PURPOSE: Analyze phenotypic data from knockout mice with late-adult retinal pathologic phenotypes to identify genes associated with development of adult-onset retinal diseases. METHODS: The International Mouse Phenotyping Consortium (IMPC) database was queried for genes associated with abnormal retinal phenotypes in the late-adult knockout mouse pipeline (49-80 weeks postnatal age). We identified human orthologs and performed protein-protein analysis and biological pathways analysis with known inherited retinal disease (IRD) and age-related macular degeneration (AMD) genes using Search Tool for the Retrieval of Interacting Genes/Proteins (STRING), PLatform for Analysis of single cell Eye in a Disk (PLAE), Protein Analysis Through Evolutionary Relationships (PANTHER), and Kyoto Encyclopedia of Genes and Genomes (KEGG). RESULTS: Screening of 587 late-adult mouse genes yielded 12 with abnormal retinal phenotypes, which corresponded to 20 human orthologs. Three of the 12 mouse genes and two of the 20 human orthologs were previously implicated in retinal pathology or physiology in a literature review. Although all of the genes demonstrated retinal pathology when deleted from the mouse genome, most do not have established roles in human retinal disease. Furthermore, human protein-protein analysis and biological pathway analysis yielded only a few relationships between the candidate gene list and that of known IRD and AMD genes, suggesting they may represent novel retinal functions. CONCLUSIONS: We identified 12 mouse genes with significant late-adult abnormal retinal pathology, eight of which have not been previously implicated in either mouse or human retinal physiology or pathology. These serve as novel retinal disease gene candidates for late-onset retinal disease.

• MeSH
• Phenotype MeSH
• Humans MeSH
• Macular Degeneration * genetics   MeSH
• Disease Models, Animal MeSH
• Mice, Knockout MeSH
• Mice MeSH
• Eye Proteins * genetics   MeSH
• Retina * pathology   metabolism   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

This paper describes a compact video-ophthalmoscope (VO) designed for capturing retinal video sequences of the optic nerve head (ONH) under flicker light stimulation. The device uses an OLED display and a fiber optic-coupled LED light source, enabling high-frame-rate video at low illumination intensity (12 μW/cm2). Retinal responses were recorded in 10 healthy subjects during flicker light exposure with a pupil irradiance of 2 μW/cm2. Following 20 s of stimulation, all subjects displayed changes in retinal reflectance and pulsation attenuation, linked to blood flow and volume variations. These findings suggest that increased blood volume leads to decreased retinal reflectance. Temporal analysis confirmed the ability to capture flicker-induced retinal reflectance changes, indicating its potential for spatial and temporal analysis. Overall, this device offers a portable approach for investigating dynamic retinal responses to light stimuli, which can aid the diagnosis of retinal diseases like diabetic retinopathy, glaucoma, or neurodegenerative diseases affecting retinal blood circulation.

• MeSH
• Video Recording * instrumentation   MeSH
• Adult MeSH
• Humans MeSH
• Young Adult MeSH
• Ophthalmoscopes * MeSH
• Retina * radiation effects   physiology   MeSH
• Photic Stimulation * MeSH
• Light * MeSH
• Check Tag
• Adult MeSH
• Humans MeSH
• Young Adult MeSH
• Male MeSH
• Female MeSH
• Publication type
• Journal Article MeSH

Retinitis pigmentosa (RP) is a hereditary disorder caused by mutations in more than 70 different genes including those that encode proteins important for pre-mRNA splicing. Most RP-associated mutations in splicing factors reduce either their expression, stability or incorporation into functional splicing complexes. However, we have previously shown that two RP mutations in PRPF8 (F2314L and Y2334N) and two in SNRNP200 (S1087L and R1090L) behaved differently, and it was still unclear how these mutations affect the functions of both proteins. To investigate this in the context of functional spliceosomes, we used iCLIP in HeLa and retinal pigment epithelial (RPE) cells. We found that both mutations in the RNA helicase SNRNP200 change its interaction with U4 and U6 snRNAs. The significantly broader binding profile of mutated SNRNP200 within the U4 region upstream of the U4/U6 stem I strongly suggests that its activity to unwind snRNAs is impaired. This was confirmed by FRAP measurements and helicase activity assays comparing mutant and WT protein. The RP variants of PRPF8 did not affect snRNAs, but showed a reduced binding to pre-mRNAs, which resulted in the slower splicing of introns and altered expression of hundreds of genes in RPE cells. This suggests that changes in the expression and splicing of specific genes are the main driver of retinal degeneration in PRPF8-linked RP.

• MeSH
• HeLa Cells MeSH
• Humans MeSH
• Ribonucleoprotein, U4-U6 Small Nuclear metabolism   genetics   MeSH
• Mutation * MeSH
• Eye Proteins genetics   metabolism   MeSH
• RNA Precursors * metabolism   genetics   MeSH
• RNA-Binding Proteins metabolism   genetics   MeSH
• Retinal Pigment Epithelium metabolism   pathology   MeSH
• Retinitis Pigmentosa * genetics   metabolism   pathology   MeSH
• Ribonucleoproteins, Small Nuclear metabolism   genetics   MeSH
• RNA, Small Nuclear genetics   metabolism   MeSH
• RNA Splicing * genetics   MeSH
• Spliceosomes metabolism   genetics   MeSH
• Protein Binding MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

Cannabidiol (CBD) is one of the principal constituents of Cannabis Sativa with no psychoactive properties. CBD is a promising neuroprotective compound bearing anti-inflammatory and antioxidant properties. However, considering its low solubility, CBD delivery to the retina represents an unresolved issue. The first aim was to investigate the potential neuroprotective effects of CBD in an in vivo model of retinal excitotoxicity induced by α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA). Rats underwent intravitreal co-injection of AMPA (42 nmol) and CBD (10-4 M). The neuroprotective effect of CBD was investigated with histology and immunohistochemical evaluation of inflammatory and oxidative stress biomarkers. CBD reversed the AMPA-induced total retinal, inner nuclear layer and inner plexiform layer shrinkage and loss of amacrine cells. Moreover, CBD decreased the AMPA induced number of cleaved caspase-3, Iba-1 and nitrotyrosine (NT) positive cells. Based on this evidence, we developed a nanotechnological formulation of CBD to overcome critical issues related to its eye delivery. Particularly, nanostructured lipid carriers (NLC) loaded with CBD were prepared, optimized and characterized. Due to the optimal physicochemical characteristics, CBD-NLC3 has been selected and the in vitro release profile has been investigated. Additionally, CBD-NLC3 was topically administered to rats, and retinal CBD levels were determined. CBD-NLC3 formulation, after a single topical administration, efficiently delivered CBD in the retina (Cmax = 98 ± 25.9 ng/mg; Tmax = 60 min), showing a high translational value. In conclusion, these findings showed a good PD/PK profile of CBD warranting further pre-clinical and clinical evaluation of the new formulation for the treatment of retinal degenerative diseases.

• MeSH
• Cannabidiol * pharmacokinetics   pharmacology   administration & dosage   MeSH
• Caspase 3 metabolism   MeSH
• Rats MeSH
• alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid toxicity   MeSH
• Microfilament Proteins metabolism   MeSH
• Disease Models, Animal MeSH
• Neuroprotective Agents * pharmacokinetics   pharmacology   administration & dosage   MeSH
• Drug Carriers chemistry   MeSH
• Oxidative Stress drug effects   MeSH
• Rats, Wistar MeSH
• Calcium-Binding Proteins MeSH
• Retina * drug effects   metabolism   pathology   MeSH
• Tyrosine analogs & derivatives   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

BACKGROUND: Large vessel carotid stenosis is a significant cause of ischaemic stroke. Indications for surgical revascularisation depend on the severity of the stenosis and clinical symptoms. However, mild symptoms such as TIA (Transient ischaemic attack), amaurosis fugax or minor stroke precede large strokes in only 15% of cases. AIM: The aim of this prospective study is to evaluate whether retinal perfusion is impacted in significant carotid stenosis. Automated retinal oximetry will be used to better assess perfusion in the post-stenotic basin. We presume the more stenotic the blood vessel, the more reduced the retinal perfusion is, resulting in adaptive changes such as greater arteriovenous saturation difference due to greater oxygen extraction. This could broaden the indication spectrum for revascularisation for carotid stenosis. METHODS: We plan to enroll yearly 50 patients with significant carotid stenosis and cross-examine them with retinal oximetry. The study group will provide stenotic vessels and, non-stenotic vessels will form the control group. Patients with significant carotid stenosis will undergo an MRI (Magnetic Resonnance imaging) examination to determine the presence of asymptomatic recent ischaemic lesions in the stenotic basin, and the correlation to oximetry parameters. STATISTICS: The stenosis severity and retinal oximetry parameters will be compared for study and control groups with a threshold of 70%, respectively 80% and 90% stenosis. Results will be then reevaluated with emphasis on MRI findings in the carotid basin. CONCLUSION: This prospective case control study protocol will be used to launch a multicentre trial assessing the relationship between significant carotid stenosis and retinal perfusion measured with automated retinal oximetry. Despite these differences, the findings indicate the potential of retinal oximetry for noninvasive real-time measurements of oxyhaemoglobin saturation in central nervous system vessels. Following calibration upgrade and technological improvement, verification retinal oximetry may potentially be applied to critically ill and anaesthesia care patients. The study on combined scanning laser ophthalmoscope and retinal oximetry supports the feasibility of the technique for oximetry analysis in newly born babies. TRIAL REGISTRATION: ClinicalTrials.gov, ID: NCT06085612.

• MeSH
• Middle Aged MeSH
• Humans MeSH
• Oximetry * methods   MeSH
• Prospective Studies MeSH
• Retina diagnostic imaging   physiopathology   MeSH
• Retinal Vessels diagnostic imaging   physiopathology   MeSH
• Aged MeSH
• Carotid Stenosis * physiopathology   surgery   complications   MeSH
• Case-Control Studies MeSH
• Check Tag
• Middle Aged MeSH
• Humans MeSH
• Male MeSH
• Aged MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Clinical Trial Protocol MeSH

PURPOSE: This study aimed to evaluate early-phase safety of subretinal application of AAVanc80.CAG.USH1Ca1 (OT_USH_101) in wild-type (WT) pigs, examining the effects of a vehicle control, low dose, and high dose. METHODS: Twelve WT pigs (24 eyes) were divided into three groups: four pigs each received bilateral subretinal injections of either vehicle, low dose (3.3 × 1010 vector genomes [vg] per eye), or high dose (1.0 × 1011 vg per eye). Total retinal thickness (TRT) was evaluated using optical coherence tomography and retinal function was assessed with full-field electroretinography (ff-ERG) at baseline and two months post-surgery. After necropsy, retinal changes were examined through histopathology, and human USH1C_a1/harmonin expression was assessed by quantitative PCR (qPCR) and Western blotting. RESULTS: OT_USH_101 led to high USH1C_a1 expression in WT pig retinas without significant TRT changes two months after subretinal injection. The qPCR revealed expression of the human USH1C_a1 transgene delivered by the adeno-associated virus vector. TRT changes were minimal across groups: vehicle (256 ± 21 to 243 ± 18 μm; P = 0.108), low dose (251 ± 32 to 258 ± 30 μm; P = 0.076), and high dose (242 ± 24 to 259 ± 28 μm; P = 0.590). The ff-ERG showed no significant changes in rod or cone responses. Histopathology indicated no severe retinal adverse effects in the vehicle and low dose groups. CONCLUSIONS: Early-phase clinical imaging, electrophysiology, and histopathological assessments indicated that subretinal administration of OT_USH_101 was well tolerated in the low-dose treatment arm. OT_USH_101 treatment resulted in high expression of human USH1C_a1. Although histopathological changes were not severe, more frequent changes were observed in the high-dose group.

• MeSH
• Cytoskeletal Proteins genetics   MeSH
• Dependovirus genetics   MeSH
• Electroretinography * MeSH
• Genetic Therapy methods   MeSH
• Genetic Vectors * MeSH
• Injections, Intraocular * MeSH
• Real-Time Polymerase Chain Reaction MeSH
• Humans MeSH
• Disease Models, Animal MeSH
• Tomography, Optical Coherence * MeSH
• Swine MeSH
• Cell Cycle Proteins genetics   MeSH
• Gene Expression Regulation MeSH
• Retina * metabolism   pathology   MeSH
• Transgenes * MeSH
• Blotting, Western MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

PURPOSE: Subretinal (SR) injection in porcine models is a promising avenue for preclinical evaluation of cell and gene therapies. Targeting of the subretinal fluid compartment (bleb) is critical to the procedure, especially if treatment of the cone-rich area centralis is required (i.e., visual streak [VS] in pigs). To our knowledge, this study is the first to investigate the influence of injection site placement on VS involvement in the pig eye. METHODS: We performed 23-gauge pars plana vitrectomy followed by SR injection in 41 eyes of 21 animals (Sus scrofa domesticus). In 27 eyes (65.9%), the injection site was placed superior to the VS, and in 14 eyes (34.1%) it was placed inferior to it. Using intraoperative imaging, blebs were classified based on their propagation behavior relative to the VS. RESULTS: In 79% of cases, blebs from inferior injection sites developed away from the VS, exhibiting a mean ± SEM vertical anisotropy (AP) of 0.67 ± 0.11. In contrast, blebs from superior injection sites tended to develop toward the VS with an AP of 1.27 ± 0.18 (P = 0.0070). Blebs developed away from the VS in only 41% of injections (P = 0.0212). Inferior blebs were orientated close to 0° (horizontal), whereas superior blebs displayed varied orientations with a mean angle of 56° (P = 0.0008). CONCLUSIONS: Bleb propagation was anisotropic (i.e., directionally biased) and dependent on injection site placement. Superior injection sites led to superior VS detachment. Morphological analysis suggested increased adhesion forces at the VS and superior vascular arcades. This study will aid the planning of surgeries for targeted subretinal delivery in pig models.

• MeSH
• Genetic Therapy * methods   MeSH
• Injections, Intraocular MeSH
• Disease Models, Animal * MeSH
• Swine MeSH
• Retina MeSH
• Subretinal Fluid metabolism   MeSH
• Sus scrofa MeSH
• Vitrectomy * methods   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH

Genetic features are currently unknown in myelinated retinal nerve fibers (MRNF). For a 20-year-old asymptomatic female with unilateral MRNF, we performed whole genome sequencing (WGS) by standard workflow protocol to produce contiguous long-read sequences with Illumina DNA PCR-Free Prep. After tagmentation, libraries were sequenced on separate runs via NovaSeq 6000 platform at 2 x 150bp read length. Gene variants included rs2248799, rs2672589, rs7555070, rs247616_T and rs2043085_C all associated with an increased macular degeneration risk, and seven novel variants of uncertain significance. For optic disc enlargement, variants rs9988687_A, rs11079419_T, rs6787363 and rs10862708_A suggested an increased risk for this condition. In contrast, modeling revealed retinal detachment risk was reduced by variants identified at rs9651980_T, rs4373767_T, and rs7940691_T which were among five other previously unreported variants. WGS data placed proband at the 66th and 64th percentiles for disc anomaly and retinal detachment risk, respectively. Additionally, risk determined from 16 loci associated with age-related macular degeneration found the patient to be at the 18th percentile for this diagnosis (i.e., below average genetic predisposition). Fundoscopic findings showed mean RNFL thickness was lower with MRNF (77 OS vs. 96?m OD) and RNFL symmetry was impaired (43 %) but stable between 2020 and 2023. Rim area and cup volume were also substantially different (2.33 OS vs. 1.34mm2 OD, and 0.001 OS vs. 0.151mm3 OD, respectively). As the first known evaluation of MRNF via WGS, these data reveal a mixed picture with variants associated with different risks for potentially related ocular pathologies. In addition, we identify multiple new variants of unknown significance. Factors affecting gene expression in MRNF require further study. Key words: Whole genome sequencing, Retina, Myelination, Anatomy, Gene variants.

• MeSH
• Genetic Predisposition to Disease MeSH
• Humans MeSH
• Young Adult MeSH
• Nerve Fibers, Myelinated * pathology   MeSH
• Retina pathology   MeSH
• Whole Genome Sequencing * MeSH
• Check Tag
• Humans MeSH
• Young Adult MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Case Reports MeSH

AIMS: To investigate the therapeutic potential of visual stimulation (VS) and BDNF in murine experimental autoimmune uveoretinitis (EAU). MAIN METHODS: Mice were immunized by subcutaneous injection of interphotoreceptor retinoid-binding protein in Freund's complete adjuvant and intravenous injection of pertussis toxin, and were then exposed to high-contrast VS 12 h/day (days 1-14 post-immunization). EAU severity was assessed by examining clinical score, visual acuity, inflammatory markers, and immune cells in the retina. The transcriptome of activated retinal cells was determined by RNA-seq using RNA immunoprecipitated in complex with phosphorylated ribosomal protein S6. The retinal levels of protein products of relevant upregulated genes were quantified. The effect of BDNF on EAU was tested in unstimulated mice by its daily topical ocular administration (days 8-14 post-immunization). KEY FINDINGS: VS attenuated EAU development and decreased the expression of pro-inflammatory cytokines/chemokines and numbers of immune cells in the retina (n = 10-20 eyes/group for each analysis). In activated retinal cells of control mice (n = 30 eyes/group), VS upregulated genes encoding immunomodulatory neuropeptides, of which BDNF and vasoactive intestinal peptide (VIP) also showed increased mRNA and protein levels in the retina of VS-treated EAU mice (n = 6-10 eyes/group for each analysis). In unstimulated EAU mice, BDNF treatment mimicked the protective effects of VS by modulating the inflammatory and stem cell properties of Müller cells (n = 5 eyes/group for each analysis). SIGNIFICANCE: VS effectively suppresses EAU, at least through enhancing retinal levels of anti-inflammatory and neuroprotective factors, VIP and BDNF. Our findings also suggest BDNF as a promising therapeutic agent for uveitis treatment.

• MeSH
• Autoimmune Diseases * immunology   metabolism   MeSH
• Cytokines metabolism   MeSH
• Disease Models, Animal MeSH
• Brain-Derived Neurotrophic Factor * metabolism   MeSH
• Mice, Inbred C57BL MeSH
• Mice MeSH
• Retina metabolism   drug effects   MeSH
• Retinitis * drug therapy   prevention & control   immunology   MeSH
• Uveitis * metabolism   drug therapy   immunology   MeSH
• Vasoactive Intestinal Peptide pharmacology   MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

Maintenance of NAD pools is critical for neuronal survival. The capacity to maintain NAD pools declines in neurodegenerative disease. We identify that low NMNAT2, the critical neuronal NAD producing enzyme, drives retinal susceptibility to neurodegenerative insults. As proof of concept, gene therapy over-expressing full length human NMNAT2 is neuroprotective. To pharmacologically target NMNAT2, we identify that epigallocatechin gallate (EGCG) can drive NAD production in neurons through an NMNAT2 and NMN dependent mechanism. We confirm this by pharmacological and genetic inhibition of the NAD-salvage pathway. EGCG is neuroprotective in rodent (mixed sex) and human models of retinal neurodegeneration. As EGCG has poor drug-like qualities, we use it as a tool compound to generate novel small molecules which drive neuronal NAD production and provide neuroprotection. This class of NMNAT2 targeted small molecules could have an important therapeutic impact for neurodegenerative disease following further drug development.

• MeSH
• Genetic Therapy methods   MeSH
• Catechin * analogs & derivatives   pharmacology   MeSH
• Rats MeSH
• Humans MeSH
• Disease Models, Animal MeSH
• Mice, Inbred C57BL MeSH
• Mice MeSH
• NAD * metabolism   MeSH
• Neurodegenerative Diseases drug therapy   metabolism   genetics   MeSH
• Neurons * metabolism   drug effects   MeSH
• Neuroprotective Agents * pharmacology   MeSH
• Nicotinamide-Nucleotide Adenylyltransferase * metabolism   genetics   MeSH
• Retina metabolism   drug effects   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Humans MeSH
• Male MeSH
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH