Single-cell RNA sequencing (scRNA-seq) has transformed the study of retinal development and diseases by enabling a detailed analysis of cellular diversity within retinal organoids (ROs). ROs generated from pluripotent stem cells mimic the essential characteristics of the human retina and provide a valuable in vitro model for investigating retinal development, cell interactions, and disease mechanisms. This review summarizes the application of scRNA-seq on RO research, emphasizing its capacity to identify distinct cell populations, uncover developmental trajectories, and reveal the molecular signatures of retinal diseases. scRNA-seq provides new insights into retinal neurogenesis, cellular diversity, and the pathophysiology of retinal degenerative diseases. This technology has enabled the identification of novel biomarkers and potential therapeutic targets. Integrating scRNA-seq with other technologies, such as spatial transcriptomics and CRISPR-based screening, can further deepen our understanding of retinal biology and improve treatment strategies.

Glaucoma is an irreversible blinding eye disease characterized by apoptosis of mature neurons-retinal ganglion cells (RGCs), visual field defect and vision loss. Regeneration of RGCs and reconstruction of the neural connections between the retina and the brain is considered an effective strategy to promote visual restoration in patients with glaucoma. However, there are currently no effective methods for regenerating RGCs to restore vision in clinical practice. Microglia are a type of glial cells that regulate the immune response in the retina and central nervous system (CNS), whether they have pluripotency and be reversed into RGCs remains unclear and challenging. This study revealed that the ectopic expression of multiple genes (Brn3b, Sox2, Cbln1, and NP1, referred to as BSCN) in microglia can promote their conversion into RGC-like cells by microglia fate lineage tracing in vivo. The regenerated RGC-like cells project axons to the distant brain and reconstruct the visual neural circuit, restoring the impaired vision in adult mice with acute glaucoma induced by retinal ischemia-reperfusion (I/R) injury. Furthermore, the regenerated RGC-like cells could survive stably for up to one year, and the same regeneration strategy was performed in older mice with acute glaucoma, which confirmed the effectiveness of the BSCN reprogramming to regenerate RGC-like cells. In summary, we have identified the microglia as a new type of reprogramming seed cells, and four key genes were found to be involved in regenerating RGC-like cells to restore vision. These findings highlight a new strategy of RGC-like cell regeneration and provide a theoretical basis for treatment of glaucoma in the future.

Optic neuropathy, characterized by the irreversible degeneration of RGCs, leads to vision loss for which treatment options remained limited. Nanoparticle-based therapeutic strategies have been developed to stimulate RGCs regeneration; however, insufficient nanoparticles delivery to RGCs and a lack of robust stimulation of RGCs intrinsic regenerative capacity, have limited their application. Here, we demonstrate that a RGC-targeting Tet1 conjugated to liposome delivery system (Lipo-T) enhances therapeutic agent delivery to RGCs while minimizing off-target effect for activating glial cells. Targeted delivery of a mTOR-specific activator MHY1485 to RGCs via Lipo-T (MHY@Lipo-T) significantly enhances in vivo RGCs survival, neurite outgrowth, and axonal regeneration without retinal inflammation from unwanted glial activation. By conferring sustained MHY1485 to degenerated retina through Lipo-T injection, and with its high target ability potential, the first use of MHY@Lipo-T expands new therapeutic option for optic nerve injury.

Leber hereditary optic neuropathy (LHON) is a paradigm for mitochondrial retinopathy due to mitochondrial DNA (mtDNA) mutations. However, the mechanism underlying retinal cell-specific effects of LHON-linked mtDNA mutations remains poorly understood, and there has been no effective treatment or cure for this disorder. Using a mouse model bearing an LHON-linked ND6P25L mutation, we demonstrated that the mutation caused retinal cell-specific deficiencies, especially in retinal ganglion cells (RGCs), rods, and Müller cells. Single-cell RNA sequencing revealed cell-specific dysregulation of oxidative phosphorylation and visual signaling pathways in the mutant retina. Strikingly, ND6 mutation-induced dysfunctions caused abnormal vitamin A (VA) metabolism essential for visual function. VA supplementation remarkably alleviated retinal deficiencies, including reduced fundus lesion and retinal thickness and increased numbers of RGCs, photoreceptors, and Müller cell neurites. The restoration of visual functions with VA treatment were further evidenced by correcting dysregulations of phototransduction cascade and neurotransmitter transmission and restoring electrophysiological properties. Interestingly, VA supplementation markedly rescued the abnormal mitochondrial morphologies and functions in the mutant retina. These findings provide insight into retina-specific pathophysiology of mitochondrial retinopathy arising from VA deficiency and mitochondrial dysfunction induced by mtDNA mutation and a step toward therapeutic intervention for LHON and other mitochondrial retinopathies.

Retinal ganglion cells (RGCs) are the only neuronal bridges connecting retinal inputs to the brain's visual processing centers, enabling visual perception. The axon of RGCs forms the optic nerve, which transmits visual information to the visual cortex. Damage to RGCs and their axons results in irreversible visual impairment. Acute retinal damage is commonly induced by conditions such as optic nerve compression, glaucoma, and optic neuritis, for which effective clinical treatments are currently unavailable. Therefore, understanding the response of RGCs and their axons to injury is crucial for the development of potential treatments. This study utilizes multiple models including optic nerve crush (ONC), acute intraocular pressure (IOP) elevation, and local lipopolysaccharide (LPS) injection into the optic nerve to mimic eye diseases. Three days post-surgery, mice underwent retinal isolation followed by bulk-RNA sequencing to analyze differential gene expression among models. Using thresholds of |Log2 fold change (FC)|> 2 and p-value < 0.05, the significant gene expression changes observed in each model were as follows: ONC (upregulated, 456; downregulated, 84), IOP (upregulated, 1946; downregulated, 655), and LPS (upregulated, 219; downregulated, 94). Gene ontology (GO) analysis of the upregulated genes unexpectedly revealed that immune system pathways were the primary shared targets across all three models. In contrast, the downregulated genes exhibited model-specific enrichment: synaptic components and functions in IOP, neurogenesis and neuronal development in ONC, and inflammation and antioxidant in LPS. These findings were further confirmed by Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. This suggests that managing immune activation is essential for treating acute retinal injury, and therapeutic strategies should address model-specific targets as well. Notably, 39 genes intersected across the models, and the protein-protein interaction (PPI) network identified Ccl5 as a key hub gene, underscoring its critical role in the pathophysiology of all three diseases.

Retinal ganglion cells (RGCs) are the visual gateway of the brain, with their axons converging to form the optic nerve, making them the most vulnerable target in diseases such as glaucoma and traumatic optic neuropathy (TON). In both diseases, the disruption of the blood-retinal barrier(BRB) is considered an important mechanism that accelerates RGC degeneration and hinders axon regeneration. The BRB consists of the inner blood-retinal barrier (iBRB) and the outer blood-retinal barrier (oBRB), which are maintained by endothelial cells(ECs), pericytes(PCs), and retinal pigment epithelial (RPE), respectively. Their functions include regulating nutrient exchange, oxidative stress, and the immune microenvironment. However, in glaucoma and TON, the structural and functional integrity of the BRB is severely damaged due to mechanical stress, inflammatory reactions, and metabolic disorders. Emerging evidence highlights that BRB disruption leads to heightened vascular permeability, immune cell infiltration, and sustained chronic inflammation, creating a hostile microenvironment for RGC survival. Furthermore, the dynamic interplay and imbalance among ECs, PCs, and glial cells within the neurovascular unit (NVU) are pivotal drivers of BRB destruction, exacerbating RGC apoptosis and limiting optic nerve regeneration. The intricate molecular and cellular mechanisms underlying these processes underscore the BRB's critical role in glaucoma and TON pathophysiology while offering a compelling foundation for therapeutic strategies targeting BRB repair and stabilization. This review provides crucial insights and lays a robust groundwork for advancing research on neural regeneration and innovative optic nerve protective strategies.

Retinal inflammatory microenvironment caused by microglia over-activation is deemed to be crucial pathological changes that lead to the massive death of retinal ganglion cells (RGCs) after traumatic optic neuropathy (TON), which then results in visual impairment and even blindness. Therefore, exploring effective targets to suppress microglia activation is a promising therapeutic strategy for TON. In the present work, we determined the roles of immune-responsive gene 1 (IRG1)/itaconate pathway on retinal microglia activation and neuroinflammation after TON, through endogenously manipulating Irg1 expression and exogenously supplementing itaconate derivatives, we evaluated its effects on RGCs survival, retinal structural damage and visual function after TON. Finally, we identified the downstream mechanism by which the Irg1/itaconate pathway regulates microglia through transcriptome analysis. We found that specifically overexpression of Irg1 in retinal microglia significantly inhibited microglia activation and alleviated neuroinflammation after TON, thereby promoting RGCs survival and improving visual function. While knockdown of Irg1 caused microglia over-activation and exacerbated neuroinflammation, thus aggravating RGCs damage and deteriorating visual function after TON. Further in vivo and in vitro experiments confirmed that itaconate derivatives significantly inhibited microglia activation and alleviated neuroinflammation, hence alleviated RGCs damage and visual impairment. Finally, transcriptome analysis indicated that complement and coagulation cascades pathway might be the crucial downstream mechanism of the Irg1/itaconate pathway. Our study identifies the Irg1/itaconate pathway as a prospective target for treating TON.

Mechanisms underlying functional axonal rewiring after adult mammalian central nervous system (CNS) injuries remain unclear partially due to limited models. Here we develop a mouse intracranial pre-olivary pretectal nucleus (OPN) optic tract injury model and demonstrate that Pten/Socs3 knockout and CNTF expression in retinal ganglion cells (RGCs) promotes optic tract regeneration and OPN reinnervation. Revealed by transmission electron microscopy, trans-synaptic labeling, and electrophysiology, functional synapses are formed in OPN mainly by intrinsically photosensitive RGCs, thereby partially restoring the pupillary light reflex (PLR). Moreover, combining with Lipin1 knockdown accelerates the recovery and achieves functional reconnection after chronic injury. PLR can be further boosted by increasing RGC photosensitivity with melanopsin overexpression, and it can also be enhanced by treatment of a voltage-gated calcium channel modulator to augment presynaptic release. These findings highlight the importance of neuronal types and presynaptic activity for functional reconnection after CNS injuries.

Retinal ganglion cells (RGCs) are the sole output neurons of the retina and convey visual information to the brain via their axons in the optic nerve. Following injury to the optic nerve, RGC axons degenerate and many cells die. For example, a model of axon injury, the optic nerve crush (ONC), kills ∼80% of RGCs after 2 weeks. Surviving cells are biased towards 'resilient' types, including several with sustained firing to light stimulation. RGC survival may depend on activity, and there is limited understanding of how or why activity changes following optic nerve injury. Here we quantified the electrophysiological properties of a highly resilient RGC type, the sustained ON-Alpha (AlphaONS) RGC, 7 days after ONC with extracellular and whole-cell patch clamp recording. Both light- and current-driven firing were reduced after ONC, but synaptic inputs were largely intact. Resting membrane potential and input resistance were relatively unchanged, while voltage-gated currents were impaired, including a reduction in voltage-gated sodium channel current and channel density in the axon initial segment. Hyperpolarization or chelation of intracellular calcium partially rescued firing rates. Extracellular recordings at 3 days following ONC showed normal light-evoked firing from AlphaONS RGCs and other Alpha RGCs, including susceptible types. These data suggest that an injured resilient RGC reduces its activity by 1 week after injury as a consequence of reduced voltage-gated current and downregulation of intrinsic excitability via a Ca2+-dependent mechanism. Reduced excitability may be due to degradation of the axon but could also be energetically beneficial, preserving energy for survival and regeneration. KEY POINTS: Retinal ganglion cell (RGC) types show diverse rates of survival after axon injury. A resilient RGC type (sustained ON-Alpha RGC) maintains its synaptic inputs 1 week after injury. The resilient RGC type shows diminished firing and reduced expression of axon initial segment genes 1 week after injury Activity deficits reflect dysfunction of intrinsic properties (Na+ channels, intracellular Ca2+), not changes to synaptic input. Both resilient and susceptible Alpha RGC types show intact firing at 3 days after injury, suggesting that activity at this time point does not predict resilience.

Retinal diseases can severely impair vision and even lead to blindness, posing significant threats to both physical and mental health. Physical retinal regenerative therapies are poised to revolutionize the treatment of various disorders associated with blindness. However, these therapies must overcome the challenges posed by the protective inner and outer blood‒retinal barriers. Nanotechnology applications in ophthalmology have shown great potential in addressing the issue of drug delivery to the eye. Moreover, nanotechnology-based therapeutics can have profound clinical impacts on retinopathy, particularly retinal regeneration, thereby improving patient outcomes. Continuous advancements in nanotechnology are being applied to regenerate lost or damaged eye tissues and to treat vision loss and blindness caused by various retinal degenerative diseases. These approaches can be categorized into three main strategies: i) nanoparticles for delivering drugs, genes, and other essential substances; ii) nanoscaffolds for providing biocompatible support; and iii) nanocomposites for enhancing the functionality of primary or stem cells. The aim of this comprehensive review is to present the current understanding of nanotechnology-based therapeutics for retinal regeneration, with a focus on the perspective functions of nanomaterials.

Failure of central nervous system (CNS) axons to regenerate after injury results in permanent disability. Several molecular neuro-protective and neuro-regenerative strategies have been proposed as potential treatments but do not provide the directional cues needed to direct target-specific axon regeneration. Here, we demonstrate that applying an external guidance cue in the form of electric field stimulation to adult rats after optic nerve crush injury was effective at directing long-distance, target-specific retinal ganglion cell (RGC) axon regeneration to native targets in the diencephalon. Stimulation was performed with asymmetric charged-balanced (ACB) waveforms that are safer than direct current and more effective than traditional, symmetric biphasic waveforms. In addition to partial anatomical restoration, ACB waveforms conferred partial restoration of visual function as measured by pattern electroretinogram recordings and local field potential recordings in the superior colliculus-and did so without the need for genetic manipulation. Our work suggests that exogenous electric field application can override cell-intrinsic and cell-extrinsic barriers to axon regeneration, and that electrical stimulation performed with specific ACB waveforms may be an effective strategy for directing anatomical and functional restoration after CNS injury.

Currently, most inherited retinal diseases lack curative interventions, and available treatment modalities are constrained to symptomatic approaches. Retinal organoid technology has emerged as a method for treating inherited retinal diseases, with growing academic interest in recent years. The purpose of this review was to systematically organize the current protocols for generating retinal organoids using induced pluripotent stem cells from patients with inherited retinal disease and to investigate the application of retinal organoids in inherited retinal disease research.

Data were collected from the PubMed, Scopus, and Web of Science databases using a keyword search. The main search term used was "retinal organoid," accompanied by secondary keywords such as "optic cup," "three-dimensional," and "self-organizing." The final search was conducted on October 2, 2024.

Of the 2,129 studies retrieved, 130 were included in the qualitative synthesis. The protocols for the generation of retinal organoids in inherited retinal disease research use five major approaches, categorized into 3D and a combination of 2D/3D approaches, implemented with modifications. Disease phenotypes have been successfully reproduced via the generation of retinal organoids from the induced pluripotent stem cells of individuals with inherited retinal diseases, facilitating the progression of research into novel therapeutic developments. Cells have been obtained from retinal organoids for cell therapy, and progress toward their potential integration into clinical practice is underway. Considering their potential applications, retinal organoid technology has shown promise across various domains.

In this systematic review, we organized protocols for generating retinal organoids using induced pluripotent stem cells from patients with inherited retinal diseases. Retinal organoid technology has various applications including disease modeling, screening for novel therapies, and cell replacement therapy. Further advancements would make this technology a clinically significant tool for patients with inherited retinal diseases.

The optic nerve, comprising axons from retinal ganglion cells (RGCs), is a component of the central nervous system (CNS) that generally exhibits a limited regeneration capacity following injury in mature mammals, resulting in permanent vision loss. Here, we summarize recent advances in interventions targeting cell-intrinsic and cell-extrinsic mechanisms to enhance RGC axon regeneration. Additionally, we summarize strategies for guiding the reconnection of regenerating axons with brain visual targets, aiming to restore partial visual function. Given the advent of high-throughput screening techniques and multiomics analyses, we discuss how these emerging methodologies deepen our understanding of regenerative mechanisms and expedite the development of innovative therapeutic approaches. Lastly, we explore the translational potential of these strategies in achieving clinically meaningful vision recovery.

This study characterizes a fluorescent Slc17a6-tdTomato neuronal reporter mouse line with strong labeling of axons throughout the optic nerve, of retinal ganglion cell (RGC) soma in the ganglion cell layer (GCL), and of RGC dendrites in the inner plexiform layer (IPL). The model facilitated assessment of RGC loss in models of degeneration and of RGC detection in mixed neural/glial cultures. The tdTomato signal showed strong overlap with >98% cells immunolabeled with RGC markers RBPMS or BRN3A, consistent with the ubiquitous presence of the vesicular glutamate transporter 2 (VGUT2, SLC17A6) in all RGC subtypes. There was no cross-labeling of ChAT-positive displaced amacrine cells in the GCL, although some signal emanated from the outer plexiform layer, consistent with horizontal cells. The fluorescence allowed rapid screening of RGC loss following optic nerve crush and intraocular pressure (IOP) elevation. The bright fluorescence also enabled non-invasive monitoring of extensive neurite networks and neuron/astrocyte interactions in culture. Robust Ca2+ responses to P2X7R agonist BzATP were detected from fluorescent RGCs using Ca2+-indicator Fura-2. Fluorescence from axons and soma was detected in vivo with a confocal scanning laser ophthalmoscope (cSLO); automatic RGC soma counts enhanced through machine learning approached the numbers found in retinal wholemounts. Controls indicated no impact of Slc17a6-tdTomato expression on light-dependent neuronal function as measured with a microelectrode array (MEA), or on retinal structure as measured with optical coherence tomography (OCT). In summary, the bright fluorescence in axons, dendrites and soma of ~all RGCs in the Slc17a6-tdTomato reporter mouse may facilitate the study of RGCs.

Glaucoma is a leading cause of irreversible blindness worldwide. It leads to the progressive degeneration of retinal ganglion cells (RGCs), the axons of which form the optic nerve. Enormous RGC apoptosis causes a lack of transfer of visual information to the brain. The RGC loss typical of the central nervous system is irreversible, and when glaucoma progresses, the total amount of RGCs in the retina enormously diminishes. The successful treatment in glaucoma patients is a direct neuroprotection by decreasing the intraocular pressure, which enables RGC protection but does not revive the lost ones. The intriguing new therapy for advanced glaucoma is the possibility of RGC replacement with new healthy cells. In this review article, the strategies regarding RGC replacement therapy are presented with the latest advances in the technique and the obstacles that it meets.

Optic nerve (ON) injury causes blindness in adult mammals as their retinal ganglion cells (RGCs) cannot regenerate axons. However, amphibian RGC axons do not experience the same regenerative failure. Studying the regeneration process of the ON in amphibians holds profound implications for regenerative medicine and human health. Using transgenic tadpoles and laser micro-optics, we developed a reproducible ON transection and regeneration model. Through microscopy of axon dynamics, functional testing to assess visual pathway recovery, TUNEL cell death and EdU cell proliferation assays, and RNA-seq of the retina and optic nerve, we characterized the optic nerve injury response and subsequent recovery. Our model suggests no chemoattractant gradient exists early in regeneration, with defasciculated axons sprouting in random directions from the globe-proximal cut end. Once individual axons reach the appropriate targets in the brain, their tract is reinforced by other regenerating axons, restoring normal ON morphology. Thus, guidance cues or scaffolding from brain-innervating axons likely support later stages of regeneration. After 14 days, the regenerated ON is morphologically indistinguishable from the naïve ON, and visual function is restored. We found no evidence of RGC death or new RGC formation in the model, suggesting that ON regeneration involves remodeling of injured axons of pre-existing RGCs.

Optic nerve regeneration remains challenging worldwide due to the limited intrinsic regenerative capacity of retinal ganglion cells (RGCs) and the inhibitory microenvironment. Oxidative stress, induced by excessive reactive oxygen species (ROS) following optic nerve injury, is associated with prolonged neuroinflammation, resulting in a secondary injury of RGCs and the impairment of axon regeneration. Herein, we developed a bionic nanocomposite scaffold (GA@PDA) with immunoregulatory ability for enhanced optic nerve regeneration. The ice-templating method was employed to fabricate biopolymer-based scaffolds with a directional porous structure, mimicking the optic nerve, which effectively guided the oriented growth of neuronal cells. The incorporation of bioinspired polydopamine nanoparticles (PDA NPs) further confers excellent ROS scavenging ability, thereby modulating the phenotype transformation of microglia/macrophages from pro-inflammatory M1 to anti-inflammatory M2. In a rat optic nerve crush model, the implantation of GA@PDA scaffold enhanced survival of RGCs and promoted axonal regeneration. Our study offers novel insights and holds promising potential for the advancement of engineered biomaterials in facilitating optic nerve regeneration.

Optic nerve (ON) injury causes blindness in adult mammals as their retinal ganglion cells (RGCs) cannot regenerate axons. However, amphibian RGC axons do not experience the same regenerative failure. Studying the regeneration process of the ON in amphibians holds profound implications for regenerative medicine and human health. Using transgenic tadpoles and laser micro-optics, we developed a reproducible ON transection and regeneration model. Through microscopy, functional testing, TUNEL, EdU assays, and RNA-seq, we characterized the ON injury response and recovery. Our model suggests no chemoattractant gradient exists early in regeneration, with defasciculated axons sprouting in random directions from the globe-proximal cut end. Once individual axons reach the appropriate anatomical insertion point in the brain, their tract is reinforced by other regenerating axons, restoring normal ON morphology. Thus, guidance cues or scaffolding from brain-innervating axons likely support later stages of regeneration. After 14 days, the regenerated ON is morphologically indistinguishable from the naïve ON, and visual function is restored. We found no evidence of RGC death or new RGC formation in the model, suggesting that only pre-existing RGCs are involved in ON regeneration.

This review explores the molecular and genetic underpinnings of axonal regeneration and functional recovery post-nerve injury, emphasizing its significance in reversing neurological deficits. It presents a systematic exploration of the roles of various genes in axonal regrowth across peripheral and central nerve injuries. Initially, it highlights genes and gene families critical for axonal growth and guidance, delving into their roles in regeneration. It then examines the regenerative microenvironment, focusing on the role of glial cells in neural repair through dedifferentiation, proliferation, and migration. The concept of "traumatic microenvironments" within the central nervous system (CNS) and peripheral nervous system (PNS) is discussed, noting their impact on regenerative capacities and their importance in therapeutic strategy development. Additionally, the review delves into axonal transport mechanisms essential for accurate growth and reinnervation, integrating insights from proteomics, genome-wide screenings, and gene editing advancements. Conclusively, it synthesizes these insights to offer a comprehensive understanding of axonal regeneration's molecular orchestration, aiming to inform effective nerve injury therapies and contribute to regenerative neuroscience.

The degeneration of retinal photoreceptors is one of the primary causes of blindness, and the implantation of retinal prostheses offers hope for vision restoration in individuals who are completely blind. Flexible bioelectronic devices present a promising avenue for the next generation of retinal prostheses owing to their soft mechanical properties and tissue friendliness. In this study, we developed flexible composite films of ferroelectric BiFeO3-BaTiO3 (BFO-BTO) particles synthesized by the hydrothermal method and ferroelectric poly(vinyldene difluoride-trifluoroethylene) (P(VDF-TrFE)) polymer and investigated their applications in artificial retinas. Owing to the coupling of the photothermal effect of BFO-BTO particles and the pyroelectric effect of the P(VDF-TrFE) polymer, the composite films demonstrate a strong photoelectric response (a maximum peak-to-peak photovoltage > 80 V under blue light of 100 mW/cm2) in a wide wavelength range of light (from visible to infrared) with the inherent flexibility and ease of preparation, making it an attractive candidate for artificial retinal applications. Experimental results showed that blind rats implanted with artificial retinas of the composites display light-responsive behavior, showcasing the effectiveness of vision restoration. This study demonstrates a novel approach for employing ferroelectric materials in vision restoration and offers insights into future artificial retina design.

Optic nerve imaging is crucial for diagnosing and understanding optic neuropathies because it provides detailed visualization of the nerve's structure and pathologies through advanced modalities. This study conducted a bibliometric analysis within the field of optic nerve imaging, aiming to pinpoint the latest research trends and focal points in optic nerve imaging.

The core literature on optic nerve imaging published between January 1991 and August 2023 was retrieved from the Web of Science Core Collection. The analysis and visualization of scientific productivity and emerging trends were facilitated through the utilization of Bibliometrix software, CiteSpace, Gephi, VOSviewer, R software, and Python.

In total, 15,247 publications on optic nerve imaging were included in the analysis. Notably, the top 3 journals contributing to this field were Investigative Ophthalmology & Visual Science, Ophthalmology, and the British Journal of Ophthalmology. This research on optic nerve imaging extended across 97 countries, with the USA leading in research endeavors. Noteworthy burst term analysis revealed that "Segmentation" and "Machine learning" are gaining attention. Additionally, the Latent Dirichlet Allocation model indicated that image processing has been a hotspot in recent years.

This study revealed the research trends, hotspots, and emerging topics in optic nerve imaging through bibliometric analysis and network visualization. At present, the research focus is directed towards employing artificial intelligence for image post-processing. The findings of this study offer valuable insights into future research direction and clinical applications.

Fish retinal ganglion cells (RGCs) can regenerate after optic nerve lesions (ONLs). We previously reported that heat shock factor 1 (HSF1) and Yamanaka factors increased in the zebrafish retina 0.5-24 h after ONLs, and they led to cell survival and the transformation of neuro-stem cells. We also showed that retinoic acid (RA) signaling and transglutaminase 2 (TG2) were activated in the fish retina, performing neurite outgrowth 5-30 days after ONLs. In this study, we found that RA signaling and TG2 increased within 0.5 h in the zebrafish retina after ONLs. We examined their interaction with the TG2-specific morpholino and inhibitor due to the significantly close initiation time of TG2 and HSF1. The inhibition of TG2 led to the complete suppression of HSF1 expression. Furthermore, the results of a ChIP assay with an anti-TG2 antibody evidenced significant anti-TG2 immunoprecipitation of HSF1 genome DNA after ONLs. The inhibition of TG2 also suppressed Yamanaka factors' gene expression. This rapid increase in TG2 expression occurred 30 min after the ONLs, and RA signaling occurred 15 min before this change. The present study demonstrates that TG2 regulates Yamanaka factors via HSF1 signals in the acute phase of fish optic nerve regeneration.

Optic nerve (ON) regeneration in mammalian systems is limited by an overshadowing dominance of inhibitory factors. This has severely hampered the identification of pro-regenerative pathways. Here, we take advantage of the regenerative capacity of larval zebrafish to identify pathways that promote ON regeneration. From a small molecule screen, we identified modulators of serotonin (5-HT) signaling that inhibit ON regeneration. We find several serotonin type-1 receptor genes are expressed in RGC neurons during regeneration and that inhibiting 5-HT1 receptors or components of the 5-HT pathway selectively impedes ON regeneration. We show that 5-HT1 receptor signaling is dispensable during ON development yet is critical for regenerating axons to emerge from the injury site. Blocking 5-HT receptors once ON axons have crossed the chiasm does not inhibit regeneration, suggesting a selective role for 5-HT receptor signaling early during ON regeneration. Finally, we show that agonist-mediated activation of 5-HT1 receptors leads to enhanced and ectopic axonal regrowth. Combined, our results provide evidence for mechanisms through which serotonin-dependent neuromodulation directs ON regeneration in vivo.

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.

How to cite this article: Dada T, Mahalingam K, Bhartiya S. Reversing Aging and Improving Health Span in Glaucoma Patients: The Next Frontier? J Curr Glaucoma Pract 2024;18(3):87-93.

This study characterizes a fluorescent Slc17a6 -tdTomato neuronal reporter mouse line offering strong labeling in axons throughout the optic nerve, dendrites and soma in 99% of retinal ganglion cells (RGCs). The model facilitates neuronal assessment ex vivo with wholemounts quantified to show neurodegeneration following optic nerve crush or elevated IOP as related to glaucoma, in vitro with robust Ca 2+ responses to P2X7 receptor stimulation in neuronal cultures, and in vivo using a confocal scanning laser ophthalmoscope (cSLO). While the tdTomato signal showed strong overlap with RGC markers, BRN3A and RBPMS, there was no cross-labeling of displaced amacrine cells in the ganglion cell layer. Controls indicated no impact of Slc17a6 -tdTomato expression on light-dependent neuronal function, as determined with a microelectrode array (MEA), or on structure, as measured with optical coherence tomography (OCT). In summary, this novel neuronal reporter mouse model offers an effective means to increase the efficiency for real-time, specific visualization of retinal ganglion cells. It holds substantial promise for enhancing our understanding of RGC pathology in glaucoma and other diseases of the optic nerve, and could facilitate the screening of targeted therapeutic interventions for neurodegeneration.

Whole-eye transplantation emerges as a frontier in ophthalmology, promising a transformative approach to irreversible blindness. Despite advancements, formidable challenges persist. Preservation of donor eye viability post-enucleation necessitates meticulous surgical techniques to optimize retinal integrity and ganglion cell survival. Overcoming the inhibitory milieu of the central nervous system for successful optic nerve regeneration remains elusive, prompting the exploration of neurotrophic support and immunomodulatory interventions. Immunological tolerance, paramount for graft acceptance, confronts the distinctive immunogenicity of ocular tissues, driving research into targeted immunosuppression strategies. Ethical and legal considerations underscore the necessity for stringent standards and ethical frameworks. Interdisciplinary collaboration and ongoing research endeavors are imperative to navigate these complexities. Biomaterials, stem cell therapies, and precision immunomodulation represent promising avenues in this pursuit. Ultimately, the aim of this review is to critically assess the current landscape of whole-eye transplantation, elucidating the challenges and advancements while delineating future directions for research and clinical practice. Through concerted efforts, whole-eye transplantation stands to revolutionize ophthalmic care, offering hope for restored vision and enhanced quality of life for those afflicted with blindness.

Extracellular vesicles (EVs), including exosomes, microvesicles, and other lipid vesicles derived from cells, play a pivotal role in intercellular communication by transferring information between cells. EVs secreted by progenitor and stem cells have been associated with the therapeutic effects observed in cell-based therapies, and they also contribute to tissue regeneration following injury, such as in orthopaedic surgery cases. This review explores the involvement of EVs in nerve regeneration, their potential as drug carriers, and their significance in stem cell research and cell-free therapies. It underscores the importance of bioengineers comprehending and manipulating EV activity to optimize the efficacy of tissue engineering and regenerative therapies.

PVD neuron of Caenorhabditis elegans is a highly polarized cell with well-defined axonal, and dendritic compartments. PVD neuron operates in multiple sensory modalities including the control of both nociceptive touch sensation and body posture. Although both the axon and dendrites of this neuron show a regeneration response following laser-assisted injury, it is rather unclear how the behavior associated with this neuron is affected by the loss of these structures. It is also unclear whether neurite regrowth would lead to functional restoration in these neurons. Upon axotomy, using a femtosecond laser, we saw that harsh touch response was specifically affected leaving the body posture unperturbed. Subsequently, recovery in the touch response is highly correlated to the axon regrowth, which was dependent on DLK-1/MLK-1 MAP Kinase. Dendrotomy of both major and minor primary dendrites affected the wavelength and amplitude of sinusoidal movement without any apparent effect on harsh touch response. We further correlated the recovery in posture behavior to the type of dendrite regeneration events. We found that dendrite regeneration through the fusion and reconnection between the proximal and distal branches of the injured dendrite corresponded to improved recovery in posture. Our data revealed that the axons and dendrites of PVD neurons regulate the nociception and proprioception in worms, respectively. It also revealed that dendrite and axon regeneration lead to the restoration of these differential sensory modalities.

Vision is formed by the transmission of light stimuli to the brain through axons extending from photoreceptor cells. Damage to these axons leads to loss of vision. Despite research on neural circuit regeneration through transplantation, achieving precise axon projection remains challenging. To achieve optic nerve regeneration by transplantation, we employed the Drosophila visual system. We previously established a transplantation method for Drosophila utilizing photoreceptor precursor cells extracted from the eye disc. However, little axonal elongation of transplanted cells into the brain, the lamina, was observed. We verified axonal elongation to the lamina by modifying the selection process for transplanted cells. Moreover, we focused on N-cadherin (Ncad), a cell adhesion factor, and Twinstar (Tsr), which has been shown to promote actin reorganization and induce axon elongation in damaged nerves. Overexpression of Ncad and tsr promoted axon elongation to the lamina, along with presynaptic structure formation in the elongating axons. Furthermore, overexpression of Neurexin-1 (Nrx-1), encoding a protein identified as a synaptic organizer, was found to not only promote presynapse formation but also enhance axon elongation. By introducing Ncad, tsr, and Nrx-1, we not only successfully achieved axonal projection of transplanted cells to the brain beyond the retina, but also confirmed the projection of transplanted cells into a deeper ganglion, the medulla. The present study offers valuable insights to realize regeneration through transplantation in a more complex nervous system.

The accuracy of diffusion MRI tractography reconstruction decreases in the white matter regions with crossing fibers. The optic pathways in rodents provide a challenging structure to test new diffusion tractography approaches because of the small crossing volume within the optic chiasm and the unbalanced 9:1 proportion between the contra- and ipsilateral neural projections from the retina to the lateral geniculate nucleus, respectively.

Common approaches based on Orientation Distribution Function (ODF) peak finding or statistical inference were compared qualitatively and quantitatively to ODF Fingerprinting (ODF-FP) for reconstruction of crossing fibers within the optic chiasm using in vivo diffusion MRI (n = 18 $$ n=18 $$ healthy C57BL/6 mice). Manganese-Enhanced MRI (MEMRI) was obtained after intravitreal injection of manganese chloride and used as a reference standard for the optic pathway anatomy.

ODF-FP outperformed by over 100% all the tested methods in terms of the ratios between the contra- and ipsilateral segments of the reconstructed optic pathways as well as the spatial overlap between tractography and MEMRI.

In this challenging model system, ODF-Fingerprinting reduced uncertainty of diffusion tractography for complex structural formations of fiber bundles.

The aim of this study was to develop and validate a test to assess visual function in pigs using the visual psychophysics contrast sensitivity function.

We utilized a touchscreen along with a pellet reward dispenser to train three Göttingen pigs on a visual psychophysics test and determined their contrast sensitivity function. Images with different contrast resolutions were used as visual stimuli and presented against a control image in a two-choice test. Following animals' acclimatization and the first phase of training, the system was arranged such that animals could self-run multiple consecutive trials without human intervention.

All animals were trained within a week and remembered the task with 1 day of reinforcement when tested 1 month after the last visual assessment. All trained animals performed well during the trial with minimal screen side bias, especially at contrast threshold above 40%.

Göttingen pigs are trainable for a visual psychophysics test and able to self-run the trial without human intervention.

Contrast sensitivity is one of the key parameters to assess visual function in humans. The possibility of measuring the same parameters in a large animal model allows for a better translation and understanding of drug safety and efficacy in preclinical ophthalmology.

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.

Leading causes of blindness worldwide include neurodegenerative diseases of the retina, which cause irreversible loss of retinal pigment epithelium (RPE) and photoreceptors, and optic neuropathies, which result in retinal ganglion cell (RGC) death. Because photoreceptor and RGCs do not spontaneously regenerate in mammals, including humans, vision loss from these conditions is, at present, permanent. Recent advances in gene and cell-based therapies have provided new hope to patients affected by these conditions. This chapter reviews the current state and future of these approaches to treating ocular neurodegenerative disease. Gene therapies for retinal degeneration and optic neuropathies primarily focus on correcting known pathogenic mutations that cause inherited conditions to halt progression. There are multiple retinal and optic neuropathy gene therapies in clinical trials, and one retinal gene therapy is approved in the United States, Canada, Europe, and Australia. Cell-based therapies are mutation agnostic and have the potential to repopulate neurons regardless of the underlying etiology of degeneration. While photoreceptor cell replacement is nearing a human clinical trial, RPE transplantation is currently in phase I/II clinical trials. RGC replacement faces numerous logistical challenges, but preclinical research has laid the foundation for functional repair of optic neuropathies to be feasible.

To investigate whether Tafluprost could promote optic nerve regeneration in mice after optic nerve crush (ONC) and determine the underlying molecular mechanism.

Tafluprost was injected into the vitreous body immediately after ONC. The level of Zn2+ in the inner plexiform layer (IPL) of the retina was stained using autometallography (AMG). The number of survival retinal ganglion cells (RGCs) was determined via dual staining with RGC markers Tuj1 and RBPMS. Individual axons that regenerated to 0.25, 0.5, 0.75 and 1 mm were manually counted in the whole-mount optic nerve labeled by cholera toxin B fragment (CTB). Immunofluorescence and Western blot were performed to detect protein expression levels. Pattern electroretinogram was used to evaluate RGCs function.

Tafluprost promoted RGC survival in a dose-dependent manner with an optimal concentration of 1 μM. Tafluprost significantly decreased ZnT-3 expression and Zn2+ accumulation in the IPL of retina. Tafluprost stimulated intense axonal regeneration and maintained RGCs function compared to control. Mechanistically, Tafluprost and Zn2+ elimination treatment (TPEN or ZnT-3 deletion) can activate the mTOR pathway with an improved percentage of pS6+ RGCs in the retina. However, rapamycin, a specific inhibitor of the mTOR1, inhibited the activation of the mTOR pathway and abolished the regenerative effect mediated by Tafluprost. Tafluprost also inhibited the upregulation of p62, LC3 and Beclin-1, attenuated the overactivation of microglia/macrophages and downregulated the expression of TNFα and IL-1β.

Our results suggest that Tafluprost promoted axon regeneration via regulation of the Zn2+-mTOR pathway, and provide novel research directions for glaucomatous optic nerve injury mechanisms.

Glaucoma, the leading cause of irreversible blindness worldwide, disproportionately affects individuals of African descent. Specifically, previous research has indicated that primary open-angle glaucoma (POAG), the most common form of disease, is more prevalent, severe, early-onset, and rapidly-progressive in populations of African ancestry. Recent studies have identified genetic variations that may contribute to the greater burden of disease in this population. In particular, mitochondrial genetics has emerged as a profoundly influential factor in multiple neurodegenerative diseases, including POAG. Several hypotheses explaining the underlying mechanisms of mitochondrial genetic contribution to disease progression have been proposed, including nuclear-mitochondrial gene mismatch. Exploring the fundamentals of mitochondrial genetics and disease pathways within the understudied African ancestry population can lead to groundbreaking advancements in the research and clinical understanding of POAG. This article discusses the currently known involvements of mitochondrial genetic factors in POAG, recent directions of study, and potential future prospects in mitochondrial genetic studies in individuals of African descent.

Purpose: There is increasing interest in nonpharmacologic approaches to protect retinal ganglion cells (RGCs) after injury and enhance the efficacy of therapeutic molecules. Accumulating evidence demonstrates neuroprotection by the high-fat low-carbohydrate ketogenic diet (KD) in humans and animal models of neurologic diseases. However, no studies to date have examined whether the KD protects RGCs and promotes axonal regrowth after traumatic injury to the optic nerve (ON) or whether it increases efficacy of experimental proregenerative molecules. In this study, we investigated whether the KD promoted RGC survival and axonal regeneration after ON injury in the presence and absence of neuroprotective Wnt3a ligand. Methods: Adult mice were placed on a KD or control diet before ON crush injury and remained on the diet until the end of the experiment. Nutritional ketosis was confirmed by measuring serum beta-hydroxybutyrate levels. Mice were intravitreally injected with Wnt3a ligand or phosphate-buffered saline (PBS), and RGC survival, function, axonal regeneration, and inflammatory responses were measured. Results: Mice fed the KD showed increased RGC survival and reduced inflammatory cells in PBS-injected mice. Also, mice fed the KD had increased RGC functional responses but not increased RGC numbers in the presence of Wnt3a, indicating that the KD did not enhance the prosurvival effect of Wnt3a. The KD did not promote axonal regeneration in the presence or absence of Wnt3a. Conclusions: The KD has a complex protective effect after ON injury and cotreatment with Wnt3a. This work sets the foundation for studies identifying underlying molecular mechanisms.

Glaucoma is a leading cause of irreversible blindness worldwide. Retinal ganglion cells (RGC), the neurons that connect the eyes to the brain, specifically die in glaucoma, leading to blindness. Elevated intraocular pressure (IOP) is the only modifiable risk factor, however, many patients progress despite excellent IOP control. Thus, alternative treatment strategies to prevent glaucoma progression are an unmet need. Citicoline has demonstrated neuroprotective properties in central neurodegenerative diseases. However, conclusive evidence of the effect of citicoline on glaucoma progression is missing. This systematic review investigates first-time the therapeutic potential of citicoline in glaucoma patients.

The present study was conducted according to the PRISMA 2020 statement. PubMed, Web of Science, Google Scholar, and Embase were accessed in July 2023 to identify all clinical studies investigating the efficacy of citicoline on IOP, the mean deviation of the 24-2 visual field testing (MD 24-2), retinal nerve fibre layer (RNFL), and the pattern electroretinogram (PERG) P50-N95 amplitude in glaucoma patients. The risk of bias was assessed using the Review Manager 5.3 software (The Nordic Cochrane Collaboration, Copenhagen) and the Risk of Bias in Non-randomised Studies of Interventions (ROBINS-I) tool.

Ten studies were eligible for this systematic review, including 424 patients. The mean length of the follow-up was 12.1 ± 11.6 months. The overall risk of bias was low to moderate. The mean age of the patients was 56.7 years. There were no significant differences in the IOP, MD 24-2, RNFL, or PERG P50-N95 amplitude between patients receiving citicoline and the control group. There was no improvement from baseline to the last follow-up in IOP, MD 24-2, RNFL, or PERG P50-N95 amplitude.

There is a lack of sufficient evidence to support that citicoline slows the progression of glaucoma.

Retinal ganglion cell (RGC) death in glaucoma and other optic neuropathies results in irreversible vision loss due to the mammalian central nervous system's limited regenerative capacity. RGC repopulation is a promising therapeutic approach to reverse vision loss from optic neuropathies if the newly introduced neurons can reestablish functional retinal and thalamic circuits. In theory, RGCs might be repopulated through the transplantation of stem cell-derived neurons or via the induction of endogenous transdifferentiation. The RGC Repopulation, Stem Cell Transplantation, and Optic Nerve Regeneration (RReSTORe) Consortium was established to address the challenges associated with the therapeutic repair of the visual pathway in optic neuropathy. In 2022, the RReSTORe Consortium initiated ongoing international collaborative discussions to advance the RGC repopulation field and has identified five critical areas of focus: (1) RGC development and differentiation, (2) Transplantation methods and models, (3) RGC survival, maturation, and host interactions, (4) Inner retinal wiring, and (5) Eye-to-brain connectivity. Here, we discuss the most pertinent questions and challenges that exist on the path to clinical translation and suggest experimental directions to propel this work going forward. Using these five subtopic discussion groups (SDGs) as a framework, we suggest multidisciplinary approaches to restore the diseased visual pathway by leveraging groundbreaking insights from developmental neuroscience, stem cell biology, molecular biology, optical imaging, animal models of optic neuropathy, immunology & immunotolerance, neuropathology & neuroprotection, materials science & biomedical engineering, and regenerative neuroscience. While significant hurdles remain, the RReSTORe Consortium's efforts provide a comprehensive roadmap for advancing the RGC repopulation field and hold potential for transformative progress in restoring vision in patients suffering from optic neuropathies.