IC100, a humanized therapeutic monoclonal anti-ASC antibody alleviates oxygen-induced retinopathy in mice.

BACKGROUND: Retinopathy of prematurity (ROP), which often presents with bronchopulmonary dysplasia (BPD), is among the most common morbidities affecting extremely premature infants and is a leading cause of severe vision impairment in children worldwide. Activations of the inflammasome cascade and microglia have been implicated in playing a role in the development of both ROP and BPD. Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) is pivotal in inflammasome assembly. Utilizing mouse models of both oxygen-induced retinopathy (OIR) and BPD, this study was designed to test the hypothesis that hyperoxia induces ASC speck formation, which leads to microglial activation and retinopathy, and that inhibition of ASC speck formation by a humanized monoclonal antibody, IC100, directed against ASC, will ameliorate microglial activation and abnormal retinal vascular formation. METHODS: We first tested ASC speck formation in the retina of ASC-citrine reporter mice expressing ASC fusion protein with a C-terminal citrine (fluorescent GFP isoform) using a BPD model that causes both lung and eye injury by exposing newborn mice to room air (RA) or 85% O2 from postnatal day (P) 1 to P14. The retinas were dissected on P14 and retinal flat mounts were used to detect vascular endothelium with AF-594-conjugated isolectin B4 (IB4) and citrine-tagged ASC specks. To assess the effects of IC100 on an OIR model, newborn ASC citrine reporter mice and wildtype mice (C57BL/6 J) were exposed to RA from P1 to P6, then 75% O2 from P7 to P11, and then to RA from P12 to P18. At P12 mice were randomized to the following groups: RA with placebo PBS (RA-PBS), O2 with PBS (O2-PBS), O2 + IC100 intravitreal injection (O2-IC100-IVT), and O2 + IC100 intraperitoneal injection (O2-IC100-IP). Retinal vascularization was evaluated by flat mount staining with IB4. Microglial activation was detected by immunofluorescence staining for allograft inflammatory factor 1 (AIF-1) and CD206. Retinal structure was analyzed on H&E-stained sections, and function was analyzed by pattern electroretinography (PERG). RNA-sequencing (RNA-seq) of the retinas was performed to determine the transcriptional effects of IC100 treatment in OIR. RESULTS: ASC specks were significantly increased in the retinas by hyperoxia exposure and colocalized with the abnormal vasculature in both BPD and OIR models, and this was associated with increased microglial activation. Treatment with IC100-IVT or IC100-IP significantly reduced vaso-obliteration and intravitreal neovascularization. IC100-IVT treatment also reduced retinal microglial activation, restored retinal structure, and improved retinal function. RNA-seq showed that IC100 treatment corrected the induction of genes associated with angiogenesis, leukocyte migration, and VEGF signaling caused by O2. IC100 also corrected the suppression of genes associated with cell junction assembly, neuron projection, and neuron recognition caused by O2. CONCLUSION: These data demonstrate the crucial role of ASC in the pathogenesis of OIR and the efficacy of a humanized therapeutic anti-ASC antibody in treating OIR mice. Thus, this anti-ASC antibody may potentially be considered in diseases associated with oxygen stresses and retinopathy, such as ROP.

Leber Hereditary Optic Neuropathy Gene Therapy: Longitudinal Relationships Among Visual Function and Anatomical Measures.

PURPOSE: To assess longitudinal relationships among visual function and anatomical measures of gene therapy in G11778A Leber hereditary optic neuropathy (LHON). DESIGN: Phase 1 clinical trial. METHODS: This was a single-institution study of patients with G11778A LHON. Patients with chronic bilateral visual loss >12 months (group 1, n = 11), acute bilateral visual loss <12 months (group 2, n = 9), or unilateral visual loss (group 3, n = 8) were administered unilateral intravitreal AAV2(Y444,500,730F)-P1ND4v2 injection with low, medium, high, and higher doses to worse eye for groups 1 and 2 and better eye for group 3. Oucome measures were best-corrected visual acuity (BCVA), visual field mean deviation (VF MD), steady-state pattern electroretinogram (SS-PERG), optical coherence tomography (OCT) retinal nerve fiber layer (RNFL) thickness and ganglion cell+inner plexiform layer (GCIPL) thickness, and National Eye Institute Visual Function Questionnaire (NEI-VFQ-25) scores. Mean follow-up was 33.6 months (range = 18-36 months). RESULTS: Baseline SS-PERG amplitude was much reduced in both eyes of all groups including asymptomatic eyes of group 3, and showed no appreciable changes irrespective of disease stage and treatment. Significant and progressive GCIPL and RNFL thinning occurred in all eyes; BCVA and VF MD fluctuated in treated and fellow eyes, with some eyes having modest improvement that may be related to natural history or to gene therapy. Mean NEI-VFQ-25 scores declined in group 3 subjects (P = .023), CONCLUSION: Asymptomatic eyes in LHON patients with unilateral visual loss may be beyond the window of effective neuroprotection given reduced GCIPL and SS-PERG. Randomization of patients to an untreated control group would help to assess treatment effect by accounting for variable natural history. NOTE: Publication of this article is sponsored by the American Ophthalmological Society.

Mitochondrially Targeted Gene Therapy Rescues Visual Loss in a Mouse Model of Leber's Hereditary Optic Neuropathy.

Leber's hereditary optic neuropathy (LHON) is a common mitochondrial genetic disease, causing irreversible blindness in young individuals. Current treatments are inadequate, and there is no definitive cure. This study evaluates the effectiveness of delivering wildtype human NADH ubiquinone oxidoreductase subunit 4 (hND4) gene using mito-targeted AAV(MTSAAV) to rescue LHOH mice. We observed a declining pattern in electroretinograms amplitudes as mice aged across all groups (p < 0.001), with significant differences among groups (p = 0.023; Control vs. LHON, p = 0.008; Control vs. Rescue, p = 0.228). Inner retinal thickness and intraocular pressure did not change significantly with age or groups. Compared to LHON mice, those rescued with wildtype hND4 exhibited improved retinal visual acuity (0.29 ± 0.1 cy/deg vs. 0.15 ± 0.1 cy/deg) and increased functional hyperemia response (effect of flicker, p < 0.001, effect of Group, p = 0.004; Interaction Flicker × Group, p < 0.001). Postmortem analysis shows a marked reduction in retinal ganglion cell density in the LHON group compared to the other groups (Effect of Group, p < 0.001, Control vs. LHON, p < 0.001, Control vs. Rescue, p = 0.106). These results suggest that MTSAAV-delivered wildtype hND4 gene rescues, at least in part, visual impairment in an LHON mouse model and has the therapeutic potential to treat this disease.

The Inflammasome-Dependent Dysfunction and Death of Retinal Ganglion Cells after Repetitive Intraocular Pressure Spikes.

The dysfunction and selective loss of retinal ganglion cells (RGCs) is a known cause of vision loss in glaucoma and other neuropathies, where ocular hypertension (OHT) is the major risk factor. We investigated the impact of transient non-ischemic OHT spikes (spOHT) on RGC function and viability in vivo to identify cellular pathways linking low-grade repetitive mechanical stress to RGC pathology. We found that repetitive spOHT had an unexpectedly high impact on intraocular homeostasis and RGC viability, while exposure to steady OHT (stOHT) of a similar intensity and duration failed to induce pathology. The repetitive spOHT induced the rapid activation of the inflammasome, marked by the upregulation of NLRP1, NLRP3, AIM2, caspases -1, -3/7, -8, and Gasdermin D (GSDMD), and the release of interleukin-1ß (IL-1ß) and other cytokines into the vitreous. Similar effects were also detected after 5 weeks of exposure to chronic OHT in an induced glaucoma model. The onset of these immune responses in both spOHT and glaucoma models preceded a 50% deficit in pattern electroretinogram (PERG) amplitude and a significant loss of RGCs 7 days post-injury. The inactivation of inflammasome complexes in Nlrp1-/-, Casp1-/-, and GsdmD-/- knockout animals significantly suppressed the spOHT-induced inflammatory response and protected RGCs. Our results demonstrate that mechanical stress produced by acute repetitive spOHT or chronic OHT is mechanistically linked to inflammasome activation, which leads to RGC dysfunction and death.

High-Throughput Binocular Pattern Electroretinograms in the Mouse.

The pattern electroretinogram (PERG) reflects the electrical activity of retinal ganglion cells (RGC) and has become the most used technology to assess RGC function in experimental models of glaucoma and optic neuropathies. We describe a novel method for obtaining user-friendly, robust PERG simultaneously from each eye using asynchronous binocular stimulation and one-channel acquisition of signals recorded from a subcutaneous needle in the snout.

Pattern Electroretinogram in Ocular Hypertension, Glaucoma Suspect and Early Manifest Glaucoma Eyes: A Systematic Review and Meta-analysis.

Topic: To provide standardized confidence limits of the transient pattern electroretinogram (tPERG) P50 and N95 and steady state pattern electroretinogram (ssPERG) amplitudes in normal controls as compared to ocular hypertension (OHT), glaucoma suspect (GS), or early manifest glaucoma (EMG) eyes. Clinical Relevance: The identification of standardized confidence limits in the context of pattern electroretinogram (PERG) might overcome the high intrinsic variability of the measure, and it might lead to a more intuitive understanding of the results as well as to an easier comparison of data from multiple tests, sites, and operators. Methods: The study protocol was prospectively registered on the International Prospective Register of Systematic Reviews (ID: CRD42022370032). A literature search was conducted on PubMed, Web of Science, and Scopus. Studies comparing PERG raw data in normal control eyes as compared to OHT, GS, or EMG were included. The risk of bias was assessed using the National Institute for Health and Clinical Excellence quality assessment tool. The main outcome was the P50, N95, and ssPERG amplitude difference between the control and the study groups' eyes. The standardized mean difference was calculated as a measure of the effect size for the primary outcome. A subanalysis was conducted based on the type of electrodes adopted for the PERG measurements (invasive vs. noninvasive). Results: Of the 4580 eligible papers, only 23 were included (1754 eyes). Statistically significant amplitude differences were found in the P50, N95, and ssPERG amplitudes between normal controls and OHT, GS, and EMG eyes. The highest standardized mean difference values were observed in the ssPERG amplitude in all 3 sets of comparison. The subanalysis did not reveal any statistically significant differences between invasive and noninvasive recording strategies. Conclusions: The use of standardized values as the main outcome measures in the context of the PERG data analysis is a valid approach, normalizing several confounding factors which have affected the clinical utility of PERG both for individual patients and in clinical trials. Steady state PERG apparently better discriminates diseased eyes compared to tPERG. The adoption of skin-active electrodes is able to adequately discriminate between healthy and diseased statuses. Financial Disclosures: Proprietary or commercial disclosure may be found after the references.

Natural History of Glaucoma Progression in the DBA/2J Model: Early Contribution of Müller Cell Gliosis.

Glaucoma is a chronic optic neuropathy characterized by progressive degeneration of retinal ganglion cells (RGCs). Elevated intraocular pressure (IOP) and the resulting mechanical stress are classically considered the main causes of RGC death. However, RGC degeneration and ensuing vision loss often occur independent of IOP, indicating a multifactorial nature of glaucoma, with the likely contribution of glial and vascular function. The aim of the present study was to provide a comprehensive evaluation of the time course of neuro-glial-vascular changes associated with glaucoma progression. We used DBA/2J mice in the age range of 2-15 months as a spontaneous model of glaucoma with progressive IOP elevation and RGC loss typical of human open-angle glaucoma. We found that the onset of RGC degeneration at 10 months of age coincided with that of IOP elevation and vascular changes such as decreased density, increased lacunarity and decreased tight-junction protein zonula occludens (ZO)-1, while hypoxia-inducible factor (HIF)-1α and vascular endothelial growth factor (VEGF) were already significantly upregulated at 6 months of age together with the onset of Müller cell gliosis. Astrocytes, however, underwent significant gliosis at 10 months. These results indicate that Müller cell activation occurs well before IOP elevation, with probable inflammatory consequences, and represents an early event in the glaucomatous process. Early upregulation of HIF-1α and VEGF is likely to contribute to blood retinal barrier failure, facilitating RGC loss. The different time courses of neuro-glial-vascular changes during glaucoma progression provide further insight into the nature of the disease and suggest potential targets for the development of efficient therapeutic intervention aside from IOP lowering.

Comparison of different gene-therapy methods to treat Leber hereditary optic neuropathy in a mouse model.

Introduction: Therapies for Leber hereditary optic neuropathy (LHON), in common with all disorders caused by mutated mitochondrial DNA, are inadequate. We have developed two gene therapy strategies for the disease: mitochondrial-targeted and allotopic expressed and compared them in a mouse model of LHON. Methods: A LHON mouse model was generated by intravitreal injection of a mitochondrialtargeted Adeno-associated virus (AAV) carrying mutant human NADH dehydrogenase 4 gene (hND4/m.11778G>A) to induce retinal ganglion cell (RGC) degeneration and axon loss, the hallmark of the human disease. We then attempted to rescue those mice using a second intravitreal injection of either mitochondrial-targeted or allotopic expressed wildtype human ND4. The rescue of RGCs and their axons were assessed using serial pattern electroretinogram (PERG) and transmission electron microscopy. Results: Compared to non-rescued LHON controls where PERG amplitude was much reduced, both strategies significantly preserved PERG amplitude over 15 months. However, the rescue effect was more marked with mitochondrial-targeted therapy than with allotopic therapy (p = 0.0128). Post-mortem analysis showed that mitochondrial-targeted human ND4 better preserved small axons that are preferentially lost in human LHON. Conclusions: These results in a pre-clinical mouse model of LHON suggest that mitochondrially-targeted AAV gene therapy, compared to allotopic AAV gene therapy, is more efficient in rescuing the LHON phenotype.

σ2R/TMEM97 in retinal ganglion cell degeneration.

The sigma 2 receptor (σ2R) was recently identified as an endoplasmic reticulum (ER) membrane protein known as transmembrane protein 97 (TMEM97). Studies have shown that σ2R/TMEM97 binding compounds are neuroprotective, suggesting a role of σ2R/TMEM97 in neurodegenerative processes. To understand the function of σ2R/TMEM97 in neurodegeneration pathways, we characterized ischemia-induced retinal ganglion cell (RGC) degeneration in TMEM97-/- mice and found that RGCs in TMEM97-/- mice are resistant to degeneration. In addition, intravitreal injection of a selective σ2R/TMEM97 ligand DKR-1677 significantly protects RGCs from ischemia-induced degeneration in wildtype mice. Our results provide conclusive evidence that σ2R/TMEM97 plays a role to facilitate RGC death following ischemic injury and that inhibiting the function of σ2R/TMEM97 is neuroprotective. This work is a breakthrough toward elucidating the biology and function of σ2R/TMEM97 in RGCs and likely in other σ2R/TMEM97 expressing neurons. Moreover, these findings support future studies to develop new neuroprotective approaches for RGC degenerative diseases by inhibiting σ2R/TMEM97.

Using Noninvasive Electrophysiology to Determine Time Windows of Neuroprotection in Optic Neuropathies.

The goal of neuroprotection in optic neuropathies is to prevent loss of retinal ganglion cells (RGCs) and spare their function. The ideal time window for initiating neuroprotective treatments should be the preclinical period at which RGCs start losing their functional integrity before dying. Noninvasive electrophysiological tests such as the Pattern Electroretinogram (PERG) can assess the ability of RGCs to generate electrical signals under a protracted degenerative process in both clinical conditions and experimental models, which may have both diagnostic and prognostic values and provide the rationale for early treatment. The PERG can be used to longitudinally monitor the acute and chronic effects of neuroprotective treatments. User-friendly versions of the PERG technology are now commercially available for both clinical and experimental use.

Gene therapy restores mitochondrial function and protects retinal ganglion cells in optic neuropathy induced by a mito-targeted mutant ND1 gene.

Therapies for genetic disorders caused by mutated mitochondrial DNA are an unmet need, in large part due barriers in delivering DNA to the organelle and the absence of relevant animal models. We injected into mouse eyes a mitochondrially targeted Adeno-Associated-Virus (MTS-AAV) to deliver the mutant human NADH ubiquinone oxidoreductase subunit I (hND1/m.3460 G > A) responsible for Leber's hereditary optic neuropathy, the most common primary mitochondrial genetic disease. We show that the expression of the mutant hND1 delivered to retinal ganglion cells (RGC) layer colocalizes with the mitochondrial marker PORIN and the assembly of the expressed hND1 protein into host respiration complex I. The hND1-injected eyes exhibit hallmarks of the human disease with progressive loss of RGC function and number, as well as optic nerve degeneration. We also show that gene therapy in the hND1 eyes by means of an injection of a second MTS-AAV vector carrying wild-type human ND1 restores mitochondrial respiratory complex I activity, the rate of ATP synthesis and protects RGCs and their axons from dysfunction and degeneration. These results prove that MTS-AAV is a highly efficient gene delivery approach with the ability to create mito-animal models and has the therapeutic potential to treat mitochondrial genetic diseases.

Leber Hereditary Optic Neuropathy Gene Therapy: Adverse Events and Visual Acuity Results of All Patient Groups.

PURPOSE: To assess safety of gene therapy in G11778A Leber hereditary optic neuropathy (LHON). DESIGN: Phase 1 clinical trial. METHODS: Setting: single institution. PARTICIPANTS: Patients with G11778A LHON and chronic bilateral visual loss >12 months (group 1, n = 11), acute bilateral visual loss <12 months (group 2, n = 9), or unilateral visual loss (group 3, n = 8). INTERVENTION: unilateral intravitreal AAV2(Y444,500,730F)-P1ND4v2 injection with low, medium, high, and higher doses to worse eye for groups 1 and 2 and better eye for group 3. OUTCOME MEASURES: Best-corrected visual acuity (BCVA), adverse events, and vector antibody responses. Mean follow-up was 24 months (range, 12-36 months); BCVAs were compared with a published prospective natural history cohort with designated surrogate study and fellow eyes. RESULTS: Incident uveitis (8 of 28, 29%), the only vector-related adverse event, resulted in no attributable vision sequelae and was related to vector dose: 5 of 7 (71%) higher-dose eyes vs 3 of 21 (14%) low-, medium-, or high-dose eyes (P < .001). Incident uveitis requiring treatment was associated with increased serum AAV2 neutralizing antibody titers (p=0.007) but not serum AAV2 polymerase chain reaction. Improvements of ≥15-letter BCVA occurred in some treated and fellow eyes of groups 1 and 2 and some surrogate study and fellow eyes of natural history subjects. All study eyes (BCVA ≥20/40) in group 3 lost ≥15 letters within the first year despite treatment. CONCLUSIONS: G11778A LHON gene therapy has a favorable safety profile. Our results suggest that if there is an efficacy effect, it is likely small and not dose related. Demonstration of efficacy requires randomization of patients to a group not receiving vector in either eye.

The Relationship Between Stage of Leber's Hereditary Optic Neuropathy and Pattern Electroretinogram Latency.

Purpose: The purpose of this study was to compare the baseline steady-state pattern electroretinogram (SS-PERG) of patients with G11778A Leber hereditary optic neuropathy (LHON) with different stages of visual acuity (VA) loss before allotopic gene therapy (GT). Methods: Patients (n = 28) were enrolled into groups (GT I: chronic bilateral VA ≤35 Early Treatment Diabetic Retinopathy Study [ETDRS]; GT II: acute bilateral VA ≤35 ETDRS; GT III: acute unilateral, VA ≤35 ETDRS, and better eye VA ≥70 ETDRS) and tested with SS-PERG together with 210 age-matched normal controls (NCs). SS-PERG amplitude (nV) and latency (ms) of each eye were averaged for groups GT I, GT II, and NC. Symptomatic eyes (GT III-S) and asymptomatic eyes (GT III-A) of group GT III were included separately and accounted for by using generalized estimating equation (GEE) methods. Results: Compared to NC, SS-PERG amplitudes were reduced similarly by approximately 50% (P < 0.001) among all GT groups (NC > GT I, GT II, GT III-S, and GT III-A). SS-PERG latencies were shorter by ≥3.5 ms in all LHON groups and differed by disease stage (G III-A < NC, P = 0.002; GT III-S < GT III-A, P = 0.01; GT II < GT III-S, P = 0.03; GT I < NC, P < 0.001, but not different from other GT groups, all P > 0.1). Conclusions: Although SS-PERG amplitude reduction did not distinguish between disease stages, SS-PERG latency shortening occurred in asymptomatic eyes and symptomatic eyes and distinguished between disease stages. Translational Relevance: SS-PERG latency shortening is consistent with primary damage of smaller/slower axons and sparing of larger/faster axons and may provide an objective staging of LHON, which may be helpful to determine efficacy in LHON trials.

Retinal microvascular and neuronal function in patients with multiple sclerosis: 2-year follow-up.

OBJECTIVE: To determine the longitudinal changes in retinal microstructure, microvasculature, microcirculation, and axonal and neuronal functions in patients with relapsing-remitting multiple sclerosis (RRMS) over the time course of about two years. METHODS: A total of 30 patients (60 eyes) with RRMS were followed for a period of 27 ± 6 months and evaluated with a battery of clinical tests including low contrast letter acuity (LCLA), intraretinal layer thicknesses by optical coherence tomography (OCT), ganglion cell function by steady-state pattern electroretinography (PERG), axonal function by polarization-sensitive OCT, volumetric vessel density (VVD) by OCT angiography, and retinal tissue perfusion (RTP) by retinal function imager. RESULTS: Axonal function measured as retinal nerve fiber layer birefringence in the temporal quadrant and vessel density in the deep vascular plexus were significantly decreased at 2-year follow-up (P < 0.05). Subgroup analyses showed that the increased retinal blood flow volume occurred in patients with no evidence of disease activity (NEDA), and with stable or improved visual function (P < 0.05). There was no significant difference in the expanded disability state scale, LCLA, RTP, VVD, or PERG measures between the two visits (P > 0.05). CONCLUSION: To our best knowledge, this is the first 2-year prospective comprehensive study with a detailed assessment of retinal microstructure and neuronal functions in patients with RRMS. The recovery of retinal microcirculation occurred in patients with NEDA, and stable or improved visual function, suggesting these measurements as potential imaging biomarkers for monitoring disease progression.

1α,25-dihydroxyvitamin D3 protects retinal ganglion cells in glaucomatous mice.

BACKGROUND: Glaucoma is an optic neuropathy characterized by loss of function and death of retinal ganglion cells (RGCs), leading to irreversible vision loss. Neuroinflammation is recognized as one of the causes of glaucoma, and currently no treatment is addressing this mechanism. We aimed to investigate the anti-inflammatory and neuroprotective effects of 1,25(OH)2D3 (1α,25-dihydroxyvitamin D3, calcitriol), in a genetic model of age-related glaucomatous neurodegeneration (DBA/2J mice). METHODS: DBA/2J mice were randomized to 1,25(OH)2D3 or vehicle treatment groups. Pattern electroretinogram, flash electroretinogram, and intraocular pressure were recorded weekly. Immunostaining for RBPMS, Iba-1, and GFAP was carried out on retinal flat mounts to assess retinal ganglion cell density and quantify microglial and astrocyte activation, respectively. Molecular biology analyses were carried out to evaluate retinal expression of pro-inflammatory cytokines, pNFκB-p65, and neuroprotective factors. Investigators that analysed the data were blind to experimental groups, which were unveiled after graph design and statistical analysis, that were carried out with GraphPad Prism. Several statistical tests and approaches were used: the generalized estimated equations (GEE) analysis, t-test, and one-way ANOVA. RESULTS: DBA/2J mice treated with 1,25(OH)2D3 for 5 weeks showed improved PERG and FERG amplitudes and reduced RGCs death, compared to vehicle-treated age-matched controls. 1,25(OH)2D3 treatment decreased microglial and astrocyte activation, as well as expression of inflammatory cytokines and pNF-κB-p65 (p < 0.05). Moreover, 1,25(OH)2D3-treated DBA/2J mice displayed increased mRNA levels of neuroprotective factors (p < 0.05), such as BDNF. CONCLUSIONS: 1,25(OH)2D3 protected RGCs preserving retinal function, reducing inflammatory cytokines, and increasing expression of neuroprotective factors. Therefore, 1,25(OH)2D3 could attenuate the retinal damage in glaucomatous patients and warrants further clinical evaluation for the treatment of optic neuropathies.

Modeling Retinal Ganglion Cell Dysfunction in Optic Neuropathies.

As in glaucoma and other optic neuropathies cellular dysfunction often precedes cell death, the assessment of retinal ganglion cell (RGC) function represents a key outcome measure for neuroprotective strategies aimed at targeting distressed but still viable cells. RGC dysfunction can be assessed with the pattern electroretinogram (PERG), a sensitive measure of electrical activity of RGCs that is recorded non-invasively in human subjects and mouse models. Here, we offer a conceptual framework based on an intuitive state-transition model used for disease management in patients to identify progressive, potentially reversible stages of RGC dysfunction leading to cell death in mouse models of glaucoma and other optic neuropathies. We provide mathematical equations to describe state-transitions with a set of modifiable parameters that alter the time course and severity of state-transitions, which can be used for hypothesis testing and fitting experimental PERG data. PERG dynamics as a function of physiological stimuli are also used to differentiate phenotypic and altered RGC response dynamics, to assess susceptibility to stressors and to assess reversible dysfunction upon pharmacological treatment.

Diabetes Exacerbates the Intraocular Pressure-Independent Retinal Ganglion Cells Degeneration in the DBA/2J Model of Glaucoma.

Purpose: Glaucoma is a multifactorial disease, causing retinal ganglion cells (RGCs) and optic nerve degeneration. The role of diabetes as a risk factor for glaucoma has been postulated but still not unequivocally demonstrated. The purpose of this study is to clarify the effect of diabetes in the early progression of glaucomatous RGC dysfunction preceding intraocular pressure (IOP) elevation, using the DBA/2J mouse (D2) model of glaucoma. Methods: D2 mice were injected with streptozotocin (STZ) obtaining a combined model of diabetes and glaucoma (D2 + STZ). D2 and D2 + STZ mice were monitored for weight, glycemia, and IOP from 3.5 to 6 months of age. In addition, the activity of RGC and outer retina were assessed using pattern electroretinogram (PERG) and flash electroretinogram (FERG), respectively. At the end point, RGC density and astrogliosis were evaluated in flat mounted retinas. In addition, Müller cell reactivity was evaluated in retinal cross-sections. Finally, the expression of inflammation and oxidative stress markers were analyzed. Results: IOP was not influenced by time or diabetes. In contrast, RGC activity resulted progressively decreased in the D2 group independently from IOP elevation and outer retinal dysfunction. Diabetes exacerbated RGC dysfunction, which resulted independent from variation in IOP and outer retinal activity. Diabetic retinas displayed decreased RGC density and increased glial reactivity given by an increment in oxidative stress and inflammation. Conclusions: Diabetes can act as an IOP-independent risk factor for the early progression of glaucoma promoting oxidative stress and inflammation-mediated RGC dysfunction, glial reactivity, and cellular death.

Longitudinal Study of Retinal Structure, Vascular, and Neuronal Function in Patients With Relapsing-Remitting Multiple Sclerosis: 1-Year Follow-Up.

Objective: The purpose of this study was to quantify retinal structural, vascular, and functional changes in patients with relapsing-remitting multiple sclerosis (RRMS) over 1 year. Methods: Eighty-eight eyes of 44 patients with RRMS underwent assessments of low contrast letter acuity (LCLA), retinal ganglion cell function detected by the steady-state pattern electroretinogram (PERG), axonal microstructural integrity measured as birefringence, intraretinal layer thicknesses by ultra-high-resolution optical coherence tomography (OCT), volumetric vessel density (VVD) by OCT angiography, and retinal tissue perfusion (RTP) by the Retinal Function Imager (RFI). All measurements were performed at baseline and 1-year follow-up. The impacts of disease activities and a history of optic neuritis (ON) were analyzed. Results: Compared to baseline, there were no significant differences in all variables (P > 0.05), except for the axonal birefringence and RTP. The birefringence's of the retinal fiber layer at the temporal and superior quadrants was significantly decreased (P < 0.05), whereas RTP was significantly increased (P < 0.05). In the subgroup with ON, significantly longer PERG latency and decreased VVD were observed at follow-up (P < 0.05). In patients with improved LCLA, significantly increased RTP and decreased VVD (P < 0.05) were also observed. Conclusions: This is the first longitudinal study that assessed the RTP and VVD, along with other retinal structural and functional parameters in MS. The recovery of retinal vascular function occurred with the improved LCLA, suggesting that these measurements may be associated with disease progression. Translational Relevance: The retinal microvascular changes could be potential biomarkers for monitoring therapeutic efficacy in MS.

Retinal and cortical visual acuity in a common inbred albino mouse.

While albino mice are widely used in research which includes the use of visually guided behavioral tests, information on their visual capability is scarce. We compared the spatial resolution (acuity) of albino mice (BALB/c) with that of pigmented mice (C57BL/6J). We used a high-throughput pattern electroretinogram (PERG) and pattern visual evoked potential (PVEP) method for objective assessment of retinal and cortical acuity, as well as optomotor head-tracking response/ reflex (OMR). We found that PERG, PVEP, and OMR acuities of C57BL/6J mice were all in the range of 0.5-0.6 cycles/degree (cyc/deg). BALB/c mice had PERG and PVEP acuities in the range of 0.1-0.2 cyc/deg but were unresponsive to OMR stimulus. Results indicate that retinal and cortical acuity can be reliably determined with electrophysiological methods in BALB/c mice, although PERG/PVEP acuities are lower than those of C57BL/6J mice. The reduced acuity of BALB/c mice appears to be primarily determined at retinal level.