Nerve Axons Research Articles

Full Recovery after Multiple Treatments with Injectable Ice Slurry.

BackgroundCryoneurolysis uses tissue cooling as an opioid-sparing, long-lasting treatment for peripheral nerve pain. A nerve-selective method for cryoneurolysis by local injection of ice-slurry was developed to allow cryoneurolysis to be performed with a standard needle and syringe, similar to peripheral nerve blocks. Since the treatment of patients with chronic pain may require repeated injections, we investigated the safety and tolerance of repeated treatments in a rat model.MethodsThree repeated ice-slurry treatments, given 6 weeks apart were performed around the rat sciatic nerve. Nerve and surrounding tissues were collected up to 4 months after the third treatment for analysis. Coherent anti-Stokes Raman scattering (CARS) microscopy was used to study effects on myelin sheaths and axon structure. Immunofluorescence (IF) staining was used to study effects on axon density. Hematoxylin and Eosin (H&E) staining was used to examine histologic effects on sciatic nerve and surrounding tissue.ResultsHistologic and CARS image analysis of nerve tissue collected months after three injections demonstrated recovery of nerve structure, myelin organization and axon density to baseline levels, without any residual inflammation, scarring or neuroma formation. No inflammation or scarring was detected in surrounding skin and muscle tissues.ConclusionRepeated ice-slurry injections cause temporary, nerve-selective and reversible changes in the peripheral nerve. There was no histologic damage to surrounding skin and muscle tissues. Repeated treatments with injectable ice-slurry for cryoneurolysis appear to be safe and well tolerated. Clinical studies for patients with chronic pain are warranted.

Abstract P245: Catheter-based Radiofrequency Renal Denervation In Normotensive Pigs Leads To Long-term Pathological Changes In Renal Nerves

Objective: Pre-clinical data of renal nerve regrowth following radiofrequency (RF) renal denervation (RDN) are lacking. In this analysis of RF RDN in normotensive pigs, we assessed the specificity and durability of RF ablation to the perivascular tissues around the renal arteries. Design and Method: RF ablations were delivered to the renal arteries in pigs. To assess specificity, serial tissue samples were harvested from 6 pigs at day 7, and 8 pigs at day 28. Surrounding tissues (ureter, kidney, lymph nodes, skeletal muscle, adrenal glands, and renal nerves) were evaluated with a semi-quantitative scoring system ( Figure ). To assess durability, RF RDN was performed in a separate group of pigs with an additional control group. Serial tissue examination at day 7 (n=6 animals per group) and day 180 (n=8 per group) included quantification of cortical norepinephrine (NE) levels and renal cortical axon density, and evaluation for fibrosis, necrosis, and nerve fiber atrophy. Results: Disruption to non-target perivascular tissues was rare, whereas artery nerve disruption and axonal loss were sustained through 28 days ( Figure ). At 180 days, there was nerve atrophy from axonal destruction around the ablation regions. Necrosis was evident at day 7 but not day 180. Fibrosis was confined to the ablated regions. Renal cortical axon density and cortical NE levels were significantly reduced at 7 and 180 days in the RDN group vs. control. Conclusion: RF RDN specifically disrupted the renal nerves leading to axonal loss. Cortical axon density, NE reduction, and nerve atrophy were sustained through 180 days. These results support the existing clinical evidence of the specificity and durability of RF RDN.

Two Weeks Of Remote Ischemic Preconditioning Improved Cutaneous Microvascular Responses To Post-occlusive Reactive Hyperemia

PURPOSE: Repeated remote ischemic preconditioning (RIPC), induced by intermittent periods of ischemia and reperfusion in a limb body, improves non-nitric oxide (NO)-mediated endothelial-dependent cutaneous vasodilation. Post-occlusive reactive hyperemia (PORH) occurs following the release of a brief arterial occlusion and is principally mediated by non-NO mediated vasodilatory factors such as prostaglandins, sensory nerve axon reflex, and endothelium-derived hyperpolarizing factors (EDHFs). However, the role of repeated bouts of RIPC on the PORH response is unknown. The purpose of this study was to test whether two weeks of RIPC improves PORH elicited cutaneous microvascular hyperemia responses. METHODS: The study was conducted with twenty-one participants (Control; 24 ± 5 yr old, 4 M/4F, n = 8, RIPC; 22 ± 2 yr old, 7 M/6F, n = 13). The RIPC group received RIPC daily sessions over a two-week period (3 intervals of 4 days with RIPC followed by a 1 day break). Each RIPC session consisted of 4 repetitions of 5-minute arm blood flow occlusion interspersed by 5-minute reperfusion. PORH was elicited by brachial artery occlusion for 5 minutes by inflating an arm cuff to 200 mmHg, and the cutaneous hyperemic response was measured after cuff release. Skin blood flow measurements were collected with laser speckle contrast image (LSCI) before and after the repeated RIPC intervention. The control group did not receive RIPC but underwent repeated PORH measurements two weeks later. RESULTS: Both Time to Peak (Tp) and max/time improved after two weeks of RIPC (Tp; 16.5 ± 2.1 vs.15.2 ± 2.9 s, Pre vs. Post, p < 0.05, max/time; 0.11 ± 0.03 vs. 0.13 ± 0.04 CVC/s, Pre vs. Post, p < 0.05). The control group did not change after 2 weeks. CONCLUSIONS: These data suggest that non-NO mediated vasodilator responses in the skin microvasculature may be affected with 2 weeks of repeated RIPC.

YR/DFO@DCNT functionalized anisotropic micro/nano composite topography scaffolds for accelerating long-distance peripheral nerve regeneration

Anisotropic topography could effectively regulate the orientation growth of nerve fibers and promote nerve regeneration process, however, their effect on repairing long-distance peripheral nerve injury is not comparable to autologous grafts and should be further enhanced. In this study, an anisotropic micro/nano composite topography scaffolds containing functionalized multi-walled carbon nanotube(CNT) with YIGSR (YR) and deferoxamine (DFO) was developed by combining use of micro-molding and electrospinning technologies, which is the so-called micro-molded electrospinning (MME). The scaffolds displayed composite topology structure, with randomly-oriented nanofibers and parallel-oriented micro-ridge/grooves, enhanced mechanical properties, and good photo-thermal responsiveness, and could guide the orientation growth of Schwann cells, promote the axon elongation of dorsal root ganglion in vitro. The in vivo evaluation indicated that the developed scaffolds obviously enhanced vascularization, inhibited inflammatory reaction and effectively accelerated the long-distance nerve regeneration with good functional recovery. Further on, the transcriptome sequencing revealed that the YR/DFO@DCNT functionalized anisotropic micro/nano composite scaffolds could upregulate the expression of MET, MPZ and TAZ, which were closely associate with neural cell migration and survival, myelination and cytoskeleton rearrangement for axon elongation and nerve regeneration. Altogether, the anisotropic micro/nano composite topography incorporated with YR/DFO@DCNT by MME may represent a promising strategy for repairing peripheral nerve injury.

Methacrylic acid-based biomaterials promote peripheral innervation in the subcutaneous space of mice

Peripheral nerve innervation is essential for regulating tissue repair and regeneration. MAA-based biomaterials have been previously shown to promote angiogenesis. Here we show a new role for MAA-based biomaterials in promoting terminal axon nerve growth. Our results demonstrate that MAA-based biomaterials promote peripheral nerve growth in an Igf-1 and Shh dependent manner. The resulting nerves increased the sensitivity of treated mice paws to nociception. iDISCO clearing showed that MAA increased the presence of peripheral nerve structures in whole explants. MAA was also able to increase the expression of key neuronal markers and growth factors in a peripheral neuropathy model, the diabetic db/db mouse, suggesting that MAA-based biomaterials may be relevant to treatment of peripheral neuropathy. Moreover, in a peripheral neuropathy model, MAA was able to up-regulate the expression of growth factors for an extended duration suggesting MAA may prevent degeneration through an effect on factors that promote survival. As all tissues are innervated, MAA-based biomaterials could have broad applications in the promoting regeneration and preventing degeneration of peripheral nerves.

3D bioprinted conductive spinal cord biomimetic scaffolds for promoting neuronal differentiation of neural stem cells and repairing of spinal cord injury

Spinal cord injury (SCI) is a terrible injury of the central nervous system for which there is no ideal treatment in clinic so far. The spinal cord biomimetic scaffolds which mimic the shape and structure of native spinal cord tissue, could effectively promote the repair of SCI. However, current biomimetic scaffolds do not mimic the biological functions, especially the electrical conductivity, of spinal cord tissue, which largely limits the therapeutic effect of SCI. In this study, we have developed novel conductive hydrogels based on gelatin methacrylate (GelMA), hyaluronic acid methacrylate (HAMA) and poly(3,4-ethylenedioxythiophene): sulfonated lignin (PEDOT:LS). The incorporation of PEDOT:LS into GelMA/HAMA hydrogel matrix significantly improved the conductivity of the hydrogels. By precisely regulating the light curing time, the conductive hydrogels showed mechanical properties similar to native spinal cord tissues. Then, the conductive biomimetic scaffolds were fabricated by 3D bioprinting, and the neural stem cells (NSCs) encapsulated in the scaffolds exhibited good survival rate (higher than 90%). Compared to the non-conductive scaffolds reported, the conductive scaffolds remarkably promoted neuronal differentiation of NSCs in vitro. In a rat spinal cord complete transection model, the conductive biomimetic scaffold dramatically promoted the recovery of hindlimb motor function. Immunofluorescence staining results further showed that the conductive biomimetic scaffold effectively promoted the regeneration of neurons at the injury site, reduced the deposition of glial scar, and promoted nerve axon regeneration and myelination. Overall, the 3D bioprinting of the conductive spinal cord biomimetic scaffolds developed in this work represents a promising approach for the stem cell-based treatment of SCI.

Pregabalin synchronizes the regeneration of nerve and muscle fibers optimizing the gait recovery of MDX dystrophic mice.

Duchenne muscular dystrophy (DMD) is an X-linked genetic disorder induced by mutations in the dystrophin gene, leading to a degeneration of muscle fibers, triggering retrograde immunomodulatory, and degenerative events in the central nervous system. Thus, neuroprotective drugs such as pregabalin (PGB) can improve motor function by modulating plasticity, together with anti-inflammatory effects. The present work aimed to study the effects of PGB on axonal regeneration after axotomy in dystrophic and non-dystrophic mice. For that, MDX and C57BL/10 mouse strains were subjected to peripheral nerve damage and were treated with PGB (30 mg/kg/day, i.p.) for 28 consecutive days. The treatment was carried out in mice as soon as they completed 5 weeks of life, 1 week before the lesion, corresponding to the peak period of muscle degeneration in the MDX strain. Six-week-old mice were submitted to unilateral sciatic nerve crush and were sacrificed in the 9th week of age. The ipsi and contralateral sciatic nerves were processed for immunohistochemistry and qRT-PCR, evaluating the expression of proteins and gene transcripts related to neuronal and Schwann cell activity. Cranial tibial muscles were dissected for evaluation of neuromuscular junctions using α-bungarotoxin, and the myelinated axons of the sciatic nerve were analyzed by morphometry. The recovery of motor function was monitored throughout the treatment through tests of forced locomotion (rotarod) and spontaneous walking track test (Catwalk system). The results show that treatment with PGB reduced the retrograde cyclic effects of muscle degeneration/regeneration on the nervous system. This fact was confirmed after peripheral nerve injury, showing better adaptation and response of neurons and glia for rapid axonal regeneration, with efficient muscle targeting and regain of function. No side effects of PGB treatment were observed, and the expression of pro-regenerative proteins in neurons and Schwann cells was upregulated. Morphometry of the axons was in line with the preservation of motor endplates, resulting in enhanced performance of dystrophic animals. Overall, the present data indicate that pregabalin is protective and enhances regeneration of the SNP during the development of DMD, improving motor function, which can, in turn, be translated to the clinic.

Mitofusin 2 mutation drives cell proliferation in Charcot-Marie-Tooth 2A fibroblasts.

Dominant mutations in ubiquitously expressed mitofusin 2 gene (MFN2) cause Charcot-Marie-Tooth type 2A (CMT2A; OMIM 609260), an inherited sensory-motor neuropathy that affects peripheral nerve axons. Mitofusin 2 protein has been found to take part in mitochondrial fusion, mitochondria-endoplasmic reticulum tethering, mitochondrial trafficking along axons, mitochondrial quality control and various types of cancer, in which MFN2 has been indicated as a tumor suppressor gene. Discordant data on the mitochondrial altered phenotypes in patient-derived fibroblasts harboring MFN2 mutations and in animal models have been reported. We addressed some of these issues by focusing on mitochondria behavior during autophagy and mitophagy in fibroblasts derived from a CMT2AMFN2 patient with an MFN2650G > T/C217F mutation in the GTPase domain. This study investigated mitochondrial dynamics, respiratory capacity and autophagy/mitophagy, to tackle the multifaceted MFN2 contribution to CMT2A pathogenesis. We found that MFN2 mutated fibroblasts showed impairment of mitochondrial morphology, bioenergetics capacity, and impairment of the early stages of autophagy, but not mitophagy. Unexpectedly, transcriptomic analysis of mutated fibroblasts highlighted marked differentially expressed pathways related to cell population proliferation and extracellular matrix organization. We consistently found the activation of mTORC2/AKT signaling and accelerated proliferation in the CMT2AMFN2 fibroblasts. In conclusion, our evidence indicates that MFN2 mutation can positively drive cell proliferation in CMT2AMFN2 fibroblasts.

Thioredoxin-interacting protein in diabetic retinal neurodegeneration: A novel potential therapeutic target for diabetic retinopathy.

Diabetic retinopathy (DR) is a common complication of diabetes mellitus and has been considered a microvascular disease for a long time. However, recent evidence suggests that diabetic retinal neurodegeneration (DRN), which manifests as neuronal apoptosis, a decrease in optic nerve axons, and reactive gliosis, occurs prior to retinal microvascular alterations. Thioredoxin-interacting protein (TXNIP) is an endogenous inhibitor of thioredoxin (Trx), and it acts by inhibiting its reducing capacity, thereby promoting cellular oxidative stress. In addition, it participates in regulating multiple signaling pathways as a member of the α-arrestin family of proteins. Accumulating evidence suggests that TXNIP is upregulated in diabetes and plays a pivotal role in the pathophysiological process of DR. In this review, we summarized the role of TXNIP in DRN, aiming to provide evidence for DR treatment in the future.

Vitamin C (Ascorbic acid) protects from neuropathy caused by cisplatin, through enhanced heat shock protein-70 and reduced oxidant effect

SUMMARYOBJECTIVE:We aimed to determine whether vitamin C has a protective effect on cisplatin-induced neuropathy in rats.METHODS:In total, 24 rats were included in the study of which 8 rats (no drug administered) were categorized as the control group. The remaining 16 rats were given a total dose of 20 mg/kg cisplatin to induce neuropathy. These drug-administered rats (16 rats) were randomly divided into two groups, namely, group-1 (n=8): cisplatin+saline and group-2 (n=8): cisplatin+vitamin C (500 mg/kg/day). All rats were tested for motor function and electromyographic activity 3 days after cisplatin. Motor performance was evaluated by an inclined-plane test. Compound muscle action potential was evaluated. Plasma malondialdehyde, glutathione, tumor necrosis factor-α, interleukin 6, and sciatic nerve HSP 70 levels were measured. Axon diameter and nerve growth factor expression levels were analyzed.RESULTS:Plasma malondialdehyde, tumor necrosis factor-α, and interleukin 6 levels were higher in the cisplatin+saline group than control group (p<0.001). But vitamin C significantly reduced malondialdehyde and inflammatory cytokine levels when compared with the cisplatin+saline group (p<0.001). Glutathione levels were lower in both cisplatin+saline and cisplatin+vitamin C groups than control group, but vitamin C significantly ameliorated the glutathione levels (p<0.05). Sciatic heat shock protein-70 levels were significantly higher in the cisplatin+vitamin C group than cisplatin+saline group. Compound muscle action potential amplitude and inclined plane test scores were significantly improved in the vitamin C group (p<0.05). Axon diameter and nerve growth factor expression ameliorated with vitamin C (p<0.05).CONCLUSIONS:We demonstrated the ameliorated effects of vitamin C on cisplatin-induced neuropathy through increased heat shock protein-70, nerve growth factor levels, and reduced inflammatory and oxidant effects. The results are promising to improve the neurotoxic effects of cisplatin in cancer patients.

Logistics over Long Distances: Nascent Proteomes in Nerve Axons

Logistics over Long Distances: Nascent Proteomes in Nerve Axons

APP Genetic Deficiency Alters Intracellular Ca2+ Homeostasis and Delays Axonal Degeneration in Dorsal Root Ganglion Sensory Neurons.

The activation of self-destructive cellular programs helps sculpt the nervous system during development, but the molecular mechanisms used are not fully understood. Prior studies have investigated the role of the APP in the developmental degeneration of sensory neurons with contradictory results. In this work, we sought to elucidate the impact of APP deletion in the development of the sensory nervous system in vivo and in vitro. Our in vivo data show an increase in the number of sciatic nerve axons in adult male and female APP-null mice, consistent with the hypothesis that APP plays a pro-degenerative role in the development of peripheral axons. In vitro, we show that genetic deletion of APP delays axonal degeneration triggered by nerve growth factor deprivation, indicating that APP does play a pro-degenerative role. Interestingly, APP depletion does not affect caspase-3 levels but significantly attenuates the rise of axoplasmic Ca2+ that occurs during degeneration. We examined intracellular Ca2+ mechanisms that could be involved and found that APP-null DRG neurons had increased Ca2+ levels within the endoplasmic reticulum and enhanced store-operated Ca2+ entry. We also observed that DRG axons lacking APP have more mitochondria than their WT counterparts, but these display a lower mitochondrial membrane potential. Finally, we present evidence that APP deficiency causes an increase in mitochondrial Ca2+ buffering capacity. Our results support the hypothesis that APP plays a pro-degenerative role in the developmental degeneration of DRG sensory neurons, and unveil the importance of APP in the regulation of calcium signaling in sensory neurons.SIGNIFICANCE STATEMENT The nervous system goes through a phase of pruning and programmed neuronal cell death during development to reach maturity. In such context, the role played by the APP in the peripheral nervous system has been controversial, ranging from pro-survival to pro-degenerative. Here we present evidence in vivo and in vitro supporting the pro-degenerative role of APP, demonstrating the ability of APP to alter intracellular Ca2+ homeostasis and mitochondria, critical players of programmed cell death. This work provides a better understanding of the physiological function of APP and its implication in developmental neuronal death in the nervous system.

Proteomics Reveals the Key Molecules Involved in Curcumin-induced Protection Against Sciatic Nerve Injury in Rats

We generated a rat model of sciatic nerve crush injury and characterized the effects of curcumin on sciatic nerve recovery by using behavioral experiments, hematoxylin-eosin staining, toluidine blue staining, and immunohistochemical. Proteomic analysis using tandem mass tagging was performed to determine differentially expressed proteins (DEPs), and GO and KEGG pathway analyses of overlapping DEPs was conducted, following which, qPCR, western blotting, and immunofluorescence were further performed to validate the proteins of interest. Finally, a Schwann cell injury model was used to verify the effect of curcumin on potential targets. The rat model was successfully established and curcumin improved the sciatic nerve function index of rats with sciatic nerve injury (SNI) and increased the number and diameter of myelinated axons in the sciatic nerve. In the Sham group versus the Injured group and in the Injured group versus the Curcumin group, we identified a total of 4,175 proteins, of which 953 were DEPs, and 218 were known overlapping DEPs. Ten associated pathways, such as calcium signaling pathway, biosynthesis of antibiotics, and long-term potentiation, were identified. The 218 overlapping DEPs were primarily involved in negative regulation of apoptotic process, biological processes, cytoplasm cellular component, and protein binding molecular function based on GO annotation. Curcumin promoted increased expression of ApoD and inhibited the expression of Cyba in vivo and in vitro. These results indicated that curcumin promoted sciatic nerve repair through regulation of various proteins, targets, and pathways. Cyba and ApoD may be potential targets of curcumin in the treatment of SNI.

ADSCs Combined with Melatonin Promote Peripheral Nerve Regeneration through Autophagy.

Background In the early stage of nerve injury, damaged tissue is cleared by autophagy. ADSCs can promote nerve axon regeneration. However, the microenvironment of the injury was changed, and ADSCs are easily apoptotic after transplantation. Mel plays a role in the apoptosis, proliferation, and differentiation of ADSCs. Therefore, we investigated whether Mel combined with ADSCs promoted peripheral nerve regeneration by enhancing early autophagy of injured nerves. Materials and Methods SD rats were randomly split into the control group, model group, Mel group, ADSCs group, ADSCs + Mel group, and 3-MA group. On day 7, autophagy was observed and gait was detected on days 7, 14, 21, and 28. On the 28th day, the sciatic nerve of rats' renewal was detected. Results After 1 w, compare with the model group, the number of autophagosomes and lysosomes and the expressions of protein of LC3-II/LC3-I and Beclin-1 in the ADSCs + Mel group were prominently increased, while the 3-MA group was significantly decreased. After 4 w, the function of the sciatic nerve in ADSCs + Mel was similar to that in the control group. Compared with the model group, the ADSCs + Mel group significantly increased myelin regeneration and the number of motor neurons and reduced gastrocnemius atrophy. Conclusions It was confirmed that ADSCs combined with Mel could promote sciatic nerve regeneration in rats by changing the early autophagy activity of the injured sciatic nerve.

Aging astrocytes metabolically support aging axon function by proficiently regulating astrocyte-neuron lactate shuttle

The astrocyte-neuron lactate shuttle (ANLS) is an essential metabolic support system that uptakes glucose, stores it as glycogen in astrocytes, and provides glycogen-derived lactate for axonal function. Aging intrinsically increases the vulnerability of white matter (WM) to injury. Therefore, we investigated the regulation of this shuttle to understand vascular-glial metabolic coupling to support axonal function during aging in two different WM tracts. Aging astrocytes displayed larger cell bodies and thicker horizontal processes in contrast to thinner vertically oriented processes of young astrocytes. Aging axons recovered less following aglycemia in mouse optic nerves (MONs) compared to young axons, although providing lactate during aglycemia equally supported young and aging axonal function. Incubating MONs in high glucose to upregulate glycogen stores in astrocytes delayed loss of function during aglycemia and improved recovery in both young and aging axons. Providing lactate during recovery from aglycemia unmasked a metabolic switch from glucose to lactate in aging axons. Young and aging corpus callosum consisting of a mixture of myelinated and unmyelinated axons sustained their function fully when lactate was available during aglycemia and surprisingly showed a greater resilience to aglycemia compared to fully myelinated axons of optic nerve. We conclude that lactate is a universal substrate for axons independent of their myelination content and age.

Absence of lenadogene nolparvovec DNA in a brain tumor biopsy from a patient in the REVERSE clinical study, a case report

BackgroundLeber Hereditary Optic Neuropathy (LHON) is a rare, maternally-inherited mitochondrial disease that primarily affects retinal ganglion cells (RGCs) and their axons in the optic nerve, leading to irreversible, bilateral severe vision loss. Lenadogene nolparvovec gene therapy was developed as a treatment for patients with vision loss from LHON caused by the most prevalent m.11778G > A mitochondrial DNA point mutation in the MT-ND4 gene. Lenadogene nolparvovec is a replication-defective recombinant adeno-associated virus vector 2 serotype 2 (AAV2/2), encoding the human wild-type MT-ND4 protein. Lenadogene nolparvovec was administered by intravitreal injection (IVT) in LHON patients harboring the m.11778G > A ND4 mutation in a clinical development program including one phase 1/2 study (REVEAL), three phase 3 pivotal studies (REVERSE, RESCUE, REFLECT), and one long-term follow-up study (RESTORE, the follow-up of REVERSE and RESCUE patients).Case presentationA 67-year-old woman with MT-ND4 LHON, included in the REVERSE clinical study, received a unilateral IVT of lenadogene nolparvovec in the right eye and a sham injection in the left eye in May 2016, 11.4 months and 8.8 months after vision loss in her right and left eyes, respectively. The patient had a normal brain magnetic resonance imaging with contrast at the time of diagnosis of LHON. Two years after treatment administration, BCVA had improved in both eyes. The product was well tolerated with mild and resolutive anterior chamber inflammation in the treated eye. In May 2019, the patient was diagnosed with a right temporal lobe glioblastoma, IDH-wildtype, World Health Organization grade 4, based on histological analysis of a tumor excision. The brain tumor was assessed for the presence of vector DNA by using a sensitive validated qPCR assay targeting the ND4 sequence of the vector.ConclusionND4 DNA was not detected (below 15.625 copies/μg of genomic DNA) in DNA extracted from the brain tumor, while a housekeeping gene DNA was detected at high levels. Taken together, this data shows the absence of detection of lenadogene nolparvovec in a brain tumor (glioblastoma) of a treated patient in the REVERSE clinical trial 3 years after gene therapy administration, supporting the long-term favorable safety of lenadogene nolparvovec.

An Open-source Computational Model of Neurostimulation of the Spinal Pudendo-Vesical Reflex for the Recovery of Bladder Control After Spinal Cord Injury.

Spinal cord stimulation (SCS) could be used to restore control of the bladder after spinal cord injury, but substantial development is still required to tailor this technology for bladder function. Computational models could be utilized to accelerate these efforts enabling in-silico optimization of stimulation parameters. However, no model of the spinal pudendo-vesical reflex can simulate the effect of stimulation amplitude on neuron recruitment. This limitation hinders accurate prediction of bladder pressure changes for different stimulation configurations. Here., we implemented an open-source realistic spiking neural network model of the pudendo-vesical reflex enabling exploration of the impact of stimulation amplitude and frequency on bladder pressure changes. We used the o2S2 PARC platform to design a parallel implementation of the bladder reflex circuits with NEURON. Our model successfully reproduced and expanded previous studies., producing a decrease in bladder pressure at low stimulation frequency (10 Hz) and excitation at high stimulation frequency (≥33 Hz) in isovolumetric experiments. We then explored the effect of mixed nerve recruitment., simulating a common case of poorly selective spinal cord stimulation. We found that high recruitments of pudendal nerve axons are necessary to maintain this bi-modal behavior., regardless of stimulation specificity. Our framework is fully open-source and can be used to simulate any type of axon stimulations such as SCS and peripheral nerve stimulation.

Sensory nerve regeneration and reinnervation in muscle following peripheral nerve injury.

Sensory afferent fibers are an important component of motor nerves and compose the majority of axons in many nerves traditionally thought of as "pure" motor nerves. These sensory afferent fibers innervate special sensory end organs in muscle, including muscle spindles that respond to changes in muscle length and Golgi tendons that detect muscle tension. Both play a major role in proprioception, sensorimotor extremity control feedback, and force regulation. After peripheral nerve injury, there is histological and electrophysiological evidence that sensory afferents can reinnervate muscle, including muscle that was not the nerve's original target. Reinnervation can occur after different nerve injury and muscle models, including muscle graft, crush, and transection injuries, and occurs in a nonspecific manner, allowing for cross-innervation to occur. Evidence of cross-innervation includes the following: muscle spindle and Golgi tendon afferent-receptor mismatch, vagal sensory fiber reinnervation of muscle, and cutaneous afferent reinnervation of muscle spindle or Golgi tendons. There are several notable clinical applications of sensory reinnervation and cross-reinnervation of muscle, including restoration of optimal motor control after peripheral nerve repair, flap sensation, sensory protection of denervated muscle, neuroma treatment and prevention, and facilitation of prosthetic sensorimotor control. This review focuses on sensory nerve regeneration and reinnervation in muscle, and the clinical applications of this phenomena. Understanding the physiology and limitations of sensory nerve regeneration and reinnervation in muscle may ultimately facilitate improvement of its clinical applications.

Neurofilament Transport Is Bidirectional In Vivo.

Neurofilaments are abundant space-filling cytoskeletal polymers that are transported into and along axons. During postnatal development, these polymers accumulate in myelinated axons causing an expansion of axon caliber, which is necessary for rapid electrical transmission. Studies on cultured nerve cells have shown that axonal neurofilaments move rapidly and intermittently along microtubule tracks in both anterograde and retrograde directions. However, it is unclear whether neurofilament transport is also bidirectional in vivo. Here, we describe a pulse-spread fluorescence photoactivation method to address this in peripheral nerves dissected from hThy1-paGFP-NFM transgenic mice, which express a photoactivatable fluorescent neurofilament protein. Neurofilaments were photoactivated in short segments of myelinated axons in tibial nerves at 2, 4, 8, and 16 weeks of age. The proximal and distal spread of the fluorescence due to the movement of the fluorescent neurofilaments was measured over time. We show that the directional bias and velocity of neurofilament transport can be calculated from these measurements. The directional bias was ∼60% anterograde and 40% retrograde and did not change significantly with age or distance along the nerve. The net velocity decreased with age and distance along the nerve, which is consistent with previous studies using radioisotopic pulse labeling. This decrease in velocity was caused by a decrease in both anterograde and retrograde movement. Thus, neurofilament transport is bidirectional in vivo, with a significant fraction of the filaments moving retrogradely in both juvenile and adult mice.

Dual Leucine Zipper Kinase Regulates Dscam Expression through a Noncanonical Function of the Cytoplasmic Poly(A)-Binding Protein.

Dual leucine zipper kinase (DLK) plays a pivotal role in the development, degeneration, and regeneration of neurons. DLK can regulate gene expression post-transcriptionally, but the underlying mechanism remains poorly understood. The Drosophila DLK, Wallenda (Wnd), regulates the expression of Down syndrome cell adhesion molecule (Dscam) to control presynaptic arbor growth. This regulation is mediated by the 3' untranslated region (3'UTR) of Dscam mRNA, which suggests that RNA binding proteins (RBPs) mediate DLK function. We performed a genome-wide cell-based RNAi screen of RBPs and identified the cytoplasmic poly(A)-binding protein, pAbp, as an RBP that mediates Wnd-induced increase in Dscam expression. Genetic analysis shows that Wnd requires pAbp for promoting presynaptic arbor growth and for enhancing Dscam expression. Our analysis revealed that Dscam mRNAs harbor short poly(A) tails. We identified a region in Dscam 3'UTR that specifically interacts with pAbp. Removing this region significantly reduced Wnd-induced increase in Dscam expression. These suggest that a noncanonical interaction of PABP with the 3'UTR of target transcripts is essential for DLK functions.SIGNIFICANCE STATEMENT The kinase DLK plays key roles in a multitude of neuronal responses, including axon development, neurodegeneration, and nerve injury. Previous studies show that DLK acts via mRNAs to regulate protein synthesis, but how DLK does so is poorly understood. This study demonstrates that DLK regulates the synthesis of Dscam through the poly(A)-binding protein PABP-C. Whereas PABP-C is known as a general translational activator, our study shows that DLK-mediated Dscam expression involves a noncanonical interaction between PABP-C and the Dscam mRNA, which leads to a selective regulation of Dscam translation by PABP-C. Thus, our study provides novel insights into the mechanisms that underlie the function of DLK and regulation of gene expression of PABP-C.