Learning models for classifying Raman spectra of genomic DNA from tumor subtypes.

An early and accurate detection of different subtypes of tumors is crucial for an effective guidance to personalized therapy and in predicting the ability of tumor to metastasize. Here we exploit the Surface Enhanced Raman Scattering (SERS) platform, based on disordered silver coated silicon nanowires (Ag/SiNWs), to efficiently discriminate genomic DNA of different subtypes of melanoma and colon tumors. The diagnostic information is obtained by performing label free Raman maps of the dried drops of DNA solutions onto the Ag/NWs mat and leveraging the classification ability of learning models to reveal the specific and distinct physico-chemical interaction of tumor DNA molecules with the Ag/NW, here supposed to be partly caused by a different DNA methylation degree.

Revealing Low Amplitude Signals of Neuroendocrine Cells through Disordered Silicon Nanowires-Based Microelectrode Array.

Today, the key methodology to study in vitro or in vivo electrical activity in a population of electrogenic cells, under physiological or pathological conditions, is by using microelectrode array (MEA). While significant efforts have been devoted to develop nanostructured MEAs for improving the electrophysiological investigation in neurons and cardiomyocytes, data on the recording of the electrical activity from neuroendocrine cells with MEA technology are scarce owing to their weaker electrical signals. Disordered silicon nanowires (SiNWs) for developing a MEA that, combined with a customized acquisition board, successfully capture the electrical signals generated by the corticotrope AtT-20 cells as a function of the extracellular calcium (Ca2+ ) concentration are reported. The recorded signals show a shape that clearly resembles the action potential waveform by suggesting a natural membrane penetration of the SiNWs. Additionally, the generation of synchronous signals observed under high Ca2+ content indicates the occurrence of a collective behavior in the AtT-20 cell population. This study extends the usefulness of MEA technology to the investigation of the electrical communication in cells of the pituitary gland, crucial in controlling several essential human functions, and provides new perspectives in recording with MEA the electrical activity of excitable cells.

Tailoring the Microarchitectures of 3D Printed Bone-like Scaffolds for Tissue Engineering Applications.

Material extrusion (MEX), commonly referred to as fused deposition modeling (FDM) or fused filament fabrication (FFF), is a versatile and cost-effective technique to fabricate suitable scaffolds for tissue engineering. Driven by a computer-aided design input, specific patterns can be easily collected in an extremely reproducible and repeatable process. Referring to possible skeletal affections, 3D-printed scaffolds can support tissue regeneration of large bone defects with complex geometries, an open major clinical challenge. In this study, polylactic acid scaffolds were printed resembling trabecular bone microarchitecture in order to deal with morphologically biomimetic features to potentially enhance the biological outcome. Three models with different pore sizes (i.e., 500, 600, and 700 µm) were prepared and evaluated by means of micro-computed tomography. The biological assessment was carried out seeding SAOS-2 cells, a bone-like cell model, on the scaffolds, which showed excellent biocompatibility, bioactivity, and osteoinductivity. The model with larger pores, characterized by improved osteoconductive properties and protein adsorption rate, was further investigated as a potential platform for bone-tissue engineering, evaluating the paracrine activity of human mesenchymal stem cells. The reported findings demonstrate that the designed microarchitecture, better mimicking the natural bone extracellular matrix, favors a greater bioactivity and can be thus regarded as an interesting option for bone-tissue engineering.

Editorial for the Special Issue on Nanostructured Surfaces and Devices for Biomedical Applications.

The ability to control and modify the surface topography of materials at the nanoscale, which produces features with a comparable size to that of biological entities, so as to effectively probe and influence processes at both the cellular and the molecular level, has facilitated incredible possibilities in the fields of biomedicine, biosensing, and diagnostics [...].

Statistical Classification for Raman Spectra of Tumoral Genomic DNA.

We exploit Surface-Enhanced Raman Scattering (SERS) to investigate aqueous droplets of genomic DNA deposited onto silver-coated silicon nanowires, and we show that it is possible to efficiently discriminate between spectra of tumoral and healthy cells. To assess the robustness of the proposed technique, we develop two different statistical approaches, one based on the Principal Components Analysis of spectral data and one based on the computation of the ℓ2 distance between spectra. Both methods prove to be highly efficient, and we test their accuracy via the Cohen's κ statistics. We show that the synergistic combination of the SERS spectroscopy and the statistical analysis methods leads to efficient and fast cancer diagnostic applications allowing rapid and unexpansive discrimination between healthy and tumoral genomic DNA alternative to the more complex and expensive DNA sequencing.

Label-Free Morpho-Molecular Imaging for Studying the Differential Interaction of Black Phosphorus with Tumor Cells.

Black phosphorus nanosheets (2D BP) are emerging as very promising, highly selective chemotherapeutic agents due to their fast degradation in the intracellular matrix of cancer cells. Here, optical diffraction tomography (ODT) and Raman spectroscopy were exploited as a powerful label-free approach to achieve integrated insights into the processes accompanying the administration of exfoliated 2D BP flakes in human prostatic adenocarcinoma and normal human prostate epithelial cells. Our ODT experiments provided unambiguous visualization of the 2D BP internalization in cancer cells and the morphological modifications of those cells in the apoptotic phase. The cellular internalization and damaging occurred, respectively, 18 h and 36-48 h after the 2D BP administration. Changes in the chemical properties of the internalized 2D BP flakes were monitored by Raman spectroscopy. Interestingly, a fast oxidation process of the 2D BP flakes was activated in the intracellular matrix of the cancer cells after 24 h of incubation. This was in sharp contrast to the low 2D BP uptake and minimal chemical changes observed in the normal cells. Along with the understanding of the 2D BP fate in the cancer cells, the proposed label-free morpho-molecular approach offers a powerful, rapid tool to study the pharmacokinetic properties of engineered nanomaterials in preclinical research.

Biological Response to Bioinspired Microporous 3D-Printed Scaffolds for Bone Tissue Engineering.

The scaffold is a key element in the field of tissue engineering, especially when large defects or substitutions of pathological tissues or organs need to be clinically addressed. The expected outcome is strongly dependent on the cell-scaffold interaction and the integration with the surrounding biological tissue. Indeed, mimicking the natural extracellular matrix (ECM) of the tissue to be healed represents a further optimization that can limit a possible morphological mismatch between the scaffold and the tissue itself. For this aim, and referring to bone tissue engineering, polylactic acid (PLA) scaffolds were 3D printed with a microstructure inspired by the trabecular architecture and biologically evaluated by means of human osteosarcoma SAOS-2 cells. The cells were seeded on two types of scaffolds differing for the designed pore size (i.e., 400 and 600 µm), showing the same growth exponential trend found in the control and no significant alterations in the actin distribution. The microporous structure of the two tested samples enhanced the protein adsorption capability and mRNA expression of markers related to protein synthesis, proliferation, and osteoblast differentiation. Our findings demonstrate that 3D-printed scaffolds support the adhesion, growth, and differentiation of osteoblast-like cells and the microporous architecture, mimicking the natural bone hierarchical structure, and favoring greater bioactivity. These bioinspired scaffolds represent an interesting new tool for bone tissue engineering and regenerative medicine applications.

High-Resolution 3D Fabrication of Glass Fiber-Reinforced Polymer Nanocomposite (FRPN) Objects by Two-Photon Direct Laser Writing.

This paper reports on the nanofabrication of a fiber-reinforced polymer nanocomposite (FRPN) by two-photon direct laser writing (TP-DLW) using silica nanowires (SiO2 NWs) as nanofillers, since they feature a refractive index very close to that of the photoresist used as a polymeric matrix. This allows for the best resolution offered by the TP-DLW technique, even with high loads of SiO2 NWs, up to 70 wt %. The FRPN presented an increase of approximately 4 times in Young's modulus (8.23 GPa) and nanohardness (120 MPa) when compared to those of the bare photoresist, indicating how the proposed technique is well-suited for applications with higher structural requirements. Moreover, three different printing configurations can be implemented thanks to the use of silicon chips, on which the SiO2 NWs are grown, as fabrication substrates. First, they can be effectively used as an adhesive layer when the laser beam is focused at the interface with the silicon substrate. Second, they can be used as a sacrificial layer, when the laser beam is focused in a plane inside the SiO2 NW layer. Third, only the outer shell of the object is printed so that the SiO2 NW tangle acts as the internal skeleton for the structure being fabricated in the so-called shell and scaffold printing strategy.

Nanowire Assisted Mechanotyping of Cellular Metastatic Potential.

Nanotechnology has provided tools for next generation biomedical devices which rely on nanostructure interfaces with living cells. In vitro biomimetic structures have enabled observation of cell response to various mechanical and chemical cues, and there is a growing interest in isolating and harnessing the specific cues that three-dimensional microenvironments can provide without the requirement for such culture and the experimental drawbacks associated with it. Here we report a randomly oriented gold coated Si nanowire substrate with patterned hydrophobic-hydrophilic areas for differentiation of isogenic breast cancer cells of varying metastatic potential. When considering synthetic surfaces for the study of cell-nanotopography interfaces, randomly oriented nanowires more closely resemble the isotropic architecture of natural extracellular matrix as compared to currently more widely used vertical nanowire arrays. In the study reported here, we show that primary cancer cells preferably attach to the hydrophilic region of randomly oriented nanowire substrate while secondary cancer cells do not adhere. Using machine learning analysis of fluorescence images, cells were found to spread and elongate on the nanowire substrates as compared to a flat substrate, where they mostly remain round, when neither surface was coated with extracellular matrix (ECM) proteins. Such platforms can not only be used for developing bioassays but also as stepping stones for tissue printing technologies where cells can be selectively patterned at desired locations.

SEM characterization and ageing analysis on two generation of invisible aligners.

BACKGROUND: The purpose of the in vitro study is to investigate and compare the morphological features and the chemical stability in weight of two different polyurethane-based blends, Smart Track (LD30) and Exceed30 (EX30), used for orthodontic aligners manufacture before and after the oral usage. METHODS: Twenty orthodontic aligners were randomly selected: 10 LD30 and 10 EX30, each group was divided in two subgroups, never used and intra-orally aged. By the employment of a Stereomicroscope, a section of 5 × 5 mm was cut from the buccal surface of the incisal region of each aligner. All samples were subjected to Scanning Electron Microscopy and Ageing tests in different solutions to simulate the hostility of the oral environment. The statistical method used was t-test. RESULTS: At SEM images, LD30 appears more homogeneous in texture respect to EX30. However, after clinical usage, both materials show significant structural alterations: findings have been supported by higher magnifications at SEM, by which it is clearly to observe many superficial cracks cross through the polymer structures of LD30U, absent in never used samples. LD30U surface becomes also smoother due to the disappearance of most of the conglomerates, but at the same time also rougher while EX30U shows a greater irregularity and porosity in which large and deep cracks are also highlighted. Although these changes occur persistently, in the aging tests no significant weight loss from both materials has been found, confirming the initial hypothesis of a good chemical stability and safety of both polyurethane mixtures even in conditions of severe hostility. CONCLUSION: LD30 is the expression of the technological evolution of EX30, this is made evident above all by its morphological architecture, more homogeneous and defined but also by the chemical stability that can be appreciated even in evident critic situations.

Raman Mapping of Biological Systems Interacting with a Disordered Nanostructured Surface: A Simple and Powerful Approach to the Label-Free Analysis of Single DNA Bases.

This article demonstrates the possibility to use a novel powerful approach based on Raman mapping of analyte solutions drop casted on a disordered array of Ag covered silicon nanowires (Ag/SiNWs), to identify the characteristic spectral signal of the four DNA bases, adenine (A), thymine (T), cytosine (C), and guanine (G), at concentration as low as 10 ng/µL, and to study their specific way of interacting with the nanostructured substrate. The results show a distinctive and amplified interaction of guanine, the base that is most susceptible to oxidation, with the nanostructured surface. Our findings explain the recently revealed diverse behaviour of cancer and normal DNA deposited on the same Ag/SiNWs, which is ascribed to mechanical deformation and base lesions present on the oxidised DNA molecule backbone and causes detectable variation in the Raman signal, usable for diagnostic purposes. The notable bio-analytical capability of the presented platform, and its sensitivity to the molecule mechanical conformation at the single-base level, thus provides a new reliable, rapid, label-free DNA diagnostic methodology alternative to more sophisticated and expensive sequencing ones.

Silver-coated silicon nanowire platform discriminates genomic DNA from normal and malignant human epithelial cells using label-free Raman spectroscopy.

Genomic deoxyribonucleic acid (DNA) stores and carries the information required to maintain and replicate cellular life. While much efforts have been devoted in decoding the sequence of DNA basis to detect the genetic mutations related to cancer disease, it is becoming clear that physical properties, like structural conformation, stiffness and shape, can play an important role to recognize DNA modifications. Here, silver-coated silicon nanowires (Ag/SiNWs) are exploited as Raman spectroscopic platform to easily discriminate healthy and cancer genomic DNA, extracted from human normal skin and malignant melanoma cells, respectively. In particular, aqueous DNA droplets are directly deposited onto a forest of Ag/SiNWs and Raman maps are acquired after sample dehydration. By applying principal component analysis (PCA) to the Raman spectra collected within the droplets, healthy and cancer cell DNA can be distinguished without false negative identifications and with few false positive results (< 2%). The discrimination occurs regardless the analysis of specific DNA sequencing, but through Raman bands strictly related to the interfacing of the DNA and the NWs. The observed phenomenon can be ascribed to conformational differences and/or diverse charge properties between healthy and cancer cell DNA determining a different arrangement of the molecules adsorbed onto the NWs upon water evaporation. The unique interaction with DNA and facile fabrication technology make Ag/SiNWs an effective platform for a robust, rapid and label-free cancer diagnosis, as well as a potential tool to investigate physical properties of DNA.

Glial Interfaces: Advanced Materials and Devices to Uncover the Role of Astroglial Cells in Brain Function and Dysfunction.

Research over the past four decades has highlighted the importance of certain brain cells, called glial cells, and has moved the neurocentric vision of structure, function, and pathology of the nervous system toward a more holistic perspective. In this view, the demand for technologies that are able to target and both selectively monitor and control glial cells is emerging as a challenge across neuroscience, engineering, chemistry, and material science. Frequently neglected or marginally considered as a barrier to be overcome between neural implants and neuronal targets, glial cells, and in particular astrocytes, are increasingly considered as active players in determining the outcomes of device implantation. This review provides a concise overview not only of the previously established but also of the emerging physiological and pathological roles of astrocytes. It also critically discusses the most recent advances in biomaterial interfaces and devices that interact with glial cells and thus have enabled scientists to reach unprecedented insights into the role of astroglial cells in brain function and dysfunction. This work proposes glial interfaces and glial engineering as multidisciplinary fields that have the potential to enable significant advancement of knowledge surrounding cognitive function and acute and chronic neuropathologies.

Silver-Coated Disordered Silicon Nanowires Provide Highly Sensitive Label-Free Glycated Albumin Detection through Molecular Trapping and Plasmonic Hotspot Formation.

Glycated albumin (GA) is rapidly emerging as a robust biomarker for screening and monitoring of diabetes. To facilitate its rapid, point-of-care measurements, a label-free surface-enhanced Raman spectroscopy (SERS) sensing platform is reported that leverages the specificity of molecular vibrations and signal amplification on silver-coated silicon nanowires (Ag/SiNWs) for highly sensitive and reproducible quantification of GA. The simulations and experimental measurements demonstrate that the disordered orientation of the nanowires coupled with the wicking of the analyte molecules during the process of solvent evaporation facilitates molecular trapping at the generated plasmonic hotspots. Highly sensitive detection of glycated albumin is shown with the ability to visually detect spectral features at as low as 500 × 10-9 m, significantly below the physiological range of GA in body fluids. Combined with chemometric regression models, the spectral data recorded on the Ag/SiNWs also allow accurate prediction of glycated concentration in mixtures of glycated and non-glycated albumin in proportions that reflect those in the bloodstream.

Cytotoxicity and internalization analysis of silicon nanowires in Buffalo Green Monkey cells: a preliminary study to evaluate the possibility of carrying viruses inside the cells.

Silicon nanowires (SiNWs) are attractive functional nanomaterials for biomedical applications. The ability to easily tune their size and density, potential biocompatibility, and knowledge of the chemical activation of SiNWs surface make them natural tools to interact with biological materials. We evaluated the possibility of exploiting SiNWs as carriers to introduce organic compounds into cells. The cellular toxicity and the internalization capacity of free-standing and label-free SiNWs were tested on Buffalo Green Monkey cells (BGM). Confocal fluorescent observation of SiNWs conjugated with fluorescein-polyethylene imine (PEI) confirmed the internalization of the NWs into the Buffalo Green Monkey Cells (BGM).

A Glial-Silicon Nanowire Electrode Junction Enabling Differentiation and Noninvasive Recording of Slow Oscillations from Primary Astrocytes.

The correct human brain function is dependent on the activity of non-neuronal cells called astrocytes. The bioelectrical properties of astrocytes in vitro do not closely resemble those displayed in vivo and the former are incapable of generating action potential; thus, reliable approaches in vitro for noninvasive electrophysiological recording of astrocytes remain challenging for biomedical engineering. Here it is found that primary astrocytes grown on a device formed by a forest of randomly oriented gold coated-silicon nanowires, resembling the complex structural and functional phenotype expressed by astrocytes in vivo. The device enables noninvasive extracellular recording of the slow-frequency oscillations generated by differentiated astrocytes, while flat electrodes failed on recording signals from undifferentiated cells. Pathophysiological concentrations of extracellular potassium, occurring during epilepsy and spreading depression, modulate the power of slow oscillations generated by astrocytes. A reliable approach to study the role of astrocytes function in brain physiology and pathologies is presented.

Array of disordered silicon nanowires coated by a gold film for combined NIR photothermal treatment of cancer cells and Raman monitoring of the process evolution.

Photothermal therapy (PTT) assisted by nanomaterials is a promising minimally invasive technique for cancer treatment. Here, we explore the PTT properties of a silicon- and gold-based nanostructured platform suitable for being directly integrated in fibre laser systems rather than injected into the human body, which occurs for the most commonly unreported PTT nanoagents. In particular, the photothermal properties of an array of disordered silicon nanowires coated by a thin gold film (Au/SiNWs) were tested on a monolayer of human colon adenocarcinoma cells (Caco-2) irradiated with a 785 nm laser. Au/SiNWs allowed an efficient photothermal action and simultaneous monitoring of the process evolution through the Raman signal coming from the irradiated cellular zone. Strong near infra-red (NIR) absorption, overlapping three biological windows, cell-friendly properties and effective fabrication technology make Au/SiNWs suitable both to be integrated in surgical laser tools and as an in vitro platform to develop novel PTT protocols using different cancer types and NIR sources.

Disordered array of Au covered Silicon nanowires for SERS biosensing combined with electrochemical detection.

We report on highly disordered array of Au coated silicon nanowires (Au/SiNWs) as surface enhanced Raman scattering (SERS) probe combined with electrochemical detection for biosensing applications. SiNWs, few microns long, were grown by plasma enhanced chemical vapor deposition on common microscope slides and covered by Au evaporated film, 150 nm thick. The capability of the resulting composite structure to act as SERS biosensor was studied via the biotin-avidin interaction: the Raman signal obtained from this structure allowed to follow each surface modification step as well as to detect efficiently avidin molecules over a broad range of concentrations from micromolar down to the nanomolar values. The metallic coverage wrapping SiNWs was exploited also to obtain a dual detection of the same bioanalyte by electrochemical impedance spectroscopy (EIS). Indeed, the SERS signal and impedance modifications induced by the biomolecule perturbations on the metalized surface of the NWs were monitored on the very same three-electrode device with the Au/SiNWs acting as both working electrode and SERS probe.

Highly Disordered Array of Silicon Nanowires: an Effective and Scalable Approach for Performing and Flexible Electrochemical Biosensors.

The direct integration of disordered arranged and randomly oriented silicon nanowires (SiNWs) into ultraflexible and transferable electronic circuits for electrochemical biosensing applications is proposed. The working electrode (WE) of a three-electrode impedance device, fabricated on a polyimide (PI) film, is modified with SiNWs covered by a thin Au layer and functionalized to bind the sensing element. The biosensing behavior is investigated through the ligand-receptor binding of biotin-avidin system. Impedance measurements show a very efficient detection of the avidin over a broad range of concentrations from hundreds of micromolar down to the picomolar values. The impedance response is modeled through a simple equivalent circuit, which takes into account the unique WE morphology and its modification with successive layers of biomolecules. This approach of exploiting highly disordered SiNW ensemble in biosensing proves to be very promising for the following three main reasons: first, the system morphology allows high sensing performance; second, these nanostructures can be built via scalable and transferable fabrication methodology allowing an easy integration on non-conventional substrates; third, reliable modeling of the sensing response can be developed by considering the morphological and surface characteristics over an ensemble of disordered NWs rather than over individual NWs.

Silicon nanotubes from sacrificial silicon nanowires: fabrication and manipulation via embedding in flexible polymers.

In the present work we report a simple method to fabricate Si nanotubes (NTs) starting from the growth of self-assembled sacrificial Si nanowires that, at the same time, embeds them into a polyimide matrix, allowing a very easy manipulation of these nano-objects, including removal, transfer and positioning. Our all-silicon fabrication method is completely compatible with the Si technology platform and is therefore implementable using the existing technology. Transferred NTs show good electrical contact with underlying electrodes, and relatively low resistance values have been measured. All these features demonstrate the effectiveness of the transfer method and the potentiality of the NTs in electronics. Finally, optical reflectivity of the NTs has been measured in the near UV-near IR spectral range.