Nerve Guidance Research Articles

DNA origami scaffold promoting nerve guidance and regeneration.

Self-assembly of biological elements into biomimetic cargo carriers for targeting and delivery is a promising approach. However, it still holds practical challenges. We developed a functionalization approach of DNA origami (DO) nanostructures with neuronal growth factor (NGF) for manipulating neuronal systems. NGF bioactivity and its interactions with the neuronal system were demonstrated in vitro and in vivo models. The DO elements fabricated by molecular self-assembly have manipulated the surrounding environment through static spatially and temporally controlled presentation of ligands to the cell surface receptors. Our data showed effective bioactivity in differentiating PC12 cells in vitro. Furthermore, the DNA origami NGF (DON) affected the growth directionality and spatial capabilities of dorsal root ganglion neurons in culture by introducing a chemotaxis effect along a gradient of functionalized DO structures. Finally, we showed that these elements provide enhanced axonal regeneration in a rat sciatic nerve injury model in vivo. This study is a proof of principle for the functionality of DO in neuronal manipulation and regeneration. The approach proposed here, of an engineered platform formed out of programmable nanoscale elements constructed of DO, could be extended beyond the nervous system and revolutionize the fields of regenerative medicine, tissue engineering, and cell biology.

Innovations in Peripheral Nerve Regeneration.

The field of peripheral nerve regeneration is a dynamic and rapidly evolving area of research that continues to captivate the attention of neuroscientists worldwide. The quest for effective treatments and therapies to enhance the healing of peripheral nerves has gained significant momentum in recent years, as evidenced by the substantial increase in publications dedicated to this field. This surge in interest reflects the growing recognition of the importance of peripheral nerve recovery and the urgent need to develop innovative strategies to address nerve injuries. In this context, this article aims to contribute to the existing knowledge by providing a comprehensive review that encompasses both biomaterial and clinical perspectives. By exploring the utilization of nerve guidance conduits and pharmacotherapy, this article seeks to shed light on the remarkable advancements made in the field of peripheral nerve regeneration. Nerve guidance conduits, which act as artificial channels to guide regenerating nerves, have shown promising results in facilitating nerve regrowth and functional recovery. Additionally, pharmacotherapy approaches have emerged as potential avenues for promoting nerve regeneration, with various therapeutic agents being investigated for their neuroprotective and regenerative properties. The pursuit of advancing the field of peripheral nerve regeneration necessitates persistent investment in research and development. Continued exploration of innovative treatments, coupled with a deeper understanding of the intricate processes involved in nerve regeneration, holds the promise of unlocking the complete potential of these groundbreaking interventions. By fostering collaboration among scientists, clinicians, and industry partners, we can accelerate progress in this field, bringing us closer to the realization of transformative therapies that restore function and quality of life for individuals affected by peripheral nerve injuries.

Constructing Nerve Guidance Conduit using dECM‐Doped Conductive Hydrogel to Promote Peripheral Nerve Regeneration

AbstractPeripheral nerve injury often leads to the loss of neurological functions due to the slow regeneration rate and inefficient functional reconstruction. Current clinical treatments using nerve guidance conduits (NGCs) still face challenges in providing a biomimetic microenvironment to promote nerve repair. Herein, decellularized extracellular matrix (dECM) is obtained from porcine Achilles tendon and crosslinked with 3‐amino‐4‐methoxybenzoic acid grafted gelatin (PAMB‐G) to obtain conductive hydrogels. Then, a novel nerve guidance conduit is developed by assembling poly(vinyl alcohol) (PVA) conduit and conductive ECM@PAMB‐G hydrogel. This bioengineered ECM@PAMB‐G/PVA conduit demonstrated excellent cytocompatibility, electrical conductivity, mechanical properties, and biodegradability. In vitro experiments confirmed that the ECM@PAMB‐G hydrogel significantly promotes the proliferation and migration of PC12 cells and primary Schwann cells, as well as the growth of dorsal root ganglion (DRG) axons. Furthermore, in vivo studies in a rat sciatic nerve model exhibited improvements in axonal regeneration, Schwann cell migration, myelin sheath formation, and functional recovery mediated by the ECM@PAMB‐G/PVA conduit. This work demonstrates the synergistic effects of extracellular matrix and electrical cues in enhancing peripheral nerve regeneration. The ECM@PAMB‐G/PVA nerve guidance conduit shows potential as an alternative to autografts for supporting peripheral nerve reconstruction.

A Gelatin/Alginate Double Network Hydrogel Nerve Guidance Conduit Fabricated by a Chemical-Free Gamma Radiation for Peripheral Nerve Regeneration.

Nerve guidance conduits (NGCs) are widely developed using various materials for the functional repair of injured or diseased peripheral nerves. Especially, hydrogels are considered highly suitable for the fabrication of NGCs due to their beneficial tissue-mimicking characteristics (e.g., high water content, softness, and porosity). However, the practical applications of hydrogel-based NGCs are hindered due to their poor mechanical properties and complicated fabrication processes. To bridge this gap, a novel double-network (DN) hydrogel using alginate and gelatin by a two-step crosslinking process involving chemical-free gamma irradiation and ionic crosslinking, is developed. DN hydrogels (1% alginate and 15% gelatin), crosslinked with 30 kGy gamma irradiation and barium ions, exhibit substantially improved mechanical properties, including tensile strength, elastic modulus, and fracture stain, compared to single network (SN) gelatin hydrogels. Additionally, the DN hydrogel NGC exhibits excellent kink resistance, mechanical stability to successive compression, suture retention, and enzymatic degradability. In vivo studies with a sciatic defect rat model indicate substantially improved nerve function recovery with the DN hydrogel NGC compared to SN gelatin and commercial silicone NGCs, as confirm footprint analysis, electromyography, and muscle weight measurement. Histological examination reveals that, in the DN NGC group, the expression of Schwann cell and neuronal markers, myelin sheath, and exon diameter are superior to the other controls. Furthermore, the DN NGC group demonstrates increased muscle fiber formation and reduced fibrotic scarring. These findings suggest that the mechanically robust, degradable, and biocompatible DN hydrogel NGC can serve as a novel platform for peripheral nerve regeneration and other biomedical applications, such as implantable tissue constructs.

A novel flexible nerve guidance conduit promotes nerve regeneration while providing excellent mechanical properties.

JOURNAL/nrgr/04.03/01300535-202507000-00029/figure1/v/2024-09-09T124005Z/r/image-tiff Autografting is the gold standard for surgical repair of nerve defects > 5 mm in length; however, autografting is associated with potential complications at the nerve donor site. As an alternative, nerve guidance conduits may be used. The ideal conduit should be flexible, resistant to kinks and lumen collapse, and provide physical cues to guide nerve regeneration. We designed a novel flexible conduit using electrospinning technology to create fibers on the innermost surface of the nerve guidance conduit and employed melt spinning to align them. Subsequently, we prepared disordered electrospun fibers outside the aligned fibers and helical melt-spun fibers on the outer wall of the electrospun fiber lumen. The presence of aligned fibers on the inner surface can promote the extension of nerve cells along the fibers. The helical melt-spun fibers on the outer surface can enhance resistance to kinking and compression and provide stability. Our novel conduit promoted nerve regeneration and functional recovery in a rat sciatic nerve defect model, suggesting that it has potential for clinical use in human nerve injuries.

Influence of Magnesium Degradation on Schwannoma Cell Responses to Nerve Injury Using an In Vitro Injury Model.

Nerve guidance conduits for peripheral nerve injuries can be improved using bioactive materials such as magnesium (Mg) and its alloys, which could provide both structural and trophic support. Therefore, we investigated whether exposure to Mg and Mg-1.6wt%Li thin films (Mg/Mg-1.6Li) would alter acute Schwann cell responses to injury. Using the RT4-D6P2T Schwannoma cell line (SCs), we tested extracts from freeze-killed cells (FKC) and nerves (FKN) as in vitro injury stimulants. Both FKC and FKN induced SC release of the macrophage chemoattractant protein 1 (MCP-1), a marker of the repair SC phenotype after injury. Next, FKC-stimulated cells exposed to Mg/Mg-1.6Li reduced MCP-1 release by 30%, suggesting that these materials could have anti-inflammatory effects. Exposing FKC-treated cells to Mg/Mg-1.6Li reduced the gene expression of the nerve growth factor (NGF), glial cell line-derived neurotrophic factor (GDNF), and myelin protein zero (MPZ), but not the p75 neurotrophin receptor. In the absence of FKC, Mg/Mg-1.6Li treatment increased the expression of NGF, p75, and MPZ, which can be beneficial to nerve regeneration. Thus, the presence of Mg can differentially alter SCs, depending on the microenvironment. These results demonstrate the applicability of this in vitro nerve injury model, and that Mg has wide-ranging effects on the repair SC phenotype.

Dual-layer conduit containing VEGF-A – Transfected Schwann cells promotes peripheral nerve regeneration via angiogenesis

Peripheral nerve injuries (PNIs) can cause neuropathies and significantly affect the patient's quality of life. Autograft transplantation is the gold standard for conventional treatment; however, its application is limited by nerve unavailability, size mismatch, and local tissue adhesion. Tissue engineering, such as nerve guidance conduits, is an alternative and promising strategy to guide nerve regeneration for peripheral nerve repair; however, only a few conduits could reach the high repair efficiency of autografts. The healing process of PNI is frequently accompanied by not only axonal and myelination regeneration but also angiogenesis, which initializes nerve regeneration through vascular endothelial growth factor A (VEGF-A). In this study, a composite nerve conduit with a poly (lactic-co-glycolic acid) (PLGA) hollow tube as the outer layer and gelatin methacryloyl (GelMA) encapsulated with VEGF-A transfected Schwann cells (SCs) as the inner layer was established to evaluate its promising ability for peripheral nerve repair. A rat model of peripheral nerve defect was used to examine the efficiency of PLGA/GelMA-SC (VA) conduits, whereas autograft, PLGA, PLGA/GelMA, and PLGA/GelMA-SC (NC) were used as controls. VEGF-A-transfected SCs can provide a stable source for VEGF-A secretion. Furthermore, encapsulation in GelMA cannot only promote proliferation and tube formation of human umbilical vein endothelial cells but also enhance dorsal root ganglia and neuronal cell extension. Previous animal studies have demonstrated that the regenerative effects of PLGA/GelMA-SC (VA) nerve conduit were similar to those of autografts and much better than those of other conduits. These findings indicate that combination of VEGF-A-overexpressing SCs and PLGA/GelMA conduit-guided peripheral nerve repair provides a promising method that enhances angiogenesis and regeneration during nerve repair. Statement of significanceNerve guidance conduits shows promise for peripheral nerve repair, while achieving the repair efficiency of autografts remains a challenge. In this study, a composite nerve conduit with a PLGA hollow tube as the outer layer and gelatin methacryloyl (GelMA) encapsulated with vascular endothelial growth factor A (VEGF-A)-transfected Schwann cells (SCs) as the inner layer was established to evaluate its potential ability for peripheral nerve repair.This approach preserves growth factor bioactivity and enhances material properties. GelMA insertion promotes Schwann cell proliferation and morphology extension. Moreover, transfected SCs serve as a stable VEGF-A source and fostering angiogenesis. This study offers a method preserving growth factor efficacy and safeguarding SCs, providing a comprehensive solution for enhanced angiogenesis and nerve regeneration.

Environment-Friendly Preparation and Characterization of Multilayered Conductive PVP/Col/CS Composite Doped with Nanoparticles as Potential Nerve Guide Conduits.

Tissue engineering constitutes the most promising method of severe peripheral nerve injuries treatment and is considered as an alternative to autografts. To provide appropriate conditions during recovery special biomaterials called nerve guide conduits are required. An ideal candidate for this purpose should not only be biocompatible and protect newly forming tissue but also promote the recovery process. In this article a novel, multilayered biomaterial based on polyvinylpyrrolidone, collagen and chitosan of gradient structure modified with conductive nanoparticles is presented. Products were obtained by the combination of electrospinning and electrospraying techniques. Nerve guide conduits were subjected to FT-IR analysis, morphology and elemental composition study using SEM/EDS as well as biodegradation. Furthermore, their effect on 1321N1 human cell line was investigated by long-term cell culture. Lack of cytotoxicity was confirmed by XTT assay and morphology study. Obtained results confirmed a high potential of newly developed biomaterials in the field of nerve tissue regeneration with a special focus on injured nerves recovery.

Rapid induction of dopaminergic neuron-like cells from human fibroblasts by autophagy activation with only 2-small molecules.

The dopaminergic neurons are responsible for the release of dopamine. Several diseases that affect motor function, including Parkinson's disease (PD), are rooted in inadequate dopamine (DA) neurotransmission. The study's goal was to create a quick way to make dopaminergic neuron-like cells from human fibroblasts (hNF) using only two small molecules: hedgehog pathway inhibitor 1 (HPI-1) and neurodazine (NZ). Two small compounds have been shown to induce the transdifferentiation of hNF cells into dopaminergic neuron-like cells. After 10 days of treatment, hNF cells had a big drop in fibroblastic markers (Col1A1, KRT18, and Elastin) and a rise in neuron marker genes (TUJ1, PAX6, and SOX1). Different proteins and factors related to dopaminergic neurons (TH, TUJ1, and dopamine) were significantly increased in cells that behave like dopaminergic neurons after treatment. A study of the autophagy signaling pathway showed that apoptotic genes were downregulated while autophagy genes (LC3, ATG5, and ATG12) were significantly upregulated. Our results showed that treating hNF cells with both HPI-1 and NZ together can quickly change them into mature neurons that have dopaminergic activity. However, the current understanding of the underlying mechanisms involved in nerve guidance remains unstable and complex. Ongoing research in this field must continue to advance for a more in-depth understanding. This is crucial for the safe and highly effective clinical application of the knowledge gained to promote neural regeneration in different neurological diseases.

Multichannel Conduits with Fascicular Complementation: Significance in Long Segmental Peripheral Nerve Injury.

Despite the advances in tissue engineering approaches, reconstruction of long segmental peripheral nerve defects remains unsatisfactory. Although autologous grafts with proper fascicular complementation have shown meaningful functional recovery according to the Medical Research Council Classification (MRCC), the lack of donor nerve for such larger defect sizes (>30 mm) has been a serious clinical issue. Further clinical use of hollow nerve conduits is limited to bridging smaller segmental defects of denuded nerve ends (<30 mm). Recently, bioinspired multichannel nerve guidance conduits (NGCs) gained attention as autograft substitutes as they mimic the fascicular connective tissue microarchitecture in promoting aligned axonal outgrowth with desirable innervation for complete sensory and motor function restoration. This review outlines the hierarchical organization of nerve bundles and their significance in the sensory and motor functions of peripheral nerves. This review also emphasizes the major challenges in addressing the longer nerve defects with the role of fascicular arrangement in the multichannel nerve guidance conduits and the need for fascicular matching to accomplish complete functional restoration, especially in treating long segmental nerve defects. Further, currently available fabrication strategies in developing multichannel nerve conduits and their inconsistency in existing preclinical outcomes captured in this review would seed a new process in designing an ideal larger nerve conduit for peripheral nerve repair.

Development of new nanofibrous nerve conduits by PCL-Chitosan-Hyaluronic acid containing Piracetam-Vitamin B12 for sciatic nerve: A rat model

Peripheral nerve injury is a critical condition that can disrupt nerve functions. Despite the progress in engineering artificial nerve guidance conduits (NGCs), nerve regeneration remains challenging. Here, we developed new nanofibrous NGCs using polycaprolactone (PCL) and chitosan (CH) containing piracetam (PIR)/vitamin B12(VITB12) with an electrospinning method. The lumen of NGCs was coated by hyaluronic acid (HA) to promote regeneration in sciatic nerve injury. The NGCs were characterized via Scanning Electron Microscopy (SEM), Fourier transform infrared (FTIR), tensile, swelling, contact angle, degradation, and drug release tests. Neuronal precursor cell line (PCL12 cell) and rat mesenchymal stem cells derived from bone marrow (MSCs) were seeded on the nanofibrous conduits. After that, the biocompatibility of the NGCs was evaluated by the 2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, 4′,6-diamidino-2-phenylindole (DAPI) staining, and SEM images. The SEM demonstrated that PCL/CH/PIR/VITB12 NGCs had nonaligned, interconnected, smooth fibers. The mechanical properties of these NGCs were similar to rat sciatic nerve. These conduits had an appropriate swelling and degradation rate. The In Vitro studies exhibited favorable biocompatibility of the PCL/CH/PIR/VITB12 NGCs towards PC12 cells and MSCs. The in vitro studies exhibited favorable biocompatibility of the PCL/CH/PIR/VIT B12 NGCs towards MSCs and PC12 cells. To analyze functional efficacy, NGCs were implanted into a 10 mm Wistar rat sciatic nerve gap and bridged the proximal and distal stump of the defect. After three months, the results of sciatic functional index (55.3 ± 1.8), hot plate latency test (5.6 ± 0.5 s), gastrocnemius muscle wet weight-loss (38.57 ± 1.6 %) and histopathological examination using hematoxylin-eosin (H&E) /toluidine blue/ Anti-Neurofilament (NF200) staining demonstrated that the produced conduit recovered motor and sensory functions and had comparable nerve regeneration compared to the autograft that can be as the gold standard to bridge the nerve gaps.

Peripheral nerve injury repair by electrical stimulation combined with graphene-based scaffolds.

Peripheral nerve injury (PNI) is a common clinical problem, which due to poor recovery often leads to limb dysfunction and sensory abnormalities in patients. Tissue-engineered nerve guidance conduits (NGCs) that are designed and fabricated from different materials are the potential alternative to nerve autografts. However, translation of these NGCs from lab to commercial scale has not been well achieved. Complete functional recovery with the aid of NGCs in PNI becomes a topic of general interest in tissue engineering and regeneration medicine. Electrical stimulation (ES) has been widely used for many years as an effective physical method to promote nerve repair in both pre-clinical and clinical settings. Similarly, ES of conductive and electroactive materials with a broad range of electrical properties has been shown to facilitate the guidance of axons and enhance the regeneration. Graphene and its derivatives possess unique physicochemical and biological properties, which make them a promising outlook for the development of synthetic scaffolds or NGCs for PNI repair, especially in combination with ES. Considering the discussion regarding ES for the treatment of PNI must continue into further detail, herein, we focus on the role of ES in PNI repair and the molecular mechanism behind the ES therapy for PNI, providing a summary of recent advances in context of graphene-based scaffolds (GBSs) in combination with ES. Future perspectives and some challenges faced in developing GBSs are also highlighted with the aim of promoting their clinical applications.

Advanced Hollow Cathode Discharge Plasma Treatment of Unique Bilayered Fibrous Nerve Guidance Conduits for Enhanced/Oriented Neurite Outgrowth.

Despite all recent progresses in nerve tissue engineering, critical-sized nerve defects are still extremely challenging to repair. Therefore, this study targets the bridging of critical nerve defects and promoting an oriented neuronal outgrowth by engineering innovative nerve guidance conduits (NGCs) synergistically possessing exclusive topographical, chemical, and mechanical cues. To do so, a mechanically adequate mixture of polycaprolactone (PCL) and polylactic-co-glycolic acid (PLGA) was first carefully selected as base material to electrospin nanofibrous NGCs simulating the extracellular matrix. The electrospinning process was performed using a newly designed 2-pole air gap collector that leads to a one-step deposition of seamless NGCs having a bilayered architecture with an inner wall composed of highly aligned fibers and an outer wall consisting of randomly oriented fibers. This architecture is envisaged to afford guidance cues for the extension of long neurites on the underlying inner fiber alignment and to concurrently provide a sufficient nutrient supply through the pores of the outer random fibers. The surface chemistry of the NGCs was then modified making use of a hollow cathode discharge (HCD) plasma reactor purposely designed to allow an effective penetration of the reactive species into the NGCs to eventually treat their inner wall. X-ray photoelectron spectroscopy (XPS) results have indeed revealed a successful O2 plasma modification of the inner wall that exhibited a significantly increased oxygen content (24 → 28%), which led to an enhanced surface wettability. The treatment increased the surface nanoroughness of the fibers forming the NGCs as a result of an etching effect. This effect reduced the ultimate tensile strength of the NGCs while preserving their high flexibility. Finally, pheochromocytoma (PC12) cells were cultured on the NGCs to monitor their ability to extend neurites which is the base of a good nerve regeneration. In addition to remarkably improved cell adhesion and proliferation on the plasma-treated NGCs, an outstanding neural differentiation occurred. In fact, PC12 cells seeded on the treated samples extended numerous long neurites eventually establishing a neural network-like morphology with an overall neurite direction following the alignment of the underlying fibers. Overall, PCL/PLGA NGCs electrospun using the 2-pole air gap collector and O2 plasma-treated using an HCD reactor are promising candidates toward a full repair of critical nerve damage.

Emulsion templated composites: Porous nerve guidance conduits for peripheral nerve regeneration

Peripheral nerve regeneration fails following more severe injuries where the gap between nerve stumps is too large. Nerve guidance conduits (NGCs) can be implanted to connect the nerve stumps and provide a regenerative microenvironment. This study shows that poly(glycerol sebacate)-methacrylate (PGS-M), a photocurable and biodegradable elastomer, can be used to fabricate porous NGCs. Further, novel developments in emulsion templating techniques are used to generate composite structures and their mechanical properties are investigated. The most appropriate composite for nerve tissue repair had a compressive and tensile Young’s modulus of 0.168 MPa and 0.694 MPa respectively, much closer to that of native nerve tissue than previously used polymers. SEM images revealed high porosity/interconnectivity of NGCs, important for the diffusion of nutrients to further aid nerve regeneration. FITC-Dextran (150 kDa) effectively diffused through conduit walls. Histological analysis demonstrated the small pore size successfully prevented infiltration of fibroblasts. In vitro studies indicated that the NGCs were cytocompatible. Ex vivo analysis showed that neurons of embryonic chick dorsal root ganglia grew on the surface of the conduits, and Schwann cells migrated effectively along the lumen. This study demonstrates a novel way of fabricating porous scaffolds and demonstrates their potential for the treatment of PNI.

Leveraging Oriented Lateral Walls of Nerve Guidance Conduit with Core-Shell MWCNTs Fibers for Peripheral Nerve Regeneration.

Peripheral nerve regeneration and functional recovery rely on the chemical, physical, and structural properties of nerve guidance conduits (NGCs). However, the limited support for long-distance nerve regeneration and axonal guidance has hindered the widespread use of NGCs. This study introduces a novel nerve guidance conduit with oriented lateral walls, incorporating multi-walled carbon nanotubes (MWCNTs) within core-shell fibers prepared in a single step using a modified electrohydrodynamic (EHD) printing technique to promote peripheral nerve regeneration. The structured conduit demonstrated exceptional stability, mechanical properties, and biocompatibility, significantly enhancing the functionality of NGCs. In vitro cell studies revealed that RSC96 cells adhered and proliferated effectively along the oriented fibers, demonstrating a favorable response to the distinctive architectures and properties. Subsequently, a rat sciatic nerve injury model demonstrated effective efficacy in promoting peripheral nerve regeneration and functional recovery. Tissue analysis and functional testing highlighted the significant impact of MWCNT concentration in enhancing peripheral nerve regeneration and confirming well-matured aligned axonal growth, muscle recovery, and higher densities of myelinated axons. These findings demonstrate the potential of oriented lateral architectures with coaxial MWCNT fibers as a promising approach to support long-distance regeneration and encourage directional nerve growth for peripheral nerve repair in clinical applications.

Application of Adipose Stem Cells in 3D Nerve Guidance Conduit Prevents Muscle Atrophy and Improves Distal Muscle Compliance in a Peripheral Nerve Regeneration Model.

Peripheral nerve injuries (PNIs) represent a significant clinical problem, and standard approaches to nerve repair have limitations. Recent breakthroughs in 3D printing and stem cell technologies offer a promising solution for nerve regeneration. The main purpose of this study was to examine the biomechanical characteristics in muscle tissue distal to a nerve defect in a murine model of peripheral nerve regeneration from physiological stress to failure. In this experimental study, we enrolled 18 Wistar rats in which we created a 10 mm sciatic nerve defect. Furthermore, we divided them into three groups as follows: in Group 1, we used 3D nerve guidance conduits (NGCs) and adipose stem cells (ASCs) in seven rats; in Group 2, we used only 3D NGCs for seven rats; and in Group 3, we created only the defect in four rats. We monitored the degree of atrophy at 4, 8, and 12 weeks by measuring the diameter of the tibialis anterior (TA) muscle. At the end of 12 weeks, we took the TA muscle and analyzed it uniaxially at 10% stretch until failure. In the group of animals with 3D NGCs and ASCs, we recorded the lowest degree of atrophy at 4 weeks, 8 weeks, and 12 weeks after nerve reconstruction. At 10% stretch, the control group had the highest Cauchy stress values compared to the 3D NGC group (0.164 MPa vs. 0.141 MPa, p = 0.007) and the 3D NGC + ASC group (0.164 MPa vs. 0.123 MPa, p = 0.007). In addition, we found that the control group (1.763 MPa) had the highest TA muscle stiffness, followed by the 3D NGC group (1.412 MPa), with the best muscle elasticity showing in the group in which we used 3D NGC + ASC (1.147 MPa). At failure, TA muscle samples from the 3D NGC + ASC group demonstrated better compliance and a higher degree of elasticity compared to the other two groups (p = 0.002 and p = 0.008). Our study demonstrates that the combination of 3D NGC and ASC increases the process of nerve regeneration and significantly improves the compliance and mechanical characteristics of muscle tissue distal to the injury site in a PNI murine model.

Advancing Peripheral Nerve Regeneration: 3D Bioprinting of GelMA-Based Cell-Laden Electroactive Bioinks for Nerve Conduits.

Peripheral nerve injuries often result in substantial impairment of the neurostimulatory organs. While the autograft is still largely used as the "gold standard" clinical treatment option, nerve guidance conduits (NGCs) are currently considered a promising approach for promoting peripheral nerve regeneration. While several attempts have been made to construct NGCs using various biomaterial combinations, a comprehensive exploration of the process science associated with three-dimensional (3D) extrusion printing of NGCs with clinically relevant sizes (length: 20 mm; diameter: 2-8 mm), while focusing on tunable buildability using electroactive biomaterial inks, remains unexplored. In addressing this gap, we present here the results of the viscoelastic properties of a range of a multifunctional gelatin methacrylate (GelMA)/poly(ethylene glycol) diacrylate (PEGDA)/carbon nanofiber (CNF)/gellan gum (GG) hydrogel bioink formulations and printability assessment using experiments and quantitative models. Our results clearly established the positive impact of the gellan gum on the enhancement of the rheological properties. Interestingly, the strategic incorporation of PEGDA as a secondary cross-linker led to a remarkable enhancement in the strength and modulus by 3 and 8-fold, respectively. Moreover, conductive CNF addition resulted in a 4-fold improvement in measured electrical conductivity. The use of four-component electroactive biomaterial ink allowed us to obtain high neural cell viability in 3D bioprinted constructs. While the conventionally cast scaffolds can support the differentiation of neuro-2a cells, the most important result has been the excellent cell viability of neural cells in 3D encapsulated structures. Taken together, our findings demonstrate the potential of 3D bioprinting and multimodal biophysical cues in developing functional yet critical-sized nerve conduits for peripheral nerve tissue regeneration.

Nerve Regeneration and Gait Function Recovery with Implantation of Glucose/Mannose Conduits Using a Rat Model: Efficacy of Glucose/Mannose as a New Neurological Guidance Material.

Therapy with clinical nerve guidance conduits often causes functional incompleteness in patients. With the aim of better therapeutic efficacy, nerve regeneration and gait function were investigated in this study using a novel nerve guidance conduit consisting of glucose/mannose. The glucose/mannose nerve guidance conduits were prepared by filling the conduits with the glucose/mannose aqueous solutions for different kinematic viscosity, which were applied to sciatic nerve defects (6 mm gap) in a rat model. The nerve regeneration effect and the gait function recovery with the fabricated nerve guidance conduits were examined. From the results of the XRD measurement, the glucose/mannose conduits were identified as crystal structures of cellulose type II. Young's modulus and the maximum tensile strength of the crystalline glucose/mannose conduits demonstrated good strength and softness for the human nerve. Above 4 weeks postoperative, macroscopic observation revealed that the nerve was regenerated in the defective area. In various staining results of the nerve tissue removed at 4 weeks postoperative, myelinated nerves contributing to gait function could not be observed in the proximal and distal sites to the central nerve. At 8-12 weeks postoperative, myelinated nerves were found at the proximal and distal sites in hematoxylin/eosin staining. Glia cells were confirmed by phosphotungstic acid-hematoxylin staining. Continuous nerve fibers were observed clearly in the sections of the regenerated nerves towards the longitudinal direction at 12 weeks postoperative. The angle between the metatarsophalangeal joint and the ground plane was approximately 93° in intact rats. At 4 weeks postoperative, walking was not possible, but at 8 weeks postoperative, the rats were able to walk, with an angle of 53°. At 12 weeks postoperative, the angle increased further, reaching 65°, confirming that the rats were able to walk more quickly than at 8 weeks postoperative. These results demonstrated that gait function in rats treated with glucose/mannose nerve guidance conduits was rapidly recovered after 8 weeks postoperative. The glucose/mannose nerve guidance conduit could be applied as a new promising candidate material for peripheral nerve regeneration.

Management of peripheral nerve injuries using natural based biomaterials and their derivatives: Advances and prospective

AbstractThe management of peripheral nerve injuries is an important concern due to the their incidence of nerve lesions and inappropriate regeneration that follows severe injuries, which ultimately reduces the lives of patients with this condition. Different strategies have been investigated to repair severe nerve injuries with the improvement of motor and sensory regeneration. Although autograft remains the gold standard technique, an emerging number of research articles concerning nerve conduit use have been reported in the last few years. Nerve conduits aim to overcome autograft disadvantages, but they satisfy some requirements to be suitable for nerve repair. A universal ideal conduit does not exist since conduit properties have to be evaluated case by case; nevertheless, because of their high biocompatibility and biodegradability, natural‐based biomaterials have great potential to be used to produce nerve guides. Although they have many characteristics with synthetic biomaterials, natural‐based biomaterials are preferable because of their extraction sources; indeed, these biomaterials are obtained from different renewable sources or food waste, thus reducing environmental impact and enhancing sustainability in comparison to synthetic ones. This review highlights the recent progress in the development of natural‐based biomaterials and their derivatives for the management of peripheral nerve injuries.

Magnetoactive Composite Conduits Based on Poly(3-hydroxybutyrate) and Magnetite Nanoparticles for Repair of Peripheral Nerve Injury.

Peripheral nerve injury poses a threat to the mobility and sensitivity of a nerve, thereby leading to permanent function loss due to the low regenerative capacity of mature neurons. To date, the most widely clinically applied approach to bridging nerve injuries is autologous nerve grafting, which faces challenges such as donor site morbidity, donor shortages, and the necessity of a second surgery. An effective therapeutic strategy is urgently needed worldwide to overcome the current limitations. Herein, a magnetic nerve guidance conduit (NGC) based on biocompatible biodegradable poly(3-hydroxybutyrate) (PHB) and 8 wt % of magnetite nanoparticles modified by citric acid (Fe3O4-CA) was fabricated by electrospinning. The crystalline structure of NGCs was studied by X-ray diffraction, which indicated an enlarged β-phase of PHB in the composite conduit compared to a pure PHB conduit. Tensile tests revealed greater ductility of PHB/Fe3O4-CA: the composite conduit has Young's modulus of 221 ± 52 MPa and an elongation at break of 28.6 ± 2.9%, comparable to clinical materials. Saturation magnetization (σs) of Fe3O4-CA and PHB/Fe3O4-CA is 61.88 ± 0.29 and 7.44 ± 0.07 emu/g, respectively. The water contact angle of the PHB/Fe3O4-CA conduit is lower as compared to pure PHB, while surface free energy (σ) is significantly higher, which was attributed to higher surface roughness and an amorphous phase as well as possible PHB/Fe3O4-CA interface interactions. In vitro, the conduits supported the proliferation of rat mesenchymal stem cells (rMSCs) and SH-SY5Y cells in a low-frequency magnetic field (0.67 Hz, 68 mT). In vivo, the conduits were used to bridge damaged sciatic nerves in rats; pure PHB and composite PHB/Fe3O4-CA conduits did not cause acute inflammation and performed a barrier function, which promotes nerve regeneration. Thus, these conduits are promising as implants for the regeneration of peripheral nerves.