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PurposeTo determine the presence of structural changes in HIV retinae (i.e., photoreceptor density and retinal thickness in the macula) compared with age-matched HIV-negative controls.MethodsCohort of patients with known HIV under CART (combination Antiretroviral Therapy) treatment were examined with a flood-illuminated retinal AO camera to assess the cone photoreceptor mosaic and spectral-domain optical coherence tomography (SD-OCT) to assess retinal layers and retinal thickness.ResultsTwenty-four eyes of 12 patients (n = 6 HIV-positive and 6 HIV-negative) were imaged with the adaptive optics camera. In each of the regions of interest studied (nasal, temporal, superior, inferior), the HIV group had significantly less mean cone photoreceptor density compared with age-matched controls (difference range, 4,308-6,872 cones/mm2). A different subset of forty eyes of 20 patients (n = 10 HIV-positive and 10 HIV-negative) was included in the retinal thickness measurements and retinal layer segmentation with the SD-OCT. We observed significant thickening in HIV positive eyes in the total retinal thickness at the foveal center, and in each of the three horizontal B-scans (through the macular center, superior, and inferior to the fovea). We also noted that the inner retina (combined thickness from ILM through RNFL to GCL layer) was also significantly thickened in all the different locations scanned compared with HIV-negative controls.ConclusionOur present study shows that the cone photoreceptor density is significantly reduced in HIV retinae compared with age-matched controls. HIV retinae also have increased macular retinal thickness that may be caused by inner retinal edema secondary to retinovascular disease in HIV. The interaction of photoreceptors with the aging RPE, as well as possible low-grade ocular inflammation causing diffuse inner retinal edema, may be the key to the progressive vision changes in HIV-positive patients without overt retinitis.

Antipsychotic drugs haloperidol and clozapine have been reported to increase the sensitivity of retinal ganglion cells (RGCs) to flashes of light in the P23H rat model of retinitis pigmentosa. In order to better understand the effects of these antipsychotic drugs on the visual responses of P23H rat RGCs, I examined the responses of RGCs to a drifting sinusoidal grating of various contrasts. In-vitro multielectrode array recordings were made from P23H rat RGCs and healthy Sprague-Dawley (SD) rat RGCs. Retinas were stimulated with a drifting sinusoidal grating with eight values of contrast (0, 4, 6, 8.5, 13, 26, 51, and 83%). Contrast response functions based on response amplitudes were fitted with a hyperbolic ratio function and contrast thresholds were determined from the fitted curves. SD rat RGCs were divided into two categories, saturating and non-saturating cells, based on whether they showed saturation of responses at high contrast levels. Most SD rat RGCs (58%) were saturating cells. Haloperidol and clozapine decreased the responses of saturating SD rat RGCs to all grating contrasts, except for the highest contrast tested. Clozapine also decreased the responses of non-saturating SD rat RGCs to all grating contrasts, except for the highest contrast tested. Haloperidol did not however significantly affect the responses of non-saturating SD rat RGCs. Haloperidol and clozapine increased the contrast thresholds of both saturating and non-saturating cells in SD rat retinas. Most (73%) P23H rat RGCs could be categorized as either saturating or non-saturating cells. The remaining ‘uncategorized’ cells were poorly responsive to the drifting grating and were analyzed separately. Haloperidol and clozapine increased the responses of non-saturating and uncategorized P23H rat RGCs to most grating contrasts, including the highest contrast tested. Haloperidol and clozapine also increased the responses of saturating P23H rat RGCs to most grating contrasts but these increases were not statistically significant. Haloperidol and clozapine decreased the contrast thresholds of saturating cells, non-saturating cells and uncategorized cells in P23H rat retinas, although the decrease in contrast thresholds of saturating cells was not found to be statistically significant. Overall, the findings show that haloperidol and clozapine have differential effects on the contrast response functions of SD and P23H rat RGCs. In contrast to the effects observed on SD rat RGCs, both haloperidol and clozapine increased the responsiveness of P23H rat RGCs to both low and high contrast visual stimuli and decreased contrast thresholds. [ABSTRACT FROM AUTHOR]

Mutations in the RHO gene encoding for the visual pigment protein, rhodopsin, are among the most common cause of autosomal dominant retinitis pigmentosa (ADRP). Previous studies of ADRP mutations in different domains of rhodopsin have indicated that changes that lead to more instability in rhodopsin structure are responsible for more severe disease in patients. Here, we further test this hypothesis by comparing side-by-side and therefore quantitatively two RHO mutations, N15S and P23H, both located in the N-terminal intradiscal domain. The in vitro biochemical properties of these two rhodopsin proteins, expressed in stably transfected tetracycline-inducible HEK293S cells, their UV-visible absorption, their Fourier transform infrared, circular dichroism and Metarhodopsin II fluorescence spectroscopy properties were characterized. As compared to the severely impaired P23H molecular function, N15S is only slightly defective in structure and stability. We propose that the molecular basis for these structural differences lies in the greater distance of the N15 residue as compared to P23 with respect to the predicted rhodopsin folding core. As described previously for WT rhodopsin, addition of the cytoplasmic allosteric modulator chlorin e6 stabilizes especially the P23H protein, suggesting that chlorin e6 may be generally beneficial in the rescue of those ADRP rhodopsin proteins whose stability is affected by amino acid replacement. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Retinitis pigmentosa (RP) is an inherited degenerative disease causing severe retinal dystrophy and visual impairment mainly with onset in infancy or adolescence. Targeted next-generation sequencing (NGS) has become an efficient tool to encounter the enormous genetic heterogeneity of diverse retinal dystrophies, including RP. To identify disease-causing mutations in unselected, consecutive RP patients, we conducted Sanger sequencing of genes commonly involved in the suspected genetic RP subtype, followed by targeted large-panel NGS if no mutation was identified, or NGS as primary analysis. A high (70%) detection rate of disease-causing mutations was achieved in a large cohort of 116 unrelated patients. About half (48%) of the solved RP cases were explained by mutations in four genes: RPGR, EYS, PRPF31 and USH2A. Overall, 110 different mutations distributed across 30 different genes were detected, and 46 of these mutations were novel. A molecular diagnosis was achieved in the majority (82–100%) of patients if the family history was suggestive for a particular mode of inheritance, but only in 60% in cases of sporadic RP. The diagnostic potential of extensive molecular analysis in a routine setting is also illustrated by the identification of unexpected genotype-phenotype correlations for RP patients with mutations in CRX, CEP290, RPGRIP1, MFSD8. Furthermore, we identified numerous mutations in autosomal dominant (PRPF31, PRPH2, CRX) and X-linked (RPGR) RP genes in patients with sporadic RP. Variants in RP2 and RPGR were also found in female RP patients with apparently sporadic or dominant disease. In summary, this study demonstrates that massively parallel sequencing of all known retinal dystrophy genes is a valuable diagnostic approach for RP patients. [ABSTRACT FROM AUTHOR]

Purpose: To compare the Esterman Disability Score (EDS) obtained with Goldmann perimetry (GP) testing and the Humphrey field analyzer (HFA) in low vision Japanese subjects. Subjects were also divided into groups by diagnosis to examine how disease influences EDS measurements. Methods: The EDS was obtained using GP (GP-EDS) and the built-in testing program of the HFA (HFA-EDS). Tests were performed within 3 months of each other. Regression analyses were used to examine the relationship between GP-EDS and HFA-EDS. Results: A total of 128 visually impaired subjects were included in this study. Subjects had low vision because of glaucoma (57 subjects), age-related macular degeneration (AMD, 17 subjects), retinitis pigmentosa (RP, 17 subjects), and other causes (37 subjects). The GP-EDS obtained was well-correlated with HFA-EDS (r = 0.87, P < 0.001) and it was possible to estimate HFA-EDS from GP-EDS. The GP-EDS was significantly lower than the HFA-EDS in eyes with glaucoma and RP. There was no significant difference between EDS values in eyes with AMD or other disease. Conclusion: The GP-EDS correlated well with the HFA-EDS. However, the relationship between the EDS measured with the two different testing modalities varies by disease. [ABSTRACT FROM AUTHOR]

Mutations in eyes shut homolog (EYS), a gene predominantly expressed in the photoreceptor cells of the retina, are among the most frequent causes of autosomal recessive (ar) retinitis pigmentosa (RP), a progressive retinal disorder. Due to the absence of EYS in several rodent species and its retina-specific expression, still little is known about the exact function of EYS and the pathogenic mechanism underlying EYS-associated RP. We characterized eys in zebrafish, by RT-PCR analysis on zebrafish eye-derived RNA, which led to the identification of a 8,715 nucleotide coding sequence that is divided over 46 exons. The transcript is predicted to encode a 2,905-aa protein that contains 39 EGF-like domains and five laminin A G-like domains, which overall shows 33% identity with human EYS. To study the function of EYS, we generated a stable eysrmc101/rmc101 mutant zebrafish model using CRISPR/Cas9 technology. The introduced lesion is predicted to result in premature termination of protein synthesis and lead to loss of Eys function. Immunohistochemistry on retinal sections revealed that Eys localizes at the region of the connecting cilium and that both rhodopsin and cone transducin are mislocalized in the absence of Eys. Electroretinogram recordings showed diminished b-wave amplitudes in eysrmc101/rmc101 zebrafish (5 dpf) compared to age- and strain-matched wild-type larvae. In addition, decreased locomotor activity in response to light stimuli was observed in eys mutant larvae. Altogether, our study shows that absence of Eys leads to a disorganized retinal architecture and causes visual dysfunction in zebrafish. [ABSTRACT FROM AUTHOR]

Introduction: Mutations in USH2A cause both isolated Retinitis Pigmentosa (RP) and Usher syndrome (that implies RP and hearing impairment). One of the most frequent variants identified in this gene and among these patients is the p.(Cys759Phe) change. However, the pathogenic role of this allele has been questioned since it was found in homozygosity in two healthy siblings of a Spanish family. To assess the causative role of USH2A p.(Cys759Phe) in autosomal recessive RP (ARRP) and Usher syndrome type II (USH2) and to establish possible genotype-phenotype correlations associated with p.(Cys759Phe), we performed a comprehensive genetic and clinical study in patients suffering from any of the two above-mentioned diseases and carrying at least one p.(Cys759Phe) allele. Materials and methods: Diagnosis was set according to previously reported protocols. Genetic analyses were performed by using classical molecular and Next-Generation Sequencing approaches. Probands of 57 unrelated families were molecularly studied and 63 patients belonging to these families were phenotypically evaluated. Results: Molecular analysis characterized 100% of the cases, identifying: 11 homozygous patients for USH2A p.(Cys759Phe), 42 compound heterozygous patients (12 of them with another missense USH2A pathogenic variant and 30 with a truncating USH2A variant), and 4 patients carrying the p.(Cys759Phe) allele and a pathogenic variant in another RP gene (PROM1, CNGB1 or RP1). No additional causative variants were identified in symptomatic homozygous patients. Statistical analysis of clinical differences between zygosity states yielded differences (p≤0.05) in age at diagnosis of RP and hypoacusis, and progression of visual field loss. Homozygosity of p.(Cys759Phe) and compound heterozygosity with another USH2A missense variant is associated with ARRP or ARRP plus late onset hypoacusis (OR = 20.62, CI = 95%, p = 0.041). Conclusions: The present study supports the role of USH2A p.(Cys759Phe) in ARRP and USH2 pathogenesis, and demonstrates the clinical differences between different zygosity states. Phenotype-genotype correlations may guide the genetic characterization based upon specific clinical signs and may advise on the clinical management and prognosis based upon a specific genotype. [ABSTRACT FROM AUTHOR]

This study’s purpose was to analyze and quantify the impact of auditory information loss versus information gain provided by electronic travel aids (ETAs) on navigation performance in people with low vision. Navigation performance of ten subjects (age: 54.9±11.2 years) with visual acuities >1.0 LogMAR was assessed via the Graz Mobility Test (GMT). Subjects passed through a maze in three different modalities: ‘Normal’ with visual and auditory information available, ‘Auditory Information Loss’ with artificially reduced hearing (leaving only visual information), and ‘ETA’ with a vibrating ETA based on ultrasonic waves, thereby facilitating visual, auditory, and tactile information. Main performance measures comprised passage time and number of contacts. Additionally, head tracking was used to relate head movements to motion direction. When comparing ‘Auditory Information Loss’ to ‘Normal’, subjects needed significantly more time (p<0.001), made more contacts (p<0.001), had higher relative viewing angles (p = 0.002), and a higher percentage of orientation losses (p = 0.011). The only significant difference when comparing ‘ETA’ to ‘Normal’ was a reduced number of contacts (p<0.001). Our study provides objective, quantifiable measures of the impact of reduced hearing on the navigation performance in low vision subjects. Significant effects of ‘Auditory Information Loss’ were found for all measures; for example, passage time increased by 17.4%. These findings show that low vision subjects rely on auditory information for navigation. In contrast, the impact of the ETA was not significant but further analysis of head movements revealed two different coping strategies: half of the subjects used the ETA to increase speed, whereas the other half aimed at avoiding contacts. [ABSTRACT FROM AUTHOR]

The purpose of this study is to determine the factors related to anxiety and depression in patients with retinitis pigmentosa (RP). The status of anxiety and depression was determined in RP patients with the Hospital Anxiety and Depression Scale (HADS) questionnaire which consisted of subscales for HADS-anxiety (HADS-A) and HADS-depression (HADS-D). The vision-specific quality of life (VSQOL) was assessed with the National Eye Institute Visual Function Questionnaire 25 (NEI-VFQ25). The correlations between the HADS-A or HADS-D scores and vision-related clinical parameters such as the best-corrected visual acuity (BCVA), Functional Acuity Score, Functional Field Score, Functional Vision Score, the NEI- VFQ25 subscale score were determined. The socioeconomic status, such as the work status and membership in the RP society, was investigated to determine the factors related to the HADS-A and HADS-D scores. One hundred and twelve RP patients (46 men and 66 women) with mean age of 60.7±15.4 (standard deviation) years were studied. The HADS-A score was not significantly correlated with any visual functions but was significantly correlated with the general health condition (r = -0.34, P<0.001) and the role limitation (r = -0.20, P = 0.03) of the NEI-VFQ25 subscale. The HADS-D score was significantly correlated with all the visual functions (r = -0.38 to 0.29, P<0.001), the NEI-VFQ25 subscale score (r = - 0.58 to -0.33, P<0.001) by Spearman’s correlations. The HADS-A score was significantly higher in the members of the RP society than in non-members (P = 0.013). The mean HADS-D score of employed individuals was significantly lower than that of unemployed ones (P = 0.001) by the Mann-Whitney U test. The results indicate that visual function impairments and vision-related quality of life are associated with a depressive state, and the general health condition is related to anxiety in RP patients. Being employed may be strongly correlated with the degree of depression in RP patients. [ABSTRACT FROM AUTHOR]

Purpose: To characterize the optical coherence tomography (OCT) appearances of photoreceptor degeneration in the rhodopsin P23H transgenic rat (line 2) in relation to the histological, ultrastructural, and electroretinography (ERG) findings. Materials and methods: Homozygous rhodopsin P23H transgenic albino rats (line 2, very-slow degeneration model) were employed. Using OCT (Micron IV®; Phoenix Research Labs, Pleasanton, CA, USA), the natural course of photoreceptor degeneration was recorded from postnatal day (P) 15 to P 287. The OCT images were qualitatively observed by comparing them to histological and ultrastructural findings at P 62 and P 169. In addition, each retinal layer was quantitatively analyzed longitudinally during degeneration, compared it to that observed in wild type Sprague-Dawley (SD) rats. The relationships between the ERG (full-field combined rod-cone response, 3.0 cds/m2 stimulation) findings and OCT images were also analyzed. Results: In the qualitative study, the two layers presumably corresponding to the photoreceptor inner segment ellipsoid zone (EZ) and interdigitation zone (IZ) were identified in the P23H rat until PN day 32. However, the photoreceptor inner and outer segment (IS/OS) layer became diffusely hyperreflective on OCT after P 46, and the EZ and IZ zones could no longer be identified on OCT. In contrast, in the SD rats, the EZ and IZ were clearly distinguished until at least P 247. The ultrastructural study showed partial disarrangements of the photoreceptor outer segment discs in the P23H rats at P 62, although a light-microscopic histological study detected almost no abnormality in the outer segment. In the quantitative study, the outer retinal layer including the outer plexiform layer (OPL) and the outer nuclear layer (ONL) became significantly thinner in the P23H rats than in the SD rats after P 71. The thickness of the IS/OS layer was maintained in the P23H rats until P 130, and it became statistically thinner than in the SD rats at P 237. The longitudinal attenuation in the amplitude of the a- and b-waves of ERG was significantly correlated with the thickness of the combined OPL and ONL but not with that of the IS/OS layer. Conclusion: OCT showed the degenerated photoreceptor IS/OS layer in rhodopsin P23H transgenic rats (line 2) as a diffuse hyperreflective zone, even in the early stage, with the partially disarranged and destabilized OS discs recognizable by ultrastructural assessment but not by a histological study. The amplitude of the a- and b-waves mainly depends on the thickness of the OPL and ONL layer rather than the thickness of the photoreceptor IS/OS layer in P23H rats. [ABSTRACT FROM AUTHOR]

Differences in regional protein expression within the human retina may explain molecular predisposition of specific regions to ophthalmic diseases like age-related macular degeneration, cystoid macular edema, retinitis pigmentosa, and diabetic retinopathy. To quantify protein levels in the human retina and identify patterns of differentially-expressed proteins, we collected foveomacular, juxta-macular, and peripheral retina punch biopsies from healthy donor eyes and analyzed protein content by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Protein expression was analyzed with 1-way ANOVA, gene ontology, pathway representation, and network analysis. We identified a mean of 1,974 proteins in the foveomacular retina, 1,999 in the juxta-macular retina, and 1,779 in the peripheral retina. Six hundred ninety-seven differentially-expressed proteins included those unique to and abundant in each anatomic region. Proteins with higher expression in each region include: heat-shock protein 90-alpha (HSP90AA1), and pyruvate kinase (PKM) in the foveomacular retina; vimentin (VIM) and fructose-bisphosphate aldolase C (ALDOC); and guanine nucleotide-binding protein subunit beta-1 (GNB1) and guanine nucleotide-binding protein subunit alpha-1 (GNAT1) in the peripheral retina. Pathway analysis identified downstream mediators of the integrin signaling pathway to be highly represented in the foveomacular region (P = 6.48 e–06). Metabolic pathways were differentially expressed among all retinal regions. Gene ontology analysis showed that proteins related to antioxidant activity were higher in the juxta-macular and the peripheral retina, but present in lower amounts in the foveomacular retina. Our proteomic analysis suggests that certain retinal regions are susceptible to different forms of metabolic and oxidative stress. The findings give mechanistic insight into retina function, reveal important molecular processes, and prioritize new pathways for therapeutic targeting. [ABSTRACT FROM AUTHOR]

The GABACR antagonist TPMPA and the mGluR1 antagonist JNJ16259685 have been shown previously to alter the sensitivity of retinal ganglion cells (RGCs) in the Sprague-Dawley (SD) rat and P23H rat (animal model of retinitis pigmentosa) to brief flashes of light. In order to better understand the effects of these antagonists on the visual responses of SD and P23H rat RGCs, I examined the responses of RGCs to a drifting sinusoidal grating of various contrasts. Multielectrode array recordings were made from RGCs to a drifting sinusoidal grating of a spatial frequency of 1 cycle/mm and a temporal frequency of 2 cycles/s. In both SD and P23H rat retinas, contrast response functions were found to have a variable shape across cells. Some cells showed saturation of responses at high contrast levels while others did not. Whereas 49% of SD rat RGCs exhibited response saturation, only 14% of P23H rat RGCs showed response saturation. TPMPA decreased the responses of saturating SD rat RGCs to low (6% to 13%) grating contrasts but increased the response to the highest contrast (83%) tested. JNJ16259685 did not significantly affect the contrast response functions of either saturating or non-saturating SD rat RGCs. In contrast, both TPMPA and JNJ16259685 increased the responses of saturating and non-saturating P23H rat RGCs to all grating contrasts. Neither TPMPA nor JNJ16259685 affected the contrast thresholds of SD rat RGCs, but both antagonists lowered the contrast thresholds of P23H rat RGCs. Overall, the findings show that GABACR and mGluR1 antagonists have differential effects on the contrast response functions of SD and P23H rat RGCs. Notably, these receptor antagonists increase the responsiveness of P23H rat RGCs to both low and high contrast visual stimuli. [ABSTRACT FROM AUTHOR]

Visual field impairment affects more than 100 million people globally. However, due to the lack of the access to appropriate ophthalmic healthcare in undeveloped regions as a result of associated costs and expertise this number may be an underestimate. Improved access to affordable diagnostic software designed for visual field examination could slow the progression of diseases, such as glaucoma, allowing for early diagnosis and intervention. We have developed Specvis, a free and open-source application written in Java programming language that can run on any personal computer to meet this requirement (). Specvis was tested on glaucomatous, retinitis pigmentosa and stroke patients and the results were compared to results using the Medmont M700 Automated Static Perimeter. The application was also tested for inter-test intrapersonal variability. The results from both validation studies indicated low inter-test intrapersonal variability, and suitable reliability for a fast and simple assessment of visual field impairment. Specvis easily identifies visual field areas of zero sensitivity and allows for evaluation of its levels throughout the visual field. Thus, Specvis is a new, reliable application that can be successfully used for visual field examination and can fill the gap between confrontation and perimetry tests. The main advantages of Specvis over existing methods are its availability (free), affordability (runs on any personal computer), and reliability (comparable to high-cost solutions). [ABSTRACT FROM AUTHOR]

Background: Carbonic anhydrase inhibitors (CAI) are often used in the treatment of cystoid macular edema (CME) in retinitis pigmentosa (RP) patients. The aim of this meta-analysis is to gain a better understanding of the overall efficacy of CAI treatment. Methods: Databases including PubMed, EMBASE, and Cochrane Library were searched to identify relevant studies. Eligible studies were clinical trials of patients with RP assigned topical or oral CAIs such as dorzolamide and acetazolamide. Changes in central macular thickness (CMT) by OCT in μm and best-corrected visual acuity (BCVA) in log MAR equivalents were extracted and results compared between baseline and after treatment. Results: 11 clinical reports were identified which included a total of 194 patients (358 eyes) available for analysis, with 59 patients (115 eyes) assigned oral CAI treatment and 135 patients (243 eyes) assigned topical CAI treatment. The combined results showed a significant reduction of macular edema, as calculated by baseline and final central macular thickness (CMT) based on OCT examination (46.02μm, 95%CI: -60.96, -31.08, I2 = 65%). However, the effect on visual acuity was inconsistent across studies. Conclusion: Based on non randomized controlled clinical studies, RP patients with CME who were treated with CAIs had better anatomical outcomes, but the effect on visual acuity was contradictory across studies. Multicenter prospective randomized controlled trials would be ideal to definitively test its clinical efficacy in RP patients. [ABSTRACT FROM AUTHOR]

Retinitis Pigmentosa (RP) begins with the death of rod photoreceptors and is slowly followed by a gradual loss of cones and a rearrangement of the remaining retinal neurons. Clusterin is a chaperone protein that protects cells and is involved in various pathophysiological stresses, including retinal degeneration. Using a well-established transgenic rat model of RP (rhodopsin S334ter), we investigated the effects of clusterin on rod photoreceptor survival. To investigate the role of clusterin in S334ter-line3 retinas, Voronoi analysis and immunohistochemistry were used to evaluate the geometry of rod distribution. Additionally, immunoblot analysis, Bax activation, STAT3 and Akt phosphorylation were used to evaluate the pathway involved in rod cell protection. In this study, clusterin (10μg/ml) intravitreal treatment produced robust preservation of rod photoreceptors in S334ter-line3 retina. The mean number of rods in 1mm2 was significantly greater in clusterin injected RP retinas (postnatal (P) 30, P45, P60, & P75) than in age-matched saline injected RP retinas (P<0.01). Clusterin activated Akt, STAT3 and significantly reduced Bax activity; in addition to inducing phosphorylated STAT3 in Müller cells, which suggests it may indirectly acts on photoreceptors. Thus, clusterin treatment may interferes with mechanisms leading to rod death by suppressing cell death through activation of Akt and STAT3, followed by Bax suppression. Novel insights into the pathway of how clusterin promotes the rod cell survival suggest this treatment may be a potential therapeutic strategy to slow progression of vision loss in human RP. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Purpose: Cytomegalovirus (CMV) retinitis remains a leading cause of blindness in countries with a high burden of AIDS. Although dilated fundus examinations are recommended for those with CD4 counts below 100 cells/μL, in practice only those with poor vision and/or symptoms are routinely referred for screening. Therefore, the predictive value of this common practice should be assessed. Methods: This is a prospective cross-sectional study. Patients with known HIV and a CD4 count of less than 100 cells/μL attending an HIV clinic in Chiang Mai, Thailand completed a standardized questionnaire about visual symptoms and underwent visual acuity testing and dilated fundus examination. Participants without CMV retinitis were invited for repeated examinations every 3 months until their CD4 count exceeded 100 cells/μL. Patient-level statistical analyses were conducted to calculate diagnostic test characteristics, with bootstrapping to account for correlated data. Results: Of 103 study participants, 16 had CMV retinitis diagnosed at some point during the study. Participants with CMV retinitis were more likely to complain of visual symptoms compared to those without CMV retinitis (p = 0.01), including scotoma (p = 0.0002), itchy or watery eyes (p < 0.0001), and eye pain (p = 0.003); they were also more likely to have visual acuity worse than Counting Fingers (p = 0.0003). However, the absence of eye symptoms and the absence of poor vision did not strongly affect the probability that a patient did not have disease (negative likelihood ratio 0.56 and 0.76, respectively). Conclusions: Ocular symptoms and poor visual acuity were poor diagnostic indicators for the presence of CMV retinitis. Systematic screening of HIV patients with CD4 counts below 100 cells/μl should be carried out to detect disease at an early stage, when blindness can still be prevented. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Purpose: To evaluate the correlation between optical coherence tomography (OCT) and the histological, ultrastructural and electroretinography (ERG) findings of retinal degeneration in Royal College of Surgeons (RCS-/-) rats. Materials and Methods: Using OCT, we qualitatively and quantitatively observed the continual retinal degeneration in RCS-/- rats, from postnatal (PN) day 17 until PN day 111. These findings were compared with the corresponding histological, electron microscopic, and ERG findings. We also compared them to OCT findings in wild type RCS+/+ rats, which were used as controls. Results: After PN day 17, the hyperreflective band at the apical side of the photoreceptor layer became blurred. The inner segment (IS) ellipsoid zone then became obscured, and the photoreceptor IS and outer segment (OS) layers became diffusely hyperreflective after PN day 21. These changes correlated with histological and electron microscopic findings showing extracellular lamellar material that accumulated in the photoreceptor OS layer. After PN day 26, the outer nuclear layer became significantly thinner (P < 0.01) and hyperreflective compared with that in the controls; conversely, the photoreceptor IS and OS layers, as well as the inner retinal layers, became significantly thicker (P < 0.001 and P = 0.05, respectively). The apical hyperreflective band, as well as the IS ellipsoid zone, gradually disappeared between PN day 20 and PN day 30; concurrently, the ERG a- and b-wave amplitudes deteriorated. In contrast, the thicknesses of the combined retinal pigment epithelium and choroid did not differ significantly between RCS-/- and RCS+/+ rats. Conclusion: Our results suggest that OCT demonstrates histologically validated photoreceptor degeneration in RCS rats, and that OCT findings partly correlate with ERG findings. We propose that OCT is a less invasive and useful method for evaluating photoreceptor degeneration in animal models of retinitis pigmentosa. [ABSTRACT FROM AUTHOR]

Purpose: Degenerative retinal diseases, especially retinitis pigmentosa (RP), lead to severe peripheral visual field loss (tunnel vision), which impairs mobility. The lack of peripheral information leads to fewer horizontal eye movements and, thus, diminished scanning in RP patients in a natural environment walking task. This randomized controlled study aimed to improve mobility and the dynamic visual field by applying a compensatory Exploratory Saccadic Training (EST). Methods: Oculomotor responses during walking and avoiding obstacles in a controlled environment were studied before and after saccade or reading training in 25 RP patients. Eye movements were recorded using a mobile infrared eye tracker (Tobii glasses) that measured a range of spatial and temporal variables. Patients were randomly assigned to two training conditions: Saccade (experimental) and reading (control) training. All subjects who first performed reading training underwent experimental training later (waiting list control group). To assess the effect of training on subjects, we measured performance in the training task and the following outcome variables related to daily life: Response Time (RT) during exploratory saccade training, Percent Preferred Walking Speed (PPWS), the number of collisions with obstacles, eye position variability, fixation duration, and the total number of fixations including the ones in the subjects' blind area of the visual field. Results: In the saccade training group, RTs on average decreased, while the PPWS significantly increased. The improvement persisted, as tested 6 weeks after the end of the training. On average, the eye movement range of RP patients before and after training was similar to that of healthy observers. In both, the experimental and reading training groups, we found many fixations outside the subjects' seeing visual field before and after training. The average fixation duration was significantly shorter after the training, but only in the experimental training condition. Conclusions: We conclude that the exploratory saccade training was beneficial for RP patients and resulted in shorter fixation durations after the training. We also found a significant improvement in relative walking speed during navigation in a real-world like controlled environment. [ABSTRACT FROM AUTHOR]

Purpose: To test the efficacy of systemic gene delivery of a mutant form of erythropoietin (EPO-R76E) that has attenuated erythropoietic activity, in a mouse model of autosomal dominant retinitis pigmentosa. Methods: Ten-day old mice carrying one copy of human rhodopsin with the P23H mutation and both copies of wild-type mouse rhodopsin (hP23H RHO+/-,mRHO+/+) were injected into the quadriceps with recombinant adeno-associated virus (rAAV) carrying either enhanced green fluorescent protein (eGFP) or EpoR76E. Visual function (electroretinogram) and retina structure (optical coherence tomography, histology, and immunohistochemistry) were assessed at 7 and 12 months of age. Results: The outer nuclear layer thickness decreased over time at a slower rate in rAAV.EpoR76E treated as compared to the rAAV.eGFP injected mice. There was a statistically significant preservation of the electroretinogram at 7, but not 12 months of age. Conclusions: Systemic EPO-R76E slows death of the photoreceptors and vision loss in hP23H RHO+/-,mRHO+/+ mice. Treatment with EPO-R76E may widen the therapeutic window for retinal degeneration patients by increasing the number of viable cells. Future studies might investigate if co-treatment with EPO-R76E and gene replacement therapy is more effective than gene replacement therapy alone. [ABSTRACT FROM AUTHOR]

Retinitis pigmentosa is an inherited blinding disorder characterized by progressive degeneration and loss of photoreceptors. The exact mechanism of degeneration and cell death of photoreceptors is not known, but is thought to involve disturbed Ca2+—signaling. Ca2+ can enter the photoreceptor cell via outer segment cyclic nucleotide-gated (CNG) channels or synaptic Cav1.4 L-type voltage-gated calcium channels (VGCC). Previously, we have shown that genetic ablation of the Cngb1 gene encoding the B subunit of the rod CNG channel delays the fast progressing degeneration in the rd1 mutant mouse model of retinitis pigmentosa. In this study, we crossbred rd1 mice with the Cacna1f-deficient mouse lacking the Cav1.4 α1 subunit of the L-type VGCC. Longitudinal in vivo examinations of photoreceptor layer thickness by optical coherence tomography revealed a significant, but not sustained delay of retinal degeneration in Cacna1f x rd1 double mutant mice compared to rd1 mice. This was accompanied by a reduction of TUNEL positive cells in the early phase of rod degeneration. Remarkably, Cacna1f x rd1 double mutant mice displayed a strong decrease in the activation of the Ca2+-dependent protease calpain during photoreceptor loss. Our results show that genetic deletion of the synaptic Cav1.4 L-type VGCCs impairs calpain activation and leads to a short-term preservation of photoreceptors in the rd1 mouse. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Background: Retinitis pigmentosa is a phenotype with diverse genetic causes. Due to this genetic heterogeneity, genome-wide identification and analysis of protein-altering DNA variants by exome sequencing is a powerful tool for novel variant and disease gene discovery. In this study, exome sequencing analysis was used to search for potentially causal DNA variants in a two-generation pedigree with apparent dominant retinitis pigmentosa. Methods: Variant identification and analysis of three affected members (mother and two affected offspring) was performed via exome sequencing. Parental samples of the index case were used to establish inheritance. Follow-up testing of 94 additional retinitis pigmentosa pedigrees was performed via retrospective analysis or Sanger sequencing. Results and Conclusions: A total of 136 high quality coding variants in 123 genes were identified which are consistent with autosomal dominant disease. Of these, one of the strongest genetic and functional candidates is a c.269A>G (p.Tyr90Cys) variant in ARL3. Follow-up testing established that this variant occurred de novo in the index case. No additional putative causal variants in ARL3 were identified in the follow-up cohort, suggesting that if ARL3 variants can cause adRP it is an extremely rare phenomenon. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

: 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 [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

For sensitive detection of rare gene repair events in terminally differentiated photoreceptors, we generated a knockin mouse model by replacing one mouse rhodopsin allele with a form of the human rhodopsin gene that causes a severe, early-onset form of retinitis pigmentosa. The human gene contains a premature stop codon at position 344 (Q344X), cDNA encoding the enhanced green fluorescent protein (EGFP) at its 3′ end, and a modified 5′ untranslated region to reduce translation rate so that the mutant protein does not induce retinal degeneration. Mutations that eliminate the stop codon express a human rhodopsin-EGFP fusion protein (hRho-GFP), which can be readily detected by fluorescence microscopy. Spontaneous mutations were observed at a frequency of about one per retina; in every case, they gave rise to single fluorescent rod cells, indicating that each mutation occurred during or after the last mitotic division. Additionally, the number of fluorescent rods did not increase with age, suggesting that the rhodopsin gene in mature rod cells is less sensitive to mutation than it is in developing rods. Thus, there is a brief developmental window, coinciding with the transcriptional activation of the rhodopsin locus, in which somatic mutations of the rhodopsin gene abruptly begin to appear. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Recent progress in molecular analysis has revealed the possible involvement of multiple inflammatory signaling pathways in pathogenesis of retinal degeneration. However, how aberrant signaling pathways cause tissue damage and dysfunction is still being elucidated. Here, we focus on 5′-adenosine monophosphate (AMP)-activated protein kinase (AMPK), originally recognized as a key regulator of energy homeostasis. AMPK is also modulated in response to inflammatory signals, although its functions in inflamed tissue are obscure. We investigated the role of activated AMPK in the retinal neural damage and visual function impairment caused by inflammation. For this purpose, we used a mouse model of lipopolysaccharide-induced inflammation in the retina, and examined the effects of an AMPK activator, 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR). During inflammation, activated AMPK in the neural retina was decreased, but AICAR treatment prevented this change. Moreover, the electroretinogram (ERG) a-wave response, representing photoreceptor function, showed visual dysfunction in this model that was prevented by AICAR. Consistently, the model showed shortened photoreceptor outer segments (OSs) with reduced levels of rhodopsin, a visual pigment concentrated in the OSs, in a post-transcriptional manner, and these effects were also prevented by AICAR. In parallel, the level of activated NF-κB increased in the retina during inflammation, and this increase was suppressed by AICAR. Treatment with an NF-κB inhibitor, dehydroxymethylepoxyquinomicin (DHMEQ) preserved the rhodopsin level during inflammation, suppressing NF-κB. These findings indicated that AMPK activation by AICAR and subsequent NF-κB inhibition had a protective effect on visual function, and that AMPK activation played a neuroprotective role during retinal inflammation. [ABSTRACT FROM AUTHOR]