Scaffolds For Nerve Tissue Engineering Research Articles

Hybrid electrospun polyhydroxybutyrate/gelatin/laminin/polyaniline scaffold for nerve tissue engineering application: Preparation, characterization, and in vitro assay

Despite the widespread central nervous system injuries, treatment of these disorders is still an issue of concern due to the complexities. Natural recovery in these patients is rarely observed, which calls for developing new methods that address these problems. In this study, natural polymers of polyhydroxybutyrate (PHB) and gelatin were electrospun into scaffolds and cross-linked. In order to modify the PHB-based scaffold for nerve tissue engineering, the scaffold surface was modified by exposure to the ammonium gas plasma under controlled conditions, and the laminin as a promoter for neural cells was coated on the sample surface. Then, polyaniline nanoparticles were inkjet-printed on a sample surface as parallel lines to induce the differentiation of stem cells into neural cells. Infrared spectroscopy, absorption of PBS, AFM, degradation rate, contact angle, electron microscopy and optical microscopy, thermal and mechanical behavior, and analysis of the viability of L929 cells were investigated for the scaffolds. The results showed gelatin decreased the contact angle from 106.2° to 38° and increased the residual weight after PBS incubation from 82 % to 38 %. The moduli of the scaffold increased from 8.78 MPa for pure PHB to 28.74 for the modified scaffold. In addition, performed methods increased cell viability from 69 % for PHB to 89 % for modified scaffold and also had a favorable effect on cell adhesion. Investigation of culturing P19 stem cells demonstrated that they successfully differentiated into neural cells. Results show that the scaffolds prepared in this study were promising for nerve tissue engineering.

The Effect of Vitamin C-Loaded Electrospun Polycaprolactone/Poly (Glycerol Sebacate) Fibers for Peripheral Nerve Tissue Engineering

Vitamin C (VC) is an essential supplement that plays a vital role in cellular processes and functions and has been applied for therapeutic purposes for many years. The beneficial effects of VC on peripheral nerve regeneration have been gained lots of attention. In this study, electrospun polycaprolactone (PCL)/polyglycerol sebacate (PGS) fibers incorporated with different concentrations of VC (5, 10, and 15 wt.%) were developed for peripheral nerve tissue engineering. The morphology of the fibers was investigated using scanning electron microscope (SEM), Fourier-transform infrared spectroscopy (FTIR), tensile analysis (Young’s modulus, ultimate tensile strength (UTS), and elongation at break), release profile of VC from the PCL/PGS fibers, in vitro degradation, water uptake behavior, and contact angle measurements were also studied. MTT assay and SEM were utilized to evaluate the attachment and viability of pheochromocytoma cells (PC12) on the scaffolds. The results showed that all scaffolds had a uniform diameter and mean diameter deceased from 1.24 to 0.88 µm followed by increasing VC. Young’s modulus and UTS enhanced with increasing in VC percentage. MTT assay demonstrated that PCL/PGS containing 5 wt.% VC had a greater viability rate among other scaffolds. Our outcome indicated possible applicability of VC containing scaffolds for nerve tissue engineering.

In vitro preparation of human Dental Pulp Stem Cell grafts with biodegradable polymer scaffolds for nerve tissue engineering.

Human Dental Pulp Stem Cells (hDPSCs) are one of the most promising stem cell sources for tissue engineering and regeneration, due to their extraordinary multi-lineage differentiation ability, ease of extraction from biological waste in dental clinics, safe non-tumorigenic phenotype, immune-tolerance upon in vivo transplantation, and great possibilities of application in autologous tissue reconstruction. The in vitro manipulation of hDPSCs paves the way for drug screening and tailor-made regeneration of damaged tissues, in the context of personalized medicine. The neural crest phenotype of these stem cells gives them the capacity to differentiate to a large variety of cell types, including neural-lineage cells. In this chapter, we describe various culture methods to generate different cellular phenotypes from hDPSCs, which can not only grow as mesenchymal-like plastic adherent cells, but also as non-adherent neurogenic dentospheres in serum-free medium. Floating dentospheres can be used to generate large populations of mature neuronal and glial marker expressing cells, which may be cultured over a substrate of nanopatterned scaffold based on biodegradable poly(lactide-co-caprolactone) (PLCL) to induce a controlled alignment of neurites and cell migration, to generate in vivo biocompatible constructs for nerve tissue bioengineering.

Proliferation and differentiation study of melatonin functionalized polycaprolactone/gelatin electrospun fibrous scaffolds for nerve tissue engineering

Melatonin (MLT), a pineal neurohormone with multiple neuroprotective, is often used for peripheral nerve recovery and regenerated nerve proliferation. In this study, Polycaprolactone/Gelatin (PG) fibrous electrospun scaffolds with various percentages of MLT (0, 1, 2, and 4%wt) were fabricated for nerve cell growth, the effects of different concentrations of MLT within PG fibers (PG, PGMLT1, PGMLT2, and PGMLT4) on the proliferation and differentiation for PC12 cells were quantitatively evaluated. The microstructures and morphologies of these scaffolds were analyzed by FE-SEM and digital camera. Fourier transform infrared spectrometer (FTIR), X-ray photoelectron spectroscopy (XPS), and Water Contact Angle (WCA) were used to study the composition, ratio and properties of MLT functionalized PG scaffolds. MTT and CLSM analysis showed that appropriate amount of MLT was beneficial to the proliferation of PC12 cell. MLT can also promote cell differentiation, neurite germination, the expression levels of MAP2 mRNA and protein were dramatically increased on the composite scaffolds with the increase of MLT content, moderate addition of MLT (PGMLT2, 2%) had a prominent enhancement for neurite length. This work would provide a more comprehensive reference for further researches on MLT functionalized composite scaffolds and suggest that high-performance PGMLT fibrous scaffolds could be a promising alternative for nerve repair.

Assessing physicochemical, mechanical, and in vitro biological properties of polycaprolactone/poly(glycerol sebacate)/hydroxyapatite composite scaffold for nerve tissue engineering

The ability to mimic the extracellular matrix (ECM) by electrospun fibers has opened new practical ways in nerve tissue engineering. In the current research, polycaprolactone (PCL)/poly(glycerol sebacate) (PGS) scaffold containing 5, 10, and 15 wt% hydroxyapatite (HAp) particles were electrospinned. The morphology, surface contact angle, mechanical properties, in vitro degradation behavior, and water uptake of the PCL/PGS/HAp fibers were studied. The viability and adhesion of the rat pheochromocytoma cell line (PC12) on each scaffold were investigated using MTT assay and scanning electron microscope (SEM), respectively. SEM images revealed that the fibers were uniform and aligned in general, while by increasing the HAp particles loading from 0 to 15%, the fiber diameter decreased from 831 to 382 nm. According to transmission electron microscopy (TEM) images, needle-like HAp particles were distributed along the fibers. Fourier-transform infrared spectroscopy (FTIR) and X-ray analyses confirmed the chemical interactions between PCL, PGS, and HAp. Young's modulus of all fibers was about 0.16–0.3 MPa, which is practically suitable for nerve tissue engineering. HAp particles displayed a positive effect on PC12 viability and adhesion. To conclude, the results suggested that electrospun PCL/PGS/HAp fibers can be a promising biodegradable scaffold for serving in nerve tissue engineering.

The influence of reduced graphene oxide on stem cells: a perspective in peripheral nerve regeneration.

Graphene and its derivatives are fascinating materials for their extraordinary electrochemical and mechanical properties. In recent decades, many researchers explored their applications in tissue engineering and regenerative medicine. Reduced graphene oxide (rGO) possesses remarkable structural and functional resemblance to graphene, although some residual oxygen-containing groups and defects exist in the structure. Such structure holds great potential since the remnant-oxygenated groups can further be functionalized or modified. Moreover, oxygen-containing groups can improve the dispersion of rGO in organic or aqueous media. Therefore, it is preferable to utilize rGO in the production of composite materials. The rGO composite scaffolds provide favorable extracellular microenvironment and affect the cellular behavior of cultured cells in the peripheral nerve regeneration. On the one hand, rGO impacts on Schwann cells and neurons which are major components of peripheral nerves. On the other hand, rGO-incorporated composite scaffolds promote the neurogenic differentiation of several stem cells, including embryonic stem cells, mesenchymal stem cells, adipose-derived stem cells and neural stem cells. This review will briefly introduce the production and major properties of rGO, and its potential in modulating the cellular behaviors of specific stem cells. Finally, we present its emerging roles in the production of composite scaffolds for nerve tissue engineering.

Fabrication and evaluation of porous and conductive nanofibrous scaffolds for nerve tissue engineering

Peripheral nerve repair is still one of the major clinical challenges which has received a great deal of attention. Nerve tissue engineering is a novel treatment approach that provides a permissive environment for neural cells to overcome the constraints of repair. Conductivity and interconnected porosity are two required characteristics for a scaffold to be effective in nerve regeneration. In this study, we aimed to fabricate a conductive scaffold with controlled porosity using polycaprolactone (PCL) and chitosan (Chit), FDA approved materials for the use in implantable medical devices. A novel method of using tetrakis (hydroxymethyl) phosphonium chloride (THPC) and formaldehyde was applied for in situ synthesis of gold nanoparticles (AuNPs) on the scaffolds. In order to achieve desirable porosity, different percentage of polyethylene oxide (PEO) was used as sacrificial fiber. Fourier transform infrared spectroscopy (FTIR) and field emission scanning electron microscopy (FE-SEM) results demonstrated the complete removing of PEO from the scaffolds after washing and construction of interconnected porosities, respectively. Elemental and electrical analysis revealed the successful synthesis of AuNPs with uniform distribution and small average diameter on the PCL/Chit scaffold. Contact angle measurements showed the effect of porosity on hydrophilic properties of the scaffolds, where the porosity of 75–80% remarkably improved surface hydrophilicity. Finally, the effect of conductive nanofibrous scaffold on Schwann cells morphology and vaibility was investigated using FE-SEM and MTT assay, respectively. The results showed that these conductive scaffolds had no cytotoxic effect and support the spindle-shaped morphology of cells with elongated process which are typical of Schwann cell cultures.

Zero-valent iron nanoparticles containing nanofiber scaffolds for nerve tissue engineering.

Regeneration of nerve tissue is a challenging issue in regenerative medicine. Especially, the peripheral nerve defects related to the accidents are one of the leading health problems. For large degeneration of peripheral nerve, nerve grafts are used in order to obtain a connection. These grafts should be biodegradable to prevent second surgical intervention. In order to make more effective nerve tissue engineering materials, nanotechnological improvements were used. Especially, the addition of electrically conductive and biocompatible metallic particles and carbon structures has essential roles in the stimulation of nerves. However, the metabolizing of these structures remains to wonder because of their nondegradable nature. In this study, biodegradable and conductive nerve tissue engineering materials containing zero-valent iron (Fe) nanoparticles were developed and investigated under in vitro conditions. By using electrospinning technique, fibrous mats composed of electrospun poly(ε-caprolactone) (PCL) nanofibers and Fe nanoparticles were obtained. Both electrical conductivity and mechanical properties increased compared with control group that does not contain nanoparticles. Conductivity of PCL/Fe5 and PCL/Fe10 increased to 0.0041 and 0.0152 from 0.0013 Scm-1 , respectively. Cytotoxicity results indicated toxicity for composite mat containing 20% Fe nanoparticles (PCL/Fe20). SH-SY5Y cells were grown on PCL/Fe10 best, which contains 10% Fe nanoparticles. Beta III tubulin staining of dorsal root ganglion neurons seeded on mats revealed higher cell number on PCL/Fe10. This study demonstrated the impact of zero-valent Fe nanoparticles on nerve regeneration. The results showed the efficacy of the conductive nanoparticles, and the amount in the composition has essential roles in the promotion of the neurites.

Physical understanding of axonal growth patterns on grooved substrates: groove ridge crossing versus longitudinal alignment

Surface topographies such as micrometric edges and grooves have been widely used to improve neuron outgrowth. However, finding the mechanism of neuron–surface interactions on grooved substrates remains a challenge. In this work, PC12 cells and chick forebrain neurons (CFNs) were cultured on grooved and smooth polyacrylonitrile substrates. It was found that CFNs showed a tendency of growing across groove ridges; while PC12 cells were only observed to grow in the longitudinal direction of grooves. To further investigate these observations, a 3D physical model of axonal outgrowth was developed. In this model, axon shafts are simulated as elastic 3D beams, accounting for the axon outgrowth as well as the focal contacts between axons and substrates. Moreover, the bending direction of axon tips during groove ridge crossing is governed by the energy minimization principle. Our physical model predicts that axonal groove ridge crossing is contributed by the bending compliance of axons, caused by lower Young’s modulus and smaller diameters. This work will aid the understanding of the mechanisms involved in axonal alignment and elongation of neurons guided by grooved substrates, and the obtained insights can be used to enhance the design of instructive scaffolds for nerve tissue engineering and regeneration applications.

Gradient degradable nerve guidance conduit with multilayer structure prepared by electrospinning

Polymeric nerve guidance conduits (NGCs) with uniform tubular structure are widely used as scaffolds in nerve tissue engineering. However, massive collapse of NGCs has greatly impeded their further application. Therefore, the gradient degradable nerve guidance conduit (GD-NGC) with multilayer structure is designed and fabricated via electrospinning technique. The GD-NGC consists of five poly(lactic-co-glycolic acid) (PLGA) layers with different polylactic acid (PLA)/polyglycolic acid (PGA) ratios. Compared to common NGCs, the novel GD-NGC degrades moderately and stepwise from inner to outer layers and also exhibits equivalent or better characteristics, such as porosity, water absorption capacity, mechanical property and biocompatibility. Collectively, GD-NGC has a promising potential for applications in nerve tissue engineering.

Beneficial effects of biodelivery of brain-derived neurotrophic factor and gold nanoparticles from functionalized electrospun PLGA scaffold for nerve tissue engineering

Despite of the pivotal role of Schwann cells (SCs) in peripheral nerve regeneration, lack of an available source has prompted researches for Schwann-like cells transdifferentiation. This study suggests an effective method for tuning the surface of aligned poly (lactic-co-glycolic acid) (PLGA) nanofibers to enhance attachment and proliferation of h-ADSCs on the scaffold through coating of laminin. Following the characterization of biofunctionalized PLGA, brain-derived neurotropic factor (BDNF) and gold nanoparticles (AuNPs) were encapsulated in chitosan nanoparticles (CSNPs), added into laminin solution, and coated on the surface of aligned PLGA scaffold. The release behavior of BDNF and AuNPs from scaffold was evaluated by Bradford assay and inductive coupled plasma optical emission spectrometry (ICP-OES) technique, respectively. Afterwards, experimental groups were investigated for potential of Schwann cell differentiation using immunocytochemical staining and real-time RT-PCR technique. Results of MTT assay showed the significantly higher proliferation of h-ADSCs on laminin-functionalized scaffold compared to PLGA scaffold (p < 0.05). Additionally, the presence of BDNF and AuNPs on scaffold significantly improved the expression of SCs markers as compared to the control group (p < 0.05). Therefore, use of biofunctionalized PLGA nanofibers can be a promising strategy for inducing the differentiation of h-ADSCs into SCs for nerve tissue engineering.

Aligned conductive core-shell biomimetic scaffolds based on nanofiber yarns/hydrogel for enhanced 3D neurite outgrowth alignment and elongation

Aligned topographical cue has been demonstrated as a critical role in neuronal guidance, and it is highly beneficial to develop a scaffold with aligned structure for peripheral nerve tissue regeneration. Although considerable efforts have been devoted to guiding neurite alignment and extension, it remains a remarkable challenge for developing a biomimetic scaffold for enhancing 3D aligned neuronal outgrowth. Herein, we present a core-shell scaffold based on aligned conductive nanofiber yarns (NFYs) within the hydrogel to mimic the 3D hierarchically aligned structure of the native nerve tissue. The aligned NFYs assembled by a bundle of aligned nanofibers composed of polycaprolactone (PCL), silk fibroin (SF), and carbon nanotubes (CNTs) are prepared by a developed dry–wet electrospinning method, which has the ability to induce neurite alignment and elongation when PC12 cells and dorsal root ganglia (DRG) cells are cultured on their 3D peripheral surface. Particularly, such an aligned nanofibrous structure also induces aligned neurite extension and cell migration from DRG explants along the direction of nanofibers. 3D core-shell scaffolds are fabricated by encapsulating NFYs within the hydrogel shell after photocrosslinking, and these 3D aligned scaffolds are able to control cellular alignment and elongation of nerve cells in this 3D environment. Our results suggest that such 3D hierarchically aligned core-shell scaffold consists of NFYs that mimic the aligned nerve fiber structure to induce neurite alignment and extension and a hydrogel shell that mimics the epineurium layer to protect nerve cell organization within a 3D environment, which is largely promising for the design of biomimetic scaffolds in nerve tissue engineering. Statement of SignificanceDesigning scaffolds with 3D aligned structure has been paid more attention for peripheral nerve tissue regeneration, because the aligned topographical cue is able to induce neurites alignment and extension. However, developing scaffolds mimicking the hierarchically aligned structure of native nerve tissue for directing 3D aligned neuronal outgrowth without external stimulation remains challenging. This work presented a simple and efficient strategy to prepare a 3D biomimetic core-shell scaffold based on electrospun aligned conductive nanofiber yarns within photocurable hydrogel shell to mimic the hierarchically aligned structure of native nerve tissue. These 3D aligned composite scaffolds performed the ability to direct 3D cellular alignment and elongation of nerve cells along with the nanofiber yarn direction, and the hydrogel shell mimicking the epineurium layer was able to protect nerve cells organization within the 3D environment, which indicated their great potential in peripheral nerve tissue engineering applications.

Fabrication of poly (trimethylene carbonate)/reduced graphene oxide-graft-poly (trimethylene carbonate) composite scaffolds for nerve regeneration

One of the key challenges for neural tissue engineering is to exploit functional materials to guide and support nerve regeneration. Currently, reduced graphene oxide (rGO), which is well-known for its unique electrical and mechanical properties, has been incorporated into biocompatible polymers to manufacture functional scaffolds for nerve tissue engineering. However, rGO has poor dispersity in polymer matrix, which limits its further application. Here, we replaced rGO with rGO-graft-PTMC. The rGO-graft-PTMC was firstly prepared by grafting trimethylene carbonate (TMC) oligomers onto rGO. Subsequently, PTMC/rGO-graft-PTMC composite fibrous mats were fabricated by electrospinning of a dispersion of PTMC and rGO-graft-PTMC. The loading of rGO-graft-PTMC could reach up to 6 wt% relative to PTMC. Scanning electron microscopy images showed that the morphologies and average diameters of PTMC/rGO-graft-PTMC composite fibrous mats were affected by the content of rGO-graft-PTMC. Additionally, the incorporation of rGO-graft-PTMC resulted in enhanced thermal stability and hydrophobicity of PTMC fibers. Biological results demonstrated that PC12 cells showed higher cell viability on PTMC/rGO-graft-PTMC fibers of 2.4, 4.0 and 6.0 wt% rGO-graft-PTMC compared to pure PTMC fibers. These results suggest that PTMC/rGO-graft-PTMC composite fibrous structures hold great potential for neural tissue engineering.

Development of three-dimensional piezoelectric polyvinylidene fluoride-graphene oxide scaffold by non-solvent induced phase separation method for nerve tissue engineering

In this work, non-solvent induced phase separation method was applied to develop polyvinylidene fluoride (PVDF)/graphene oxide (GO) scaffold for nerve tissue engineering. Furthermore, the effects of GO concentration (0, 0.5, 1, 3 and 5 wt%) on the electrical, mechanical, physical and biological properties of scaffolds were also evaluated. Results demonstrated that, incorporation of GO nanosheets in the PVDF matrix decreased water contact angle, while enhanced the hydrophilicity, water absorption and water flux of the scaffolds. Moreover, mechanical properties of the nanocomposite scaffolds improved in the presence of GO nanosheets. Significantly, increasing GO content up to 3 wt% enhanced tensile modulus and strength of PVDF scaffold from 8.1 ± 1.4 and 0.8 ± 0.2 MPa to 17.0 ± 3.7 and 1.4 ± 0.4 MPa, respectively. Incorporation of GO nanosheets into the PVDF scaffold simultaneously enhanced β phase fraction, piezoelectricity and electrical conductivity of all nanocomposite scaffolds. Furthermore, PVDF-GO scaffolds significantly promoted cell proliferation, compared to PVDF scaffold, depending on the GO content. Finally, PVDF-GO scaffold could easily be converted in to a nerve guidance conduit with 4 internal longitudinally aligned channels making it appropriate for the nerve regeneration applications.

Melanin incorporated electroactive and antioxidant silk fibroin nanofibrous scaffolds for nerve tissue engineering

Nerve restoration and repair in the central nervous system is complicated and requires several factors to be considered while designing the scaffolds like being bioactive as well as having neuroinductive, neuroconductive and antioxidant properties. Aligned electrospun nanofibers provide necessary guidance and topographical cues required for directing the axonal and neurite outgrowth during regeneration. Conduction of nerve impulses is a mandatory feature of a typical nerve. The neuro-conductive property can be imparted by blending the biodegradable, bioactive polymers with conductive polymers. This will provide additional features, i.e., electrical cues to the already existing topographical and bioactive cues in order to make it a more multifaceted neuroregenerative approach. Hence in the present study, we used a combination of silk fibroin and melanin for the fabrication of random and aligned electrospun nanofibrous composite scaffolds. We performed the physico-chemical characterization and also assessed their antioxidant properties. We also evaluated their neurogenic potential using human neuroblastoma cells (SH-SY5Y) for their cellular viability, proliferation, adhesion and differentiation levels. Designed nanofibrous scaffolds had adequate physical properties suitable as neural substrates to promote neuronal growth and regeneration. They stimulated the neuroblastoma cell attachment and viability indicating their biocompatible nature. Silk/melanin composite scaffolds have specifically exhibited high antioxidant nature proven by the radical scavenging activity. Additionally, the melanin incorporated aligned silk fibroin scaffolds promoted the cell differentiation into neurons and orientation along their axis. Our results confirmed the potential of melanin incorporated aligned silk fibroin scaffolds as the promising candidates for effective nerve regeneration and recovery.

Fabrication and characterization of gold nanoparticle-doped electrospun PCL/chitosan nanofibrous scaffolds for nerve tissue engineering.

In the field of nerve tissue engineering, nanofibrous scaffolds could be a promising candidate when they are incorporated with electrical cues. Unique physico-chemical properties of gold nanoparticles (AuNPs) make them an appropriate component for increasing the conductivity of scaffolds to enhance the electrical signal transfer between neural cells. The aim of this study was fabrication of AuNPs-doped nanofibrous scaffolds for peripheral nerve tissue engineering. Polycaprolactone (PCL)/chitosan mixtures with different concentrations of chitosan (0.5, 1 and 1.5) were electrospun to obtain nanofibrous scaffolds. AuNPs were synthesized by the reduction of HAuCl4 using chitosan as a reducing/stabilizing agent. A uniform distribution of AuNPs with spherical shape was achieved throughout the PCL/chitosan matrix. The UV-Vis spectrum revealed that the amount of gold ions absorbed by nanofibrous scaffolds is in direct relationship with their chitosan content. Evaluation of electrical property showed that inclusion of AuNPs significantly enhanced the conductivity of scaffolds. Finally, after 5 days of culture, biological response of Schwann cells on the AuNPs-doped scaffolds was superior to that on as-prepared scaffolds in terms of improved cell attachment and higher proliferation. It can be concluded that the prepared AuNPs-doped scaffolds can be used to promote peripheral nerve regeneration.

Electrodeless coating polypyrrole on chitosan grafted polyurethane with functionalized multiwall carbon nanotubes electrospun scaffold for nerve tissue engineering

Herein, we engineered a self-electrical stimulated double-layered nerve guidance conduit (NGC) assembled from electrospun mats with an aligned oriented inward layer covered with a random oriented outer layer. The biomimetic NGC can be achieved from chitosan grafted polyurethane with well-dispersed functionalized multiwall carbon nanotubes (fMWCNTs) nanofibrous mats after a uniform coating of polypyrrole (PPy). The structural framework of interconnected NGC exhibited cellular biomaterial interface and improved the physicochemical properties, including electrical conductivity, mechanical strength, and cytocompatibility, serving as natural hosting substrate to natural extracellular matrices (ECM) for vital roles in nerve tissue engineering. The regrowth, proliferation, and migration, of Schwann cells (S42) and the differentiation of rat pheochromocytoma cells (PC12) were greatly accelerated on the aligned oriented mats as compared to the randomly oriented mats during in vitro cell cultures. The morphology of the spontaneous outgrowth and phenotype of neurite bundles were preferentially guided along the axis of the aligned oriented nanofibers, which maintains a strong adaptability in axonal regeneration. In addition, the differentiation of PC12 cells cultured on as-fabricated NGCs were evaluated from cDNA gene expression. It is hoped that the results will contribute to the efficient application of designed NGCs and can be used in therapeutic strategies for treating injured sites and stimulate recovery from substantial damage to nerve cells.

Development of a novel porous polyvinyl formal (PVF) microfibrous scaffold for nerve tissue engineering

Tissue engineering strategies are constantly in search of novel scaffold materials that can accelerate cell adhesion, growth and proliferation. In this context, poly(vinyl formal) (PVF), a biodegradable and water absorbing polymer derived from poly(vinyl alcohol) was explored as a potential scaffold material for neural regeneration. No prior attempts have been made to fabricate electrospun fibrous scaffolds from PVF for tissue engineering applications and this forms the focus of the present study. The electrospinning conditions were optimized to obtain defect-free PVF microfibres with an average diameter of 4.4 ± 0.7 μm. The microfibres exhibited a uniform nanoporous surface and a solid core that augmented neuronal cell adhesion and neurite extension. The fibres were brittle and exhibited a 50% degradation in physiological pH and temperature. The viability of hippocampal neuronal cells cultured on the scaffold showed a progressive increase with time revealing its promise as a scaffold material for neural tissue engineering.

A facile one-step strategy for development of a double network fibrous scaffold for nerve tissue engineering

The aim of this study was to develop a novel double network scaffold composed of polycaprolactone fumarate (PCLF) and eggshell membrane (ESM) (ESM:PCLF) by using the vacuum infiltration method. Compared to ESM, the mechanical properties of double network scaffold were significantly improved, depending on the solvents applied for double network scaffold formation; acetic acid and dichloromethane. Noticeably, the toughness and strength of double network scaffold prepared using acetic acid were significantly improved compared to ESM (26.6 and 25 times, respectively) attributed to the existence of hydrophilic functional groups in acetic acid which made ESM flexible to absorb further PCLF solution. To assess the effect of double network formation on the biological behavior of ESM, the attachment, proliferation and spreading of PC12 cells cultured on the ESM:PCLF scaffolds were evaluated. Results revealed that the number of cells attached on double network ESM:PCLF scaffold were nearly similar to ESM and significantly higher than that of on the tissue culture plate (2.6 times) and PCLF film (1.7 times). It is envisioned that the offered ESM:PCLF double network scaffold might have great potential to develop the constructs for nerve regeneration.

Heparin/collagen encapsulating nerve growth factor multilayers coated aligned PLLA nanofibrous scaffolds for nerve tissue engineering.

Biomimicing topological structure of natural nerve tissue to direct axon growth and controlling sustained release of moderate neurotrophic factors are extremely propitious to the functional recovery of damaged nervous systems. In this study, the heparin/collagen encapsulating nerve growth factor (NGF) multilayers were coated onto the aligned poly-L-lactide (PLLA) nanofibrous scaffolds via a layer-by-layer (LbL) self-assembly technique to combine biomolecular signals, and physical guidance cues for peripheral nerve regeneration. Scanning electronic microscopy (SEM) revealed that the surface of aligned PLLA nanofibrous scaffolds coated with heparin/collagen multilayers became rougher and appeared some net-like filaments and protuberances in comparison with PLLA nanofibrous scaffolds. The heparin/collagen multilayers did not destroy the alignment of nanofibers. X-ray photoelectron spectroscopy and water contact angles displayed that heparin and collagen were successfully coated onto the aligned PLLA nanofibrous scaffolds and improved its hydrophilicity. Three-dimensional (3 D) confocal microscopy images further demonstrated that collagen, heparin, and NGF were not only coated onto the surface of aligned PLLA nanofibrous scaffolds but also permeated into the inner of scaffolds. Moreover, NGF presented a sustained release for 2 weeks from aligned nanofibrous scaffolds coated with 5.5 bilayers or above and remained good bioactivity. The heparin/collagen encapsulating NGF multilayers coated aligned nanofibrous scaffolds, in particular 5.5 bilayers or above, was more beneficial to Schwann cells (SCs) proliferation and PC12 cells differentiation as well as the SC cytoskeleton and neurite growth along the direction of nanofibrous alignment compared to the aligned PLLA nanofibrous scaffolds. This novel scaffolds combining sustained release of bioactive NGF and aligned nanofibrous topography presented an excellent potential in peripheral nerve regeneration. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1900-1910, 2017.