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 ‘through’ 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 “recipe” 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.

La rigenerazione tissutale che occorre in seguito all’insorgere di malattie e/o lesioni attraverso l’organizzazione delle cellule in organi/tessuti, è fortemente indebolita dagli stimoli meccanici e biochimici esercitati dall’ambiente extracellulare danneggiato, i quali impattano definitivamente sul destino cellulare. Per guidare l’abilità rigenerativa dei tessuti, possono essere sfruttati biomateriali mimanti la complessa architettura fisiologica. La matrice extracellulare (ECM), tuttavia, è 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. Poiché le cellule normalmente interagiscono con queste nanostrutture presenti nel loro ambiente, un requisito fondamentale per i biomateriali mimetici è il fatto di possederli. Non è sorprendente, quindi, il grande interesse rivolto ai nanotubi di carbonio (CNTs) negli ultimi decenni, grazie alle tante similarità con la ECM, morfologiche e dimensionali, oltre alle loro peculiari proprietà chimico-fisiche e meccaniche. Essi hanno inequivocabilmente dimostrato la loro abilità di potenziare l’attività 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, però, di poter modulare facilmente le proprietà della matrice a base di carbonio che possono essere utilizzate senza la necessità di alcuna purificazione chimica e/o funzionalizzazione semplificando notevolmente il loro uso. Con lo scopo di sfruttare le potenzialità 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’uso di tutte quelle tecniche di caratterizzazione che richiedono di visualizzare le cellule ‘attraverso’ il campione (l’elettrofisiologia 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 abilità di supportare la crescita del complesso tessuto neuronale come le colture Entorinali-Ippocampali (EHCs) dimostrando, per la prima volta, che il nostro nanomateriale può 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 è stato rivolto alla calcificazione della valvola aortica (CAVD), legata a variazioni della ECM, in termini di composizione, organizzazione e proprietà meccaniche. Di conseguenza, sulla base del ruolo cruciale della ECM in questa malattia e considerando, inoltre, le similarità 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 ‘sviluppo’ delle VICs in cui la quantità dei miofibroblasti, legato alla CAVD, è simile a quello caratterizzante le valvole sane.

Tuning cellular functionality and mechanobiology via carbon nanotubes based scaffolds / Rago, ILARIA, CARMELA. - (2018 Feb 28).

Tuning cellular functionality and mechanobiology via carbon nanotubes based scaffolds

RAGO, ILARIA, CARMELA
2018-02-28

Abstract

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 ‘through’ 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 “recipe” 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.
28-feb-2018
SCAINI, DENIS
30
2016/2017
Settore FIS/03 - Fisica della Materia
Università degli Studi di Trieste
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11368/2919812
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