Peripheral Nervous System Injury Research Articles

Novel AAV variants with improved tropism for human Schwann cells

Gene therapies and associated technologies are transforming biomedical research and enabling novel therapeutic options for patients living with debilitating and incurable genetic disorders. The vector system based on recombinant adeno-associated viral vectors (rAAV) has shown great promise in recent clinical trials for genetic diseases of multiple organs, such as the liver and the nervous system. Despite recent successes towards development of novel bioengineered AAV variants for improved transduction of primary human tissues and cells, vectors that can efficiently transduce human Schwann cells (hSCs) have yet to be identified. Here, we report the application of the functional transduction (FT)-RNA selection method in primary hSCs for the development of bespoke AAV variants for specific and efficient transgene delivery to hSCs. The two identified capsid variants, Pep2hSC1 and Pep2hSC2, show conserved potency for delivery across various in vitro, in vivo and ex vivo models of hSCs. These novel AAV capsids will serve as valuable research tools, forming the basis for therapeutic solutions for both SC-related disorders or peripheral nervous system injury.

Binding and Activation of LRP1-Dependent Cell Signaling in Schwann Cells Using a Peptide Derived from the Hemopexin Domain of MMP-9.

Schwann cells (SCs) undergo phenotypic transformation and then orchestrate nerve repair following a peripheral nervous system injury. The low-density lipoprotein receptor-related protein-1 (LRP1) is significantly upregulated in SCs in response to acute injury, activating cJun and promoting SC survival. Matrix-metalloproteinase-9 (MMP-9) is an LRP1 ligand that binds LRP1 through its hemopexin domain (PEX) and activates SC survival signaling and migration. To identify novel peptide mimetics within the hemopexin domain of MMP-9, we examined the crystal structure of PEX, synthesized four peptides, and examined their potential to bind and activate LRP1. We demonstrate that a 22 amino acid peptide, peptide 2, was the only peptide that activated Akt and ERK1/2 signaling in SCs, similar to a glutathione s-transferase (GST)-fused holoprotein, GST-PEX. Intraneural injection of peptide 2, but not vehicle, into crush-injured sciatic nerves activated cJun greater than 2.5-fold in wild-type mice, supporting that peptide 2 can activate the SC repair signaling in vivo. Peptide 2 also bound to Fc-fusion proteins containing the ligand-binding motifs of LRP1, clusters of complement-like repeats (CCRII and CCRIV). Pulldown and computational studies of alanine mutants of peptide 2 showed that positively charged lysine and arginine amino acids within the peptide are critical for stability and binding to CCRII. Collectively, these studies demonstrate that a novel peptide derived from PEX can serve as an LRP1 agonist and possesses qualities previously associated with LRP1 binding and SC signaling in vitro and in vivo.

Performance of biomarkers NF-L, NSE, Tau and GFAP in blood and cerebrospinal fluid in rat for the detection of nervous system injury.

Target organ toxicity is often a reason for attritions in nonclinical and clinical drug development. Leveraging emerging safety biomarkers in nonclinical studies provides an opportunity to monitor such toxicities early and efficiently, potentially translating to early clinical trials. As a part of the European Union's Innovative Medicines Initiative (IMI), two projects have focused on evaluating safety biomarkers of nervous system (NS) toxicity: Translational Safety Biomarker Pipeline (TransBioLine) and Neurotoxicity De-Risking in Preclinical Drug Discovery (NeuroDeRisk). Performance of fluid-based NS injury biomarker candidates neurofilament light chain (NF-L), glial fibrillary acidic protein (GFAP), neuron specific enolase (NSE) and total Tau in plasma and cerebrospinal fluid (CSF) was evaluated in 15 rat in vivo studies. Model nervous system toxicants as well as other compounds were used to evaluate sensitivity and specificity. Histopathologic assessments of nervous tissues and behavioral observations were conducted to detect and characterize NS injuries. Receiver operator characteristic (ROC) curves were generated to compare the relative performance of the biomarkers in their ability to detect NS injury. NF-L was the best performer in detecting both peripheral nervous system (PNS) and CNS injury in plasma, (AUC of 0.97-0.99; respectively). In CSF, Tau correlated the best with CNS (AUC 0.97), but not PNS injury. NSE and GFAP were suitable for monitoring CNS injury, but with lesser sensitivity. In summary, NF-L is a sensitive and specific biomarker in rats for detecting compound-induced central and peripheral NS injuries. While NF-L measurement alone cannot inform the site of the injury, addition of biomarkers like Tau and NSE and analysis in both blood and CSF can provide additional information about the origin of the NS injury. These results demonstrate the utility of emerging safety biomarkers of drug-induced NS injury in rats and provide additional supporting evidence for biomarker translation across species and potential use in clinical settings to monitor drug-induced NS injury in patients.

Guiding axon regeneration: Instructions from blood vessels

In this issue of Neuron, Bhat etal.1 unveil the temporary reawakening of an embryonic guidance program, which facilitates the alignment of blood neovessels, creating a supportive "bridge" microenvironment for axon regrowth and tissue regeneration after peripheral nervous system (PNS) injury.

Does retinal microglia respond to peripheral nervous system injury?

The nervous system is the principal integration and coordination centre of the organism receiving, processing and transmitting information to produce a response that regulates and maintains homeostasis. In the central nervous system (CNS), homeostasis is regulated by haemato‐neuronal barriers that prevent free entry to immune system cells, although there is increasing evidence that neuro‐immunological interactions during injury, inflammation and chronic peripheral disease may contribute to the establishment and development of chronic neurodegenerative pathologies at the CNS level.Inflammation plays a significant role in the resolution and sequelae of CNS damage. In recent years, special attention has been dedicated to the role and phenotype of phagocytic cells acting in the CNS and, in particular, to the study of which macrophage subpopulations contribute to excessive or detrimental inflammation or, on the contrary, to beneficial repair or resolution of the lesion.All this raises the question of how local peripheral neuroimmunological interaction induces an inflammatory response in the CNS and how this can be better understood. The study of this phenomenon in the retina is of particular interest, as the retina does not receive neural afferents from other areas of the CNS.The use of experimental models of peripheral lesions of the peripheral nervous system, such as sciatic nerve trauma, or inflammatory lesions, such as the induction of a peripheral arthritic joint process, aims to investigate the time course of the response in the CNS to local peripheral inflammation in the retina of adult mice. The results reveal qualitative and quantitative cellular changes compatible with microglial activation processes in the retina, but no significant changes occur in its neuronal population. The presence and activation of haematic mononuclear‐phagocytic cells of myelopoietic origin are also observed in the retina, where they differentiate into microglia.In the absence of local retinal injury, we postulate that the detected neuroinflammatory reaction may be due to a preventive CNS defence reaction favouring neuroprotection since microglial cells derived from the haematic monocyte/macrophage cell population may consider within the M2 macrophage subtype, characterized by their ability to suppress proinflammatory immune responses and promote the expression of neuroprotective and repair genes.On the other hand, we must consider that activation of the mononuclear phagocytic system in the retina caused by a peripheral inflammatory process could potentially be detrimental to patients with neurodegenerative retinal diseases or contribute to their development.

LKB1 and CRMP1 cooperatively promote the repair of the sciatic nerve injury.

After peripheral nervous system injury, Schwann cells (SCs) can repair axons by providing a growth-promoting microenvironment. The aim of this study is to explore the effects and mechanisms of LKB1 and CRMP1 on the repair of sciatic nerve injury (SNI). The expressions of LKB1 and CRMP1 were changed in rats with SNI from 12h to 4 weeks by hematoxylin-eosin staining, RT-PCR assay, immunohistochemical staining, and western blotting. Immunofluorescence results show that LKB1 and CRMP1 are co-localized in the regenerated axons of the sciatic nerve tissue of SNI rats. Co-immunoprecipitation indicates that LKB1 interacts with CRMP1. LKB1 interference suppresses the phosphorylation level of CRMP1. Overexpression of LKB1 and CRMP1 promotes the invasion and migration of SCs, and nerve cell protuberance extends. The structure of the myelin sheath in the sciatic nerve of the model group was found to be loose and disordered. Rats in the model group had higher pain thresholds and heat sensitivity response times than those in the control group. Nerve conduction velocity, the latency of action potential, and the peak value of compound muscle action potential in the SNI group were significantly lower than those in the control group, and the muscle atrophy was severe. Overexpression of LKB1 may significantly improve the above conditions. However, the function of LKB1 to improve SNI is abolished by the interference of CRMP1. In summary, the interaction between LKB1 and CRMP promotes the migration and differentiation of SCs and the extension of neurons, thereby improving the repair of nerve injury.

Ultrasound-Activated Piezoelectric Polyvinylidene Fluoride-Trifluoroethylene Scaffolds for Tissue Engineering Applications.

Severe peripheral nervous system (PNS) injuries have limited options for therapeutic solutions to regain functional recovery. This can be attributed in part to the lack of regeneration pathways promoted by recapitulating chemical, physical, and electrical cues to direct nerve guidance. To address this, we examined ultrasonic stimulation of a piezoelectric polyvinylidene fluoride-triflouroethylene (PVDF-TrFE) scaffold as a potentially clinically relevant therapy for PNS regeneration. Owing to the piezoelectric modality of PVDF-TrFE, we hypothesize that ultrasound stimulation will activate the scaffold to electrically stimulate cells in response to the mechanical deformation mediated by sound waves. Biocompatible PVDF-TrFE scaffolds were fabricated to be used as an ultrasound-activated, piezoelectric biomaterial to enhance cellular activity for PNS applications. NIH-3T3 fibroblasts were cultured on PVDF-TrFE nanofibers and stimulated with low-, medium-, or high-powered ultrasound. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assays were performed on fibroblasts to measure the metabolic activity of the cells following stimulation. MTT assays showed that ultrasound-stimulated fibroblasts on PVDF-TrFE scaffolds had increased metabolic activity as power was increased, whereas on plain polystyrene, an opposite trend was observed where cells had a decreased metabolic activity with ascending levels of ultrasound power. Ultrasound-stimulated PVDF-TrFE nanofibers hold exciting potential as a therapy for PNS injuries by promoting increased metabolic activity and proliferation. The ability to noninvasively stimulate implantable piezoelectric nanofibers to promote mechanical and electrical stimulation for nerve repair offers a promising benefit to severe trauma patients.

Surgical Management of Iatrogenic Nerve Root Palsy: Cervical Root

Management of permanent iatrogenic cervical nerve root palsy is a challenge faced by spine surgeons. Reconstructive surgery using nerve transfer has shown great hope in restoring somatosensory injuries such as iatrogenic cervical nerve root palsy. Nerve transfer is justified by using less important donor nerves to regain major function. Nerve transfers are cost-effective procedures increasingly used to treat central and peripheral nervous system injuries. This article provides a comprehensive reanimation strategy to restore lost functions after iatrogenic cervical nerve root palsy. After reading this article, the trainers should explain the importance of surgical reconstruction after iatrogenic cervical nerve root palsy; they should manage preoperative assessments, address various nerve transfers based on the level of injury and pattern of paralysis, and plan for postoperative rehabilitation strategy.

First person – Clara Mutschler

ABSTRACT First Person is a series of interviews with the first authors of a selection of papers published in Journal of Cell Science, helping researchers promote themselves alongside their papers. Clara Mutschler is first author on ‘Schwann cells are axo-protective after injury irrespective of myelination status in mouse Schwann cell–neuron cocultures’, published in JCS. Clara is a PhD student in the lab of Peter Arthur-Farraj at the John Van Geest Centre for Brain Repair, University of Cambridge, UK, investigating axon–Schwann cell interactions after peripheral nervous system injury.

Schwann cells are axo-protective after injury irrespective of myelination status in mouse Schwann cell-neuron cocultures.

Myelinating Schwann cell (SC)-dorsal root ganglion (DRG) neuron cocultures are an important technique for understanding cell-cell signalling and interactions during peripheral nervous system (PNS) myelination, injury, and regeneration. Although methods using rat SCs and neurons or mouse DRG explants are commonplace, there are no established protocols for compartmentalised myelinating cocultures with dissociated mouse cells. There consequently is a need for a coculture protocol that allows separate genetic manipulation of mouse SCs or neurons, or use of cells from different transgenic animals to complement in vivo mouse experiments. However, inducing myelination of dissociated mouse SCs in culture is challenging. Here, we describe a new method to coculture dissociated mouse SCs and DRG neurons in microfluidic chambers and induce robust myelination. Cocultures can be axotomised to study injury and used for drug treatments, and cells can be lentivirally transduced for live imaging. We used this model to investigate axon degeneration after traumatic axotomy and find that SCs, irrespective of myelination status, are axo-protective. At later timepoints after injury, live imaging of cocultures shows that SCs break up, ingest and clear axonal debris.

Neutrophil biology in injuries and diseases of the central and peripheral nervous systems.

The role of inflammation in nervous system injury and disease is attracting increased attention. Much of that research has focused on microglia in the central nervous system (CNS) and macrophages in the peripheral nervous system (PNS). Much less attention has been paid to the roles played by neutrophils. Neutrophils are part of the granulocyte subtype of myeloid cells. These cells, like macrophages, originate and differentiate in the bone marrow from which they enter the circulation. After tissue damage or infection, neutrophils are the first immune cells to infiltrate into tissues and are directed there by specific chemokines, which act on chemokine receptors on neutrophils. We have reviewed here the basic biology of these cells, including their differentiation, the types of granules they contain, the chemokines that act on them, the subpopulations of neutrophils that exist, and their functions. We also discuss tools available for identification and further study of neutrophils. We then turn to a review of what is known about the role of neutrophils in CNS and PNS diseases and injury, including stroke, Alzheimer's disease, multiple sclerosis, amyotrophic lateral sclerosis, spinal cord and traumatic brain injuries, CNS and PNS axon regeneration, and neuropathic pain. While in the past studies have focused on neutrophils deleterious effects, we will highlight new findings about their benefits. Studies on their actions should lead to identification of ways to modify neutrophil effects to improve health.

Quantifying Hand Strength and Isometric Pinch Individuation Using a Flexible Pressure Sensor Grid.

Modulating force between the thumb and another digit, or isometric pinch individuation, is critical for daily tasks and can be impaired due to central or peripheral nervous system injury. Because surgical and rehabilitative efforts often focus on regaining this dexterous ability, we need to be able to consistently quantify pinch individuation across time and facilities. Currently, a standardized metric for such an assessment does not exist. Therefore, we tested whether we could use a commercially available flexible pressure sensor grid (Tekscan F-Socket [Tekscan Inc., Norwood, MA, USA]) to repeatedly measure isometric pinch individuation and maximum voluntary contraction (MVC) in twenty right-handed healthy volunteers at two visits. We developed a novel equation informed by the prior literature to calculate isometric individuation scores that quantified percentage of force on the grid generated by the indicated digit. MVC intra-class correlation coefficients (ICCs) for the left and right hands were 0.86 (p < 0.0001) and 0.88 (p < 0.0001), respectively, suggesting MVC measurements were consistent over time. However, individuation score ICCs, were poorer (left index ICC 0.41, p = 0.28; right index ICC -0.02, p = 0.51), indicating that this protocol did not provide a sufficiently repeatable individuation assessment. These data support the need to develop novel platforms specifically for repeatable and objective isometric hand dexterity assessments.

The Synergistic Healing Function of Regular Treadmill Exercise and Nanofibrous Scaffolds on Peripheral Nervous System Injury: An In Vitro and In Vivo Study

In the current study, injured peripheral nerves were treated with a regular treadmill exercise program in combination with a nanofibrous neural channels loaded with arctium lappa extract (ALE). In Vitro studies showed that the neural channels were biocompatible and potentially facilitated gas exchange through the scaffolds. In addition, the produced scaffolds modulated the inflammatory responses. In Vivo study showed that the animals treated with both exercise and neural channels had the highest rate of functional recovery and histopathological restorations.

Morbidity of single-fraction spine radiosurgery for metastatic disease.

e24116 Background: The optimal management of metastatic spine disease involves radiation therapy, with stereotactic radiosurgery (SRS) utilization having increased in adoption while proving to be efficacious and safe. Spine SRS morbidity (fatigue, dermatitis, nausea and esophagitis) is generally mild and depends on the spinal vertebral level(s) treated. More severe adverse events such as vertebral compression fracture (VCF), radiation-induced myelopathy, and tracheoesophageal (TE) fistula have been documented. The potential benefits of single-fraction SRS (sfSRS) over multi-fraction SRS include increased patient convenience, decreased financial toxicity, and increased treatment completion rates. These benefits may potentially be overshadowed by the morbidity of sfSRS versus multifraction SRS. In this analysis, we perform the first comprehensive examination of sfSRS morbidity in the treatment of spinal lesions. Methods: An extensive search of the PubMed database was undertaken through 2/2/23 to find studies examining spinal sfSRS. Search parameters included the terms “single-fraction, spine, radiation SRS OR SBRT” Articles including multi-fractionated regimens only, reviewed sites outside of the spine, or did not report on clinical outcomes/toxicities were excluded. Results: 17 articles met initial inclusion criteria; 9 were added from references for a total of 26 articles comprising at least 2,699 patients (one study reported the number of tumors, not patients). Dose ranges for single-fraction SRS were 8-26 Gy. Commonly noted toxicities were fatigue, nausea/vomiting, and dermatitis which were no worse than moderate/minimal, not requiring invasive intervention. The overall rate of VCF was 16.2%, with an increased propensity at &gt;20 Gy. The proportion of reported segments requiring surgical intervention (operative stabilization, vertebroplasty/kyphoplasty) for VCF after single-fraction SRS was 20.2% (33/163). The rate of TE fistula at 25 Gy was 8.3%, with an overall rate (dose range = 10-28 Gy) of 1.3% with up to 12-years of follow-up. In patients receiving median dose of 24 Gy, the rate of severe esophagitis was 6.8%, myositis was 1.3% (at 1-year) and 8.3% (at 5-years). The rate of peripheral nervous system injury was 2.5% (at 18-26 Gy) and pain flare was 8% (at 16-24 Gy). The rate of ureteral and esophageal stricture was 1.9% each in patients receiving 12-24 Gy. Conclusions: VCF is the most commonly reported severe adverse events with single-fraction spine SRS, particularly for doses &gt; 20 Gy. 80% of VCFs can be successfully managed without invasive intervention. Other severe adverse events are rare – particularly for &lt; 20 Gy – but require post-SRS monitoring to expediently and efficiently intervene if necessary. These findings indicate that single-fraction spine SRS to &lt; 20 Gy is safe with minimal morbidity. Further study regarding the efficacy of such dosing is being evaluated in ongoing clinical trials.

206 High Resolution Ultrasound for Evaluation of Peripheral Nerve Injuries: Retrospective Examination of an Emerging Technology

INTRODUCTION: Recovery from peripheral nervous system injury (PNI) after trauma has been shown to improve with earlier detection and treatment. Unfortunately, surgical treatments are usually postponed several months after injury due to the need to evaluate for potential recovery without intervention. If diagnostic modalities are able to identify which nerve injuries will require surgical intervention sooner, we may be able to significantly improve outcomes after peripheral nerve injury. High-resolution ultrasound (HRUS) is an imaging modality that potentially could provide this in an economic manner. METHODS: A retrospective chart review was conducted for all patients who received an HRUS of their PNI at our institution from 01/01/2020-01/31/2022. RESULTS: A total of 32 patients were included. The mean age was 45 years with a male predominance (69%). Neuronal and surrounding tissue pathology greater than or equal to the fascicular level could be identified with HRUS. EDx studies were available for 28 (88%) and clinical exams were available for 32 (100%) of patients. Comparing HRUS to the gold standard of a combination of clinical exam and EMG findings, HRUS had a sensitivity of 94% at identifying PNI. Ultimately, 19 (59%) of patients evaluated using HRUS underwent operative intervention. CONCLUSIONS: HRUS is an important emerging imaging modality that can identify and characterize PNI. Our study demonstrates the high sensitivity of HRUS in detecting PNI. Additionally, HRUS in our study identified a number of features that may serve as surrogate markers of injury useful in prognosticating recovery. Further work is needed to define the earliest time point HRUS can be used to identify a PNI.

Study on antibacterial activity and repair of peripheral nervous system injury by Zno nanocomposites

In this paper, three samples of ZnO/PPy, ZnO/CNTs and ZnO/PPy/CNTs nano- composites were studied in the aspects of antimicrobial properties, cell imaging and tumor targeting. Firstly, Escherichia coli strains were selected and controlled test method was used. Colony data showed that the inhibition rate of ZnO/PPy was 73.06%, the inhibition rate of ZnO /CNTs was 99.44%, and the inhibition rate of ZnO/PPy/CNTs composite was 99.99%. The results showed that the three kinds of composites had certain antibacterial effect. At the same time, in order to optimize the application of nano-sized zinc oxide (ZnOs), ZnOs nanocomposite microspheres were prepared by emulsification and dispersion method with polypyrrole and carbon nanotubes as microballoon walls, and the animal model of peripheral nervous system (PNS) injury was established. ZnOs nanocomposite microspheres and control samples were implanted into the injured peripheral nervous system (sciatic nerve was used as the site). The experimental results show that ZnOs nanocomposite microspheres can repair peripheral nervous system injury in animals.

The best of both worlds: mastering nerve regeneration combining biological and nanotechnological tools.

The best of both worlds: mastering nerve regeneration combining biological and nanotechnological tools.

All the PNS is a Stage: Transplanted Bone Marrow Cells Play an Immunomodulatory Role in Peripheral Nerve Regeneration.

Bone marrow cell transplant has proven to be an effective therapeutic approach to treat peripheral nervous system injuries as it not only promoted regeneration and remyelination of the injured nerve but also had a potent effect on neuropathic pain.

Regeneration of sciatic nerve injury through nanofiber neural guidance channels containing collagen hydrogel and acetyl L carnitine: An in vitro and in vivo study

Unlike the central nervous system (CNS), peripheral nervous system (PNS) injuries are partially repairable. Nerve guidance channels (NGCs) have been shown to improve the level of nerve repair after injury. In the present study, we developed a nanofiber NGC for the delivery of acetyl L carnitine (ALC) in a rat model of sciatic nerve injury. NGCs were produced by electrospinning a polymer blend of polycaprolacton and gelatin. The physicochemical and biological properties of developed scaffolds were investigated using Scanning electron microscopy, surface hydrophilicity measurement, porosity measurement, tensile strength studies, cell viability assay, and cell attachment assay. ALC was included in the collagen hydrogels at three weight ratios of 1%, 3%, and 5%. Cell viability assay showed that the hydrogels containing 5% ALC demonstrated a more favorable effect on PC-12 metabolic activity. Therefore, this concentration was chosen to treat PNS injury. The NGCs were implanted in rats and then their lumen was filled with collagen hydrogel + 5%ALC. The results of histopathological examinations and functional recovery studies showed that NGCs filled with ALC containing hydrogel have significant recovery potential compared to NGCs loaded with collagen hydrogels without ALC. Our results support the potential use of ALC-delivering NGCs in the treatment of peripheral nerve injury in the clinic.

The impact of physical, biochemical, and electrical signaling on Schwann cell plasticity

Peripheral nervous system (PNS) injuries are an ongoing health care concern. While autografts and allografts are regarded as the current clinical standard for traumatic injury, there are inherent limitations that suggest alternative remedies should be considered for therapeutic purposes. In recent years, nerve guidance conduits (NGCs) have become increasingly popular as surgical repair devices, with a multitude of various natural and synthetic biomaterials offering potential to enhance the design of conduits or supplant existing technologies entirely. From a cellular perspective, it has become increasingly evident that Schwann cells (SCs), the primary glia of the PNS, are a predominant factor mediating nerve regeneration. Thus, the development of severe nerve trauma therapies requires a deep understanding of how SCs interact with their environment, and how SC microenvironmental cues may be engineered to enhance regeneration. Here we review the most recent advancements in biomaterials development and cell stimulation strategies, with a specific focus on how the microenvironment influences the behavior of SCs and can potentially lead to functional repair. We focus on microenvironmental cues that modulate SC morphology, proliferation, migration, and differentiation to alternative phenotypes. Promotion of regenerative phenotypic responses in SCs and other non-neuronal cells that can augment the regenerative capacity of multiple biomaterials is considered along with innovations and technologies for traumatic injury.