ResearchPad - retinitis-pigmentosa https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[The qualitative assessment of optical coherence tomography and the central retinal sensitivity in patients with retinitis pigmentosa]]> https://www.researchpad.co/article/elastic_article_7697 To analyze the relationships between qualitative and quantitative parameters of spectral-domain optical coherence tomography (SD-OCT) and the central retinal sensitivity in patients with retinitis pigmentosa (RP).Materials and methodsNinety-three eyes of 93 patients were finally enrolled, with a median age (quartile) of 58 (24.5) years. We assessed the patients using SD-OCT and the 10–2 program of a Humphry Field Analyzer (HFA). As a qualitative parameter, two graders independently classified the patients’ SD-OCT images into five severity grades (grades 1–5) based on the severity of damage to the photoreceptor inner and outer segments (IS/OS) layer. As quantitative parameters, we measured the IS-ellipsoid zone (IS-EZ) width, IS/OS thickness, outer nuclear layer (ONL) thickness, central macular thickness (CMT, 1 and 3 mm) and macular cube (6 × 6 mm) volume and thickness. The central retinal sensitivity was defined by the best-corrected visual acuity (BCVA; logMAR), average sensitivities of the central 4 (foveal sensitivity [FS]) and 12 (macular sensitivity [MS]) points of the HFA 10–2 program and the mean deviation (MD) of the 10–2 program. Spearman’s correlation was used to assess the association between both qualitative and quantitative parameters and variables of the central retinal sensitivity. In addition, we performed a multiple regression analysis using these parameters to identify the parameters most strongly influencing the central retinal sensitivity.ResultsThe IS/OS severity grade was significantly correlated with the BCVA (ρ = 0.741, P < 0.001), FS (ρ = −0.844, P < 0.001), MS (ρ = −0.820, P < 0.001) and MD (ρ = −0.681, P < 0.001) and showed stronger correlations to them than any other quantitative parameters including the IS-EZ width, IS/OS thickness, ONL thickness, CMTs and macular cube volume/thickness. Furthermore, a step-wise multiple regression analysis indicated that the IS/OS severity grade was more strongly associated with the BCVA (β = 0.659, P < 0.001), FS (β = −0.820, P < 0.001), MS (β = −0.820, P < 0.001) and MD (β = −0.674, P < 0.001) than any other quantitative parameters. The intraclass correlation coefficient between two graders indicated substantial correlation (κ = 0.70).DiscussionThe qualitative grading of OCT based on the severity of the IS/OS layer was simple and strongly correlated with the central retinal sensitivity in patients with RP. It may be useful to assess the central visual function in patients with RP, although there is some variation in severity within the same severity grade. ]]> <![CDATA[The findings of optical coherence tomography of retinal degeneration in relation to the morphological and electroretinographic features in RPE65−/− mice]]> https://www.researchpad.co/article/5c59ff13d5eed0c484135aa6

Purpose

Mutations of the gene encoding RPE65 cause Leber congenital amaurosis (LCA) retinitis pigmentosa (RP). The optical coherence tomography (OCT) is increasingly utilized to noninvasively evaluate various types of retinal diseases, including RP. The present study was conducted to characterize the OCT findings of the RPE65−/− mice—an animal model of LCA and RP—in relation to the morphological features based on histological and electron microscopic findings as well as electroretinography (ERG) features.

Materials and methods

RPE65−/− mice were employed as a model of retinal degeneration. C57BL/6J mice were used as a wild-type control. OCT was performed on the RPE65−/− mice from postnatal day (P) 22 to 170. The longitudinal changes in the OCT images and fundus pictures were analyzed both qualitatively and quantitatively in comparison to those of C57BL/6J mice. The OCT images were also compared to the histological and electron microscopic findings. Full field combined rod and cone ERG was performed to analyze the relationship between morphology based on OCT and the amplitudes of the a- and b-waves.

Results

In the RPE65−/− mice, the photoreceptor rod and cone layer appeared as a diffuse hyperreflective zone contiguous with the inner segment ellipsoid zone (IS-EZ) on OCT, even on P22, whereas the IS-EZ and interdigitation zone were clearly identified in the age-matched C57BL/6J mice. The histological analyses revealed that the regular arrangement of the photoreceptor inner and outer segments was gradually lost in the RPE65-/- mice. On electron microscopy, most of the rod outer segments were degenerated from P21 to P35, whereas outer segments became variably shorter after P49 although ultrastructure appeared to normalize. The thickness of the outer nuclear layer of RPE65−/− mice was slowly and progressively reduced in comparison to C57BL/6J mice. Although the thickness of the inner and outer segment layer of RPE65−/− mice was significantly decreased in comparison to C57BL/6J mice, the change was not progressive, at least until P170. Even at P35, the amplitudes of both a- and b-waves on ERG were severely deteriorated in comparison to those of C57BL/6J mice. Mottled depigmented spots appeared throughout the fundus in RPE65−/− mice after P72, and were detected as hyperreflective deposits under the retinal pigment epithelium on OCT.

Discussion

The pathological changes in the inner and outer segments layer of RPE65−/− mice were identified as diffuse hyperreflective changes on OCT. The rod outer segments showed degeneration in the early postnatal periods but became morphologically normalized in the disc structure after P49, although the sizes of the length of the rod outer segments were variable. OCT could not qualitatively differentiate the early degeneration of rods from the late variability in size of rods. Although the morphology of the photoreceptor outer segments was relatively preserved in the RPE65−/− mice, the amplitudes of ERG were severely disturbed. These structural and functional deficits may be derived from the defective supply of 11-cis-retinol to the photoreceptors.

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<![CDATA[Molecular and clinical analysis of 27 German patients with Leber congenital amaurosis]]> https://www.researchpad.co/article/5c269745d5eed0c48470f15a

Leber congenital amaurosis (LCA) is the earliest and most severe form of all inherited retinal dystrophies (IRD) and the most frequent cause of inherited blindness in children. The phenotypic overlap with other early-onset and severe IRDs as well as difficulties associated with the ophthalmic examination of infants can complicate the clinical diagnosis. To date, 25 genes have been implicated in the pathogenesis of LCA. The disorder is usually inherited in an autosomal recessive fashion, although rare dominant cases have been reported. We report the mutation spectra and frequency of genes in 27 German index patients initially diagnosed with LCA. A total of 108 LCA- and other genes implicated in IRD were analysed using a cost-effective targeted next-generation sequencing procedure based on molecular inversion probes (MIPs). Sequencing and variant filtering led to the identification of putative pathogenic variants in 25 cases, thereby leading to a detection rate of 93%. The mutation spectrum comprises 34 different alleles, 17 of which are novel. In line with previous studies, the genetic results led to a revision of the initial clinical diagnosis in a substantial proportion of cases, demonstrating the importance of genetic testing in IRD. In addition, our detection rate of 93% shows that MIPs are a cost-efficient and sensitive tool for targeted next-generation sequencing in IRD.

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<![CDATA[First-in-Human Trial of a Novel Suprachoroidal Retinal Prosthesis]]> https://www.researchpad.co/article/5989db10ab0ee8fa60bcbf1b

Retinal visual prostheses (“bionic eyes”) have the potential to restore vision to blind or profoundly vision-impaired patients. The medical bionic technology used to design, manufacture and implant such prostheses is still in its relative infancy, with various technologies and surgical approaches being evaluated. We hypothesised that a suprachoroidal implant location (between the sclera and choroid of the eye) would provide significant surgical and safety benefits for patients, allowing them to maintain preoperative residual vision as well as gaining prosthetic vision input from the device. This report details the first-in-human Phase 1 trial to investigate the use of retinal implants in the suprachoroidal space in three human subjects with end-stage retinitis pigmentosa. The success of the suprachoroidal surgical approach and its associated safety benefits, coupled with twelve-month post-operative efficacy data, holds promise for the field of vision restoration.

Trial Registration

Clinicaltrials.gov NCT01603576

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<![CDATA[Structure-Function Modeling of Optical Coherence Tomography and Standard Automated Perimetry in the Retina of Patients with Autosomal Dominant Retinitis Pigmentosa]]> https://www.researchpad.co/article/5989daa3ab0ee8fa60ba6815

Purpose

To assess relationships between structural and functional biomarkers, including new topographic measures of visual field sensitivity, in patients with autosomal dominant retinitis pigmentosa.

Methods

Spectral domain optical coherence tomography line scans and hill of vision (HOV) sensitivity surfaces from full-field standard automated perimetry were semi-automatically aligned for 60 eyes of 35 patients. Structural biomarkers were extracted from outer retina b-scans along horizontal and vertical midlines. Functional biomarkers were extracted from local sensitivity profiles along the b-scans and from the full visual field. These included topographic measures of functional transition such as the contour of most rapid sensitivity decline around the HOV, herein called HOV slope for convenience. Biomarker relationships were assessed pairwise by coefficients of determination (R2) from mixed-effects analysis with automatic model selection.

Results

Structure-function relationships were accurately modeled (conditional R2>0.8 in most cases). The best-fit relationship models and correlation patterns for horizontally oriented biomarkers were different than vertically oriented ones. The structural biomarker with the largest number of significant functional correlates was the ellipsoid zone (EZ) width, followed by the total photoreceptor layer thickness. The strongest correlation observed was between EZ width and HOV slope distance (marginal R2 = 0.85, p<10−10). The mean sensitivity defect at the EZ edge was 7.6 dB. Among all functional biomarkers, the HOV slope mean value, HOV slope mean distance, and maximum sensitivity along the b-scan had the largest number of significant structural correlates.

Conclusions

Topographic slope metrics show promise as functional biomarkers relevant to the transition zone. EZ width is strongly associated with the location of most rapid HOV decline.

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<![CDATA[Retinal Diseases Caused by Mutations in Genes Not Specifically Associated with the Clinical Diagnosis]]> https://www.researchpad.co/article/5989da0fab0ee8fa60b78ec0

Purpose

When seeking a confirmed molecular diagnosis in the research setting, patients with one descriptive diagnosis of retinal disease could carry pathogenic variants in genes not specifically associated with that description. However, this event has not been evaluated systematically in clinical diagnostic laboratories that validate fully all target genes to minimize false negatives/positives.

Methods

We performed targeted next-generation sequencing analysis on 207 ocular disease-related genes for 42 patients whose DNA had been tested negative for disease-specific panels of genes known to be associated with retinitis pigmentosa, Leber congenital amaurosis, or exudative vitreoretinopathy.

Results

Pathogenic variants, including single nucleotide variations and copy number variations, were identified in 9 patients, including 6 with variants in syndromic retinal disease genes and 3 whose molecular diagnosis could not be distinguished easily from their submitted clinical diagnosis, accounting for 21% (9/42) of the unsolved cases.

Conclusion

Our study underscores the clinical and genetic heterogeneity of retinal disorders and provides valuable reference to estimate the fraction of clinical samples whose retinal disorders could be explained by genes not specifically associated with the corresponding clinical diagnosis. Our data suggest that sequencing a larger set of retinal disorder related genes can increase the molecular diagnostic yield, especially for clinically hard-to-distinguish cases.

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<![CDATA[Increased Risk of Acute Angle Closure in Retinitis Pigmentosa: A Population-Based Case-Control Study]]> https://www.researchpad.co/article/5989da34ab0ee8fa60b857e9

Purpose

To investigate the association between retinitis pigmentosa (RP) and acute angle closure during a 15-year follow-up period.

Methods

Using the Taiwan Longitudinal Health Insurance Database 2000, we identified 382 RP patients based on the diagnostic code of RP (International Classification of Diseases, 9th Revision, Clinical Modification (ICD-9-CM) 362.74) made during 1996–2010, excluding subjects under age of 20 years at diagnosis or subjects undergoing lens extraction before the index date. The control group included 3820 randomly selected non-RP subjects matched with the RP patients in age, gender and the index date of diagnosis. The incidence of acute angle closure during the study period was observed based on an ICD-9-CM code of 365.22. Cochran-Mantel-Haenszel test was used to determine the odds ratio (OR) of having acute angle closure in RP patients.

Results

The mean age at the diagnosis of RP was 51.1years (standard deviation [SD] 16.7). Acute angle closure occurred in 5 RP patients (1.3%) and in 15 controls (0.4%). The mean age with the acute angle closure was 53.3 years (SD 8.0) in RP patients and 64.6 years (SD 8.4) in controls (P = 0.015). After adjusting for age, gender and comorbid disorders, RP patients had 3.64-fold (95% confidence interval [CI], 1.29–10.25, P<0.001) greater odds of having acute angle closure. After stratification for gender and age, the risk of acute angle closure in RP was higher in patients under age of 60 years (adjusted OR 11.84; 95% CI, 2.84–49.48) and male patients (adjusted OR 19.36; 95% CI, 3.43–109.40)(both P = 0.001).

Conclusions

RP patients had increased risk of acute angle closure than controls. Contrary to the fact that angle closure disease is more prevalent in elderly females in general population, acute angle closure attack occurred earlier in life and the risk was higher in males among RP patients.

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<![CDATA[Retinotopic Distribution of Structural and Functional Damages following Bright Light Exposure of Juvenile Rats]]> https://www.researchpad.co/article/5989da5bab0ee8fa60b8ffe8

In the present study, we aimed at better understanding the short (acute) and long term (chronic) degenerative processes characterizing the juvenile rat model of light-induced retinopathy. Electroretinograms, visual evoked potentials (VEP), retinal histology and western blots were obtained from juvenile albino Sprague-Dawley rats at preselected postnatal ages (from P30 to P400) following exposure to 10,000 lux from P14 to P28. Our results show that while immediately following the cessation of exposure, photoreceptor degeneration was concentrated within a well delineated area of the superior retina (i.e. the photoreceptor hole), with time, this hole continued to expand to form an almost photoreceptor-free region covering most of superior-inferior axis. By the end of the first year of life, only few photoreceptors remained in the far periphery of the superior hemiretina. Interestingly, despite a significant impairment of the outer retinal function, the retinal output (VEP) was maintained in the early phase of this retinopathy. Our findings thus suggest that postnatal exposure to a bright luminous environment triggers a degenerative process that continues to impair the retinal structure and function (mostly at the photoreceptor level) long after the cessation of the exposure regimen (more than 1 year documented herein). Given the slow degenerative process triggered following postnatal bright light exposure, we believe that our model represents an attractive alternative (to other more genetic models) to study the pathophysiology of photoreceptor-induced retinal degeneration as well as therapeutic strategies to counteract it.

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<![CDATA[Targeted Next-Generation Sequencing Reveals Novel USH2A Mutations Associated with Diverse Disease Phenotypes: Implications for Clinical and Molecular Diagnosis]]> https://www.researchpad.co/article/5989daefab0ee8fa60bc0970

USH2A mutations have been implicated in the disease etiology of several inherited diseases, including Usher syndrome type 2 (USH2), nonsyndromic retinitis pigmentosa (RP), and nonsyndromic deafness. The complex genetic and phenotypic spectrums relevant to USH2A defects make it difficult to manage patients with such mutations. In the present study, we aim to determine the genetic etiology and to characterize the correlated clinical phenotypes for three Chinese pedigrees with nonsyndromic RP, one with RP sine pigmento (RPSP), and one with USH2. Family histories and clinical details for all included patients were reviewed. Ophthalmic examinations included best corrected visual acuities, visual field measurements, funduscopy, and electroretinography. Targeted next-generation sequencing (NGS) was applied using two sequence capture arrays to reveal the disease causative mutations for each family. Genotype-phenotype correlations were also annotated. Seven USH2A mutations, including four missense substitutions (p.P2762A, p.G3320C, p.R3719H, and p.G4763R), two splice site variants (c.8223+1G>A and c.8559-2T>C), and a nonsense mutation (p.Y3745*), were identified as disease causative in the five investigated families, of which three reported to have consanguineous marriage. Among all seven mutations, six were novel, and one was recurrent. Two homozygous missense mutations (p.P2762A and p.G3320C) were found in one individual family suggesting a potential double hit effect. Significant phenotypic divergences were revealed among the five families. Three families of the five families were affected with early, moderated, or late onset RP, one with RPSP, and the other one with USH2. Our study expands the genotypic and phenotypic variability relevant to USH2A mutations, which would help with a clear insight into the complex genetic and phenotypic spectrums relevant to USH2A defects, and is complementary for a better management of patients with such mutations. We have also demonstrated that a targeted NGS approach is a valuable tool for the genetic diagnosis of USH2 and RP.

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<![CDATA[Alternative Isoform Analysis of Ttc8 Expression in the Rat Pineal Gland Using a Multi-Platform Sequencing Approach Reveals Neural Regulation]]> https://www.researchpad.co/article/5989da2bab0ee8fa60b829fb

Alternative isoform regulation (AIR) vastly increases transcriptome diversity and plays an important role in numerous biological processes and pathologies. However, the detection and analysis of isoform-level differential regulation is difficult, particularly in the face of complex and incompletely-annotated transcriptomes. Here we have used Illumina short-read/high-throughput RNA-Seq to identify 55 genes that exhibit neurally-regulated AIR in the pineal gland, and then used two other complementary experimental platforms to further study and characterize the Ttc8 gene, which is involved in Bardet-Biedl syndrome and non-syndromic retinitis pigmentosa. Use of the JunctionSeq analysis tool led to the detection of several novel exons and splice junctions in this gene, including two novel alternative transcription start sites which were found to display disproportionately strong neurally-regulated differential expression in several independent experiments. These high-throughput sequencing results were validated and augmented via targeted qPCR and long-read Pacific Biosciences SMRT sequencing. We confirmed the existence of numerous novel splice junctions and the selective upregulation of the two novel start sites. In addition, we identified more than 20 novel isoforms of the Ttc8 gene that are co-expressed in this tissue. By using information from multiple independent platforms we not only greatly reduce the risk of errors, biases, and artifacts influencing our results, we also are able to characterize the regulation and splicing of the Ttc8 gene more deeply and more precisely than would be possible via any single platform. The hybrid method outlined here represents a powerful strategy in the study of the transcriptome.

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<![CDATA[Reshaping the Cone-Mosaic in a Rat Model of Retinitis Pigmentosa: Modulatory Role of ZO-1 Expression in DL-Alpha-Aminoadipic Acid Reshaping]]> https://www.researchpad.co/article/5989db04ab0ee8fa60bc7a4b

In S334ter-line-3 rat model of Retinitis Pigmentosa (RP), rod cell death induces the rearrangement of cones into mosaics of rings while the fibrotic processes of Müller cells remodel to fill the center of the rings. In contrast, previous work established that DL-alpha-aminoadipic-acid (AAA), a compound that transiently blocks Müller cell metabolism, abolishes these highly structured cone rings. Simultaneously, adherens-junction associated protein, Zonula occludens-1 (ZO-1) expression forms in a network between the photoreceptor segments and Müller cells processes. Thus, we hypothesized that AAA treatment alters the cone mosaic rings by disrupting the distal sealing formed by these fibrotic processes, either directly or indirectly, by down regulating the expression of ZO-1. Therefore, we examined these processes and ZO-1 expression at the outer retina after intravitreal injection of AAA and observed that AAA treatment transiently disrupts the distal glial sealing in RP retina, plus induces cones in rings to become more homogeneous. Moreover, ZO-1 expression is actively suppressed after 3 days of AAA treatment, which coincided with cone ring disruption. Similar modifications of glial sealing and cone distribution were observed after injection of siRNA to inhibit ZO-1 expression. These findings support our hypothesis and provide additional information about the critical role played by ZO-1 in glial sealing and shaping the ring mosaic in RP retina. These studies represent important advancements in the understanding of retinal degeneration’s etiology and pathophysiology.

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<![CDATA[Assessing Visual Fields in Patients with Retinitis Pigmentosa Using a Novel Microperimeter with Eye Tracking: The MP-3]]> https://www.researchpad.co/article/5989da84ab0ee8fa60b9be2c

Purpose

The purpose of the current study is to investigate the test-retest reproducibility of visual fields (VFs) measured with the MP-3 microperimeter, in patients with retinitis pigmentosa (RP).

Method

VFs were twice measured with the MP-3 and also the Humphrey Field Analyzer, using the 10–2 test grid pattern in both perimeters, in 30 eyes (15 right and 15 left eyes) of 18 RP patients (11 males and 7 females). Test-retest reproducibility was assessed using the mean absolute deviation (MAD) measure at all 68 points in the test grid. Reproducibility was also evaluated using the intraclass correlation coefficient (ICC) of VF sensitivities.

Result

The mean sensitivity measured in the HFA 10–2 was significantly higher than that measured in the MP-3 in both the first and second VF tests (p <0.0001, linear mixed model). The MAD was 2.4±0.6 [1.1 to 3.6] dB for MP-3 and 2.4±0.9 [1.1 to 5.1] dB for HFA 10–2, which was not significantly different (p = 0.76, linear mixed model). The ICC value associated with the MP-3 VFs was 0.81±0.13 [0.49 to 0.98], which was significantly larger than that observed for the HFA 10–2 VFs: 0.77±0.19 [0.20 to 0.94] (p = 0.043, linear mixed model).

Conclusion

The MP-3 microperimeter appears to be useful to evaluate central visual function in RP eyes, exhibiting test-retest reproducibility that is equal to, or better than, that observed in HFA 10–2 VFs.

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<![CDATA[Inhibition of Matrix Metalloproteinase 9 Enhances Rod Survival in the S334ter-line3 Retinitis Pigmentosa Model]]> https://www.researchpad.co/article/5989da8dab0ee8fa60b9eacd

Retinitis Pigmentosa (RP) is one of the most common forms of inherited visual loss with the initial degeneration of rod photoreceptors, followed by a progressive cone photoreceptor deterioration. Coinciding with this visual loss, the extracellular matrix (ECM) is reorganized, which alters matrix metalloproteinase (MMP) activity levels. A potential pathological role of MMPs, MMP-9 in particular, involves an excitotoxicity-mediated physiological response. In the current study, we examine the MMP-9 and MMP-2 expression levels in the rhodopsin S334ter-line3 RP rat model and investigate the impact of treatment with SB-3CT, a specific MMP-9 and MMP-2 inhibitor, on rod cell survival was tested. Retinal MMP-9 and MMP-2 expression levels were quantified by immunoblot analysis from S334ter-line3 rats compared to controls. Gelatinolytic activities of MMP-9 and MMP-2 by zymography were examined. The geometry of rod death was further evaluated using Voronoi analysis. Our results revealed that MMP-9 was elevated while MMP-2 was relatively unchanged when S334ter-line 3 retinas were compared to controls. With SB-3CT treatment, we observed gelatinolytic activity of both MMPs was decreased and diminished clustering associated with rod death, in addition to a robust preservation of rod photoreceptors. These results demonstrate that up-regulation of MMP-9 in retinas of S334ter-line3 are associated with rod death. The application of SB-3CT dramatically interferes with mechanisms leading to apoptosis in an MMP-9-dependent manner. Future studies will determine the feasibility of using SB-3CT as a potential therapeutic strategy to slow progression of vision loss in genetic inherited forms of human RP.

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<![CDATA[Axial Spondylometaphyseal Dysplasia Is Caused by C21orf2 Mutations]]> https://www.researchpad.co/article/5989db1eab0ee8fa60bceaaa

Axial spondylometaphyseal dysplasia (axial SMD) is an autosomal recessive disease characterized by dysplasia of axial skeleton and retinal dystrophy. We conducted whole exome sequencing and identified C21orf2 (chromosome 21 open reading frame 2) as a disease gene for axial SMD. C21orf2 mutations have been recently found to cause isolated retinal degeneration and Jeune syndrome. We found a total of five biallelic C21orf2 mutations in six families out of nine: three missense and two splicing mutations in patients with various ethnic backgrounds. The pathogenic effects of the splicing (splice-site and branch-point) mutations were confirmed on RNA level, which showed complex patterns of abnormal splicing. C21orf2 mutations presented with a wide range of skeletal phenotypes, including cupped and flared anterior ends of ribs, lacy ilia and metaphyseal dysplasia of proximal femora. Analysis of patients without C21orf2 mutation indicated genetic heterogeneity of axial SMD. Functional data in chondrocyte suggest C21orf2 is implicated in cartilage differentiation. C21orf2 protein was localized to the connecting cilium of the cone and rod photoreceptors, confirming its significance in retinal function. Our study indicates that axial SMD is a member of a unique group of ciliopathy affecting skeleton and retina.

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<![CDATA[Whole Exome Analysis Identifies Frequent CNGA1 Mutations in Japanese Population with Autosomal Recessive Retinitis Pigmentosa]]> https://www.researchpad.co/article/5989d9e2ab0ee8fa60b69e4f

Objective

The purpose of this study was to investigate frequent disease-causing gene mutations in autosomal recessive retinitis pigmentosa (arRP) in the Japanese population.

Methods

In total, 99 Japanese patients with non-syndromic and unrelated arRP or sporadic RP (spRP) were recruited in this study and ophthalmic examinations were conducted for the diagnosis of RP. Among these patients, whole exome sequencing analysis of 30 RP patients and direct sequencing screening of all CNGA1 exons of the other 69 RP patients were performed.

Results

Whole exome sequencing of 30 arRP/spRP patients identified disease-causing gene mutations of CNGA1 (four patients), EYS (three patients) and SAG (one patient) in eight patients and potential disease-causing gene variants of USH2A (two patients), EYS (one patient), TULP1 (one patient) and C2orf71 (one patient) in five patients. Screening of an additional 69 arRP/spRP patients for the CNGA1 gene mutation revealed one patient with a homozygous mutation.

Conclusions

This is the first identification of CNGA1 mutations in arRP Japanese patients. The frequency of CNGA1 gene mutation was 5.1% (5/99 patients). CNGA1 mutations are one of the most frequent arRP-causing mutations in Japanese patients.

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<![CDATA[Dominant Retinitis Pigmentosa, p.Gly56Arg Mutation in NR2E3: Phenotype in a Large Cohort of 24 Cases]]> https://www.researchpad.co/article/5989daf8ab0ee8fa60bc39e6

Importance

This research is the single largest NR2E3 genotype-phenotype correlation study performed to date in autosomal dominant Retinitis Pigmentosa.

Objective

The aim of this study is to analyse the frequency of the p.Gly56Arg mutation in NR2E3 for the largest cohort of autosomal dominant Retinitis Pigmentosa patients to date and its associated phenotype.

Patients and Methods

A cohort of 201 unrelated Spanish families affected by autosomal dominant Retinitis Pigmentosa. The p.Gly56Arg mutation in the NR2E3 (NM_014249.2) gene was analysed in 201 families. In the 24 cases where the mutation had been detected, a haplotype analysis linked to the p.Gly56Arg families was performed, using four extragenic polymorphic markers D15S967, D15S1050, D15S204 and D15S188. Phenotype study included presence and age of onset of night blindness, visual field loss and cataracts; and an ophthalmoscopic examination after pupillary dilation and electroretinogram for the 24 cases.

Results

Seven of the 201 analyzed families were positive for the p.Gly56Arg, leading to a prevalence of 3.5%. Clinical data were available for 24 subjects. Night blindness was the first noticeable symptom (mean 15.9 years). Visual field loss onset was variable (23.3 ± 11.9 years). Loss of visual acuity appeared late in the disease´s evolution. Most of the patients with cataracts (50%) presented it from the third decade of life. Fundus changes showed inter and intrafamiliar variability, but most of the patients showed typical RP changes and it was common to find macular affectation (47.4%). Electroretinogram was impaired from the beginning of the disease. Two families shared a common haplotype. Additionally, all patients shared a 104Kb region between D15S1050 and the NR2E3 gene.

Conclusions

This study highlights the importance of p.Gly56Arg in the NR2E3 gene as a common mutation associated with adRP, and provides new clues to its phenotype, which can allow for a better clinical management and genetic counselling of patients and their families.

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<![CDATA[Identification of a PRPF4 Loss-of-Function Variant That Abrogates U4/U6.U5 Tri-snRNP Integration and Is Associated with Retinitis Pigmentosa]]> https://www.researchpad.co/article/5989da79ab0ee8fa60b97d05

Pre-mRNA splicing by the spliceosome is an essential step in the maturation of nearly all human mRNAs. Mutations in six spliceosomal proteins, PRPF3, PRPF4, PRPF6, PRPF8, PRPF31 and SNRNP200, cause retinitis pigmentosa (RP), a disease characterized by progressive photoreceptor degeneration. All splicing factors linked to RP are constituents of the U4/U6.U5 tri-snRNP subunit of the spliceosome, suggesting that the compromised function of this particle may lead to RP. Here, we report the identification of the p.R192H variant of the tri-snRNP factor PRPF4 in a patient with RP. The mutation affects a highly conserved arginine residue that is crucial for PRPF4 function. Introduction of a corresponding mutation into the zebrafish homolog of PRPF4 resulted in a complete loss of function in vivo. A series of biochemical experiments suggested that p.R192H disrupts the binding interface between PRPF4 and its interactor PRPF3. This interferes with the ability of PRPF4 to integrate into the tri-snRNP, as shown in a human cell line and in zebrafish embryos. These data suggest that the p.R192H variant of PRPF4 represents a functional null allele. The resulting haploinsufficiency of PRPF4 compromises the function of the tri-snRNP, reinforcing the notion that this spliceosomal particle is of crucial importance in the physiology of the retina.

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<![CDATA[Controlled delivery of tauroursodeoxycholic acid from biodegradable microspheres slows retinal degeneration and vision loss in P23H rats]]> https://www.researchpad.co/article/5989db5cab0ee8fa60be0057

Successful drug therapies for treating ocular diseases require effective concentrations of neuroprotective compounds maintained over time at the site of action. The purpose of this work was to assess the efficacy of intravitreal controlled delivery of tauroursodeoxycholic acid (TUDCA) encapsulated in poly(D,L-lactic-co-glycolic acid) (PLGA) microspheres for the treatment of the retina in a rat model of retinitis pigmentosa. PLGA microspheres (MSs) containing TUDCA were produced by the O/W emulsion-solvent evaporation technique. Particle size and morphology were assessed by light scattering and scanning electronic microscopy, respectively. Homozygous P23H line 3 rats received a treatment of intravitreal injections of TUDCA-PLGA MSs. Retinal function was assessed by electroretinography at P30, P60, P90 and P120. The density, structure and synaptic contacts of retinal neurons were analyzed using immunofluorescence and confocal microscopy at P90 and P120. TUDCA-loaded PLGA MSs were spherical, with a smooth surface. The production yield was 78%, the MSs mean particle size was 23 μm and the drug loading resulted 12.5 ± 0.8 μg TUDCA/mg MSs. MSs were able to deliver the loaded active compound in a gradual and progressive manner over the 28-day in vitro release study. Scotopic electroretinografic responses showed increased ERG a- and b-wave amplitudes in TUDCA-PLGA-MSs-treated eyes as compared to those injected with unloaded PLGA particles. TUDCA-PLGA-MSs-treated eyes showed more photoreceptor rows than controls. The synaptic contacts of photoreceptors with bipolar and horizontal cells were also preserved in P23H rats treated with TUDCA-PLGA MSs. This work indicates that the slow and continuous delivery of TUDCA from PLGA-MSs has potential neuroprotective effects that could constitute a suitable therapy to prevent neurodegeneration and visual loss in retinitis pigmentosa.

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<![CDATA[Multimodal Imaging of Photoreceptor Structure in Choroideremia]]> https://www.researchpad.co/article/5989d9ffab0ee8fa60b735ae

Purpose

Choroideremia is a progressive X-linked recessive dystrophy, characterized by degeneration of the retinal pigment epithelium (RPE), choroid, choriocapillaris, and photoreceptors. We examined photoreceptor structure in a series of subjects with choroideremia with particular attention to areas bordering atrophic lesions.

Methods

Twelve males with clinically-diagnosed choroideremia and confirmed hemizygous mutations in the CHM gene were examined. High-resolution images of the retina were obtained using spectral domain optical coherence tomography (SD-OCT) and both confocal and non-confocal split-detector adaptive optics scanning light ophthalmoscope (AOSLO) techniques.

Results

Eleven CHM gene mutations (3 novel) were identified; three subjects had the same mutation and one subject had two mutations. SD-OCT findings included interdigitation zone (IZ) attenuation or loss in 10/12 subjects, often in areas with intact ellipsoid zones; RPE thinning in all subjects; interlaminar bridges in the imaged areas of 10/12 subjects; and outer retinal tubulations (ORTs) in 10/12 subjects. Only split-detector AOSLO could reliably resolve cones near lesion borders, and such cones were abnormally heterogeneous in morphology, diameter and density. On split-detector imaging, the cone mosaic terminated sharply at lesion borders in 5/5 cases examined. Split-detector imaging detected remnant cone inner segments within ORTs, which were generally contiguous with a central patch of preserved retina.

Conclusions

Early IZ dropout and RPE thinning on SD-OCT are consistent with previously published results. Evidence of remnant cone inner segments within ORTs and the continuity of the ORTs with preserved retina suggests that these may represent an intermediate state of retinal degeneration prior to complete atrophy. Taken together, these results supports a model of choroideremia in which the RPE degenerates before photoreceptors.

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<![CDATA[The Time Course of Deafness and Retinal Degeneration in a Kunming Mouse Model for Usher Syndrome]]> https://www.researchpad.co/article/5989da14ab0ee8fa60b7a8ad

Usher syndrome is a group of autosomal recessive diseases characterized by congenital deafness and retinitis pigmentosa. In a mouse model for Usher syndrome, KMush/ush, discovered in our laboratory, we measured the phenotypes, characterized the architecture and morphology of the retina, and quantified the level of expression of pde6b and ush2a between postnatal (P) days 7, and 56. Electroretinograms and auditory brainstem response were used to measure visual and auditory phenotypes. Fundus photography and light microscopy were used to measure the architecture and morphology of the retina. Quantitative real-time PCR was used to measure the expression levels of mRNA. KMush/ush mice had low amplitudes and no obvious waveforms of Electroretinograms after P14 compared with controls. Thresholds of auditory brainstem response in our model were higher than those of controls after P14. By P21, the retinal vessels of KMush/ush mice were attenuated and their optic discs had a waxy pallor. The retinas of KMush/ush mice atrophied and the choroidal vessels were clearly visible. Notably, the architecture of each retinal layer was not different as compared with control mice at P7, while the outer nuclear layer (ONL) and other retinal layers of KMush/ush mice were attenuated significantly between P14 and P21. ONL cells were barely seen in KMush/ush mice at P56. As compared with control mice, the expression of pde6b and ush2a in KMush/ush mice declined significantly after P7. This study is a first step toward characterizing the progression of disease in our mouse model. Future studies using this model may provide insights about the etiology of the disease and the relationships between genotypes and phenotypes providing a valuable resource that could contribute to the foundation of knowledge necessary to develop therapies to prevent the retinal degeneration in patients with Usher Syndrome.

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