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1.
PLoS Genet ; 18(3): e1010129, 2022 03.
Artículo en Inglés | MEDLINE | ID: mdl-35353811

RESUMEN

Over 1,500 variants in the ABCA4 locus cause phenotypes ranging from severe, early-onset retinal degeneration to very late-onset maculopathies. The resulting ABCA4/Stargardt disease is the most prevalent Mendelian eye disorder, although its underlying clinical heterogeneity, including penetrance of many alleles, are not well-understood. We hypothesized that a share of this complexity is explained by trans-modifiers, i.e., variants in unlinked loci, which are currently unknown. We sought to identify these by performing exome sequencing in a large cohort for a rare disease of 622 cases and compared variation in seven genes known to clinically phenocopy ABCA4 disease to cohorts of ethnically matched controls. We identified a significant enrichment of variants in 2 out of the 7 genes. Moderately rare, likely functional, variants, at the minor allele frequency (MAF) <0.005 and CADD>25, were enriched in ROM1, where 1.3% of 622 patients harbored a ROM1 variant compared to 0.3% of 10,865 controls (p = 2.41E04; OR 3.81 95% CI [1.77; 8.22]). More importantly, analysis of common variants (MAF>0.1) identified a frequent haplotype in PRPH2, tagged by the p.Asp338 variant with MAF = 0.21 in the matched general population that was significantly increased in the patient cohort, MAF 0.25, p = 0.0014. Significant differences were also observed between ABCA4 disease subgroups. In the late-onset subgroup, defined by the hypomorphic p.Asn1868Ile variant and including c.4253+43G>A, the allele frequency for the PRPH2 p.Asp338 variant was 0.15 vs 0.27 in the remaining cohort, p = 0.00057. Known functional data allowed suggesting a mechanism by which the PRPH2 haplotype influences the ABCA4 disease penetrance. These associations were replicated in an independent cohort of 408 patients. The association was highly statistically significant in the combined cohorts of 1,030 cases, p = 4.00E-05 for all patients and p = 0.00014 for the hypomorph subgroup, suggesting a substantial trans-modifying role in ABCA4 disease for both rare and common variants in two unlinked loci.


Asunto(s)
Transportadoras de Casetes de Unión a ATP , Degeneración Macular , Transportadoras de Casetes de Unión a ATP/genética , Proteínas del Ojo/genética , Frecuencia de los Genes , Humanos , Degeneración Macular/genética , Mutación , Linaje , Fenotipo , Enfermedad de Stargardt/genética , Tetraspaninas/genética
2.
Hum Mol Genet ; 30(14): 1293-1304, 2021 06 26.
Artículo en Inglés | MEDLINE | ID: mdl-33909047

RESUMEN

Over 1200 variants in the ABCA4 gene cause a wide variety of retinal disease phenotypes, the best known of which is autosomal recessive Stargardt disease (STGD1). Disease-causing variation encompasses all mutation categories, from large copy number variants to very mild, hypomorphic missense variants. The most prevalent disease-causing ABCA4 variant, present in ~ 20% of cases of European descent, c.5882G > A p.(Gly1961Glu), has been a subject of controversy since its minor allele frequency (MAF) is as high as ~ 0.1 in certain populations, questioning its pathogenicity, especially in homozygous individuals. We sequenced the entire ~140Kb ABCA4 genomic locus in an extensive cohort of 644 bi-allelic, i.e. genetically confirmed, patients with ABCA4 disease and analyzed all variants in 140 compound heterozygous and 10 homozygous cases for the p.(Gly1961Glu) variant. A total of 23 patients in this cohort additionally harbored the deep intronic c.769-784C > T variant on the p.(Gly1961Glu) allele, which appears on a specific haplotype in ~ 15% of p.(Gly1961Glu) alleles. This haplotype was present in 5/7 of homozygous cases, where the p.(Gly1961Glu) was the only known pathogenic variant. Three cases had an exonic variant on the same allele with the p.(Gly1961Glu). Patients with the c.[769-784C > T;5882G > A] complex allele exhibit a more severe clinical phenotype, as seen in compound heterozygotes with some more frequent ABCA4 mutations, e.g. p.(Pro1380Leu). Our findings indicate that the c.769-784C > T variant is major cis-acting modifier of the p.(Gly1961Glu) allele. The absence of such additional allelic variation on most p.(Gly1961Glu) alleles largely explains the observed paucity of affected homozygotes in the population.


Asunto(s)
Transportadoras de Casetes de Unión a ATP , Transportadoras de Casetes de Unión a ATP/genética , Alelos , Frecuencia de los Genes , Humanos , Mutación , Penetrancia , Fenotipo , Enfermedad de Stargardt/genética
3.
Mol Vis ; 27: 95-106, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33907365

RESUMEN

Purpose: Despite the extensive use of next-generation sequencing (NGS) technology to identify disease-causing genomic variations, a major gap in our understanding of Mendelian diseases is the unidentified molecular lesion in a significant portion of patients. For inherited retinal degenerations (IRDs), although currently close to 300 disease-associated genes have been identified, the mutations in approximately one-third of patients remain unknown. With mounting evidence that noncoding mutations might contribute significantly to disease burden, we aimed to systematically investigate the contributions of noncoding regions in the genome to IRDs. Methods: In this study, we focused on RPGRIP1, which has been linked to various IRD phenotypes, including Leber congenital amaurosis (LCA), retinitis pigmentosa (RP), and macular dystrophy (MD). As several noncoding mutant alleles have been reported in RPGRIP1, and we observed that the mutation carrier frequency of RPGRIP1 is higher in patient cohorts with unsolved IRDs, we hypothesized that mutations in the noncoding regions of RPGRIP1 might be a significant contributor to pathogenicity. To test this hypothesis, we performed whole-genome sequencing (WGS) for 25 patients with unassigned IRD who carry a single mutation in RPGRIP1. Results: Three noncoding variants in RPGRIP1, including a 2,890 bp deletion and two deep-intronic variants (c.2710+233G>A and c.1468-263G>C), were identified as putative second hits of RPGRIP1 in three patients with LCA. The mutant alleles were validated with direct sequencing or in vitro assays. Conclusions: The results highlight the significance of the contribution of noncoding pathogenic variants to unsolved IRD cases.


Asunto(s)
Proteínas del Citoesqueleto/genética , Mutación/genética , ARN no Traducido/genética , Degeneración Retiniana/genética , Adulto , Alelos , Preescolar , Clonación Molecular , Electrorretinografía , Femenino , Humanos , Masculino , Fenotipo , Reacción en Cadena en Tiempo Real de la Polimerasa , Retina/fisiopatología , Degeneración Retiniana/diagnóstico , Degeneración Retiniana/fisiopatología , Tomografía de Coherencia Óptica , Transfección , Agudeza Visual/fisiología , Secuenciación Completa del Genoma
4.
Retina ; 40(8): 1630-1633, 2020 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-31568063

RESUMEN

PURPOSE: Retinitis pigmentosa (RP) is an inherited pigmentary retinal dystrophy where patients experience poor peripheral, night, and eventually central vision. There are statements in the literature which suggest visual acuity loss can progress to total blindness in these patients. This study sought to examine these statements by performing a retrospective analysis of the visual acuity measured in a large cohort of RP patients. METHODS: The charts of 1,095 RP patients were reviewed in this retrospective cross-sectional analysis. They included all of the RP patients examined by one of the authors (G.A.F.). Patients with sector RP or a delimited form of this disease were not included. The review was focused on the analysis of patients with 20/200 or worse vision in the better-seeing eye (N = 215). RESULTS: We determined that 0.46% of the enrolled patients progressed to no light perception in each eye. Ninety-two percent of the 1,095 patients examined were able to read a visual acuity chart. There were 6.8% who saw only hand motion, count fingers, or light perception. CONCLUSION: No light perception was measured in only 0.46% of patients. Thus, only a very small number of the RP patients in our cohort progressed to total blindness.


Asunto(s)
Ceguera/fisiopatología , Retinitis Pigmentosa/fisiopatología , Baja Visión/fisiopatología , Personas con Daño Visual/clasificación , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Niño , Estudios Transversales , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Agudeza Visual/fisiología , Campos Visuales/fisiología , Adulto Joven
5.
Doc Ophthalmol ; 138(2): 161-166, 2019 04.
Artículo en Inglés | MEDLINE | ID: mdl-30796641

RESUMEN

PURPOSE: The Usher syndrome phenotype is comprised of ocular and audiologic anomalies. Patients characteristically experience congenital hearing loss, nyctalopia, reduced visual fields, and ultimately decreased visual acuity. However, diagnosis may initially be more difficult in cases with limited ocular findings. Here, we present a case in which an adult patient had neither subjective visual complaints nor ocular findings at the time of diagnosis aside from a moderate reduction in rod and cone function on electroretinogram testing. Nevertheless, 43 years after his initial examination, he showed severe degenerative changes in the retina. METHODS: A 63-year-old man with Usher syndrome type 2 underwent ophthalmic examination that included visual acuity, optical coherence tomography (OCT), electroretinogram (ERG), fundus photography, and Goldmann visual field testing. The patient also had genetic testing performed. We additionally reviewed the ocular findings on two of his siblings also afflicted with Usher syndrome type 2. RESULTS: Our findings documented the long-term progression of Usher syndrome in this patient. They showed that the patient was asymptomatic with only a moderate reduction on ERG testing at the time of diagnosis, but subsequently progressed to an advanced stage of retinal disease with severe visual loss. CONCLUSIONS: The patient demonstrated that the absence of visual symptoms and favorable findings on functional testing on initial presentation might yet belie a future for austere visual loss. Caution is thus warranted when predicting a visual prognosis in such a patient. Further, the value in electroretinographic testing for diagnosis is demonstrated.


Asunto(s)
Retinitis Pigmentosa/diagnóstico , Síndromes de Usher/diagnóstico , Adulto , Electrorretinografía , Femenino , Humanos , Masculino , Persona de Mediana Edad , Fenotipo , Pronóstico , Retina/fisiopatología , Retinitis Pigmentosa/fisiopatología , Tomografía de Coherencia Óptica , Síndromes de Usher/fisiopatología , Agudeza Visual/fisiología , Pruebas del Campo Visual , Campos Visuales/fisiología , Adulto Joven
6.
Doc Ophthalmol ; 139(1): 11-20, 2019 08.
Artículo en Inglés | MEDLINE | ID: mdl-30927187

RESUMEN

PURPOSE: To investigate receptor and post-receptor function in KCNV2 retinopathy [cone dystrophy with supernormal rod electroretinogram (ERG)], using the pupillary light reflex (PLR) and the ERG. METHODS: Two unrelated patients (1 male and 1 female) with molecularly confirmed KCNV2 retinopathy underwent full-field two-color pupillometry testing in one eye, with monitoring of the stimulated eye by an infrared digital camera. Pupillometry stimuli consisted of 1-s duration, short-wavelength (465-nm, blue) and long-wavelength (642-nm, red) stimuli. Pupillometry intensity series were performed under both a dark-adapted condition and a light-adapted condition (on a 0.76-log cd m-2 blue background). The transient PLR, defined as the maximum constriction following flash onset, was measured under all conditions. The melanopsin-mediated sustained constriction was measured 5-7 s following flash offset for the highest flash luminance presented in the dark. Both patients were also tested in one eye with the full-field ERG, including a dark-adapted intensity series and ISCEV standard stimuli. RESULTS: Dark-adapted PLRs were markedly attenuated or extinguished for low-luminance stimuli, but the responses to higher-luminance blue stimuli were within normal limits. Light-adapted PLRs to blue stimuli were generally within normal limits, exceeding the responses to photopically matched red stimuli. Thus, light-adapted responses were consistent with either rod or S-cone mediation of the PLR. Melanopsin-mediated sustained PLRs were within normal limits. ERG showed the characteristic findings previously reported in this condition. Cone-mediated ERG responses were markedly decreased in amplitude. Rod-mediated ERG responses were absent for low-luminance stimuli (- 3 log cd s m-2), but had normal amplitude for stimuli of - 2 log cd s m-2 and above (although none were "supernormal"). The b-wave for the dark-adapted ISCEV standard - 2 log cd s m-2 stimulus was markedly delayed, whereas the b-wave timing was generally normal for higher flash luminances. CONCLUSIONS: The abnormalities measured by pupillometry have a similar pattern to the outer-retinal abnormalities measured by ERG in KCNV2 retinopathy. These findings as well as the normal sustained PLR suggest that inner-retinal function may be preserved in KCNV2 retinopathy and highlight the potential for therapies designed to restore outer-retinal function in these individuals.


Asunto(s)
Células Fotorreceptoras de Vertebrados/fisiología , Reflejo Pupilar/fisiología , Retinitis Pigmentosa/fisiopatología , Adulto , Consanguinidad , Adaptación a la Oscuridad , Electrorretinografía , Femenino , Técnicas de Genotipaje , Humanos , Masculino , Estimulación Luminosa , Canales de Potasio con Entrada de Voltaje/genética , Retinitis Pigmentosa/genética , Opsinas de Bastones/metabolismo , Adulto Joven
7.
Retina ; 39(12): 2311-2325, 2019 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-30204727

RESUMEN

PURPOSE: To investigate the Stargardt disease phenotype associated with an unusually common and "extremely hypomorphic" ABCA4 variant, p.N1868I. METHODS: The charts of 27 patients with p.N1868I on one allele and a severe/deleterious mutation on the other allele were reviewed. Subjective age of onset, best-corrected visual acuity, and stage of disease were recorded for all 27 patients, 18 of whom had multiple visits. When available, fundus photography, spectral domain optical coherence tomography, fundus autofluorescence, full-field electroretinograms, Goldmann visual fields, and fluorescein angiography were included. Five families with multiple affected members were analyzed. RESULTS: The median age at symptom onset was 41.5 years, and 3 p.N1868I patients had not developed visual symptoms as of the most recent eye examination. Median best-corrected visual acuity in the better-seeing eye at baseline was 20/25, and the median duration from symptom onset to legal blindness was 25 years. The five families described in this study demonstrated clinically significant intrafamilial variability, and affected family members who did not share the p.N1868I variant had relatively more severe phenotypes. CONCLUSION: This study demonstrates the consistency of foveal sparing, the variation in age at onset, the intrafamilial variability, and the prognosis with regard to visual acuity in p.N1868I-associated Stargardt disease.


Asunto(s)
Transportadoras de Casetes de Unión a ATP/genética , Mutación , Enfermedad de Stargardt/diagnóstico , Enfermedad de Stargardt/genética , Adolescente , Adulto , Edad de Inicio , Anciano , Alelos , Electrorretinografía , Femenino , Angiografía con Fluoresceína , Fóvea Central , Humanos , Masculino , Persona de Mediana Edad , Fenotipo , Fotograbar , Estudios Retrospectivos , Enfermedad de Stargardt/fisiopatología , Tomografía de Coherencia Óptica , Agudeza Visual/fisiología , Pruebas del Campo Visual , Adulto Joven
8.
J Med Genet ; 54(6): 404-412, 2017 06.
Artículo en Inglés | MEDLINE | ID: mdl-28446513

RESUMEN

BACKGROUND: Variation in the ABCA4 gene is causal for, or associated with, a wide range of phenotypes from early onset Mendelian retinal dystrophies to late-onset complex disorders such as age-related macular degeneration (AMD). Despite substantial progress in determining the causal genetic variation, even complete sequencing of the entire open reading frame and splice sites of ABCA4 identifies biallelic mutations in only 60%-70% of cases; 20%-25% remain with one mutation and no mutations are found in 10%-15% of cases with clinically confirmed ABCA4 disease. This study was designed to identify missing causal variants specifically in monoallelic cases of ABCA4 disease. METHODS: Direct sequencing and analysis were performed in a large familial ABCA4 disease cohort of predominately European descent (n=643). Patient phenotypes were assessed from clinical and retinal imaging data. RESULTS: We determined that a hypomorphic ABCA4 variant c.5603A>T (p.Asn1868Ile), previously considered benign due to high minor allele frequency (MAF) (~7%) in the general population, accounts for 10% of the disease, >50% of the missing causal alleles in monoallelic cases, ~80% of late-onset cases and distinguishes ABCA4 disease from AMD. It results in a distinct clinical phenotype characterised by late-onset of symptoms (4th decade) and foveal sparing (85%). Intragenic modifying effects involving this variant and another, c.2588G>C (p.Gly863Ala) allele, were also identified. CONCLUSIONS: These findings substantiate the causality of frequent missense variants and their phenotypic outcomes as a significant contribution to ABCA4 disease, particularly the late-onset phenotype, and its clinical variation. They also suggest a significant revision of diagnostic screening and assessment of ABCA4 variation in aetiology of retinal diseases.


Asunto(s)
Transportadoras de Casetes de Unión a ATP/genética , Frecuencia de los Genes/genética , Degeneración Macular/genética , Adulto , Alelos , Estudios de Cohortes , Variación Genética/genética , Humanos , Mutación/genética , Fenotipo , Distrofias Retinianas/genética
9.
Retina ; 38(12): 2387-2394, 2018 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-29068916

RESUMEN

PURPOSE: To better define visual acuity loss in patients with Stargardt disease later in life. METHODS: The most recent best-corrected visual acuities in the better-seeing eye of 221 patients with Stargardt disease over 40 years of age were recorded. Also included were the age at subjective onset for symptoms and duration of symptoms. Juvenile onset was defined as onset before age 21; adult onset was defined as onset between 21 and 40 years; and late onset was defined as onset at age 41 or later. RESULTS: The median age of the patients with Stargardt disease was 53.1 years. Twenty-four patients (10.9%) had worse than 20/400 best-corrected visual acuity, and none had either light perception or no light perception vision. Whereas 17 of the 52 juvenile onset patients had best-corrected visual acuity worse than 20/400, only 4 of 80 adult-onset patients and 1 of 70 late-onset patients reached this level of acuity loss. CONCLUSION: Although many patients with Stargardt disease lose visual acuity to the 20/200 to 20/400 range, and some lose visual acuity beyond 20/400, none of these patients reached either light perception or no light perception. The numbers found in this study will be valuable in counseling patients with Stargardt disease and could have value in planning treatment trials.


Asunto(s)
Degeneración Macular/congénito , Grupos Raciales , Agudeza Visual/fisiología , Transportadoras de Casetes de Unión a ATP/genética , Adulto , Distribución por Edad , Factores de Edad , Anciano , Estudios Transversales , ADN/genética , Análisis Mutacional de ADN , Femenino , Genotipo , Humanos , Illinois/epidemiología , Incidencia , Degeneración Macular/etnología , Degeneración Macular/genética , Degeneración Macular/fisiopatología , Masculino , Persona de Mediana Edad , Mutación , Pronóstico , Estudios Retrospectivos , Enfermedad de Stargardt
10.
Retina ; 37(8): 1555-1561, 2017 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-27828908

RESUMEN

PURPOSE: To evaluate changes in cystic-appearing macular lesions and visual acuity in patients with X-linked retinoschisis while being treated with a carbonic anhydrase inhibitor. METHODS: A retrospective analysis of 68 eyes from 36 patients between the ages of 5 years and 61 years with X-linked retinoschisis were monitored while on a carbonic anhydrase inhibitor. Macular cystic-appearing lesions were monitored with optical coherence tomography. Snellen visual acuity measurements were converted to logarithm of the minimum angle of resolution equivalent Early Treatment Diabetic Retinopathy Study letters for analysis. Analyses for changes in both visual acuity and macular cysts included comparisons between treatment and pretreatment segments. RESULTS: Forty-five eyes (66%) had a reduction of their cysts while on a carbonic anhydrase inhibitor. Twenty eyes (29%) showed no cystic change, whereas 3 eyes (4%) demonstrated worsening of their cysts with treatment when compared with pretreatment. There was a statistically significant improvement in logarithm of the minimum angle of resolution visual acuity while on treatment relative to pretreatment (P < 0.0001). The estimated average Early Treatment Diabetic Retinopathy Study equivalent improvement was 0.09 (slightly less than one line on the Early Treatment Diabetic Retinopathy Study chart) with a 95% confidence interval of 0.08 to 0.11. CONCLUSION: Considering the entire 36 patients in this cohort, while statistically significant, the average improvement in visual acuity was modest. Nonetheless, in individual patients, the improvement was more substantial. Improvement in the extent of cystic macular lesions was observed in a high percentage of cases.


Asunto(s)
Acetazolamida/administración & dosificación , Inhibidores de Anhidrasa Carbónica/administración & dosificación , Edema Macular/diagnóstico , Retinosquisis/tratamiento farmacológico , Sulfonamidas/administración & dosificación , Tiazinas/administración & dosificación , Tomografía de Coherencia Óptica/métodos , Agudeza Visual , Adolescente , Adulto , Niño , Preescolar , Relación Dosis-Respuesta a Droga , Esquema de Medicación , Electrorretinografía , Femenino , Estudios de Seguimiento , Humanos , Edema Macular/tratamiento farmacológico , Edema Macular/etiología , Masculino , Persona de Mediana Edad , Soluciones Oftálmicas/administración & dosificación , Retinosquisis/complicaciones , Retinosquisis/diagnóstico , Estudios Retrospectivos , Factores de Tiempo , Resultado del Tratamiento , Adulto Joven
11.
Retina ; 37(10): 1956-1966, 2017 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-28145975

RESUMEN

PURPOSE: Congenital achromatopsia is an autosomal recessive disease causing substantial reduction or complete absence of cone function. Although believed to be a relatively stationary disorder, questions remain regarding the stability of cone structure over time. In this study, the authors sought to assess the repeatability of and examine longitudinal changes in measurements of central cone structure in patients with achromatopsia. METHODS: Forty-one subjects with CNGB3-associated achromatopsia were imaged over a period of between 6 and 26 months using optical coherence tomography and adaptive optics scanning light ophthalmoscopy. Outer nuclear layer (ONL) thickness, ellipsoid zone (EZ) disruption, and peak foveal cone density were assessed. RESULTS: ONL thickness increased slightly compared with baseline (0.184 µm/month, P = 0.02). The EZ grade remained unchanged for 34/41 subjects. Peak foveal cone density did not significantly change over time (mean change 1% per 6 months, P = 0.126). CONCLUSION: Foveal cone structure showed little or no change in this group of subjects with CNGB3-associated achromatopsia. Over the time scales investigated (6-26 months), achromatopsia seems to be a structurally stable condition, although longer-term follow-up is needed. These data will be useful in assessing foveal cone structure after therapeutic intervention.


Asunto(s)
Defectos de la Visión Cromática/genética , Canales Catiónicos Regulados por Nucleótidos Cíclicos/genética , ADN/genética , Fóvea Central/patología , Mutación , Células Fotorreceptoras Retinianas Conos/patología , Agudeza Visual , Adolescente , Adulto , Niño , Defectos de la Visión Cromática/diagnóstico , Defectos de la Visión Cromática/fisiopatología , Canales Catiónicos Regulados por Nucleótidos Cíclicos/metabolismo , Análisis Mutacional de ADN , Electrorretinografía , Femenino , Fóvea Central/fisiopatología , Humanos , Estudios Longitudinales , Masculino , Oftalmoscopía/métodos , Células Fotorreceptoras Retinianas Conos/fisiología , Tomografía de Coherencia Óptica/métodos , Adulto Joven
12.
Hum Mol Genet ; 23(25): 6797-806, 2014 Dec 20.
Artículo en Inglés | MEDLINE | ID: mdl-25082829

RESUMEN

Autosomal recessive Stargardt disease (STGD1, MIM 248200) is caused by mutations in the ABCA4 gene. Complete sequencing of ABCA4 in STGD patients identifies compound heterozygous or homozygous disease-associated alleles in 65-70% of patients and only one mutation in 15-20% of patients. This study was designed to find the missing disease-causing ABCA4 variation by a combination of next-generation sequencing (NGS), array-Comparative Genome Hybridization (aCGH) screening, familial segregation and in silico analyses. The entire 140 kb ABCA4 genomic locus was sequenced in 114 STGD patients with one known ABCA4 exonic mutation revealing, on average, 200 intronic variants per sample. Filtering of these data resulted in 141 candidates for new mutations. Two variants were detected in four samples, two in three samples, and 20 variants in two samples, the remaining 117 new variants were detected only once. Multimodal analysis suggested 12 new likely pathogenic intronic ABCA4 variants, some of which were specific to (isolated) ethnic groups. No copy number variation (large deletions and insertions) was detected in any patient suggesting that it is a very rare event in the ABCA4 locus. Many variants were excluded since they were not conserved in non-human primates, were frequent in African populations and, therefore, represented ancestral, and not disease-associated, variants. The sequence variability in the ABCA4 locus is extensive and the non-coding sequences do not harbor frequent mutations in STGD patients of European-American descent. Defining disease-associated alleles in the ABCA4 locus requires exceptionally well characterized large cohorts and extensive analyses by a combination of various approaches.


Asunto(s)
Transportadoras de Casetes de Unión a ATP/genética , Sitios Genéticos , Variación Genética , Degeneración Macular/congénito , Mutación , Alelos , Población Negra , Estudios de Casos y Controles , Hibridación Genómica Comparativa , Exones , Femenino , Expresión Génica , Genes Recesivos , Heterocigoto , Secuenciación de Nucleótidos de Alto Rendimiento , Homocigoto , Humanos , Intrones , Degeneración Macular/etnología , Degeneración Macular/genética , Degeneración Macular/patología , Masculino , Linaje , Enfermedad de Stargardt , Población Blanca
13.
Hum Mol Genet ; 23(21): 5827-37, 2014 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-24899048

RESUMEN

Neurodegenerative diseases affecting the macula constitute a major cause of incurable vision loss and exhibit considerable clinical and genetic heterogeneity, from early-onset monogenic disease to multifactorial late-onset age-related macular degeneration (AMD). As part of our continued efforts to define genetic causes of macular degeneration, we performed whole exome sequencing in four individuals of a two-generation family with autosomal dominant maculopathy and identified a rare variant p.Glu1144Lys in Fibrillin 2 (FBN2), a glycoprotein of the elastin-rich extracellular matrix (ECM). Sanger sequencing validated the segregation of this variant in the complete pedigree, including two additional affected and one unaffected individual. Sequencing of 192 maculopathy patients revealed additional rare variants, predicted to disrupt FBN2 function. We then undertook additional studies to explore the relationship of FBN2 to macular disease. We show that FBN2 localizes to Bruch's membrane and its expression appears to be reduced in aging and AMD eyes, prompting us to examine its relationship with AMD. We detect suggestive association of a common FBN2 non-synonymous variant, rs154001 (p.Val965Ile) with AMD in 10 337 cases and 11 174 controls (OR = 1.10; P-value = 3.79 × 10(-5)). Thus, it appears that rare and common variants in a single gene--FBN2--can contribute to Mendelian and complex forms of macular degeneration. Our studies provide genetic evidence for a key role of elastin microfibers and Bruch's membrane in maintaining blood-retina homeostasis and establish the importance of studying orphan diseases for understanding more common clinical phenotypes.


Asunto(s)
Estudios de Asociación Genética , Variación Genética , Degeneración Macular/genética , Proteínas de Microfilamentos/genética , Adulto , Anciano , Secuencia de Aminoácidos , Lámina Basal de la Coroides/metabolismo , Análisis Mutacional de ADN , Exoma , Matriz Extracelular/metabolismo , Fibrilina-2 , Fibrilinas , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Degeneración Macular/diagnóstico , Masculino , Metaanálisis como Asunto , Proteínas de Microfilamentos/metabolismo , Persona de Mediana Edad , Modelos Moleculares , Datos de Secuencia Molecular , Mutación , Linaje , Conformación Proteica , Estabilidad Proteica , Retina/metabolismo , Retina/patología , Alineación de Secuencia
14.
Ophthalmology ; 123(1): 9-18, 2016 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-26507665

RESUMEN

PURPOSE: To identify specific mutations causing North Carolina macular dystrophy (NCMD). DESIGN: Whole-genome sequencing coupled with reverse transcription polymerase chain reaction (RT-PCR) analysis of gene expression in human retinal cells. PARTICIPANTS: A total of 141 members of 12 families with NCMD and 261 unrelated control individuals. METHODS: Genome sequencing was performed on 8 affected individuals from 3 families affected with chromosome 6-linked NCMD (MCDR1) and 2 individuals affected with chromosome 5-linked NCMD (MCDR3). Variants observed in the MCDR1 locus with frequencies <1% in published databases were confirmed using Sanger sequencing. Confirmed variants absent from all published databases were sought in 8 additional MCDR1 families and 261 controls. The RT-PCR analysis of selected genes was performed in stem cell-derived human retinal cells. MAIN OUTCOME MEASURES: Co-segregation of rare genetic variants with disease phenotype. RESULTS: Five sequenced individuals with MCDR1-linked NCMD shared a haplotype of 14 rare variants spanning 1 Mb of the disease-causing allele. One of these variants (V1) was absent from all published databases and all 261 controls, but was found in 5 additional NCMD kindreds. This variant lies in a DNase 1 hypersensitivity site (DHS) upstream of both the PRDM13 and CCNC genes. Sanger sequencing of 1 kb centered on V1 was performed in the remaining 4 NCMD probands, and 2 additional novel single nucleotide variants (V2 in 3 families and V3 in 1 family) were identified in the DHS within 134 bp of the location of V1. A complete duplication of the PRDM13 gene was also discovered in a single family (V4). The RT-PCR analysis of PRDM13 expression in developing retinal cells revealed marked developmental regulation. Next-generation sequencing of 2 individuals with MCDR3-linked NCMD revealed a 900-kb duplication that included the entire IRX1 gene (V5). The 5 mutations V1 to V5 segregated perfectly in the 102 affected and 39 unaffected members of the 12 NCMD families. CONCLUSIONS: We identified 5 rare mutations, each capable of arresting human macular development. Four of these strongly implicate the involvement of PRDM13 in macular development, whereas the pathophysiologic mechanism of the fifth remains unknown but may involve the developmental dysregulation of IRX1.


Asunto(s)
Cromosomas Humanos Par 6/genética , Distrofias Hereditarias de la Córnea/genética , Proteínas del Ojo/genética , Polimorfismo Genético , ARN/genética , Adolescente , Adulto , Niño , Preescolar , Distrofias Hereditarias de la Córnea/diagnóstico , Distrofias Hereditarias de la Córnea/metabolismo , Proteínas del Ojo/metabolismo , Familia , Femenino , Angiografía con Fluoresceína , Fondo de Ojo , Ligamiento Genético , Humanos , Inmunohistoquímica , Masculino , Linaje , Fenotipo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Tomografía de Coherencia Óptica , Adulto Joven
15.
Doc Ophthalmol ; 132(3): 157-66, 2016 06.
Artículo en Inglés | MEDLINE | ID: mdl-27033713

RESUMEN

PURPOSE: The purpose of this study was to evaluate pupillary light reflexes (PLRs) mediated by rod, cone, and intrinsically photosensitive retinal ganglion cell pathways as indices of outer- and inner-retinal function in patients who have enhanced S-cone syndrome (ESCS) due to NR2E3 mutations. METHODS: Four patients with ESCS (ages 16-23 years) participated in the study. Subjects were tested with long- and short-wavelength single-flash full-field ERG stimuli under light-adapted conditions. They were also tested with an established pupillometry protocol involving 1-s duration, long- and short-wavelength stimuli under dark- and light-adapted conditions. The PLR was measured as a function of stimulus luminance. Transient PLRs were measured under all conditions, and sustained PLRs were measured under the highest luminance dark-adapted condition. RESULTS: Two-color light-adapted full-field ERGs demonstrated larger amplitude responses for short-wavelength stimuli relative to long-wavelength stimuli of the same photopic luminance, with three of four ESCS patients having super-normal a-wave amplitudes to the short-wavelength stimulus. b/a wave ratios were reduced in all four cases. Transient PLRs elicited by low-luminance stimuli under dark-adapted conditions (rod-mediated) were unrecordable, whereas the sustained PLRs elicited by high-luminance stimuli (melanopsin-mediated) were normal. Cone-mediated PLRs were recordable for all four patients, but generally lower than normal in amplitude. However, the cone-mediated PLR was larger for the short-wavelength stimulus compared to the photopically matched long-wavelength stimulus at high luminances, a pattern that was not observed for control subjects. None of the PLR conditions demonstrated "super-normal" responses. CONCLUSION: ESCS patients appear to have generally well-preserved cone- and melanopsin-mediated PLRs, indicating intact inner-retinal function. Two-color pupillometry demonstrates greater sensitivity to short-wavelength light under higher-luminance conditions and could complement the ERG as a tool for evaluating retinal function in ESCS.


Asunto(s)
Visión de Colores/fisiología , Enfermedades Hereditarias del Ojo/fisiopatología , Luz , Receptores Nucleares Huérfanos/genética , Reflejo Pupilar/fisiología , Retina/fisiopatología , Degeneración Retiniana/fisiopatología , Trastornos de la Visión/fisiopatología , Adolescente , Adulto , Adaptación a la Oscuridad/fisiología , Electrorretinografía/métodos , Enfermedades Hereditarias del Ojo/genética , Femenino , Humanos , Masculino , Mutación , Estimulación Luminosa , Reflejo Pupilar/efectos de la radiación , Células Fotorreceptoras Retinianas Conos/fisiología , Degeneración Retiniana/genética , Células Ganglionares de la Retina/fisiología , Células Fotorreceptoras Retinianas Bastones/fisiología , Trastornos de la Visión/genética , Adulto Joven
16.
Doc Ophthalmol ; 133(1): 61-70, 2016 08.
Artículo en Inglés | MEDLINE | ID: mdl-27369766

RESUMEN

PURPOSE: To evaluate rod-isolated, cone-isolated, and combined rod and cone flicker electroretinograms (ERGs) as a possible means to identify electrophysiological abnormalities in carriers of X-linked retinoschisis (XLRS). METHODS: Full-field ERGs were recorded from six carriers of XLRS (aged 34-66 years) and eight normally sighted subjects (aged 27-59 years) under rod-isolated (ERGR), cone-isolated (ERGC), and combined rod and cone (ERGR+C) conditions. ERGs were obtained using a four-primary LED-based ganzfeld photostimulator and standard recording techniques. The four primaries were modulated sinusoidally in phase to achieve combined rod and cone activation (ERGR+C) or in different phases to achieve ERGR and ERGC by means of triple silent substitution. After 30 min of dark adaptation, 8- and 15-Hz ERGR, ERGC, and ERGR+C responses were obtained at a mean luminance level of 24 scot. cd/m(2). Standard ISCEV ERGs were also obtained from each subject. RESULTS: The ISCEV and 15-Hz flicker ERGs were generally within the normal range for the carriers. The 8-Hz ERGR, ERGC, and ERGR+C amplitudes were also generally normal. In contrast, the carriers had ERGR, ERGC, and ERGR+C timing abnormalities, with phase advances beyond the range of normal for the ERGR (four carriers), ERGC (four carriers), and ERGR+C (three carriers). Only one carrier had normal 8-Hz responses under all conditions. CONCLUSIONS: The 8-Hz ERG timing abnormalities in five of six carriers indicate that retinal function is not necessarily normal in carriers of XLRS. The 8-Hz flicker ERG may be useful for studying retinal function in these individuals.


Asunto(s)
Electrorretinografía/métodos , Células Fotorreceptoras Retinianas Conos/fisiología , Células Fotorreceptoras Retinianas Bastones/fisiología , Retinosquisis/fisiopatología , Visión Ocular/fisiología , Adulto , Anciano , Estudios de Casos y Controles , Adaptación a la Oscuridad/fisiología , Femenino , Fusión de Flicker/fisiología , Humanos , Masculino , Persona de Mediana Edad , Estimulación Luminosa/métodos , Retina
17.
Hum Mol Genet ; 22(25): 5136-45, 2013 Dec 20.
Artículo en Inglés | MEDLINE | ID: mdl-23918662

RESUMEN

Mutations in ABCA4 cause Stargardt disease and other blinding autosomal recessive retinal disorders. However, sequencing of the complete coding sequence in patients with clinical features of Stargardt disease sometimes fails to detect one or both mutations. For example, among 208 individuals with clear clinical evidence of ABCA4 disease ascertained at a single institution, 28 had only one disease-causing allele identified in the exons and splice junctions of the primary retinal transcript of the gene. Haplotype analysis of these 28 probands revealed 3 haplotypes shared among ten families, suggesting that 18 of the 28 missing alleles were rare enough to be present only once in the cohort. We hypothesized that mutations near rare alternate splice junctions in ABCA4 might cause disease by increasing the probability of mis-splicing at these sites. Next-generation sequencing of RNA extracted from human donor eyes revealed more than a dozen alternate exons that are occasionally incorporated into the ABCA4 transcript in normal human retina. We sequenced the genomic DNA containing 15 of these minor exons in the 28 one-allele subjects and observed five instances of two different variations in the splice signals of exon 36.1 that were not present in normal individuals (P < 10(-6)). Analysis of RNA obtained from the keratinocytes of patients with these mutations revealed the predicted alternate transcript. This study illustrates the utility of RNA sequence analysis of human donor tissue and patient-derived cell lines to identify mutations that would be undetectable by exome sequencing.


Asunto(s)
Transportadoras de Casetes de Unión a ATP/genética , Empalme Alternativo/genética , Retina/patología , Adulto , Anciano de 80 o más Años , Alelos , Exoma/genética , Exones/genética , Femenino , Haplotipos , Humanos , Degeneración Macular/genética , Degeneración Macular/fisiopatología , Masculino , Mutación , Linaje , Sitios de Empalme de ARN/genética , Enfermedad de Stargardt
18.
Hum Mol Genet ; 22(3): 593-607, 2013 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-23139242

RESUMEN

Best disease (BD) is an inherited degenerative disease of the human macula that results in progressive and irreversible central vision loss. It is caused by mutations in the retinal pigment epithelium (RPE) gene BESTROPHIN1 (BEST1), which, through mechanism(s) that remain unclear, lead to the accumulation of subretinal fluid and autofluorescent waste products from shed photoreceptor outer segments (POSs). We employed human iPS cell (hiPSC) technology to generate RPE from BD patients and unaffected siblings in order to examine the cellular and molecular processes underlying this disease. Consistent with the clinical phenotype of BD, RPE from mutant hiPSCs displayed disrupted fluid flux and increased accrual of autofluorescent material after long-term POS feeding when compared with hiPSC-RPE from unaffected siblings. On a molecular level, RHODOPSIN degradation after POS feeding was delayed in BD hiPSC-RPE relative to unaffected sibling hiPSC-RPE, directly implicating impaired POS handling in the pathophysiology of the disease. In addition, stimulated calcium responses differed between BD and normal sibling hiPSC-RPE, as did oxidative stress levels after chronic POS feeding. Subcellular localization, fractionation and co-immunoprecipitation experiments in hiPSC-RPE and human prenatal RPE further linked BEST1 to the regulation and release of endoplasmic reticulum calcium stores. Since calcium signaling and oxidative stress are critical regulators of fluid flow and protein degradation, these findings likely contribute to the clinical picture of BD. In a larger context, this report demonstrates the potential to use patient-specific hiPSCs to model and study maculopathies, an important class of blinding disorders in humans.


Asunto(s)
Células Madre Pluripotentes Inducidas/citología , Distrofia Macular Viteliforme/genética , Distrofia Macular Viteliforme/fisiopatología , Animales , Bestrofinas , Calcio/metabolismo , Bovinos , Diferenciación Celular , Línea Celular , Canales de Cloruro/genética , Canales de Cloruro/metabolismo , Proteínas del Ojo/genética , Proteínas del Ojo/metabolismo , Regulación de la Expresión Génica , Homeostasis , Humanos , Inmunohistoquímica , Inmunoprecipitación , Mácula Lútea/patología , Microscopía Electrónica de Transmisión , Estrés Oxidativo , Fagocitosis , Segmento Externo de las Células Fotorreceptoras Retinianas/metabolismo , Epitelio Pigmentado de la Retina/citología , Epitelio Pigmentado de la Retina/patología , Distrofia Macular Viteliforme/metabolismo
19.
Doc Ophthalmol ; 130(2): 111-9, 2015 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-25579805

RESUMEN

PURPOSE: To evaluate rod and cone contributions to the dark-adapted 15-Hz flicker electroretinogram (ERG) across a broad range of stimulus luminances by comparing rod-isolating (ERGR), cone-isolating (ERGC), and non-receptor-specific (ERGR+C) responses. METHODS: Dark-adapted, full-field 15-Hz ERGs were obtained from four normally sighted subjects (ages 29-36 years) using a four-primary LED-based stimulating system. The primaries were either modulated sinusoidally in phase (ERGR+C) or were modulated in counter-phase to achieve rod isolation (ERGR) or cone isolation (ERGC) by means of triple silent substitution. Measurements were made for a broad range of luminances (-2.5 to 1.8 log scot. cd/m(2) in 0.2 log unit steps). Fourier analysis was used to obtain the amplitude and phase of the fundamental response component at each stimulus luminance. RESULTS: Stimulus luminance had different effects on response amplitudes and phases under the three paradigms. Specifically, ERGC amplitude and phase increased monotonically as luminance increased. The effects on ERGR+C and ERGR were complex: ERGR+C and ERGR amplitude was small and the phase decreased for low luminances, whereas amplitude and phase increased sharply at moderate luminances. For high luminances, ERGR+C amplitude and phase increased, whereas ERGR amplitude decreased and phase was approximately constant. CONCLUSIONS: At low luminances, the ERGR+C and ERGR functions can be attributed to interactions between two rod pathways. At high luminances, the functions can be accounted for by interactions between rod and cone pathways (ERGR+C) or rod insensitivity (ERGR). The ERGR paradigm minimizes cone intrusion, permitting assessment of rod function over a large range of luminance levels.


Asunto(s)
Adaptación a la Oscuridad/fisiología , Electrorretinografía , Células Fotorreceptoras Retinianas Conos/fisiología , Células Fotorreceptoras Retinianas Bastones/fisiología , Adulto , Femenino , Análisis de Fourier , Voluntarios Sanos , Humanos , Masculino , Estimulación Luminosa
20.
Hum Mutat ; 35(10): 1187-94, 2014 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-25066811

RESUMEN

Autosomal recessive Stargardt disease (STGD1) is caused by hundreds of mutations in the ABCA4 gene, which are often specific to racial and ethnic groups. Here, we investigated the ABCA4 variation and their phenotypic expression in a cohort of 44 patients of African American descent, a previously under-characterized racial group. Patients were screened for mutations in ABCA4 by next-generation sequencing and array-comparative genomic hybridization (aCGH), followed by analyses for pathogenicity by in silico programs. Thorough ophthalmic examination was performed on all patients. At least two (expected) disease-causing alleles in the ABCA4 gene were identified in 27 (61.4%) patients, one allele in 11 (25%) patients, and no ABCA4 mutations were found in six (13.6%) patients. Altogether, 39 different disease-causing ABCA4 variants, including seven new, were identified on 65 (74%) chromosomes, most of which were unique for this racial group. The most frequent ABCA4 mutation in this cohort was c.6320G>A (p.(R2107H)), representing 19.3% of all disease-associated alleles. No large copy number variants were identified in any patient. Most patients reported later onset of symptoms. In summary, the ABCA4 mutation spectrum in patients of West African descent differs significantly from that in patients of European descent, resulting in a later onset and "milder" disease.


Asunto(s)
Transportadoras de Casetes de Unión a ATP/genética , Mutación , Adulto , Negro o Afroamericano , Hibridación Genómica Comparativa , Análisis Mutacional de ADN , Electrorretinografía , Femenino , Predisposición Genética a la Enfermedad , Humanos , Degeneración Macular/etnología , Degeneración Macular/genética , Masculino , Persona de Mediana Edad , Enfermedad de Stargardt , Estados Unidos , Población Blanca
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