Neuroprotective effects of glial mediators in interactions between retinal neurons and Müller cells.

Progressive retinal ganglion cell (RGC) loss underlies a number of retinal neurodegenerative disorders, which may lead to permanent vision loss. However, secreted neuroprotective factors, such as PEDF, VEGF and IL-6, which are produced by Müller cells, have been shown to promote RGC survival. Assuming that the communication of RGCs with Müller cells involves a release of glioactive substances we sought to determine whether retinal neurons are able to modulate expression levels of Müller cell-derived PEDF, VEGF and IL-6. We demonstrate elevated mRNA levels of these factors in Müller cells in co-cultures with RGCs or R28 cells when compared to homotypic Müller cell cultures. Furthermore, R28 cells were more protected from apoptosis when co-cultured with Müller cells. IL-6 and VEGF were upregulated in Müller cells under hypoxia. Both cytokines, as well as PEDF, induced an altered neuronal expression of members of the Bcl-2 family, which are central molecules in the regulation of apoptosis. These results suggest that in retinal ischemia, via own secreted mediators, RGCs can resist a potential demise by stimulating Müller cells to increase production of neuroprotective factors, which counteract RGC apoptosis.

Retinal Stem Cell 'Retirement Plans': Growth, Regulation and Species Adaptations in the Retinal Ciliary Marginal Zone.

The vertebrate retina develops from a specified group of precursor cells that adopt distinct identities and generate lineages of either the neural retina, retinal pigmented epithelium, or ciliary body. In some species, including teleost fish and amphibians, proliferative cells with stem-cell-like properties capable of continuously supplying new retinal cells post-embryonically have been characterized and extensively studied. This region, termed the ciliary or circumferential marginal zone (CMZ), possibly represents a conserved retinal stem cell niche. In this review, we highlight the research characterizing similar CMZ-like regions, or stem-like cells located at the peripheral margin, across multiple different species. We discuss the proliferative parameters, multipotency and growth mechanisms of these cells to understand how they behave in vivo and how different molecular factors and signalling networks converge at the CMZ niche to regulate their activity. The evidence suggests that the mature retina may have a conserved propensity for homeostatic growth and plasticity and that dysfunction in the regulation of CMZ activity may partially account for dystrophic eye growth diseases such as myopia and hyperopia. A better understanding of the properties of CMZ cells will enable important insight into how an endogenous generative tissue compartment can adapt to altered retinal physiology and potentially even restore vision loss caused by retinal degenerative conditions.

A single-cell guide to retinal development: Cell fate decisions of multipotent retinal progenitors in scRNA-seq.

Recent advances in high throughput single-cell RNA sequencing (scRNA-seq) technology have enabled the simultaneous transcriptomic profiling of thousands of individual cells in a single experiment. To investigate the intrinsic process of retinal development, researchers have leveraged this technology to quantify gene expression in retinal cells across development, in multiple species, and from numerous important models of human disease. In this review, we summarize recent applications of scRNA-seq and discuss how these datasets have complemented and advanced our understanding of retinal progenitor cell competence, cell fate specification, and differentiation. Finally, we also highlight the outstanding questions in the field that advances in single-cell data generation and analysis will soon be able to answer.

Determinants of peripapillary retinal nerve fiber layer's grayscale value in normal eyes by spectral domain optical coherence tomography.

To determine and evaluate the distribution, variation, and determinants of peripapillary retinal nerve fiber layer (pRNFL) grayscale value with spectral-domain optical coherence tomography (SD-OCT) in normal eyes. In this cross-sectional study, three hundred ninety-seven normal eyes from 397 healthy Chinese adults aged 18-80 were consecutively recruited from a tertiary eye care center. An SD-OCT instrument took pRNFL imaging. We used a customized software to measure pRNFL parameters, including thickness and grayscale value. Univariable and multiple linear regression analyses were performed to examine the relationship between pRNFL grayscale value with ocular (e.g., axial length [A.L.], spherical equivalent [S.E.], intraocular pressure [IOP]), and systemic (e.g., age, sex) factors. A total of 397 eyes from 397 healthy subjects were included in the final analysis with mean (± SD) age 44.63 ± 16.43 years (range 18-80 years) and 196 (49.4%) males. The mean average of pRNFL grayscale value and thickness 164.82 ± 5.69 and 106.68 ± 8.89 µm, respectively. pRNFL grayscale value in nasal sectors (163.26 ± 9.31) was significantly lower comparing those in all other five sectors (all with p < 0.001)]. In multivariable analysis, average pRNFL grayscale value was independently correlated to older age (ß = - 0.053, p = 0.002), longer axial length (ß = - 0.664, p = 0.003), lower RPE grayscale value (ß = 0.372, p < 0.001) and lower ImageQ (ß = 0.658, p < 0.001). In this study, we provided normative SD-OCT data on the pRNFL grayscale value profile in nonglaucomatous eyes. Lower average pRNFL grayscale value was independently correlated to older age, longer axial length, lower RPE grayscale value, and lower ImageQ. These determinants should be considered when interpreting pRNFL grayscale value in glaucoma assessment.

Uptake and Distribution of Administered Bone Marrow Mesenchymal Stem Cell Extracellular Vesicles in Retina.

Cell replacement therapy using mesenchymal (MSC) and other stem cells has been evaluated for diabetic retinopathy and glaucoma. This approach has significant limitations, including few cells integrated, aberrant growth, and surgical complications. Mesenchymal Stem Cell Exosomes/Extracellular Vesicles (MSC EVs), which include exosomes and microvesicles, are an emerging alternative, promoting immunomodulation, repair, and regeneration by mediating MSC's paracrine effects. For the clinical translation of EV therapy, it is important to determine the cellular destination and time course of EV uptake in the retina following administration. Here, we tested the cellular fate of EVs using in vivo rat retinas, ex vivo retinal explant, and primary retinal cells. Intravitreally administered fluorescent EVs were rapidly cleared from the vitreous. Retinal ganglion cells (RGCs) had maximal EV fluorescence at 14 days post administration, and microglia at 7 days. Both in vivo and in the explant model, most EVs were no deeper than the inner nuclear layer. Retinal astrocytes, microglia, and mixed neurons in vitro endocytosed EVs in a dose-dependent manner. Thus, our results indicate that intravitreal EVs are suited for the treatment of retinal diseases affecting the inner retina. Modification of the EV surface should be considered for maintaining EVs in the vitreous for prolonged delivery.

Novel Technique for Retinal Nerve Cell Regeneration with Electrophysiological Functions Using Human Iris-Derived iPS Cells.

Regenerative medicine in ophthalmology that uses induced pluripotent stem cells (iPS) cells has been described, but those studies used iPS cells derived from fibroblasts. Here, we generated iPS cells derived from iris cells that develop from the same inner layer of the optic cup as the retina, to regenerate retinal nerves. We first identified cells positive for p75NTR, a marker of retinal tissue stem and progenitor cells, in human iris tissue. We then reprogrammed the cultured p75NTR-positive iris tissue stem/progenitor (H-iris stem/progenitor) cells to create iris-derived iPS (H-iris iPS) cells for the first time. These cells were positive for iPS cell markers and showed pluripotency to differentiate into three germ layers. When H-iris iPS cells were pre-differentiated into neural stem/progenitor cells, not all cells became positive for neural stem/progenitor and nerve cell markers. When these cells were pre-differentiated into neural stem/progenitor cells, sorted with p75NTR, and used as a medium for differentiating into retinal nerve cells, the cells differentiated into Recoverin-positive cells with electrophysiological functions. In a different medium, H-iris iPS cells differentiated into retinal ganglion cell marker-positive cells with electrophysiological functions. This is the first demonstration of H-iris iPS cells differentiating into retinal neurons that function physiologically as neurons.

Prpf31 is essential for the survival and differentiation of retinal progenitor cells by modulating alternative splicing.

Dysfunction of splicing factors often result in abnormal cell differentiation and apoptosis, especially in neural tissues. Mutations in pre-mRNAs processing factor 31 (PRPF31) cause autosomal dominant retinitis pigmentosa, a progressive retinal degeneration disease. The transcriptome-wide splicing events specifically regulated by PRPF31 and their biological roles in the development and maintenance of retina are still unclear. Here, we showed that the differentiation and viability of retinal progenitor cells (RPCs) are severely perturbed in prpf31 knockout zebrafish when compared with other tissues at an early embryonic stage. At the cellular level, significant mitotic arrest and DNA damage were observed. These defects could be rescued by the wild-type human PRPF31 rather than the disease-associated mutants. Further bioinformatic analysis and experimental verification uncovered that Prpf31 deletion predominantly causes the skipping of exons with a weak 5' splicing site. Moreover, genes necessary for DNA repair and mitotic progression are most enriched among the differentially spliced events, which may explain the cellular and tissular defects in prpf31 mutant retinas. This is the first time that Prpf31 is demonstrated to be essential for the survival and differentiation of RPCs during retinal neurogenesis by specifically modulating the alternative splicing of genes involved in DNA repair and mitosis.

New Optical Tools to Study Neural Circuit Assembly in the Retina.

During development, neurons navigate a tangled thicket of thousands of axons and dendrites to synapse with just a few specific targets. This phenomenon termed wiring specificity, is critical to the assembly of neural circuits and the way neurons manage this feat is only now becoming clear. Recent studies in the mouse retina are shedding new insight into this process. They show that specific wiring arises through a series of stages that include: directed axonal and dendritic growth, the formation of neuropil layers, positioning of such layers, and matching of co-laminar synaptic partners. Each stage appears to be directed by a distinct family of recognition molecules, suggesting that the combinatorial expression of such family members might act as a blueprint for retinal connectivity. By reviewing the evidence in support of each stage, and by considering their underlying molecular mechanisms, we attempt to synthesize these results into a wiring model which generates testable predictions for future studies. Finally, we conclude by highlighting new optical methods that could be used to address such predictions and gain further insight into this fundamental process.

dnmt1 function is required to maintain retinal stem cells within the ciliary marginal zone of the zebrafish eye.

The ciliary marginal zone (CMZ) of the zebrafish retina contains a population of actively proliferating resident stem cells, which generate retinal neurons throughout life. The maintenance methyltransferase, dnmt1, is expressed within the CMZ. Loss of dnmt1 function results in gene misregulation and cell death in a variety of developmental contexts, however, its role in retinal stem cell (RSC) maintenance is currently unknown. Here, we demonstrate that zebrafish dnmt1s872 mutants possess severe defects in RSC maintenance within the CMZ. Using a combination of immunohistochemistry, in situ hybridization, and a transgenic reporter assay, our results demonstrate a requirement for dnmt1 activity in the regulation of RSC proliferation, gene expression and in the repression of endogenous retroelements (REs). Ultimately, cell death is elevated in the dnmt1-/- CMZ, but in a p53-independent manner. Using a transgenic reporter for RE transposition activity, we demonstrate increased transposition in the dnmt1-/- CMZ. Taken together our data identify a critical role for dnmt1 function in RSC maintenance in the vertebrate eye.

Reprogramming Müller Glia to Regenerate Retinal Neurons.

In humans, various genetic defects or age-related diseases, such as diabetic retinopathies, glaucoma, and macular degeneration, cause the death of retinal neurons and profound vision loss. One approach to treating these diseases is to utilize stem and progenitor cells to replace neurons in situ, with the expectation that new neurons will create new synaptic circuits or integrate into existing ones. Reprogramming non-neuronal cells in vivo into stem or progenitor cells is one strategy for replacing lost neurons. Zebrafish have become a valuable model for investigating cellular reprogramming and retinal regeneration. This review summarizes our current knowledge regarding spontaneous reprogramming of Müller glia in zebrafish and compares this knowledge to research efforts directed toward reprogramming Müller glia in mammals. Intensive research using these animal models has revealed shared molecular mechanisms that make Müller glia attractive targets for cellular reprogramming and highlighted the potential for curing degenerative retinal diseases from intrinsic cellular sources.

Physiologic maturation is both extrinsically and intrinsically regulated in progenitor-derived neurons.

During development, newly-differentiated neurons undergo several morphological and physiological changes to become functional, mature neurons. Physiologic maturation of neuronal cells derived from isolated stem or progenitor cells may provide insight into maturation in vivo but is not well studied. As a step towards understanding how neuronal maturation is regulated, we studied the developmental switch of response to the neurotransmitter GABA, from excitatory depolarization to inhibitory hyperpolarization. We compared acutely isolated retinal ganglion cells (RGCs) at various developmental stages and RGCs differentiated in vitro from embryonic retinal progenitors for the effects of aging and, independently, of retinal environment age on their GABAA receptor (GABAAR) responses, elicited by muscimol. We found that neurons generated in vitro from progenitors exhibited depolarizing, immature GABA responses, like those of early postnatal RGCs. As progenitor-derived neurons aged from 1 to 3 weeks, their GABA responses matured. Interestingly, signals secreted by the early postnatal retina suppressed acquisition of mature GABA responses. This suppression was not associated with changes in expression of GABAAR or of the chloride co-transporter KCC2, but rather with inhibition of KCC2 dimerization in differentiating neurons. Taken together, these data indicate GABA response maturation depends on release of inhibition by developmentally regulated diffusible signals from the retina.

In-vivo longitudinal changes in thickness of the postnatal canine retina.

The objectives of the present work were to assess by spectral domain optical coherence tomography (OCT) the changes in thickness of the outer nuclear layer (ONL), the ONL + photoreceptor inner segment (IS), and the retinal thickness, as a function of age in the normal canine retina. OCT retinal scans extending from the edge of the optic nerve head (ONH) along the superior and inferior meridians were captured in both eyes of 17 normal dogs at age ranging from 4 to 119 weeks. The different parameters along the superior and the inferior regions were determined following manual segmentation using the Heidelberg Eye Explorer software. Changes in thickness with age were modeled using one-phase exponential decay models. In vivo OCT imaging results showed no interocular statistically significant differences in ONL, ONL + IS, and retinal thickness at any age. All three parameters were however found to be statistically significantly thicker in the superior vs inferior retina. A rapid thinning of the three layers occurs in both the superior and inferior retina between 4 and 12 weeks of age, before reaching a plateau at around 20 weeks of age. In conclusion, the ONL, ONL + IS, and retinal thickness of the normal canine retina decrease significantly during the first three postnatal months, and is likely attributed to an overall increase in the eye volume and tangential dispersion of the photoreceptor since early photoreceptor developmental cell death is very limited at that age. Establishment of the natural history of ONL, ONL + IS, and retinal thinning will allow a more accurate assessment of the progression of a retinal degenerative condition as well as facilitate the detection of positive rescue effect of novel retinal therapies evaluated in this large animal model.

Spatiotemporal gene expression patterns reveal molecular relatedness between retinal laminae.

In several areas of the central nervous system, neurons are regionally organized into groups or layers that carry out specific activities. In this form of patterning, neurons of distinct types localize their cell bodies to just one or a few of the layers within a structure. However, little is known about whether diverse neuron types within a lamina share molecular features that coordinate their organization. To begin to identify such candidates, we used the laminated murine retina to screen 92 lacZ reporter lines available through the Knockout Mouse Project. Thirty-two of these displayed reporter expression in restricted subsets of inner retina neurons. We then identified the spatiotemporal expression patterns of these genes at key developmental stages. This uncovered several that were heavily enriched in development but reduced in adulthood, including the transcriptional regulator Hmga1. An additional set of genes displayed maturation associated laminar enrichment. Among these, we identified Bbox1 as a novel gene that specifically labels all neurons in the ganglion cell layer but is largely excluded from otherwise molecularly similar neurons in the inner retina. Finally, we established Dbn1 as a new marker enriched in amacrines and Fmnl3 as a marker for subsets of αRGCs. Together, these data provide a spatiotemporal map for laminae-specific molecules and suggest that diverse neuron types within a lamina share coordinating molecular features that may inform their fate or function.

Pigment Epithelium-Derived Factor (PEDF) Receptors Are Involved in Survival of Retinal Neurons.

The demise of retinal ganglion cells (RGCs) is characteristic of diseases of the retina such as glaucoma and diabetic or ischemic retinopathies. Pigment epithelium-derived factor (PEDF) is a multifunctional secreted protein that mediates neuroprotection and inhibition of angiogenesis in the retina. We have studied expression and regulation of two of several receptors for PEDF, patatin-like phospholipase 2 gene product/PEDF-R and laminin receptor (LR), in serum-starved RGC under normoxia and hypoxia and investigated their involvement in the survival of retinal neuronal cells. We show that PEDF-R and LR are co-expressed in RGC and R28 retinal precursor cells. Expression of both receptors was enhanced in the presence of complex secretions from retinal glial (Müller) cells and upregulated by VEGF and under hypoxic conditions. PEDF-R- and LR-knocked-down cells demonstrated a markedly attenuated expression of anti-apoptotic Bcl-2 family members (Bcl-2, Bcl-xL) and neuroprotective mediators (PEDF, VEGF, BDNF) suggesting that both PEDF-R and LR mediate pro-survival effects of PEDF on RGC. While this study does not provide evidence for a differential survival-promoting influence of either PEDF-R or LR, it nevertheless highlights the importance of both PEDF receptors for the viability of retinal neurons.

AUTOMATED RETINAL LAYER SEGMENTATION AND THEIR THICKNESS PROFILES IN HEALTHY SUBJECTS: A Comparison of 55° Wide-field and Conventional 30° Spectral Domain-Optical Coherence Tomography.

PURPOSE: To assess whether retinal thickness measurements with a standard 30° spectral domain optical coherence tomography (SD-OCT) are comparable with wide-field 55° SD-OCT. METHODS: Thirty-three healthy individuals were scanned using 55° as well as 30° SD-OCT according to a standardized protocol. Automated retinal layer segmentation of standard and wide-field SD-OCTs was assessed using customized software. RESULTS: Both lenses showed a high correlation when analyzing total retinal thickness within the central, the inner, and the outer retinal ring (r = > 0.9). Automated thickness measurements with the 55° system were marginally higher compared with the 30° lens. The thickness of each separate retinal layer using automated segmentation showed excellent correlations within the inner and outer rings (range: r = 0.6-r = 0.9 for the inner ring and range: r = 0.9-r = 1.0 for the outer ring). CONCLUSION: Fifty-five degree wide-field SD-OCT provides a good overview of the posterior pole and presents similar quantitative values as a standard 30° OCT lens. Therefore, thickness values are comparable when switching between these two lenses.

IPL Sublamination in Chicken Retinal Spheroids Is Initiated via Müller Cells and Cholinergic Differentiation, and Is Disrupted by NMDA Signaling.

Purpose: Reaggregates from E6 embryonic chicken retina exhibit areas corresponding to an inner plexiform layer (IPL), which presents an ideal in vitro model to test conditions and constraints of cholinergic and glutamatergic network formation, providing a basis for retinal tissue engineering. Here, we show that ipl formation is regulated by cholinergic starburst amacrine cells (SACs), a glial scaffold and by L-glutamate. Methods: Rosetted spheroids were cultured in absence or presence of 0.2 to 0.4 mM L-glutamate and analyzed by immuno- and enzyme histochemistry, proliferation, and apoptosis assays. Results: After 2 days in vitro (div), ipl formation was announced by acetylcholinesterase+ (AChE) and choline acetyltransferase+ (ChAT) cells. Individual vimentin+ or transitin+ Müller glial cell precursors (MCPs) in ipl centers coexpressed ChAT. Comparable to in vivo, pairwise arranged ChAT+ SACs formed two laminar subbands. Projections of calretinin+ amacrine cells (ACs) into ipl associated with MCP processes. In L-glutamate-, or NMDA-treated spheroids ipls were disrupted, including loss of SACs and MCs; coincubation with NMDA receptor inhibitor MK-801 prevented these effects. Also, many Pax6+ cells, comprising most ACs, were lost, while rho4D2+ rod photoreceptors were increased. Cell proliferation was slightly increased, while apoptosis remained unaffected. Conclusions: This demonstrated: (1) a far-advanced differentiation of an IPL in retinal spheroids, as never described before; (2) ipl sublamination was initiated by cholinergic precursor cells, which-functioning as "ipl founder cells"-(3) gave rise to neurons and glial cells; (4) these SACs and MCPs together organized ipl formation; and (5) this process was counteracted by NMDA-dependent glutamate actions.

Application of Hanging Drop Culture for Retinal Precursor-Like Cells Differentiation of Human Adipose-Derived Stem Cells Using Small Molecules.

Retinal degenerative diseases lead to blindness due to poorly regenerative potential of the retina. Recently, cell therapy is more considered for degenerative diseases. Autologous mesenchymal stem cells derived from adipose tissue are a suitable source for this purpose. Therefore, we conducted a stepwise efficient method to differentiate human adipose-derived stem cells (hADSCs) into retinal precursor-like cells in vitro. We compared two differentiation protocols, monolayer and hanging drop cultures. Through the defined medium and 3D hanging drop culture method, we could achieve up to 75% retinal precursor gene expression profile (PAX6, RAX, CHX10, and CRX) from hADSCs. By imitation of in vivo development, for direct conversion of stem cells into retinal cells, the suppression of the BMP, Nodal, and Wnt signaling pathways was carried out by using three small molecules. The hADSCs were primarily differentiated into anterior neuroectodermal cells by expression of OTX2, SIX3, and Β-TUB III and then the differentiated cells were propelled into the retinal cells. According to our data from real-time PCR, RT-PCR, immunocytochemistry, and functional assay, it seems that the hanging drop method improved retinal precursor differentiation yield which these precursor-like cells respond to glutamate neurotransmitter. Regarding the easy accessibility and immunosuppressive properties of hADSCs and more efficient hanging drop method, this study may be useful for future autologous cell therapy of retinal degenerative disorders.

Topography of Neurons in the Rod Pathway of Human Retina.

Purpose: The objective of this study was to map the distribution and density of the three major components of the classical scotopic "night vision" pathway (rods, rod bipolar, and AII amacrine cells) in postmortem human retinas. Methods: Four postmortem donor eyes (male and female, aged 44-56 years) were used to cut vertical sections through the temporal horizontal meridian. The sections were processed for immunohistochemistry and imaged using high-resolution multichannel confocal microscopy. Rods, rod bipolar, and AII amacrine cells were counted along the temporal horizontal meridian. Two additional retinas were used for intracellular injections. Results: Rod peak density is close to 150,000 cells/mm2 at 4 to 5 mm (15° to 20°) eccentricity, declining to below 70,000 cells/mm2 in peripheral retina. Rod bipolar density is lower but follows a similar distribution with peak density near 10,000 cells/mm2 between 2 and 4 mm (7° to 15°) eccentricity declining to below 4000 cells/mm2 in peripheral retina. The peak density of AII amacrine cells (near 4000 cells/mm2) is located close to the fovea, at 0.5- to 2 mm-eccentricity (2° to 7°) and declines to below 1000 cells/mm2 in the periphery. Thus, convergence between rods and AII cells increases from central to peripheral retina. Conclusions: Comparison with human psychophysics and ganglion cell density indicates that the spatial resolution of scotopic vision is limited by the AII mosaic at eccentricities below 15° and by the midget ganglion cell mosaic at eccentricities above 15°.

Regeneration associated transcriptional signature of retinal microglia and macrophages.

Zebrafish have the remarkable capacity to regenerate retinal neurons following a variety of damage paradigms. Following initial tissue insult and a period of cell death, a proliferative phase ensues that generates neuronal progenitors, which ultimately regenerate damaged neurons. Recent work has revealed that Müller glia are the source of regenerated neurons in zebrafish. However, the roles of another important class of glia present in the retina, microglia, during this regenerative phase remain elusive. Here, we examine retinal tissue and perform QuantSeq. 3'mRNA sequencing/transcriptome analysis to reveal localization and putative functions, respectively, of mpeg1 expressing cells (microglia/macrophages) during Müller glia-mediated regeneration, corresponding to a time of progenitor proliferation and production of new neurons. Our results indicate that in this regenerative state, mpeg1-expressing cells are located in regions containing regenerative Müller glia and are likely engaged in active vesicle trafficking. Further, mpeg1+ cells congregate at and around the optic nerve head. Our transcriptome analysis reveals several novel genes not previously described in microglia. This dataset represents the first report, to our knowledge, to use RNA sequencing to probe the microglial transcriptome in such context, and therefore provides a resource towards understanding microglia/macrophage function during successful retinal (and central nervous tissue) regeneration.

Peripapillary Retinal Nerve Fiber Layer Thickness in Patients with Alzheimer's Disease: A Comparison of Eyes of Patients with Alzheimer's Disease, Primary Open-Angle Glaucoma, and Preperimetric Glaucoma and Healthy Controls.

BACKGROUND The aim of this study was to assess and compare peripapillary retinal nerve fiber layer (RNFL) thickness in patients with Alzheimer's disease (AD), primary open-angle glaucoma (POAG), preperimetric glaucoma (PPG), and healthy controls with the use of Spectral Domain Optical Coherence Tomography (SD-OCT). MATERIAL AND METHODS Thirty patients with AD, 30 patients with POAG, 30 patients with PPG, and 30 healthy controls were enrolled in this cross-sectional study. Only 1 randomly selected eye of each patient was analyzed. Every subject underwent a thorough ophthalmological examination and OCT of the optic disc. The peripapillary RNFL thickness in each of the 6 sectors and globally was analyzed. RESULTS The RNFL was thinnest in patients with POAG. The mean RNFL thickness value was 60.97±12.97 µm and it was significantly lower than in healthy controls (106.30±8.95 µm), patients with PPG (93.20±12.04 µm), and AD patients (95.73±13.52 µm). Mean RNFL thickness in patients with AD was significantly lower when compared to healthy controls, and was higher compared to eyes with POAG, while there were no significant differences compared to patients with PPG. CONCLUSIONS Neuronal damage in the central nervous system (CNS) also affects to retinal axons. A major problem is to distinguish the cause for a moderate decrease in the RNFL thickness. This is particularly true for patients with glaucoma who have not been diagnosed with changes in the visual field. It is not possible to distinguish the cause of a mild decrease in the RNFL thickness based on the SD-OCT. This may result in misdiagnosis of glaucoma, unnecessary use of anti-glaucoma eye drops, and a delayed diagnosis of AD.