Organic electrochemical transistor: a tool for cell tissue monitoring

Questa tesi si inserisce nel ramo di ricerca della Bioelettronica organica, in particolare l'obiettivo è quello di monitorare per via elettrica la formazione e la rottura di ricoprimenti e barriere cellulari. Queste sono di particolare importanza dal punto di vista biologico per il loro ruolo di protezione e per l'azione regolatrice nel passaggio di ioni e macromolecole necessarie al benessere dell'organo. Per fare questo sono stati utilizzati gli Organic ElectroChemical Transistor (OECT) basati sul polimero organico biocompatibile PEDOT:PSS. Per poter portare a termine questo lavoro il primo passo è stato lo sviluppo di un apparato sperimentale, detto TE-OECT (Tissue Engineering-Organic ElectroChemical Transistor), che permettesse l'acquisizione delle misure a bassa intensità di segnale dall'interno di un incubatore oltre alla trasparenza necessaria all'acquisizione di misure ottiche utilizzate come riferimento. Oltre allo sviluppo, l'ottimizzazione e la calibrazione dei dispositivi e del TE-OECT, è stato sviluppato un programma per l'elaborazione dei dati. Sono state misurate le risposte degli OECT in differenti fasi della crescita di due ricoprimenti cellulari (HeLa e NIH-3T3). Per validare le misure elettriche nelle varie fasi della crescita cellulare sono state acquisite immagini al microscopio dei ricoprimenti studiati. Come ulteriore conferma di quanto osservato è stata utilizzata la Tripsina per provocare il distacco dei ricoprimenti. Sono state eseguite misure elettriche durante il distacco per verificare se fosse possibile monitorare in tempo reale l'integrità dei layer cellulari. Questo tipo di analisi permette di ottenere utili informazioni aggiuntive sullo stato del ricoprimento cellulare e rende possibile svincolare l'efficacia di un agente patogeno o l'efficacia di un enzima utilizzato per il distacco cellulare dalle analisi di tipo ottico

The physics of plasma membrane photostimulation

Cell membrane perturbation is a common way to stimulate cells by using external actuators. Recently, nanotechnology has added a number of new strategies for doing this, enlarging the scope and the range of mechanisms involved. Here, we describe a number of possible perturbation actions that are driven by light, and we try to capture the underlying phenomena. The discussion is based on the simple equivalent circuit model for the cell membrane

Modulation of Mechanosensitive Potassium Channels by a Membrane-targeted Nongenetic Photoswitch

Mechanosensitive ion channels are present in the plasma membranes of all cells. They play a fundamental role in converting mechanical stimuli into biochemical signals and are involved in several physiological processes such as touch sensation, hearing, and blood pressure regulation. This protein family includes TWIK-related arachidonic acid-stimulated K+ channel (TRAAK), which is specifically implicated in the maintenance of the resting membrane potential and in the regulation of a variety of important neurobiological functions. Dysregulation of these channels has been linked to various diseases, including blindness, epilepsy, cardiac arrhythmia, and chronic pain. For these reasons, mechanosensitive channels are targets for the treatment of several diseases. Here, we propose a new approach to investigate TRAAK ion channel modulation that is based on nongenetic photostimulation. We employed an amphiphilic azobenzene, named Ziapin2. In the dark, Ziapin2 preferentially dwells in the plasma membrane, causing a thinning of the membrane. Upon light irradiation, an isomerization occurs, breaking the dimers and inducing membrane relaxation. To study the effect of Ziapin2 on the mechanosensitive channels, we expressed human TRAAK (hTRAAK) channels in HEK293T cells. We observed that Ziapin2 insertion in the membrane is able per se to recruit hTRAAK, permitting the exit of K+ ions outside the cells with a consequent hyperpolarization of the cell membrane. During light stimulation, membrane relaxation induces hTRAAK closure, generating a consistent and compensatory depolarization. These results add information to the Ziapin2 mechanism and suggest that membrane deformation can be a tool for the nonselective modulation of mechanosensitive channels

Skeletal muscle cells opto-stimulation by intramembrane molecular transducers

Optical stimulation and control of muscle cell contraction opens up a number of interesting applications in hybrid robotic and medicine. Here we show that recently designed molecular phototransducer can be used to stimulate C2C12 skeletal muscle cells, properly grown to exhibit collective behaviour. C2C12 is a skeletal muscle cell line that does not require animal sacrifice Furthermore, it is an ideal cell model for evaluating the phototransducer pacing ability due to its negligible spontaneous activity. We study the stimulation process and analyse the distribution of responses in multinuclear cells, in particular looking at the consistency between stimulus and contraction. Contractions are detected by using an imaging software for object recognition. We find a deterministic response to light stimuli, yet with a certain distribution of erratic behaviour that is quantified and correlated to light intensity or stimulation frequency. Finally, we compare our optical stimulation with electrical stimulation showing advantages of the optical approach, like the reduced cell stress.A study on the stimulation of cells with light, thanks to a photochromic molecule, called Ziapin2. It shows the difference between the light stimulation and the electrical stimulation in terms of cell viability and performance

Stable and Solution-Processable Cumulenic sp-Carbon Wires: A New Paradigm for Organic Electronics

[EN] Solution-processed, large-area, and flexible electronics largely relies on the excellent electronic properties of sp(2)-hybridized carbon molecules, either in the form of pi-conjugated small molecules and polymers or graphene and carbon nanotubes. Carbon with sp-hybridization, the foundation of the elusive allotrope carbyne, offers vast opportunities for functionalized molecules in the form of linear carbon atomic wires (CAWs), with intriguing and even superior predicted electronic properties. While CAWs represent a vibrant field of research, to date, they have only been applied sparingly to molecular devices. The recent observation of the field-effect in microcrystalline cumulenes suggests their potential applications in solution-processed thin-film transistors but concerns surrounding the stability and electronic performance have precluded developments in this direction. In the present study, ideal field-effect characteristics are demonstrated for solution-processed thin films of tetraphenyl[3]cumulene, the shortest semiconducting CAW. Films are deposited through a scalable, large-area, meniscus-coating technique, providing transistors with hole mobilities in excess of 0.1 cm(2 )V(-1 )s(-1), as well as promising operational stability under dark conditions. These results offer a solid foundation for the exploitation of a vast class of molecular semiconductors for organic electronics based on sp-hybridized carbon systems and create a previously unexplored paradigm.E.G.F. acknowledges the support through the EU Horizon 2020 research and innovation program, H2020-FETOPEN-01-2018-2020 (FET-Open Challenging Current Thinking), "LION-HEARTED", grant agreement no. 828984. C.S.C. acknowledges funding from the European Research Council (ERC) under the European Union's Horizon 2020 research and innovation program ERC-Consolidator Grant (ERC CoG 2016 EspLORE grant agreement no. 724610, website: ). R.R.T. acknowledges funding from the Natural Sciences and Engineering Research Council of Canada (NSERC) and the Canada Foundation for Innovation (CFI). This work was partially supported by the European Union's H2020-EU.4.b. - Twinning of research institutions "GREENELIT", grant agreement number 951747. GIWAXS experiments were performed at BL11 NCD-SWEET beamline at ALBA Synchrotron (Spain) with the collaboration of ALBA staff. This work was in part carried out at Polifab, the micro- and nanotechnology centre of the Politecnico di Milano. Open access funding provided by Istituto Italiano di Tecnologia within the CRUI-CARE Agreement

Optical modulation of excitation-contraction coupling in human-induced pluripotent stem cell-derived cardiomyocytes

Non-genetic photostimulation is a novel and rapidly growing multidisciplinary field that aims to induce light-sensitivity in living systems by exploiting exogeneous phototransducers. Here, we propose an intramembrane photoswitch, based on an azobenzene derivative (Ziapin2), for optical pacing of human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). The light-mediated stimulation process has been studied by applying several techniques to detect the effect on the cell properties. In particular, we recorded changes in membrane capacitance, in membrane potential (V-m), andmodulation of intracellular Ca2+ dynamics. Finally, cell contractility was analyzed using a custom MATLAB algorithm. Photostimulation of intramembrane Ziapin2 causes a transient V-m hyperpolarization followed by a delayed depolarization and action potential firing. The observed initial electrical modulation nicely correlates with changes in Ca2+ dynamics and contraction rate. This work represents the proof of principle that Ziapin2 can modulate electrical activity and contractility in hiPSC-CMs, opening up a future development in cardiac physiology

Phytotoxic Effects of Essential Oils from Six Lamiaceae Species

Essential oils produced by plants, and their components, could be sources of new natural herbicidal compounds. Thirteen oils extracted from six wild Lamiaceae species (namely Clinopodium suaveolens (Sm.) Kuntze, Satureja montana L. subsp. montana, Thymbra capitata (L.) Cav., Salvia fruticosa Mill. subsp. thomasii (Lacaita) Brullo, Guglielmo, Pavone & Terrasi, Satureja cuneifolia Ten., and Thymus spinulosus Ten.) from South Italy were tested in vitro for the phytotoxic activity to cress and branched broomrape seeds, tomato radicles, and lambsquarters leaf disks. Moreover, the possible correlation between oil composition and biological activity was evaluated. One of the oils from T. capitata inhibited cress germination by 96.4% at the lowest tested concentration (100 ppm) and reduced both chlorophyll and carotenoid content in lambsquarters leaf disks by around 50%. Some oils, particularly those from T. spinulosus, inhibited tomato radicle elongation by 85% at 1000 ppm. Many oils inhibited broomrape seed germination up to 100% when tested in solution at 1000 ppm or released as vapors. Among the oil components, α-terpinene, p-cymene, β-cis-ocimene, cis-sabinene hydrate, carvacrol methyl ether, and thymol were mostly correlated to the inhibition of cress seeds germination and tomato radicle elongation. The presence of thymol and p-cymene was also correlated to the inhibition of broomrape seed germination. Some of the tested essential oils or their components could have potential as pre-emergence herbicides and could be useful in the development of new weed control strategies

Gold-chlorophyll a-hybrid nanoparticles and chlorophyll a/cetyltrimethylammonium chloride self-assembled-suprastructures as novel carriers for chlorophyll a delivery in water medium: Photoactivity and photostability

The stability of Chlorophyll a in water during prolonged exposure, at room temperature, to a neon lamp has been investigated by means of UV–vis and fluorescence spectroscopies. In addition, the Chlorophyll a (photo)stability evaluation in presence of suitable carriers has been performed in order to investigate its reactivity under the same conditions, for possible and future applications in Antimicrobial Photodynamic Therapy. Cetyltrimethylammonium chloride was chosen to solubilize Chlorophyll a in water. While, cetyltrimethylammonium chloride-capped gold nanoparticles offer a great opportunity because combine the Chlorophyll a action, used as a photosensitizer in Antimicrobial Photodynamic Therapy, with gold nanoparticles effect used in photothermal therapy. Indeed, the latter ones have exhibited an interesting rise of temperature if irradiated with visible light. Overall, both examined systems, cetyltrimethylammonium chloride/Chlorophyll a and gold nanoparticles/Chlorophyll a, were able to induce the Reactive Oxygen Species formation fundamental for a potential application in Antimicrobial Photodynamic Therapy

Skeletal muscle cells opto-stimulation by intramembrane molecular transducers

Data of contracting muscle cells upon optical and electrical stimulatio