Survival Of Retinal Neurons Research Articles

Pigment Epithelium-Derived Factor Binding to VEGFR-1 (Flt-1) Increases the Survival of Retinal Neurons.

The purpose of this study was to examine possible involvement of vascular endothelial growth factor (VEGF) receptor (VEGFR)-1/Flt-1 in pigment epithelium-derived factor (PEDF)-promoted survival of retinal neurons. Survival of growth factor-deprived retinal ganglion cells (RGCs) and R28 cells and activation of ERK-1/-2 MAP kinases were assessed in the presence of PEDF, placental growth factor (PlGF), and VEGF using cell cultures, viability assays and quantitation of ERK-1/-2 phosphorylation. VEGFR-1/Flt-1 expression was determined using quantitative PCR (qPCR) and Western blotting. VEGFR-1/Flt-1 was knocked down in R28 cells by small interfering RNA (siRNA). Binding of a PEDF-IgG Fc fusion protein (PEDF-Fc) to retinal neurons, immobilized VEGFR-1/Flt-1 and VEGFR-1/Flt-1-derived peptides was studied using binding assays and peptide scanning. PEDF in combination with PlGF stimulated increased cell survival and ERK-1/-2 MAP kinase activation compared to effects of either factor alone. VEGFR-1/Flt-1 expression in RGCs and R28 cells was significantly upregulated by hypoxia, VEGF, and PEDF. VEGFR-1/Flt-1 ligands (VEGF and PlGF) or soluble VEGFR-1 (sflt-1) competed with PEDF-Fc for binding to R28 cells. Depleting R28 cells of VEGFR-1/Flt-1 resulted in reduced PEDF-Fc binding when comparing VEGFR-1/Flt-1 siRNA- and control siRNA-treated cells. PEDF-Fc interacted with immobilized sflt-1, which was specifically blocked by VEGF and PlGF. PEDF-Fc binding sites were mapped to VEGFR-1/Flt-1 extracellular domains D3 and D4. Peptides corresponding to D3 and D4 specifically inhibited PEDF-Fc binding to R28 cells. These peptides and sflt-1 significantly inhibited PEDF-promoted survival of R28 cells. These results suggest that PEDF can target VEGFR-1/Flt-1 and this interaction plays a significant role in PEDF-mediated neuroprotection in the retina.

The Circadian Clock of Müller Glia Is Necessary for Retinal Homeostasis and Neuronal Survival

Biological processes throughout the body are orchestrated in time through the regulation of local circadian clocks. The retina is among the most metabolically active tissues, with demands depending greatly on the light/dark cycle. Most cell types within the retina are known to express the circadian clock in rodents; however, retinal clock expression in the human has not previously been localized. Moreover, the effect of local circadian clock dysfunction on retinal homeostasis is incompletely understood. We demonstrate an age-dependent decline in circadian clock gene and protein expression in the human retina. Using an animal model of targeted Bmal1 deficiency, we identify the circadian clock of the retinal Müller glia as essential for neuronal survival, vascular integrity, and retinal function. These results suggest a potential role for the local retinal circadian clock within the Müller glia in age-related retinal disease and retinal degeneration.

Fasting pancreatic polypeptide predicts incident microvascular and macrovascular complications of type 2 diabetes: An observational study.

Pancreatic polypeptide (PP) is elevated in people with vascular risk factors such as type 2 diabetes or increased visceral fat. We investigated potential relationships between PP and microvascular and macrovascular complications of diabetes. Animal study: Subcutaneous PP infusion for 4weeks in high fat diet mouse model. Retinal mRNA submitted for Ingenuity Pathway Analysis. Human study: fasting PP measured in 1478 participants and vascular complications recorded over median 5.5 (IQR 4.9-5.8) years follow-up. Animal study: The retinal transcriptional response to PP was indicative of cellular stress and damage, and this footprint matched responses described in previously published studies of retinal disease. Of mechanistic importance the transcriptional landscape was consistent with upregulation of folliculin, a recently identified susceptibility gene for diabetic retinopathy. Human study: Adjusting for established risk factors, PP was associated with prevalent and incident clinically significant retinopathy (odds ratio (OR) 1.289 (1.107-1.501) p=0.001; hazard ratio (HR) 1.259 (1.035-1.531) p=0.0213), albuminuria (OR 1.277 (1.124-1.454), p=0.0002; HR 1.608 (1.208-2.141) p=0.0011), and macrovascular disease (OR 1.021 (1.006-1.037) p=0.0068; HR 1.324 (1.089-1.61), p=0.0049), in individuals with type 2 diabetes, and progression to diabetes in non-diabetic individuals (HR 1.402 (1.081-1.818), p=0.0109). Elevated fasting PP is independently associated with vascular complications of diabetes and affects retinal pathways potentially influencing retinal neuronal survival. Our results suggest possible new roles for PP-fold peptides in the pathophysiology of diabetes complications and vascular risk stratification.

A New Modulator of Neuroinflammation in Diabetic Retinopathy: USP25

Diabetic retinopathy (DR) is a diabetes-associated complication that poses a threat to vision, distinguished by persistent and mild inflammation of the retinal microvasculature. The activation of microglia plays a crucial role in driving this pathological progression. Previous investigations have demonstrated that ubiquitin-specific peptidase 25 (USP25), a deubiquitinating enzyme, is involved in the regulation of immune cell activity. Nevertheless, the precise mechanisms through which USP25 contributes to the development of DR remain incompletely elucidated. Firstly, we have demonstrated the potential mechanism by which ROCKs can facilitate microglial activation and augment the synthesis of inflammatory mediators through the modulation of NF-κB signaling pathways in a high-glucose milieu. Furthermore, our study has provided novel insights by demonstrating that the regulatory role of USP25 in the secretion of proinflammatory factors is mediated through the involvement of ROCK in modulating the expression of NF-κB and facilitating the nuclear translocation of the phosphatase NF-κB. This regulatory mechanism plays a crucial role in modulating the activation of microglial cells within a high-glycemic environment. Hence, USP25 emerges as a pivotal determinant for the inflammatory activation of microglial cells, and its inhibition exhibits a dual effect of promoting retinal neuron survival while suppressing the inflammatory response in the retina. In conclusion, the promotion of diabetic retinopathy (DR) progression by USP25 is attributed to its facilitation of microglial activation induced by high glucose levels, a process mediated by the ROCK pathway. These findings highlight the importance of considering USP25 as a potential therapeutic target for the management of diabetic neuroinflammation.

On the potential therapeutic role of taurine in retinal degenerative diseases

Taurine, considered a non‐essential amino acid that differs from other amino acids in that it contains a sulfonic acid instead of a carboxyl group, is the most abundant amino acid in the retina. Taurine plays a critical role in maintaining retinal health and the survival of retinal neurons, although its exact function in the retina is not yet fully understood. Interestingly, taurine shares structural similarities with β‐alanine, another non‐proteinogenic amino acid that can act as a competitive inhibitor of taurine receptors, allowing β‐alanine to be used to induce taurine depletion. β‐alanine induced taurine depletion results in the activation and migration of microglial cells in the retina, oxidative stress in the inner and outer nuclear and ganglion cell layers, loss of retinal synaptic connectivity, and impaired phagocytic capacity of retinal pigment epithelial cells. In addition, taurine depletion leads to photoreceptor and retinal ganglion cell loss. Most of these detrimental events are exacerbated by light exposure. These findings suggest that taurine levels are associated with retinal sensitivity to light.Given the important role of taurine in the retina, it has been proposed as a promising therapeutic candidate for retinal degenerative diseases. These diseases, which can be inherited or acquired, are a leading cause of irreversible blindness worldwide. Age‐related macular degeneration is the most common acquired cause of photoreceptor degeneration and a major public health concern due to the aging population. Retinitis pigmentosa is the most common inherited retinal degenerative disease, which is a heterogeneous group of sight‐threatening disorders. These diseases share photoreceptor degeneration as a common feature, typically due to genetic defects affecting the photoreceptors or the retinal pigment epithelium but can also be influenced by environmental factors. Additionally, inflammation and oxidative stress contribute to their development and progression. These diseases represent a major economic burden to healthcare systems and society. Therefore, it is imperative to develop preventive and/or therapeutic strategies that can impede or halt their progression.Taurine supplementation is a promising therapeutic approach for these diseases due to its antioxidant, anti‐apoptotic, and anti‐inflammatory properties. Our group has investigated the effects of taurine supplementation in the Royal College of Surgeons (RCS) rat, an established model of inherited retinal degeneration that manifests an impaired phagocytic capacity of the retinal pigment epithelium due to a MERKT gene mutation. Our findings demonstrate that taurine supplementation improves functional photoreceptor survival, decreases glial activation (including decreased GFAP expression in Müller cells and inhibition of microglial activation and migration), reduces oxidative stress in the outer and inner nuclear layers, suppresses apoptosis, and preserves synaptic connections in the RCS rat retina. Therefore, taurine is essential for retinal health and may provide neuroprotection against retinal degeneration thanks to its multifaceted properties.

Lithium Chloride Exerts Anti-Inflammatory and Neuroprotective Effects by Inhibiting Microglial Activation in LPS-Induced Retinal Injury.

To explore the anti-inflammatory and neuroprotective effects of lithium chloride (LiCl) in LPS-induced retinal injury. In vitro, primary retinal microglia were pretreated with LiCl and stimulated with lipopolysaccharide (LPS). Pro-inflammatory cytokine production, microglial morphological changes, and inflammation-associated signaling pathways were measured by real-time PCR (RT-PCR), western blotting, and immunofluorescence. Primary retinal neurons were cultured with microglial-derived conditioned medium in the absence or presence of LiCl. Neurotoxicity was evaluated by Cell Counting Kit-8 (CCK-8), terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay, and γ-H2AX detection. In vivo, an endotoxin-induced uveitis mice model was established, and each animal was given intraperitoneal injection of LiCl or vehicle. The retinal inflammatory response was measured by hematoxylin and eosin and fluorescent staining, RT-PCR, western blotting, and TUNEL assay. Retinal thickness and function were evaluated by spectral-domain optical coherence tomography and electroretinography. In vitro, LiCl exerted no obvious toxic effects on microglia and significantly decreased proinflammatory factor (inducible nitric oxide synthase, tumor necrosis factor α, interleukin 6) production, inhibited microglial activation in morphology, and suppressed nuclear factor kappa B (NF-κB), Akt, and phosphatidylinositol 3-kinase (PI3K) phosphorylation. Moreover, LiCl promoted retinal neuron survival and reduced cell apoptosis and the expression of γ-H2AX. In vivo, LiCl reduced inflammatory infiltrating cells in the vitreous cavity and decreased proinflammatory cytokine expression in retinas. LiCl suppressed LPS-induced microglial activation, proliferation, and migration. Additionally, LiCl reduced LPS-induced apoptosis of ganglion cells and retinal edema and rescued retinal functional damage. This study demonstrates that LiCl exerts anti-inflammatory and neuroprotective effects by inhibiting microglial activation via the PI3K/Akt/NF-κB pathway in LPS-induced retinal injury. LiCl provides a novel and promising option to treat retinal inflammatory diseases.

The novel visual cycle inhibitor (±)-RPE65-61 protects retinal photoreceptors from light-induced degeneration.

The visual cycle refers to a series of biochemical reactions of retinoids in ocular tissues and supports the vision in vertebrates. The visual cycle regenerates visual pigments chromophore, 11-cis-retinal, and eliminates its toxic byproducts from the retina, supporting visual function and retinal neuron survival. Unfortunately, during the visual cycle, when 11-cis-retinal is being regenerated in the retina, toxic byproducts, such as all-trans-retinal and bis-retinoid is N-retinylidene-N-retinylethanolamine (A2E), are produced, which are proposed to contribute to the pathogenesis of the dry form of age-related macular degeneration (AMD). The primary biochemical defect in Stargardt disease (STGD1) is the accelerated synthesis of cytotoxic lipofuscin bisretinoids, such as A2E, in the retinal pigment epithelium (RPE) due to mutations in the ABCA4 gene. To prevent all-trans-retinal—and bisretinoid-mediated retinal degeneration, slowing down the retinoid flow by modulating the visual cycle with a small molecule has been proposed as a therapeutic strategy. The present study describes RPE65-61, a novel, non-retinoid compound, as an inhibitor of RPE65 (a key enzyme in the visual cycle), intended to modulate the excessive activity of the visual cycle to protect the retina from harm degenerative diseases. Our data demonstrated that (±)-RPE65-61 selectively inhibited retinoid isomerase activity of RPE65, with an IC50 of 80 nM. Furthermore, (±)-RPE65-61 inhibited RPE65 via an uncompetitive mechanism. Systemic administration of (±)-RPE65-61 in mice resulted in slower chromophore regeneration after light bleach, confirming in vivo target engagement and visual cycle modulation. Concomitant protection of the mouse retina from high-intensity light damage was also observed. Furthermore, RPE65-61 down-regulated the cyclic GMP-AMP synthase stimulator of interferon genes (cGAS-STING) pathway, decreased the inflammatory factor, and attenuated retinal apoptosis caused by light-induced retinal damage (LIRD), which led to the preservation of the retinal function. Taken together, (±)-RPE65-61 is a potent visual cycle modulator that may provide a neuroprotective therapeutic benefit for patients with STGD and AMD.

Curcumin protects retinal neuronal cells against oxidative stress-induced damage by regulating mitochondrial dynamics

Oxidative stress plays a crucial role in the damage of retinal neuronal cells. Curcumin, the phytocompound, has anti-inflammatory and antioxidative properties. It was shown that curcumin exerted a beneficial effect on retinal neuronal cell survival. However, the role of mitochondrial dynamics in curcumin-mediated protective effect on retinal neuronal cells remains to be elucidated. Here, H2O2 was used to mimic the oxidative stress in retinal neuronal R28 cells. Drp1 and Mfn2 are key regulators of mitochondrial fission and fusion. 100 μM of H2O2 significantly increased the cleavage of caspase-3 and Drp1 expression, but downregulated the expression of Mfn2. Pretreatment with 5 μM curcumin effectively alleviated H2O2-induced alterations in the expression of Drp1 and Mfn2 and mitochondrial fission in R28 cells. In addition, curcumin and Drp1 knockdown prevented H2O2-induced intracellular ROS increment and mitochondrial membrane potential disruption. On the contrary, knockdown of Mfn2 diminished curcumin-mediated protection against ROS increment and mitochondrial membrane potential disruption after H2O2. Moreover, curcumin protected R28 cells against H2O2-induced PINK1 expression, mitophagy, caspase-3 cleavage and apoptosis. Knockdown of Mfn2 significantly alleviated the protective effect of curcumin on R28 cells after H2O2. Taken together, our data indicate that curcumin protects against oxidative stress-induced injury in retinal neuronal cells by promoting mitochondrial fusion.

High-Contrast Stimulation Potentiates the Neurotrophic Properties of Müller Cells and Suppresses Their Pro-Inflammatory Phenotype.

High-contrast visual stimulation promotes retinal regeneration and visual function, but the underlying mechanism is not fully understood. Here, we hypothesized that Müller cells (MCs), which express neurotrophins such as brain-derived neurotrophic factor (BDNF), could be key players in this retinal plasticity process. This hypothesis was tested by conducting in vivo and in vitro high-contrast stimulation of adult mice and MCs. Following stimulation, we examined the expression of BDNF and its inducible factor, VGF, in the retina and MCs. We also investigated the alterations in the expression of VGF, nuclear factor kappa B (NF-κB) and pro-inflammatory mediators in MCs, as well as their capacity to proliferate and develop a neurogenic or reactive gliosis phenotype after high-contrast stimulation and treatment with BDNF. Our results showed that high-contrast stimulation upregulated BDNF levels in MCs in vivo and in vitro. The additional BDNF treatment significantly augmented VGF production in MCs and their neuroprotective features, as evidenced by increased MC proliferation, neurodifferentiation, and decreased expression of the pro-inflammatory factors and the reactive gliosis marker GFAP. These results demonstrate that high-contrast stimulation activates the neurotrophic and neuroprotective properties of MCs, suggesting their possible direct involvement in retinal neuronal survival and improved functional outcomes in response to visual stimulation.

B cell-dependent EAE induces visual deficits in the mouse with similarities to human autoimmune demyelinating diseases

BackgroundIn the field of autoimmune demyelinating diseases, visual impairments have extensively been studied using the experimental autoimmune encephalomyelitis (EAE) mouse model, which is classically induced by immunization with myelin oligodendrocyte glycoprotein peptide (MOG35–55). However, this model does not involve B cells like its human analogs. New antigens have thus been developed to induce a B cell-dependent form of EAE that better mimics human diseases.MethodsThe present study aimed to characterize the visual symptoms of EAE induced with such an antigen called bMOG. After the induction of EAE with bMOG in C57BL/6J mice, visual function changes were studied by electroretinography and optomotor acuity tests. Motor deficits were assessed in parallel with a standard clinical scoring method. Histological examinations and Western blot analyses allowed to follow retinal neuron survival, gliosis, microglia activation, opsin photopigment expression in photoreceptors and optic nerve demyelination. Disease effects on retinal gene expression were established by RNA sequencing.ResultsWe observed that bMOG EAE mice exhibited persistent loss of visual acuity, despite partial recovery of electroretinogram and motor functions. This loss was likely due to retinal inflammation, gliosis and synaptic impairments, as evidenced by histological and transcriptomic data. Further analysis suggests that the M-cone photoreceptor pathway was also affected.ConclusionTherefore, by documenting visual changes induced by bMOG and showing similarities to those seen in diseases such as multiple sclerosis and neuromyelitis optica, this study offers a new approach to test protective or restorative ophthalmic treatments.

Nuclear NAD+-biosynthetic enzyme NMNAT1 facilitates development and early survival of retinal neurons.

Despite mounting evidence that the mammalian retina is exceptionally reliant on proper NAD+ homeostasis for health and function, the specific roles of subcellular NAD+ pools in retinal development, maintenance, and disease remain obscure. Here, we show that deletion of the nuclear-localized NAD+ synthase nicotinamide mononucleotide adenylyltransferase-1 (NMNAT1) in the developing murine retina causes early and severe degeneration of photoreceptors and select inner retinal neurons via multiple distinct cell death pathways. This severe phenotype is associated with disruptions to retinal central carbon metabolism, purine nucleotide synthesis, and amino acid pathways. Furthermore, transcriptomic and immunostaining approaches reveal dysregulation of a collection of photoreceptor and synapse-specific genes in NMNAT1 knockout retinas prior to detectable morphological or metabolic alterations. Collectively, our study reveals previously unrecognized complexity in NMNAT1-associated retinal degeneration and suggests a yet-undescribed role for NMNAT1 in gene regulation during photoreceptor terminal differentiation.

Dpp/TGFβ-superfamily play a dual conserved role in mediating the damage response in the retina.

Retinal homeostasis relies on intricate coordination of cell death and survival in response to stress and damage. Signaling mechanisms that coordinate this process in the adult retina remain poorly understood. Here we identify Decapentaplegic (Dpp) signaling in Drosophila and its mammalian homologue Transforming Growth Factor-beta (TGFβ) superfamily, that includes TGFβ and Bone Morphogenetic Protein (BMP) signaling arms, as central mediators of retinal neuronal death and tissue survival following acute damage. Using a Drosophila model for UV-induced retinal damage, we show that Dpp released from immune cells promotes tissue loss after UV-induced retinal damage. Interestingly, we find a dynamic response of retinal cells to this signal: in an early phase, Dpp-mediated stimulation of Saxophone/Smox signaling promotes apoptosis, while at a later stage, stimulation of the Thickveins/Mad axis promotes tissue repair and survival. This dual role is conserved in the mammalian retina through the TGFβ/BMP signaling, as supplementation of BMP4 or inhibition of TGFβ using small molecules promotes retinal cell survival, while inhibition of BMP negatively affects cell survival after light-induced photoreceptor damage and NMDA induced inner retinal neuronal damage. Our data identify key evolutionarily conserved mechanisms by which retinal homeostasis is maintained.

Implantation and long-term assessment of the stability and biocompatibility of a novel 98 channel suprachoroidal visual prosthesis in sheep

Severe visual impairment can result from retinal degenerative diseases such as retinitis pigmentosa, which lead to photoreceptor cell death. These pathologies result in extensive neural and glial remodelling, with survival of excitable retinal neurons that can be electrically stimulated to elicit visual percepts and restore a form of useful vision. The Phoenix99 Bionic Eye is a fully implantable visual prosthesis, designed to stimulate the retina from the suprachoroidal space. In the current study, nine passive devices were implanted in an ovine model from two days to three months. The impact of the intervention and implant stability were assessed using indirect ophthalmoscopy, infrared imaging, and optical coherence tomography to establish the safety profile of the surgery and the device. The biocompatibility of the device was evaluated using histopathological analysis of the tissue surrounding the electrode array, with a focus on the health of the retinal cells required to convey signals to the brain. Appropriate stability of the electrode array was demonstrated, and histological analysis shows that the fibrotic and inflammatory response to the array was mild. Promising evidence of the safety and potential of the Phoenix99 Bionic Eye to restore a sense of vision to the severely visually impaired was obtained.

Plasma Rich in Growth Factors (PRGF) Increases the Number of Retinal Müller Glia in Culture but Not the Survival of Retinal Neurons.

Plasma rich in growth factors (PRGF) is a subtype of platelet-rich plasma (PRP) that stimulates tissue regeneration and may promote neuronal survival. It has been employed in ophthalmology to achieve tissue repair in some retinal pathologies, although how PRGF acts in the retina is still poorly understood. As a part of the central nervous system, the retina has limited capacity for repair capacity following damage, and retinal insult can provoke the death of retinal ganglion cells (RGCs), potentially producing irreversible blindness. RGCs are in close contact with glial cells, such as Müller cells, that help maintain homeostasis in the retina. In this study, the aim was to determine whether PRGF can protect RGCs and whether it increases the number of Müller cells. Therefore, PRGF were tested on primary cell cultures of porcine RGCs and Müller cells, as well as on co-cultures of these two cell types. Moreover, the inflammatory component of PRGF was analyzed and the cytokines in the different PRGFs were quantified. In addition, we set out to determine if blocking the inflammatory components of PRGF alters its effect on the cells in culture. The presence of PRGF compromises RGC survival in pure cultures and in co-culture with Müller cells, but this effect was reversed by heat-inactivation of the PRGF. The detrimental effect of PRGF on RGCs could be in part due to the presence of cytokines and specifically, to the presence of pro-inflammatory cytokines that compromise their survival. However, other factors are likely to be present in the PRGF that have a deleterious effect on the RGCs since the exposure to antibodies against these cytokines were insufficient to protect RGCs. Moreover, PRGF promotes Müller cell survival. In conclusion, PRGF hinders the survival of RGCs in the presence or absence of Müller cells, yet it promotes Müller cell survival that could be the reason of retina healing observed in the in vivo treatments, with some cytokines possibly implicated. Although PRGF could stimulate tissue regeneration, further studies should be performed to evaluate the effect of PRGF on neurons and the implication of its potential inflammatory role in such processes.

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.

Tau Gene Deletion does not Influence Axonal Regeneration and Retinal Neuron Survival in the Injured Mouse Visual System.

In the present study, we hypothesized that the microtubule-associated protein Tau may influence retinal neuron survival and axonal regeneration after optic nerve injury. To test this hypothesis, the density of retinal ganglion cells was evaluated by immunostaining retinal flat-mounts for RNA-binding protein with multiple splicing (RBPMS) two weeks after optic nerve micro-crush lesion in Tau-deprived (Tau knock-out (KO)) and wild-type (WT) mice. Axon growth was determined on longitudinal sections of optic nerves after anterograde tracing. Our results showed that the number of surviving retinal ganglion cells and growing axons did not significantly vary between WT and Tau KO animals. Moreover, sustained activation of the neuronal growth program with ciliary neurotrophic factor (CNTF) resulted in a similar increase in surviving neurons and in growing axons in WT and Tau KO mice. Taken together, our data suggest that Tau does not influence axonal regeneration or neuronal survival.

Loss of XBP1 Leads to Early-Onset Retinal Neurodegeneration in a Mouse Model of Type I Diabetes.

Retinal neuronal injury and degeneration is one of the primary manifestations of diabetic retinopathy, a leading cause of vision loss in working age adults. In pathological conditions, including diabetes and some physiological conditions such as aging, protein homeostasis can become disrupted, leading to endoplasmic reticulum (ER) stress. Severe or unmitigated ER stress can lead to cell death, which in retinal neurons results in irreversible loss of visual function. X-box binding protein 1 (XBP1) is a major transcription factor responsible for the adaptive unfolded protein response (UPR) to maintain protein homeostasis in cells undergoing ER stress. The purpose of this study is to determine the role of XBP1-mediated UPR in retinal neuronal survival and function in a mouse model of type 1 diabetes. Using a conditional retina-specific XBP1 knockout mouse line, we demonstrate that depletion of XBP1 in retinal neurons results in early onset retinal function decline, loss of retinal ganglion cells and photoreceptors, disrupted photoreceptor ribbon synapses, and Müller cell activation after induction of diabetes. Our findings suggest an important role of XBP1-mediated adaptive UPR in retinal neuronal survival and function in diabetes.

Reactive microglia and IL1\u03b2/IL-1R1-signaling mediate neuroprotection in excitotoxin-damaged mouse retina

BackgroundMicroglia and inflammation have context-specific impacts upon neuronal survival in different models of central nervous system (CNS) disease. Herein, we investigate how inflammatory mediators, including microglia, interleukin 1 beta (IL1β), and signaling through interleukin 1 receptor type 1 (IL-1R1), influence the survival of retinal neurons in response to excitotoxic damage.MethodsExcitotoxic retinal damage was induced via intraocular injections of NMDA. Microglial phenotype and neuronal survival were assessed by immunohistochemistry. Single-cell RNA sequencing was performed to obtain transcriptomic profiles. Microglia were ablated by using clodronate liposome or PLX5622. Retinas were treated with IL1β prior to NMDA damage and cell death was assessed in wild type, IL-1R1 null mice, and mice expressing IL-1R1 only in astrocytes.ResultsNMDA-induced damage included neuronal cell death, microglial reactivity, upregulation of pro-inflammatory cytokines, and genes associated with IL1β-signaling in different types of retinal neurons and glia. Expression of the IL1β receptor, IL-1R1, was evident in astrocytes, endothelial cells, some Müller glia, and OFF bipolar cells. Ablation of microglia with clodronate liposomes or Csf1r antagonist (PLX5622) resulted in elevated cell death and diminished neuronal survival in excitotoxin-damaged retinas. Exogenous IL1β stimulated the proliferation and reactivity of microglia in the absence of damage, reduced numbers of dying cells in damaged retinas, and increased neuronal survival following an insult. IL1β failed to provide neuroprotection in the IL-1R1-null retina, but IL1β-mediated neuroprotection was rescued when expression of IL-1R1 was restored in astrocytes.ConclusionsWe conclude that reactive microglia provide protection to retinal neurons, since the absence of microglia is detrimental to survival. We propose that, at least in part, the survival-influencing effects of microglia may be mediated by IL1β, IL-1R1, and interactions of microglia and other macroglia.

The M1 muscarinic acetylcholine receptor subtype is important for retinal neuron survival in aging mice

Muscarinic acetylcholine receptors have been implicated as potential neuroprotective targets for glaucoma. We tested the hypothesis that the lack of a single muscarinic receptor subtype leads to age-dependent neuron reduction in the retinal ganglion cell layer. Mice with targeted disruption of single muscarinic acetylcholine receptor subtype genes (M1 to M5) and wild-type controls were examined at two age categories, 5 and 15 months, respectively. We found no differences in intraocular pressure between individual mouse groups. Remarkably, in 15-month-old mice devoid of the M1 receptor, neuron number in the retinal ganglion cell layer and axon number in the optic nerve were markedly reduced. Moreover, mRNA expression for the prooxidative enzyme, NOX2, was increased, while mRNA expression for the antioxidative enzymes, SOD1, GPx1 and HO-1, was reduced in aged M1 receptor-deficient mice compared to age-matched wild-type mice. In line with these findings, the reactive oxygen species level was also elevated in the retinal ganglion cell layer of aged M1 receptor-deficient mice. In conclusion, M1 receptor deficiency results in retinal ganglion cell loss in aged mice via involvement of oxidative stress. Based on these findings, activation of M1 receptor signaling may become therapeutically useful to promote retinal ganglion cell survival.

The agonistic TSPO ligand XBD173 attenuates the glial response thereby protecting inner retinal neurons in a murine model of retinal ischemia

BackgroundLigand-driven modulation of the mitochondrial translocator protein 18 kDa (TSPO) was recently described to dampen the neuroinflammatory response of microglia in a retinal light damage model resulting in protective effects on photoreceptors. We characterized the effects of the TSPO ligand XBD173 in the postischemic retina focusing on changes in the response pattern of the major glial cell types of the retina—microglia and Müller cells.MethodsRetinal ischemia was induced by increasing the intraocular pressure for 60 min followed by reperfusion of the tissue in mice. On retinal cell types enriched via immunomagnetic separation expression analysis of TSPO, its ligand diazepam-binding inhibitor (DBI) and markers of glial activation were performed at transcript and protein level using RNA sequencing, qRT-PCR, lipid chromatography-mass spectrometry, and immunofluorescent labeling. Data on cell morphology and numbers were assessed in retinal slice and flatmount preparations. The retinal functional integrity was determined by electroretinogram recordings.ResultsWe demonstrate that TSPO is expressed by Müller cells, microglia, vascular cells, retinal pigment epithelium (RPE) of the healthy and postischemic retina, but only at low levels in retinal neurons. While an alleviated neurodegeneration upon XBD173 treatment was found in postischemic retinae as compared to vehicle controls, this neuroprotective effect of XBD173 is mediated putatively by its action on retinal glia. After transient ischemia, TSPO as a marker of activation was upregulated to similar levels in microglia as compared to their counterparts in healthy retinae irrespective of the treatment regimen. However, less microglia were found in XBD173-treated postischemic retinae at 3 days post-surgery (dps) which displayed a more ramified morphology than in retinae of vehicle-treated mice indicating a dampened microglia activation. Müller cells, the major retinal macroglia, show upregulation of the typical gliosis marker GFAP. Importantly, glutamine synthetase was more stably expressed in Müller glia of XBD173-treated postischemic retinae and homeostatic functions such as cellular volume regulation typically diminished in gliotic Müller cells remained functional.ConclusionsIn sum, our data imply that beneficial effects of XBD173 treatment on the postischemic survival of inner retinal neurons were primarily mediated by stabilizing neurosupportive functions of glial cells.