Genetic and Clinical Profile of Retinopathies Due to Disease-Causing Variants in Leber Congenital Amaurosis (LCA)-Associated Genes in a Large German Cohort.

To report the spectrum of Leber congenital amaurosis (LCA) associated genes in a large German cohort and to delineate their associated phenotype. Local databases were screened for patients with a clinical diagnosis of LCA and for patients with disease-causing variants in known LCA-associated genes independent of their clinical diagnosis. Patients with a mere clinical diagnosis were invited for genetic testing. Genomic DNA was either analyzed in a diagnostic-genetic or research setup using various capture panels for syndromic and non-syndromic IRD (inherited retinal dystrophy) genes. Clinical data was obtained mainly retrospectively. Patients with genetic and phenotypic information were eventually included. Descriptive statistical data analysis was performed. A total of 105 patients (53 female, 52 male, age 3-76 years at the time of data collection) with disease-causing variants in 16 LCA-associated genes were included. The genetic spectrum displayed variants in the following genes: CEP290 (21%), CRB1 (21%), RPE65 (14%), RDH12 (13%), AIPL1 (6%), TULP1 (6%), and IQCB1 (5%), and few cases harbored pathogenic variants in LRAT, CABP4, NMNAT1, RPGRIP1, SPATA7, CRX, IFT140, LCA5, and RD3 (altogether accounting for 14%). The most common clinical diagnosis was LCA (53%, 56/105) followed by retinitis pigmentosa (RP, 40%, 42/105), but also other IRDs were seen (cone-rod dystrophy, 5%; congenital stationary night blindness, 2%). Among LCA patients, 50% were caused by variants in CEP290 (29%) and RPE65 (21%), whereas variants in other genes were much less frequent (CRB1 11%, AIPL1 11%, IQCB1 9%, and RDH12 7%, and sporadically LRAT, NMNAT1, CRX, RD3, and RPGRIP1). In general, the patients showed a severe phenotype hallmarked by severely reduced visual acuity, concentric narrowing of the visual field, and extinguished electroretinograms. However, there were also exceptional cases with best corrected visual acuity as high as 0.8 (Snellen), well-preserved visual fields, and preserved photoreceptors in spectral domain optical coherence tomography. Phenotypic variability was seen between and within genetic subgroups. The study we are presenting pertains to a considerable LCA group, furnishing valuable comprehension of the genetic and phenotypic spectrum. This knowledge holds significance for impending gene therapeutic trials. In this German cohort, CEP290 and CRB1 are the most frequently mutated genes. However, LCA is genetically highly heterogeneous and exhibits clinical variability, showing overlap with other IRDs. For any therapeutic gene intervention, the disease-causing genotype is the primary criterion for treatment access, but the clinical diagnosis, state of the retina, number of to be treated target cells, and the time point of treatment will be crucial.

Comprehensive variant spectrum of the CNGA3 gene in patients affected by achromatopsia.

Achromatopsia (ACHM) is a congenital cone photoreceptor disorder characterized by impaired color discrimination, low visual acuity, photosensitivity, and nystagmus. To date, six genes have been associated with ACHM (CNGA3, CNGB3, GNAT2, PDE6C, PDE6H, and ATF6), the majority of these being implicated in the cone phototransduction cascade. CNGA3 encodes the CNGA3 subunit of the cyclic nucleotide-gated ion channel in cone photoreceptors and is one of the major disease-associated genes for ACHM. Herein, we provide a comprehensive overview of the CNGA3 variant spectrum in a cohort of 1060 genetically confirmed ACHM patients, 385 (36.3%) of these carrying "likely disease-causing" variants in CNGA3. Compiling our own genetic data with those reported in the literature and in public databases, we further extend the CNGA3 variant spectrum to a total of 316 variants, 244 of which we interpreted as "likely disease-causing" according to ACMG/AMP criteria. We report 48 novel "likely disease-causing" variants, 24 of which are missense substitutions underlining the predominant role of this mutation class in the CNGA3 variant spectrum. In addition, we provide extensive in silico analyses and summarize reported functional data of previously analyzed missense, nonsense and splicing variants to further advance the pathogenicity assessment of the identified variants.

Comparison of CRT and LCD monitors for objective estimation of visual acuity using the sweep VEP.

PURPOSE: To investigate the applicability of liquid crystal displays (LCD) as suitable replacement for cathode ray tube monitors (CRT) as stimulator for the sweep VEP for estimating visual acuity. METHODS: In a first experiment, sweep VEPs were recorded in 13 healthy volunteers with best-corrected visual acuity with an LCD and a CRT monitor, respectively. Time-to-peak after stimulus and peak-to-trough amplitudes as well as the visual acuity, estimated using a second-order polynomial and the modified Ricker model, were compared between both monitor types. In a second experiment, sweep VEPs were recorded in six healthy volunteers with two levels of stimulus contrast using artificially reduced visual acuities as well as best-corrected with the same monitors as in the first experiment and additionally, a modern LCD gaming monitor with a response time of 1 ms. Time-to-peak after stimulus and peak-to-trough amplitudes were compared between the different combinations of monitors and contrasts. Finally, visual acuities estimated using the modified Ricker model were compared to subjective visual acuities determined using the Freiburg Visual Acuity and Contrast Test (FrACT). RESULTS: In the first experiment, the time-to-peak after stimulus presentation was statistically significantly delayed for LCD displays (mean difference [confidence interval]: 60.0 [54.0, 65.9] ms; t(516) = 19.7096, p < 0.0001). Likewise, peak-to-trough amplitudes were statistically significantly smaller for the LCD stimulator, however, not clinically relevant (mean difference [confidence interval]: - 0.89 [- 1.59, - 0.20] µV; t(516) = - 2.5351, p = 0.0115). No statistically significant effect of the monitor type on the estimated visual acuity was found for neither method, second-order polynomial, nor the modified Ricker model. In the second experiment, statistically significant delays of the time-to-peak after stimulus onset were found for all combinations of monitor and contrast compared to the CRT monitor. A statistically significant, but not clinically relevant, difference of the peak-to-trough amplitudes was only found between the CRT monitor and the LCD gaming monitor (mean difference [confidence interval]: 2.6 [1.2, 4.0] µV; t(814) = 4.66, p < 0.0001). Visual acuities estimated from LCD stimulation significantly underestimated the subjective visual acuity up to 0.2 logMAR using the conversion formula of the first experiment. No statistically significant difference was found when using conversion formulas adjusted for each combination of monitor and contrast. CONCLUSIONS: Based on the results of this study, LCD monitors may substitute CRT monitors for presenting the stimuli for the sweep VEP to objectively estimate visual acuity. Nevertheless, it is advisable to perform a calibration and to collect normative data of healthy volunteers using best-corrected and artificially reduced visual acuity for establishing a conversion formula between sweep VEP outcome and the subjective visual acuity before replacing a CRT with an LCD stimulator.

Characteristics of Retinitis Pigmentosa Associated with ADGRV1 and Comparison with USH2A in Patients from a Multicentric Usher Syndrome Study Treatrush.

In contrast to USH2A, variants in ADGRV1 are a minor cause of Usher syndrome type 2, and the associated phenotype is less known. The purpose of the study was to characterize the retinal phenotype of 18 ADGRV1 patients (9 male, 9 female; median age 52 years) and compare it with that of 204 USH2A patients (111 male, 93 female; median age 43 years) in terms of nyctalopia onset, best corrected visual acuity (BCVA), fundus autofluorescence (FAF), and optical coherence tomography (OCT) features. There was no statistical difference in the median age at onset (30 and 18 years; Mann-Whitney U test, p = 0.13); the mean age when 50% of the patients reached legal blindness (≥1.0 log MAR) based on visual acuity (64 years for both groups; log-rank, p = 0.3); the risk of developing advanced retinal degeneration (patch or atrophy) with age (multiple logistic regression, p = 0.8); or the frequency of cystoid macular edema (31% vs. 26%, Fisher's exact test, p = 0.4). ADGRV1 and USH2A retinopathy were indistinguishable in all major functional and structural characteristics, suggesting that the loss of function of the corresponding proteins produces similar effects in the retina. The results are important for counseling ADGRV1 patients, who represent the minor patient subgroup.

Clinical Phenotype of PDE6B-Associated Retinitis Pigmentosa.

In this retrospective, longitudinal, observational cohort study, we investigated the phenotypic and genotypic features of retinitis pigmentosa associated with variants in the PDE6B gene. Patients underwent clinical examination and genetic testing at a single tertiary referral center, including best-corrected visual acuity (BCVA), kinetic visual field (VF), full-field electroretinography, full-field stimulus threshold, spectral domain optical coherence tomography, and fundus autofluorescence imaging. The genetic testing comprised candidate gene sequencing, inherited retinal disease gene panel sequencing, whole-genome sequencing, and testing for familial variants by Sanger sequencing. Twenty-four patients with mutations in PDE6B from 21 families were included in the study (mean age at the first visit: 32.1 ± 13.5 years). The majority of variants were putative splicing defects (8/23) and missense (7/23) mutations. Seventy-nine percent (38/48) of eyes had no visual acuity impairment at the first visit. Visual acuity impairment was mild in 4% (2/48), moderate in 13% (6/48), and severe in 4% (2/48). BCVA was symmetrical in the right and left eyes. The kinetic VF measurements were highly symmetrical in the right and left eyes, as was the horizontal ellipsoid zone (EZ) width. Regarding the genetic findings, 43% of the PDE6B variants found in our patients were novel. Thus, this study contributed substantially to the PDE6B mutation spectrum. The visual acuity impairment was mild in 83% of eyes, providing a window of opportunity for investigational new drugs. The EZ width was reduced in all patients and was highly symmetric between the eyes, making it a promising outcome measure. We expect these findings to have implications on the design of future PDE6B-related retinitis pigmentosa (RP) clinical trials.

Genetic architecture of inherited retinal degeneration in Germany: A large cohort study from a single diagnostic center over a 9-year period.

We aimed to unravel the molecular genetic basis of inherited retinal degeneration (IRD) in a comprehensive cohort of patients diagnosed in the largest center for IRD in Germany. A cohort of 2,158 affected patients from 1,785 families diagnosed with IRD was analyzed by targeted next-generation sequencing (NGS). Patients with single-gene disorders (i.e., choroideremia and retinoschisis) were analyzed by Sanger sequencing and multiplex ligation-dependent probe amplification. Our study cohort accounts for ∼7% of the estimated 30,000 patients with IRD in Germany, thereby providing representative data for both the prevalence of IRDs and the mutation spectrum of IRD genes for the population in Germany. We achieved a molecular diagnostic rate of 35-95%, depending on the clinical entities, with a high detection rate for achromatopsia, retinoschisis, and choroideremia, and a low detection rate for central areolar choroidal dystrophy and macular dystrophy. A total of 1,161 distinct variants were identified, including 501 novel variants, reaffirming the known vast genetic heterogeneity of IRD in a mainly outbred European population. This study demonstrates the clinical utility of panel-based NGS in a large and highly heterogeneous cohort from an outbred population and for the first time gives a comprehensive representation of the genetic landscape of IRDs in Germany. The data are valuable and crucial for the scientific community and healthcare providers, but also for the pharmaceutical industry in the progressing field of personalized medicine and gene therapy.

Objective assessment of visual acuity: a refined model for analyzing the sweep VEP.

PURPOSE: The aim of this study was to develop a simple and reliable method for the objective assessment of visual acuity by optimizing the stimulus used in commercially available systems and by improving the methods of evaluation using a nonlinear function, the modified Ricker model. METHODS: Subjective visual acuity in the normal subjects was measured with Snellen targets, best-corrected, and in some cases also uncorrected and with plus lenses (+ 1 D, + 2 D, + 3 D). In patients, subjective visual acuity was measured best-corrected using the Freiburg Visual Acuity Test. Sweep VEP recordings to 11 spatial frequencies, with check sizes in logarithmically equidistant steps (0.6, 0.9, 1.4, 2.1, 3.3, 4.9, 7.3, 10.4, 18.2, 24.4, and 36.5 cpd), were obtained from 56 healthy subjects aged between 17 and 69 years (mean 42.5 ± 15.3 SD years) and 20 patients with diseases of the lens (n = 6), retina (n = 8) or optic nerve (n = 6). The results were fit by a multiple linear regression (2nd-order polynomial) or a nonlinear regression (modified Ricker model) and parameters compared (limiting spatial frequency (sflimiting) and the spatial frequency of the vertex (sfvertex) of the parabola for the 2nd-order polynomial fitting, and the maximal spatial frequency (sfmax), and the spatial frequency where the amplitude is 2 dB higher than the level of noise (sfthreshold) for the modified Ricker model. RESULTS: Recording with 11 spatial frequencies allows a more accurate determination of acuities above 1.0 logMAR. Tuning curves fitted to the results show that compared to the normal 2nd-order polynomial analysis, the modified Ricker model is able to describe closely the amplitudes of the sweep VEP in relation to the spatial frequencies of the presented checkerboards. In patients with a visual acuity better than about 0.5 (decimal), the predicted acuities based on the different parameters show a good match of the predicted visual acuities based on the models established in healthy volunteers to the subjective visual acuities. However, for lower visual acuities, both models tend to overestimate the visual acuity (up to ~ 0.4 logMAR), especially in patients suffering from AMD. CONCLUSIONS: Both models, the 2nd-order polynomial and the modified Ricker model performed equally well in the prediction of the visual acuity based on the amplitudes recorded using the sweep VEP. However, the modified Ricker model does not require the exclusion of data points from the fit, as necessary when fitting the 2nd-order polynomial model making it more reliable and robust against outliers, and, in addition, provides a measure for the noise of the recorded results.

Full-field electroretinography, visual acuity and visual fields in Usher syndrome: a multicentre European study.

PURPOSE: Usher syndrome (USH) is a multisensory deficiency involving vision, hearing and the vestibular system. The purpose of this study is to report on the functional data (i.e. electroretinography, visual fields, visual acuity) of patients with retinitis pigmentosa (RP) due to Usher syndrome that were collected in a multicentre European study (TREATRUSH). METHODS: A total of 268 genetically confirmed USH patients underwent electrophysiological examinations in the context of multimodal ophthalmological examination in the study (75 USH1, 189 USH2 and four USH3). Full-field electroretinography (ERG) was performed according to ISCEV standards, visual field determination was carried out with either the Octopus or Goldmann perimeters and visual acuity was examined with either ETDRS or Snellen charts. The data were compared between USH subtypes (USH1/USH2/USH3) and correlated with age. RESULTS: Visual acuity decreases significantly with age for both USH1 and USH2 (p < 0.001), without a difference between the two cohorts. When corrected for age, the preserved kinetic visual field was significantly larger in USH2 than in USH1 (p = 0.04). Furthermore, the preserved kinetic visual field area showed a significant decrease with age (based on an exponential fit) in both USH1 and USH2 (p < 0.001). In USH1 patients, however, the visual field was already vastly reduced at an early age. The ERG results were abnormal in all patients. Detectable data for scotopic ERG were obtained from nine patients, and data of photopic ERG were obtained from 24 patients, without a difference between USH1 and USH2 subtypes. CONCLUSIONS: There are differences in the phenotypes of RP in USH subtypes, most visible in the progression of visual fields between USH1 and USH2. The perimetric reduction occurs earlier in USH1 than in USH2. In both subtypes, visual acuity decreases significantly with age and the ERG is not detectable already at early ages.

Novel homozygous large deletion including the 5' part of the SPATA7 gene in a consanguineous Israeli Muslim Arab family.

PURPOSE: To identify the genetic defect in a consanguineous Israeli Muslim Arab family with juvenile retinitis pigmentosa (RP). METHODS: DNA samples were collected from the index patient, her parents, her affected sister, and two non-affected siblings. Genome-wide linkage analysis with 250 K single nucleotide polymorphism (SNP) arrays was performed using DNA from the two affected patients. Owing to consanguinity in the family, we applied homozygosity mapping to identify the disease-causing gene. The candidate gene SPATA7 was screened for mutations with PCR amplifications and direct Sanger sequencing. RESULTS: Following high-density SNP arrays, we identified several homozygous genomic regions one of which included the SPATA7 gene. Several mutations in SPATA7 have been reported for various forms of retinal dystrophy, including Leber congenital amaurosis (LCA) and juvenile RP. PCR-based sequence content mapping, long-distance PCR amplifications, and subsequent sequencing analysis revealed a homozygous 63.4 kb large deletion that encompasses the 5' part of the SPATA7 gene including exons 1-5. The mutation showed concordant segregation with the phenotype in the family as expected for autosomal recessive mode of inheritance and is consistent with a diagnosis of juvenile RP. CONCLUSIONS: We report a novel homozygous large deletion in SPATA7 associated with juvenile RP in a consanguineous Israeli Muslim Arab family. This is the first larger deletion mutation reported for SPATA7.

Ophthalmological assessment of cannabis-induced persisting perception disorder: is there a direct retinal effect?

PURPOSE: Cannabis is a psychotomimetic agent that induces impairment of sensory perception. We present detailed clinical and electrophysiological data of patients with hallucinogen persisting perception disorder (HPPD) after marijuana consumption. METHODS: A HPPD patient and four heavy cannabis smokers with no visual disturbances (controls) underwent complete ophthalmological examination including psychophysical tests (visual acuity, color vision, visual field, and dark adaptation) and detailed electrophysiological examinations, including extended Ganzfeld ERG, multifocal ERG, and electrooculography (EOG). Furthermore, electrically evoked phosphene thresholds (EPTs) were measured to further evaluate retinal function. RESULTS: Ophthalmological and most electrophysiological examinations were within normal limits for the HPPD patient and for all control subjects. Interestingly, EOG results of the HPPD patient showed a slightly reduced fast oscillation ratio, diminished standing potentials of the slow oscillations, and a light peak within normal range resulting in higher Arden ratios. The EPTs of the patient were reduced, in particular for pulses with long durations (50 ms) causing visual sensations even at lowest possible currents of the neurostimulator. The control subjects did not reveal such alterations. CONCLUSIONS: Our findings suggest a direct effect of cannabinoids on the retina and retinal pigment epithelium function, which may be involved in disturbances of the visual function experienced after drug consumption. The observations presented here may contribute to the elucidation of the detailed mechanism. Furthermore, EOG and EPT measurements may be useful tools to demonstrate long-term retinal alterations in cannabis-induced HPPD in patients.

Achromatopsia: on the doorstep of a possible therapy.

Achromatopsia (ACHM) is a rare autosomal recessive inherited retinal disorder with an incidence of approximately 1 in 30,000. It presents at birth or early infancy and is typically characterized by reduced visual acuity, nystagmus, photophobia, and very poor or absent color vision. The symptoms arise from isolated cone dysfunction, which can be caused by mutations in the crucial components of the cone phototransduction cascade. Although ACHM is considered a functionally nonprogressive disease affecting only the cone system, recent studies have described progressive age-dependent changes in retinal architecture. Currently, no specific therapy is available for ACHM; however, gene replacement therapy performed on animal models for three ACHM genes has shown promising results. Accurate genetic and clinical diagnosis of patients may therefore enhance and enable therapeutic intervention in the near future. This short review summarizes the genetic background, pathophysiology, clinical findings, diagnostics, and therapeutic perspectives in ACHM.

Homozygosity mapping reveals new nonsense mutation in the FAM161A gene causing autosomal recessive retinitis pigmentosa in a Palestinian family.

PURPOSE: Retinitis pigmentosa (RP) is a heterogenous group of inherited retinal degenerations caused by mutations in at least 45 genes. Recently, the FAM161A gene was identified as the causative gene for RP28, an autosomal recessive form of RP. METHODS: We performed a clinical and molecular genetic study of a consanguineous Palestinian family with two three siblings affected with retinitis pigmentosa. DNA samples were collected from the index patient, his father, his affected sister, and two non-affected brothers. DNA sample from the index was subjected to high resolution genome-wide SNP array. Assuming identity-by-descent in this consanguineous family we applied homozygosity mapping to identify disease causing genes. RESULTS: The index patient reported night blindness since the age of 20 years, followed by moderate disease progression with decrease of peripheral vision, the development of photophobia and later on reduced central vision. At the age of 40 his visual acuity was counting fingers (CF) for both eyes, color discrimination was not possible and his visual fields were severely constricted. Funduscopic examination revealed a typical appearance of advanced RP with optic disc pallor, narrowed retinal vessels, bone-spicule like pigmentary changes in the mid-periphery and atrophic changes in the macula. His younger affected brother (37 years) was reported with overall milder symptoms, while the youngest sister (21 years) reported problems only with night vision. Applying high-density SNP arrays we identified several homozygous genomic regions one of which included the recently identified FAM161A gene mutated in RP28-linked autosomal recessive RP. Sequencing analysis revealed the presence of a novel homozygous nonsense mutation, c.1003C>T/p.R335X in the index patient and the affected sister. CONCLUSION: We identified an RP28-linked RP family in the Palestinian population caused by a novel nonsense mutation in FAM161A. RP in this family shows a typical disease onset with moderate to rapid progression into severe visual impairment including central vision in the index and overall milder symptoms in the younger brother and sister.

Novel compound heterozygous NMNAT1 variants associated with Leber congenital amaurosis.

PURPOSE: The gene encoding nicotinamide nucleotide adenylyltransferase 1 (NMNAT1) was recently found to be mutated in a subset of patients with Leber congenital amaurosis (LCA) with macular atrophy. The aim of this study was to determine the occurrence and frequency of NMNAT1 mutations and associated phenotypes in different types of inherited retinal dystrophies. METHODS: DNA samples of 161 patients with LCA without genetic diagnosis were analyzed for variants in NMNAT1 using Sanger sequencing. Variants in exon 5 of NMNAT1, which harbors the majority of the previously identified mutations, were screened in 532 additional patients with retinal dystrophies. This cohort encompassed 108 persons with isolated or autosomal recessive cone-rod dystrophy (CRD), 271 with isolated or autosomal recessive retinitis pigmentosa (RP), and 49 with autosomal dominant RP, as well as 104 persons with LCA in whom the causative mutation was previously identified. RESULTS: Compound heterozygous alterations were found in six patients with LCA and in one person with early-onset RP. All except one carried the common p.E257K variant on one allele. Macular atrophy was absent in one patient, who carried this variant in combination with a truncating mutation on the other allele. The p.E257K alteration was also found in a heterozygous state in five individuals with LCA and one with RP while no mutation was detected on the other allele. Two individuals with LCA carried other NMNAT1 variants in a heterozygous state, whereas no NMNAT1 variants in exon 5 were identified in individuals with CRD. The p.E257K variant was found to be enriched in a heterozygous state in individuals with LCA (0.94%) compared to Caucasian controls (0.18%), although the difference was statistically insignificant (p=0.12). CONCLUSIONS: Although macular atrophy can occur in LCA and CRD, no NMNAT1 mutations were found in the latter cohort. NMNAT1 variants were also not found in a large group of patients with sporadic or autosomal recessive RP. The enrichment of p.E257K in a heterozygous state in patients with LCA versus controls suggests that this allele could act as a modifier in other genetic subtypes of LCA.

The progressive rod-cone degeneration (PRCD) protein is secreted through the conventional ER/Golgi-dependent pathway.

Retinitis pigmentosa (RP) is the most common form of hereditary retinal degeneration. Mutations of the PRCD gene are associated with RP in both dogs and humans. To date, four distinct PRCD mutations have been reported worldwide. Here we report the clinical phenotype of another patient with PRCD mutations, carrying the known p.R18X mutation and a novel missense mutation, p.P25T. This mutation affects a highly conserved amino acid, is predicted to be damaging by several prediction tools, and was not found in the public databases or in 115 ethnically-matched control individuals. The phenotype of this patient resembles that of previously reported patients with PRCD mutations, including bull's eye maculopathy, which appears to be a hallmark of the PRCD-induced phenotype. PRCD encodes for a 54 amino acids long protein with unknown function. The first 20 amino acids appear to encode for a signal peptide (SP), suggesting that PRCD is a secreted protein. To study PRCD secretion, C-terminally myc-tagged PRCD was expressed in cultured cells. Cells and conditioned media were analyzed by Western blot. PRCD was found in both cell extracts and media. However, a truncated PRCD protein lacking the first 20 amino acids was present only in cell extracts and not in media, confirming that PRCD extracellular secretion is mediated by its N-terminal SP. To characterize the secretory pathway of PRCD, various pharmacological agents which interfere with transport of proteins through the ER and Golgi to the plasma membrane were used. PRCD secretion was significantly inhibited by all tested pharmacological agents, confirming that it is secreted through the classic ER/Golgi-dependent secretory pathway. We tested the effect of two mutations on the PRCD protein, and found that p.C2Y, but not p.P25T, affects protein stability, and that neither mutation affects secretion. Our data suggest that PRCD functions as a secreted protein. These findings shed a new light on PRCD function and the etiology of RP.

Early postoperative changes of the foveal surface in epiretinal membranes: comparison of 23-gauge macular surgery with air vs. balanced salt solution.

OBJECTIVE: To analyze the foveal surface using binary image analysis after spectral-domain optical coherence tomography (SD-OCT) following 23-gauge macular surgery in epiretinal membranes (ERM) using either air tamponade (AIR) or balanced salt solution (BSS). METHODS: One hundred twenty-four eyes (124 patients) with ERM that had undergone membrane peeling with installation of air or BSS were analyzed retrospectively. Ophthalmic examination was performed at baseline and 3 months. OUTCOME MEASURES: The foveal area and surface symmetry, area matched thickness, area matched contour, and best-corrected visual acuity (BCVA). The OCT images were analyzed after binary conversion with ImageJ software. RESULTS: Eighty eyes (80 patients) of 124 screened patients were included (AIR group: 39 patients, BSS group: 41 patients). Median follow-up time was 14 weeks (range, 9-19 weeks). Three months after surgery, the median horizontal area decreased significantly in both groups (p < 0.0001). At follow-up, the foveal surface symmetry values for the BSS group (median, 22.73 µm, range, 0-153) were significantly lower than for the AIR group (median, 23.95 µm, range, 0-160.43) (p < 0.0001). The area-matched thickness increased significantly in both groups (p < 0.001). The AIR group showed a significant increase of the area matched contour for the nasal located measurement-areas N1 (p < 0.0003), N2 (p < 0.0079), N3 (p < 0.007). The BSS group showed a significant increase of the area-matched contour for the measurement areas N1 (p < 0.019), N2 (p < 0.0014), and N4 (p < 0.022). After surgery, median BCVA for both groups increased significantly to 0.3 logMAR. CONCLUSIONS: The analysis of early contour changes after ERM surgery was technically possible. Long-term data have to be looked at before the clinical impact of this methodology can be estimated. Although there were no big differences between both groups (AIR vs. BSS), this could change within a longer and more representative follow-up.

GUCY2D- or GUCA1A-related autosomal dominant cone-rod dystrophy: is there a phenotypic difference?

PURPOSE: To compare the phenotype of patients with heterozygous mutation in GUCY2D or GUCA1A causing autosomal dominant cone or cone-rod dystrophies. METHODS: Five patients from one family with GUCA1A and nine patients from four families with GUCY2D mutations were included. Psychophysical and electrophysiological examinations were performed to study retinal function. Fundus autofluorescence imaging and spectral domain optical coherence tomography were performed for morphologic characterization. RESULTS: Genetic analysis revealed the mutation c.451C>T (p.L151F) in the GUCA1A family. In the GUCY2D group, c.2512C>T (p.R838C) was the most frequent (2 families), c.2512C>G (p.R838G) and c.2513G>A (p.R838H) were found in one family each. Visual acuity was reduced to 0.04 to 0.7 in GUCA1A and to 0.014 to 0.5 in patients with GUCY2D. Dark adaptation showed elevated thresholds in the GUCY2D group. Scotopic electroretinography revealed a tendency to a more affected rod function in the GUCY2D group. Photopic electroretinography showed residual or absent responses in both groups. Fundus alterations were confined to the macula in both groups. CONCLUSION: GUCA1A and GUCY2D mutations are both accompanied by similar pattern of generalized cone dysfunction with a tendency to less involvement of the rod photoreceptors and a less severe phenotype in patients with GUCA1A.

Visual cortex organisation in a macaque monkey with macular degeneration.

The visual field is retinotopically represented in early visual areas. It has been suggested that when adult primary visual cortex (V1) is deprived of normal retinal input it is capable of large-scale reorganisation, with neurons inside the lesion projection zone (LPZ) being visually driven by inputs from intact retinal regions. Early functional magnetic resonance imaging (fMRI) studies in humans with macular degeneration (MD) report > 1 cm spread of activity inside the LPZ border, whereas recent results report no shift of the LPZ border. Here, we used fMRI population receptive field measurements to study, for the first time, the visual cortex organisation of one macaque monkey with MD and to compare it with normal controls. Our results showed that the border of the V1 LPZ remained stable, suggesting that the deafferented area V1 zone of the MD animal has limited capacity for reorganisation. Interestingly, the pRF size of non-deafferented V1 voxels increased slightly (~20% on average), although this effect appears weaker than that in previous single-unit recording reports. Area V2 also showed limited reorganisation. Remarkably, area V5/MT of the MD animal showed extensive activation compared to controls stimulated over the part of the visual field that was spared in the MD animal. Furthermore, population receptive field size distributions differed markedly in area V5/MT of the MD animal. Taken together, these results suggest that V5/MT has a higher potential for reorganisation after MD than earlier visual cortex.

Identification of a novel mutation in the PRCD gene causing autosomal recessive retinitis pigmentosa in a Turkish family.

PURPOSE: Progressive rod-cone degeneration (PRCD) is a canine form of autosomal recessive photoreceptor degeneration and serves as an animal model for human retinitis pigmentosa (RP). To date, only two RP-causing mutations of the PRCD gene have been reported in humans. We found a novel mutation in PRCD (c.52C>T, p.R18X) in three siblings affected by RP and present detailed morphologic and functional parameters. METHODS: A complete ophthalmological examination was performed including psychophysical tests (best-corrected visual acuity, Lanthony Panel D-15 color vision test, and visual field) and electrophysiology (ganzfeld and multifocal electroretinogram). Additionally, color and infrared fundus photography, autofluorescence, and spectral domain optical coherence tomography recordings were performed. Genomic DNA of the three affected individuals was analyzed with high-throughput sequencing for all RP-related genes in a diagnostic set-up. RESULTS: We identified a novel homozygous mutation in PRCD (c.52C>T, p.R18X) with diagnostic high-throughput panel sequencing. All three patients showed an advanced stage of retinitis pigmentosa with reduced visual acuity (mean: 20/80), small residual visual fields (mean for target III4e: 1134.35 deg²), and non-detectable electrophysiological responses. Myopia, posterior subcapsular cataract, bone spicule-like pigmentation, and attenuated arterioles were typical findings. Interestingly, bull's eye maculopathy due to patchy retinal pigment epithelium atrophy was also present in all patients. The mean central retinal thickness observed in optical coherence tomography was 148 µm. CONCLUSIONS: The identification of a third mutation in PRCD confirms its role in the pathogenesis of RP. Clinical findings were in line with the morphological changes observed in previous studies. Bull's eye maculopathy seems to be a hallmark of RP due to mutations in the PRCD gene.

Detailed functional and structural characterization of a macular lesion in a rhesus macaque.

PURPOSE: Animal models are powerful tools to broaden our understanding of disease mechanisms and to develop future treatment strategies. Here we present detailed structural and functional findings of a rhesus macaque suffering from a naturally occurring bilateral macular dystrophy (BMD), partial optic atrophy and corresponding reduction of central V1 signals in visual fMRI experiments when compared to data in a healthy macaque (CTRL) of similar age. METHODS: Retinal imaging included infrared and autofluorescence recordings, fluorescein and indocyanine green angiography and spectral domain optical coherence tomography (OCT) on the Spectralis HRA + OCT platform. Electroretinography included multifocal and Ganzfeld-ERG recordings. Animals were killed and eyes analyzed by immunohistochemistry. RESULTS: Angiography showed reduced macular vascularization with significantly larger foveal avascular zones (FAZ) in the affected animal (FAZBMD = 8.85 mm(2) vs. FAZCTRL = 0.32 mm(2)). OCT showed bilateral thinning of the macula within the FAZ (total retinal thickness, TRTBMD = 174 ± 9 µm) and partial optic nerve atrophy when compared to control (TRTCTRL = 303 ± 45 µm). Segmentation analysis revealed that inner retinal layers were primarily affected (inner retinal thickness, IRTBMD = 33 ± 9 µm vs. IRTCTRL = 143 ± 45 µm), while the outer retina essentially maintained its thickness (ORTBMD = 141 ± 7 µm vs. ORTCTRL = 160 ± 11 µm). Altered macular morphology corresponded to a preferential reduction of central signals in the multifocal electroretinography and to a specific attenuation of cone-derived responses in the Ganzfeld electroretinography, while rod function remained normal. CONCLUSION: We provided detailed characterization of a primate macular disorder. This study aims to stimulate awareness and further investigation in primates with macular disorders eventually leading to the identification of a primate animal model and facilitating the preclinical development of therapeutic strategies.

Central Visual Function and Genotype-Phenotype Correlations in PDE6A-Associated Retinitis Pigmentosa.

Purpose: Autosomal recessive retinitis pigmentosa (arRP) can be caused by mutations in the phosphodiesterase 6A (PDE6A) gene. Here, we describe the natural course of disease progression with respect to central retinal function (i.e., visual acuity, contrast sensitivity, and color vision) and establish a detailed genotype--phenotype correlation. Methods: Forty-four patients (26 females; mean age ± SD, 43 ± 13 years) with a confirmed genetic diagnosis of PDE6A-associated arRP underwent comprehensive ophthalmological examinations including best-corrected visual acuity (BCVA) with Early Treatment Diabetic Retinopathy Study charts, contrast sensitivity (CS) with Pelli-Robson charts at distances of 3 m and 1 m, and color vision testing using Roth 28-Hue and Panel D-15 saturated color cups. Results: The most frequently observed variants were c.998+1G>A/p.?, c.304C>A/p.R102S, and c.2053G>A/p.V685M. Central retinal function in patients homozygous for variant c.304C>A/p.R102S was better when compared to patients homozygous for variant c.998+1G>A/p.?, although the former were older at baseline. Central retinal function was similar in patients homozygous for variant c.304C>A/p.R102S and patients heterozygous for variants c.304C>A/p.R102S and c.2053G>A/p.V685M, although the latter were younger at baseline. Annual decline rates in central retinal function were small. Conclusions: We conclude that the severity of the different disease-causing PDE6A mutations in humans with respect to central visual function may be ranked as follows: c.2053G>A/p.V685M in homozygous state (most severe) > c.998+1G>A/p.? in homozygous state > c.304C>A/p.R102S and c.2053G>A/p.V685M in compound-heterozygous state > c.304C>A/p.R102S in homozygous state (mildest). The assessment of treatment efficacy in interventional trials will remain challenging due to small annual decline rates in central retinal function.