Anisotropic Scaffolds Research Articles

Anisotropic electrospun honeycomb polycaprolactone scaffolds: Elaboration, morphological and mechanical properties

Tissue engineering technology requires porous scaffolds, based on biomaterials, which have to mimic as closely as possible the morphological and anisotropic mechanical properties of the native tissue to substitute. Anisotropic fibrous scaffolds fabricated by template-assisted electrospinning are investigated in this study. Fibers of electrospun Polycaprolactone (PCL) were successfully arranged spatially into honeycomb structures by using well-shaped 3D micro-architected metal collectors. Fibrous scaffolds present 2 × 4 mm2 wide elementary patterns with low and high fiber density areas. Distinct regions of the honeycomb patterns were analyzed through SEM images revealing different fiber diameters with specific fiber orientation depending on the regions of interest. Tensile test experiments were carried out with an optical observation of the local deformation at the pattern scale, allowing the determination and analysis, at small and large deformation, of the axial and transverse local strains. The honeycomb patterned mats showed significantly different mechanical properties along the two orthogonal directions probing an anisotropic ratio of 4.2. Stress relaxation test was performed on scaffolds at 15% of strain. This measurement pointed out the low contribution of the viscosity of about 20% in the mechanical response of the scaffold. An orthotropic linear elastic model was consequently proposed to characterize the anisotropic behavior of the produced patterned membranes. This new versatile method to produce architected porous materials, adjustable to several polymers and structures, will provide appealing benefits for soft regenerative medicine application and the development of custom-made scaffolds.

Optimizing Bifurcated Channels within an Anisotropic Scaffold for Engineering Vascularized Oriented Tissues.

Despite progress in engineering both vascularized tissues and oriented tissues, the fabrication of 3D vascularized oriented tissues remains a challenge due to an inability to successfully integrate vascular and anisotropic structures that can support mass transfer and guide cell alignment, respectively. More importantly, there is a lack of an effective approach to guiding the scaffold design bearing both structural features. Here, an approach is presented to optimize the bifurcated channels within an anisotropic scaffold based on oxygen transport simulation and biological experiments. The oxygen transport simulation is performed using the experimentally measured effective oxygen diffusion coefficient and hydraulic permeability of the anisotropic scaffolds, which are also seeded with muscle precursor cells and cultured in a custom-made perfusion bioreactor. Symmetric bifurcation model is used as fractal unit to design the channel network based on biomimetic principles. The bifurcation level of channel network is further optimized based on the oxygen transport simulation, which is then validated by DNA quantification assay and pimonidazole immunostaining. This study provides a practical guide to optimizing bifurcated channels in anisotropic scaffolds for oriented tissue engineering.

Development of orthophosphosilicate glass/poly(lactic acid) composite anisotropic scaffolds for simultaneous reconstruction of bone quality and quantity.

Reconstruction of organ‐specific architecture is necessary to recover the original organ function. The anisotropic structure of bone tissue is strongly related to the collagen fibril alignment and bone apatite crystal direction. Bone regeneration indicates following two main process; first, restoration of bone mineral density (BMD; bone quantity), and second, restoring bone apatite c‐axis orientation (bone quality). In addition to BMD, bone quality is the most important factor among bone mechanical properties. Recovery of the original bone function requires development of novel scaffolds with simultaneous reconstruction of bone quality and quantity. Herein, novel orthophosphosilicate glass (PSG)/poly(lactic acid) composite anisotropic scaffolds were developed to control cell alignment and enhance bone formation, which are important for the simultaneous reconstruction of bone quality and quantity. The strategy to control cell alignment and bone formation involved designing anisotropic scaffolds in combination with the release of therapeutic ions by PSGs. The morphology of fibrous scaffolds containing PSGs was quantitatively designed using electrospinning. This successfully modulated cell alignment and subsequent bone apatite c‐axis orientation along the fiber‐oriented direction. The released silicate and Mg2+ ions from PSGs in scaffolds improved cell adhesion, proliferation, and calcification. To best of our knowledge, this is the first report demonstrating that the anisotropic scaffolds containing bioactive glasses regenerate bone tissues with simultaneous reconstruction of bone quality and quantity via stimulating osteoblasts by inorganic ions and designing morphology of scaffolds.

Rapidly formed stable and aligned dense collagen gels seeded with Schwann cells support peripheral nerve regeneration

Objective. Gel aspiration-ejection (GAE) has recently been developed for the rapid production of dense, anisotropic collagen gel scaffolds with adjustable collagen fibrillar densities. In this study, a GAE system was applied to produce aligned Schwann cells within a type-1 collagen matrix to generate GAE-engineered neural tissues (GAE-EngNT) for potential nerve tissue engineering applications. Approach. The stability and mechanical properties of the constructs were investigated along with the viability, morphology and distribution of Schwann cells. Having established the methodology to construct stable robust Schwann cell-loaded engineered neural tissues using GAE (GAE-EngNTs), the potential of these constructs in supporting and guiding neuronal regeneration, was assessed both in vitro and in vivo. Main results. Dynamic mechanical analysis strain and frequency sweeps revealed that the GAE-EngNT produced via cannula gauge number 16 G (∼1.2 mm diameter) exhibited similar linear viscoelastic behaviors to rat sciatic nerves. The viability and alignment of seeded Schwann cells in GAE-EngNT were maintained over time post GAE, supporting and guiding neuronal growth in vitro with an optimal cell density of 2.0 × 106 cells ml−1. An in vivo test of the GAE-EngNTs implanted within silicone conduits to bridge a 10 mm gap in rat sciatic nerves for 4 weeks revealed that the constructs significantly promoted axonal regeneration and vascularization across the gap, as compared with the empty conduits although less effective regeneration compared with the autograft groups. Significance. Therefore, this is a promising approach for generating anisotropic and robust engineered tissue which can be used with Schwann cells for peripheral nerve repair.

Engineered dual-scale poly (ε-caprolactone) scaffolds using 3D printing and rotational electrospinning for bone tissue regeneration

Large bone defects due to trauma or disease present a significant clinical challenge with limited efficacy of current therapies. A key aim is to develop biomimetic scaffolds that reflect the native tissue structure with 3D printing being an important enabling technology. However, the incorporation of multiple length scales and anisotropic features, mimicking the native architecture, is difficult with current processes. In this study, we propose a simple and versatile hybrid printing process using a screw-assisted additive manufacturing technique combined with rotational electrospinning to fabricate dual-scale anisotropic scaffolds. 3D microscale porous polycaprolactone (PCL) structures with highly aligned nanoscale fibres were successfully produced layer-by-layer. The scaffolds were morphological, mechanical and biological characterised. Human adipose-derived stem cells (hADSCs) were seeded on the hybrid scaffold to evaluate the effects of structural and anisotropic topographic cues on cell attachment, proliferation and osteogenesis differentiation. Results show that the 3D printed microscale structures have uniform and well-defined geometries and the alignment of nanoscale electrospun fibres increases by increasing the electrospinning rotational velocity. Mechanical results show that there is no significant difference between 3D printed scaffolds with or without electrospun meshes. In vitro results show higher cell seeding efficiency and proliferation in dual-scale scaffolds with high density electrospun meshes. A more stretched and elongated cell morphology was observed in aligned nanofibre scaffolds showing higher anisotropic cytoskeletal organization than 3D printed PCL scaffolds without electrospun meshes. The dual-scale scaffolds present improved overall osteogenic markers expressions (COL-1, ALP and OCN). However, no statistical difference between normalised osteogenic marker expressions were observed between dual-scale scaffolds and 3D printed scaffolds. This might be attributed to the poor bioactivity of the substrate material, PCL, suggesting topographical cues might not be sufficient to stimulate cell fate towards to an osteogenic linage. The results suggest that the proposed fabrication strategy is a promising approach for the design of novel bone scaffolds to modulate cell fates by integrating the topographic cue reported in this paper with biochemical cues associated to the use of more bioactive materials.

An “all-in-one” scaffold targeting macrophages to direct endogenous bone repair in situ

Scaffolds for tissue repair are designed in an increasingly complicated manner to meet multi-facet biological needs during the healing process. However, overly sophisticated design, especially the use of multiple components and delivery of exogenous cells, hampers the bench-to-bedside translation. Here, a multi-functional – yet mono-compositional – bioactive scaffold is devised to mediate the full-range, endogenous bone repair. Based on immunoactivity screening, a chemically-modified glucomannan polysaccharide is selected and processed into an anisotropic porous scaffold, which accurately stimulates macrophages to produce pro-regenerative cytokines. These cytokines effectively enhance the recruitment (“R”) and induced osteogenesis (“IO”) of the bone progenitor cells in situ. Meanwhile, the anisotropic porosity and carbohydrate signal of the scaffold facilitate differential adhesion (“A”) and distribution (“D”) of the macrophages and bone progenitor cells – enabling the former's accumulation at the surface while encouraging the latter's infiltration into the scaffold. Implanted in a rat calvarial defect model, this “RADIO” system effectively promotes healing over 12 weeks, with the obvious formation of hard callus through the scaffold. In summary, RADIO integrates multiple functions into one single scalable system (“all-in-one”) to govern the dynamic bone-repair process, by harnessing the power of host macrophages. RADIO represents an open platform to solving the long-lasting complexity-versus-simplicity dilemma in biomaterials design. Statement of SignificanceBiomaterials as versatile tools for tissue repair are becoming increasingly complicated, yet overly sophisticated design – especially the use of multiple components, exogenous cells, and overdosed growth factors – hampers their clinical application. The pre-requisite for designing a successful integrative scaffold is to identify an inherent biological target responding to biomaterial signals, thereby efficiently and safely promoting tissue repair via the endogenous healing capability instead of extra multifarious biochemical components. For bone regeneration, the pivotal regulator is macrophages. Through activating host macrophages, our single-component scaffold system coordinates the entire bone regenerative cascade in situ and induces successful bone regeneration in a calvarial defect model. This scaffold represents a scalable and multi-functional approach to effectively simplify the sophisticated design in regenerative medicine.

Optimizing Anisotropic Polyurethane Scaffolds to Mechanically Match with Native Myocardium.

Biodegradable cardiac patch is desirable to possess mechanical properties mimicking native myocardium for heart infarction treatment. We fabricated a series of anisotropic and biodegradable polyurethane porous scaffolds via thermally induced phase separation (TIPS) and tailored their mechanical properties by using various polyurethanes with different soft segments and varying polymer concentrations. The uniaxial mechanical properties, suture retention strength, ball-burst strength, and biaxial mechanical properties of the anisotropic porous scaffolds were optimized to mechanically match native myocardium. The optimal anisotropic scaffold had a ball burst strength (20.7 ± 1.5 N) comparable to that of native porcine myocardium (20.4 ± 6.0 N) and showed anisotropic behavior close to biaxial stretching behavior of the native porcine myocardium. Furthermore, the optimized porous scaffold was combined with a porcine myocardium-derived hydrogel to form a biohybrid scaffold. The biohybrid scaffold showed morphologies similar to the decellularized porcine myocardial matrix. This combination did not affect the mechanical properties of the synthetic scaffold alone. After in vivo rat subcutaneous implantation, the biohybrid scaffolds showed minimal immune response and exhibited higher cell penetration than the polyurethane scaffold alone. This biohybrid scaffold with biomimetic mechanics and good tissue compatibility would have great potential to be applied as a biodegradable acellular cardiac patch for myocardial infarction treatment.

Nonmulberry Silk Based Ink for Fabricating Mechanically Robust Cardiac Patches and Endothelialized Myocardium-on-a-Chip Application.

Bioprinting holds great promise towards engineering functional cardiac tissue constructs for regenerative medicine and as drug test models. However, it is highly limited by the choice of inks that require maintaining a balance between the structure and functional properties associated with the cardiac tissue. In this regard, we have developed a novel and mechanically robust biomaterial-ink based on non-mulberry silk fibroin protein. The silk-based ink demonstrated suitable mechanical properties required in terms of elasticity and stiffness (~40 kPa) for developing clinically relevant cardiac tissue constructs. The ink allowed the fabrication of stable anisotropic scaffolds using a dual crosslinking method, which were able to support formation of aligned sarcomeres, high expression of gap junction proteins as connexin-43, and maintain synchronously beating of cardiomyocytes. The printed constructs were found to be non-immunogenic in vitro and in vivo. Furthermore, delving into an innovative method for fabricating a vascularized myocardial tissue-on-a-chip, the silk-based ink was used as supporting hydrogel for encapsulating human induced pluripotent stem cell derived cardiac spheroids (hiPSC-CSs) and creating perfusable vascularized channels via an embedded bioprinting technique. We confirmed the ability of silk-based supporting hydrogel towards maturation and viability of hiPSC-CSs and endothelial cells, and for applications in evaluating drug toxicity.

Valve leaflet‐inspired elastomeric scaffolds with tunable and anisotropic mechanical properties

Fibrous scaffolds, which can mimic the elastic and anisotropic mechanical properties of native tissues, hold great promise in recapitulating the native tissue microenvironment. We previously fabricated electrospun fibrous scaffolds made of hybrid synthetic elastomers (poly(1,3‐diamino‐2‐hydroxypropane‐co‐glycerol sebacate)‐co‐poly (ethylene glycol) (APS‐co‐PEG) and polycaprolactone (PCL)) to obtain uniaxial mechanical properties similar to those of human aortic valve leaflets. However, conventional electrospinning process often yields scaffolds with random alignment, which fails to recreate the anisotropic nature of most of the soft tissues such as native heart valves. Inspired by the structure of native valve leaflet, we designed a novel valve leaflet‐inspired ring‐shaped collector to modulate the electrospun fiber alignment and studied the effect of polymer formulation (PEG amount [mole %] in APS‐co‐PEG; ratio between APS‐co‐PEG and PCL; and total polymer concentration) in tuning the biaxial mechanical properties of the fibrous scaffolds. The fibrous scaffolds collected on the ring‐shaped collector displayed anisotropic biaxial mechanical properties, suggesting that their biaxial mechanical properties are closely associated with the fiber alignment in the scaffold. Additionally, the scaffold stiffness was easily tuned by changing the composition and concentration of the polymer blend. Human valvular interstitial cells (hVICs) cultured on these anisotropic scaffolds displayed aligned morphology as instructed by the fiber alignment. Overall, we generated a library of biologically relevant fibrous scaffolds with tunable mechanical properties, which will guide the cellular alignment.

Inverse Nacre-like Epoxy-Graphene Layered Nanocomposites with Integration of High Toughness and Self-Monitoring

Summary Epoxy nanocomposites have many promising applications in the fields of aerospace and aeronautics, as well as many others. Achieving tough epoxy nanocomposites remains a great challenge, however. One inspiration for improving the mechanical properties of epoxy nanocomposites is nacre, which has remarkable fracture toughness for its layered “brick-and-mortar” architecture. Inspired by this, we fabricated a lamellar graphene scaffold by the freeze-casting technique. An alternating-layered epoxy-graphene nanocomposite was made by infiltrating epoxy into this graphene scaffold. As our epoxy-graphene nanocomposite consists of ∼99 wt % organic epoxy, in contrast to nacre containing ∼96 wt % inorganic aragonite, we call it an “inverse nacre-like” epoxy-graphene layered nanocomposite. It exhibits exceptional fracture toughness, 3.61 times that of pure epoxy, and demonstrates anisotropic conductivity due to the anisotropic graphene scaffold, which can be used to detect cracks. Our bioinspired strategy provides a promising approach to combine excellent mechanical properties with functional properties to fabricate high-performance nanocomposites.

Anisotropic Cryostructured Collagen Scaffolds for Efficient Delivery of RhBMP-2 and Enhanced Bone Regeneration.

In the treatment of bone non-unions, an alternative to bone autografts is the use of bone morphogenetic proteins (BMPs), e.g., BMP–2, BMP–7, with powerful osteoinductive and osteogenic properties. In clinical settings, these osteogenic factors are applied using absorbable collagen sponges for local controlled delivery. Major side effects of this strategy are derived from the supraphysiological doses of BMPs needed, which may induce ectopic bone formation, chronic inflammation, and excessive bone resorption. In order to increase the efficiency of the delivered BMPs, we designed cryostructured collagen scaffolds functionalized with hydroxyapatite, mimicking the structure of cortical bone (aligned porosity, anisotropic) or trabecular bone (random distributed porosity, isotropic). We hypothesize that an anisotropic structure would enhance the osteoconductive properties of the scaffolds by increasing the regenerative performance of the provided rhBMP–2. In vitro, both scaffolds presented similar mechanical properties, rhBMP–2 retention and delivery capacity, as well as scaffold degradation time. In vivo, anisotropic scaffolds demonstrated better bone regeneration capabilities in a rat femoral critical-size defect model by increasing the defect bridging. In conclusion, anisotropic cryostructured collagen scaffolds improve bone regeneration by increasing the efficiency of rhBMP–2 mediated bone healing.

Use of Aligned Microscale Sacrificial Fibers in Creating Biomimetic, Anisotropic Poly(glycerol sebacate) Scaffolds.

Poly(glycerol sebacate) (PGS) is a biocompatible, biodegradable elastomer that has been shown promise as a scaffolding material for tissue engineering; it is still challenging, however, to produce anisotropic scaffolds by using a thermoset polymer, such as PGS. Previously, we have used aligned sacrificial poly(vinyl alcohol) (PVA) fibers to help produce an anisotropic PGS membrane; a composite membrane, formed by embedding aligned PVA fibers in PGS prepolymer, was subjected to curing and subsequent PVA removal, resulting in aligned grooves and cylindrical pores on the surface of and within the membrane, respectively. PVA, however, appeared to react with PGS during its curing, altering the mechanical characteristics of PGS. In this study, aligned sacrificial fibers made of polylactide (PLA) were used instead. Specifically, PLA was blend-electrospun with polyethylene oxide to increase the sacrificial fiber diameter, which in turn increased the size of the grooves and cylindrical pores. The resultant PGS membrane was shown to be in vitro cyto-compatible and mechanically anisotropic. The membrane’s Young’s modulus was 1–2 MPa, similar to many soft tissues. In particular, the microscale grooves on the membrane surface were found to be capable of directing cell alignment. Finally, based on the same approach, we fabricated a biomimetic, anisotropic, PGS tubular scaffold. The compliance of the tubular scaffold was comparable to native arteries and in the range of 2% to 8% per 100 mmHg, depending on the orientations of the sacrificial fibers. The anisotropic PGS tubular scaffolds can potentially be used in vascular tissue engineering.

Anisotropy Properties of Tissues: A Basis for Fabrication of Biomimetic Anisotropic Scaffolds for Tissue Engineering

Tissue engineering has been a subject of extensive scientific exploration in the last two decades making gradual inroads into clinical studies as well. Along with regenerative cells and growth factors, biomaterial scaffolds are integral to the development of a tissue engineered construct. It is now appreciated that scaffolds should mimic the target tissue properties intimately in order to provide a micro-environment milieu that allows the seeded cells to differentiate into the desired tissue. Even from a structural viewpoint, mismatch between scaffold and native matrix properties can cause cell necrosis through mechanisms such as stress shielding. One of the key properties of most body tissues is that they exhibit anisotropy. However, most fabrication methods generate isotropic scaffolds and require specific modifications to produce anisotropic scaffolds. In the last decade, the advent of additive manufacturing and bioprinting has provided facile tools to fabricate scaffolds with desired anisotropy. On the other hand, a biomimetic scaffold can be designed only when target tissue anisotropy is well known to the tissue engineer. This review presents an overview of the anisotropic properties of different tissues, which will be critical for developing biomimetic engineered constructs. The traditional anatomical records do not adequately present these properties from the perspective of designing tissue engineering scaffolds. Subsequently, present state-of-the art in development of anisotropic scaffolds as well as tissue constructs using different conventional and emerging fabrication techniques is discussed. It is expected that the readers will obtain a comprehensive reference on the research area by examining these two aspects juxtaposed to each other and gain key trends for fabrication of anisotropic scaffolds, plausibly with improved regenerative outcomes.

4D anisotropic skeletal muscle tissue constructs fabricated by staircase effect strategy

Like the morphology of native tissue fiber arrangement (such as skeletal muscle), unidirectional anisotropic scaffolds are highly desired as a means to guide cell behavior in anisotropic tissue engineering. In contrast, contour-like staircases exhibit directional topographical cues and are judged as an inevitable defect of fused deposition modeling (FDM). In this study, we will translate this staircase defect into an effective bioengineering strategy by integrating FDM with surface coating technique (FCT) to investigate the effect of topographical cues on regulating behaviors of human mesenchymal stem cells (hMSCs) toward skeletal muscle tissues. This integrated approach serves to fabricate shape-specific, multiple dimensional, anisotropic scaffolds using different biomaterials. 2D anisotropic scaffolds, first demonstrated with different polycaprolactone concentrations herein, efficiently direct hMSC alignment, especially when the scaffold is immobilized on a support ring. By surface coating the polymer solution inside FDM-printed sacrificial structures, 3D anisotropic scaffolds with thin wall features are developed and used to regulate seeded hMSCs through a self-established rotating bioreactor. Using layer-by-layer coating, along with a shape memory polymer, smart constructs exhibiting shape fix and recovery processes are prepared, bringing this study into the realm of 4D printing. Immunofluorescence staining and real-time quantitative polymerase chain reaction analysis confirm that the topographical cues created via FCT significantly enhance the expression of myogenic genes, including myoblast differentiation protein-1, desmin, and myosin heavy chain-2. We conclude that there are broad application potentials for this FCT strategy in tissue engineering as many tissues and organs, including skeletal muscle, possess highly organized and anisotropic extracellular matrix components.

3D printing algorithm of anisotropic biological scaffold with oxidized nanocellulose and gelatin

Tissue scaffolds need to have anisotropic mechanical properties and a porous structure to meet the needs of different tissues and organs. This report presents results of a study on an especially-designed 3D printing method with oxidized nanocellulose and gelatin, analyzes the servo principle of pneumatic condensing extrusion 3D printer, and proposes a hexagonal algorithm. For the purpose of this study, a printing process file was written by G code, physical and mechanical performance of the 3D scaffolds was evaluated with Solidworks simulation, the porous structure and pressure-pull performance of the printed 3D scaffolds was observed by SEM, and experiments were conducted to measure their bio-compatibility. The study draws the conclusion that scaffolds thus printed have a highly porous structure and anisotropic mechanical properties.

Cylindrical Layered Bone Scaffolds with Anisotropic Mechanical Properties as Potential Drug Delivery Systems.

3D cylindrical layered scaffolds with anisotropic mechanical properties were prepared according to a new and simple method, which involves gelatin foaming, deposition of foamed strips, in situ crosslinking, strip rolling and lyophilization. Different genipin concentrations were tested in order to obtain strips with different crosslinking degrees and a tunable stability in biological environment. Before lyophilization, the strips were curled in a concentric structure to generate anisotropic spiral-cylindrical scaffolds. The scaffolds displayed significantly higher values of stress at break and of the Young modulus in compression along the longitudinal than the transverse direction. Further improvement of the mechanical properties was achieved by adding strontium-substituted hydroxyapatite (Sr-HA) to the scaffold composition and by increasing genipin concentration. Moreover, composition modulated also water uptake ability and degradation behavior. The scaffolds showed a sustained strontium release, suggesting possible applications for the local treatment of abnormally high bone resorption. This study demonstrates that assembly of layers of different composition can be used as a tool to obtain scaffolds with modulated properties, which can be loaded with drugs or biologically active molecules providing properties tailored upon the needs.

Analysis of biomechanical behavior of 3D printed mandibular graft with porous scaffold structure designed by topological optimization

BackgroundOur long-term goal is to design and manufacture a customized graft with porous scaffold structure for repairing large mandibular defects using topological optimization and 3D printing technology. The purpose of this study is to characterize the mechanical behavior of 3D printed anisotropic scaffolds as bone analogs by fused deposition modeling (FDM).MethodsCone beam computed tomography (CBCT) images were used to reconstruct a 3D mandible and finite element models. A virtual sectioned-block of the mandible was used as the control group and the trabecular portion of the block was modified by topological optimization methods as experimental groups. FDM (FDM) printed samples at 0, 45 and 90 degrees with Poly-lactic acid (PLA) material under a three-point bending test. Finite element analysis was also used to validate the data obtained from the physical model tests.ResultsThe ultimate load, yield load, failure deflection, yield deflection, stress, strain distribution, and porosity of scaffold structures were compared. The results show that the topological optimized graft had the best mechanical properties.ConclusionsThe results from mechanical tests on physical models and numerical simulations from this study show a great potential for topological optimization and 3D printing technology to be served in design and rapidly manufacturing of artificial porous grafts.

Tissue Engineered Neural Constructs Composed of Neural Precursor Cells, Recombinant Spidroin and PRP for Neural Tissue Regeneration

We have designed a novel two-component matrix (SPRPix) for the encapsulation of directly reprogrammed human neural precursor cells (drNPC). The matrix is comprised of 1) a solid anisotropic complex scaffold prepared by electrospinning a mixture of recombinant analogues of the spider dragline silk proteins – spidroin 1 (rS1/9) and spidroin 2 (rS2/12) - and polycaprolactone (PCL) (rSS-PCL), and 2) a “liquid matrix” based on platelet-rich plasma (PRP). The combination of PRP and spidroin promoted drNPC proliferation with the formation of neural tissue organoids and dramatically activated neurogenesis. Differentiation of drNPCs generated large numbers of βIII-tubulin and MAP2 positive neurons as well as some GFAP-positive astrocytes, which likely had a neuronal supporting function. Interestingly the SPRPix microfibrils appeared to provide strong guidance cues as the differentiating neurons oriented their processes parallel to them. Implantation of the SPRPix matrix containing human drNPC into the brain and spinal cord of two healthy Rhesus macaque monkeys showed good biocompatibility: no astroglial and microglial reaction was present around the implanted construct. Importantly, the human drNPCs survived for the 3 month study period and differentiated into MAP2 positive neurons. Tissue engineered constructs based on SPRPix exhibits important attributes that warrant further examination in spinal cord injury treatment.

Bioadhesive anisotropic nanogrooved microfibers directing three-dimensional neurite extension.

Neurodegenerative diseases and acute nerve injuries are becoming global clinical problems. Engineering three-dimensional (3D), anisotropic neural cellular structures in vitro is therefore desirable in the regenerative medicine research community. Here, we present, for the first time, a single-step, facile but delicate, fabrication of a 3D macroporous microfibrous scaffold with both anisotropic nanogrooved topography and in situ functionalization with a mussel inspired bioadhesive, poly(norepinephrine) (pNE). Specifically, immiscible blends of polycaprolactone (PCL) and polyethylene oxide (PEO) were electrospun into a grounded coagulation bath containing the precursor of pNE. Upon jet entrance in the bath, both phase-separation-driven longitudinal nanotopography and in situ pNE surface functionalization were introduced on individual microfibers that were packed into a 3D macroporous structure. The resulting scaffold significantly promoted 3D neurite extension capacity, 8-fold higher neurite extension over the isotropic counterpart, demonstrating that such a scaffold has great promise in 3D neural cell culture for nerve tissue modelling and engineering.

Microarchitecture and Radiological Flow Pattern of Cocoon-like Nanocellulose Hydrogels

Abstract This work investigates fluid mechanics and structural properties inside bacterial cellulose hydrogels in from of macroscopic anisotropic ellipsoid body cocoon-like 3D scaffolds. The hydrogels were evaluated by injections of iopamidol, a radiopaque medium used in medical radiological procedures. Results showed that hydromechanical behavior of BC hydrogels may be used to characterize its internal structure. It is suggested that internal barriers and flow distribution anisotropy may be used to develop novel therapeutic applications based on the tissue engineered multifunctional possibilities of the biomaterial.