Transforming growth factor beta 1, a cytokine with regenerative functions

We review the biology and role of transforming growth factor beta 1(TGF-β1) in peripheral nerve injury and regeneration, as it relates to injuries to large nerve trunks(i.e., sciatic nerve, brachial plexus), which often leads to suboptimal functional recovery. Experimental studies have suggested that the reason for the lack of functional recovery resides in the lack of sufficient mature axons reaching their targets, which is a result of the loss of the growth-supportive environment provided by the Schwann cells in the distal stump of injured nerves. Using an established chronic nerve injury and delayed repair animal model that accurately mimics chronic nerve injuries in humans, we summarize our key findings as well as others to better understand the pathophysiology of poor functional recovery. We demonstrated that 6 month TGF-β1 treatment for chronic nerve injury significantly improved Schwann cell capacity to support axonal regeneration. When combined with forskolin, the effect was additive, as evidenced by a near doubling of regenerated axons proximal to the repair site. We showed that in vivo application of TGF-β1 and forskolin directly onto chronically injured nerves reactivated chronically denervated Schwann cells, induced their proliferation, and upregulated the expression of regeneration-associated proteins. The effect of TGF-β1 and forskolin on old nerve injuries is quite impressive and the treatment regiment appears to mediate a growth-supportive milieu in the injured peripheral nerves. In summary, TGF-β1 and forskolin treatment reactivates chronically denervated Schwann cells and could potentially be used to extend and prolong the regenerative responses to promote axonal regeneration.     

Classic axon guidance molecules control correct nerve bridge tissue formation and precise axon regeneration

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.     

Mesenchymal stem cell treatment for peripheral nerve injury: a narrative review

Peripheral nerve injuries occur as the result of sudden trauma and lead to reduced quality of life.The peripheral nervous system has an inherent capability to regenerate axons.However,peripheral nerve regeneration following injury is generally slow and incomplete that results in poor functional outcomes such as muscle atrophy.Although conventional surgical procedures for peripheral nerve injuries present many benefits,there are still several limitations including scarring,difficult accessibility to donor nerve,neuroma formation and a need to sacrifice the autologous nerve.For many years,other therapeutic approaches for peripheral nerve injuries have been explored,the most notable being the replacement of Schwann cells,the glial cells responsible for clearing out debris from the site of injury.Introducing cultured Schwann cells to the injured sites showed great benefits in promoting axonal regeneration and functional recovery.However,there are limited sources of Schwann cells for extraction and difficulties in culturing Schwann cells in vitro.Therefore,novel therapeutic avenues that offer maximum benefits for the treatment of peripheral nerve injuries should be investigated.This review focused on strategies using mesenchymal stem cells to promote peripheral nerve regeneration including exosomes of mesenchymal stem cells,nerve engineering using the nerve guidance conduits containing mesenchymal stem cells,and genetically engineered mesenchymal stem cells.We present the current progress of mesenchymal stem cell treatment of peripheral nerve injuries.     

Sleeve bridging of the rhesus monkey ulnar nerve with muscular branches of the pronator teres: multiple amplification of axonal regeneration

Multiple-bud regeneration, i.e., multiple amplification, has been shown to exist in peripheral nerve regeneration. Multiple buds grow towards the distal nerve stump during proximal nerve fiber regeneration. Our previous studies have verified the limit and validity of multiple amplification of peripheral nerve regeneration using small gap sleeve bridging of small donor nerves to repair large receptor nerves in rodents. The present study sought to observe multiple amplification of myelinated nerve fiber regeneration in the primate peripheral nerve. Rhesus monkey models of distal ulnar nerve defects were established and repaired using muscular branches of the right forearm pronator teres. Proximal muscular branches of the pronator teres were sutured into the distal ulnar nerve using the small gap sleeve bridging method. At 6 months after suture, two-finger flexion and mild wrist flexion were restored in the ulnar-sided injured limbs of rhesus monkey. Neurophysiological examination showed that motor nerve conduction velocity reached 22.63 ± 6.34 m/s on the affected side of rhesus monkey. Osmium tetroxide staining demonstrated that the number of myelinated nerve fibers was 1,657 ± 652 in the branches of pronator teres of donor, and 2,661 ± 843 in the repaired ulnar nerve. The rate of multiple amplification of regenerating myelinated nerve fibers was 1.61. These data showed that when muscular branches of the pronator teres were used to repair ulnar nerve in primates, effective regeneration was observed in regenerating nerve fibers, and functions of the injured ulnar nerve were restored to a certain extent. Moreover, multiple amplification was subsequently detected in ulnar nerve axons.     

功能性电刺激干预损伤动眼神经的肌电图评估

Functional electrical stimulation delivered early after injury to the proximal nerve stump has been proposed as a therapeutic approach for enhancing the speed and specificity of axonal regeneration following nerve injury. In this study, the injured oculomotor nerve was stimulated functionally by an implantable electrode. Electromyographic monitoring of the motor unit potential of the inferior oblique muscle was conducted for 12 weeks in two injury groups, one with and one without electric stimulation. The results revealed that, at 2, 4, 6, 8 weeks after functional electric stimulation of the injured oculomotor nerve, motor unit potentials significantly increased, such that amplitude was longer and spike duration gradually shortened. These findings indicate that the injured oculomotor nerve has the potential for regeneration and repair, but this ability is not sufficient for full functional recovery to occur. Importantly, the current results indicated that recovery and regeneration of the injured oculomotor nerve can be promoted with functional electrical stimulation.     

New insights on the molecular mechanisms of collateral sprouting after peripheral nerve injury

正Axonal regeneration after injuries to the nervous system has been extensively studied due to its implication in motor and sensory functional recovery. Distinct types of regeneration has been identified, such as canonical axonal regeneration, defined as the growth of axons from the transected axonal stump to reinnervate the original target, or regenerative sprouting, in which the growth occurs from a region of the damaged axon either close or far from the injury site(Tuszynski and Steward, 2012).     

Myelin-associated glycoprotein combined with chitin conduit inhibits painful neuroma formation after sciatic nerve transection

Studies have shown that myelin-associated glycoprotein(MAG)can inhibit axon regeneration after nerve injury.However,the effects of MAG on neuroma formation after peripheral nerve injury remain poorly understood.In this study,local injection of MAG combined with nerve cap made of chitin conduit was used to intervene with the formation of painful neuroma after sciatic nerve transfection in rats.After 8 weeks of combined treatment,the autotomy behaviors were reduced in rats subjected to sciatic nerve transfection,the mRNA expression of nerve growth factor,a pain marker,in the proximal nerve stump was decreased,the density of regenerated axons was decreased,the thickness of the myelin sheath was increased,and the ratio of unmyelinated to myelinated axons was reduced.Moereover,the percentage of collagen fiber area and the percentage of fibrosis marker alpha-smooth muscle actin positive staining area in the proximal nerve stump were decreased.The combined treatment exhibited superior effects in these measures to chitin conduit treatment alone.These findings suggest that MAG combined with chitin conduit synergistically inhibits the formation of painful neuroma after sciatic nerve transection and alleviates neuropathic pain.This study was approved by the Animal Ethics Committee of Peking University People’s Hospital(approval No.2019PHE027)on December 5,2019.     

Biological conduit small gap sleeve bridging method for peripheral nerve injury: regeneration law of nerve fibers in the conduit

The clinical effects of 2-mm small gap sleeve bridging of the biological conduit to repair peripheral nerve injury are better than in the traditional epineurium suture, so it is possible to replace the epineurium suture in the treatment of peripheral nerve injury. This study sought to identify the regeneration law of nerve fibers in the biological conduit. A nerve regeneration chamber was constructed in models of sciatic nerve injury using 2-mm small gap sleeve bridging of a biodegradable biological conduit. The results showed that the biological conduit had good histocompatibility. Tissue and cell apoptosis in the conduit apparently lessened, and regenerating nerve fibers were common. The degeneration regeneration law of Schwann cells and axons in the conduit was quite different from that in traditional epineurium suture. During the prime period for nerve fiber regeneration(2–8 weeks), the number of Schwann cells and nerve fibers was higher in both proximal and distal ends, and the effects of the small gap sleeve bridging method were better than those of the traditional epineurium suture. The above results provide an objective and reliable theoretical basis for the clinical application of the biological conduit small gap sleeve bridging method to repair peripheral nerve injury.     

Visualization of peripheral nerve regeneration

<正>Peripheral nerve injuries are often caused by trauma and they may result in a partial or total loss of motor function or sensory perception.After nerve injuries,peripheral axons have the ability to regenerate and reconnect the proximal and distal ends of severed nerve axons if the nerve gap is small.For larger nerve gaps,surgical treatments are often required to repair the injured nerves.     

Effect of the combination of high-frequency repetitive magnetic stimulation and neurotropin on injured sciatic nerve regeneration in rats

Repetitive magnetic stimulation is effective for treating posttraumatic neuropathies following spinal or axonal injury.Neurotropin is a potential treatment for nerve injuries like demyelinating diseases.This study sought to observe the effects of high-frequency repetitive magnetic stimulation,neurotropin and their combined use in the treatment of peripheral nerve injury in 32 adult male Sprague-Dawley rats.To create a sciatic nerve injury model,a 10 mm-nerve segment of the left sciatic nerve was cut and rotated through 180°and each end restored continuously with interrupted sutures.The rats were randomly divided into four groups.The control group received only a reversed autograft in the left sciatic nerve with no treatment.In the high-frequency repetitive magnetic stimulation group,peripheral high-frequency repetitive magnetic stimulation treatment(20 Hz,20 min/d)was delivered for 10 consecutive days after auto-grafting.In the neurotropin group,neurotropin therapy(0.96 NU/kg per day)was administrated for 10 consecutive days after surgery.In the combined group,the combination of peripheral high-frequency repetitive magnetic stimulation(20 Hz,20 min/d)and neurotropin(0.96 NU/kg per day)was given for 10 consecutive days after the operation.The Basso-Beattie-Bresnahan locomotor rating scale was used to assess the behavioral recovery of the injured nerve.The sciatic functional index was used to evaluate the recovery of motor functions.Toluidine blue staining was performed to determine the number of myelinated fibers in the distal and proximal grafts.Immunohistochemistry staining was used to detect the length of axons marked by neurofilament 200.Our results reveal that the Basso-Beattie-Bresnahan locomotor rating scale scores,sciatic functional index,the number of myelinated fibers in distal and proximal grafts were higher and axon lengths were longer in the high-frequency repetitive magnetic stimulation,neurotropin and combined groups compared with the control group.These measures were not significantly different among the high-frequency repetitive magnetic stimulation,neurotropin and combined groups.Therefore,our results suggest that peripheral high-frequency repetitive magnetic stimulation or neurotropin can promote the repair of injured sciatic nerves,but their combined use seems to offer no significant advantage.This study was approved by the Animal Ethics Committee of the Affiliated Changzhou No.2 People’s Hospital of Nanjing Medical University,China on December 23,2014(approval No.2014keyan002-01).     

GSK3β inhibitor promotes myelination and mitigates muscle atrophy after peripheral nerve injury

Delay of axon regeneration after peripheral nerve injury usually leads to progressive muscle atrophy and poor functional recovery. The Wnt/β-catenin signaling pathway is considered to be one of the main molecular mechanisms that lead to skeletal muscle atrophy in the elderly. We hold the hypothesis that the innervation of target muscle can be promoted by accelerating axon regeneration and decelerating muscle cell degeneration so as to improve functional recovery of skeletal muscle following peripheral nerve injury. This process may be associated with the Wnt/β-catenin signaling pathway. Our study designed in vitro cell models to simulate myelin regeneration and muscle atrophy. We investigated the effects of SB216763, a glycogen synthase kinase 3 beta inhibitor, on the two major murine cell lines RSC96 and C2C12 derived from Schwann cells and muscle satellite cells. The results showed that SB216763 stimulated the Schwann cell migration and myotube contraction. Quantitative polymerase chain reaction results demonstrated that myelin related genes, myelin associated glycoprotein and cyclin-D1, muscle related gene myogenin and endplate-associated gene nicotinic acetylcholine receptors levels were stimulated by SB216763. Immunocytochemical staining revealed that the expressions of β-catenin in the RSC96 and C2C12 cytosolic and nuclear compartments were increased in the SB216763-treated cells. These findings confirm that the glycogen synthase kinase 3 beta inhibitor, SB216763, promoted the myelination and myotube differentiation through the Wnt/β-catenin signaling pathway and contributed to nerve remyelination and reduced denervated muscle atrophy after peripheral nerve injury.     

Bridging peripheral nerves using a deacetyl chitin conduit combined with short-term electrical stimulation

Previous studies have demonstrated that deacetyl chitin conduit nerve bridging or electrical stimulation can effectively promote the regeneration of the injured peripheral nerve. We hypoth-esized that the combination of these two approaches could result in enhanced regeneration. Rats with right sciatic nerve injury were subjected to deacetyl chitin conduit bridging combined with electrical stimulation （0.1 ms, 3 V, 20 Hz, for 1 hour）. At 6 and 12 weeks after treatment, nerve conduction velocity, myelinated axon number, ifber diameter, axon diameter and the thickness of the myelin sheath in the stimulation group were better than in the non-stimulation group. The results indicate that deacetyl chitin conduit bridging combined with temporary electrical stimu-lation can promote peripheral nerve repair.     

Role of microtubule dynamics in Wallerian degeneration and nerve regeneration after peripheral nerve injury

Wallerian degeneration,the progressive disintegration of distal axons and myelin that occurs after peripheral nerve injury,is essential for creating a permissive microenvironment for nerve regeneration,and involves cytoskeletal reconstruction.However,it is unclear whether microtubule dynamics play a role in this process.To address this,we treated cultured sciatic nerve explants,an in vitro model of Wallerian degeneration,with the microtubule-targeting agents paclitaxel and nocodazole.We found that paclitaxel-induced microtubule stabilization promoted axon and myelin degeneration and Schwann cell dedifferentiation,whereas nocodazole-induced microtubule destabilization inhibited these processes.Evaluation of an in vivo model of peripheral nerve injury showed that treatment with paclitaxel or nocodazole accelerated or attenuated axonal regeneration,as well as functional recovery of nerve conduction and target muscle and motor behavior,respectively.These results suggest that microtubule dynamics participate in peripheral nerve regeneration after injury by affecting Wallerian degeneration.This study was approved by the Animal Care and Use Committee of Southern Medical University,China(approval No.SMUL2015081) on October 15,2015.     

Hydrogen sulfide controls peripheral nerve degeneration and regeneration： a novel therapeutic strategy for peripheral demyelinating disorders or nerve degenerative diseases

<正>After peripheral nerve injury,the process of Wallerian degeneration is initiated in the distal stump of injured nerves.Wallerian degeneration in peripheral nerves involves axonal degeneration and degradation of the myelin sheath in Schwann cells.This provides the necessary conditions for axonal regeneration and remyelination.After nerve injury,macrophages are also recruited to the distal nerve stump and,together with Schwann cells,play a role in the clearance of     

Exenatide promotes regeneration of injured rat sciatic nerve

Damage to peripheral nerves results in partial or complete dysfunction. After peripheral nerve injuries, a full functional recovery usually cannot be achieved despite the standard surgical repairs. Neurotrophic factors and growth factors stimulate axonal growth and support the viability of nerve cells. The objective of this study is to investigate the neurotrophic effect of exenatide(glucagon like peptide-1 analog) in a rat sciatic nerve neurotmesis model. We injected 10 μg/d exenatide for 12 weeks in the experimental group(n = 12) and 0.1 m L/d saline for 12 weeks in the control group(n = 12). We evaluated nerve regeneration by conducting electrophysiological and motor functional tests. Histological changes were evaluated at weeks 1, 3, 6, and 9. Nerve regeneration was monitored using stereomicroscopy. The electrophysiological and motor functions in rats treated with exenatide were improved at 12 weeks after surgery. Histological examination revealed a significant increase in the number of axons in injured sciatic nerve following exenatide treatment confirmed by stereomicroscopy. In an experimentally induced neurotmesis model in rats, exenatide had a positive effect on nerve regeneration evidenced by electromyography, functional motor tests, histological and stereomicroscopic findings.     

Axonal protein synthesis in central nervous system regeneration:is building an axon a local matter?

Neurons of the mature central nervous system(CNS,mainly the brain and spinal cord)are unable to regenerate spontaneously after a lesion,in contrast to neurons of the peripheral nervous system(PNS).While the extraneuronal environment was long thought to be limiting,evidence was given less than 15 years ago that neurons themselves are critical players of their own regeneration(Park et al.,2008).Indeed,CNS neurons show a decline of axon growth capacity as they mature and after an injury.Today,the role of axonal translation is actively explored in the paradigm of embryonic neuronal growth and in peripheral nerve injury and regeneration,but less is known about the role of local protein synthesis in regrowth of adult CNS axons.Here we discuss how the current understanding of axonal translation in the CNS may contribute to the development of novel strategies to enhance axon regeneration in the injured CNS.     

Femoral nerve regeneration and its accuracy under different injury mechanisms

Surgical accuracy has greatly improved with the advent of microsurgical techniques. However, complete functional recovery after peripheral nerve injury has not been achieved to date. The mechanisms hindering accurate regeneration of damaged axons after peripheral nerve injury are in urgent need of exploration. The present study was designed to explore the mechanisms of peripheral nerve regeneration after different types of injury. Femoral nerves of rats were injured by crushing or freezing. At 2, 3, 6, and 12 weeks after injury, axons were retrogradely labeled using 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate(Dil) and True Blue, and motor and sensory axons that had regenerated at the site of injury were counted. The number and percentage of Dil-labeled neurons in the anterior horn of the spinal cord increased over time. No significant differences were found in the number of labeled neurons between the freeze and crush injury groups at any time point. Our results confirmed that the accuracy of peripheral nerve regeneration increased with time, after both crush and freeze injury, and indicated that axonal regeneration accuracy was still satisfactory after freezing, despite the prolonged damage.     

Functional and immunological peculiarities of peripheral nerve allografts

This review addresses the accumulating evidence that live(not decellularized)allogeneic peripheral nerves are functionally and immunologically peculiar in comparison with many other transplanted allogeneic tissues.This is relevant because live peripheral nerve allografts are very effective at promoting recovery after segmental peripheral nerve injury via axonal regeneration and axon fusion.Understanding the immunological peculiarities of peripheral nerve allografts may also be of interest to the field of transplantation in general.Three topics are addressed:The first discusses peripheral nerve injury and the potential utility of peripheral nerve allografts for bridging segmental peripheral nerve defects via axon fusion and axon regeneration.The second reviews evidence that peripheral nerve allografts elicit a more gradual and less severe host immune response allowing for prolonged survival and function of allogeneic peripheral nerve cells and structures.Lastly,potential mechanisms that may account for the immunological differences of peripheral nerve allografts are discussed.     

TC10 as an essential molecule in axon regeneration through membrane supply and microtubule stabilization

Mammalian central nervous system(CNS)neurons lose axon regenerative ability as they mature.This failure to regenerate shows a clear contrast to a remarkable potential of axon growth during embryonic development and after an injury in the peripheral nervous system(PNS)(Hilton and Bradke,2017).The absence of regeneration in the mature CNS neurons is caused by an inhibitory influence of the environment of the injured axons and the deficit of intrinsic factors that enable regeneration in the PNS(He and Jin,2016).     

Nerve function restoration following targeted muscle reinnervation after varying delayed periods

Targeted muscle reinnervation has been proposed for reconstruction of neuromuscular function in amputees. However, it is unknown whether performing delayed targeted muscle reinnervation after nerve injury will affect restoration of function. In this rat nerve injury study, the median and musculocutaneous nerves of the forelimb were transected. The proximal median nerve stump was sutured to the distal musculocutaneous nerve stump immediately and 2 and 4 weeks after surgery to reinnervate the bice...