Interleukin-4 protects retinal ganglion cells and promotes axon regeneration.

BACKGROUND: The preservation of retinal ganglion cells (RGCs) and the facilitation of axon regeneration are crucial considerations in the management of various vision-threatening disorders. Therefore, we investigate the efficacy of interleukin-4 (IL-4), a potential therapeutic agent, in promoting neuroprotection and axon regeneration of retinal ganglion cells (RGCs) as identified through whole transcriptome sequencing in an in vitro axon growth model. METHODS: A low concentration of staurosporine (STS) was employed to induce in vitro axon growth. Whole transcriptome sequencing was utilized to identify key target factors involved in the molecular mechanism underlying axon growth. The efficacy of recombinant IL-4 protein on promoting RGC axon growth was validated through in vitro experiments. The protective effect of recombinant IL-4 protein on somas of RGCs was assessed using RBPMS-specific immunofluorescent staining in mouse models with optic nerve crush (ONC) and N-methyl-D-aspartic acid (NMDA) injury. The protective effect on RGC axons was evaluated by anterograde labeling of cholera toxin subunit B (CTB), while the promotion of RGC axon regeneration was assessed through both anterograde labeling of CTB and immunofluorescent staining for growth associated protein-43 (GAP43). RESULTS: Whole-transcriptome sequencing of staurosporine-treated 661 W cells revealed a significant upregulation in intracellular IL-4 transcription levels during the process of axon regeneration. In vitro experiments demonstrated that recombinant IL-4 protein effectively stimulated axon outgrowth. Subsequent immunostaining with RBPMS revealed a significantly higher survival rate of RGCs in the rIL-4 group compared to the vehicle group in both NMDA and ONC injury models. Axonal tracing with CTB confirmed that recombinant IL-4 protein preserved long-distance projection of RGC axons, and there was a notably higher number of surviving axons in the rIL-4 group compared to the vehicle group following NMDA-induced injury. Moreover, intravitreal delivery of recombinant IL-4 protein substantially facilitated RGC axon regeneration after ONC injury. CONCLUSION: The recombinant IL-4 protein exhibits the potential to enhance the survival rate of RGCs, protect RGC axons against NMDA-induced injury, and facilitate axon regeneration following ONC. This study provides an experimental foundation for further investigation and development of therapeutic agents aimed at protecting the optic nerve and promoting axon regeneration.

Disturbed brain energy metabolism in a rodent model of DYT-TOR1A dystonia.

DYT-TOR1A (DYT1) dystonia, characterized by reduced penetrance and suspected environmental triggers, is explored using a "second hit" DYT-TOR1A rat model. We aim to investigate the biological mechanisms driving the conversion into a dystonic phenotype, focusing on the striatum's role in dystonia pathophysiology. Sciatic nerve crush injury was induced in ∆ETorA rats, lacking spontaneous motor abnormalities, and wild-type (wt) rats. Twelve weeks post-injury, unbiased RNA-sequencing was performed on the striatum to identify differentially expressed genes (DEGs) and pathways. Fenofibrate, a PPARα agonist, was introduced to assess its effects on gene expression. 18F-FDG autoradiography explored metabolic alterations in brain networks. Low transcriptomic variability existed between naïve wt and ∆ETorA rats (17 DEGs). Sciatic nerve injury significantly impacted ∆ETorA rats (1009 DEGs) compared to wt rats (216 DEGs). Pathway analyses revealed disruptions in energy metabolism, specifically in fatty acid ß-oxidation and glucose metabolism. Fenofibrate induced gene expression changes in wt rats but failed in ∆ETorA rats. Fenofibrate increased dystonia-like movements in wt rats but reduced them in ∆ETorA rats. 18F-FDG autoradiography indicated modified glucose metabolism in motor and somatosensory cortices and striatum in both ∆ETorA and wt rats post-injury. Our findings highlight perturbed energy metabolism pathways in DYT-TOR1A dystonia, emphasizing compromised PPARα agonist efficacy in the striatum. Furthermore, we identify impaired glucose metabolism in the brain network, suggesting a potential shift in energy substrate utilization in dystonic DYT-TOR1A rats. These results contribute to understanding the pathophysiology and potential therapeutic targets for DYT-TOR1A dystonia.

Enriching new transplantable RGC-like cells from retinal organoids for RGC replacement therapy.

Optic neuropathies, such as glaucoma, are due to progressive retinal ganglion cells (RGCs) degeneration, result in irreversible vision loss. The promising RGCs replacement therapy for restoring vision are impeded by insufficient RGC-like cells sources. The present work was enriched one new type RGC-like cells using two surface markers CD184 and CD171 from human induced pluripotent stem cells (hiPSCs) by FACS sorting firstly. These new kind cells have well proliferation ability and possessed passage tolerance in vitro 2D or 3D spheroids culture, which kept expressing Pax6, Brn3b and ßIII-Tubulin and so on. The transplanted CD184+CD171+ RGC-like cells could survive and integrate into the normal and optic nerve crush (ONC) mice retina, especially they were more inclined to across the optic nerve head and extend to the damaged optic nerve. These data support the feasible application for cell replacement therapy in RGC degenerative diseases, as well as help to develop new commercial cells sorting reagents and establish good manufacturing practice (GMP) grade RGC-like donor cells for further clinical application.

Brain injury drives optic glioma formation through neuron-glia signaling.

Tissue injury and tumorigenesis share many cellular and molecular features, including immune cell (T cells, monocytes) infiltration and inflammatory factor (cytokines, chemokines) elaboration. Their common pathobiology raises the intriguing possibility that brain injury could create a tissue microenvironment permissive for tumor formation. Leveraging several murine models of the Neurofibromatosis type 1 (NF1) cancer predisposition syndrome and two experimental methods of brain injury, we demonstrate that both optic nerve crush and diffuse traumatic brain injury induce optic glioma (OPG) formation in mice harboring Nf1-deficient preneoplastic progenitors. We further elucidate the underlying molecular and cellular mechanisms, whereby glutamate released from damaged neurons stimulates IL-1ß release by oligodendrocytes to induce microglia expression of Ccl5, a growth factor critical for Nf1-OPG formation. Interruption of this cellular circuit using glutamate receptor, IL-1ß or Ccl5 inhibitors abrogates injury-induced glioma progression, thus establishing a causative relationship between injury and tumorigenesis.

Sleep disturbance correlated with severity of neuropathic pain in sciatic nerve crush injury model.

The association between sleep and pain has been investigated widely. However, inconsistent results from animal studies compared with human data show the need for a validated animal model in the sleep-pain association field. Our study aims to validate common neuropathic pain models as a tool for evaluating the sleep-pain association. Electrodes electroencephalogram (EEG) and electromyogram (EMG) were surgically implanted to measure sleep. The von Frey test was used to measure pain sensitivity. Following the baseline data acquisition, two pain-modelling procedures were performed: sciatic nerve crush injury (SCI) and common peroneal nerve ligation (CPL). Post-injury measurements were performed on days 1, 5, 10, and 15 post-surgery. The results presented decreased paw withdrawal thresholds and reduced NREM sleep duration in both models on the first post-surgery day. In the SCI model, NREM sleep duration was negatively correlated with paw withdrawal thresholds (p = 0.0466), but not in the CPL model. Wake alpha and theta EEG powers were also correlated with the pain threshold. The results confirm that the SCI model shows disturbed sleep patterns associated with increased pain sensitivity, suggesting it is a reliable tool for investigating sleep disturbances associated with neuropathic pain.

Identification of circular RNA-Dcaf6 as a therapeutic target for optic nerve crush-induced RGC degeneration.

The death of retinal ganglion cells (RGCs) can cause irreversible injury in visual function. Clarifying the mechanism of RGC degeneration is critical for the development of therapeutic strategies. Circular RNAs (circRNAs) are important regulators in many biological and pathological processes. Herein, we performed circRNA microarrays to identify dysregulated circRNAs following optic nerve crush (ONC). The results showed that 221 circRNAs were differentially expressed between ONC retinas and normal retinas. Notably, the levels of circular RNA-Dcaf6 (cDcaf6) expression in aqueous humor of glaucoma patients were higher than that in cataract patients. cDcaf6 silencing could reduce oxidative stress-induced RGC apoptosis in vitro and alleviate retinal neurodegeneration in vivo as shown by increased neuronal nuclei antigen (NeuN, neuronal bodies) and beta-III-tubulin (TUBB3, neuronal filaments) staining and reduced glial fibrillary acidic protein (GFAP, activated glial cells) and vimentin (activated glial cells) staining. Collectively, this study identifies a promising target for treating retinal neurodegeneration.

Glaucoma Animal Models beyond Chronic IOP Increase.

Glaucoma is a complex and multifactorial disease defined as the loss of retinal ganglion cells (RGCs) and their axons. Besides an elevated intraocular pressure (IOP), other mechanisms play a pivotal role in glaucoma onset and progression. For example, it is known that excitotoxicity, immunological alterations, ischemia, and oxidative stress contribute to the neurodegeneration in glaucoma disease. To study these effects and to discover novel therapeutic approaches, appropriate animal models are needed. In this review, we focus on various glaucoma animal models beyond an elevated IOP. We introduce genetically modified mice, e.g., the optineurin E50K knock-in or the glutamate aspartate transporter (GLAST)-deficient mouse. Excitotoxicity can be mimicked by injecting the glutamate analogue N-methyl-D-aspartate intravitreally, which leads to rapid RGC degeneration. To explore the contribution of the immune system, the experimental autoimmune glaucoma model can serve as a useful tool. Here, immunization with antigens led to glaucoma-like damage. The ischemic mechanism can be mimicked by inducing a high IOP for a certain amount of time in rodents, followed by reperfusion. Thereby, damage to the retina and the optic nerve occurs rapidly after ischemia/reperfusion. Lastly, we discuss the importance of optic nerve crush models as model systems for normal-tension glaucoma. In summary, various glaucoma models beyond IOP increase can be utilized.

Reduced Zn2+ promotes retinal ganglion cells survival and optic nerve regeneration after injury through inhibiting autophagy mediated by ROS/Nrf2.

The molecular mechanism of how reduced mobile zinc (Zn2+) affected retinal ganglion cell (RGC) survival and optic nerve regeneration after optic nerve crush (ONC) injury remains unclear. Here, we used conditionally knocked out ZnT-3 in the amacrine cells (ACs) of mice (CKO) in order to explore the role of reactive oxygen species (ROS), nuclear factor erythroid 2-related factor 2 (NFE2L2, Nrf2) and autophagy in the protection of RGCs and axon regeneration after ONC injury. We found that reduced Zn2+ can promote RGC survival and axonal regeneration by decreasing ROS, activating Nrf2, and inhibiting autophagy. Additionally, autophagy after ONC is regulated by ROS and Nrf2. Visual function in mice after ONC injury was partially recovered through the reduction of Zn2+, achieved by using a Zn2+ specific chelator N,N,N',N'-tetrakis-(2-Pyridylmethyl) ethylenediamine (TPEN) or through CKO mice. Overall, our data reveal the crosstalk between Zn2+, ROS, Nrf2 and autophagy following ONC injury. This study verified that TPEN or knocking out ZnT-3 in ACs is a promising therapeutic option for the treatment of optic nerve damage and elucidated the postsynaptic molecular mechanism of Zn2+-triggered damage to RGCs after ONC injury.

Parvalbumin expression changes with retinal ganglion cell degeneration.

Background: Glaucoma is one of the main causes of irreversible visual field loss and blindness worldwide. Vision loss in this multifactorial neurodegenerative disease results from progressive degeneration of retinal ganglion cells (RGCs) and their axons. Identifying molecular markers that can be measured objectively and quantitatively may provide essential insights into glaucoma diagnosis and enhance pathophysiology understanding. Methods: The chronic, progressive DBA/2J glaucomatous mouse model of glaucoma and C57BL6/J optic nerve crush (ONC) mouse model were used in this study. Changes in PVALB expression with RGC and optic nerve degeneration were assessed via gene expression microarray analysis, quantitative real-time polymerase chain reaction (qRT-PCR), Western blot and immunohistochemistry. Results: Microarray analysis of the retinal gene expression in the DBA/2J mice at different ages showed that the expression of PVALB was downregulated as the mice aged and developed glaucoma with retinal ganglion cell loss. Analysis of qRT-PCR results demonstrated PVALB at the mRNA level was reduced in the retinas and optic nerves of old DBA/2J mice and in those after ONC compared to baseline young DBA2/J mice. PVALB protein expression measured by Western blot was also significantly reduced signal in the retinas and optic nerves of old DBA/2J mice and those eyes with crushed nerves. Immunohistochemical staining results demonstrated that there were fewer PVALB-positive cells in the ganglion cell layer (GCL) of the retina and staining pattern changed in the optic nerve from old DBA/2J mice as well as in mice eyes following ONC. Conclusion: PVALB is abundantly expressed both by RGCs' soma in the retinas and RGCs' axons in the optic nerves of C57BL/6J. Furthermore, the expression level of PVALB decreases with RGC degeneration in the glaucomatous DBA/2J mice and after ONC injury of C57BL6/6J, indicating that PVALB is a reliable RGC molecular marker that can be used to study retinal and optic nerve degeneration.

Activation of multiple Eph receptors on neuronal membranes correlates with the onset of optic neuropathy.

BACKGROUND: Optic neuropathy is a major cause of irreversible blindness, yet the molecular determinants that contribute to neuronal demise have not been fully elucidated. Several studies have identified 'ephrin signaling' as one of the most dysregulated pathways in the early pathophysiology of optic neuropathy with varied etiologies. Developmentally, gradients in ephrin signaling coordinate retinotopic mapping via repulsive modulation of cytoskeletal dynamics in neuronal membranes. Little is known about the role ephrin signaling plays in the post-natal visual system and its correlation with the onset of optic neuropathy. METHODS: Postnatal mouse retinas were collected for mass spectrometry analysis for erythropoietin-producing human hepatocellular (Eph) receptors. Optic nerve crush (ONC) model was employed to induce optic neuropathy, and proteomic changes during the acute phase of neuropathic onset were analyzed. Confocal and super-resolution microscopy determined the cellular localization of activated Eph receptors after ONC injury. Eph receptor inhibitors assessed the neuroprotective effect of ephrin signaling modulation. RESULTS: Mass spectrometry revealed expression of seven Eph receptors (EphA2, A4, A5, B1, B2, B3, and B6) in postnatal mouse retinal tissue. Immunoblotting analysis indicated a significant increase in phosphorylation of these Eph receptors 48 h after ONC. Confocal microscopy demonstrated the presence of both subclasses of Eph receptors within the retina. Stochastic optical reconstruction microscopy (STORM) super-resolution imaging combined with optimal transport colocalization analysis revealed a significant co-localization of activated Eph receptors with injured neuronal cells, compared to uninjured neuronal and/or injured glial cells, 48 h post-ONC. Eph receptor inhibitors displayed notable neuroprotective effects for retinal ganglion cells (RGCs) after six days of ONC injury. CONCLUSIONS: Our findings demonstrate the functional presence of diverse Eph receptors in the postnatal mammalian retina, capable of modulating multiple biological processes. Pan-Eph receptor activation contributes to the onset of neuropathy in optic neuropathies, with preferential activation of Eph receptors on neuronal processes in the inner retina following optic nerve injury. Notably, Eph receptor activation precedes neuronal loss. We observed a neuroprotective effect on RGCs upon inhibiting Eph receptors. Our study highlights the importance of investigating this repulsive pathway in early optic neuropathies and provides a comprehensive characterization of the receptors present in the developed retina of mice, relevant to both homeostasis and disease processes.

Unveiling Neuroprotection and Regeneration Mechanisms in Optic Nerve Injury: Insight from Neural Progenitor Cell Therapy with Focus on Vps35 and Syntaxin12.

Axonal degeneration resulting from optic nerve damage can lead to the progressive death of retinal ganglion cells (RGCs), culminating in irreversible vision loss. We contrasted two methods for inducing optic nerve damage: optic nerve compression (ONCo) and optic nerve crush (ONCr). These were assessed for their respective merits in simulating traumatic optic neuropathies and neurodegeneration. We also administered neural progenitor cells (NPCs) into the subtenon space to validate their potential in mitigating optic nerve damage. Our findings indicate that both ONCo and ONCr successfully induced optic nerve damage, as shown by increases in ischemia and expression of genes linked to neuronal regeneration. Post NPC injection, recovery in the expression of neuronal regeneration-related genes was more pronounced in the ONCo model than in the ONCr model, while inflammation-related gene expression saw a better recovery in ONCr. In addition, the proteomic analysis of R28 cells in hypoxic conditions identified Vps35 and Syntaxin12 genes. Vps35 preserved the mitochondrial function in ONCo, while Syntaxin12 appeared to restrain inflammation via the Wnt/ß-catenin signaling pathway in ONCr. NPCs managed to restore damaged RGCs by elevating neuroprotection factors and controlling inflammation through mitochondrial homeostasis and Wnt/ß-catenin signaling in hypoxia-injured R28 cells and in both animal models. Our results suggest that ischemic injury and crush injury cause optic nerve damage via different mechanisms, which can be effectively simulated using ONCo and ONCr, respectively. Moreover, cell-based therapies such as NPCs may offer promising avenues for treating various optic neuropathies, including ischemic and crush injuries.

Type I Collagen-Adhesive and ROS-Scavenging Nanoreactors Enhanced Retinal Ganglion Cell Survival in an Experimental Optic Nerve Crush Model.

Traumatic optic neuropathy (TON) is a severe condition characterized by retinal ganglion cell (RGC) death, often leading to irreversible vision loss, and the death of RGCs is closely associated with oxidative stress. Unfortunately, effective treatment options for TON are lacking. To address this, catalase (CAT) is encapsulated in a tannic acid (TA)/poly(ethylenimine)-crosslinked hollow nanoreactor (CAT@PTP), which exhibited enhanced anchoring in the retina due to TA-collagen adhesion. The antioxidative activity of both CAT and TA synergistically eliminated reactive oxygen species (ROS) to save RGCs in the retina, thereby treating TON. In vitro experiments demonstrated that the nanoreactors preserve the enzymatic activity of CAT and exhibit high adhesion to type I collagen. The combination of CAT and TA-based nanoreactors enhanced ROS elimination while maintaining high biocompatibility. In an optic nerve crush rat model, CAT@PTP is effectively anchored to the retina via TA-collagen adhesion after a single vitreous injection, and RGCs are significantly preserved without adverse events. CAT@PTP exhibited a protective effect on retinal function. Given the abundance of collagen that exists in ocular tissues, these findings may contribute to the further application of this multifunctional nanoreactor in ocular diseases to improve therapeutic efficacy and reduce adverse effects.

Involvement of Glycogen Synthase Kinase 3ß (GSK3ß) in Formation of Phosphorylated Tau and Death of Retinal Ganglion Cells of Rats Caused by Optic Nerve Crush.

Tauopathy is a neurodegenerative condition associated with oligomeric tau formation through abnormal phosphorylation. We previously showed that tauopathy is involved in death of retinal ganglion cells (RGCs) after optic nerve crush (ONC). It has been proposed that glycogen synthase kinase 3ß (GSK3ß) is involved in the hyperphosphorylation of tau in Alzheimer's disease. To determine the roles of GSK3ß in tauopathy-related death of RGCs, lithium chloride (LiCl), a GSK3ß inhibitor, was injected intravitreally just after ONC. The neuroprotective effects of LiCl were determined by counting Tuj-1-stained RGCs on day 7. Changes of phosphorylated (ser 396) tau in the retina were determined by Simple Western analysis (WES) on day 3. Retinal GSK3ß levels were determined by immunohistochemistry (IHC) and an ELISA. There was a 1.9- and 2.1-fold increase in the levels of phosphorylated tau monomers and dimers on day 3 after ONC. LiCl significantly suppressed the increase in the levels of phosphorylated tau induced by ONC. GSK3ß was mainly present in somas of RGCs, and ELISA showed that retinal levels increased to 2.0-fold on day 7. IHC showed that the GSK3ß expression increased over time and remained in RGCs that were poorly stained by Tuj-1. The GSK3ß and tau expression was colocalized in RGCs. The number of RGCs decreased from 1881 ± 188 (sham control) to 1150 ± 192 cells/mm2 on day 7, and LiCl preserved the levels at 1548 ± 173 cells/mm2. Accordingly, GSK3ß may be a promising target for some optic nerve injuries.

Diversity in homeostatic calcium set points predicts retinal ganglion cell survival following optic nerve injury in vivo.

Retinal ganglion cell (RGC) degeneration drives vision loss in blinding conditions. RGC death is often triggered by axon degeneration in the optic nerve. Here, we study the contributions of dynamic and homeostatic Ca2+ levels to RGC death from axon injury. We find that axonal Ca2+ elevations from optic nerve injury do not propagate over distance or reach RGC somas, and acute and chronic Ca2+ dynamics do not affect RGC survival. Instead, we discover that baseline Ca2+ levels vary widely between RGCs and predict their survival after axon injury, and that lowering these levels reduces RGC survival. Further, we find that well-surviving RGC types have higher baseline Ca2+ levels than poorly surviving types. Finally, we observe considerable variation in the baseline Ca2+ levels of different RGCs of the same type, which are predictive of within-type differences in survival.

Regional Gene Expression in the Retina, Optic Nerve Head, and Optic Nerve of Mice with Optic Nerve Crush and Experimental Glaucoma.

A major risk factor for glaucomatous optic neuropathy is the level of intraocular pressure (IOP), which can lead to retinal ganglion cell axon injury and cell death. The optic nerve has a rostral unmyelinated portion at the optic nerve head followed by a caudal myelinated region. The unmyelinated region is differentially susceptible to IOP-induced damage in rodent models and human glaucoma. While several studies have analyzed gene expression changes in the mouse optic nerve following optic nerve injury, few were designed to consider the regional gene expression differences that exist between these distinct areas. We performed bulk RNA-sequencing on the retina and separately micro-dissected unmyelinated and myelinated optic nerve regions from naïve C57BL/6 mice, mice after optic nerve crush, and mice with microbead-induced experimental glaucoma (total = 36). Gene expression patterns in the naïve unmyelinated optic nerve showed significant enrichment of the Wnt, Hippo, PI3K-Akt, and transforming growth factor ß pathways, as well as extracellular matrix-receptor and cell membrane signaling pathways, compared to the myelinated optic nerve and retina. Gene expression changes induced by both injuries were more extensive in the myelinated optic nerve than the unmyelinated region, and greater after nerve crush than glaucoma. Changes present three and fourteen days after injury largely subsided by six weeks. Gene markers of reactive astrocytes did not consistently differ between injury states. Overall, the transcriptomic phenotype of the mouse unmyelinated optic nerve was significantly different from immediately adjacent tissues, likely dominated by expression in astrocytes, whose junctional complexes are inherently important in responding to IOP elevation.

Osteopontin drives retinal ganglion cell resiliency in glaucomatous optic neuropathy.

Chronic neurodegeneration and acute injuries lead to neuron losses via diverse processes. We compared retinal ganglion cell (RGC) responses between chronic glaucomatous conditions and the acute injury model. Among major RGC subclasses, αRGCs and intrinsically photosensitive RGCs (ipRGCs) preferentially survive glaucomatous conditions, similar to findings in the retina subject to axotomy. Focusing on an αRGC intrinsic factor, Osteopontin (secreted phosphoprotein 1 [Spp1]), we found an ectopic neuronal expression of Osteopontin (Spp1) in other RGCs subject to glaucomatous conditions. This contrasted with the Spp1 downregulation subject to axotomy. αRGC-specific Spp1 elimination led to significant αRGC loss, diminishing their resiliency. Spp1 overexpression led to robust neuroprotection of susceptible RGC subclasses under glaucomatous conditions. In contrast, Spp1 overexpression did not significantly protect RGCs subject to axotomy. Additionally, SPP1 marked adult human RGC subsets with large somata and SPP1 expression in the aqueous humor correlated with glaucoma severity. Our study reveals Spp1's role in mediating neuronal resiliency in glaucoma.

Neuroinflammation and gliosis in the injured and contralateral retinas after unilateral optic nerve crush.

The main purpose of this study is to analyze the effects of unilateral optic nerve crush in the gene expression of pro- and anti-inflammatory mediators, and gliosis markers in injured and contralateral retinas. Retinas from intact, unilaterally optic nerve injured or sham-operated C57BL/6J mice were analyzed 1, 3, 9 and 30 days after the surgery (n = 5/group and time point) and the relative expression of TGF-ß1, IL-1ß, TNF-α, Iba1, AQP4, GFAP, MHCII, and TSPO was analyzed in injured and contralateral using qPCR. The results indicated that compared with intact retinas, sham-operated animals showed an early (day 1) upregulation of IL-1ß, TNF-α and TSPO and a late (day 30) upregulation of TNF-α. In sham-contralateral retinas, TNF-α and TSPO mRNA expression were upregulated and day 30 while GFAP, Iba1, AQP4 and MHCII downregulated at day 9. Compared with sham-operated animals, in retinas affected by optic nerve crush GFAP and TSPO upregulated at day 1 and TNF-α, Iba1, AQP4 and MHCII at day 3. In the crushed-contralateral retinas, TGF-ß1, TNF-α, Iba1 and MHCII were upregulated at day 1. TSPO was upregulated up to day 30 whereas TGF-ß1 and Iba1 downregulated after day 9. In conclusion, both sham surgery and optic nerve crush changed the profile of inflammatory and gliosis markers in the injured and contralateral retinas, changes that were more pronounced for optic nerve crush when compared to sham.

Optic Nerve Crush Injury in Rodents to Study Retinal Ganglion Cell Neuroprotection and Regeneration.

Optic nerve crush injury is a useful model for studying the response of central nervous system neurons (CNS) to injury. A particular focus of this model has been to elucidate therapeutic factors in promoting neuroprotection and axon regeneration after injury. Here we describe a step-by-step protocol in accessing the optic nerve and creating a crush injury. This can be used to create a reproducible model to study the response of retinal ganglion cells (RGC), the main projection neurons of the eye, to injury.

Whole body vibration accelerates the functional recovery of motor nerve components in sciatic nerve-crush injury model rats.

This study aimed to investigate the effect of whole body vibration (WBV) on the sensory and motor nerve components with sciatic nerve injury model rats. Surgery was performed on 21 female Wister rats (6-8 weeks) under intraperitoneal anesthesia. The nerve-crush injuries for the left sciatic nerve were inflicted using a Sugita aneurysm clip. The sciatic nerve model rats were randomly divided into two groups (n=9; control group, n=12; WBV group). The rats in the WBV group walked in the cage with a vibratory stimulus (frequency 50 Hz, 20 min/day, 5 times/wk), while those in the control group walked in the cage without any vibratory stimulus. We used heat stimulation-induced sensory threshold and lumbar magnetic stimulation-induced motor-evoked potentials (MEPs) to measure the sensory and motor nerve components, respectively. Further, morphological measurements, bilateral hind-limb dimension, bilateral gastrocnemius dimension, and weight were evaluated. Consequently, there were no significant differences in the sensory threshold at the injury side between the control and WBV groups. However, at 4 and 6 weeks postoperatively, MEPs latencies in the WBV group were significantly shorter than those in the control group. Furthermore, both sides of the hind-limb dimension at 6 weeks postoperatively, the left side of the gastrocnemius dimension, and both sides of the gastrocnemius weight significantly increased. In conclusion, WBV especially accelerates the functional recovery of motor nerve components in sciatic nerve-crush injury model rats.

Mass spectrometry dataset of LC-MS lipidomics analysis of Xenopus laevis optic nerve.

CNS injuries of the anuran amphibian, Xenopus laevis, are uniquely suited for studying the molecular compositions of neuronal regeneration of retinal ganglion cells (RGC) due to a functional recovery of optic axons disparate to adult mammalian analogues. RGCs and their optic nerve axons undergo irreversible neurodegeneration in glaucoma and associated optic neuropathies, resulting in blindness in mammals. Conversely, Xenopus demonstrates RGC lifetime-spanning regenerative capabilities after optic nerve crush [1], inciting opportunities to compare de novo regeneration and develop efficient pharmaceutical approaches for vision restoration. Studies revealing lipidome alterations during optic nerve regeneration are sparse and could serve as a solid foundation for these underlying molecular changes. We profile the lipid changes in a transgenic line of 1 year old Xenopus laevis Tg(islet2b:gfp) frogs that were either left untreated (naïve) or had a monocular surgery of either a left optic crush injury (crush) or sham surgery (sham). Matching controls of uninjured right optic nerves were also collected (control). Tg(islet2b:gfp) frogs were allowed to recover for 7,12,18, and 27 days post optic nerve crush. Following euthanasia, the optic nerves were collected for lipidomic analysis. A modified Bligh and Dyer method [2] was used for lipid extraction, followed by untargeted mass spectrometry lipid profiling with a Q Exactive Orbitrap Mass Spectrometer coupled with a Vanquish Horizon Binary UHPLC LC-MS system (LC MS-MS). The raw scans were analyzed and quantified with LipidSearch 5.0 and the statistical analysis was conducted through Metaboanalyst 5.0. This data is available at Metabolomics Workbench, study ID [ST002414].