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Interfaces between conjugated polymers and aqueous electrolytes are gaining increasing interest for a number of different applications, including electrochemical cells, biosensors and functional organic devices for recording and stimulating the bioelectrical activity. In this field, devices sensitive to visible light are particularly interesting, both for inducing solar-driven photo-electrochemical reactions and for spatially selective, not-invasive optical control of living systems. We demonstrate that photoexcitation of the conjugated polymer leads to both capacitive and faradaic electrical signals at the interface with water. We introduce here a mesoporous, light sensitive polymer/electrolyte interface, we characterize its behavior and we reproduce the experimental data with an equivalent circuit model. Interestingly, we show that the use of polymer-based mesoporous devices offers a practical and simple handle for the modulation of the interface behavior. Device engineering allows to control the sign of charges at the polymer surface and to balance faradaic and capacitive currents established at the interface with water. This work opens up interesting opportunities towards applications in both photo-electrochemistry and biology.

Hybrid interfaces between living cells and nano/microstructured scaffolds have huge application potential in biotechnology, spanning from regenerative medicine and stem cell therapies to localized drug delivery and from biosensing and tissue engineering to neural computing. However, 3D architectures based on semiconducting polymers, endowed with responsivity to visible light, have never been considered. Here, we apply for the first time a push-coating technique to realize high aspect ratio polymeric pillars, based on polythiophene, showing optimal biocompatibility and allowing for the realization of soft, 3D cell cultures of both primary neurons and cell line models. HEK-293 cells cultured on top of polymer pillars display a remarkable change in the cell morphology and a sizable enhancement of the membrane capacitance due to the cell membrane thinning in correspondence to the pillars' top surface, without negatively affecting cell proliferation. Electrophysiology properties and synapse number of primary neurons are also very well preserved. In perspective, high aspect ratio semiconducting polymer pillars may find interesting applications as soft, photoactive elements for cell activity sensing and modulation. Hybrid interfaces between living cells and nano/microstructured scaffolds have huge application potential in biotechnology, spanning from regenerative medicine and stem cell therapies to localized drug delivery and from biosensing and tissue engineering to neural computing. However, 3D architectures based on semiconducting polymers, endowed with responsivity to visible light, have never been considered. Here, we apply for the first time a push-coating technique to realize high aspect ratio polymeric pillars, based on polythiophene, showing optimal biocompatibility and allowing for the realization of soft, 3D cell cultures of both primary neurons and cell line models. HEK-293 cells cultured on top of polymer pillars display a remarkable change in the cell morphology and a sizable enhancement of the membrane capacitance due to the cell membrane thinning in correspondence to the pillars' top surface, without negatively affecting cell proliferation. Electrophysiology properties and synapse number of primary neurons are also very well preserved. In perspective, high aspect ratio semiconducting polymer pillars may find interesting applications as soft, photoactive elements for cell activity sensing and modulation.

The possibility to optically excite the electrical activity of living cells by using exogenous absorbers is gaining more and more interest in the neuroscience and biotechnology community. Conjugated polymers, inherently sensitive to visible light, were recently proposed as candidates to this goal. To date, however, only one polymer type, namely regio-regular poly-3-hexylthiophene, has been tested as the active material. In this work four different conjugated polymers, regarded as prototypes of their category, are investigated as photoactive bio-interfaces. The selected materials have different absorption spectra, morphology, light emission efficiency and charge transport properties. We analyze their key-enabling properties, such as electrochemical stability, surface morphology, wettability, sterilization compatibility, interaction with protein adhesion layers and toxicity, throughout all the necessary steps for the realization of an efficient bio-optical interface. We demonstrate that all considered polymers are characterized by good biocompatibility and cell seeding properties, and can optimally sustain thermal sterilization. Conversely, electrochemical stability and cell photostimulation efficacy can vary a lot among different materials, and should be carefully evaluated case by case. Reported results represent the starting point for the implementation of bio-polymer interfaces sensitive to different colors and, in perspective, for the realization of a three-chromatic artificial visual prosthesis.

Optical modulation of living cells activity by light-absorbing exogenous materials is gaining increasing interest, due to the possibility both to achieve high spatial and temporal resolution with a minimally invasive and reversible technique and to avoid the need of viral transfection with light-sensitive proteins. In this context, conjugated polymers represent ideal candidates for photo-transduction, due to their excellent optoelectronic and biocompatibility properties. In this work, we demonstrate that organic polymer nanoparticles, based on poly(3-hexylthiophene) conjugated polymer, establish a functional interaction with an in vitro cell model (Human Embryonic Kidney cells, HEK-293). They display photocatalytic activity in aqueous environment and, once internalized within the cell cytosol, efficiently generate reactive oxygen species (ROS) upon visible light excitation, without affecting cell viability. Interestingly, light-activated ROS generation deterministically triggers modulation of intracellular calcium ion flux, successfully controlled at the single cell level. In perspective, the capability of polymer NPs to produce ROS and to modulate Ca2+ dynamics by illumination on-demand, at non-toxic levels, may open the path to the study of biological processes with a gene-less approach and unprecedented spatio-temporal resolution, as well as to the development of new biotechnology tools for cell optical modulation.

We report that nanoparticles prepared from appropriately functionalized polythiophenes once administered to live cells can acquire phototransduction properties under illumination, becoming photoactive sites able to absorb visible light and convert it to an electrical signal through cell membrane polarization. Amine-reactive fluorescent nanoparticles with pendant N-succinimidyl-ester groups (NPs-NHS) are prepared from polythiophenes alternating unsubstituted and 3-(2,5-dioxopyrrolidin-1-yl-8-octanoate)-substituted thiophenes by a nanoprecipitation method. By 1H NMR of nanoparticles prepared using THF-d8/D2O (solvent/non-solvent) we demonstrate that the hydrolysis of the N-succinimidyl-ester group to free N-hydroxysuccinimide takes place slowly over several hours. NPs-NHS reactivity towards primary amine groups is tested towards the NH2 of d- and l-enantiomers of tryptophan. We show that the formation of a tryptophan-nanoparticle amidic bond creates a chiral shell displaying opposite CD signals for the nanoparticles bound to d or l enantiomers. The interaction of NPs-NHS with live HEK-293 cells is monitored via LSCM. We show that the NPs-NHS are not internalized but remain docked on the cell membrane. We assume that this is mainly the result of the reaction of the NHS groups in the external layer with NH2 groups present in cell membrane proteins, although the contribution of alternative mechanisms cannot be excluded. To support this assumption LSCM experiments show that nanoparticles of comparable size obtained from poly(3-hexylthiophene), NPs-P3HT, are rapidly internalized by live HEK-293 cells. Finally, using the whole-cell current clamp technique under light illumination we demonstrate that NPs-NHS can polarize the cell membrane upon light irradiation while NPs-P3HT cannot.

Hybrid interfaces between organic semiconductors and living tissues represent a new tool for in-vitro and in-vivo applications. In particular, conjugated polymers display several optimal properties as substrates for biological systems, such as good biocompatibility, excellent mechanical properties, cheap and easy processing technology, and possibility of deposition on light, thin and flexible substrates. These materials have been employed for cellular interfaces like neural probes, transistors for excitation and recording of neural activity, biosensors and actuators for drug release. Recent experiments have also demonstrated the possibility to use conjugated polymers for all-optical modulation of the electrical activity of cells. Several in-vitro study cases have been reported, including primary neuronal networks, astrocytes and secondary line cells. Moreover, signal photo-transduction mediated by organic polymers has been shown to restore light sensitivity in degenerated retinas, suggesting that these devices may be used for artificial retinal prosthesis in the future. All in all, light sensitive conjugated polymers represent a new approach for optical modulation of cellular activity. In this work, all the steps required to fabricate a bio-polymer interface for optical excitation of living cells are described. The function of the active interface is to transduce the light stimulus into a modulation of the cell membrane potential. As a study case, useful for in-vitro studies, a polythiophene thin film is used as the functional, light absorbing layer, and Human Embryonic Kidney (HEK-293) cells are employed as the biological component of the interface. Practical examples of successful control of the cell membrane potential upon stimulation with light pulses of different duration are provided. In particular, it is shown that both depolarizing and hyperpolarizing effects on the cell membrane can be achieved depending on the duration of the light stimulus. The reported protocol is of general validity and can be straightforwardly extended to other biological preparations.

Purpose To speed up the osteointegration process, surface-treated titanium has been widely used in dental and orthopedic applications. The present work describes a new silicon-based anodic spark deposition (ASD) treatment and investigates the properties of the surfaces obtained, focusing on their capability to modulate the osteogenic differentiation potential of adult mesenchymal stem cells (MSCs). Methods The surfaces examined were obtained from commercially pure grade 2 titanium by a single-step ASD (SUM) eventually followed by a thermal treatment in alkali solution (SUMNa), while acid-etched titanium (AE; NextMaterials s.r.l.) was selected as a control. Their morphology, elemental composition, crystallographic structure of the TiO2 layer, wettability and topography were evaluated by scanning electron microscopy, energy-dispersive X-ray spectroscopy, thin-film X-ray diffraction, contact angle measurements and laser profilometry, respectively. MSCs' response to surface properties was assessed by examining cell morphology and viability by scanning electron microscopy and Alamar Blue™ assay, while their osteogenic differentiation potential was investigated by evaluating the levels of the enzyme alkaline phosphatase (ALP) and the degree of calcium accumulation by Alizarin Red-S (AR-S) staining. Results The proposed ASD treatment has allowed the obtaining of surfaces with round-shaped micrometric pores, enriched in calcium, phosphorus and silicon and significantly more wettable than controls; furthermore, the treatment has been shown to promote MSC proliferation and the degree of in vitro mineralization. Conclusions The described ASD treatment may be an effective technique to modify the surface cues of titanium implants, aiming at enhancing the conveying of osteoprogenitor cells and their functional differentiation in bone cells.

Hybrid interfaces between organic semiconductors and living tissues represent a new tool for in-vitro and in-vivo applications, bearing a huge potential, from basic researches to clinical applications. In particular, light sensitive conjugated polymers can be exploited as a new approach for optical modulation of cellular activity. In this work we focus on light-induced changes in the membrane potential of Human Embryonic Kidney (HEK-293) cells grown on top of a poly(3-hexylthiophene) (P3HT) thin film. On top of a capacitive charging of the polymer interface, we identify and fully characterize two concomitant mechanisms, leading to membrane depolarization and hyperpolarisation, both mediated by a thermal effect. Our results can be usefully exploited in the creation of a new platform for light-controlled cell manipulation, with possible applications in neuroscience and medicine.