based on neurovascular human cells.In this work, we fabricated a BBB-oC with human astrocytes, pericytes and endothelial cells and a TEER measuring system (TEER-BBB-oC) integrated at a micrometric distance of the endothelial barrier. The characterization displayed a neurovascular network and the expression of tight junctions in the endothelium. We produced GNR-PEG-Ang2/D1 and determined its non-cytotoxic range (0.05-0.4 nM) for plated cells included in the BBB-oC and confirmed its harmless effect at the highest concentration (0.4 nM) in the microfluidic device. The permeability assays revealed that GNR-PEG-Ang2/D1 cross the BBB and this entry is facilitated by Ang2 peptide. Parallel to the permeability analysis of GNR-PEG-Ang2/D1, an interesting behavior of the TJs expression was observed after its administration probably related to the ligands on the nanoparticle surface.BBB-oC with a novel TEER integrated setup which allow a correct read-out and cell imaging monitoring was proven as a functional and throughput platform to evaluate the brain permeability performance of nanotherapeutics in a physiological environment with human cells, putting forward a viable alternative to animal experimentation.© 2023. The Author(s).

JTD Keywords: alzheimer disease (ad), cell-culture, cytotoxicity, endothelial-cells, gold nanoparticles, microfluidic platform, model, organ-on-a-chip (ooc), peptide, tight junction, trans-endothelial electrical resistance (teer), transport, Alzheimer disease (ad), Blood-brain barrier (bbb), Blood-brain-barrier, Blood–brain barrier (bbb), Gold nanoparticles, Organ-on-a-chip (ooc), Trans-endothelial electrical resistance (teer)

The quest for novel nanoscale materials for different applications necessitates that they are easy to obtain and have excellent physical properties and low toxicity. Moreover, considering the ongoing environmental impact of noxious chemical waste products, it is important to adopt eco-friendly approaches for nanoparticle synthesis. In this work, a natural polymer (medium molecular weight chitosan) derived from chitin was employed as a reducing agent to obtain gold nanoparticles (AuNPs) with a chitosan shell (AuNPs@CS) by a microwave oven. The chitosan is economically viable and cost-competitive in the market showing also nontoxic behavior in the environment and living organisms. The synthesized AuNPs@CS-FITC NPs were fully characterized by spectroscopic and microscopic characterization techniques. The size distribution of NPs was about 15 nm, which is a suitable dimension to use in biomedical applications due to their high tissue penetration, great circulation in blood, and optimal clearance as well as low toxicity. The prepared polymer-capped NPs were further functionalized with a fluorescent molecule, i.e., Fluorescein-5-isothiocyanate (FITC), to perform imaging in the cell. The results highlighted the goodness of the synthesis procedure, as well as the high internalization rate that resulted in an optimal fluorescence intensity. Thus, this work presents a good sustainable/green approach-mediated polymer nanocomposite for various applications in the field of diagnostic imaging.

JTD Keywords: Agent, Biomedical, Fluorescent, Gold nanoparticles, Green synthesis, Nanomaterials, Silver nanoparticles

Polypropylene (PP) surgical meshes, with different knitted architectures, were chemically functionalized with gold nanoparticles (AuNPs) and 4-mercaptothiazole (4-MB) to transform their fibers into a surface enhanced Raman scattering (SERS) detectable plastic material. The application of a thin layer of poly[N-isopropylacrylamide-co-N,N'-methylene bis(acrylamide)] (PNIPAAm-co-MBA) graft copolymer, covalently polymerized to the mesh-gold substrate, caused the conversion of the inert plastic into a thermoresponsive material, resulting in the first PP implantable mesh with both SERS and temperature stimulus responses. AuNPs were homogeneously distributed over the PP yarns, offering a clear SERS recognition together with higher PNIPAAm lower critical solution temperature (LCST ∼ 37 °C) than without the metallic particles (LCST ∼ 32 °C). An infrared thermographic camera was used to observe the polymer-hydrogel folding-unfolding process and to identify the new value of the LCST, connected with the heat generation by plasmonic-resonance gold NPs. The development of SERS PP prosthesis will be relevant for the bioimaging and biomarker detection of the implant by using the plasmonic effect and Raman vibrational spectroscopy for minimally invasive interventions (such as laparoscopy), to prevent patient inflammatory processes. Furthermore, Raman sources have been proved to not damage the cells, like happens with near-infrared irradiation, representing another advantage of moving to SERS approaches. The findings reported here offer unprecedented application possibilities in the biomedical field by extrapolating the material functionalization to other nonabsorbable polymer made devices (e.g., surgical sutures, grapes, wound dressings, among others).

JTD Keywords: gold nanoparticles, poly(n-isopropylacrylamide), polymers, polypropylene, raman-spectroscopy, reduction, resonance, sers spectroscopy, size, surface functionalization, Gold nanoparticles, Polypropylene, Surface functionalization

Quantum dots (QDs) are semiconductor nanoparticles with unique optical and electronic properties, whose interest as potential nano-theranostic platforms for imaging and sensing is increasing. The design and use of QDs requires the understanding of cell-nanoparticle interactions at a microscopic and nanoscale level. Model systems such as supported lipid bilayers (SLBs) are useful, less complex platforms mimicking physico-chemical properties of cell membranes. In this work, we investigated the effect of topographical homogeneity of SLBs bearing different surface charge in the adsorption of hydrophilic QDs. Using quartz-crystal microbalance, a label-free surface sensitive technique, we show significant differences in the interactions of QDs onto homogeneous and inhomogeneous SLBs formed following different strategies. Within short time scales, QDs adsorb onto topographically homogeneous, defect-free SLBs is driven by electrostatic interactions, leading to no layer disruption. After prolonged QD exposure, the nanomechanical stability of the SLB decreases suggesting nanoparticle insertion. In the case of inhomogeneous, defect containing layers, QDs target preferentially membrane defects, driven by a subtle interplay of electrostatic and entropic effects, inducing local vesicle rupture and QD insertion at membrane edges. © 2021

JTD Keywords: adsorption, atomic force microscopy, bilayer formation, gold nanoparticles, hydrophilic quantum dots, lipid membrane defects, model, nanomechanics, quartz crystal microbalance with dissipation, size, supported lipid bilayers, surfaces, Atomic force microscopy, Atomic-force-microscopy, Cytology, Defect-free, Electronic properties, Electrostatics, Hydrophilic quantum dot, Hydrophilic quantum dots, Hydrophilicity, Hydrophilics, Lipid bilayers, Lipid membrane defect, Lipid membrane defects, Lipid membranes, Lipids, Nanocrystals, Nanomechanics, Optical and electronic properties, Quartz, Quartz crystal microbalance with dissipation, Quartz crystal microbalances, Quartz-crystal microbalance, Semiconductor nanoparticles, Semiconductor quantum dots, Supported lipid bilayers

The use of silver nanoparticles (Ag NPs) in the biomedical field deserves a mindful analysis of the possible inflammatory response which could limit their use in the clinic. Despite the anti-cancer properties of Ag NPs having been widely demonstrated, there are still few studies concerning their involvement in the activation of specific inflammatory pathways. The inflammatory outcome depends on the synthetic route used in the NPs production, in which toxic reagents are employed. In this work, we compared two types of Ag NPs, obtained by two different chemical routes: conventional synthesis using sodium citrate and a green protocol based on leaf extracts as a source of reduction and capping agents. A careful physicochemical characterization was carried out showing spherical and stable Ag NPs with an average size between 20 nm and 35 nm for conventional and green Ag NPs respectively. Then, we evaluated their ability to induce the activation of inflammation in Human Leukemic Monocytes (THP-1) differentiated into M0 macrophages using 1 µM and 2 µM NPs concentrations (corresponded to 0.1 µg/mL and 0.2 µg/mL respectively) and two-time points (24 h and 48 h). Our results showed a clear difference in Nuclear Factor ?B (NF-?b) activation, Interleukins 6–8 (IL-6, IL-8) secretion, Tumor Necrosis Factor-? (TNF-?) and Cyclooxygenase-2 (COX-2) expression exerted by the two kinds of Ag NPs. Green Ag NPs were definitely tolerated by macrophages compared to conventional Ag NPs which induced the activation of all the factors mentioned above. Subsequently, the exposure of breast cancer cell line (MCF-7) to the green Ag NPs showed that they exhibited antitumor activity like the conventional ones, but surprisingly, using the MCF-10A line (not tumoral breast cells) the green Ag NPs did not cause a significant decrease in cell viability. © 2022 by the authors. Licensee MDPI, Basel, Switzerland.

JTD Keywords: activation, biosynthesis, gold nanoparticles, green route, inflammation response, mechanism, metal, nanotechnology, physico-chemical properties, raman-spectroscopy, resonance, silver nanoparticles, surface, Biomedical fields, Cell culture, Cell death, Chemical activation, Chemical routes, Conventional synthesis, Diseases, Green route, Inflammation response, Inflammatory response, Macrophages, Metal nanoparticles, Nf-kappa-b, Pathology, Physico-chemical properties, Physicochemical property, Property, Silver nanoparticles, Sodium compounds, Synthetic routes, Toxic reagents

The SARS-CoV-2 outbreaks highlighted the need for effective, reliable, fast, easy-to-do and cheap diagnostics procedures. We pragmatically experienced that an early positive-case detection, inevitably coupled with a mass vaccination campaign, is a milestone to control the COVID-19 pandemic. Gold nanoparticles (AuNPs) can indeed play a crucial role in this context, as their physicochemical, optics and electronics properties are being extensively used in photothermal therapy (PTT), radiation therapy (RT), drug delivery and diagnostic. AuNPs can be synthesized by several approaches to obtain different sizes and shapes that can be easily functionalized with many kinds of molecules such as antibodies, proteins, probes, and lipids. In addition, AuNPs showed high biocompatibility making them useful tool in medicine field. We thus reviewed here the most relevant evidence on AuNPs as effective way to detect the presence of SARS-CoV-2 antigens. We trust future diagnostic efforts must take this 'old-fashioned' nanotechnology tool into consideration for the development and commercialization of reliable and feasible detection kits.

JTD Keywords: Aggregation, Antibodies, Assay, Covid-19, Diagnosis, Enhanced raman-scattering, Gold nanoparticles, Immunoassay, Pandemic disease, Physicochemical properties, Rapid detection, Sars-cov-2, Sensors, Surface-plasmon resonance, Therapy

© 2020 Elsevier Ltd The use of nanoparticles in consumer products is currently on the rise, so it is important to have reliable methods to predict any associated toxicity effects. Traditional in vitro assays fail to mimic true physiological responses of living organisms against nanoparticles whereas murine in vivo models are costly and ethically controversial. For these reasons, this study aimed to evaluate the efficacy of Galleria mellonella as an alternative, non-rodent in vivo model for examining nanoparticle toxicity. Silver, selenium, and functionalized gold nanoparticles were synthesized, and their toxicity was assessed in G. mellonella larvae. The degree of acute toxicity effects caused by each type of NP was efficiently detected by an array of indicators within the larvae: LD50 calculation, hemocyte proliferation, NP distribution, behavioral changes, and histological alterations. G. mellonella larvae are proposed as a nanotoxicological model that can be used as a bridge between in vitro and in vivo murine assays in order to obtain better predictions of NP toxicity.

JTD Keywords: cellular uptake, cytotoxicity, galleria mellonella, gold nanoparticles, hemocytes, nanoparticles, nanotoxicity, non-rodent in vivo model, non-rodent in vivo model, oxidative stress, selenium-compounds, silica nanoparticles, silver nanoparticles, toxicity, toxicity screening, vitro, Galleria mellonella, Hemocytes, In-vivo model, Nanoparticles, Nanotoxicity, Non-rodent in vivo model, Toxicity screening

© 2020 The Authors Although the concept of selective delivery has been postulated over 100 years ago, no targeted nanomedicine has been clinically approved so far. Nanoparticles modified with targeting ligands to promote the selective delivery of therapeutics towards a specific cell population have been extensively reported. However, the rational design of selective particles is still challenging. One of the main reasons for this is the lack of quantitative theoretical and experimental understanding of the interactions involved in cell targeting. In this review, we discuss new theoretical models and experimental methods that provide a quantitative view of targeting. We show the new advancements in multivalency theory enabling the rational design of super-selective nanoparticles. Furthermore, we present the innovative approaches to obtain key targeting parameters at the single-cell and single molecule level and their role in the design of targeting nanoparticles. We believe that the combination of new theoretical multivalent design and experimental methods to quantify receptors and ligands aids in the rational design and clinical translation of targeted nanomedicines.

JTD Keywords: binding-kinetics, biological identity, biomolecular corona, blood-brain-barrier, drug-delivery, gold nanoparticles, multivalency, nanotechnology, protein corona, quantitative characterization, rational design, super-selectivity, superresolution microscopy, tumor heterogeneity, Ligand-receptor interactions, Multivalency, Nanotechnology, Quantitative characterization, Rational design, Super-selectivity

Engineered immunoglobulin-G molecules (IgGs) are of wide interest for the development of detection elements in protein-based biosensors with clinical applications. The strategy usually employed for the de novo design of such engineered IgGs consists on merging fragments of the three-dimensional structure of a native IgG, which is immobilized on the biosensor surface, and of an antibody with an exquisite target specificity and affinity. In this work conventional and accelerated classical molecular dynamics (cMD and aMD, respectively) simulations have been used to propose two IgG-like antibodies for COVID-19 detection. More specifically, the crystal structure of the IgG1 B12 antibody, which inactivates the human immunodeficiency virus-1, has been merged with the structure of the antibody CR3022 Fab tightly bounded to SARS-CoV-2 receptor-binding domain (RBD) and the structure of the 5309 antibody Fab fragment complexed with SARS-CoV-2 RBD. The two constructed antibodies, named IgG1-CR3022 and IgG1-S309, respectively, have been immobilized on a stable gold surface through a linker. Analyses of the influence of both the merging strategy and the substrate on the stability of the two constructs indicate that the IgG1-S309 antibody better preserves the neutralizing structure than the IgG1-CR3022 one. Overall, results indicate that the IgG1-S309 is appropriated for the generation of antibody based sensors for COVID-19 diagnosis. (C) 2021 The Author(s). Published by Elsevier B.V. on behalf of Research Network of Computational and Structural Biotechnology.

JTD Keywords: cr3022, igg1, molecular engineering, s309, Antibodies, Antibody engineering, Biosensors, Chemical detection, Clinical application, Cov, Cr3022, Crystal structure, Design, Diseases, Gold nanoparticles, Igg1, Igg1 antibody, Immobilization, Immunoglobulin g, Immunosensor, In-silico, Merging, Molecular dynamics, Molecular engineering, Orientation, Protein-based biosensors, Receptor-binding domains, S309, Sars, Sensor, Spike protein, Target, Vaccine, Viruses

The inclusion compound (IC) of cyclodextrin (CD) containing the antitumor drug Methotrexate (MTX) as a guest molecule was obtained to increase the solubility of MTX and decrease its inherent toxic effects in nonspecific cells. The IC was conjugated with gold nanoparticles (AuNPs), obtained by a chemical method, creating a ternary intelligent delivery system for MTX molecules, based on the plasmonic properties of the AuNPs. Irradiation of the ternary system, with a laser wavelength tunable with the corresponding surface plasmon of AuNPs, causes local energy dissipation, producing the controlled release of the guest from CD cavities. Finally, cell viability was evaluated using MTS assays for β-CD/MTX and AuNPs + β-CD/MTX samples, with and without irradiation, against HeLa tumor cells. The irradiated sample of the ternary system AuNPs + β-CD/MTX produced a diminution in cell viability attributed to the photothermal release of MTX.

JTD Keywords: Cyclodextrin, Delivery system, Gold nanoparticles, Inclusion compound, Irradiation, Laser, Methotrexate, Photothermal release

JTD Keywords: DNA nanotechnology, Gold nanoparticles, Lithography, Metasurfaces, Nanopatterning

We report the first DNA-based origami technique that can print addressable patterns on surfaces with sub-10 nm resolution. Specifically, we have used a two-dimensional DNA origami as a template (DNA origami stamp) to transfer DNA with pre-programmed patterns (DNA ink) on gold surfaces. The DNA ink is composed of thiol-modified staple strands incorporated at specific positions of the DNA origami stamp to create patterns upon thiol-gold bond formation on the surface (DNA ink). The DNA pattern formed is composed of unique oligonucleotide sequences, each of which is individually addressable. As a proof-of-concept, we created a linear pattern of oligonucleotide-modified gold nanoparticles complementary to the DNA ink pattern. We have developed an in silico model to identify key elements in the formation of our DNA origami-driven lithography and nanoparticle patterning as well as simulate more complex nanoparticle patterns on surfaces.

JTD Keywords: DNA nanotechnology, Lithography, Nanopatterning, Gold nanoparticles, Metasurfaces

Evidence of guest migration in α-cyclodextrin-octylamine (α-CD-OA) inclusion compound (IC) generated via plasmonic heating of gold nanoparticles (AuNPs) has been studied. In this report, we demonstrate local effects generated by laser-mediated irradiation of a sample of AuNPs covered with inclusion compounds on surface-derivatized glass under liquid conditions by atomic force microscopy (AFM). Functionalized AuNPs on the glass and covered by the ICs were monitored by recording images by AFM during 5 h of irradiation, and images showed that after irradiation, a drastic decrease in the height of the AuNPs occurred. The absorption spectrum of the irradiated sample showed a hypsochromic shift from 542 to 536 nm, evidence suggesting that much of the population of nanoparticles lost all of the parts of the overlay of ICs due to the plasmonic heat generated by the irradiation. Mass spectrometry matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF) performed on a sample containing a collection of drops obtained from the surface of the functionalized glass provided evidence that the irradiation lead to disintegration of the ICs and therefore exit of the octylamine molecule (the guest) from the cyclodextrin cavity (the matrix).

JTD Keywords: Cyclodextrin inclusion compound, Gold nanoparticles, Guest migration, Plasmonic heating