Defects in the Cell Signaling Mediator ß-Catenin Cause the Retinal Vascular Condition FEVR.

Familial exudative vitreoretinopathy (FEVR) is an inherited blinding disorder characterized by the abnormal development of the retinal vasculature. The majority of mutations identified in FEVR are found within four genes that encode the receptor complex (FZD4, LRP5, and TSPAN12) and ligand (NDP) of a molecular pathway that controls angiogenesis, the Norrin-ß-catenin signaling pathway. However, half of all FEVR-affected case subjects do not harbor mutations in these genes, indicating that further mutated genes remain to be identified. Here we report the identification of mutations in CTNNB1, the gene encoding ß-catenin, as a cause of FEVR. We describe heterozygous mutations (c.2142_2157dup [p.His720∗] and c.2128C>T [p.Arg710Cys]) in two dominant FEVR-affected families and a de novo mutation (c.1434_1435insC [p.Glu479Argfs∗18]) in a simplex case subject. Previous studies have reported heterozygous de novo CTNNB1 mutations as a cause of syndromic intellectual disability (ID) and autism spectrum disorder, and somatic mutations are linked to many cancers. However, in this study we show that Mendelian inherited CTNNB1 mutations can cause non-syndromic FEVR and that FEVR can be a part of the syndromic ID phenotype, further establishing the role that ß-catenin signaling plays in the development of the retinal vasculature.

Mutations in TSPAN12 cause autosomal-dominant familial exudative vitreoretinopathy.

Familial exudative vitreoretinopathy (FEVR) is an inherited blinding disorder of the retinal vascular system. Although mutations in three genes (LRP5, FZD4, and NDP) are known to cause FEVR, these account for only a fraction of FEVR cases. The proteins encoded by these FEVR genes form part of a signaling complex that activates the Norrin-beta-catenin signaling pathway. Recently, through a large-scale reverse genetic screen in mice, Junge and colleagues identified an additional member of this signaling complex, Tspan12. Here, we report that mutations in TSPAN12 also cause autosomal-dominant FEVR. We describe seven mutations identified in a cohort of 70 FEVR patients in whom we had already excluded the known FEVR genes. This study provides further evidence for the importance of the Norrin-beta-catenin signaling pathway in the development of the retinal vasculature and also indicates that more FEVR genes remain to be identified.

Mutations in CNNM4 cause Jalili syndrome, consisting of autosomal-recessive cone-rod dystrophy and amelogenesis imperfecta.

The combination of recessively inherited cone-rod dystrophy (CRD) and amelogenesis imperfecta (AI) was first reported by Jalili and Smith in 1988 in a family subsequently linked to a locus on chromosome 2q11, and it has since been reported in a second small family. We have identified five further ethnically diverse families cosegregating CRD and AI. Phenotypic characterization of teeth and visual function in the published and new families reveals a consistent syndrome in all seven families, and all link or are consistent with linkage to 2q11, confirming the existence of a genetically homogenous condition that we now propose to call Jalili syndrome. Using a positional-candidate approach, we have identified mutations in the CNNM4 gene, encoding a putative metal transporter, accounting for the condition in all seven families. Nine mutations are described in all, three missense, three terminations, two large deletions, and a single base insertion. We confirmed expression of Cnnm4 in the neural retina and in ameloblasts in the developing tooth, suggesting a hitherto unknown connection between tooth biomineralization and retinal function. The identification of CNNM4 as the causative gene for Jalili syndrome, characterized by syndromic CRD with AI, has the potential to provide new insights into the roles of metal transport in visual function and biomineralization.

Importance of Anatomical Efficacy for Disease Control in Neovascular AMD: An Expert Opinion.

BACKGROUND: Neovascular age-related macular degeneration (nAMD) presents a significant treatment burden for patients, carers and medical retina services. However, significant debate remains regarding how best to manage nAMD when assessing disease activity by optical coherence tomography (OCT), and particularly the significance of different types of fluid and how the understanding of anatomical efficacy can influence treatment strategies. This article provides opinion on the practical implications of anatomical efficacy and significance of fluid in the management of nAMD and proposes recommendations for healthcare professionals (HCPs) to improve understanding and promote best practice to achieve disease control. METHODS: An evidence-based review was performed and an expert panel debate from the Retina Outcomes Group (ROG), a forum of retinal specialists, provided insights and recommendations on the definition, role and practical implications of anatomical efficacy and the significance of fluid at the macula in the management of nAMD. RESULTS: The ROG has developed recommendations for achieving disease control through a zero-tolerance approach to the presence of fluid in nAMD as patients who avoid fluctuations in fluid at the macula have better visual outcomes. Recommendations cover five key areas: service protocol, training, regimen, multidisciplinary teams and engagement. This approach facilitates more standardised protocol-based treatment strategies. CONCLUSIONS: Targeting a fluid-free macula and aiming for disease control are essential to improve outcomes. As new therapies and technologies become available, drying the macula and maintaining disease control will become even more achievable. The outlined recommendations aim to promote best practice among HCPs and medical retina services to improve patient outcomes.

Familial exudative vitreoretinopathy and DiGeorge syndrome: a new locus for familial exudative vitreoretinopathy on chromosome 22q11.2?

PURPOSE: To describe a patient with DiGeorge syndrome in association with familial exudative vitreoretinopathy (FEVR). DESIGN: Observational case report. PARTICIPANTS: A newborn female and her parents. METHODS: Family members were examined by slit-lamp biomicroscopy and indirect ophthalmoscopy. Deletion mapping was performed by fluorescent in situ hybridization and genotyping. Mutation screening was undertaken by direct sequencing. MAIN OUTCOME MEASURES: The presence or absence of a microdeletion on chromosome 22q11.2 in the patient and her parents and mutation screening of FZD4 and LRP5 in the patient. RESULTS: The patient had classical features of DiGeorge syndrome and FEVR. A de novo microdeletion on chromosome 22q11.2 was found in the patient, confirming the diagnosis of DiGeorge syndrome. No mutations were identified in the known FEVR genes. CONCLUSIONS: Patients with DiGeorge syndrome should have a dilated retinal examination to look for signs of FEVR. Chromosome 22q11.2 may represent a novel locus for FEVR.

Identification of a locus on chromosome 2q11 at which recessive amelogenesis imperfecta and cone-rod dystrophy cosegregate.

A consanguineous Arab pedigree in which recessive amelogenesis imperfecta (AI) and cone-rod dystrophy cosegregate, was screened for linkage to known retinal dystrophy and tooth abnormality loci by genotyping neighbouring microsatellite markers. This analysis resulted in linkage with a maximum lod score of 7.03 to the marker D2S2187 at the achromatopsia locus on chromosome 2q11, and haplotype analysis placed the gene(s) involved in a 2 cM/5 Mb interval between markers D2S2209 and D2S373. The CNGA3 gene, known to be involved in achromatopsia, lies in this interval but thorough analysis of its coding sequence revealed no mutation. Furthermore, affected individuals in four consanguineous recessive pedigrees with AI but without CRD were heterozygous at this locus, excluding it as a common cause of non-syndromic recessive AI. It remains to be established whether this pedigree is segregating two closely linked mutations causing disparate phenotypes or whether a single defect is causing pathology in both teeth and eyes.

Spectrum and frequency of FZD4 mutations in familial exudative vitreoretinopathy.

PURPOSE: Mutations in the frizzled-4 gene (FZD4) have recently been associated with autosomal dominant familial exudative vitreoretinopathy (FEVR) in families linking to the EVR1 locus on the long arm of chromosome 11. The purpose of this study was to screen FZD4 in a panel of 40 patients with FEVR to identify the types and location of mutations and to calculate what proportion of this heterogeneous condition is attributable to FZD4 mutations. METHODS: PCR products were generated from genomic DNA with primers designed to amplify the coding sequence of FZD4. The PCR products were screened for mutations by single-strand conformational polymorphism-heteroduplex analysis (SSCP-HA) and by direct sequencing. RESULTS: In total, eight mutations were identified, seven of which were novel. Three were deletions (c957delG, c1498delA, and c1501-1502delCT), one was a nonsense mutation (Q505X), and four were missense mutations (G36D, M105T, M157V, and S497F). CONCLUSIONS: Eight mutations have been identified in the FZD4 gene in a cohort of 40 unrelated patients with FEVR. This result indicates that FZD4 mutations are responsible for only 20% of FEVR index cases and suggests that the other FEVR loci may account for more cases than previously anticipated.

Identification of a fourth locus (EVR4) for familial exudative vitreoretinopathy (FEVR).

PURPOSE: Familial exudative vitreoretinopathy (FEVR) is a genetically heterogeneous inherited blinding disorder of the retinal vascular system. To date three loci have been mapped: EVR1 on chromosome 11q, EVR2 on chromosome Xp, and EVR3 on chromosome 11p. The gene underlying EVR3 remains unidentified whilst the EVR2 gene, which encodes the Norrie disease protein (NDP), was identified over a decade ago. More recently, FZD4, the gene that encodes the Wnt receptor Frizzled-4, was identified as the mutated gene at the EVR1 locus. The purpose of this study was to screen FZD4 in a large family previously proven to be linked to the EVR1 locus. METHODS: PCR products were generated using genomic DNA from affected family members with primers designed to amplify the coding sequence of FZD4. The PCR products were screened for mutations by direct sequencing. Genotyping was performed in all available family members using fluorescently labeled microsatellite markers from chromosome 11q. RESULTS: Sequencing of the EVR1 gene, FZD4, in this family identified no mutation. To investigate this family further we performed high-resolution genotyping with markers spanning chromosome 11q. Haplotype analysis excluded FZD4 as the mutated gene in this family and identified a candidate region approximately 10 cM centromeric to EVR1. This new FEVR locus is flanked by markers D11S1368 (centromeric) and D11S937 (telomeric) and spans approximately 15 cM. CONCLUSIONS: High-resolution genotyping and haplotype analysis excluded FZD4 as the defective gene in a family previously linked to the EVR1 locus. The results indicate that the gene mutated in this family lies centromeric to the EVR1 gene, FZD4, and is also genetically distinct from the EVR3 locus. This new locus has been designated EVR4 and is the fourth FEVR locus to be described.

Analysis of the rdd locus in chicken: a model for human retinitis pigmentosa.

PURPOSE: To identify the locus responsible for the blind mutation rdd (retinal dysplasia and degeneration) in chickens and to further characterise the rdd phenotype. METHODS: The eyes of blind and sighted birds were subjected to ophthalmic, morphometric and histopathological examination to confirm and extend published observations. Electroretinography was used to determine age of onset. Birds were crossed to create pedigrees suitable for genetic mapping. DNA samples were obtained and subjected to a linkage search. RESULTS: Measurement of IOP, axial length, corneal diameter, and eye weight revealed no gross morphological changes in the rdd eye. However, on ophthalmic examination, rdd homozygotes have a sluggish pupillary response, atrophic pecten, and widespread pigmentary disturbance that becomes more pronounced with age. Older birds also have posterior subcapsular cataracts. At three weeks of age, homozygotes have a flat ERG indicating severe loss of visual function. Pathological examination shows thinning of the RPE, ONL, photoreceptors and INL, and attenuation of the ganglion cell layer. From 77 classified backcross progeny, 39 birds were blind and 38 sighted. The rdd mutation was shown to be sex-linked and not autosomal as previously described. Linkage analysis mapped the rdd locus to a small region of the chicken Z chromosome with homologies to human chromosomes 5q and 9p. CONCLUSIONS: Ophthalmic, histopathologic, and electrophysiological observations suggest rdd is similar to human recessive retinitis pigmentosa. Linkage mapping places rdd in a region homologous to human chromosomes 9p and 5q. Candidate disease genes or loci include PDE6A, WGN1, and USH2C. This is the first use of genetic mapping in a chicken model of human disease.

Genetic, ophthalmic, morphometric and histopathological analysis of the Retinopathy Globe Enlarged (rge) chicken.

PURPOSE: To identify the locus responsible for rge (retinopathy globe enlarged) in chickens and further characterise the rge phenotype. METHODS: A colony of chickens carrying the rge mutation was rederived from a single heterozygous animal of the original line. The eyes of blind, heterozygous and normal birds were subjected to ophthalmic, morphometric and histopathological examination to confirm and extend published observations. DNA samples were obtained and subjected to a whole genome linkage search. RESULTS: From 138 classified backcross progeny, 56 birds were blind and 82 sighted. Heterozygous birds were indistinguishable from wild type, but homozygotes had sluggish or unresponsive pupils, posterior sub-capsular lens opacities and an atrophic pecten. The fundus appeared normal with no significant pigmentary disturbance, but axial length and eye weight were increased. Pathology revealed focal retinal lesions. Linkage analysis placed the rge locus in a small region of chicken chromosome 1. CONCLUSIONS: rge is a severe recessive retinal dystrophy in chickens, with associated globe enlargement. Linkage mapping has highlighted chicken chromosome 1 in a region most probably homologous to human chromosomes 7q31-35, 21q21 or 22q12-21. Candidate disease loci include RP10 (IMPDH1) and uncharacterised Ushers (USH1E) and glaucoma (GLC1F) loci.

Recessive mutations in TSPAN12 cause retinal dysplasia and severe familial exudative vitreoretinopathy (FEVR).

PURPOSE: Familial exudative vitreoretinopathy (FEVR) is an inherited disorder that disrupts the development of the retinal vasculature and can result in blindness. FEVR is genetically heterogeneous and mutations in four genes, NDP, FZD4, LRP5, and TSPAN12, encoding components of a novel ligand-receptor complex that activates the Norrin-ß-catenin signaling pathway, account for approximately 50% of cases. We recently identified mutations in TSPAN12 as a cause of dominant FEVR. The purpose of this study was to identify recessive TSPAN12 mutations in FEVR patients. METHODS: Mutation screening was performed by directly sequencing PCR products generated from genomic DNA with primers designed to amplify the coding sequence of TSPAN12. Splicing defects were verified by reverse transcriptase PCR of leukocyte cDNA. RESULTS: TSPAN12 screening in a large dominant FEVR family unexpectedly led to the identification of homozygous mutations in severely affected family members, whereas mildly affected family members were heterozygous. Further screening in a cohort of 10 retinal dysplasia/severe FEVR patients identified an additional three cases with recessive TSPAN12 mutations. In all examined cases, single mutation carriers were mildly affected compared to patients harboring two TSPAN12 mutations. CONCLUSIONS: We report for the first time recessive mutations in TSPAN12 and describe the first genetic cause for the clinical variation seen in FEVR families. Our data raise the possibility that patients with severe FEVR actually may harbor two mutant alleles, derived either from the same gene or potentially from other genes encoding components of the Norrin-ß-catenin signaling pathway.

Mutations in LRP5 or FZD4 underlie the common familial exudative vitreoretinopathy locus on chromosome 11q.

Familial exudative vitreoretinopathy (FEVR) is an inherited blinding disorder of the retinal vascular system. Autosomal dominant FEVR is genetically heterogeneous, but its principal locus, EVR1, is on chromosome 11q13-q23. The gene encoding the Wnt receptor frizzled-4 (FZD4) was recently reported to be the EVR1 gene, but our mutation screen revealed fewer patients harboring mutations than expected. Here, we describe mutations in a second gene at the EVR1 locus, low-density-lipoprotein receptor-related protein 5 (LRP5), a Wnt coreceptor. This finding further underlines the significance of Wnt signaling in the vascularization of the eye and highlights the potential dangers of using multiple families to refine genetic intervals in gene-identification studies.