Modification of Living Diatom, Thalassiosira weissflogii, with a Calcium Precursor through a Calcium Uptake Mechanism: A Next Generation Biomaterial for Advanced Delivery Systems.

The diatom's frustule, characterized by its rugged and porous exterior, exhibits a remarkable biomimetic morphology attributable to its highly ordered pores, extensive surface area, and unique architecture. Despite these advantages, the toxicity and nonbiodegradable nature of silica-based organisms pose a significant challenge when attempting to utilize these organisms as nanotopographically functionalized microparticles in the realm of biomedicine. In this study, we addressed this limitation by modulating the chemical composition of diatom microparticles by modulating the active silica metabolic uptake mechanism while maintaining their intricate three-dimensional architecture through calcium incorporation into living diatoms. Here, the diatom Thalassiosira weissflogii was chemically modified to replace its silica composition with a biodegradable calcium template, while simultaneously preserving the unique three-dimensional (3D) frustule structure with hierarchical patterns of pores and nanoscale architectural features, which was evident by the deposition of calcium as calcium carbonate. Calcium hydroxide is incorporated into the exoskeleton through the active mechanism of calcium uptake via a carbon-concentrating mechanism, without altering the microstructure. Our findings suggest that calcium-modified diatoms hold potential as a nature-inspired delivery system for immunotherapy through antibody-specific binding.

Flexible, Transparent, and Cytocompatible Nanostructured Indium Tin Oxide Thin Films for Bio-optoelectronic Applications.

Electrical stimulation has been used successfully for several decades for the treatment of neurodegenerative disorders, including motor disorders, pain, and psychiatric disorders. These technologies typically rely on the modulation of neural activity through the focused delivery of electrical pulses. Recent research, however, has shown that electrically triggered neuromodulation can be further enhanced when coupled with optical stimulation, an approach that can benefit from the development of novel electrode materials that combine transparency with excellent electrochemical and biological performance. In this study, we describe an electrochemically modified, nanostructured indium tin oxide/poly(ethylene terephthalate) (ITO/PET) surface as a flexible, transparent, and cytocompatible electrode material. Electrochemical oxidation and reduction of ITO/PET electrodes in the presence of an ionic liquid based on d-glucopyranoside and bistriflamide units were performed, and the electrochemical behavior, conductivity, capacitance, charge transport processes, surface morphology, optical properties, and cytocompatibility were assessed in vitro. It has been shown that under selected conditions, electrochemically modified ITO/PET films remained transparent and highly conductive and were able to enhance neural cell survival and neurite outgrowth. Consequently, electrochemical modification of ITO/PET electrodes in the presence of an ionic liquid is introduced as an effective approach for tailoring the properties of ITO for advanced bio-optoelectronic applications.

Physicochemical cues are not potent regulators of human dermal fibroblast trans-differentiation.

Due to their inherent plasticity, dermal fibroblasts hold great promise in regenerative medicine. Although biological signals have been well-established as potent regulators of dermal fibroblast function, it is still unclear whether physiochemical cues can induce dermal fibroblast trans-differentiation. Herein, we evaluated the combined effect of surface topography, substrate rigidity, collagen type I coating and macromolecular crowding in human dermal fibroblast cultures. Our data indicate that tissue culture plastic and collagen type I coating increased cell proliferation and metabolic activity. None of the assessed in vitro microenvironment modulators affected cell viability. Anisotropic surface topography induced bidirectional cell morphology, especially on more rigid (1,000 kPa and 130 kPa) substrates. Macromolecular crowding increased various collagen types, but not fibronectin, deposition. Macromolecular crowding induced globular extracellular matrix deposition, independently of the properties of the substrate. At day 14 (longest time point assessed), macromolecular crowding downregulated tenascin C (in 9 out of the 14 groups), aggrecan (in 13 out of the 14 groups), osteonectin (in 13 out of the 14 groups), and collagen type I (in all groups). Overall, our data suggest that physicochemical cues (such surface topography, substrate rigidity, collagen coating and macromolecular crowding) are not as potent as biological signals in inducing dermal fibroblast trans-differentiation.

The synergistic effect of physicochemical in vitro microenvironment modulators in human bone marrow stem cell cultures.

Modern bioengineering utilises biomimetic cell culture approaches to control cell fate during in vitro expansion. In this spirit, herein we assessed the influence of bidirectional surface topography, substrate rigidity, collagen type I coating and macromolecular crowding (MMC) in human bone marrow stem cell cultures. In the absence of MMC, surface topography was a strong modulator of cell morphology. MMC significantly increased extracellular matrix deposition, albeit in a globular manner, independently of the surface topography, substrate rigidity and collagen type I coating. Collagen type I coating significantly increased cell metabolic activity and none of the assessed parameters affected cell viability. At day 14, in the absence of MMC, none of the assessed genes was affected by surface topography, substrate rigidity and collagen type I coating, whilst in the presence of MMC, in general, collagen type I α1 chain, tenascin C, osteonectin, bone sialoprotein, aggrecan, cartilage oligomeric protein and runt-related transcription factor were downregulated. Interestingly, in the presence of the MMC, the 1000 kPa grooved substrate without collagen type I coating upregulated aggrecan, cartilage oligomeric protein, scleraxis homolog A, tenomodulin and thrombospondin 4, indicative of tenogenic differentiation. This study further supports the notion for multifactorial bioengineering to control cell fate in culture.

A Self-Powered Piezo-Bioelectric Device Regulates Tendon Repair-Associated Signaling Pathways through Modulation of Mechanosensitive Ion Channels.

Tendon disease constitutes an unmet clinical need and remains a critical challenge in the field of orthopaedic surgery. Innovative solutions are required to overcome the limitations of current tendon grafting approaches, and bioelectronic therapies show promise in treating musculoskeletal diseases, accelerating functional recovery through the activation of tissue regeneration-specific signaling pathways. Self-powered bioelectronic devices, particularly piezoelectric materials, represent a paradigm shift in biomedicine, negating the need for battery or external powering and complementing existing mechanotherapy to accelerate the repair processes. Here, the dynamic response of tendon cells to a piezoelectric collagen-analogue scaffold comprised of aligned nanoscale fibers made of the ferroelectric material poly(vinylidene fluoride-co-trifluoroethylene) is shown. It is demonstrated that motion-powered electromechanical stimulation of tendon tissue through piezo-bioelectric device results in ion channel modulation in vitro and regulates specific tissue regeneration signaling pathways. Finally, the potential of the piezo-bioelectronic device in modulating the progression of tendinopathy-associated processes in vivo, using a rat Achilles acute injury model is shown. This study indicates that electromechanical stimulation regulates mechanosensitive ion channel sensitivity and promotes tendon-specific over non-tenogenic tissue repair processes.

In vitro analysis of a physiological strain sensor formulated from a PEDOT:PSS functionalized carbon nanotube-poly(glycerol sebacate urethane) composite.

Biodegradable strain sensors able to undergo controlled degradation following implantation have recently received significant interest as novel approaches to detect pathological tissue swelling or non-physiological stresses. In this study, the physicomechanical, electrochemical and active pressure sensing behavior of an electrically conductive and biodegradable poly(glycerol sebacate urethane) (PGSU) composite, reinforced with poly(3,4-ethylenedioxythiophene) polystyrene sulfonate (PEDOT:PSS) functionalized carbon nanotubes (CNTs), was evaluated in vitro. Analysis of these PGSU-CNTs composites demonstrated that the incorporation of functionalized CNTs into a biodegradable elastomer resulted in enhanced mechanical strength, conductivity and tailored matrix biodegradation. PGSU-CNT composites were subsequently formulated into flexible and active pressure sensors which demonstrated optimal sensitivity to applied 1% uniaxial tensile strains. Finally, cytocompatibility analysis a with primary neural culture confirmed that PGSU-CNT composites exhibited low cytotoxicity, and supported neuron adhesion, viability, and proliferation in vitro.

Electrical percolation in extrinsically conducting, poly(ε-decalactone) composite neural interface materials.

By providing a bidirectional communication channel between neural tissues and a biomedical device, it is envisaged that neural interfaces will be fundamental in the future diagnosis and treatment of neurological disorders. Due to the mechanical mismatch between neural tissue and metallic neural electrodes, soft electrically conducting materials are of great benefit in promoting chronic device functionality. In this study, carbon nanotubes (CNT), silver nanowires (AgNW) and poly(hydroxymethyl 3,4-ethylenedioxythiophene) microspheres (MSP) were employed as conducting fillers within a poly(ε-decalactone) (EDL) matrix, to form a soft and electrically conducting composite. The effect of a filler type on the electrical percolation threshold, and composite biocompatibility was investigated in vitro. EDL-based composites exhibited favourable electrochemical characteristics: EDL/CNT-the lowest film resistance (1.2 ± 0.3 kΩ), EDL/AgNW-the highest charge storage capacity (10.7 ± 0.3 mC cm- 2), and EDL/MSP-the highest interphase capacitance (1478.4 ± 92.4 µF cm-2). All investigated composite surfaces were found to be biocompatible, and to reduce the presence of reactive astrocytes relative to control electrodes. The results of this work clearly demonstrated the ability of high aspect ratio structures to form an extended percolation network within a polyester matrix, resulting in the formulation of composites with advantageous mechanical, electrochemical and biocompatibility properties.

Nanovibrational Stimulation of Mesenchymal Stem Cells Induces Therapeutic Reactive Oxygen Species and Inflammation for Three-Dimensional Bone Tissue Engineering.

There is a pressing clinical need to develop cell-based bone therapies due to a lack of viable, autologous bone grafts and a growing demand for bone grafts in musculoskeletal surgery. Such therapies can be tissue engineered and cellular, such as osteoblasts, combined with a material scaffold. Because mesenchymal stem cells (MSCs) are both available and fast growing compared to mature osteoblasts, therapies that utilize these progenitor cells are particularly promising. We have developed a nanovibrational bioreactor that can convert MSCs into bone-forming osteoblasts in two- and three-dimensional, but the mechanisms involved in this osteoinduction process remain unclear. Here, to elucidate this mechanism, we use increasing vibrational amplitude, from 30 nm (N30) to 90 nm (N90) amplitudes at 1000 Hz and assess MSC metabolite, gene, and protein changes. These approaches reveal that dose-dependent changes occur in MSCs' responses to increased vibrational amplitude, particularly in adhesion and mechanosensitive ion channel expression and that energetic metabolic pathways are activated, leading to low-level reactive oxygen species (ROS) production and to low-level inflammation as well as to ROS- and inflammation-balancing pathways. These events are analogous to those that occur in the natural bone-healing processes. We have also developed a tissue engineered MSC-laden scaffold designed using cells' mechanical memory, driven by the stronger N90 stimulation. These mechanistic insights and cell-scaffold design are underpinned by a process that is free of inductive chemicals.

In vitro attenuation of astrocyte activation and neuroinflammation through ibuprofen-doping of poly(3,4-ethylenedioxypyrrole) formulations.

Neuroinflammation is often associated with poor functional recovery and may contribute to or initiate the development of severe neurological disorders, such as epilepsy, Parkinson's disease or Alzheimer's disease. Ibuprofen (IBU), being one of the most commonly used non-steroidal anti-inflammatory drugs, is known to possess neuroprotective activity and serve as a promising therapeutic for the treatment of neuroinflammation. In this study, the potential of an IBU-loaded poly(3,4-ethylenedioxypyrrole) (PEDOP) matrix has been assessed as a neural interface material with an aim to control astrocyte activation and suppress neuroinflammation in vitro. Three types of drug immobilization protocols were investigated, leading to the fabrication of IBU-loaded PEDOP matrices exhibiting a broad spectrum of electrical characteristics, drug release profiles, as well as biological responses. Among all investigated PEDOP formulations, PEDOP matrices formed through a three-step immobilization protocol exhibited the highest charge storage capacity (30 ± 1 mC/cm2) as well as a double layer capacitance of 645.0 ± 51.1 µF, associated with a relatively enlarged surface area. Demonstrating a total drug loading capacity of 150 µg/ml and a release rate constant of 0.15 1/h, this coating formulation may be employed as a safe electrical conducting drug eluting system.

Laser-Induced Periodic Surface Structure Enhances Neuroelectrode Charge Transfer Capabilities and Modulates Astrocyte Function.

The brain machine interface (BMI) describes a group of technologies capable of communicating with excitable nervous tissue within the central nervous system (CNS). BMIs have seen major advances in recent years, but these advances have been impeded because of a temporal deterioration in the signal to noise ratio of recording electrodes following insertion into the CNS. This deterioration has been attributed to an intrinsic host tissue response, namely, reactive gliosis, which involves a complex series of immune mediators, resulting in implant encapsulation via the synthesis of pro-inflammatory signaling molecules and the recruitment of glial cells. There is a clinical need to reduce tissue encapsulation in situ and improve long-term neuroelectrode functionality. Physical modification of the electrode surface at the nanoscale could satisfy these requirements by integrating electrochemical and topographical signals to modulate neural cell behavior. In this study, commercially available platinum iridium (Pt/Ir) microelectrode probes were nanotopographically functionalized using femto/picosecond laser processing to generate laser-induced periodic surface structures (LIPSS). Three different topographies and their physical properties were assessed by scanning electron microscopy and atomic force microscopy. The electrochemical properties of these interfaces were investigated using electrochemical impedance spectroscopy and cyclic voltammetry. The in vitro response of mixed cortical cultures (embryonic rat E14/E17) was subsequently assessed by confocal microscopy, ELISA, and multiplex protein array analysis. Overall LIPSS features improved the electrochemical properties of the electrodes, promoted cell alignment, and modulated the expression of multiple ion channels involved in key neuronal functions.

PEDOT:PSS interfaces stabilised using a PEGylated crosslinker yield improved conductivity and biocompatibility.

The rapidly expanding fields of bioelectronics, and biological interfaces with electronic sensors and stimulators, are placing an increasing demand on candidate materials to serve as robust surfaces that are both biocompatible, stable and electroconductive. Amongst conductive polymers, poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) is a promising material in biomedical research due to its appropriate stability and high conductivity, however its intrinsic solubility requires a crosslinking process that can limit its conductivity and biocompatibility. Poly(ethylene glycol) is known to be a suitably anti-immunogenic moiety and its derivatives have been widely used for biomedical applications. In this study we investigate the application of poly(ethylene glycol)diglycidyl ether (PEGDE) as an effective crosslinker and conductive filler for PEDOT:PSS. From our interpretation of XPS analysis we hypothesise that the crosslinking reaction is occurring via the epoxy ring of PEGDE interacting with the sulfonic groups of excel PSS chains, which reaches a saturation at 3 w/v% PEGDE concentration. PEGDE crosslinked films did not disperse in aqueous environments, had enhanced electrical conductivity and imparted a significant degree of hydrophilicity to PEDOT:PSS films. This hydrophilicity and the presence of biocompatible PEGDE led to good cell viability and a significantly increased degree of cell spreading on PEDOT:PSS films. In comparison to widely reported chemical crosslinking via glycidoxy propyltrimethoxysilane (GOPS), this original crosslinking yields a highly hydrophilic 2D film substrate with increased electroconductive and biocompatibility properties, resulting in a next-generation formulation for bioengineering applications.

Boron Nitride Nanotube Addition Enhances the Crystallinity and Cytocompatibility of PVDF-TrFE.

Analysis of the cellular response to piezoelectric materials has been driven by the discovery that many tissue components exhibit piezoelectric behavior ex vivo. In particular, polyvinylidene fluoride and the trifluoroethylene co-polymer (PVDF-TrFE) have been identified as promising piezo and ferroelectric materials with applications in energy harvesting and biosensor devices. Critically, the modulation of the structural and crystalline properties of PVDF-TrFE through annealing processes and the addition of particulate or fibrous fillers has been shown to modulate significantly the materials electromechanical properties. In this study, a PVDF-TrFE/boron-nitride nanotube composite was evaluated by modulated differential scanning calorimetry to assess the effects of boron nitride nanotube addition and thermal annealing on the composite structure and crystal behavior. An increased beta crystal formation [f(ß) = 0.71] was observed following PVDF-TrFE annealing at the first crystallization temperature of 120°C. In addition, the inclusion of boron nitride nanotubes significantly increased the crystal formation behavior [f(ß) = 0.76] and the mechanical properties of the material. Finally, it was observed that BNNT incorporation enhance the adherence and proliferation of human tenocyte cells in vitro.

The Synergistic Effects of Gold Particles and Dexamethasone on the Electrochemical and Biological Performance of PEDOT Neural Interfaces.

Although neural devices have shown efficacy in the treatment of neurodegenerative diseases, their functionality is limited by the inflammatory state and glial scar formation associated with chronic implantation. The aim of this study was to investigate neural electrode performance following functionalization with an anti-inflammatory coating derived from a conducting polymer poly(3,4-ethylenedioxythiophene) (PEDOT) matrix doped with dexamethasone (Dex) and decorated with Au particles. Pristine PEDOT, PEDOT-Dex and their gold-decorated analogues (PEDOT/Au and PEDOT-Dex/Au) were formulated by electrochemical deposition and characterized with respect to electrode electrochemical properties, surface morphology and biocompatibility towards primary neural cells. Through a process of gold deposition, it was possible to eliminate the initial burst release observed in PEDOT-Dex and maintain a stable, stepwise increase in Dex elution over 7 days. The released amounts of Dex exceeded the concentrations considered as therapeutic for both PEDOT-Dex and PEDOT-Dex/Au. The results clearly indicated that the presence of either Dex or Au particles facilitated the outgrowth of neurites. Finally, it was shown that the application of composite materials, such as PEDOT-Dex/Au, is an efficient way to improve the efficacy of neural interfaces in vitro.

Analysis of a poly(ε-decalactone)/silver nanowire composite as an electrically conducting neural interface biomaterial.

BACKGROUND: Advancement in polymer technologies, facilitated predominantly through chemical engineering approaches or through the identification and utilization of novel renewable resources, has been a steady focus of biomaterials research for the past 50 years. Aliphatic polyesters have been exploited in numerous biomedical applications including the formulation of soft-tissue sutures, bone fixation devices, cardiovascular stents etc. Biomimetic 'soft' polymer formulations are of interest in the design of biological interfaces and specifically, in the development of implantable neuroelectrode systems intended to interface with neural tissues. Critically, soft polymer formulations have been shown to address the challenges associated with the disregulation of mechanotransductive processes and micro-motion induced inflammation at the electrode/tissue interface. In this study, a polyester-based poly(ε-decalactone)/silver nanowire (EDL:Ag) composite was investigated as a novel electrically active biomaterial with neural applications.Neural interfaces were formulated through spin coating of a polymer/nanowire formulation onto the surface of a Pt electrode to form a biocompatible EDL matrix supported by a percolated network of silver nanowires. As-formed EDL:Ag composites were characterized by means of infrared spectroscopy, scanning electron microscopy and electrochemical methods, with their cytocompatibility assessed using primary cultures of a mixed neural population obtained from the ventral mesencephalon of Sprague-Dawley rat embryos. RESULTS: Electrochemical characterization of various EDL:Ag composites indicated EDL:Ag 10:1 as the most favourable formulation, exhibiting high charge storage capacity (8.7 ± 1.0 mC/cm2), charge injection capacity (84.3 ± 1.4 µC/cm2) and low impedance at 1 kHz (194 ± 28 Ω), outperforming both pristine EDL and bare Pt electrodes. The in vitro biological evaluation showed that EDL:Ag supported significant neuron viability in culture and to promote neurite outgrowth, which had the average length of 2300 ± 6 µm following 14 days in culture, 60% longer than pristine EDL and 120% longer than bare Pt control substrates. CONCLUSIONS: EDL:Ag nanocomposites are shown to serve as robust neural interface materials, possessing favourable electrochemical characteristics together with high neural cytocompatibility.

Chirality-sorted carbon nanotube films as high capacity electrode materials.

Carbon nanomaterials show great promise for a wide range of applications due to their excellent physicochemical and electrical properties. Since their discovery, the state-of-the-art has expanded the scope of their application from scientific curiosity to impactful solutions. Due to their tunability, carbon nanomaterials can be processed into a wide range of formulations and significant scope exists to couple carbon structures to electronic and electrochemical applications. In this paper, the electrochemical performance of various types of CNT films, which differ by the number of walls, diameter, chirality and surface chemistry is presented. Especially, chirality-sorted (6,5)- and (7,6)-based CNT films are shown to possess a high charge storage capacity (up to 621.91 mC cm-2), areal capacitance (262 mF cm-2), significantly increased effective surface area and advantageous charge/discharge characteristics without addition of any external species, and outperform many other high capacity materials reported in the literature. The results suggest that the control over the CNT structure can lead to the manufacture of macroscopic CNT devices precisely tailored for a wide range of applications, with the focus on energy storage devices and supercapacitors. The sorted CNT macroassemblies show great potential for energy storage technologies to come from R&D laboratories into real life.

Nanocellulose reinforced gellan-gum hydrogels as potential biological substitutes for annulus fibrosus tissue regeneration.

Intervertebral disc (IVD) degeneration is associated with both structural damage and aging related degeneration. Annulus fibrosus (AF) defects such as annular tears, herniation and discectomy require novel tissue engineering strategies to functionally repair AF tissue. An ideal construct will repair the AF by providing physical and biological support, facilitating regeneration. The presented strategy herein proposes a gellan gum-based construct reinforced with cellulose nanocrystals (nCell) as a biological self-gelling AF substitute. Nanocomposite hydrogels were fabricated and characterized with respect to hydrogel swelling capacity, degradation rate in vitro and mechanical properties. Rheological evaluation on the nanocomposites demonstrated the GGMA reinforcement with nCell promoted matrix entanglement with higher scaffold stiffness observed upon ionic crosslinking. Compressive mechanical tests demonstrated compressive modulus values close to those of the human AF tissue. Furthermore, cell culture studies with encapsulated bovine AF cells indicated that nanocomposite constructs promoted cell viability and a physiologically relevant cell morphology for up to fourteen days in vitro.

The Functional Response of Mesenchymal Stem Cells to Electron-Beam Patterned Elastomeric Surfaces Presenting Micrometer to Nanoscale Heterogeneous Rigidity.

Cells directly probe and respond to the physicomechanical properties of their extracellular environment, a dynamic process which has been shown to play a key role in regulating both cellular adhesive processes and differential cellular function. Recent studies indicate that stem cells show lineage-specific differentiation when cultured on substrates approximating the stiffness profiles of specific tissues. Although tissues are associated with a range of Young's modulus values for bulk rigidity, at the subcellular level, tissues are comprised of heterogeneous distributions of rigidity. Lithographic processes have been widely explored in cell biology for the generation of analytical substrates to probe cellular physicomechanical responses. In this work, it is shown for the first time that that direct-write e-beam exposure can significantly alter the rigidity of elastomeric poly(dimethylsiloxane) substrates and a new class of 2D elastomeric substrates with controlled patterned rigidity ranging from the micrometer to the nanoscale is described. The mechanoresponse of human mesenchymal stem cells to e-beam patterned substrates was subsequently probed in vitro and significant modulation of focal adhesion formation and osteochondral lineage commitment was observed as a function of both feature diameter and rigidity, establishing the groundwork for a new generation of biomimetic material interfaces.

Stimulation of 3D osteogenesis by mesenchymal stem cells using a nanovibrational bioreactor.

Bone grafts are one of the most commonly transplanted tissues. However, autologous grafts are in short supply, and can be associated with pain and donor-site morbidity. The creation of tissue-engineered bone grafts could help to fulfil clinical demand and provide a crucial resource for drug screening. Here, we show that vibrations of nanoscale amplitude provided by a newly developed bioreactor can differentiate a potential autologous cell source, mesenchymal stem cells (MSCs), into mineralized tissue in 3D. We demonstrate that nanoscale mechanotransduction can stimulate osteogenesis independently of other environmental factors, such as matrix rigidity. We show this by generating mineralized matrix from MSCs seeded in collagen gels with stiffness an order of magnitude below the stiffness of gels needed to induce bone formation in vitro. Our approach is scalable and can be compatible with 3D scaffolds.

Publisher Correction: Stimulation of 3D osteogenesis by mesenchymal stem cells using a nanovibrational bioreactor.

In the version of this Article originally published, in Fig. 4f, the asterisk was missing; in Fig. 6a-c, the labels 'Wnt/ß-catenin signalling', 'Wnt/Ca+ pathway' and 'ERK' and their associated lines/arrows were missing; and in Fig. 6d and in the sentence beginning "In MSCs that were...", 'myosin' and 'nanostimulated', respectively, were spelt incorrectly. These errors have now been corrected in all versions of the Article.