Tuning cellular functionality and mechanobiology via carbon nanotubes based scaffolds

La rigenerazione tissutale che occorre in seguito all\u2019insorgere di malattie e/o lesioni attraverso l\u2019organizzazione delle cellule in organi/tessuti, \ue8 fortemente indebolita dagli stimoli meccanici e biochimici esercitati dall\u2019ambiente extracellulare danneggiato, i quali impattano definitivamente sul destino cellulare. Per guidare l\u2019abilit\ue0 rigenerativa dei tessuti, possono essere sfruttati biomateriali mimanti la complessa architettura fisiologica. La matrice extracellulare (ECM), tuttavia, \ue8 caratterizzata da un intricato network di elementi nanostrutturati che si adattano agli input cellulari fornendo gli stimoli attraverso i quali le cellule attivano i percorsi di meccanotrasduzione necessari per la modulazione delle loro funzioni. Poich\ue9 le cellule normalmente interagiscono con queste nanostrutture presenti nel loro ambiente, un requisito fondamentale per i biomateriali mimetici \ue8 il fatto di possederli. Non \ue8 sorprendente, quindi, il grande interesse rivolto ai nanotubi di carbonio (CNTs) negli ultimi decenni, grazie alle tante similarit\ue0 con la ECM, morfologiche e dimensionali, oltre alle loro peculiari propriet\ue0 chimico-fisiche e meccaniche. Essi hanno inequivocabilmente dimostrato la loro abilit\ue0 di potenziare l\u2019attivit\ue0 elettrica della rete neurale. Negli studi precedenti, CNTs purificati erano depositati su substrato di vetro mediante drop-casting. Qui, abbiamo dimostrato, per la prima volta, che i CNTs cresciuti direttamente su substrati di silicio tramite deposizione chimica da fase vapore catalitica (CCVD) preservano lo stesso effetto di potenziamento con il vantaggio, per\uf2, di poter modulare facilmente le propriet\ue0 della matrice a base di carbonio che possono essere utilizzate senza la necessit\ue0 di alcuna purificazione chimica e/o funzionalizzazione semplificando notevolmente il loro uso. Con lo scopo di sfruttare le potenzialit\ue0 del nostro tappeto di CNTs come biomateriale artificiale per la rigenerazione tissutale, sono richiesti risultati sperimentali da tecniche complementari. Tuttavia, i nostri substrati cresciuti su silicio, mancano della trasparenza ottica, primariamente a causa del silicio stesso. Questo limita l\u2019uso di tutte quelle tecniche di caratterizzazione che richiedono di visualizzare le cellule \u2018attraverso\u2019 il campione (l\u2019elettrofisiologia e la microscopia in campo chiaro). Con tale scopo, abbiamo sviluppato una nuova strategia per fabbricare substrati trasparenti di CNTs (tCNTs) attraverso il CCVD direttamente su un substrato trasparente e controllando accuratamente la loro lunghezza. Abbiamo dimostrato che tali supporti a base di carbonio inducono su colture ippocampali lo stesso potenziamento sinaptico, in precedenza osservato per i substrati tradizionali (drop-casted-CNTs). Abbiamo investigato, inoltre, la loro abilit\ue0 di supportare la crescita del complesso tessuto neuronale come le colture Entorinali-Ippocampali (EHCs) dimostrando, per la prima volta, che il nostro nanomateriale pu\uf2 aiutare riconnessione funzionale dei tessuti neuronali lesionati. I tCNTs possono anche essere sfruttati per migliorare le attuali strategie adottate per i trattamenti delle malattie cardiovascolari (CAVD) che ad oggi non determinano una soluzione a lungo termine. In particolare, il nostro interesse \ue8 stato rivolto alla calcificazione della valvola aortica (CAVD), legata a variazioni della ECM, in termini di composizione, organizzazione e propriet\ue0 meccaniche. Di conseguenza, sulla base del ruolo cruciale della ECM in questa malattia e considerando, inoltre, le similarit\ue0 tra CNTs ed ECM, abbiamo studiato il loro effetto sulle cellule valvolari interstiziali (pVICs), costituenti principali della valvola aortica. Abbiamo dimostrato che essi possono fornire un ambiente fisiologico per lo \u2018sviluppo\u2019 delle VICs in cui la quantit\ue0 dei miofibroblasti, legato alla CAVD, \ue8 simile a quello caratterizzante le valvole sane.Natural tissue self-regeneration, occurring at the onset of injury or disease through the self-organization of cells into organs/tissues, is strongly impaired by mechanical and biochemical cues from the damaged extracellular environment, which impact cell fate. To drive tissue self-renewal ability, artificial biomaterials mimicking the complex architecture of the physiological cell microenvironment are highly desired. The natural extracellular matrix (ECM), however, displays an intricate network of nanoscale structures, whose morphology adapts to cell input, providing in turn mechanical cues to the surrounding cells which activate the biochemical and mechano-transduction pathways necessary for the modulation of their functions. Since cells normally interact with typical nanometer-scale elements present in their environment, nanoscale features are the first essential requirement for the design of biomimetic scaffolds. In this context, it is not surprising that carbon nanotubes (CNTs), owning various similarities with the native ECM, physico-chemical and mechanical properties, have captured increased attention. CNTs unequivocally demonstrated their ability to perturb electrical activity of neuronal networks. In previous studies, cell cultures were grown on purified CNTs deposited on supporting surfaces via drop casting. Here, we demonstrate that CNTs directly grown on a supporting silicon surface by catalytic chemical vapor deposition (CCVD) technique bear the same potentiating effect, with the added value of easy modulation of the CNT matrix properties. In our approach we developed a novel and well-controllable synthesis method leading to the realization of various CNTs-based architectures which could be employed as-produced, without the necessity of any chemical purification /functionalization, thus significantly simplifying their use. To further exploit the potential of our CNTs for tissue regeneration, experimental results from complementary techniques are required. Such substrates grown on silicon surfaces, however, lack of optical transparency, preventing its exploitation with all the investigation techniques requiring to optically visualize cells \u2018through\u2019 the specimens (electrophysiology and bright field microscopy). Therefore, we developed a novel strategy to fabricate transparent carbon nanotubes substrates (tCNTs) by synthesizing these carbon nanostructure via CCVD directly on a transparent substrate (i.e. fused silica) and finely controlling their length. We demonstrated that this original fabrication \u201crecipe\u201d gives rise to CNT carpet able to induce the same synaptic potentiation in hippocampal cells we observed in the case of opaque CNT films and drop-casted layers. We further investigated the ability of tCNTs to support the growth of complex neuronal tissues as intact and lesioned Entorhinal-Hippocampal slice cultures (EHCs), demonstrating that our nanomaterial can help in promoting a successful reconnection and functional cross talk between the two slices after the lesion. CNTs-based scaffolds can be exploited also to improve the standard strategies adopted for the treatments of cardiovascular diseases (CVD) which currently do not lead to a long-term solution. In particular, our interest has been directed towards calcific aortic valve diseases (CAVD), strongly related to significant changes in ECM organization, composition and mechanical properties. Therefore, based on the crucial role of ECM properties have on the progression of this disease and considering also the peculiar CNTs ability to structurally emulate the native ECM, we interfaced our novel tCNTs scaffold with porcine valve interstitial cells (pVICs), the predominant constituent of aortic valve. We demonstrated that tCNTs substrates can provide a physiological environment for VICs development in which the amount of myofibroblasts, related to CAVD, is similar to that characterizing healthy valves

Carbon nanostructures for directional light dark matter detection

Carbon nanostructures offer exciting new possibilities in the detection of light dark matter. A darkmatter particle with mass between 1 MeV and 1 GeV scattering off an electron in the carbon wouldtransfer sufficient energy to extract the electron from the lattice. In 2D materials, such as grapheneor carbon nanotubes, these electrons would be released directly into the vacuum, avoiding theirre-absorption in the medium. We present two novel detector concepts: a ’Graphene-FET’ design,based on graphene sheets, developed at Princeton University; and a ’Dark-PMT’ based on alignedcarbon nanotubes, developed in University of Rome Sapienza. We discuss their light dark matterdiscovery potential, the status of the RD, and the recent commissioning of a state-of-the-art carbonnanotube growing facility in Rome

Searching for Light Dark Matter with Aligned Carbon Nanotubes: The ANDROMeDa Project

The ANDROMeDa (Aligned Nanotube Detector for Research On MeV Dark matter) project aims to develop a novel Dark Matter detector based on carbon nanotubes: the “Dark-PMT”. The detector is designed to be sensitive to dark matter particles with mass between 1 MeV and 1 GeV. The detection scheme is based on dark matter-electron scattering inside a target made of vertically-aligned carbon nanotubes. Vertically-aligned carbon nanotubes have reduced density in the direction of the tube axis, therefore the scattered electrons are expected to leave the target without being re-absorbed only if their momentum has a small enough angle with that direction, which is what happens when the tubes are parallel to the dark matter wind. This grants directional sensitivity to the detector, a unique feature in this dark matter mass range

Transmission through graphene of electrons in the 30 – 900 eV range

Here, we report on accurate transmission measurements of electrons below 1 keV through suspended monolayer graphene. Monolayer graphene was grown via chemical vapor deposition and transferred onto transmission electron microscopy (TEM) grids. A monochromatic electron gun has been employed to perform the measurements in the 30 – 900 eV range in ultra-high vacuum. The graphene transparency is obtained from the absolute measurement of the direct beam current and the transmitted one, by means of a Faraday cup. We observed a transmission going from 20 to 80% for monolayer graphene within the experimental electron energy range. The high quality and the grid coverage of the suspended graphene has been proved via micro-Raman, X-ray photoemission, electron energy loss spectroscopies and field-emission scanning electron microscopy. After a 550 °C in-vacuum annealing of the samples, the main contribution to the C 1s spectrum is due to the component and the evidence of suspended monolayer graphene has been observed through the -plasmon excitation

Graphene Oxide Nanosheets Reshape Synaptic Function in Cultured Brain Networks

Graphene offers promising advantages for biomedical applications. However, adoption of graphene technology in biomedicine also poses important challenges in terms of understanding cell responses, cellular uptake, or the intracellular fate of soluble graphene derivatives. In the biological microenvironment, graphene nanosheets might interact with exposed cellular and subcellular structures, resulting in unexpected regulation of sophisticated biological signaling. More broadly, biomedical devices based on the design of these 2D planar nanostructures for interventions in the central nervous system require an accurate understanding of their interactions with the neuronal milieu. Here, we describe the ability of graphene oxide nanosheets to down-regulate neuronal signaling without affecting cell viability

Transparent carbon nanotubes promote the outgrowth of enthorino-dentate projections in lesioned organ slice cultures

The increasing engineering of carbon-based nanomaterials as components of neuro-regenerative interfaces is motivated by their dimensional compatibility with subcellular compartments of excitable cells, such as axons and synapses. In neuroscience applications, carbon nanotubes (CNTs) have been used to improve electronic device performance by exploiting their physical properties. Besides, when manufactured to interface neuronal networks formation in vitro, CNT carpets have shown their unique ability to potentiate synaptic networks formation and function. Due to the low optical transparency of CNTs films, further developments of these materials in neural prosthesis fabrication or in implementing interfacing devices to be paired with in vivo imaging or in vitro optogenetic approaches are currently limited. In the present work, we exploit a new method to fabricate CNTs by growing them on a fused silica surface, which results in a transparent CNT-based substrate (tCNTs). We show that tCNTs favour dissociated primary neurons network formation and function, an effect comparable to the one observed for their dark counterparts. We further adopt tCNTs to support the growth of intact or lesioned Entorhinal-Hippocampal Complex organotypic cultures (EHCs). Through immunocytochemistry and electrophysiological field potential recordings, we show here that tCNTs platforms are suitable substrates for the growth of EHCs and we unmask their ability to significantly increase the signal synchronization and fibre sprouting between the cortex and the hippocampus with respect to Controls. tCNTs transparency and ability to enhance recovery of lesioned brain cultures, make them optimal candidates to implement implantable devices in regenerative medicine and tissue engineering. This article is protected by copyright. All rights reserved

The Gaia mission

Gaia is a cornerstone mission in the science programme of the EuropeanSpace Agency (ESA). The spacecraft construction was approved in 2006, following a study in which the original interferometric concept was changed to a direct-imaging approach. Both the spacecraft and the payload were built by European industry. The involvement of the scientific community focusses on data processing for which the international Gaia Data Processing and Analysis Consortium (DPAC) was selected in 2007. Gaia was launched on 19 December 2013 and arrived at its operating point, the second Lagrange point of the Sun-Earth-Moon system, a few weeks later. The commissioning of the spacecraft and payload was completed on 19 July 2014. The nominal five-year mission started with four weeks of special, ecliptic-pole scanning and subsequently transferred into full-sky scanning mode. We recall the scientific goals of Gaia and give a description of the as-built spacecraft that is currently (mid-2016) being operated to achieve these goals. We pay special attention to the payload module, the performance of which is closely related to the scientific performance of the mission. We provide a summary of the commissioning activities and findings, followed by a description of the routine operational mode. We summarise scientific performance estimates on the basis of in-orbit operations. Several intermediate Gaia data releases are planned and the data can be retrieved from the Gaia Archive, which is available through the Gaia home page. http://www.cosmos.esa.int/gai

The dark-PMT: A novel directional light dark matter detector based on vertically-aligned carbon nanotubes

The ANDROMeDa project, recently funded by the italian ministry of research with a 1M€ grant, has the objective of developing, over the course of the next three years, a novel dark matter (DM) detector based on carbon nanotubes: the ‘dark-PMT’. Such a detector would be sensitive to DM-electron recoils in the eV energy range, and could have world-leading sensitivity for DM masses below 30 MeV with an exposure of only . Significant R&D is needed to produce carbon nanotubes with ideal properties for a DM target. In particular two by-products of synthesis (non-aligned crust layer and the sub-m ‘waviness’) are expected to reduce electron transmission probability, and therefore need to be minimized. This will be done via a precise tuning of the evaporation and growth parameters, and of post-growth plasma etching

Searching for Light Dark Matter with Aligned Carbon Nanotubes: The ANDROMeDa Project

The ANDROMeDa (Aligned Nanotube Detector for Research On MeV Dark matter) project aims to develop a novel Dark Matter detector based on carbon nanotubes: the “Dark-PMT”. The detector is designed to be sensitive to dark matter particles with mass between 1 MeV and 1 GeV. The detection scheme is based on dark matter-electron scattering inside a target made of vertically-aligned carbon nanotubes. Vertically-aligned carbon nanotubes have reduced density in the direction of the tube axis, therefore the scattered electrons are expected to leave the target without being re-absorbed only if their momentum has a small enough angle with that direction, which is what happens when the tubes are parallel to the dark matter wind. This grants directional sensitivity to the detector, a unique feature in this dark matter mass range

Response of Windowless Silicon Avalanche Photo-Diodes to Electrons in the 90-900 eV Range

We report on the characterization of the response of windowless silicon avalanche photo-diodes to electrons in the 90-900 eV energy range. The electrons were provided by a monoenergetic electron gun present in the LASEC laboratories of University of Roma Tre. We find that the avalanche photo-diode generates a current proportional to the current of electrons hitting its active surface. The gain is found to depend on the electron energy $E_e$, and varies from $2.147 \pm 0.027$ (for $E_e = 90$ eV) to $385.8 \pm 3.3$ (for $E_e = 900$ eV), when operating the diode at a bias of $V_{apd} = 350$ V.} This is the first time silicon avalanche photo-diodes are employed to measure electrons with $E_e < 1$ keV