Axonal Fusion Research Articles

Polyethylene glycol has immunoprotective effects on sciatic allografts, but behavioral recovery and graft tolerance require neurorrhaphy and axonal fusion.

JOURNAL/nrgr/04.03/01300535-202504000-00033/figure1/v/2024-07-06T104127Z/r/image-tiff Behavioral recovery using (viable) peripheral nerve allografts to repair ablation-type (segmental-loss) peripheral nerve injuries is delayed or poor due to slow and inaccurate axonal regeneration. Furthermore, such peripheral nerve allografts undergo immunological rejection by the host immune system. In contrast, peripheral nerve injuries repaired by polyethylene glycol fusion of peripheral nerve allografts exhibit excellent behavioral recovery within weeks, reduced immune responses, and many axons do not undergo Wallerian degeneration. The relative contribution of neurorrhaphy and polyethylene glycol-fusion of axons versus the effects of polyethylene glycol per se was unknown prior to this study. We hypothesized that polyethylene glycol might have some immune-protective effects, but polyethylene glycol-fusion was necessary to prevent Wallerian degeneration and functional/behavioral recovery. We examined how polyethylene glycol solutions per se affect functional and behavioral recovery and peripheral nerve allograft morphological and immunological responses in the absence of polyethylene glycol-induced axonal fusion. Ablation-type sciatic nerve injuries in outbred Sprague-Dawley rats were repaired according to a modified protocol using the same solutions as polyethylene glycol-fused peripheral nerve allografts, but peripheral nerve allografts were loose-sutured (loose-sutured polyethylene glycol) with an intentional gap of 1-2 mm to prevent fusion by polyethylene glycol of peripheral nerve allograft axons with host axons. Similar to negative control peripheral nerve allografts not treated by polyethylene glycol and in contrast to polyethylene glycol-fused peripheral nerve allografts, animals with loose-sutured polyethylene glycol peripheral nerve allografts exhibited Wallerian degeneration for all axons and myelin degeneration by 7 days postoperatively and did not recover sciatic-mediated behavioral functions by 42 days postoperatively. Other morphological signs of rejection, such as collapsed Schwann cell basal lamina tubes, were absent in polyethylene glycol-fused peripheral nerve allografts but commonly observed in negative control and loose-sutured polyethylene glycol peripheral nerve allografts at 21 days postoperatively. Loose-sutured polyethylene glycol peripheral nerve allografts had more pro-inflammatory and less anti-inflammatory macrophages than negative control peripheral nerve allografts. While T cell counts were similarly high in loose-sutured-polyethylene glycol and negative control peripheral nerve allografts, loose-sutured polyethylene glycol peripheral nerve allografts expressed some cytokines/chemokines important for T cell activation at much lower levels at 14 days postoperatively. MHCI expression was elevated in loose-sutured polyethylene glycol peripheral nerve allografts, but MHCII expression was modestly lower compared to negative control at 21 days postoperatively. We conclude that, while polyethylene glycol per se reduces some immune responses of peripheral nerve allografts, successful polyethylene glycol-fusion repair of some axons is necessary to prevent Wallerian degeneration of those axons and immune rejection of peripheral nerve allografts, and produce recovery of sensory/motor functions and voluntary behaviors. Translation of polyethylene glycol-fusion technologies would produce a paradigm shift from the current clinical practice of waiting days to months to repair ablation peripheral nerve injuries.

Polyethylene Glycol-Mediated Axonal Fusion Promotes Early Sensory Recovery after Digital Nerve Injury: A Randomized Clinical Trial.

Peripheral nerve repair is limited by Wallerian degeneration coupled with the slow and inconsistent rates of nerve regrowth. In more proximal injuries, delayed nerve regeneration can cause debilitating muscle atrophy. Topical application of polyethylene glycol (PEG) during neurorrhaphy facilitates the fusion of severed axonal membranes, immediately restoring action potentials across the coaptation site. In preclinical animal models, PEG fusion resulted in remarkable early functional recovery. This is the first randomized clinical trial comparing functional outcomes between PEG fusion and standard neurorrhaphy. Participants with digital nerve transections were followed up at 2 weeks, 1 month, and 3 months postoperatively. The primary outcome was assessed using the Medical Research Council Classification (MRCC) rating for sensory recovery at each time point. Semmes-Weinstein monofilaments and static 2-point discrimination determined MRCC ratings. Postoperative quality of life was measured using the Michigan Hand Outcomes Questionnaire. Forty-eight transected digital nerves (25 control and 23 PEG) across 22 patients were analyzed. PEG-fused nerves demonstrated significantly higher MRCC scores at 2 weeks (OR, 16.95; 95% CI, 1.79 to 160.38; P = 0.008) and 1 month (OR, 13.40; 95% CI, 1.64 to 109.77; P = 0.009). Participants in the PEG cohort also had significantly higher average Michigan Hand Outcomes Questionnaire scores at 2 weeks (Hodge g , 1.28; 95% CI, 0.23 to 2.30; P = 0.0163) and 1 month (Hodge g , 1.02; 95% CI, 0.04 to 1.99; P = 0.049). No participants had adverse events related to the study drug. PEG fusion promotes early sensory recovery and improved patient well-being following peripheral nerve repair of digital nerves. Therapeutic, II.

The dynamin GTPase mediates regenerative axonal fusion in Caenorhabditis elegans by regulating fusogen levels.

Axonal fusion is a neuronal repair mechanism that results in the reconnection of severed axon fragments, leading to the restoration of cytoplasmic continuity and neuronal function. While synaptic vesicle recycling has been linked to axonal regeneration, its role in axonal fusion remains unknown. Dynamin proteins are large GTPases that hydrolyze lipid-binding membranes to carry out clathrin-mediated synaptic vesicle recycling. Here, we show that the Caenorhabditis elegans dynamin protein DYN-1 is a key component of the axonal fusion machinery. Animals carrying a temperature-sensitive allele of dyn-1(ky51) displayed wild-type levels of axonal fusion at the permissive temperature (15°C) but presented strongly reduced levels at the restrictive temperature (25°C). Furthermore, the average length of regrowth was significantly diminished in dyn-1(ky51) animals at the restrictive temperature. The expression of wild-type DYN-1 cell-autonomously into dyn-1(ky51) mutant animals rescued both the axonal fusion and regrowth defects. Furthermore, DYN-1 was not required prior to axonal injury, suggesting that it functions specifically after injury to control axonal fusion. Finally, using epistatic analyses and superresolution imaging, we demonstrate that DYN-1 regulates the levels of the fusogen protein EFF-1 post-injury to mediate axonal fusion. Together, these results establish DYN-1 as a novel regulator of axonal fusion.

Coordinated NADPH oxidase/hydrogen peroxide functions regulate cutaneous sensory axon de- and regeneration

Tissue wounding induces cutaneous sensory axon regeneration via hydrogen peroxide (H2O2) that is produced by the epithelial NADPH oxidase, Duox1. Sciatic nerve injury instead induces axon regeneration through neuronal uptake of the NADPH oxidase, Nox2, from macrophages. We therefore reasoned that the tissue environment in which axons are damaged stimulates distinct regenerative mechanisms. Here, we show that cutaneous axon regeneration induced by tissue wounding depends on both neuronal and keratinocyte-specific mechanisms involving H2O2 signaling. Genetic depletion of H2O2 in sensory neurons abolishes axon regeneration, whereas keratinocyte-specific H2O2 depletion promotes axonal repulsion, a phenotype mirrored in duox1 mutants. Intriguingly, cyba mutants, deficient in the essential Nox subunit, p22Phox, retain limited axon regenerative capacity but display delayed Wallerian degeneration and axonal fusion, observed so far only in invertebrates. We further show that keratinocyte-specific oxidation of the epidermal growth factor receptor (EGFR) at a conserved cysteine thiol (C797) serves as an attractive cue for regenerating axons, leading to EGFR-dependent localized epidermal matrix remodeling via the matrix-metalloproteinase, MMP-13. Therefore, wound-induced cutaneous axon de- and regeneration depend on the coordinated functions of NADPH oxidases mediating distinct processes following injury.

The metalloprotease ADM-4/ADAM17 promotes axonal repair.

Axonal fusion is an efficient means of repair following axonal transection, whereby the regenerating axon fuses with its own separated axonal fragment to restore neuronal function. Despite being described over 50 years ago, its molecular mechanisms remain poorly understood. Here, we demonstrate that the Caenorhabditis elegans metalloprotease ADM-4, an ortholog of human ADAM17, is essential for axonal fusion. We reveal that animals lacking ADM-4 cannot repair their axons by fusion, and that ADM-4 has a cell-autonomous function within injured neurons, localizing at the tip of regrowing axon and fusion sites. We demonstrate that ADM-4 overexpression enhances fusion to levels higher than wild type, and that the metalloprotease and phosphatidylserine-binding domains are essential for its function. Last, we show that ADM-4 interacts with and stabilizes the fusogen EFF-1 to allow membranes to merge. Our results uncover a key role for ADM-4 in axonal fusion, exposing a molecular target for axonal repair.

Neurorrhaphy in Presence of Polyethylene Glycol Enables Immediate Electrophysiological Conduction in Porcine Model of Facial Nerve Injury.

Facial nerve trauma often leads to disfiguring facial muscle paralysis. Despite several promising advancements, facial nerve repair procedures often do not lead to complete functional recovery. Development of novel repair strategies requires testing in relevant preclinical models that replicate key clinical features. Several studies have reported that fusogens, such as polyethylene glycol (PEG), can improve functional recovery by enabling immediate reconnection of injured axons; however, these findings have yet to be demonstrated in a large animal model. We first describe a porcine model of facial nerve injury and repair, including the relevant anatomy, surgical approach, and naive nerve morphometry. Next, we report positive findings from a proof-of-concept experiment testing whether a neurorrhaphy performed in conjunction with a PEG solution maintained electrophysiological nerve conduction at an acute time point in a large animal model. The buccal branch of the facial nerve was transected and then immediately repaired by direct anastomosis and PEG application. Immediate electrical conduction was recorded in the PEG-fused nerves (n = 9/9), whereas no signal was obtained in a control cohort lacking calcium chelating agent in one step (n = 0/3) and in the no PEG control group (n = 0/5). Nerve histology revealed putative-fused axons across the repair site, whereas no positive signal was observed in the controls. Rapid electrophysiological recovery following nerve fusion in a highly translatable porcine model of nerve injury supports previous studies suggesting neurorrhaphy supplemented with PEG may be a promising strategy for severe nerve injury. While acute PEG-mediated axon conduction is promising, additional work is necessary to determine if physical axon fusion occurs and the longer-term fate of distal axon segments as related to functional recovery.

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.

Closing the Gap Between Mammalian and Invertebrate Peripheral Nerve Injury: Protocol for a Novel Nerve Repair.

BackgroundOutcomes after peripheral nerve injuries are poor despite current nerve repair techniques. Currently, there is no conclusive evidence that mammalian axons are capable of spontaneous fusion after transection. Notably, certain invertebrate species are able to auto-fuse after transection. Although mammalian axonal auto-fusion has not been observed experimentally, no mammalian study to date has demonstrated regenerating axolemmal membranes contacting intact distal segment axolemmal membranes to determine whether mammalian peripheral nerve axons have the intrinsic mechanisms necessary to auto-fuse after transection.ObjectiveThis study aims to assess fusion competence between regenerating axons and intact distal segment axons by enhancing axon regeneration, delaying Wallerian degeneration, limiting the immune response, and preventing myelin obstruction.MethodsThis study will use a rat sciatic nerve model to evaluate the effects of a novel peripheral nerve repair protocol on behavioral, electrophysiologic, and morphologic parameters. This protocol consists of a variety of preoperative, intraoperative, and postoperative interventions. Fusion will be assessed with electrophysiological conduction of action potentials across the repaired transection site. Axon-axon contact will be assessed with transmission electron microscopy. Behavioral recovery will be analyzed with the sciatic functional index. A total of 36 rats will be used for this study. The experimental group will use 24 rats and the negative control group will use 12 rats. For both the experimental and negative control groups, there will be both a behavior group and another group that will undergo electrophysiological and morphological analysis. The primary end point will be the presence or absence of action potentials across the lesion site. Secondary end points will include behavioral recovery with the sciatic functional index and morphological analysis of axon-axon contact between regenerating axons and intact distal segment axons.ResultsThe author is in the process of grant funding and institutional review board approval as of March 2020. The final follow-up will be completed by December 2021.ConclusionsIn this study, the efficacy of the proposed novel peripheral nerve repair protocol will be evaluated using behavioral and electrophysiologic parameters. The author believes this study will provide information regarding whether spontaneous axon fusion is possible in mammals under the proper conditions. This information could potentially be translated to clinical trials if successful to improve outcomes after peripheral nerve injury.International Registered Report Identifier (IRRID)PRR1-10.2196/18706

Stabilin Receptors: Role as Phosphatidylserine Receptors.

Phosphatidylserine is a membrane phospholipid that is localized to the inner leaflet of the plasma membrane. Phosphatidylserine externalization to the outer leaflet of the plasma membrane is an important signal for various physiological processes, including apoptosis, platelet activation, cell fusion, lymphocyte activation, and regenerative axonal fusion. Stabilin-1 and stabilin-2 are membrane receptors that recognize phosphatidylserine on the cell surface. Here, we discuss the functions of Stabilin-1 and stabilin-2 as phosphatidylserine receptors in apoptotic cell clearance (efferocytosis) and cell fusion, and their ligand-recognition and signaling pathways.

Disruption of RAB-5 Increases EFF-1 Fusogen Availability at the Cell Surface and Promotes the Regenerative Axonal Fusion Capacity of the Neuron.

Following a transection injury to the axon, neurons from a number of species have the ability to undergo spontaneous repair via fusion of the two separated axonal fragments. In the nematode Caenorhabditis elegans, this highly efficient regenerative axonal fusion is mediated by epithelial fusion failure-1 (EFF-1), a fusogenic protein that functions at the membrane to merge the two axonal fragments. Identifying modulators of axonal fusion and EFF-1 is an important step toward a better understanding of this repair process. Here, we present evidence that the small GTPase RAB-5 acts to inhibit axonal fusion, a function achieved via endocytosis of EFF-1 within the injured neuron. Therefore, we find that perturbing RAB-5 activity is sufficient to restore axonal fusion in mutant animals with decreased axonal fusion capacity. This is accompanied by enhanced membranous localization of EFF-1 and the production of extracellular EFF-1-containing vesicles. These findings identify RAB-5 as a novel regulator of axonal fusion in C. elegans hermaphrodites and the first regulator of EFF-1 in neurons.SIGNIFICANCE STATEMENT Peripheral and central nerve injuries cause life-long disabilities due to the fact that repair rarely leads to reinnervation of the target tissue. In the nematode Caenorhabditis elegans, axonal regeneration can proceed through axonal fusion, whereby a regrowing axon reconnects and fuses with its own separated distal fragment, restoring the original axonal tract. We have characterized axonal fusion and established that the fusogen epithelial fusion failure-1 (EFF-1) is a key element for fusing the two separated axonal fragments back together. Here, we show that the small GTPase RAB-5 is a key cell-intrinsic regulator of the fusogen EFF-1 and can in turn regulate axonal fusion. Our findings expand the possibility for this process to be controlled and exploited to facilitate axonal repair in medical applications.

Optimization and evaluation of an in vitro model of PEG-mediated fusion of nerve cell bodies

Polyethylene glycol (PEG) is a cell membrane fusogen (de StGroth et al., 1980; Hoehn et al., 1978 [2]; Pontecorvo et al., 1975 [3]). In clinical application PEG allows some axonal fusion in peripheral nerve repairs, resulting in retention of some mobility and sensation during the regeneration process and reducing muscular atrophy. Several manuscripts exist concerning in vivo models of PEG fusion, while there is a distinct lack of in vitro studies. This study optimizes an experimental in vitro PEG fusion procedure in B35 rat neuroblastoma cells, through alteration of PEG and dimethyl sulfoxide (DMSO) concentrations and utilizes flow cytometry for assessment. The optimized procedure was then tested on B104-1-1 cells, a mouse neuroblastoma line expressing P2X7R, to validate our in vitro procedure/model against previous in vivo testing of P2X7R modulators, Brilliant Blue FCF (FCF) and bzATP. In brief, two cell populations were differentially stained, mixed, pre-treated (if applicable) and PEG-fused. Initial optimization generated the highest fusion rates with 70% PEG + 0% DMSO. Subsequent testing of the model with FCF and bzATP proved that assessment over time is required when evaluating potential modulators of PEG fusion. Our conclusion, at 72 h post PEG fusion, is that the optimized procedure is suitable for initial candidate testing of modulators of PEG fusion. In vitro experiments with FCF and bzATP corroborated previous in vivo findings and confirmed that DMSO is not required to aid PEG fusion in the in vitro model. The development of this cellular based in vitro model will be invaluable as a substitute for initial assessment of chemical modulators of PEG fusion.

Effect of exogenous spastin combined with polyethylene glycol on sciatic nerve injury.

Polyethylene glycol can connect the distal and proximal ends of an injured nerve at the cellular level through axonal fusion to avoid Wallerian degeneration of the injured distal nerve and promote peripheral nerve regeneration. However, this method can only prevent Wallerian degeneration in 10% of axons because the cytoskeleton is not repaired in a timely fashion. Reconstruction of the cytoskeletal trunk and microtubule network has been suggested to be the key for improving the efficiency of axonal fusion. As a microtubule-severing protein, spastin has been used to enhance cytoskeletal reconstruction. Therefore, we hypothesized that spastin combined with polyethylene glycol can more effectively promote peripheral nerve regeneration. A total of 120 male Sprague-Dawley rats were randomly divided into sham, suture, polyethylene glycol, and polyethylene glycol + spastin groups. In suture group rats, only traditional nerve anastomosis of the end-to-end suture was performed after transection of the sciatic nerve. In polyethylene glycol and polyethylene glycol + spastin groups, 50 μL of polyethylene glycol or 25 μL of polyethylene glycol + 25 μL of spastin, respectively, were injected immediately under the epineurium of the distal suture. Sensory fiber regeneration distance, which was used to assess early nerve regeneration at 1 week after surgery, was shortest in the suture group, followed by polyethylene glycol group and greatest in the polyethylene glycol + spastin group. Behavioral assessment of motor function recovery in rats showed that limb function was restored in polyethylene glycol and polyethylene glycol + spastin groups at 8 weeks after surgery. At 1, 2, 4 and 8 weeks after surgery, sciatic functional index values and percentages of gastrocnemius muscle wet weight were highest in the sham group, followed by polyethylene glycol + spastin and polyethylene glycol groups, and lowest in the suture group. Masson staining was utilized to assess the morphology of muscle tissue. Morphological changes in skeletal muscle were detectable in suture, polyethylene glycol, and polyethylene glycol + spastin groups at 1, 2, 4, and 8 weeks after surgery. Among them, muscular atrophy of the suture group was most serious, followed by polyethylene glycol and polyethylene glycol + spastin groups. Ultrastructure of distal sciatic nerve tissue, as detected by transmission electron microscopy, showed a pattern of initial destruction, subsequent disintegration, and gradual repair in suture, polyethylene glycol, and polyethylene glycol + spastin groups at 1, 2, 4, and 8 weeks after surgery. As time proceeded, axonal ultrastructure gradually recovered. Indeed, the polyethylene glycol + spastin group was similar to the sham group at 8 weeks after surgery. Our findings indicate that the combination of polyethylene glycol and spastin can promote peripheral nerve regeneration. Moreover, the effect of this combination was better than that of polyethylene glycol alone, and both were superior to the traditional neurorrhaphy. This study was approved by the Animal Ethics Committee of the Second Military Medical University, China (approval No. CZ20170216) on March 16, 2017.

Bridging the gap: axonal fusion drives rapid functional recovery of the nervous system.

Injuries to the central or peripheral nervous system frequently cause long-term disabilities because damaged neurons are unable to efficiently self-repair. This inherent deficiency necessitates the need for new treatment options aimed at restoring lost function to patients. Compared to humans, a number of species possess far greater regenerative capabilities, and can therefore provide important insights into how our own nervous systems can be repaired. In particular, several invertebrate species have been shown to rapidly initiate regeneration post-injury, allowing separated axon segments to re-join. This process, known as axonal fusion, represents a highly efficient repair mechanism as a regrowing axon needs to only bridge the site of damage and fuse with its separated counterpart in order to re-establish its original structure. Our recent findings in the nematode Caenorhabditis elegans have expanded the promise of axonal fusion by demonstrating that it can restore complete function to damaged neurons. Moreover, we revealed the importance of injury-induced changes in the composition of the axonal membrane for mediating axonal fusion, and discovered that the level of axonal fusion can be enhanced by promoting a neuron's intrinsic growth potential. A complete understanding of the molecular mechanisms controlling axonal fusion may permit similar approaches to be applied in a clinical setting.

Phosphatidylserine save-me signals drive functional recovery of severed axons in Caenorhabditis elegans

Functional regeneration after axonal injury requires transected axons to regrow and reestablish connection with their original target tissue. The spontaneous regenerative mechanism known as axonal fusion provides a highly efficient means of achieving targeted reconnection, as a regrowing axon is able to recognize and fuse with its own detached axon segment, thereby rapidly reestablishing the original axonal tract. Here, we use behavioral assays and fluorescent reporters to show that axonal fusion enables full recovery of function after axotomy of Caenorhabditis elegans mechanosensory neurons. Furthermore, we reveal that the phospholipid phosphatidylserine, which becomes exposed on the damaged axon to function as a "save-me" signal, defines the level of axonal fusion. We also show that successful axonal fusion correlates with the regrowth potential and branching of the proximal fragment and with the retraction length and degeneration of the separated segment. Finally, we identify discrete axonal domains that vary in their propensity to regrow through fusion and show that the level of axonal fusion can be genetically modulated. Taken together, our results reveal that axonal fusion restores full function to injured neurons, is dependent on exposure of phospholipid signals, and is achieved through the balance between regenerative potential and level of degeneration.

Let-7 miRNA controls CED-7 homotypic adhesion and EFF-1–mediated axonal self-fusion to restore touch sensation following injury

Neuronal injury often leads to devastating consequences such as loss of senses or locomotion. Restoration of function after injury relies on whether the injured axons can find their target cells. Although fusion between injured proximal axon and distal fragment has been observed in many organisms, its functional significance is not clear. Here, using Caenorhabditis elegans mechanosensory neurons, we address this question. Using two femtosecond lasers simultaneously, we could scan and sever posterior lateral microtubule neurons [posterior lateral microtubules (PLMs)] on both sides of the worm. We showed that axotomy of both PLMs leads to a dramatic loss of posterior touch sensation. During the regenerative phase, only axons that fuse to their distal counterparts contribute to functional recovery. Loss of let-7 miRNA promotes functional restoration in both larval and adult stages. In the L4 stage, loss of let-7 increases fusion events by increasing the mRNA level of one of the cell-recognition molecules, CED-7. The ability to establish cytoplasmic continuity between the proximal and distal ends declines with age. Loss of let-7 overcomes this barrier by promoting axonal transport and enrichment of the EFF-1 fusogen at the growing tip of cut processes. Our data reveal the functional property of a regenerating neuron.

Immediate Enhancement of Nerve Function Using a Novel Axonal Fusion Device After Neurotmesis.

BackgroundThe management of peripheral nerve injuries remains a large challenge for plastic surgeons. With the inability to fuse axonal endings, results after microsurgical nerve repair have been inconsistent. Our current nerve repair strategies rely upon the slow and lengthy process of axonal regeneration (~1 mm/d). Polyethylene glycol (PEG) has been investigated as a potential axonal fusion agent; however, the percentage of axonal fusion has been inconsistent. The purpose of this study was to identify a PEG delivery device to standardize outcomes after attempted axonal fusion with PEG.Materials and MethodsWe used a rat sciatic nerve injury model in which we completely transected and repaired the left sciatic nerve to evaluate the efficacy of PEG fusion over a span of 12 weeks. In addition, we evaluated the effectiveness of a delivery device's ability to optimize results after PEG fusion.ResultsWe found that PEG rapidly (within minutes) restores axonal continuity as assessed by electrophysiology, fluorescent retrograde tracer, and diffusion tensor imaging. Immunohistochemical analysis shows that motor axon counts are significantly increased at 1 week, 4 weeks, and 12 weeks postoperatively in PEG-treated animals. Furthermore, PEG restored behavioral functions up to 50% compared with animals that received the criterion standard epineurial repair (control animals).ConclusionsThe ability of PEG to rapidly restore nerve function after neurotmesis could have vast implications on the clinical management of traumatic injuries to peripheral nerves.

Polyethylene glycol restores axonal conduction after corpus callosum transection.

Polyethylene glycol (PEG) has been shown to restore axonal continuity after peripheral nerve transection in animal models. We hypothesized that PEG can also restore axonal continuity in the central nervous system. In this current experiment, coronal sectioning of the brains of Sprague-Dawley rats was performed after animal sacrifice. 3Brain high-resolution microelectrode arrays (MEA) were used to measure mean firing rate (MFR) and peak amplitude across the corpus callosum of the ex-vivo brain slices. The corpus callosum was subsequently transected and repeated measurements were performed. The cut ends of the corpus callosum were still apposite at this time. A PEG solution was applied to the injury site and repeated measurements were performed. MEA measurements showed that PEG was capable of restoring electrophysiology signaling after transection of central nerves. Before injury, the average MFRs at the ipsilateral, midline, and contralateral corpus callosum were 0.76, 0.66, and 0.65 spikes/second, respectively, and the average peak amplitudes were 69.79, 58.68, and 49.60 μV, respectively. After injury, the average MFRs were 0.71, 0.14, and 0.25 spikes/second, respectively and peak amplitudes were 52.11, 8.98, and 16.09 μV, respectively. After application of PEG, there were spikes in MFR and peak amplitude at the injury site and contralaterally. The average MFRs were 0.75, 0.55, and 0.47 spikes/second at the ipsilateral, midline, and contralateral corpus callosum, respectively and peak amplitudes were 59.44, 45.33, 40.02 μV, respectively. There were statistically differences in the average MFRs and peak amplitudes between the midline and non-midline corpus callosum groups (P < 0.01, P < 0.05). These findings suggest that PEG restores axonal conduction between severed central nerves, potentially representing axonal fusion.

A novel therapy to promote axonal fusion in human digital nerves.

Peripheral nerve injury can have a devastating impact on our military and veteran population. Current strategies for peripheral nerve repair include techniques such as nerve tubes, nerve grafts, tissue matrices, and nerve growth guides to enhance the number of regenerating axons. Even with such advanced techniques, it takes months to regain function. In animal models, polyethylene glycol (PEG) therapy has shown to improve both physiologic and behavioral outcomes after nerve transection by fusion of a portion of the proximal axons to the distal axon stumps. The objective of this study was to show the efficacy of PEG fusion in humans and to retrospectively compare PEG fusion to standard nerve repair. Patients with traumatic lacerations involving digital nerves were treated with PEG after standard microsurgical neurorrhaphy. Sensory assessment after injury was performed at 1 week, 2 weeks, 1 month, and 2 months using static two-point discrimination and Semmes-Weinstein monofilament testing. The Medical Research Council Classification (MRCC) for Sensory Recovery Scale was used to evaluate the level of injury. The PEG fusion group was compared to patient-matched controls whose data were retrospectively collected. Four PEG fusions were performed on four nerve transections in two patients. Polyethylene glycol therapy improves functional outcomes and speed of nerve recovery in clinical setting assessed by average MRCC score in week 1 (2.8 vs 1.0, p = 0.03). At 4 weeks, MRCC remained superior in the PEG fusion group (3.8 vs 1.3, p = 0.01). At 8 weeks, there was improvement in both groups with the PEG fusion cohort remaining statistically better (4.0 vs 1.7, p = 0.01). Polyethylene glycol fusion is a novel therapy for peripheral nerve repair with proven effectiveness in animal models. Clinical studies are still in early stages but have had encouraging results. Polyethylene glycol fusion is a potential revolutionary therapy in peripheral nerve repair but needs further investigation. Therapeutic study, level IV.

A novel technique using hydrophilic polymers to promote axonal fusion.

The management of traumatic peripheral nerve injury remains a considerable concern for clinicians. With minimal innovations in surgical technique and a limited number of specialists trained to treat peripheral nerve injury, outcomes of surgical intervention have been unpredictable. The inability to manipulate the pathophysiology of nerve injury (i.e., Wallerian degeneration) has left scientists and clinicians depending on the slow and lengthy process of axonal regeneration (~1 mm/day). When axons are severed, the endings undergo calcium-mediated plasmalemmal sealing, which limits the ability of the axon to be primarily repaired. Polythethylene glycol (PEG) in combination with a bioengineered process overcomes the inability to fuse axons. The mechanism for PEG axonal fusion is not clearly understood, but multiple studies have shown that a providing a calcium-free environment is essential to the process known as PEG fusion. The proposed mechanism is PEG-induced lipid bilayer fusion by removing the hydration barrier surrounding the axolemma and reducing the activation energy required for membrane fusion to occur. This review highlights PEG fusion, its past and current studies, and future directions in PEG fusion.

Molecular characterisation of axonal fusion, a highly efficient means of nervous system repair

Molecular characterisation of axonal fusion, a highly efficient means of nervous system repair