Publications by authors named "Paolo Matteini"

44 Publications

Probing the Structure of Toxic Amyloid-β Oligomers with Electron Spin Resonance and Molecular Modeling.

ACS Chem Neurosci 2021 04 16;12(7):1150-1161. Epub 2021 Mar 16.

National Research Council of Italy (CNR), Institute of Chemistry of Organometallic Compounds (ICCOM), I-56124 Pisa, Italy.

Structural models of the toxic species involved in the development of Alzheimer's disease are of utmost importance to understand the molecular mechanism and to describe early biomarkers of the disease. Among toxic species, soluble oligomers of amyloid-β (Aβ) peptides are particularly important, because they are responsible for spreading cell damages over brain regions, thus rapidly impairing brain functions. In this work we obtain structural information on a carefully prepared Aβ(1-42) sample, representing a toxic state for cell cultures, by combining electron spin resonance spectroscopy and computational models. We exploited the binding of Cu to Aβ(1-42) and used copper as a probe for estimating Cu-Cu distances in the oligomers by applying double electron-electron resonance (DEER) pulse sequence. The DEER trace of this sample displays a unique feature that fits well with structural models of oligomers formed by Cu-cross-linked peptide dimers. Because Cu is bound to the Aβ(1-42) N-terminus, for the first time structural constraints that are missing in reported studies are provided at physiological conditions for the Aβ N-termini. These constraints suggest the Aβ(1-42) dimer as the building block of soluble oligomers, thus changing the scenario for any kinetic model of Aβ(1-42) aggregation.
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http://dx.doi.org/10.1021/acschemneuro.0c00714DOI Listing
April 2021

Photobiomodulation of Human Fibroblasts and Keratinocytes with Blue Light: Implications in Wound Healing.

Biomedicines 2021 Jan 5;9(1). Epub 2021 Jan 5.

Istituto di Fisica Applicata "Nello Carrara", Consiglio Nazionale delle Ricerche (CNR-IFAC), 50019 Florence, Italy.

In recent years, photobiomodulation (PBM) has been recognized as a physical therapy in wound management. Despite several published research papers, the mechanism underlying photobiomodulation is still not completely understood. The investigation about application of blue light to improve wound healing is a relatively new research area. Tests in selected patients evidenced a stimulation of the healing process in superficial and chronic wounds treated with a blue LED light emitting at 420 nm; a study in animal model pointed out a faster healing process in superficial wound, with an important role of fibroblasts and myofibroblasts. Here, we present a study aiming at evidencing the effects of blue light on the proliferation and metabolism in fibroblasts from healthy skin and keratinocytes. Different light doses (3.43, 6.87, 13.7, 20.6, 30.9 and 41.2 J/cm2) were used to treat the cells, evidencing inhibitory and stimulatory effects following a biphasic dose behavior. Electrophysiology was used to investigate the effects on membrane currents: healthy fibroblasts and keratinocytes showed no significant differences between treated and not treated cells. Raman spectroscopy revealed the mitochondrial Cytochrome C (Cyt C) oxidase dependence on blue light irradiation: a significant decrease in peak intensity of healthy fibroblast was evidenced, while it is less pronounced in keratinocytes. In conclusion, we observed that the blue LED light can be used to modulate metabolism and proliferation of human fibroblasts, and the effects in wound healing are particularly evident when studying the fibroblasts and keratinocytes co-cultures.
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http://dx.doi.org/10.3390/biomedicines9010041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7824830PMC
January 2021

Experimental Study on Blue Light Interaction with Human Keloid-Derived Fibroblasts.

Biomedicines 2020 Dec 6;8(12). Epub 2020 Dec 6.

Istituto di Fisica Applicata "Nello Carrara", Consiglio Nazionale delle Ricerche (CNR-IFAC), 50019 Florence, Italy.

Keloids are an exuberant response to wound healing, characterized by an exaggerated synthesis of collagen, probably due to the increase of fibroblasts activity and to the reduction of their apoptosis rate: currently no standard treatments or pharmacological therapies are able to prevent keloid recurrence. To reach this goal, in recent years some physical treatments have been proposed, and among them the PhotoBioModulation therapy (PBM). This work analyses the effects of a blue LED light irradiation (410-430 nm, 0.69 W/cm power density) on human fibroblasts, isolated from both keloids and perilesional tissues. Different light doses (3.43-6.87-13.7-20.6-30.9 and 41.2 J/cm) were tested. Biochemical assays and specific staining were used to assess cell metabolism, proliferation and viability. Micro-Raman spectroscopy was used to explore direct effects of the blue LED light on the Cytochrome C (Cyt C) oxidase. We also investigated the effects of the irradiation on ionic membrane currents by patch-clamp recordings. Our results showed that the blue LED light can modulate cell metabolism and proliferation, with a dose-dependent behavior and that these effects persist at least till 48 h after treatment. Furthermore, we demonstrated that the highest fluence value can reduce cell viability 24 h after irradiation in keloid-derived fibroblasts, while the same effect is observed 48 h after treatment in perilesional fibroblasts. Electrophysiological recordings showed that the medium dose (20.6 J/cm) of blue LED light induces an enhancement of voltage-dependent outward currents elicited by a depolarizing ramp protocol. Overall, these data demonstrate the potentials that PBM shows as an innovative and minimally-invasive approach in the management of hypertrophic scars and keloids, in association with current treatments.
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http://dx.doi.org/10.3390/biomedicines8120573DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7762279PMC
December 2020

Label-free SERS detection of proteins based on machine learning classification of chemo-structural determinants.

Analyst 2021 Jan 19;146(2):674-682. Epub 2020 Nov 19.

Institute of Applied Physics "Nello Carrara", Italian National Research Council, via Madonna del Piano 10, Sesto Fiorentino, I-50019, Italy.

Establishing standardized methods for a consistent analysis of spectral data remains a largely underexplored aspect in surface-enhanced Raman spectroscopy (SERS), particularly applied to biological and biomedical research. Here we propose an effective machine learning classification of protein species with closely resembled spectral profiles by a mixed data processing based on principal component analysis (PCA) applied to multipeak fitting on SERS spectra. This strategy simultaneously assures a successful discrimination of proteins and a thorough characterization of the chemostructural differences among them, ultimately opening up new routes for SERS evolution toward sensing applications and diagnostics of interest in life sciences.
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http://dx.doi.org/10.1039/d0an02137gDOI Listing
January 2021

Emergence of Barrel Motif in Amyloid-β Trimer: A Computational Study.

J Phys Chem B 2020 11 12;124(47):10617-10631. Epub 2020 Nov 12.

National Research Council of Italy (CNR), Institute for Chemistry of Organometallic Compounds (ICCOM), 50019 Florence, Italy.

Amyloid-β (Aβ) peptides form assemblies that are pathological hallmarks of Alzheimer's disease. Aβ oligomers are soluble, mobile, and toxic forms of the peptide that act in the extracellular space before assembling into protofibrils and fibrils. Therefore, oligomers play an important role in the mechanism of Alzheimer's disease. Since it is difficult to determine by experiment the atomic structures of oligomers, which accumulate fast and are polymorphic, computer simulation is a useful tool to investigate elusive oligomers' structures. In this work, we report extended all-atom molecular dynamics simulations, both canonical and replica exchange, of Aβ(1-42) trimer starting from two different initial conformations: (i) the pose produced by the best docking of a monomer aside of a dimer (simulation ), representing oligomers freshly formed by assembling monomers, and (ii) a configuration extracted from an experimental mature fibril structure (simulation ), representing settled oligomers in equilibrium with extended fibrils. We showed that in simulation , regions with small β-barrels are populated, indicating the chance of spontaneous formation of domains resembling channel-like structures. These structural domains are alternative to those more representative of mature fibrils (simulation ), the latter showing a stable bundle of C-termini that is not sampled in simulation . Moreover, trimer of Aβ(1-42) can form internal pores that are large enough to be accessed by water molecules and Ca ions.
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http://dx.doi.org/10.1021/acs.jpcb.0c05508DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7735726PMC
November 2020

Contributions of Molecular and Optical Techniques to the Clinical Diagnosis of Alzheimer's Disease.

Brain Sci 2020 Nov 3;10(11). Epub 2020 Nov 3.

Fondazione IRCCS Istituto Neurologico Carlo Besta, Division of Neurology 5 and Neuropathology, 20133 Milan, Italy.

Alzheimer's disease (AD) is the most common neurodegenerative disorder worldwide. The distinctive neuropathological feature of AD is the intracerebral accumulation of two abnormally folded proteins: β-amyloid (Aβ) in the form of extracellular plaques, and tau in the form of intracellular neurofibrillary tangles. These proteins are considered disease-specific biomarkers, and the definite diagnosis of AD relies on their post-mortem identification in the brain. The clinical diagnosis of AD is challenging, especially in the early stages. The disease is highly heterogeneous in terms of clinical presentation and neuropathological features. This phenotypic variability seems to be partially due to the presence of distinct Aβ conformers, referred to as strains. With the development of an innovative technique named Real-Time Quaking-Induced Conversion (RT-QuIC), traces of Aβ strains were found in the cerebrospinal fluid of AD patients. Emerging evidence suggests that different conformers may transmit their strain signature to the RT-QuIC reaction products. In this review, we describe the current challenges for the clinical diagnosis of AD and describe how the RT-QuIC products could be analyzed by a surface-enhanced Raman spectroscopy (SERS)-based systems to reveal the presence of strain signatures, eventually leading to early diagnosis of AD with the recognition of individual disease phenotype.
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http://dx.doi.org/10.3390/brainsci10110815DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7692713PMC
November 2020

Hollow core photonic crystal fiber-assisted Raman spectroscopy as a tool for the detection of Alzheimer's disease biomarkers.

J Biomed Opt 2020 07;25(7):1-10

Tel Aviv Univ., Israel.

Significance: Alzheimer's disease (AD) is an irreversible and progressive disorder that damages brain cells and impairs the cognitive abilities of the affected. Developing a sensitive and cost-effective method to detect Alzheimer's biomarkers appears vital in both a diagnostic and therapeutic perspective.

Aim: Our goal is to develop a sensitive and reliable tool for detection of amyloid β (1-42) peptide (Aβ42), a major AD biomarker, using fiber-enhanced Raman spectroscopy (FERS).

Approach: A hollow core photonic crystal fiber (HCPCF) was integrated with a conventional Raman spectroscopic setup to perform FERS measurements. FERS was then coupled with surface-enhanced Raman spectroscopy (SERS) to further amplify the Raman signal thanks to a combined FERS-SERS assay.

Results: A minimum 20-fold enhancement of the Raman signal of Aβ42 as compared to a conventional Raman spectroscopy scheme was observed using the HCPCF-based light delivery system. The signal was further boosted by decorating the fiber core with gold bipyramids generating an additional SERS effect, resulting in an overall 200 times amplification.

Conclusions: The results demonstrate that the use of an HCPCF-based platform can provide sharp and intense Raman signals of Aβ42, in turn paving the way toward the development of a sensitive label-free detection tool for early diagnosis of AD.
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http://dx.doi.org/10.1117/1.JBO.25.7.077001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7330420PMC
July 2020

Biosensor surface functionalization by a simple photochemical immobilization of antibodies: experimental characterization by mass spectrometry and surface enhanced Raman spectroscopy.

Analyst 2019 Nov;144(23):6871-6880

Dipartimento di Fisica, Politecnico di Milano, Piazza Leonardo da Vinci 32, 20133 - Milano, Italy.

Surface functionalization is a key step in biosensing since it is the basis of an effective analyte recognition. Among all the bioreceptors, antibodies (Abs) play a key role thanks to their superior specificity, although the available immobilization strategies suffer from several drawbacks. When gold is the interacting surface, the recently introduced Photochemical Immobilization Technique (PIT) has been shown to be a quick, easy-to-use and very effective method to tether Abs oriented upright by means of thiols produced via tryptophan mediated disulphide bridge reduction. Although the molecular mechanism of this process is quite well identified, the detailed morphology of the immobilized antibodies is still elusive due to inherent difficulties related to the microscopy imaging of Abs. The combination of Mass Spectrometry, Surface-Enhanced Raman Spectroscopy and Ellman's assay demonstrates that Abs irradiated under the conditions in which PIT is realized show only two effective disulphide bridges available for binding. They are located in the constant region of the immunoglobulin light chain so that the most likely position Ab assumes is side-on, i.e. with one Fab (i.e. the antigen binding portion of the antibody) exposed to the solution. This is not a limitation of the recognition efficiency in view of the intrinsic flexibility of the Ab structure, which makes the free Fab able to sway in the solution, a feature of great importance in many biosensing applications.
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http://dx.doi.org/10.1039/c9an00443bDOI Listing
November 2019

Black Phosphorus/Palladium Nanohybrid: Unraveling the Nature of P-Pd Interaction and Application in Selective Hydrogenation.

Chem Mater 2019 Jul 3;31(14):5075-5080. Epub 2019 Jul 3.

CNR-ICCOM, Via Madonna del Piano10, 50019 Sesto Fiorentino, Italy.

The burgeoning interest in two-dimensional (2D) black phosphorus (bP) contributes to the expansion of its applications in numerous fields. In the present study, 2D bP is used as a support for homogeneously dispersed palladium nanoparticles directly grown on it by a wet chemical process. Electron energy loss spectroscopy-scanning transmission electron microscopy analysis evidences a strong interaction between palladium and P atoms of the bP nanosheets. A quantitative evaluation of this interaction comes from the X-ray absorption spectroscopy measurements that show a very short Pd-P distance of 2.26 Å, proving for the first time the existence of an unprecedented Pd-P coordination bond of a covalent nature. Additionally, the average Pd-P coordination number of about 1.7 reveals that bP acts as a polydentate phosphine ligand toward the surface of the Pd atoms of the nanoparticles, thus preventing their agglomeration and inferring with structural stability. These unique properties result in a superior performance in the catalytic hydrogenation of chloronitroarenes to chloroanilines, where a higher chemoselectivity in comparison to other heterogeneous catalyst based on palladium has been observed.
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http://dx.doi.org/10.1021/acs.chemmater.9b00851DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6804426PMC
July 2019

Seeding variability of different alpha synuclein strains in synucleinopathies.

Ann Neurol 2019 05 27;85(5):691-703. Epub 2019 Mar 27.

Department of Neurology, University Medicine Goettingen and the German Center for Neurodegenerative Diseases (DZNE), Göttingen, Germany.

Objectives: Currently, the exact reasons why different α-synucleinopathies exhibit variable pathologies and phenotypes are still unknown. A potential explanation may be the existence of distinctive α-synuclein conformers or strains. Here, we intend to analyze the seeding activity of dementia with Lewy bodies (DLB) and Parkinson's disease (PD) brain-derived α-synuclein seeds by real-time quaking-induced conversion (RT-QuIC) and to investigate the structure and morphology of the α-synuclein aggregates generated by RT-QuIC.

Methods: A misfolded α-synuclein-enriched brain fraction from frontal cortex and substantia nigra pars compacta tissue, isolated by several filtration and centrifugation steps, was subjected to α-synuclein/RT-QuIC analysis. Our study included neuropathologically well-characterized cases with DLB, PD, and controls (Ctrl). Biochemical and morphological analyses of RT-QuIC products were conducted by western blot, dot blot analysis, Raman spectroscopy, atomic force microscopy, and transmission electron microscopy.

Results: Independently from the brain region, we observed different seeding kinetics of α-synuclein in the RT-QuIC in patients with DLB compared to PD and Ctrl. Biochemical characterization of the RT-QuIC product indicated the generation of a proteinase K-resistant and fibrillary α-synuclein species in DLB-seeded reactions, whereas PD and control seeds failed in the conversion of wild-type α-synuclein substrate.

Interpretation: Structural variances of α-synuclein seeding kinetics and products in DLB and PD indicated, for the first time, the existence of different α-synuclein strains in these groups. Therefore, our study contributes to a better understanding of the clinical heterogeneity among α-synucleinopathies, offers an opportunity for a specific diagnosis, and opens new avenues for the future development of strain-specific therapies. Ann Neurol 2019;85:691-703.
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http://dx.doi.org/10.1002/ana.25446DOI Listing
May 2019

Low cost tips for tip-enhanced Raman spectroscopy fabricated by two-step electrochemical etching of 125 µm diameter gold wires.

Beilstein J Nanotechnol 2018 22;9:2718-2729. Epub 2018 Oct 22.

CNR-IPCF, Istituto per i Processi Chimico-Fisici, Viale F. Stagno D'Alcontres 37, 98168 Messina, Italy.

Tip-enhanced Raman spectroscopy (TERS) has become a well-applied technique for nanospectroscopy, allowing for single molecule sensitivity with sub-nanometer spatial resolution. The demand for efficient, reproducible and cost-effective probes for TERS is increasing. Here we report on a new electrochemical etching protocol to fabricate TERS tips starting from 125 µm diameter gold wires in a reproducible way. The process is reliable (50% of the tips have radius of curvature <35 nm, 66% <80 nm), fast (less than 2 min) and 2.5 times cheaper than the etching of standard 250 µm diameter wires. The TERS performance of the tips is tested on dyes, pigments and biomolecules and enhancement factors higher than 10 are observed. TERS mapping with a spatial resolution of 5 nm is demonstrated.
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http://dx.doi.org/10.3762/bjnano.9.254DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6204785PMC
October 2018

Nanoscale Discrimination between Toxic and Nontoxic Protein Misfolded Oligomers with Tip-Enhanced Raman Spectroscopy.

Small 2018 09 9;14(36):e1800890. Epub 2018 Aug 9.

IFAC-CNR, Institute of Applied Physics "Nello Carrara,", National Research Council, Via Madonna del Piano 10, I-50019, Sesto Fiorentino, Italy.

Highly toxic protein misfolded oligomers associated with neurological disorders such as Alzheimer's and Parkinson's diseases are nowadays considered primarily responsible for promoting synaptic failure and neuronal death. Unraveling the relationship between structure and neurotoxicity of protein oligomers appears pivotal in understanding the causes of the pathological process, as well as in designing novel diagnostic and therapeutic strategies tuned toward the earliest and presymptomatic stages of the disease. Here, it is benefited from tip-enhanced Raman spectroscopy (TERS) as a surface-sensitive tool with spatial resolution on the nanoscale, to inspect the spatial organization and surface character of individual protein oligomers from two samples formed by the same polypeptide sequence and different toxicity levels. TERS provides direct assignment of specific amino acid residues that are exposed to a large extent on the surface of toxic species and buried in nontoxic oligomers. These residues, thanks to their outward disposition, might represent structural factors driving the pathogenic behavior exhibited by protein misfolded oligomers, including affecting cell membrane integrity and specific signaling pathways in neurodegenerative conditions.
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http://dx.doi.org/10.1002/smll.201800890DOI Listing
September 2018

Structural differences between toxic and nontoxic HypF-N oligomers.

Chem Commun (Camb) 2018 Aug 18;54(62):8637-8640. Epub 2018 Jul 18.

Department of Experimental and Clinical Biomedical Sciences, University of Florence, Viale Morgagni 50, 50134 Firenze, Italy.

We have studied two misfolded oligomeric forms of the protein HypF-N, which show similar morphologies but very different toxicities. We measured over 80 intermolecular distance-dependent parameters for each oligomer type using FRET, in conjunction with solution- and solid-state NMR and other biophysical techniques. The results indicate that the formation of a highly organised hydrogen bonded core in the toxic oligomers results in the exposure of a larger number of hydrophobic residues than in the nontoxic species, causing the former to form aberrant interactions with cellular components.
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http://dx.doi.org/10.1039/c8cc03446jDOI Listing
August 2018

Triggering molecular assembly at the mesoscale for advanced Raman detection of proteins in liquid.

Sci Rep 2018 01 18;8(1):1033. Epub 2018 Jan 18.

Institute of Applied Physics 'Nello Carrara', National Research Council (IFAC-CNR), via Madonna del Piano 10, Sesto Fiorentino, Italy.

An advanced optofluidic system for protein detection based on Raman signal amplification via dewetting and molecular gathering within temporary mesoscale assemblies is presented. The evaporation of a microliter volume of protein solution deposited in a circular microwell precisely follows an outward-receding geometry. Herein the combination of liquid withdrawal with intermolecular interactions induces the formation of self-assembled molecular domains at the solid-liquid interface. Through proper control of the evaporation rate, amplitude of the assemblies and time for spectral collection at the liquid edge are extensively raised, resulting in a local enhancement and refinement of the Raman response, respectively. Further signal amplification is obtained by taking advantage of the intense local electromagnetic fields generated upon adding a plasmonic coating to the microwell. Major advantages of this optofluidic method lie in the obtainment of high-quality, high-sensitivity Raman spectra with detection limit down to sub-micromolar values. Peculiarly, the assembled proteins in the liquid edge region maintain their native-like state without displaying spectral changes usually occurring when dried drop deposits are considered.
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http://dx.doi.org/10.1038/s41598-018-19558-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5773671PMC
January 2018

Computational Insight into the Interaction of Cytochrome C with Wet and PVP-Coated Ag Surfaces.

J Phys Chem B 2017 10 6;121(41):9532-9540. Epub 2017 Oct 6.

Department of Chemical and Geological Sciences, University of Modena and Reggio Emilia , Via G. Campi 103, 41125 Modena, Italy.

In this work, the adsorption of cytochrome C (CytC) on wet {100}, {111}, {110}, and {120} silver surfaces has been investigated by computational simulations. The effect of polyvinylpyrrolidone (PVP) coating has also been studied. The main results obtained can be summarized as follow: (a) CytC strongly interacts with wet bare high index facets, while the adsorption over the {100} surface is disfavored due to the strong water structuring at the surface; (b) a nonselective protein adsorption mechanism is highlighted; (c) the native structure of CytC is well preserved during adsorption; (d) the heme group of CytC is never found to interact directly with the surface; (e) the interactions with the PVP-capped {100} surface is weak and specific. These results can be exploited to better control biological responses at engineered nanosurface, allowing the development of improved diagnostic tools.
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http://dx.doi.org/10.1021/acs.jpcb.7b07492DOI Listing
October 2017

Site-Selective Surface-Enhanced Raman Detection of Proteins.

ACS Nano 2017 01 20;11(1):918-926. Epub 2016 Dec 20.

Institute of Applied Physics "Nello Carrara", National Research Council , via Madonna del Piano 10, 50019 Sesto Fiorentino, Italy.

Strategies for protein detection via surface-enhanced Raman spectroscopy (SERS) currently exploit the formation of randomly generated hot spots at the interfaces of metal colloidal nanoparticles, which are clustered together by intrusive chemical or physical processes in the presence of the target biomolecule. We propose a different approach based on selective and quantitative gathering of protein molecules at regular hot spots generated on the corners of individual silver nanocubes in aqueous medium at physiological pH. Here, the protein, while keeping its native configuration, experiences an intense local E-field, which boosts SERS efficiency and detection sensitivity. Uncontrolled signal fluctuations caused by variable molecular adsorption to different particle areas or inside clustered nanoparticles are circumvented. Advanced electron microscopy analyses and computational simulations outline a strategy relying on a site-selective mechanism with superior Raman signal enhancement, which offers the perspective of highly controlled and reproducible routine SERS detection of proteins.
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http://dx.doi.org/10.1021/acsnano.6b07523DOI Listing
January 2017

Preparation and Photoacoustic Analysis of Cellular Vehicles Containing Gold Nanorods.

J Vis Exp 2016 05 2(111). Epub 2016 May 2.

Institute of Applied Physics, Italian National Research Council.

Gold nanorods are attractive for a range of biomedical applications, such as the photothermal ablation and the photoacoustic imaging of cancer, thanks to their intense optical absorbance in the near-infrared window, low cytotoxicity and potential to home into tumors. However, their delivery to tumors still remains an issue. An innovative approach consists of the exploitation of the tropism of tumor-associated macrophages that may be loaded with gold nanorods in vitro. Here, we describe the preparation and the photoacoustic inspection of cellular vehicles containing gold nanorods. PEGylated gold nanorods are modified with quaternary ammonium compounds, in order to achieve a cationic profile. On contact with murine macrophages in ordinary Petri dishes, these particles are found to undergo massive uptake into endocytic vesicles. Then these cells are embedded in biopolymeric hydrogels, which are used to verify that the stability of photoacoustic conversion of the particles is retained in their inclusion into cellular vehicles. We are confident that these results may provide new inspiration for the development of novel strategies to deliver plasmonic particles to tumors.
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http://dx.doi.org/10.3791/53328DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4942024PMC
May 2016

Controlled graphene oxide assembly on silver nanocube monolayers for SERS detection: dependence on nanocube packing procedure.

Beilstein J Nanotechnol 2016 6;7:9-21. Epub 2016 Jan 6.

Department of Chemistry and CSGI, University of Florence, Via della Lastruccia 3-13, I-50019 Sesto Fiorentino, Italy.

Hybrid graphene oxide/silver nanocubes (GO/AgNCs) arrays for surface-enhanced Raman spectroscopy (SERS) applications were prepared by means of two procedures differing for the method used in the assembly of the silver nanocubes onto the surface: Langmuir-Blodgett (LB) transfer and direct sequential physisorption of silver nanocubes (AgNCs). Adsorption of graphene oxide (GO) flakes on the AgNC assemblies obtained with both procedures was monitored by quartz crystal microbalance (QCM) technique as a function of GO bulk concentration. The experiment provided values of the adsorbed GO mass on the AgNC array and the GO saturation limit as well as the thickness and the viscoelastic properties of the GO film. Atomic force microscopy (AFM) measurements of the resulting samples revealed that a similar surface coverage was achieved with both procedures but with a different distribution of silver nanoparticles. In the GO covered LB film, the AgNC distribution is characterized by densely packed regions alternating with empty surface areas. On the other hand, AgNCs are more homogeneously dispersed over the entire sensor surface when the nanocubes spontaneously adsorb from solution. In this case, the assembly results in less-packed silver nanostructures with higher inter-cube distance. For the two assembled substrates, AFM of silver nanocubes layers fully covered with GO revealed the presence of a homogeneous, flexible and smooth GO sheet folding over the silver nanocubes and extending onto the bare surface. Preliminary SERS experiments on adenine showed a higher SERS enhancement factor for GO on Langmuir-Blodgett films of AgNCs with respect to bare AgNC systems. Conversely, poor SERS enhancement for adenine resulted for GO-covered AgNCs obtained by spontaneous adsorption. This indicated that the assembly and packing of AgNCs obtained in this way, although more homogeneous over the substrate surface, is not as effective for SERS analysis.
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http://dx.doi.org/10.3762/bjnano.7.2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4734306PMC
February 2016

One-pot synthesis of magnesium nanoparticles embedded in a chitosan microparticle matrix: a highly biocompatible tool for in vivo cancer treatment.

J Mater Chem B 2016 Jan 15;4(2):207-211. Epub 2015 Dec 15.

Division of Surgical Oncology, Department of Surgery, University of Louisville, Louisville, KY 40202, USA.

A novel highly biocompatible nanosystem made up of a chitosan matrix and filled with magnesium nanoparticles was synthesized using a simple and one-pot strategy, and tested as a promising, well-tolerated tool for photothermal therapy. Moreover, in vivo a proof of concept on hepatocarcinoma-bearing mice is presented.
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http://dx.doi.org/10.1039/c5tb02499dDOI Listing
January 2016

Controlled Veiling of Silver Nanocubes with Graphene Oxide for Improved Surface-Enhanced Raman Scattering Detection.

ACS Appl Mater Interfaces 2016 Feb 21;8(4):2628-34. Epub 2016 Jan 21.

Institute of Applied Physics , National Research Council, via Madonna del Piano 10, I-50019 Sesto Fiorentino, Italy.

Hybrid graphene oxide (GO)/metal nanocomposites have been recently proposed as novel surface-enhanced Raman scattering (SERS) substrates. Despite an increasing interest in these systems, standardization in their fabrication process is still lacking but urgently required to support their use for real-life applications. In this work we investigate how the assembly of GO should be conducted to control adsorption geometry and optical properties at the interface with plasmonic nanostructures as monolayer assemblies of silver nanocubes, by tuning main experimental parameters including GO concentration and self-assembly time. We finally identified the experimental conditions for building up a close-fitting soft dressing of the plasmonic surface, which shows optimal characteristics for flexible and reliable SERS detection.
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http://dx.doi.org/10.1021/acsami.5b10438DOI Listing
February 2016

SERS Detection of Amyloid Oligomers on Metallorganic-Decorated Plasmonic Beads.

ACS Appl Mater Interfaces 2015 May 1;7(18):9420-8. Epub 2015 May 1.

†Universitat Rovira i Virgili and Centro de Tecnologia Quimica de Cataluña, C/de Marcel·lí Domingo s/n, N5, 43007 Tarragona, Spain.

Protein misfolded proteins are among the most toxic endogenous species of macromolecules. These chemical entities are responsible for neurodegenerative disorders such as Alzheimer's, Parkinson's, Creutzfeldt-Jakob's and different non-neurophatic amyloidosis. Notably, these oligomers show a combination of marked heterogeneity and low abundance in body fluids, which have prevented a reliable detection by immunological methods so far. Herein we exploit the selectivity of proteins to react with metallic ions and the sensitivity of surface-enhanced Raman spectroscopy (SERS) toward small electronic changes in coordination compounds to design and engineer a reliable optical sensor for protein misfolded oligomers. Our strategy relies on the functionalization of Au nanoparticle-decorated polystyrene beads with an effective metallorganic Raman chemoreceptor, composed by Al(3+) ions coordinated to 4-mercaptobenzoic acid (MBA) with high Raman cross-section, that selectively binds aberrant protein oligomers. The mechanical deformations of the MBA phenyl ring upon complexation with the oligomeric species are registered in its SERS spectrum and can be quantitatively correlated with the concentration of the target biomolecule. The SERS platform used here appears promising for future implementation of diagnostic tools of aberrant species associated with protein deposition diseases, including those with a strong social and economic impact, such as Alzheimer's and Parkinson's diseases.
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http://dx.doi.org/10.1021/acsami.5b01056DOI Listing
May 2015

Concave gold nanocube assemblies as nanotraps for surface-enhanced Raman scattering-based detection of proteins.

Nanoscale 2015 Feb;7(8):3474-80

Institute of Applied Physics "Nello Carrara", National Research Council, via Madonna del Piano 10, I-50019 Sesto Fiorentino, Italy.

SERS detection of proteins is typically performed by using labeling agents with stable and high Raman scattering cross sections. This is a valuable approach for trace detection and quantification of a target protein but is unsuitable for inspecting its inherent structural and functional properties. On the other hand, direct SERS of proteins has been mainly devoted to the study of short peptides and aminoacid sequences or of prosthetic groups with intense Raman signals, which is of scarce interest for a thorough characterization of most proteins. Here we try to overcome these limitations by setting-up an effective platform for the structural SERS analysis of proteins. The platform consists of an extended bidimensional array of gold concave nanocubes (CNCs) supported on a PDMS film. CNCs are closely-packed through face-face and face-corner interactions generating a monolayered arrangement featuring well distributed nanoholes. Here the protein homogeneously experiences an E-field enhancement outward from the metal surfaces surrounding it, which causes a large number of vibrations to be contemporarily amplified. The proposed platform provides stable and detailed SERS spectra and confers rapidity and reproducibility to the analysis.
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http://dx.doi.org/10.1039/c4nr05704jDOI Listing
February 2015

Graphene as a photothermal switch for controlled drug release.

Nanoscale 2014 Jul;6(14):7947-53

Institute of Applied Physics "Nello Carrara", National Research Council, via Madonna del Piano 10, I-50019 Sesto Fiorentino, Italy.

Graphene has recently emerged as a novel material in the biomedical field owing to its optical properties, biocompatibility, large specific surface area and low cost. In this paper, we provide the first demonstration of the possibility of using light to remotely trigger the release of drugs from graphene in a highly controlled manner. Different drugs including chemotherapeutics and proteins are firmly adsorbed onto reduced graphene oxide (rGO) nanosheets dispersed in a biopolymer film and then released by individual millisecond-long light pulses generated by a near infrared (NIR) laser. Here graphene plays the dual role of a versatile substrate for temporary storage of drugs and an effective transducer of NIR-light into heat. Drug release appears to be narrowly confined within the size of the laser spot under noninvasive conditions and can be precisely dosed depending on the number of pulses. The approach proposed paves the way for tailor-made pharmacological treatments of chronic diseases, including cancer, anaemia and diabetes.
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http://dx.doi.org/10.1039/c4nr01622jDOI Listing
July 2014

Fluorescence approach for measuring anthocyanins and derived pigments in red wine.

J Agric Food Chem 2013 Oct 9;61(42):10156-62. Epub 2013 Oct 9.

Institute of Applied Physics, National Research Council, Sesto Fiorentino (Florence), Italy.

A novel fluorescence approach to monitor the evolution of anthocyanins and derivatives in red wine was developed. Some red table wines and Port wine with different vintage years were first tested with the aim to determine the ideal fluorescent conditions. The fluorescence contribution of both monomeric and polymeric anthocyanins was studied by comparing their emission spectra. By measuring the F700/F560 ratio on different wine samples and applying an inverted exponential function, it was possible to estimate the monomeric/polymeric anthocyanin absorbance ratio that is proportional to the relative content of the two classes of compounds. The methodology was further developed by using pure compounds representative of monomeric anthocyanins and anthocyanin-pyruvic acid adducts, namely, by using malvidin-3-O-glucoside and vitisin A. A fluorescence excitation ratio (FER350/550) was considered for estimating the absorbance ratio between vitisin A and malvidin-3-O-glucoside. Overall, this work aims to use fluorescence to monitor the evolution of anthocyanin derivatives and to distinguish them from their anthocyanin precursors, thereby allowing the evolution of anthocyanin pigments during wine aging to be monitored, but it also may be useful to determine age markers or even geographical markers.
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http://dx.doi.org/10.1021/jf402398aDOI Listing
October 2013

Quantitative readout of optically encoded gold nanorods using an ordinary dark-field microscope.

Nanoscale 2013 Oct;5(20):9645-50

INO-CNR, National Institute of Optics-CNR, Largo E. Fermi 6, Florence, Italy.

In this paper we report on a new use for dark-field microscopy in order to retrieve two-dimensional maps of optical parameters of a thin sample such as a cryptograph, a histological section, or a cell monolayer. In particular, we discuss the construction of quantitative charts of light absorbance and scattering coefficients of a polyvinyl alcohol film that was embedded with gold nanorods and then etched using a focused mode-locked Ti:Sapphire oscillator. Individual pulses from this laser excite plasmonic oscillations of the gold nanorods, thus triggering plastic deformations of the particles and their environment, which are confined within a few hundred nm of the light focus. In turn, these deformations modify the light absorbance and scattering landscape, which can be measured with optical resolution in a dark-field microscope equipped with an objective of tuneable numerical aperture. This technique may prove to be valuable for various applications, such as the fast readout of optically encoded data or to model functional interactions between light and biological tissue at the level of cellular organelles, including the photothermolysis of cancer.
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http://dx.doi.org/10.1039/c3nr00726jDOI Listing
October 2013

In vivo laser assisted microvascular repair and end-to-end anastomosis by means of indocyanine green-infused chitosan patches: a pilot study.

Lasers Surg Med 2013 Jul 5;45(5):318-25. Epub 2013 Jun 5.

Institute of Neurosurgery, Catholic University School of Medicine, Rome, 00100, Italy.

Background And Objectives: Laser-based repairing techniques offer several advantages respect to standard suturing in microsurgery. In this work we evaluate the applicability and feasibility of two innovative laser-based approaches for microvascular repair and anastomoses: (1) laser-assisted vascular repair (LAVR); (2) laser-assisted end-to-end vascular anastomosis (LAVA). All these procedures have been executed by the use of diode laser irradiation and chitosan-patches infused with Indocyanine Green (ICG).

Study Design/materials And Methods: Experiments were performed on 30 rabbits. Twenty animals underwent LAVR and 10 end-to-end LAVA procedures. In the LAVR group, a 5-mm longitudinal cut was performed on the common carotid artery (CCA), then an ICG-infused chitosan patch was topically applied and laser-soldered over the arterial lesion. In the LAVA group the end-to-end anastomosis was executed on CCA by means of application of the three interrupted sutures and subsequent laser soldering of the ICG-infused patch. Animals underwent different follow-up periods (2, 7, 30, and 90 days). At the end of every follow-up, the animals were re-anesthetized and a microdoppler analysis was performed in order to check patency of the treated vessels. Then soldered segments were excised and subjected to histological and ultrastructural evaluations.

Results: At the end of surgery no bleeding from the treated segment was observed; all the treated vessels were patent. At the end of follow-up periods, no signs of perivascular haemorrhage were found. An intraoperative microdoppler evaluation assessed the patency of all the treated vessels. Histology showed a good reorganization of the vascular wall structures and an early endothelial regeneration was observed by SEM.

Conclusions: Our study demonstrated the efficacy of laser tissue soldering by means of ICG-infused chitosan patches for the in vivo repairing of microvascular lesions and end-to-end anastomoses. This approach offers several advantages over conventional suturing methods and is technically easy to perform, minimizing the surgical trauma to vessels.
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http://dx.doi.org/10.1002/lsm.22145DOI Listing
July 2013

Photothermally activated hybrid films for quantitative confined release of chemical species.

Angew Chem Int Ed Engl 2013 Jun 9;52(23):5956-60. Epub 2013 May 9.

Institute of Applied Physics Nello Carrara, National Research Council via Madonna del Piano 10, 50019, Italy.

Illuminating films of a porous chitosan matrix containing gold nanorods and thermosensitive micelles loaded with a chemical stimulates local photothermal conversion of the gold nanorods. The heat produced activates the ejection of the chemical from the micelles (see scheme), and causes the transient permeabilization of adjacent cell membranes, resulting in a selective cellular uptake of the released chemical with control over spatiotemporal parameters and dosage.
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http://dx.doi.org/10.1002/anie.201207986DOI Listing
June 2013

Light-responsive nanocomposite sponges for on demand chemical release with high spatial and dosage control.

J Mater Chem B 2013 Feb 7;1(8):1096-1100. Epub 2013 Jan 7.

Institute of Applied Physics "Nello Carrara", National Research Council, via Madonna del Piano 10, 50019, Sesto Fiorentino, Italy.

We present a biocompatible device for on demand chemical release in the form of a light-activated sponge-like nanocomposite scaffold, which ensures excellent control over the principal parameters of chemical release and dosage in order to sustain effective therapeutic action. The sponge consists of a porous biopolymer scaffold containing a dispersion of gold nanorods, which acts as an absorber of the incoming laser light, and of thermosensitive micelles, which serve as a reservoir for the drug molecules to be released. The photothermal response of the nanoparticles contained inside the sponge triggers a contraction in proximal micelles, thus promoting the expulsion of the drug that in turn is released from the sponge to the external environment. The peculiar physiochemical and structural properties of the nanocomposite sponges impart a number of interesting features to the proposed drug release system, including the possibility of spatially confining the therapeutic treatment as well as precise control of the amount of released drug as a function of duration and power of the excitation light.
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http://dx.doi.org/10.1039/c2tb00310dDOI Listing
February 2013

Thermal transitions of fibrillar collagen unveiled by second-harmonic generation microscopy of corneal stroma.

Biophys J 2012 Sep;103(6):1179-87

Institute of Applied Physics "Nello Carrara", National Research Council, Sesto Fiorentino, Italy.

The thermal transitions of fibrillar collagen are investigated with second-harmonic generation polarization anisotropy microscopy. Second-harmonic generation images and polarization anisotropy profiles of corneal stroma heated in the 35-80°C range are analyzed by means of a theoretical model that is suitable to probe principal intramolecular and interfibrillar parameters of immediate physiological interest. Our results depict the tissue modification with temperature as the interplay of three destructuration stages at different hierarchical levels of collagen assembly including its tertiary structure and interfibrillar alignment, thus supporting and extending previous findings. This method holds the promise of a quantitative inspection of fundamental biophysical and biochemical processes and may find future applications in real-time and postsurgical functional imaging of collagen-rich tissues subjected to thermal treatments.
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http://dx.doi.org/10.1016/j.bpj.2012.07.055DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3446693PMC
September 2012

Hybrid nanocomposite films for laser-activated tissue bonding.

J Biophotonics 2012 Nov 17;5(11-12):868-77. Epub 2012 Aug 17.

Istituto di Fisica Applicata Nello Carrara, Consiglio Nazionale delle Ricerche, Sesto Fiorentino, Italy.

We report new advancements in the biomedical exploitation of plasmonic nanoparticles as an effective platform for the photothermal repair of biological tissue. Chitosan films are loaded with gold nanorods with intense optical absorption in the "therapeutic window" of deepest light penetration through the body, and then activated by near infrared laser excitation to give adhesion with adjacent connective tissues. The adhesion consists of 0.07 mm(2) welds of ~20 kPa tensile strength at the film/tissue interface, which are obtained by administration of pulses with duration in the hundreds of millisecond timescale from a diode laser at ~130 J cm(-2). We investigate the adhesive effect as a function of pulse power and duration and identify an optimal operative window to achieve effective and reproducible welds with minimal detrimental superheating. These results may prove valuable to standardize laser bonding techniques and meet current needs for new knowledge which is urged by the penetration of nanotechnology into biomedical optics.
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http://dx.doi.org/10.1002/jbio.201200115DOI Listing
November 2012