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Potassium channels and aquaporins expressed by astrocytes are key players in the maintenance of cerebral homeostasis and in brain pathophysiologies. One major challenge in the study of astrocyte membrane channels in vitro, is that their expression pattern does not resemble the one observed in vivo. Nanostructured interfaces represent a significant resource to control the cellular behaviour and functionalities at micro and nanoscale as well as to generate novel and more reliable models to study astrocytes in vitro. However, the potential of nanotechnologies in the manipulation of astrocytes ion channels and aquaporins has never been previously reported. Hydrotalcite-like compounds (HTlc) are layered materials with increasing potential as biocompatible nanoscale interface. Here, we evaluate the effect of the interaction of HTlc nanoparticles films with primary rat neocortical astrocytes. We show that HTlc films are biocompatible and do not promote gliotic reaction, while favouring astrocytes differentiation by induction of F-actin fibre alignment and vinculin polarization. Western Blot, Immunofluorescence and patch-clamp revealed that differentiation was accompanied by molecular and functional up-regulation of both inward rectifying potassium channel Kir 4.1 and aquaporin 4, AQP4. The reported results pave the way to engineering novel in vitro models to study astrocytes in a in vivo like condition.
Assuntos
Astrócitos/citologia , Materiais Biocompatíveis/química , Nanoestruturas , Actinas/metabolismo , Hidróxido de Alumínio/química , Animais , Aquaporina 4/metabolismo , Astrócitos/metabolismo , Técnicas de Cultura de Células , Sobrevivência Celular , Células Cultivadas , Citoesqueleto/metabolismo , Gliose/metabolismo , Hidróxido de Magnésio/química , Teste de Materiais , Microscopia Confocal , Microscopia Eletrônica de Varredura , Microscopia de Fluorescência , Técnicas de Patch-Clamp , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Ratos , Vinculina/metabolismoRESUMO
Photoactivatable keratin sponges were prepared from protein aqueous solutions by the freeze-drying method, followed by photofunctionalization with two different photosensitizers (PS): Azure A (AzA) and 5,10,15,20-tetrakis [4-(2-N,N,N-trimethylethylthio)-2,3,5,6-tetrafluorophenyl]porphyrin tetraiodide salt (TTFAP). The prepared sponges have a porosity between 49% and 80% and a mean pore size in the 37-80 µm range. As compared to AzA, TTFAP interacts more strongly with the sponges as demonstrated by a lower PS release (6% vs 20%), a decreased swelling ratio (1.6 vs 7.4), and a slower biodegradation rate. Nevertheless, AzA-loaded sponges showed the highest photoactivity, as also demonstrated by their higher antibactericidal activity toward both Gram-positive and Gram-negative bacteria. The obtained results suggest that the antimicrobial photodynamic effect can be finely triggered through a proper selection of the amount and type of photosensitizer, as well as through the irradiation time. Finally, all the prepared sponges support human fibroblast cells growth, while no significant cell viability impairment is observed upon light irradiation.
Assuntos
Anti-Infecciosos/farmacologia , Queratinas/química , Queratinas/farmacologia , Fármacos Fotossensibilizantes/farmacologia , Pseudomonas aeruginosa/efeitos dos fármacos , Staphylococcus aureus/efeitos dos fármacos , Lã/química , Animais , Anti-Infecciosos/química , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Células Cultivadas , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/efeitos da radiação , Humanos , Luz , Pseudomonas aeruginosa/efeitos da radiação , Staphylococcus aureus/efeitos da radiaçãoRESUMO
The use of doped silk fibroin (SF) films and substrates from Bombyx mori cocoons for green nanotechnology and biomedical applications has been recently highlighted. Cocoons from coloured strains of B. mori, such as Golden-Yellow, contain high levels of pigments that could have a huge potential for the fabrication of SF based biomaterials targeted to photonics, optoelectronics and neuroregenerative medicine. However, the features of extracted and regenerated SF from cocoons of B. mori Golden-Yellow strain have never been reported. Here we provide a chemophysical characterization of regenerated silk fibroin (RSF) fibers, solution, and films obtained from cocoons of a Golden-Yellow strain of B. mori, by SEM, (1) H-NMR, HPLC, FT-IR, Raman and UV-Vis spectroscopy. We found that the extracted solution and films from B. mori Golden-Yellow fibroin displayed typical Raman spectroscopic and optical features of carotenoids. HPLC-analyses revealed that lutein was the carotenoid contained in the fiber and RSF biopolymer from yellow cocoons. Notably, primary neurons cultured on yellow SF displayed a threefold higher neurite length than those grown of white SF films. The results we report pave the way to expand the potential use of yellow SF in the field of neuroregenerative medicine and provide green chemistry approaches in biomedicine.
Assuntos
Axônios , Materiais Biocompatíveis , Fibroínas/química , Luteína/química , Neurônios/citologia , Seda/química , Animais , Bombyx , Espectrofotometria Ultravioleta , Espectroscopia de Infravermelho com Transformada de FourierRESUMO
We report the design, synthesis and structure-property investigation of a new perylene diimide material (PDI-Lys) bearing lysine end substituents. Water processed films of PDI-Lys were prepared and their self-assembly, morphology and electrical properties in both inert and air environments were theoretically and experimentally investigated. With the aim of evaluating the potential of PDI-Lys as a biocompatible and functional neural interface for organic bioelectronic applications, its electrochemical impedance as well as the adhesion and viability properties of primary neurons on the PDI-Lys films were studied. By combining theoretical calculations and electrical measurements we show that due to conversion between neutral and zwitterionic anions, the PDI-Lys film conductivity increased significantly upon passing from air to an inert atmosphere, reaching a maximum value of 6.3 S m-1. We also show that the PDI-Lys film allows neural cell adhesion and neuron differentiation and decreases up to 5 times the electrode/solution impedance in comparison to a naked gold electrode. The present study introduces an innovative, water processable conductive film usable in organic electronics and as a putative neural interface.
RESUMO
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.
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Materiais Biocompatíveis/química , Semicondutores , Tiofenos/química , Animais , Adesão Celular , Células Cultivadas , Gânglios Espinais/citologia , Gânglios Espinais/metabolismo , Lisina/química , Microscopia de Força Atômica , Microscopia Confocal , Estrutura Molecular , Neuritos/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Ratos , Propriedades de SuperfícieRESUMO
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.
Assuntos
Hidróxido de Alumínio/química , Fibroínas/química , Hidróxido de Magnésio/química , Nanocompostos/química , Seda/química , Animais , Bombyx , Sinergismo Farmacológico , Fibroínas/fisiologia , Seda/fisiologiaRESUMO
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.
RESUMO
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.
Assuntos
Potenciais de Ação/fisiologia , Estimulação Elétrica/instrumentação , Eletrodos Implantados , Microeletrodos , Neurônios/fisiologia , Transistores Eletrônicos , Animais , Células Cultivadas , Desenho de Equipamento , Análise de Falha de Equipamento , Compostos Orgânicos/química , Ratos , Refratometria/instrumentaçãoRESUMO
The bioelectrical signalling within neural networks has to be monitored in real-time and localized in space in order to unravel the mechanisms behind pathologies and diseases of the nervous systems. Organic materials have significant potential for bio-functional neural interfacing given that their "soft" nature offers better mechanical compatibility with the nerve tissues than conventional semiconductors, and their flexibility allows realization of the non-planar forms typically required for biomedical implants. The integration of living cells into organic semiconductors is an important step towards the development of bio-organic electronic transducers of cellular activity from neurons. Here, we report on the use and characterization of n-type perylene derivatives as a suitable interface platform for organic neuro-electronic devices. We demonstrate that primary neurons can adhere, grow and differentiate on a suitably engineered perylene-based field-effect transistor platform, while maintaining their firing properties even after a prolonged time of cell-culturing. It is noteworthy that the field-effect transistors preserve their electrical characteristics even after 10 days of incubation in cell culture media. These results validate n-type perylene derivatives as a suitable long-term interface platform for organic neuro-electronic devices, which is particularly relevant in view of the recently reported perylene-based field-effect transistor structure capable of providing bidirectional stimulation and recording of primary neurons.
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Ahead of print article withdrawn by publisher
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The objective of this research was to realize a new oral solid dosage form in order to improve the release of furosemide (FURO) in its preferential absorption region. In fact FURO is a drug labeled in class IV of the Biopharmaceutical Classification System (BCS) characterized by low and variable bioavailability due to both low solubility and low permeability and because of its weakly acid nature is preferentially absorbed in the stomach whereas its solubility is hampered. FURO was included in the mesoporous silica material SBA-15 obtaining an inorganic-organic compound fully characterized by: thermogravimetric analysis (TGA), X-ray Powder Diffraction (XRPD), Differential Scanning Calorimetry (DSC), Fourier Transform Infrared Spectroscopy (FT-IR) and nitrogen adsorption-desorption analysis and then submitted to in vitro dissolution. The results showed a remarkable dissolution rate improvement in comparison to the crystalline drug and to the marketed product Lasix®. The inclusion product was also submitted to physical stability studies that revealed the matrix ability to prevent re-organization in crystal nucleus of the drug molecules.