A functional tacrolimus-releasing nerve wrap for enhancing nerve regeneration following surgical nerve repair.

JOURNAL/nrgr/04.03/01300535-202501000-00036/figure1/v/2024-05-14T021156Z/r/image-tiff Axonal regeneration following surgical nerve repair is slow and often incomplete, resulting in poor functional recovery which sometimes contributes to lifelong disability. Currently, there are no FDA-approved therapies available to promote nerve regeneration. Tacrolimus accelerates axonal regeneration, but systemic side effects presently outweigh its potential benefits for peripheral nerve surgery. The authors describe herein a biodegradable polyurethane-based drug delivery system for the sustained local release of tacrolimus at the nerve repair site, with suitable properties for scalable production and clinical application, aiming to promote nerve regeneration and functional recovery with minimal systemic drug exposure. Tacrolimus is encapsulated into co-axially electrospun polycarbonate-urethane nanofibers to generate an implantable nerve wrap that releases therapeutic doses of bioactive tacrolimus over 31 days. Size and drug loading are adjustable for applications in small and large caliber nerves, and the wrap degrades within 120 days into biocompatible byproducts. Tacrolimus released from the nerve wrap promotes axon elongation in vitro and accelerates nerve regeneration and functional recovery in preclinical nerve repair models while off-target systemic drug exposure is reduced by 80% compared with systemic delivery. Given its surgical suitability and preclinical efficacy and safety, this system may provide a readily translatable approach to support axonal regeneration and recovery in patients undergoing nerve surgery.

Physiology of Nerve Regeneration: Key Factors Affecting Clinical Outcomes.

Functional recovery after peripheral nerve injuries is disappointing despite surgical advances in nerve repair. This review summarizes the relatively short window of opportunity for successful nerve regeneration due to the decline in the expression of growth-associated genes and in turn, the decline in regenerative capacity of the injured neurons and the support provided by the denervated Schwann cells, and the atrophy of denervated muscles. Brief, low-frequency electrical stimulation and post-injury exercise regimes ameliorate these deficits in animal models and patients, but the misdirection of regenerating nerve fibers compromises functional recovery and remains an important area of future research.

Appropriate dosage, timing, and site of intramuscular injections of brain-derived neurotrophic factor (BDNF) promote motor recovery after facial nerve injury in rats.

INTRODUCTION/AIMS: Daily intramuscular injections of fibroblast growth factor 2 (FGF2) but not of brain-derived neurotrophic factor (BDNF) significantly improve whisking behavior and mono-innervation of the rat levator labii superioris (LLS) muscle 56 days after buccal nerve transection and suture (buccal-buccal anastomosis, BBA). We explored the dose-response of BDNF, FGF2, and insulin growth factor 2 (IGF2) on the same parameters, asking whether higher doses of BDNF would promote recovery. METHODS: After BBA, growth factors were injected (30 µL volume) daily into the LLS muscle over 14, 28, or 56 days. At 56 days, video-based motion analysis of vibrissal whisking was performed and the extent of mono- and poly-reinnervation of the reinnervated neuromuscular junctions (NMJs) of the muscle determined with immunostaining of the nerve with ß-tubulin and histochemical staining of the endplates with Alexa Fluor 488-conjugated α-bungarotoxin. RESULTS: The dose-response curve demonstrated significantly higher whisking amplitudes and corresponding increased mono-innervation of the NMJ in the reinnervated LLS muscle at concentrations of 20-30 µg/mL BDNF administered daily for 14-28 days after BBA surgery. In contrast, high doses of IGF2 and FGF2, or doses of 20 and 40 µg/mL of BDNF administered for 14-56 days had no effect on either whisking behavior or in reducing poly-reinnervation of endplates in the muscle. DISCUSSION: These data suggest that the re-establishment of mono-innervation of whiskerpad muscles and the improved motor function by injections of BDNF into the paralyzed vibrissal musculature after facial nerve injury have translation potential and promote clinical application.

Brief Electrical Stimulation Promotes Recovery after Surgical Repair of Injured Peripheral Nerves.

Injured peripheral nerves regenerate their axons in contrast to those in the central nervous system. Yet, functional recovery after surgical repair is often disappointing. The basis for poor recovery is progressive deterioration with time and distance of the growth capacity of the neurons that lose their contact with targets (chronic axotomy) and the growth support of the chronically denervated Schwann cells (SC) in the distal nerve stumps. Nonetheless, chronically denervated atrophic muscle retains the capacity for reinnervation. Declining electrical activity of motoneurons accompanies the progressive fall in axotomized neuronal and denervated SC expression of regeneration-associated-genes and declining regenerative success. Reduced motoneuronal activity is due to the withdrawal of synaptic contacts from the soma. Exogenous neurotrophic factors that promote nerve regeneration can replace the endogenous factors whose expression declines with time. But the profuse axonal outgrowth they provoke and the difficulties in their delivery hinder their efficacy. Brief (1 h) low-frequency (20 Hz) electrical stimulation (ES) proximal to the injury site promotes the expression of endogenous growth factors and, in turn, dramatically accelerates axon outgrowth and target reinnervation. The latter ES effect has been demonstrated in both rats and humans. A conditioning ES of intact nerve days prior to nerve injury increases axonal outgrowth and regeneration rate. Thereby, this form of ES is amenable for nerve transfer surgeries and end-to-side neurorrhaphies. However, additional surgery for applying the required electrodes may be a hurdle. ES is applicable in all surgeries with excellent outcomes.

The Role of Sensory Innervation in Homeostatic and Injury-Induced Corneal Epithelial Renewal.

The cornea is the window through which we see the world. Corneal clarity is required for vision, and blindness occurs when the cornea becomes opaque. The cornea is covered by unique transparent epithelial cells that serve as an outermost cellular barrier bordering between the cornea and the external environment. Corneal sensory nerves protect the cornea from injury by triggering tearing and blink reflexes, and are also thought to regulate corneal epithelial renewal via unknown mechanism(s). When protective corneal sensory innervation is absent due to infection, trauma, intracranial tumors, surgery, or congenital causes, permanent blindness results from repetitive epithelial microtraumas and failure to heal. The condition is termed neurotrophic keratopathy (NK), with an incidence of 5:10,000 people worldwide. In this report, we review the currently available therapeutic solutions for NK and discuss the progress in our understanding of how the sensory nerves induce corneal epithelial renewal.

Advancing Nerve Regeneration: Translational Perspectives of Tacrolimus (FK506).

Peripheral nerve injuries have far-reaching implications for individuals and society, leading to functional impairments, prolonged rehabilitation, and substantial socioeconomic burdens. Tacrolimus, a potent immunosuppressive drug known for its neuroregenerative properties, has emerged in experimental studies as a promising candidate to accelerate nerve fiber regeneration. This review investigates the therapeutic potential of tacrolimus by exploring the postulated mechanisms of action in relation to biological barriers to nerve injury recovery. By mapping both the preclinical and clinical evidence, the benefits and drawbacks of systemic tacrolimus administration and novel delivery systems for localized tacrolimus delivery after nerve injury are elucidated. Through synthesizing the current evidence, identifying practical barriers for clinical translation, and discussing potential strategies to overcome the translational gap, this review provides insights into the translational perspectives of tacrolimus as an adjunct therapy for nerve regeneration.

Schwann Cells Are Key Regulators of Corneal Epithelial Renewal.

Purpose: Corneal sensory nerves protect the cornea from injury. They are also thought to stimulate limbal stem cells (LSCs) to produce transparent epithelial cells constantly, enabling vision. In other organs, Schwann cells (SCs) associated with tissue-innervating axon terminals mediate tissue regeneration. This study defines the critical role of the corneal axon-ensheathing SCs in homeostatic and regenerative corneal epithelial cell renewal. Methods: SC localization in the cornea was determined by in situ hybridization and immunohistochemistry with SC markers. In vivo SC visualization and/or ablation were performed in mice with inducible corneal SC-specific expression of tdTomato and/or Diphtheria toxin, respectively. The relative locations of SCs and LSCs were observed with immunohistochemical analysis of harvested genetically SC-prelabeled mouse corneas with LSC-specific antibodies. The correlation between cornea-innervating axons and the appearance of SCs was ascertained using corneal denervation in rats. To determine the limbal niche cellular composition and gene expression changes associated with innervation-dependent epithelial renewal, single-cell RNA sequencing (scRNA-seq) of dissociated healthy, de-epithelized, and denervated cornea limbi was performed. Results: We observed limbal enrichment of corneal axon-associated myelinating and non-myelinating SCs. Induced local genetic ablation of SCs, although leaving corneal sensory innervation intact, markedly inhibited corneal epithelial renewal. scRNA-seq analysis (1) highlighted the transcriptional heterogenicity of cells populating the limbal niche, and (2) identified transcriptional changes associated with corneal innervation and during wound healing that model potential regulatory paracrine interactions between SCs and LSCs. Conclusions: Limbal SCs are required for innervation-dependent corneal epithelial renewal.

Peripheral Nerve Matrix Hydrogel Promotes Recovery after Nerve Transection and Repair.

BACKGROUND: Nerve transection is the most common form of peripheral nerve injury. Treatment of peripheral nerve injury has primarily focused on stabilization and mechanical cues to guide extension of the regenerating growth cone across the site of transection. The authors investigated the effects of a peripheral nerve matrix (PNM) hydrogel on recovery after nerve transection. METHODS: The authors used rodent models to determine the effect of PNM on axon extension, electrophysiologic nerve conduction, force generation, and neuromuscular junction formation after nerve transection and repair. The authors complemented this work with in vivo and in vitro fluorescence-activated cell sorting and immunohistochemistry approaches to determine the effects of PNM on critical cell populations early after repair. RESULTS: Extension of axons from the proximal stump and overall green fluorescent protein-positive axon volume within the regenerative bridge were increased in the presence of PNM compared with an empty conduit ( P < 0.005) 21 days after repair. PNM increased electrophysiologic conduction (compound muscle action potential amplitude) across the repair site ( P < 0.05) and neuromuscular junction formation ( P = 0.04) 56 days after repair. PNM produced a shift in macrophage phenotype in vitro and in vivo ( P < 0.05) and promoted regeneration in a murine model used to characterize the early immune response to PNM ( P < 0.05). CONCLUSION: PNM, delivered by subepineural injection, promoted recovery after nerve transection with immediate repair, supporting a beneficial macrophage response, axon extension, and downstream remodeling using a range of clinically relevant outcome measures. CLINICAL RELEVANCE STATEMENT: This article describes an approach for subepineural injection at the site of nerve coaptation to modulate the response to injury and improve outcomes.

Delay modulates the immune response to nerve repair.

Effective regeneration after peripheral nerve injury requires macrophage recruitment. We investigated the activation of remodeling pathways within the macrophage population when repair is delayed and identified alteration of key upstream regulators of the inflammatory response. We then targeted one of these regulators, using exogenous IL10 to manipulate the response to injury at the repair site. We demonstrate that this approach alters macrophage polarization, promotes macrophage recruitment, axon extension, neuromuscular junction formation, and increases the number of regenerating motor units reaching their target. We also demonstrate that this approach can rescue the effects of delayed nerve graft.

Electrical stimulation to promote muscle and motor unit force and endurance after spinal cord injury.

Fatigue is a common feature of paralysed skeletal muscle, hindering performance when subjected to functional electrical stimulation (ES) for movement. We asked whether (1) 20 Hz ES for 5% of each day (2.5 s on and 2.5 s off for 3 h) increases tibialis anterior and medial gastrocnemius muscle and motor unit (MU) endurance after paralysis by hemisection and deafferentation (HSDA), and (2) muscle length or loading affects their isometric contractile properties. The daily 5% ES increased muscle endurance, largely independent of muscle length or loading, but to a lesser extent than the daily 50% ES (2.5 s on and 2.5 s off for 24 h). The former was effective in counteracting the decline and slowing of muscle force promoted by the 50% ES. The altered muscle properties were confirmed at the MU level in final experiments once the properties had plateaued. Fast-fatigable MUs were converted to fatigue-intermediate and -resistant MUs that finally comprised ∼80% as compared to ∼10% of the total MU number in the daily 5% ES and the control normal groups, respectively. We conclude that the daily 5% ES regimen counteracts the fatigue of paralysed muscle without compromising contractile force, and thereby, is effective in conditioning muscle for effective movement. KEY POINTS: We asked whether 20 Hz electrical stimulation (ES) for 5% of each day (2.5 s on and 2.5 s off for 3 h; 5% ES) preserves medial gastrocnemius and tibialis anterior muscle and MU isometric contractile forces and increases their endurance after paralysis. Daily 5% ES promoted increased muscle endurance irrespective of the muscle length or loading but to a lesser extent than daily 50% ES (20 Hz ES 2.5 s on and 2.5 s off for 24 h). 5% ES was effective in counteracting decline and slowing of muscle force that resulted from 50% ES. Motor units (MUs) were converted from fast fatigable to fatigue intermediate and resistant MUs, comprising ∼80% as compared to ∼10% in the control normal groups. We conclude that the 5% ES regimen counteracts the fatigue of paralysed muscle without compromising contractile force, and thereby is effective in conditioning the muscle for effective movement.

Trigeminal Sensory Supply Is Essential for Motor Recovery after Facial Nerve Injury.

Recovery of mimic function after facial nerve transection is poor. The successful regrowth of regenerating motor nerve fibers to reinnervate their targets is compromised by (i) poor axonal navigation and excessive collateral branching, (ii) abnormal exchange of nerve impulses between adjacent regrowing axons, namely axonal crosstalk, and (iii) insufficient synaptic input to the axotomized facial motoneurons. As a result, axotomized motoneurons become hyperexcitable but unable to discharge. We review our findings, which have addressed the poor return of mimic function after facial nerve injuries, by testing the hypothesized detrimental component, and we propose that intensifying the trigeminal sensory input to axotomized and electrophysiologically silent facial motoneurons improves the specificity of the reinnervation of appropriate targets. We compared behavioral, functional, and morphological parameters after single reconstructive surgery of the facial nerve (or its buccal branch) with those obtained after identical facial nerve surgery, but combined with direct or indirect stimulation of the ipsilateral infraorbital nerve. We found that both methods of trigeminal sensory stimulation, i.e., stimulation of the vibrissal hairs and manual stimulation of the whisker pad, were beneficial for the outcome through improvement of the quality of target reinnervation and recovery of vibrissal motor performance.

Enlargement of the Nerve Fibers of Silenced Lumbosacral Motoneurons in Cats.

Whether neuromuscular activity influences the size of motor nerves is controversial. All neuromuscular activity in cat hindlimbs was eliminated by spinal cord isolation (SCI), namely, spinal cord transection above and below the medial gastrocnemius (MG) and soleus (SOL) motoneuron pools and L5-S3 dorsal root transection. MG, SOL and sural (SUR) nerves were removed for size measurements, eight months after SCI surgery and from age-matched control cats. Nerve fiber number, the linear relationship between axon size and myelin thickness, and the bimodal distributions of nerve fiber area and diameter were maintained in all three nerves after SCI. The distributions of myelinated sensory fibers were unchanged in SUR nerves in contrast to the myelinated motor fibers in the MG and SOL nerves that were significantly larger. These findings provide evidence that all lumbar motoneurons survive SCI and that their nerve fibers enlarge. Thus, motor nerve fiber size in addition to the properties of the motoneurons and their muscle fibers is dynamic, responding to neuromuscular activity.

Sustained Release of Tacrolimus From a Topical Drug Delivery System Promotes Corneal Reinnervation.

Purpose: Corneal nerve fibers provide sensation and maintain the epithelial renewal process. Insufficient corneal innervation can cause neurotrophic keratopathy. Here, topically delivered tacrolimus is evaluated for its therapeutic potential to promote corneal reinnervation in rats. Methods: A compartmentalized neuronal cell culture was used to determine the effect of locally delivered tacrolimus on sensory axon regeneration in vitro. The regenerating axons but not the cell bodies were exposed to tacrolimus (50 ng/mL), nerve growth factor (50 ng/mL), or a vehicle control. Axon area and length were measured after 48 hours. Then, a biodegradable nanofiber drug delivery system was fabricated via electrospinning of a tacrolimus-loaded polycarbonate-urethane polymer. Biocompatibility, degradation, drug biodistribution, and therapeutic effectiveness were tested in a rat model of neurotrophic keratopathy induced by stereotactic trigeminal nerve ablation. Results: Sensory neurons whose axons were exposed to tacrolimus regenerated significantly more and longer axons compared to vehicle-treated cultures. Trigeminal nerve ablation in rats reliably induced corneal denervation. Four weeks after denervation, rats that had received tacrolimus topically showed similar limbal innervation but a significantly higher nerve fiber density in the center of the cornea compared to the non-treated control. Topically applied tacrolimus was detectable in the ipsilateral vitreal body, the plasma, and the ipsilateral trigeminal ganglion but not in their contralateral counterparts and vital organs after 4 weeks of topical release. Conclusions: Locally delivered tacrolimus promotes axonal regeneration in vitro and corneal reinnervation in vivo with minimal systemic drug exposure. Translational Relevance: Topically applied tacrolimus may provide a readily translatable approach to promote corneal reinnervation.

Foretinib mitigates cutaneous nerve fiber loss in experimental diabetic neuropathy.

Diabetes is by far, the most common cause of neuropathy, inducing neurodegeneration of terminal sensory nerve fibers associated with loss of sensation, paresthesia, and persistent pain. Foretinib prevents die-back degeneration in cultured sensory and sympathetic neurons by rescuing mitochondrial activity and has been proven safe in prospective clinical trials. Here we aimed at investigating a potential neuroprotective effect of Foretinib in experimental diabetic neuropathy. A mouse model of streptozotocin induced diabetes was used that expresses yellow fluorescent protein (YFP) in peripheral nerve fibers under the thy-1 promoter. Streptozotocin-injected mice developed a stable diabetic state (blood glucose > 270 mg/dl), with a significant reduction of intraepidermal nerve fiber density by 25% at 5 weeks compared to the non-diabetic controls. When diabetic mice were treated with Foretinib, a significantly greater volume of the cutaneous nerve fibers (67.3%) in the plantar skin was preserved compared to vehicle treated (37.8%) and non-treated (44.9%) diabetic mice while proximal nerve fiber morphology was not affected. Our results indicate a neuroprotective effect of Foretinib on cutaneous nerve fibers in experimental diabetic neuropathy. As Foretinib treated mice showed greater weight loss compared to vehicle treated controls, future studies may define more sustainable treatment regimen and thereby may allow patients to take advantage of this neuroprotective drug in chronic neurodegenerative diseases like diabetic neuropathy.

Rapid, automated nerve histomorphometry through open-source artificial intelligence.

We aimed to develop and validate a deep learning model for automated segmentation and histomorphometry of myelinated peripheral nerve fibers from light microscopic images. A convolutional neural network integrated in the AxonDeepSeg framework was trained for automated axon/myelin segmentation using a dataset of light-microscopic cross-sectional images of osmium tetroxide-stained rat nerves including various axonal regeneration stages. In a second dataset, accuracy of automated segmentation was determined against manual axon/myelin labels. Automated morphometry results, including axon diameter, myelin sheath thickness and g-ratio were compared against manual straight-line measurements and morphometrics extracted from manual labels with AxonDeepSeg as a reference standard. The neural network achieved high pixel-wise accuracy for nerve fiber segmentations with a mean (± standard deviation) ground truth overlap of 0.93 (± 0.03) for axons and 0.99 (± 0.01) for myelin sheaths, respectively. Nerve fibers were identified with a sensitivity of 0.99 and a precision of 0.97. For each nerve fiber, the myelin thickness, axon diameter, g-ratio, solidity, eccentricity, orientation, and individual x -and y-coordinates were determined automatically. Compared to manual morphometry, automated histomorphometry showed superior agreement with the reference standard while reducing the analysis time to below 2.5% of the time needed for manual morphometry. This open-source convolutional neural network provides rapid and accurate morphometry of entire peripheral nerve cross-sections. Given its easy applicability, it could contribute to significant time savings in biomedical research while extracting unprecedented amounts of objective morphologic information from large image datasets.

Optical tissue clearing enables rapid, precise and comprehensive assessment of three-dimensional morphology in experimental nerve regeneration research.

Morphological analyses are key outcome assessments for nerve regeneration studies but are historically limited to tissue sections. Novel optical tissue clearing techniques enabling three-dimensional imaging of entire organs at a subcellular resolution have revolutionized morphological studies of the brain. To extend their applicability to experimental nerve repair studies we adapted these techniques to nerves and their motor and sensory targets in rats. The solvent-based protocols rendered harvested peripheral nerves and their target organs transparent within 24 hours while preserving tissue architecture and fluorescence. The optical clearing was compatible with conventional laboratory techniques, including retrograde labeling studies, and computational image segmentation, providing fast and precise cell quantitation. Further, optically cleared organs enabled three-dimensional morphometry at an unprecedented scale including dermatome-wide innervation studies, tracing of intramuscular nerve branches or mapping of neurovascular networks. Given their wide-ranging applicability, rapid processing times, and low costs, tissue clearing techniques are likely to be a key technology for next-generation nerve repair studies. All procedures were approved by the Hospital for Sick Children's Laboratory Animal Services Committee (49871/9) on November 9, 2019.

Peripheral nerves preferentially regenerate in intramuscular endoneurial tubes to reinnervate denervated skeletal muscles.

Schwann cells are essential for peripheral nerve regeneration but, over short distances in acellular nerve grafts, extracellular matrix (ECM) molecules can support growth. The ECM molecules are present also on denervated muscle surfaces where they can support nerve growth. In this study, we addressed the efficacy of the ECM molecules of denervated muscle to support nerve fiber regeneration and muscle reinnervation. In the hindlimb of Sprague-Dawley rats, the proximal stump of the transected posterior tibial nerve, was cross-sutured to the distal nerve stump (NN) of each of three denervated muscles, tibialis anterior, extensor digitorum longus, and soleus, or implanted onto the denervated muscles' surfaces (N-M), proximal or distal to the endplate zone. Recordings of muscle and motor unit (MU) isometric forces and silver/cholinesterase histochemical staining of longitudinal muscle cryosections were used to determine the numbers of reinnervated MUs and the spatial course of regenerating nerve fibers, respectively. MU numbers declined significantly after N-M (>50%) as compared to those after NN. Muscle forces were reduced despite each nerve reinnervating up to three times the normal MU muscle fiber number. Regenerating nerves 'streamed' from the N-M site either proximal or distal to endplate zones toward the denervated intramuscular endoneurial tubes, with reduced numbers reinnervating endplates. We conclude that there is preferential reinnervation through the endoneurial tube and that it is important to drive implanted nerve fibers to enter endoneurial tubes for optimal muscle reinnervation. Schwann cells play the essential role in guiding regenerating nerve fibers to reinnervate denervated muscle fibers.

Local FK506 drug delivery enhances nerve regeneration through fresh, unprocessed peripheral nerve allografts.

OBJECTIVE: Nerve allografts offer many advantages in the reconstruction of peripheral nerve gaps: they retain their native microstructure, contain pro-regenerative Schwann cells, are widely available, and avoid donor site morbidity. Unfortunately, clinical use of nerve allografts is limited by the need for systemic immunosuppression and its adverse effects. To eliminate the toxicity of the systemic immunosuppressant FK506, we developed a local FK506 drug delivery system (DDS) to provide drug release over 28 days. The study objective was to investigate if the local FK506 DDS enhances nerve regeneration in a rodent model of nerve gap defect reconstruction with immunologically-disparate nerve allografts. METHODS: In male Lewis rats, a common peroneal nerve gap defect was reconstructed with either a 20 mm nerve isograft from a donor Lewis rat or a 20 mm fresh, unprocessed nerve allograft from an immunologically incompatible donor ACI rat. After 4 weeks of survival, nerve regeneration was evaluated using retrograde neuronal labelling, quantitative histomorphometry, and serum cytokine profile. RESULTS: Treatment with both systemic FK506 and the local FK506 DDS significantly improved motor and sensory neuronal regeneration, as well as histomorphometric indices including myelinated axon number. Rats with nerve allografts treated with either systemic or local FK506 had significantly reduced serum concentrations of the pro-inflammatory cytokine IL-12 compared to untreated vehicle control rats with nerve allografts. Serum FK506 levels were undetectable in rats with local FK506 DDS. INTERPRETATION: The local FK506 DDS improved motor and sensory nerve regeneration through fresh nerve allografts to a level equal to that of either systemic FK506 or nerve isografting. This treatment may be clinically translatable in peripheral nerve reconstruction or vascularized composite allotransplantation.

Failures of nerve regeneration caused by aging or chronic denervation are rescued by restoring Schwann cell c-Jun.

After nerve injury, myelin and Remak Schwann cells reprogram to repair cells specialized for regeneration. Normally providing strong regenerative support, these cells fail in aging animals, and during chronic denervation that results from slow axon growth. This impairs axonal regeneration and causes significant clinical problems. In mice, we find that repair cells express reduced c-Jun protein as regenerative support provided by these cells declines during aging and chronic denervation. In both cases, genetically restoring Schwann cell c-Jun levels restores regeneration to control levels. We identify potential gene candidates mediating this effect and implicate Shh in the control of Schwann cell c-Jun levels. This establishes that a common mechanism, reduced c-Jun in Schwann cells, regulates success and failure of nerve repair both during aging and chronic denervation. This provides a molecular framework for addressing important clinical problems, suggesting molecular pathways that can be targeted to promote repair in the PNS.