Assunta Pistone

A transparent organic transistor structure for bidirectional stimulation and recording of primary neurons

Real-time stimulation and recording of neural cell bioelectrical activity could provide an unprecedented insight in understanding the functions of the nervous system, and it is crucial for developing advanced in vitro drug screening approaches. Among organic materials, suitable candidates for cell interfacing can be found that combine long-term biocompatibility and mechanical flexibility. Here, we report on transparent organic cell stimulating and sensing transistors (O-CSTs), which provide bidirectional stimulation and recording of primary neurons. We demonstrate that the device enables depolarization and hyperpolarization of the primary neuron membrane potential. The transparency of the device also allows the optical imaging of the modulation of the neuron bioelectrical activity. The maximal amplitude-to-noise ratio of the extracellular recording achieved by the O-CST device exceeds that of a microelectrode array system on the same neuronal preparation by a factor of 16. Our organic cell stimulating and sensing device paves the way to a new generation of devices for stimulation, manipulation and recording of cell bioelectrical activity in vitro and in vivo.

Photostimulation of Whole-Cell Conductance in Primary Rat Neocortical Astrocytes Mediated by Organic Semiconducting Thin Films

Astroglial ion channels are fundamental molecular targets in the study of brain physiology and pathophysiology. Novel tools and devices intended for stimulation and control of astrocytes ion channel activity are therefore highly desirable. The study of the interactions between astrocytes and biomaterials is also essential to control and minimize reactive astrogliosis, in view of the development of implantable functional devices. Here, the growth of rat primary neocortical astrocytes on the top of a light sensitive, organic polymer film is reported; by means of patch-clamp analyses, the effect of the visible light stimulation on membrane conductance is then determined. Photoexcitation of the active material causes a significant depolarization of the astroglial resting membrane potential: the effect is associated to an increase in whole-cell conductance at negative potentials. The magnitude of the evoked inward current density is proportional to the illumination intensity. Biophysical and pharmacological characterization suggests that the ion channel mediating the photo-transduction mechanism is a chloride channel, the ClC-2 channel. These results open interesting perspectives for the selective manipulation of astrocyte bioelectrical activity by non-invasive, label-free, organic-based, photostimulation approaches.

Innovative multifunctional silk fibroin and hydrotalcite nanocomposites: a synergic effect of the components.

Novel hybrid functional materials are formed by combining hydrotalcite-like compounds and silk fibroin (SF-HTlc) via an environmental friendly aqueous process. The nanocomposites can be prepared with different weight ratio of the constituting components and preserve the conformational properties of the silk protein and the lamellar structure of hydrotalcites. Optical microscopy, scanning electron microscopy, and atomic force microscopy analyses show a good dispersion degree of the inorganic nanoparticles into the organic silk matrix. A mutual benefit on the stability of both organic and inorganic components was observed in the nanocomposites. SF-HTlc displayed limited dissolution of hydrotalcite in acidic medium, enhanced mechanical properties, and higher protease resistance of silk protein. The transparency, flexibility, and acidic environment resistance of silk fibroin combined to the protective and reinforcing properties of hydrotalcites generate a hybrid material, which is very attractive for applications in recently reported silk based opto-electronic and photonics technologies.

A nanostructured conductive bio-composite of silk fibroin–single walled carbon nanotubes

Silk fibroin (SF), a protein core fibre from the silkworm Bombyx mori, has huge potential to become a sustainable, biocompatible, and biodegradable material platform that can pave the way towards the replacement of plastic in the fabrication of bio-derived materials for a variety of technological and biomedical applications. SF has remarkable mechanical flexibility, controllable biodegradability, biocompatibility and is capable of drug/doping inclusion, stabilization and release. However, the dielectric properties of SF limit its potential as a direct bioelectronic interface in biomedical devices intended to control the bioelectrical activity of the cell for regenerative purposes. In this work, a novel wet templating method is proposed to generate nanostructured, conductive Silk Fibroin (SF) composite films. We combine the unusual properties of SF, such as its mechanical properties, its convenience and biocompatibility with the electrical conductivity and stiffness of Single Walled Carbon Nanotubes (SWCNTs). The presented SF-SWCNT composite displays a periodic architecture where SWCNTs are regularly and homogeneously distributed in the SF protein matrix. The morphological and chemo-physical properties of the nanocomposite are analysed and defined by SEM, Raman Spectroscopy, ATR-IR, UFM and contact angle analyses. Notably, the SF-SWCNT composite film is conductive, showing additional functionality compared to the dielectric properties of the bare SF film. Finally, SF-SWCNT is biocompatible and enables the growth of primary rat Dorsal Root Ganglion (DRG) neurons. Collectively our results demonstrate that the nanostructured, conductive, robust and biocompatible SF-SWCNT composite can be fabricated using a wet templating method, paving the way towards the fabrication and development of silk-based electronic devices for use in bioelectronic and biomedical applications.

A lysinated thiophene-based semiconductor as a multifunctional neural bioorganic interface.

Lysinated molecular organic semiconductors are introduced as valuable multifunctional platforms for neural cells growth and interfacing. Cast films of quaterthiophene (T4) semiconductor covalently modified with lysine-end moieties (T4Lys) are fabricated and their stability, morphology, optical/electrical, and biocompatibility properties are characterized. T4Lys films exhibit fluorescence and electronic transport as generally observed for unsubstituted oligothiophenes combined to humidity-activated ionic conduction promoted by the charged lysine-end moieties. The Lys insertion in T4 enables adhesion of primary culture of rat dorsal root ganglion (DRG), which is not achievable by plating cells on T4. Notably, on T4Lys, the number on adhering neurons/area is higher and displays a twofold longer neurite length than neurons plated on glass coated with poly-l-lysine. Finally, by whole-cell patch-clamp, it is shown that the biofunctionality of neurons cultured on T4Lys is preserved. The present study introduces an innovative concept for organic material neural interface that combines optical and iono-electronic functionalities with improved biocompatibility and neuron affinity promoted by Lys linkage and the softness of organic semiconductors. Lysinated organic semiconductors could set the scene for the fabrication of simplified bioorganic devices geometry for cells bidirectional communication or optoelectronic control of neural cells biofunctionality.

Sulfonates-PMMA nanoparticles conjugates: A versatile system for multimodal application

We report herein the viability of a novel nanoparticles (NPs) conjugated system, namely the attachment, based on ionic and hydrophobic interactions, of different sulfonated organic salts to positively charged poly(methylmethacrylate) (PMMA)-based core-shell nanoparticles (EA0) having an high density of ammonium groups on their shells. In this context three different applications of the sulfonates@EA0 systems have been described. In detail, their ability as cytotoxic drugs and pro-drugs carriers was evaluated in vitro on NCI-H460 cell line and in vivo against human ovarian carcinoma IGROV-1 cells. Besides, 8-hydroxypyrene-1,3,6-trisulfonic acid, trisodium salt (HPTS) was chosen for NPs loading, and its internalization as bioimaging probe was evaluated on Hep G2 cells. Overall, the available data support the interest for these PMMA NPs@sulfonates systems as a promising formulation for theranostic applications. In vivo biological data strongly support the potential value of these core-shell NPs as delivery system for negatively charged drugs or biologically active molecules. Additionally, we have demonstrated the ability of these PMMA core-shell nanoparticles to act as efficient carriers of fluorophores. In principle, thanks to the high PMMA NPs external charge density, sequential and very easy post-loading of different sulfonates is achievable, thus allowing the preparation of nanocarriers either with bi-modal drug delivery behaviour or as theranostic systems.

Bio-doping of regenerated silk fibroin solution and films: a green route for biomanufacturing

Silk fibroin (SF) is a natural biocompatible material that can be integrated in a variety of photonic systems and optoelectronics: i.e. organic lasing from dye-doped nano-structured silk film. In this context, biological incorporation of doping molecules into SF by means of feeding silk worms with dyes to their diet could be an innovative and eco-sustainable approach to obtain doped SF substrates, thus avoiding additional chemical processes and post-treatments of the protein solution. In the present work, we demonstrated that SF regenerated solutions and films containing rhodamine B (RhB) could be successfully obtained from the cocoons of Bombyx mori fed with a RhB-added diet (RhB-md-SF). Comparative analyses of optical and vibration characteristics of the RhB-md-SF solution and films with those of white SF blended with RhB (RhB-d-SF) revealed significant differences, suggesting that the silkworms metabolism could be involved in the binding mechanism of SF with RhB. In conclusion, we observed that the doping diet is a promising method for the green fabrication of SF-based optically active materials, and it opens novel routes for silk-based biophotonics.

Effect of different fabrication methods on the chemo-physical properties of silk fibroin films and on their interaction with neural cells

In this study, we investigated the influence of processing methods on the chemo-physical properties of silk fibroin (SF) film and on their interaction with neural cells. Structural, thermal and morphological analysis revealed a strong correlation between the conformation, stability and texture of silk films and the fabrication conditions. An increase in temperature, methanol treatment and the use of a microfluidic approach led to an improvement in SF film stability in terms of β-sheet content, mechanical resistance, dissolution and enzymatic degradation. An effect on the interaction of SF films with neural cells, through a modulation of the surface properties, was also observed. In particular, hydrophobic surfaces induce proliferation of astrocytes and neuron adhesion whereas hydrophilic surfaces promote a remarkable neurite outgrowth. A detailed knowledge of the effect of manufacturing parameters on SF film properties can facilitate and extend the applications of silk-based biomaterials in tissue engineering and drug release systems.

Core–shell poly-methylmethacrylate nanoparticles as effective carriers of electrostatically loaded anionic porphyrin

Among the medical applications of nanoparticles, their usage as photosensitizer (PS) carriers for photodynamic therapy (PDT) has attracted increasing attention. In the present study we explored the morphological and photophysical properties of core-shell PMMA nanoparticles (PMMA-NPs) electrostatically post-loaded with the synthetic, water soluble 5,10,15,20-tetrakis(4-sulphonatophenyl)-porphyrin (TPPS4). pH response and singlet oxygen analyses of differently loaded samples proved the high capability of the PMMA-NPs to shield the PS from the environment, while retaining the PS singlet oxygen production capability. Preliminary in vitro imaging and phototoxicity experiments on HepG2 cells demonstrated the efficacy of the system to trigger photoinduced cell death in the culture.

Selective MW-assisted surface chemical tailoring of hydrotalcites for fluorescent and biocompatible nanocomposites

ZnAl based hydrotalcite nanoparticles (ZnAl-HTlc NPs) were covalently modified by an organic oligothiophene fluorescent compound (T4Si) by using direct microwave (MW)-assisted silylation. Morphological and optical characterization proved that the MW-assisted method enables efficient grafting of the target fluorescent dye on the nanoparticles (NPs) surface in a few minutes with a predefined loading ratio only depending by the MW irradiation time. Moreover, the presented approach preserved the HTlc interlayer region, allowing further functionalization. Filmability, fluorescent properties, and biocompatibility of the silylated compound was also demonstrated highlighting the potential of the so-obtained lamellar NPs in applications broadening from diagnostic biomedical tools to photonics and sensing.