Recombinant COL6 α2 as a Self-Organization Factor That Triggers Orderly Nerve Regeneration Without Guidance Cues.
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ABSTRACT:
SUBMITTER: Fang Z
PROVIDER: S-EPMC8732766 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Nerve guidance conduits with multifunctional features could offer microenvironments for improved nerve regeneration and functional recovery. However, the challenge remains to optimize multiple cues in nerve conduit systems due to the interplay of these factors during fabrication. Here, a modular assembly for the fabrication of nerve conduits is utilized to address the goal of incorporating multifunctional guidance cues for nerve regeneration. Silk-based hollow conduits with suitable size and mechanical properties, along with silk nanofiber fillers with tunable hierarchical anisotropic architectures and microporous structures, are developed and assembled into conduits. These conduits supported improves nerve regeneration in terms of cell proliferation (Schwann and PC12 cells) and growth factor secretion (BDNF, brain-derived neurotrophic factor) in vitro, and the in vivo repair and functional recovery of rat sciatic nerve defects. Nerve regeneration using these new conduit designs is comparable to autografts, providing a path towards future clinical impact.
| S-EPMC8295195 | biostudies-literature
Project description:Nerve guidance conduits (NGCs) are used as an alternative to the "gold standard" nerve autografting, preventing the need for surgical intervention required to harvest autologous nerves. However, the regeneration outcomes achieved with the current NGCs are only comparable with autografting when the gap is short (less than 10 mm). In the present study, we have developed NGCs made from a blend of polyhydroxyalkanoates, a family of natural resorbable polymers. Hollow NGCs made from a 75:25 poly(3-hydroxyoctanoate)/poly(3-hydroxybutyrate) blend (PHA-NGCs) were manufactured using dip-molding. These PHA-NGCs showed appropriate flexibility for peripheral nerve regeneration. In vitro cell studies performed using RT4-D6P2T rat Schwann cell line confirmed that the material is capable of sustaining cell proliferation and adhesion. PHA-NGCs were then implanted in vivo to repair 10 mm gaps of the median nerve of female Wistar rats for 12 weeks. Functional evaluation of the regenerated nerve using the grasping test showed that PHA-NGCs displayed similar motor recovery as the autograft, starting from week 7. Additionally, nerve cross-sectional area, density and number of myelinated cells, as well as axon diameter, fiber diameter, myelin thickness and g-ratio obtained using the PHA-NGCs were found comparable to an autograft. This preclinical data confirmed that the PHA-NGCs are indeed highly promising candidates for peripheral nerve regeneration.
| S-EPMC8459605 | biostudies-literature
Project description:Despite microsurgical repair, recovery of function following peripheral nerve injury is slow and often incomplete. Outcomes could be improved by an increased understanding of the molecular biology of regeneration and by translation of experimental bioengineering strategies. Topographical cues have been shown to be powerful regulators of the rate and directionality of neurite regeneration, and in this study we investigated the downstream molecular effects of linear micropatterned structures in an organotypic explant model. Linear topographical cues enhanced neurite outgrowth and our results demonstrated that the mTOR pathway is important in regulating these responses. mTOR gene expression peaked between 48 and 72h, coincident with the onset of rapid neurite outgrowth and glial migration, and correlated with neurite length at 48h. mTOR protein was located to glia and in a punctate distribution along neurites. mTOR levels peaked at 72h and were significantly increased by patterned topography (p<0.05). Furthermore, the topographical cues could override pharmacological inhibition. Downstream phosphorylation assays and inhibition of mTORC1 using rapamycin highlighted mTORC2 as an important mediator, and more specific therapeutic target. Quantitative immunohistochemistry confirmed the presence of the mTORC2 component rictor at the regenerating front where it co-localised with F-actin and vinculin. Collectively, these results provide a deeper understanding of the mechanism of action of topography on neural regeneration, and support the incorporation of topographical patterning in combination with pharmacological mTORC2 potentiation within biomaterial constructs used to repair peripheral nerves.Statement of significancePeripheral nerve injury is common and functionally devastating. Despite microsurgical repair, healing is slow and incomplete, with lasting functional deficit. There is a clear need to translate bioengineering approaches and increase our knowledge of the molecular processes controlling nerve regeneration to improve the rate and success of healing. Topographical cues are powerful determinants of neurite outgrowth and represent a highly translatable engineering strategy. Here we demonstrate, for the first time, that microtopography potentiates neurite outgrowth via the mTOR pathway, with the mTORC2 subtype being of particular importance. These results give further evidence for the incorporation of microtopographical cues into peripheral nerve regeneration conduits and indicate that mTORC2 may be a suitable therapeutic target to potentiate nerve regeneration.
| S-EPMC5593812 | biostudies-literature
Project description:Neural interfaces are designed to decode motor intent and evoke sensory precepts in amputees. In peripheral nerves, recording movement intent is challenging because motor axons are only a small fraction compared to sensory fibers and are heterogeneously mixed particularly at proximal levels. We previously reported that pain and myelinated axons regenerating through a Y-shaped nerve guide with sealed ends, can be modulated by luminar release of nerve growth factor (NGF) and neurotrophin-3 (NT-3), respectively. Here, we evaluate the differential potency of NGF, glial cell line-derived neurotrophic factor (GDNF), brain-derived neurotrophic factor (BDNF), pleiotrophin (PTN), and NT-3 in asymmetrically guiding the regeneration of sensory and motor neurons. We report that, in the absence of distal target organs, molecular guidance cues can mediate the growth of electrically conductive fascicles with normal microanatomy. Compared to Y-tube compartments with bovine serum albumin (BSA), GDNF and NGF increased the motor and sensory axon content, respectively. In addition, the sensory to motor ratio was significantly increased by PTN (12.7:1) when compared to a BDNF?+?GDNF choice. The differential content of motor and sensory axons modulated by selective guidance cues may provide a strategy to better define axon types in peripheral nerve interfaces.
| S-EPMC5662603 | biostudies-literature
Project description:Peripheral nerve repair and regeneration remains among the greatest challenges in tissue engineering and regenerative medicine. Even though peripheral nerve injuries (PNIs) are capable of some degree of regeneration, frail recovery is seen even when the best microsurgical technique is applied. PNIs are known to be very incapacitating for the patient, due to the deprivation of motor and sensory abilities. Since there is no optimal solution for tackling this problem up to this day, the evolution in the field is constant, with innovative designs of advanced nerve guidance conduits (NGCs) being reported every day. As a basic concept, a NGC should act as a physical barrier from the external environment, concomitantly acting as physical guidance for the regenerative axons across the gap lesion. NGCs should also be able to retain the naturally released nerve growth factors secreted by the damaged nerve stumps, as well as reducing the invasion of scar tissue-forming fibroblasts to the injury site. Based on the neurobiological knowledge related to the events that succeed after a nerve injury, neuronal subsistence is subjected to the existence of an ideal environment of growth factors, hormones, cytokines, and extracellular matrix (ECM) factors. Therefore, it is known that multifunctional NGCs fabricated through combinatorial approaches are needed to improve the functional and clinical outcomes after PNIs. The present work overviews the current reports dealing with the several features that can be used to improve peripheral nerve regeneration (PNR), ranging from the simple use of hollow NGCs to tissue engineered intraluminal fillers, or to even more advanced strategies, comprising the molecular and gene therapies as well as cell-based therapies.
| S-EPMC6882937 | biostudies-literature
Project description:Nerve guidance conduits (NGCs) have become a promising alternative for peripheral nerve regeneration; however, the outcome of nerve regeneration and functional recovery is greatly affected by the physical, chemical, and electrical properties of NGCs. In this study, a conductive multiscale filled NGC (MF-NGC) consisting of electrospun poly(lactide-co-caprolactone) (PCL)/collagen nanofibers as the sheath, reduced graphene oxide /PCL microfibers as the backbone, and PCL microfibers as the internal structure for peripheral nerve regeneration is developed. The printed MF-NGCs presented good permeability, mechanical stability, and electrical conductivity, which further promoted the elongation and growth of Schwann cells and neurite outgrowth of PC12 neuronal cells. Animal studies using a rat sciatic nerve injury model reveal that the MF-NGCs promote neovascularization and M2 transition through the rapid recruitment of vascular cells and macrophages. Histological and functional assessments of the regenerated nerves confirm that the conductive MF-NGCs significantly enhance peripheral nerve regeneration, as indicated by improved axon myelination, muscle weight increase, and sciatic nerve function index. This study demonstrates the feasibility of using 3D-printed conductive MF-NGCs with hierarchically oriented fibers as functional conduits that can significantly enhance peripheral nerve regeneration.
| S-EPMC10131803 | biostudies-literature
Project description:Synthetic nerve guidance conduits (NGCs) offer an alternative to harvested nerve grafts for treating peripheral nerve injury (PNI). NGCs have been made from both naturally derived and synthesized materials. While naturally derived materials typically have an increased capacity for bioactivity, synthesized materials have better material control, including tunability and reproducibility. Protein engineering is an alternative strategy that can bridge the benefits of these two classes of materials by designing cell-responsive materials that are also systematically tunable and consistent. Here, we tested a recombinantly derived elastin-like protein (ELP) hydrogel as an intraluminal filler in a rat sciatic nerve injury model. We demonstrated that ELPs enhance the probability of forming a tissue bridge between the proximal and distal nerve stumps compared to an empty silicone conduit across the length of a 10 mm nerve gap. These tissue bridges have evidence of myelinated axons, and electrophysiology demonstrated that regenerated axons innervated distal muscle groups. Animals implanted with an ELP-filled conduit had statistically higher functional control at 6 weeks than those that had received an empty silicone conduit, as evaluated by the sciatic functional index. Taken together, our data support the conclusion that ELPs support peripheral nerve regeneration in acute complete transection injuries when used as an intraluminal filler. These results support the further study of protein engineered recombinant ELP hydrogels as a reproducible, off-the-shelf alternative for regeneration of peripheral nerves.
| S-EPMC8438224 | biostudies-literature
Project description:BackgroundEfficient and stable delivery of neurotrophic factors (NTFs) is crucial to provide suitable microenvironment for peripheral nerve regeneration. Neurotrophin-3 (NT-3) is an important NTF during peripheral nerve regeneration which is scarce in the first few weeks of nerve defect. Exosomes are nanovesicles and have been served as promising candidate for biocarrier. In this work, NT-3 mRNA was encapsulated in adipose-derived stem cell (ADSC)-derived exosomes (ExoNT-3). These engineered exosomes were applied as NT-3 mRNA carrier and then were loaded in nerve guidance conduit (ExoNT-3-NGC) to bridge rat sciatic nerve defect.MethodNT-3 mRNA was encapsulated in exosomes by forcedly expression of NT-3 mRNA in the donor ADSCs. ExoNT-3 were co-cultured with SCs in vitro; after 24 h of culture, the efficiency of NT-3 mRNA delivery was evaluated by qPCR, western blotting and ELISA. Then, ExoNT-3 were loaded in alginate hydrogel to construct the nerve guidance conduits (ExoNT-3-NGC). ExoNT-3-NGC were implanted in vivo to reconstruct 10 mm rat sciatic nerve defect. The expression of NT-3 was measured 2 weeks after the implantation operation. The sciatic nerve functional index (SFI) was examined at 2 and 8 weeks after the operation. Moreover, the therapeutic effect of ExoNT-3-NGC was also evaluated by morphology assay, immunofluorescence staining of regenerated nerves, function evaluation of gastrocnemius muscles after 8 weeks of implantation.ResultsThe engineered exosomes could deliver NT-3 mRNA to the recipient cells efficiently and translated into functional protein. The constructed NGC could realize stable release of exosomes at least for 2 weeks. After NGC implantation in vivo, ExoNT-3-NGC group significantly promote nerve regeneration and improve the function recovery of gastrocnemius muscles compared with control exosomes (Exoempty-NGC) group.ConclusionIn this work, NGC was constructed to allow exosome-mediated NT-3 mRNA delivery. After ExoNT-3-NGC implantation in vivo, the level of NT-3 could restore which enhance the nerve regeneration. Our study provide a potential approach to improve nerve regeneration.
| S-EPMC8343914 | biostudies-literature
Project description:Vocal cord paralysis caused by recurrent laryngeal nerve (RLN) injury during thyroidectomy results in hoarseness, aspiration, and dyspnea. We evaluated the usefulness of nerve guidance conduits (NGCs) constructed from an asymmetric polycaprolactone (PCL)/Pluronic F127 porous membrane and filled with platelet-rich plasma (PRP) for functional RLN regeneration. We evaluated the proliferation and migration of Schwann cells (SCs) after PRP treatment in vitro. For the in vivo study, rabbits were divided into a non-loaded NGC group and a PRP-loaded NGC group. The left RLNs were resected and interposed with the NGCs. Functional and histological examinations of the vocal cords were performed. SC proliferation and migration increased in a PRP dose-dependent manner, with the PRP increasing the levels of neurotrophic factors, myelin-associated glycoprotein, and ERK. In vivo, the PRP group showed significantly better vocal cord mobility and less vocalis muscle atrophy than the non-loaded NGC group. Histologically, the ingrowth of nerve endings occurred more rapidly in the PRP group, and acetylcholinesterase, neurofilament, and S-100 expression in neural endings were significantly higher in the PRP group. Furthermore, transmission electron microscopy showed that myelinated axons were more tightly packed in the PRP group. This study shows that PRP-loaded NGCs provide a favorable environment for neural regeneration and suggests that this technique has therapeutic potential for promoting RLN recovery.
| S-EPMC9474804 | biostudies-literature
Project description:The peripheral nervous system has an astonishing ability to regenerate following a compression or crush injury; however, the potential for full repair following a transection injury is much less. Currently, the major clinical challenge for peripheral nerve repair come from long gaps between the proximal and distal nerve stumps, which prevent regenerating axons reaching the distal nerve. Precise axon targeting during nervous system development is controlled by families of axon guidance molecules including Netrins, Slits, Ephrins and Semaphorins. Several recent studies have indicated key roles of Netrin1, Slit3 and EphrinB2 signalling in controlling the formation of new nerve bridge tissue and precise axon regeneration after peripheral nerve transection injury. Inside the nerve bridge, nerve fibroblasts express EphrinB2 while migrating Schwann cells express the receptor EphB2. EphrinB2/EphB2 signalling between nerve fibroblasts and migrating Schwann cells is required for Sox2 upregulation in Schwann cells and the formation of Schwann cell cords within the nerve bridge to allow directional axon growth to the distal nerve stump. Macrophages in the outermost layer of the nerve bridge express Slit3 while migrating Schwann cells and regenerating axons express the receptor Robo1; within Schwann cells, Robo1 expression is also Sox2-dependent. Slit3/Robo1 signalling is required to keep migrating Schwann cells and regenerating axons inside the nerve bridge. In addition to the Slit3/Robo1 signalling system, migrating Schwann cells also express Netrin1 and regenerating axons express the DCC receptor. It appears that migrating Schwann cells could also use Netrin1 as a guidance cue to direct regenerating axons across the peripheral nerve gap. Engineered neural tissues have been suggested as promising alternatives for the repair of large peripheral nerve gaps. Therefore, understanding the function of classic axon guidance molecules in nerve bridge formation and their roles in axon regeneration could be highly beneficial in developing engineered neural tissue for more effective peripheral nerve repair.
| S-EPMC6862410 | biostudies-literature