Nerve Crush Research Articles

NFATc4 Knockout Promotes Neuroprotection and Retinal Ganglion Cell Regeneration After Optic Nerve Injury.

Retinal ganglion cells (RGCs), neurons transmitting visual information via the optic nerve, fail to regenerate their axons after injury. The progressive loss of RGC function underlies the pathophysiology of glaucoma and other optic neuropathies, often leading to irreversible blindness. Therefore, there is an urgent need to identify the regulators of RGC survival and the regenerative program. In this study, we investigated the role of the family of transcription factors known as nuclear factor of activated T cells (NFAT), which are expressed in the retina; however, their role in RGC survival after injury is unknown. Using the optic nerve crush (ONC) model, widely employed to study optic neuropathies and central nervous system axon injury, we found that NFATc4 is specifically but transiently up-regulated in response to mechanical injury. In the injured retina, NFATc4 immunolocalized primarily to the ganglionic cell layer. Utilizing NFATc4-/- and NFATc3-/- mice, we demonstrated that NFATc4, but not NFATc3, knockout increased RGC survival, improved retina function, and delayed axonal degeneration. Microarray screening data, along with decreased immunostaining of cleaved caspase-3, revealed that NFATc4 knockout was protective against ONC-induced degeneration by suppressing pro-apoptotic signaling. Finally, we used lentiviral-mediated NFATc4 delivery to the retina of NFATc4-/- mice and reversed the pro-survival effect of NFATc4 knockout, conclusively linking the enhanced survival of injured RGCs to NFATc4-dependent mechanisms. In summary, this study is the first to demonstrate that NFATc4 knockout may confer transient RGC neuroprotection and decelerate axonal degeneration after injury, providing a potent therapeutic strategy for optic neuropathies.

RIP1 inhibition protects retinal ganglion cells in glaucoma models of ocular injury

AbstractReceptor-interacting protein 1 (RIP1, RIPK1) is a critical mediator of multiple signaling pathways that promote inflammatory responses and cell death. The kinase activity of RIP1 contributes to the pathogenesis of a number of inflammatory and neurodegenerative diseases. However, the role of RIP1 in retinopathies remains unclear. This study demonstrates that RIP1 inhibition protects retinal ganglion cells (RGCs) in preclinical glaucoma models. Genetic inactivation of RIP1 improves RGC survival and preserves retinal function in the preclinical glaucoma models of optic nerve crush (ONC) and ischemia–reperfusion injury (IRI). In addition, the involvement of necroptosis in ONC and IRI glaucoma models was examined by utilizing RIP1 kinase-dead (RIP1-KD), RIP3 knockout (RIP3-KO), and MLKL knockout (MLKL-KO) mice. The number of RGCs, retinal thickness, and visual acuity were rescued in RIP1-kinase-dead (RIP1-KD) mice in both models, while wild-type (WT) mice experienced significant retinal thinning, RGC loss, and vision impairment. RIP3-KO and MLKL-KO mice showed moderate protective effects in the IRI model and limited in the ONC model. Furthermore, we confirmed that a glaucoma causative mutation in optineurin, OPTN-E50K, sensitizes cells to RIP1-mediated inflammatory cell death. RIP1 inhibition reduces RGC death and axonal degeneration following IRI in mice expressing OPTN-WT and OPTN-E50K variant mice. We demonstrate that RIP1 inactivation suppressed microglial infiltration in the RGC layer following glaucomatous damage. Finally, this study highlights that human glaucomatous retinas exhibit elevated levels of TNF and RIP3 mRNA and microglia infiltration, thus demonstrating the role of neuroinflammation in glaucoma pathogenesis. Altogether, these data indicate that RIP1 plays an important role in modulating neuroinflammation and that inhibiting RIP1 activity may provide a neuroprotective therapy for glaucoma.

Citrus extraction provides neuroprotective effect in optic nerve crush injury mice through P53 signaling pathway

Optic nerve injury induces the loss of retinal ganglion cells and optic nerve atrophy. Our previous study demonstrated the neuroprotective effects of Citrus in the optic nerve crush (ONC) injury model. To explore the underlying neuroprotective mechanism, we focus on the changes in gene expression profile pre- and post-Citrus treatment in the ONC injury model. The results suggested that the Citrus regulated the chemokine signaling pathway and P53 signaling pathway, etc. Four genes were identified as potential target genes (Melk, Mki67, Ccnb1, and Cenpf). Subsequent qRT-PCR and western blot confirmed that Citrus regulated cell apoptosis and inflammation-related gene expression, and inhibited P53 activation-induced cell death. The present study demonstrated that the neuroprotective effect of Citrus was achieved through the regulation of the P53/BCL-2/BAX pathway and the expression regulation of four critical genes, which provide novel therapeutic targets for the therapy of optic nerve injury.

IL-33 protects retinal structure and function via mTOR/S6 signaling pathway in optic nerve crush

This study demonstrated the functions and molecular mechanisms of the IL-33/ST2 axis in experimental optic neuropathy. C57BL/6J mice were used to establish an optic nerve crush (ONC) model. ONC mice were administered with IL-33 intraperitoneal injection, with PBS vehicle as control. Immunofluorescence, quantitative RT-PCR, and western blot techniques were utilized to assess the expression of the IL-33/ST2 axis. The electroretinography (ERG), optical coherence tomography (OCT), H&E, and luxol fast blue were used to assess the structural and functional changes. Western blot was employed to detect the activation of the mTOR/S6 pathway. The IL-33 expression level in the inner nuclear layer of the retina in ONC mice reached its peak on day 3, accompanied by a significant increase in IL-33 receptor ST2 expression. IL-33 treatment promoted the survival of retinal ganglion cells, restored the thickness of inner retina layer (IRL), alleviated the demyelination of the optic nerve, and recovered the decreased amplitude of b-wave in ONC mice. Furthermore, administration of IL-33 activated the mTOR/S6 signaling pathway in RGCs, which was significantly suppressed in the ONC condition. This study indicated that boosting the IL-33/ST2/mTOR/S6 pathway can protect against structural and functional damage to the retina and optic nerve induced by ONC. As a result, the IL-33/ST2 axis holds potential as a therapeutic option for treating various optic neuropathies.

Dnajc3 (HSP40) Modulates Axon Regeneration in the Mouse Optic Nerve.

The present study is designed to identify the genes modulating optic nerve regeneration in the mouse. Using the BXD mouse strains as a genetic mapping panel, we examined differential responses to axon regeneration in order to map genomic loci modulating axonal regeneration. To study regeneration in the optic nerve, Pten was knocked down in the retinal ganglion cells using adeno-associated virus (AAV) delivery of an shRNA, followed by the induction of a mild inflammatory response by an intravitreal injection of Zymosan with CPT-cAMP. The axons of the retinal ganglion cells were damaged by optic nerve crush (ONC). Following a 12-day survival period, regenerating axons were labeled by Cholera Toxin B. Two days later, the regenerating axons within the optic nerve were examined to determine the number of regenerating axons and the distance traveled down the optic nerve. An integral genomic map was made using the regenerative response. Candidate genes were tested by knocking down expression using shRNA or by overexpressing the gene in AAV vectors. The analysis revealed a considerable amount of differential axonal regeneration across all 33 BXD strains, demonstrated by the number of axons regenerating and the length of the regenerating axons. Some strains (BXD99, BXD90, and BXD29) demonstrated significant axonal regeneration; while other strains (BXD13, BXD18, and BXD34) had very little axon regrowth. Within the regenerative data, there was a 4-fold increase in distance regenerated and a 7.5-fold difference in the number of regenerating axons. These data were used to map a quantitative trait locus modulating axonal regeneration to Chromosome 14 (115 to 119 Mb). Within this locus were 16 annotated genes. Subsequent testing revealed that one candidate gene, Dnajc3 , modulates axonal regeneration. Knocking down of Dnajc3 led to a decreased regeneration response in the high regenerative strains (BXD90), while overexpression of Dnajc3 resulted in an increased regeneration response in C57BL/6J and a low regenerative strain (BXD34). In this study, Dnajc3 (encodes Heat Shock Protein 40, HSP40, a molecular chaperone) was identified as a modulator of axon regeneration in mice. This is the first report defining the role of Dnajc3 (HSP40) in axon regeneration.

Differential evaluation of neuromuscular injuries to understand re-innervation at the neuromuscular junction

Peripheral nerve-crush injury is a well-established model of neuromuscular junction (NMJ) denervation and subsequent re-innervation. Functionally, the skeletal muscle follows a similar pattern as neural recovery, with immediate loss of force production that steadily improves in parallel with rates of re-innervation. On the other hand, traumatic injury to the muscle itself, specifically volumetric muscle loss (VML), results in an irrecoverable loss of muscle function. Recent work has indicated significant impairments to the NMJ following this injury that appear chronic in nature, alongside the lack of functional recovery. Thus, the goal of this study was to compare the effects of nerve and muscle injury on NMJ remodeling. Even numbers of adult male and female mice were used with three experimental groups: injury Naïve, nerve crush, and VML injury; and three terminal timepoints: 3-, 48-, and 112-days post-injury. Confirming the assumed recoverability of the two injury models, we found in vivo maximal torque was fully restored following nerve-crush injury but remained at a significant deficit following VML. Compared to injury Naïve and nerve-crush injury, we found VML results in aberrantly high trophic signaling (e.g., neuregulin-1) and numbers of supporting cells, including terminal Schwann cells and sub-synaptic nuclei. In some cases, sex differences were detected, including higher rates of innervation in females than males. Both nerve crush and VML injury display chronic changes to NMJ morphology, such as increased fragmentation and nerve sprouting, highlighting the potential of VML for modeling NMJ regeneration in adulthood, alongside the established nerve-injury models.

The regenerative effect of exosomes isolated from mouse embryonic fibroblasts in mice created as a sciatic nerve crush injury model.

Exosomes (Exos) are candidates for functional recovery and regeneration following sciatic nerve crushed (SNC) injury due to their composition which can accelerate tissue regeneration. Therefore, mouse embryonic fibroblast-derived exosomes were evaluated for their regenerative capacity in SNC injury. In the study, 40Balb/c males (20 ± 5g) and two pregnant mice (for embryonic fibroblast tissue) were used and crushed injury was induced in the left sciatic nerve with an aneurysm clamp. Sciatic nerve model mice were randomly divided into 5 groups (n = 8; control, n = 8; sham, n = 8; SNC, n = 8; Mouse embryonic fibroblast exosome (mExo), n = 8; SNC + Mouse embryonic fibroblast exosome (SNC + mExo). Rotarod tests for motor functions and hot plate and von Frey tests for sensory functions were analyzed in the groups. Expression changes of exosome genes (RARRES1, NAGS, HOXA13, and MEIS1) immunohistochemical analysis of these gene proteins, and structural exosome NF-200 and S100 proteins were evaluated by confocal microscopy. Behavioral analyses showed that the damage in SNC was significant between groups on day14 and day28 (P < 0.05). In behavioral analyses, it was determined that motor functions and mechanical sensitivity lost in SNC were regained after mExo treatment. While expression of all genes was detected in MEF-derived exosomes, the high expression was MESI1 and the low expression was HOXA13. NF200, an indicator of axon number and neurofilament density, was found to decrease in SNC (P < 0.001) and increase after treatment, but not significantly. The decreased S100 protein levels in SNC and the increase detected after treatment were not significant. The expression of four mRNAs in mExos indicates that these genes may have an effect on regenerative processes after SNC injury. The regenerative process supported by tissue protein expressions demonstrates the therapeutic potential of mExo treatment.

CXCR2 mediated trafficking of neutrophils and neutrophil extracellular traps are required for myelin clearance after a peripheral nerve injury

Neutrophils are a vital part of the innate immune system. Many of their functions eliminate bacteria & viruses, like neutrophil extracellular traps (NETs), which trap bacteria, enhancing macrophage phagocytosis. It was surprising when it was demonstrated that neutrophils are a part of Wallerian degeneration, a process that is essential for nerve regeneration after a nerve injury. It is not known what signals attract neutrophils into the nerve and how they aid Wallerian degeneration. Neutrophils accumulate in the distal nerve within one day after an injury and are found in the nerve from one to three days. We demonstrate that CXCR2 mediates the trafficking of neutrophils into the distal nerve, and without CXCR2 Wallerian degeneration, as indicated by luxol fast blue staining, was reduced seven days after a sciatic nerve crush or transection injury. NETs were detected in the distal nerve after a sciatic nerve transection. NET formation has been shown to require protein arginine deiminase 4 (PAD4), which citrullinates histone 3. Inhibiting PAD4 reduced NET formation significantly in the distal nerve at two days and myelin clearance at seven days indicating that NETs aid myelin clearance. These results demonstrate another function for NETs other than clearing pathogens. Neutrophils have been detected after injuries to the central nervous system and diseases in humans and animal models. Our results demonstrate neutrophils aid myelin clearance, suggesting a role for their presence in central nervous system injuries and diseases.

Glycyrrhizic acid promote remyelination after peripheral nerve injury by reducing NF-κB activation

BackgroundPeripheral nerve injury (PNI) causes motor and sensory defects, has strong impact on life quality and still has no effective therapy. Glycyrrhizic acid (GA) is one of the most widely used in traditional Chinese prescriptions and as a flavoring additive in the food industry; the aims of the study were to investigate the effects of GA during sciatic nerve regeneration in a mouse model of sciatic nerve crush injury. MethodsWe established peripheral nerve crush model and investigated the effects of GA. We further studied the potential mechanism of action of GA by Western blotting, fluorescence immunohistochemistry, and PCR analysis. ResultsGA improves the sensory and motor functions of crushed nerve by preventing Schwann cell loss, axonal loss and promoting remyelination of sciatic nerve. Affected by GA, the inflammatory response in the distal part of the sciatic nerve was reduced. Finally, the neuroprotective properties of GA may be regulated by the nuclear factor (NF)‐κB pathway. ConclusionsOur data suggest that GA can effectively alleviate PNI, and the mechanism involves mediating inflammatory response by suppressing NF-κB pathway activation. Thus, GA may represent a potential therapeutic intervention for nerve crush injury.

Local glycolysis supports injury-induced axonal regeneration.

Successful axonal regeneration following injury requires the effective allocation of energy. How axons withstand the initial disruption in mitochondrial energy production caused by the injury and subsequently initiate regrowth is poorly understood. Transcriptomic data showed increased expression of glycolytic genes after optic nerve crush in retinal ganglion cells with the co-deletion of Pten and Socs3. Using retinal cultures in a multicompartment microfluidic device, we observed increased regrowth and enhanced mitochondrial trafficking in the axons of Pten and Socs3 co-deleted neurons. While wild-type axons relied on mitochondrial metabolism, after injury, in the absence of Pten and Socs3, energy production was supported by local glycolysis. Specific inhibition of lactate production hindered injury survival and the initiation of regrowth while slowing down glycolysis upstream impaired regrowth initiation, axonal elongation, and energy production. Together, these observations reveal that glycolytic ATP, combined with sustained mitochondrial transport, is essential for injury-induced axonal regrowth, providing new insights into the metabolic underpinnings of axonal regeneration.

FGF21 inhibits ferroptosis caused by mitochondrial damage to promote the repair of peripheral nerve injury.

Ferroptosis is a new type of cell death characterized by lipid peroxidation and iron dependency, representing an emerging disease regulation mechanism. The limited understanding of ferroptosis in peripheral nerve injury (PNI) complicates the management of such injuries. Mitochondrial dysfunction, which contributes to ferroptosis, further exacerbates the challenges of peripheral nerve repair. In this study, we established an in vitro model of Schwann cells model treated with TBHP and an in vivo sciatic nerve crush injury model in rats. These models were used to investigate the effects of fibroblast growth factor 21 (FGF21) on PNI, both in vitro and in vivo, and to explore the potential mechanisms linking injury-induced ferroptosis and mitochondrial dysfunction. Our findings reveal that PNI triggers abnormal accumulation of lipid reactive oxygen species (ROS) and inactivates mitochondrial respiratory chain complex III, leading to mitochondrial dysfunction. This dysfunction catalyzes the oxidation of excessive polyunsaturated fatty acids, resulting in antioxidant imbalance and loss of ferroptosis suppressor protein 1 (FSP1), which drives lipid peroxidation. Additionally, irregular iron metabolism, defective mitophagy, and other factors contribute to the induction of ferroptosis. Importantly, we found that FGF21 attenuates the abnormal accumulation of lipid ROS, restores mitochondrial function, and suppresses ferroptosis, thus promoting PNI repair. Notably, glutathione peroxidase 4 (GPX4), a downstream target of nuclear factor E2-related factor 2 (Nrf2), and the ERK/Nrf2 pathway are involved in the regulation of ferroptosis by FGF21. FGF21 promotes peripheral nerve repair by inhibiting ferroptosis caused by mitochondrial dysfunction. Therefore, targeting mitochondria and ferroptosis represents a promising therapeutic strategy for effective PNI repair.

DNL343 is an investigational CNS penetrant eukaryotic initiation factor 2B activator that prevents and reverses the effects of neurodegeneration caused by the integrated stress response.

The integrated stress response (ISR) is a conserved pathway in eukaryotic cells that is activated in response to multiple sources of cellular stress. Although acute activation of this pathway restores cellular homeostasis, intense or prolonged ISR activation perturbs cell function and may contribute to neurodegeneration. DNL343 is an investigational CNS-penetrant small-molecule ISR inhibitor designed to activate the eukaryotic initiation factor 2B (eIF2B) and suppress aberrant ISR activation. DNL343 reduced CNS ISR activity and neurodegeneration in a dose-dependent manner in two established in vivo models - the optic nerve crush injury and an eIF2B loss of function (LOF) mutant - demonstrating neuroprotection in both and preventing motor dysfunction in the LOF mutant mouse. Treatment with DNL343 at a late stage of disease in the LOF model reversed elevation in plasma biomarkers of neuroinflammation and neurodegeneration and prevented premature mortality. Several proteins and metabolites that are dysregulated in the LOF mouse brains were normalized by DNL343 treatment, and this response is detectable in human biofluids. Several of these biomarkers show differential levels in CSF and plasma from patients with vanishing white matter disease (VWMD), a neurodegenerative disease that is driven by eIF2B LOF and chronic ISR activation, supporting their potential translational relevance. This study demonstrates that DNL343 is a brain-penetrant ISR inhibitor capable of attenuating neurodegeneration in mouse models and identifies several biomarker candidates that may be used to assess treatment responses in the clinic.

Inhibition of CRMP2 Phosphorylation Suppresses Microglia Activation in the Retina and Optic Nerve and Promotes Optic Nerve Regeneration After Optic Nerve Injury

As the primary connection between the eye and brain, the optic nerve plays a pivotal role in visual information transmission. Injuries to the optic nerve can occur for various reasons, including trauma, glaucoma, and neurodegenerative diseases. Retinal ganglion cells (RGCs), a type of neurons that extend axons through the optic nerve, can rapidly respond to injury and initiate cell death. Additionally, following optic nerve injury microglia, which serve as markers of neuroinflammation, transition from a resting state to an activated state. The phosphorylation of collapsin response mediator protein2 (CRMP2) in the semaphorin 3A (Sema3A) signalling pathway affects several processes, including axon guidance and neuron regeneration. In this study, we used an optic nerve crush (ONC) mouse model to investigate the effects of suppressing CRMP2 phosphorylation on microglia activation. We found that CRMP2 phosphorylation inhibitor suppressed RGCs loss and promoted neuronal regeneration following ONC. In addition, CRMP2 S522A mutant (CRMP2 KI) mice exhibited decreased microglial activation in both the retina and optic nerve following ONC. These results suggest that inhibiting the phosphorylation of CRMP2 can alleviate the loss of RGCs and microglial activation after optic nerve injury, providing insight into the development of treatments for optical neuropathies and neurodegenerative diseases.

The effectiveness of concentrated growth factor in facial nerve crush injury

AimTo evaluate the effect of concentrated growth factor (CGF) on regeneration of facial nerve after crush injury. Materials and MethodsFourteen rats were randomized into two groups. The control group (CG) (n = 7) received a crush injury to the right facial nerve. The CGF group (CGFG) (n = 7) received a crush injury to the right facial nerve and concentrated growth factor prepared from their own blood thereafter. Left facial nerves were used for functional comparison. Nerve function was evaluated using whisker movements and electromyography. Histologic properties were evaluated using hematoxylin and eosin and Masson-trichrome staining, and immunohistochemical properties were evaluated using Neurofilament-H and Anti-Tau degeneration markers. ResultsIn the CGFG, whisker functions began to recover earlier and recovered more quickly compared with the CG. The CG showed significantly prolonged latency and reduced amplitudes in the first week compared with the CGFG (p < 0.05). Recordings of 4th-week latency and amplitudes were similar to the preoperative period in the CGFG (p > 0.05), whereas recordings of the same week were significantly worse in the CG (p < 0.05). Edema and fibrosis were also more pronounced in the CG compared with the CGFG. Neurofilament-H and Anti-Tau were at significantly high levels in the CG (p < 0.05). ConclusionConcentrated growth factor promotes recovery in facial crush injury and may prove a cost-effective, practical, and effective treatment choice in peripheral nerve injury.

Electrical Stimulation: Enhancing Axonal Growth following Peripheral Nerve Injury.

Electrical stimulation has been integrated in recent decades into rehabilitation protocols following neuromuscular injuries. Existing literature supports the utilisation of prolonged or continuous stimulation generated by implantable or transcutaneous devices for chronic pain subsidence and muscle trophism maintenance, which improve outcomes following microsurgical interventions. Newer uses include brief electrical stimulation for peripheral nerve injury. Brief electrical stimulation has shown promise in expediting regeneration of both torn and crushed nerve axons in the murine model and has been incorporated into a limited number of clinical studies. Augmentation of the natural response of an injured peripheral nerve by electrical stimulation has the potential to accelerate regeneration, presumably leading to improved function and clinical outcomes. We review the existing literature on intraoperative utilisation of electrical stimulation to enhance regeneration, such as neural mechanisms of action and their microscopic effect in animal models, as well as results from initial human studies. Level of Evidence: Level V (Therapeutic).

Enhancement of Heme-Oxygenase 1 in the Injured Peripheral Nerve Following Sulforaphane Administration Fosters Regeneration via Proliferation and Maintenance of Repair Schwann Cells.

Nuclear erythroid 2-related factor 2 (Nrf2) and its downstream effector heme oxygenase 1 (HO-1) are commonly activated in response to cellular stresses. The elevated expression of HO-1 has been associated with markedly accelerated peripheral nerve regeneration. This study aimed to evaluate the impact of a naturally occurring dietary Nrf2/HO-1 activator-sulforaphane (SFN)-on regeneration in a murine sciatic nerve crush model. The beneficial safety profile of SFN has been thoroughly investigated and confirmed several times. Here, SFN was administered daily, starting immediately after C57BL/6 mice were subjected to sciatic nerve crush injury. Injured sciatic nerves were excised at various time points post injury for molecular, immunohistochemical and morphometric analyses. Moreover, functional assessment was performed by grip strength analysis and electrophysiology. Following SFN treatment, the early response to injury includes a modulation of autophagic pathways and marked upregulation of Nrf2/HO-1 expression. This enhancement of HO-1 expression was maintained throughout the regeneration phase and accompanied by a significant increase in repair Schwann cells. In these cells, elevated proliferation rates were observed. Significant improvements in grip strength test performance, nerve conduction velocity and remyelination were also noted following SFN treatment. Collectively, SFN modulates cytoprotective and autophagic pathways in the injured nerve, increasing the number of repair Schwann cells and contributing to effective nerve regeneration. Given the availability of SFN as a nutritional supplement, this compound might constitute a novel regenerative approach with broad patient accessibility and further studies on this topic are warranted.

RXR agonist, 9-cis-13,14-dihydroretinoic acid (9CDHRA), reduces damage and protects from demyelination in transsynaptic degeneration model

Neurodegenerative and demyelinating disease, such as multiple sclerosis (MS) are at the forefront of medical research and the discovery of new drugs and therapeutics. One phenomenon of degeneration seen in these diseases is transsynaptic degeneration (TSD), where damage from one axon spreads to the other axons that are connected to it synaptically. It has previously been found that demyelination occurs prior to neuronal loss in an experimental form of induced TSD. Retinoid-x receptor (RXR) agonists have been shown to promote remyelination. Therefore, this study aimed to reveal the effects of a novel endogenous RXR-γ agonist, 9-cis-13,14-dihydroretinoic acid (9CDHRA), on preventing or restoring the effects of TSD. 9CDHRA was administered to mice following optic nerve crush (ONC) procedures, and electrophysiology (visual evoked potential, VEP) and histological (immunofluorescent) assessments were performed. It was found that 9CDHRA treatment effectively delayed glial activation and reduced the presence of apoptosis at the site of injury and further anterogradely in the visual system, including the lateral geniculate nucleus (LGN) and primary visual cortex (V1). Most notably, 9CDHRA was able to maintain myelin levels following ONC, and effectively protected from demyelination. This was corroborated by VEP recordings with improved P1 latency. The promising findings regarding the injury attenuating and myelin protecting properties of 9CDHRA necessitates further investigations into the potential therapeutic uses of this compound.

Comparative Proteomic Study of Retinal Ganglion Cells Undergoing Various Types of Cellular Stressors.

Retinal ganglion cell (RGC) damage serves as a key indicator of various retinal degenerative diseases, including diabetic retinopathy (DR), glaucoma, retinal arterial and retinal vein occlusions, as well as inflammatory and traumatic optic neuropathies. Despite the growing body of data on the RGC proteomics associated with these conditions, there has been no dedicated study conducted to compare the molecular signaling pathways involved in the mechanism of neuronal cell death. Therefore, we launched the study using two different insults leading to RGC death: glutamate excitotoxicity and optic nerve crush (ONC). C57BL/6 mice were used for the study and underwent NMDA- and ONC-induced damages. Twenty-four hours after ONC and 1 hour after NMDA injection, we collected RGCs using CD90.2 coupled magnetic beads, prepared protein extracts, and employed LC-MS for the global proteomic analysis of RGCs. Statistically significant changes in proteins were analyzed using the Shiny Go program to identify GO biological processes and molecular functions resulting from the treatment. We identified unique and common alterations in protein profiles in RGCs undergoing different types of cellular stressors. Additionally, we observed the absence of certain proteins in treated RGCs compared to the control group. Our study not only identified both unique and shared proteomic changes but also laid the groundwork for the future development of a therapeutic platform for testing gene candidates for DR and glaucoma.

Modulating amacrine cell-derived dopamine signaling promotes optic nerve regeneration and preserves visual function.

As part of the central nervous system, the optic nerve, composed of axons from retinal ganglion cells (RGCs), generally fails to regenerate on its own when injured in adult mammals. An innovative approach to promoting optic nerve regeneration involves manipulating the interactions between amacrine cells (ACs) and RGCs. Here, we identified a unique AC subtype, dopaminergic ACs (DACs), that responded early after optic nerve crush by down-regulating neuronal activity and reducing retinal dopamine (DA) release. Activating DACs or augmenting DA release with levodopa demonstrated neuroprotective effects and modestly enhanced axon regeneration. Within this context, we pinpointed the DA receptor D1 (DRD1) as a critical mediator of DAC-derived DA and showed that RGC-specific Drd1 overexpression effectively overcame subtype-specific barriers to regeneration. This strategy markedly boosted RGC survival and axon regeneration after crush and preserved vision in a glaucoma model. This study unveils the crucial role of DAC-derived DA signaling in optic nerve regeneration, holding promise for therapeutic insights into neural repair.

Reversal of injury-associated retinal ganglion cell gene expression by a phosphodiesterase anchoring disruptor peptide

Loss of retinal ganglion cells (RGCs) is central to the pathogenesis of optic neuropathies such as glaucoma. Increased RGC cAMP signaling is neuroprotective. We have shown that displacement of the cAMP-specific phosphodiesterase PDE4D3 from an RGC perinuclear compartment by expression of the modified PDE4D3 N-terminal peptide 4D3(E) increases perinuclear cAMP and protein kinase A activity in cultured neurons and in vivo RGC survival after optic nerve crush (ONC) injury. To explore mechanisms by which PDE4D3 displacement promotes neuroprotection, in this study mice intravitreally injected with an adeno-associated virus to express an mCherry-tagged 4D3(E) peptide were subjected to ONC injury and analyzed by single cell RNA-sequencing (scRNA-seq). 4D3(E)-mCherry expression was associated with an attenuation of injury-induced changes in gene expression, thereby supporting the hypothesis that enhanced perinuclear PKA signaling promotes neuroprotective RGC gene expression.