Translational Retinal Research and Therapies.

The following review summarizes the state of the art in representative aspects of gene therapy/translational medicine and evolves from a symposium held at the School of Veterinary Medicine, University of Pennsylvania on November 16, 2017 honoring Dr. Gustavo Aguirre, recipient of ARVO's 2017 Proctor Medal. Focusing on the retina, speakers highlighted current work on moving therapies for inherited retinal degenerative diseases from the laboratory bench to the clinic

The Relationship Between Ambient Atmospheric Fine Particulate Matter (PM2.5) and Glaucoma in a Large Community Cohort.

Purpose: Glaucoma is more common in urban populations than in others. Ninety percent of the world's population are exposed to air pollution above World Health Organization (WHO) recommended limits. Few studies have examined the association between air pollution and glaucoma. Methods: Questionnaire data, ophthalmic measures, and ambient residential area air quality data for 111,370 UK Biobank participants were analyzed. Particulate matter with an aerodynamic diameter < 2.5 μm (PM2.5) was selected as the air quality exposure of interest. Eye measures included self-reported glaucoma, intraocular pressure (IOP), and average thickness of macular ganglion cell-inner plexiform layer (GCIPL) across nine Early Treatment Diabetic Retinopathy Study (ETDRS) retinal subfields as obtained from spectral-domain optical coherence tomography. We examined the associations of PM2.5 concentration with self-reported glaucoma, IOP, and GCIPL. Results: Participants resident in areas with higher PM2.5 concentration were more likely to report a diagnosis of glaucoma (odds ratio = 1.06, 95% confidence interval [CI] = 1.01-1.12, per interquartile range [IQR] increase P = 0.02). Higher PM2.5 concentration was also associated with thinner GCIPL (β = -0.56 μm, 95% CI = -0.63 to -0.49, per IQR increase, P = 1.2 × 10-53). A dose-response relationship was observed between higher levels of PM2.5 and thinner GCIPL (P < 0.001). There was no clinically relevant relationship between PM2.5 concentration and IOP. Conclusions: Greater exposure to PM2.5 is associated with both self-reported glaucoma and adverse structural characteristics of the disease. The absence of an association between PM2.5 and IOP suggests the relationship may occur through a non-pressure-dependent mechanism, possibly neurotoxic and/or vascular effects

RP2-associated X-linked Retinopathy: Clinical Findings, Molecular Genetics, and Natural History

PURPOSE: To review and describe in detail the clinical course, functional and anatomical characteristics of RP2-associated retinal degeneration. DESIGN: Retrospective case series. PARTICIPANTS: Males with disease-causing variants in the RP2 gene. METHODS: Review of all case notes and results of molecular genetic testing, retinal imaging (fundus autofluorescence (FAF) imaging, optical coherence tomography (OCT)) and electrophysiology assessment. MAIN OUTCOME MEASURES: Molecular genetic testing, clinical findings including best-corrected visual acuity (BCVA), qualitative and quantitative retinal imaging analysis, and electrophysiology parameters. RESULTS: Fifty-four molecularly confirmed patients were identified, from 38 pedigrees. Twenty-eight disease-causing variants were identified; with 20 not previously clinically characterized. Fifty-three patients (98.1%) presented with retinitis pigmentosa. The mean age of onset (range, ±SD) was 9.6 years of age (1-57 years, ± 9.2 years). Forty-four patients (91.7%) had childhood-onset disease, with mean age of onset of 7.6 years. The commonest first symptom was night blindness (68.8%). Mean BCVA (range, ±SD) was 0.91 LogMAR (0-2.7, ±0.80) and 0.94 LogMAR (0-2.7, ±0.78) for right and left eyes respectively. Based on the WHO visual impairment criteria, 18 patients (34%) had low vision. The majority (17/22) showed ERG evidence of a rod-cone dystrophy. Pattern ERG P50 was undetectable in all but 2 patients. A range of FAF findings was observed, from normal to advanced atrophy. There were no statistically significant differences between right and left eyes for ellipsoid zone (EZ) width and outer nuclear layer (ONL) thickness. The mean annual rate of EZ width loss was 219 μm/year and the mean annual decrease in ONL thickness was 4.93 μm/year. No patient with childhood-onset disease had identifiable EZ after the age of 26 years at baseline or follow-up. Four patients had adulthood-onset disease and a less severe phenotype. CONCLUSIONS: This study details the clinical phenotype of RP2 retinopathy in a large cohort. The majority presented with early-onset severe retinal degeneration, with early macular involvement and complete loss of the foveal photoreceptor layer by the third decade of life. Full-field ERGs revealed rod-cone dystrophy in the vast majority, but with generalised (peripheral) cone system involvement of widely varying severity in the first two decades of life

Cone photoreceptor structure in patients with x-linked cone dysfunction and red-green color vision deficiency

PURPOSE: Mutations in the coding sequence of the L and M opsin genes are often associated with X-linked cone dysfunction (such as Bornholm Eye Disease, BED), though the exact color vision phenotype associated with these disorders is variable. We examined individuals with L/M opsin gene mutations to clarify the link between color vision deficiency and cone dysfunction. METHODS: We recruited 17 males for imaging. The thickness and integrity of the photoreceptor layers were evaluated using spectral-domain optical coherence tomography. Cone density was measured using high-resolution images of the cone mosaic obtained with adaptive optics scanning light ophthalmoscopy. The L/M opsin gene array was characterized in 16 subjects, including at least one subject from each family. RESULTS: There were six subjects with the LVAVA haplotype encoded by exon 3, seven with LIAVA, two with the Cys203Arg mutation encoded by exon 4, and two with a novel insertion in exon 2. Foveal cone structure and retinal thickness was disrupted to a variable degree, even among related individuals with the same L/M array. CONCLUSIONS: Our findings provide a direct link between disruption of the cone mosaic and L/M opsin variants. We hypothesize that, in addition to large phenotypic differences between different L/M opsin variants, the ratio of expression of first versus downstream genes in the L/M array contributes to phenotypic diversity. While the L/M opsin mutations underlie the cone dysfunction in all of the subjects tested, the color vision defect can be caused either by the same mutation or a gene rearrangement at the same locus

Substitution of a single non-coding nucleotide upstream of TMEM216 causes non-syndromic retinitis pigmentosa and is associated with reduced TMEM216 expression

Genome analysis of individuals affected by retinitis pigmentosa (RP) identified two rare nucleotide substitutions at the same genomic location on chromosome 11 (g.61392563 [GRCh38]), 69 base pairs upstream of the start codon of the ciliopathy gene TMEM216 (c.-69G&gt;A, c.-69G&gt;T [GenBank: NM_001173991.3]), in individuals of South Asian and African ancestry, respectively. Genotypes included 71 homozygotes and 3 mixed heterozygotes in trans with a predicted loss-of-function allele. Haplotype analysis showed single-nucleotide variants (SNVs) common across families, suggesting ancestral alleles within the two distinct ethnic populations. Clinical phenotype analysis of 62 available individuals from 49 families indicated a similar clinical presentation with night blindness in the first decade and progressive peripheral field loss thereafter. No evident systemic ciliopathy features were noted. Functional characterization of these variants by luciferase reporter gene assay showed reduced promotor activity. Nanopore sequencing confirmed the lower transcription of the TMEM216 c.-69G&gt;T allele in blood-derived RNA from a heterozygous carrier, and reduced expression was further recapitulated by qPCR, using both leukocytes-derived RNA of c.-69G&gt;T homozygotes and total RNA from genome-edited hTERT-RPE1 cells carrying homozygous TMEM216 c.-69G&gt;A. In conclusion, these variants explain a significant proportion of unsolved cases, specifically in individuals of African ancestry, suggesting that reduced TMEM216 expression might lead to abnormal ciliogenesis and photoreceptor degeneration

Substitution of a single non-coding nucleotide upstream of TMEM216 causes non-syndromic retinitis pigmentosa and is associated with reduced TMEM216 expression

Genome analysis of individuals affected by retinitis pigmentosa (RP) identified two rare nucleotide substitutions at the same genomic location on chromosome 11 (g.61392563 [GRCh38]), 69 base pairs upstream of the start codon of the ciliopathy gene TMEM216 (c.−69G>A, c.−69G>T [GenBank: NM_001173991.3]), in individuals of South Asian and African ancestry, respectively. Genotypes included 71 homozygotes and 3 mixed heterozygotes in trans with a predicted loss-of-function allele. Haplotype analysis showed single-nucleotide variants (SNVs) common across families, suggesting ancestral alleles within the two distinct ethnic populations. Clinical phenotype analysis of 62 available individuals from 49 families indicated a similar clinical presentation with night blindness in the first decade and progressive peripheral field loss thereafter. No evident systemic ciliopathy features were noted. Functional characterization of these variants by luciferase reporter gene assay showed reduced promotor activity. Nanopore sequencing confirmed the lower transcription of the TMEM216 c.−69G>T allele in blood-derived RNA from a heterozygous carrier, and reduced expression was further recapitulated by qPCR, using both leukocytes-derived RNA of c.−69G>T homozygotes and total RNA from genome-edited hTERT-RPE1 cells carrying homozygous TMEM216 c.−69G>A. In conclusion, these variants explain a significant proportion of unsolved cases, specifically in individuals of African ancestry, suggesting that reduced TMEM216 expression might lead to abnormal ciliogenesis and photoreceptor degeneration

Investigation of associations between retinal microvascular parameters and albuminuria in UK Biobank: a cross-sectional case-control study.

BACKGROUND: Associations between microvascular variation and chronic kidney disease (CKD) have been reported previously. Non-invasive retinal fundus imaging enables evaluation of the microvascular network and may offer insight to systemic risk associated with CKD. METHODS: Retinal microvascular parameters (fractal dimension [FD] - a measure of the complexity of the vascular network, tortuosity, and retinal arteriolar and venular calibre) were quantified from macula-centred fundus images using the Vessel Assessment and Measurement Platform for Images of the REtina (VAMPIRE) version 3.1 (VAMPIRE group, Universities of Dundee and Edinburgh, Scotland) and assessed for associations with renal damage in a case-control study nested within the multi-centre UK Biobank cohort study. Participants were designated cases or controls based on urinary albumin to creatinine ratio (ACR) thresholds. Participants with ACR ≥ 3 mg/mmol (ACR stages A2-A3) were characterised as cases, and those with an ACR < 3 mg/mmol (ACR stage A1) were categorised as controls. Participants were matched on age, sex and ethnic background. RESULTS: Lower FD (less extensive microvascular branching) was associated with a small increase in odds of albuminuria independent of blood pressure, diabetes and other potential confounding variables (odds ratio [OR] 1.18, 95% confidence interval [CI] 1.03-1.34 for arterioles and OR 1.24, CI 1.05-1.47 for venules). Measures of tortuosity or retinal arteriolar and venular calibre were not significantly associated with ACR. CONCLUSIONS: This study supports previously reported associations between retinal microvascular FD and other metabolic disturbances affecting the systemic vasculature. The association between retinal microvascular FD and albuminuria, independent of diabetes and blood pressure, may represent a useful indicator of systemic vascular damage associated with albuminuria

Associations with corneal hysteresis in a population cohort: results from 96 010 UK Biobank participants

Purpose To describe the distribution of corneal hysteresis (CH) in a large cohort and explore its associated factors and possible clinical applications. Design Cross-sectional study within the UK Biobank, a large cohort study in the United Kingdom. Participants We analyzed CH data from 93 345 eligible participants in the UK Biobank cohort, aged 40 to 69 years. Methods All analyses were performed using left eye data. Linear regression models were used to evaluate associations between CH and demographic, lifestyle, ocular, and systemic variables. Piecewise logistic regression models were used to explore the relationship between self-reported glaucoma and CH. Main Outcome Measures Corneal hysteresis (mmHg). Results The mean CH was 10.6 mmHg (10.4 mmHg in male and 10.8 mmHg in female participants). After adjusting for covariables, CH was significantly negatively associated with male sex, age, black ethnicity, self-reported glaucoma, diastolic blood pressure, and height. Corneal hysteresis was significantly positively associated with smoking, hyperopia, diabetes, systemic lupus erythematosus (SLE), greater deprivation (Townsend index), and Goldmann-correlated intraocular pressure (IOPg). Self-reported glaucoma and CH were significantly associated when CH was less than 10.1 mmHg (odds ratio, 0.86; 95% confidence interval, 0.79–0.94 per mmHg CH increase) after adjusting for covariables. When CH exceeded 10.1 mmHg, there was no significant association between CH and self-reported glaucoma. Conclusions In our analyses, CH was significantly associated with factors including age, sex, and ethnicity, which should be taken into account when interpreting CH values. In our cohort, lower CH was significantly associated with a higher prevalence of self-reported glaucoma when CH was less than 10.1 mmHg. Corneal hysteresis may serve as a biomarker aiding glaucoma case detection

A multi-ethnic genome-wide association study implicates collagen matrix integrity and cell differentiation pathways in keratoconus

Keratoconus is characterised by reduced rigidity of the cornea with distortion and focal thinning that causes blurred vision, however, the pathogenetic mechanisms are unknown. It can lead to severe visual morbidity in children and young adults and is a common indication for corneal transplantation worldwide. Here we report the first large scale genome-wide association study of keratoconus including 4,669 cases and 116,547 controls. We have identified significant association with 36 genomic loci that, for the first time, implicate both dysregulation of corneal collagen matrix integrity and cell differentiation pathways as primary disease-causing mechanisms. The results also suggest pleiotropy, with some disease mechanisms shared with other corneal diseases, such as Fuchs endothelial corneal dystrophy. The common variants associated with keratoconus explain 12.5% of the genetic variance, which shows potential for the future development of a diagnostic test to detect susceptibility to disease

Multi-trait Genome-wide Association Study Identifies New Loci Associated With Optic Disc Parameters

A new avenue of mining published genome-wide association studies includes the joint analysis of related traits. The power of this approach depends on the genetic correlation of traits, which reflects the number of pleiotropic loci, i.e. genetic loci influencing multiple traits. Here, we applied new meta-analyses of optic nerve head (ONH) related traits implicated in primary open-angle glaucoma (POAG); intraocular pressure and central corneal thickness using Haplotype reference consortium imputations. We performed a multi-trait analysis of ONH parameters cup area, disc area and vertical cup-disc ratio. We uncover new variants; rs11158547 in PPP1R36-PLEKHG3 and rs1028727 near SERPINE3 at genome-wide significance that replicate in independent Asian cohorts imputed to 1000 Genomes. At this point, validation of these variants in POAG cohorts is hampered by the high degree of heterogeneity. Our results show that multi-trait analysis is a valid approach to identify novel pleiotropic variants for ONH