A Dinuclear Osmium(II) Complex Near-Infrared Nanoscopy Probe for Nuclear DNA.

With the aim of developing photostable near-infrared cell imaging probes, a convenient route to the synthesis of heteroleptic OsII complexes containing the Os(TAP)2 fragment is reported. This method was used to synthesize the dinuclear OsII complex, [{Os(TAP)2}2tpphz]4+ (where tpphz = tetrapyrido[3,2-a:2',3'-c:3″,2''-h:2‴,3'''-j]phenazine and TAP = 1,4,5,8- tetraazaphenanthrene). Using a combination of resonance Raman and time-resolved absorption spectroscopy, as well as computational studies, the excited state dynamics of the new complex were dissected. These studies revealed that, although the complex has several close lying excited states, its near-infrared, NIR, emission (λmax = 780 nm) is due to a low-lying Os → TAP based 3MCLT state. Cell-based studies revealed that unlike its RuII analogue, the new complex is neither cytotoxic nor photocytotoxic. However, as it is highly photostable as well as live-cell permeant and displays NIR luminescence within the biological optical window, its properties make it an ideal probe for optical microscopy, demonstrated by its use as a super-resolution NIR STED probe for nuclear DNA.

A Tuneable, Photocurable, Poly(Caprolactone)-Based Resin for Tissue Engineering-Synthesis, Characterisation and Use in Stereolithography.

Stereolithography is a useful additive manufacturing technique for the production of scaffolds for tissue engineering. Here we present a tuneable, easy-to-manufacture, photocurable resin for use in stereolithography, based on the widely used biomaterial, poly(caprolactone) (PCL). PCL triol was methacrylated to varying degrees and mixed with photoinitiator to produce a photocurable prepolymer resin, which cured under UV light to produce a cytocompatible material. This study demonstrates that poly(caprolactone) methacrylate (PCLMA) can be produced with a range of mechanical properties and degradation rates. By increasing the degree of methacrylation (DM) of the prepolymer, the Young's modulus of the crosslinked PCLMA could be varied from 0.12-3.51 MPa. The accelerated degradation rate was also reduced from complete degradation in 17 days to non-significant degradation in 21 days. The additive manufacturing capabilities of the resin were demonstrated by the production of a variety of different 3D structures using micro-stereolithography. Here, ß-carotene was used as a novel, cytocompatible photoabsorber and enabled the production of complex geometries by giving control over cure depth. The PCLMA presented here offers an attractive, tuneable biomaterial for the production of tissue engineering scaffolds for a wide range of applications.

A Dinuclear Ruthenium(II) Complex Excited by Near-Infrared Light through Two-Photon Absorption Induces Phototoxicity Deep within Hypoxic Regions of Melanoma Cancer Spheroids.

The dinuclear photo-oxidizing RuII complex [{Ru(TAP2)}2(tpphz)]4+ (TAP = 1,4,5,8- tetraazaphenanthrene, tpphz = tetrapyrido[3,2-a:2',3'-c:3″,2''-h:2‴,3'''-j]phenazine), 14+, is readily taken up by live cells localizing in mitochondria and nuclei. In this study, the two-photon absorption cross section of 14+ is quantified and its use as a two-photon absorbing phototherapeutic is reported. It was confirmed that the complex is readily photoexcited using near-infrared, NIR, and light through two-photon absorption, TPA. In 2-D cell cultures, irradiation with NIR light at low power results in precisely focused phototoxicity effects in which human melanoma cells were killed after 5 min of light exposure. Similar experiments were then carried out in human cancer spheroids that provide a realistic tumor model for the development of therapeutics and phototherapeutics. Using the characteristic emission of the complex as a probe, its uptake into 280 µm spheroids was investigated and confirmed that the spheroid takes up the complex. Notably TPA excitation results in more intense luminescence being observed throughout the depth of the spheroids, although emission intensity still drops off toward the necrotic core. As 14+ can directly photo-oxidize DNA without the mediation of singlet oxygen or other reactive oxygen species, phototoxicity within the deeper, hypoxic layers of the spheroids was also investigated. To quantify the penetration of these phototoxic effects, 14+ was photoexcited through TPA at a power of 60 mW, which was progressively focused in 10 µm steps throughout the entire z-axis of individual spheroids. These experiments revealed that, in irradiated spheroids treated with 14+, acute and rapid photoinduced cell death was observed throughout their depth, including the hypoxic region.

Biomimetic surface delivery of NGF and BDNF to enhance neurite outgrowth.

Treatment for peripheral nerve injuries includes the use of autografts and nerve guide conduits (NGCs). However, outcomes are limited, and full recovery is rarely achieved. The use of nerve scaffolds as a platform to surface immobilize neurotrophic factors and deliver locally is a promising approach to support neurite and nerve outgrowth after injury. We report on a bioactive surface using functional amine groups, to which heparin binds electrostatically. X-ray photoelectron spectroscopy analysis was used to characterize the presence of nitrogen and sulfur. Nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) were bound by electrostatic interaction to heparin, and the release profile evaluated by enzyme-linked immunosorbent assay, which showed that ca. 1% of NGF was released from each of the bioactive surface within 7 days. Furthermore, each surface showed a maximum release of 97% of BDNF. Neurotrophin release on neurite outgrowth was evaluated by primary dorsal root ganglion with a maximum neurite growth response in vitro of 1,075 µm detected for surfaces immobilized with NGF at 1 ng/ml. In summary, the study reports on the design and construction of a biomimetic platform to deliver NGF and BDNF using physiologically low concentrations of neurotrophin. The platform is directly applicable and scalable for improving the regenerative ability of existing NGCs and scaffolds.

Decellularisation and histological characterisation of porcine peripheral nerves.

Peripheral nerve injuries affect a large proportion of the global population, often causing significant morbidity and loss of function. Current treatment strategies include the use of implantable nerve guide conduits (NGC's) to direct regenerating axons between the proximal and distal ends of the nerve gap. However, NGC's are limited in their effectiveness at promoting regeneration Current NGCs are not suitable as substrates for supporting either neuronal or Schwann cell growth, as they lack an architecture similar to that of the native extracellular matrix (ECM) of the nerve. The aim of this study was to create an acellular porcine peripheral nerve using a novel decellularisation protocol, in order to eliminate the immunogenic cellular components of the tissue, while preserving the three-dimensional histoarchitecture and ECM components. Porcine peripheral nerve (sciatic branches were decellularised using a low concentration (0.1%; w/v) sodium dodecyl sulphate in conjunction with hypotonic buffers and protease inhibitors, and then sterilised using 0.1% (v/v) peracetic acid. Quantitative and qualitative analysis revealed a ≥95% (w/w) reduction in DNA content as well as preservation of the nerve fascicles and connective tissue. Acellular nerves were shown to have retained key ECM components such as collagen, laminin and fibronectin. Slow strain rate to failure testing demonstrated the biomechanical properties of acellular nerves to be comparable to fresh controls. In conclusion, we report the production of a biocompatible, biomechanically functional acellular scaffold, which may have use in peripheral nerve repair. Biotechnol. Bioeng. 2016;113: 2041-2053. © 2016 The Authors. Biotechnology and Bioengineering published by Wiley Periodicals, Inc.

An anatomical study of porcine peripheral nerve and its potential use in nerve tissue engineering.

Current nerve tissue engineering applications are adopting xenogeneic nerve tissue as potential nerve grafts to help aid nerve regeneration. However, there is little literature that describes the exact location, anatomy and physiology of these nerves to highlight their potential as a donor graft. The aim of this study was to identify and characterise the structural and extracellular matrix (ECM) components of porcine peripheral nerves in the hind leg. Methods included the dissection of porcine nerves, localisation, characterisation and quantification of the ECM components and identification of nerve cells. Results showed a noticeable variance between porcine and rat nerve (a commonly studied species) in terms of fascicle number. The study also revealed that when porcine peripheral nerves branch, a decrease in fascicle number and size was evident. Porcine ECM and nerve fascicles were found to be predominately comprised of collagen together with glycosaminoglycans, laminin and fibronectin. Immunolabelling for nerve growth factor receptor p75 also revealed the localisation of Schwann cells around and inside the fascicles. In conclusion, it is shown that porcine peripheral nerves possess a microstructure similar to that found in rat, and is not dissimilar to human. This finding could extend to the suggestion that due to the similarities in anatomy to human nerve, porcine nerves may have utility as a nerve graft providing guidance and support to regenerating axons.

Investigating NF-κB signaling in lung fibroblasts in 2D and 3D culture systems.

BACKGROUND: Inflammatory respiratory diseases are amongst major global health challenges. Lung fibroblasts have been shown to play a key role in lung inflammatory responses. However, their exact role in initiation and maintenance of lung diseases has remained elusive partly due to the limited availability of physiologically relevant in vitro models. Therefore, developing new tools that enable investigating the molecular pathways (e.g. nuclear factor-kappa B (NF-κB) activation) that underpin inflammatory responses in fibroblasts could be a valuable resource for scientists working in this area of research. RESULTS: In order to investigate NF-κB activation in response to pro-inflammatory stimuli in real-time, we first developed two detection systems based on nuclear localization of NF-κB by immunostaining and luciferase reporter assay system. Furthermore using electrospun porous scaffolds, with similar geometry to human lung extracellular matrix, we developed 3D cultures of lung fibroblasts allowing comparing NF-κB activation in response to pro-inflammatory stimuli (i.e. TNF-α) in 2D and 3D. Our data clearly show that the magnitude of NF-κB activation in 2D cultures is substantially higher than 3D cultures. However, unlike 2D cultures, cells in the 3D model remained responsive to TNF-α at higher concentrations. The more subdued and wider dynamic range of NF-κB responses in 3D culture system was associated with a different expression pattern for TNF receptor I in 3D versus 2D cultures collectively reflecting a more in vivo like TNF receptor I expression and NF-κB activation pattern in the 3D system. CONCLUSION: Our data suggest that lung fibroblasts are actively involved in the pathogenesis of lung inflammation by activation of NF-κB signaling pathway. The 3D culture detection system provides a sensitive and biologically relevant tool for investigating different pro-inflammatory events involving lung fibroblasts.

Arginine-glycine-aspartic acid functional branched semi-interpenetrating hydrogels.

For the first time a series of functional hydrogels based on semi-interpenetrating networks with both branched and crosslinked polymer components have been prepared and we show the successful use of these materials as substrates for cell culture. The materials consist of highly branched poly(N-isopropyl acrylamide)s with peptide functionalised end groups in a continuous phase of crosslinked poly(vinyl pyrrolidone). Functionalisation of the end groups of the branched polymer component with the GRGDS peptide produces a hydrogel that supports cell adhesion and proliferation. The materials provide a new synthetic functional biomaterial that has many of the features of extracellular matrix, and as such can be used to support tissue regeneration and cell culture. This class of high water content hydrogel material has important advantages over other functional hydrogels in its synthesis and does not require post-processing modifications nor are functional-monomers, which change the polymerisation process, required. Thus, the systems are amenable to large scale and bespoke manufacturing using conventional moulding or additive manufacturing techniques. Processing using additive manufacturing is exemplified by producing tubes using microstereolithography.

Immunocompetent 3D model of human upper airway for disease modeling and in vitro drug evaluation.

The development of more complex in vitro models for the assessment of novel drugs and chemicals is needed because of the limited biological relevance of animal models to humans as well as ethical considerations. Although some human-cell-based assays exist, they are usually 2D, consist of single cell type, and have limited cellular and functional representation of the native tissue. In this study, we have used biomimetic porous electrospun scaffolds to develop an immunocompetent 3D model of the human respiratory tract comprised of three key cell types present in upper airway epithelium. The three cell types, namely, epithelial cells (providing a physical barrier), fibroblasts (extracellular matrix production), and dendritic cells (immune sensing), were initially grown on individual scaffolds and then assembled into the 3D multicell tissue model. The epithelial layer was cultured at the air-liquid interface for up to four weeks, leading to formation of a functional barrier as evidenced by an increase in transepithelial electrical resistance (TEER) and tight junction formation. The response of epithelial cells to allergen exposure was monitored by quantifying changes in TEER readings and by assessment of cellular tight junctions using immunostaining. It was found that epithelial cells cocultured with fibroblasts formed a functional epithelial barrier at a quicker rate than single cultures of epithelial cells and that the recovery from allergen exposure was also more rapid. Also, our data show that dendritic cells within this model remain viable and responsive to external stimulation as evidenced by their migration within the 3D construct in response to allergen challenge. This model provides an easy to assemble and physiologically relevant 3D model of human airway epithelium that can be used for studies aiming at better understanding lung biology, the cross-talk between immune cells, and airborne allergens and pathogens as well as drug delivery.

Dinuclear ruthenium(II) complexes as two-photon, time-resolved emission microscopy probes for cellular DNA.

The first transition-metal complex-based two-photon absorbing luminescence lifetime probes for cellular DNA are presented. This allows cell imaging of DNA free from endogenous fluorophores and potentially facilitates deep tissue imaging. In this initial study, ruthenium(II) luminophores are used as phosphorescent lifetime imaging microscopy (PLIM) probes for nuclear DNA in both live and fixed cells. The DNA-bound probes display characteristic emission lifetimes of more than 160 ns, while shorter-lived cytoplasmic emission is also observed. These timescales are orders of magnitude longer than conventional FLIM, leading to previously unattainable levels of sensitivity, and autofluorescence-free imaging.

Laser exposure of gold nanorods can increase neuronal cell outgrowth.

The usage of gold nanoparticles (Au NPs) in biological applications has risen significantly over the last 10 years. With the wide variety of chemical and biological functionalization available and their distinctive optical properties, Au NPs are currently used in a range of biological applications including sensing, labeling, drug delivery, and imaging applications. Among the available particles, gold nanorods (Au NRs) are particularly useful because their optical absorption can be tuned across the visible to near infrared region. Here, we present a novel application of Au NRs associated with low power laser exposure of NG108-15 neuronal cells. When cells were irradiated with a 780 nm laser, the average number of neurons with neurites increased. A similar stimulatory effect was observed for cells that were cultured with poly-(4-styrenesulfonic acid)-coated and silica-coated Au NRs. Furthermore, when the NG108-15 cells were cultured with both bare and coated Au NRs and then irradiated with 1.2-7.5 W/cm(2) at 780 nm, they showed a neurite length increase of up to 25 µm versus control. To the best of our knowledge, this effect has never been reported before. While the pathways of the stimulation is not yet clear, the data presented here demonstrates that it is linked to the absorption of light by the Au NRs. These initial results open up new opportunities for peripheral nerve regeneration treatments and for novel approaches to addressing central nervous system axons following spinal cord injury.

Selectively inhibiting malignant melanoma migration and invasion in an engineered skin model using actin-targeting dinuclear RuII-complexes.

Due to the poor prognosis of metastatic cancers, there is a clinical need for agents with anti-metastatic activity. Here we report on the anti-metastatic effect of a previously reported Ru(ii) complex [{(phen)2Ru}2(tpphz)]4+, 14+, that has recently been shown to disrupt actin fiber assembly. In this study, we investigated the anti-migratory effect of +14+ and a close structural analogue+, 24+, on two highly invasive, metastatic human melanoma cell lines. Laser scanning confocal imaging was used to investigate the structure of actin filament and adhesion molecule vinculin and results show disassembly of central actin filaments and focal adhesions. The effect of both compounds on actin filaments was also found to be reversible. As these results revealed that the complexes were cytostatic and produced a significant inhibitory effect on the migration of both melanoma cell lines but not human dermal fibroblasts their effect on 3D-spheroids and a tissue-engineered living skin model were also investigated. These experiments demonstrated that the compounds inhibited the growth and invasiveness of the melanoma-based spheroidal tumor model and both complexes were found to penetrate the epidermis of the skin tissue model and inhibit the invasion of melanoma cells. Taken together, the cytostatic and antimigratory effects of the complexes results in an antimetastatic effect that totally prevent invasion of malignant melanoma into skin tissue.

Aligned Polyhydroxyalkanoate Blend Electrospun Fibers as Intraluminal Guidance Scaffolds for Peripheral Nerve Repair.

The use of nerve guidance conduits (NGCs) to treat peripheral nerve injuries is a favorable approach to the current "gold standard" of autografting. However, as simple hollow tubes, they lack specific topographical and mechanical guidance cues present in nerve grafts and therefore are not suitable for treating large gap injuries (30-50 mm). The incorporation of intraluminal guidance scaffolds, such as aligned fibers, has been shown to increase neuronal cell neurite outgrowth and Schwann cell migration distances. A novel blend of PHAs, P(3HO)/P(3HB) (50:50), was investigated for its potential as an intraluminal aligned fiber guidance scaffold. Aligned fibers of 5 and 8 µm diameter were manufactured by electrospinning and characterized using SEM. Fibers were investigated for their effect on neuronal cell differentiation, Schwann cell phenotype, and cell viability in vitro. Overall, P(3HO)/P(3HB) (50:50) fibers supported higher neuronal and Schwann cell adhesion compared to PCL fibers. The 5 µm PHA blend fibers also supported significantly higher DRG neurite outgrowth and Schwann cell migration distance using a 3D ex vivo nerve injury model.

Medium chain length polyhydroxyalkanoates as potential matrix materials for peripheral nerve regeneration.

Polyhydroxyalkanoates are natural, biodegradable, thermoplastic and sustainable polymers with a huge potential in fabrication of bioresorbable implantable devices for tissue engineering. We describe a comparative evaluation of three medium chain length polyhydroxyalkanoates (mcl-PHAs), namely poly(3-hydroxyoctanoate), poly(3-hydroxyoctanoate-co-3-hydoxydecanoate) and poly(3-hydroxyoctanoate-co-3-hydroxydecanoate-co-3-hydroxydodecanoate), one short chain length polyhydroxyalkanoate, poly(3-hydroxybutyrate), P(3HB) and synthetic aliphatic polyesters (polycaprolactone and polylactide) with a specific focus on nerve regeneration, due to mechanical properties of mcl-PHAs closely matching nerve tissues. In vitro biological studies with NG108-15 neuronal cell and primary Schwann cells did not show a cytotoxic effect of the materials on both cell types. All mcl-PHAs supported cell adhesion and viability. Among the three mcl-PHAs, P(3HO-co-3HD) exhibited superior properties with regards to numbers of cells adhered and viable cells for both cell types, number of neurite extensions from NG108-15 cells, average length of neurite extensions and Schwann cells. Although, similar characteristics were observed for flat P(3HB) surfaces, high rigidity of this biomaterial, and FDA-approved polymers such as PLLA, limits their applications in peripheral nerve regeneration. Therefore, we have designed, synthesized and evaluated these materials for nerve tissue engineering and regenerative medicine, the interaction of mcl-PHAs with neuronal and Schwann cells, identifying mcl-PHAs as excellent materials to enhance nerve regeneration and potentially their clinical application in peripheral nerve repair.

Aminosilane Functionalized Aligned Fiber PCL Scaffolds for Peripheral Nerve Repair.

Silane modification is a simple and cost-effective tool to modify existing biomaterials for tissue engineering applications. Aminosilane layer deposition has previously been shown to control NG108-15 neuronal cell and primary Schwann cell adhesion and differentiation by controlling deposition of ─NH2 groups at the submicron scale across the entirety of a surface by varying silane chain length. This is the first study toreport depositing 11-aminoundecyltriethoxysilane (CL11) onto aligned Polycaprolactone (PCL) scaffolds for peripheral nerve regeneration. Fibers are manufactured via electrospinning and characterized using water contact angle measurements, atomic force microscopy (AFM), and X-ray photoelectron spectroscopy (XPS). Confirmed modified fibers are investigated using in vitro cell culture of NG108-15 neuronal cells and primary Schwann cells to determine cell viability, cell differentiation, and phenotype. CL11-modified fibers significantly support NG108-15 neuronal cell and Schwann cell viability. NG108-15 neuronal cell differentiation maintains Schwann cell phenotype compared to unmodified PCL fiber scaffolds. 3D ex vivo culture of Dorsal root ganglion explants (DRGs) confirms further Schwann cell migration and longer neurite outgrowth from DRG explants cultured on CL11 fiber scaffolds compared to unmodified scaffolds. Thus, a reproducible and cost-effective tool is reported to modify biomaterials with functional amine groups that can significantly improve nerve guidance devices and enhance nerve regeneration.

Human hair follicle dermal cells and skin fibroblasts show differential activation of NF-κB in response to pro-inflammatory challenge.

The underlying mechanism of immune privilege in hair follicle cell dermal papilla (DP) and sheath (DS) populations is not well understood, and the responsiveness of hair follicle dermal cells to pro-inflammatory challenge presently remains unknown. In this work, we describe acute NF-κB activation in human DS, DP and dermal fibroblast (DF) cells challenged with TNF-alpha and IL1-beta. In contrast, the DS and DP cells revealed an unexpected tolerance to bacterial LPS challenge relative to DF cells. Understanding follicle cell responses to typical pro-inflammatory stimuli is critical for diseases where collapse of hair follicle immune privilege is observed, and to further applications in autologous stem cell/wound healing therapeutics.

Polyhydroxyalkanoates and their advances for biomedical applications.

Polyhydroxyalkanoates (PHAs) are sustainable, versatile, biocompatible, and bioresorbable polymers that are suitable for biomedical applications. Produced via bacterial fermentation under nutrient-limiting conditions, they are uncovering a new horizon for devices in biomedical applications. A wide range of cell types including bone, cartilage, nerve, cardiac, and pancreatic cells, readily attach grow and are functional on PHAs. The tuneable physical properties and resorption rates of PHAs provide a toolbox for biomedical engineers in developing devices for hard and soft tissue engineering applications and drug delivery. The versatility of PHAs and the vast range of different PHA-based prototypes are discussed. Current in vitro, ex vivo, and in vivo development work are described and their regulatory approvals are reviewed.

Anatomical site influences the differentiation of adipose-derived stem cells for Schwann-cell phenotype and function.

Considerable attention has recently been given to adipose-derived stem cells (ASCs) as an important source for differentiation to Schwann cells in the treatment of peripheral nerve injury, with considerable clinical advantages over the use of mesenchymal stem cells derived from bone marrow or autologous Schwann cells. However, the relationship between adipose donor site and differentiated ASC phenotype and function is presently unknown. This work systematically studied the differentiation of ASCs harvested from three anatomical sites: (i) subcutaneous; (ii) perinephric; and (iii) epididymal adipose tissue. We show that ASC source is a major determining factor of immunophenotype, multilineage differentiation, Schwann-cell protein expression, and paracrine ability to stimulate neuronal growth. Upregulation of S100ß, glial fibrillary acidic protein (GFAP), and p75NGFR was observed in differentiated ASCs from perinephric fat tissue, while only the expression of S100ß or GFAP and p75NGFR was elevated in differentiated ASCs from subcutaneous or epididymal fat tissue. Although the co-culture of differentiated ASCs with NG108-15 neuronal cells demonstrated that ASCs from each source could stimulate neurite outgrowth and number, differentiated ASCs from subcutaneous and perinephric fat versus epididymal fat were most effective, which was attributed to high-brain-derived neurotropic factor/nerve growth factor and low-neurotrophin-3 levels. Thus, ASCs can be obtained from different anatomical locations, and this determines Schwann-cell phenotype upon differentiation and extent of function. This work is therefore of relevance in local therapeutic delivery of ASCs for the repair of peripheral nerve injury, but also in the broader context of ASC use in related stem-cell therapies.

Time-resolved and two-photon emission imaging microscopy of live cells with inert platinum complexes.

This work explores time-resolved emission imaging microscopy (TREM) for noninvasive imaging and mapping of live cells on a hitherto uncharted microsecond time scale. Simple robust molecules for this purpose have long been sought. We have developed highly emissive, synthetically versatile, and photostable platinum(II) complexes that make TREM a practicable reality. [PtLCl], {HL = 1,3-di(2-pyridyl)benzene and derivatives}, are charge-neutral, small molecules that have low cytotoxicity and accumulate intracellularly within a remarkably short incubation time of 5 min, apparently under diffusion control. Their microsecond lifetimes and emission quantum yields of up to 70% are exceptionally high for transition metal complexes and permit the application of TREM to be demonstrated in a range of live cell types-normal human dermal fibroblast, neoplastic C8161 and CHO cells. [PtLCl] are thus likely to be suitable emission labels for any eukaryotic cell types. The high photostability of [PtLCl] under intense prolonged irradiation has allowed the development of tissue-friendly NIR two-photon excitation (TPE) in conjunction with transition metal complexes in live cells. A combination of confocal one-photon excitation, nonlinear TPE, and microsecond time-resolved imaging has revealed (i) preferential localization of the complexes to intracellular nucleic acid structures, in particular the nucleoli and (ii) the possibility of measuring intracellular emission lifetimes in the microsecond range. The combination of TREM, TPE, and Pt(II) complexes will be a powerful tool for investigating intracellular processes in vivo, because the long lifetimes allow discrimination from autofluorescence and open up the use of commonplace technology.

Cost effective optimised synthetic surface modification strategies for enhanced control of neuronal cell differentiation and supporting neuronal and Schwann cell viability.

Enriching a biomaterial surface with specific chemical groups has previously been considered for producing surfaces that influence cell response. Silane layer deposition has previously been shown to control mesenchymal stem cell adhesion and differentiation. However, it has not been used to investigate neuronal or Schwann cell responses in vitro to date. We report on the deposition of aminosilane groups for peripheral neurons and Schwann cells studying two chain lengths: (a) 3-aminopropyl triethoxysilane (short chain-SC) and (b) 11-aminoundecyltriethoxysilane (long chain-LC) by coating glass substrates. Surfaces were characterised by water contact angle, AFM and XPS. LC-NH2 was produced reproducibly as a homogenous surface with controlled nanotopography. Primary neuron and NG108-15 neuronal cell differentiation and primary Schwann cell responses were investigated in vitro by S100ß, p75, and GFAP antigen expression. Both amine silane surface supported neuronal and Schwann cell growth; however, neuronal differentiation was greater on LC aminosilanes versus SC. Thus, we report that silane surfaces with an optimal chain length may have potential in peripheral nerve repair for the modification and improvement of nerve guidance devices.