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Aniridia is a rare congenital condition of abnormal eye development arising principally from heterozygous mutation of the PAX6 gene. Among the multiple complications arising in the eye, aniridia-associated keratopathy (AAK) is a severe vision-impairing condition of the cornea associated with a progressive limbal stem cell deficiency that lacks suitable treatment options. Current mouse models of aniridia do not accurately represent the onset and progression dynamics of human AAK, hindering therapy development. Here, we performed deep phenotyping of a haploinsufficient Pax6+/- small-eye (Sey) mouse model on the 129S1/SvImJ background, which exhibits key features of mild presentation at birth and progressive AAK with aging, mimicking human disease. The model exhibits a slowly progressing AAK phenotype and provides insights into the disease, including disturbed basal epithelial cell organization, function, and marker expression; persistent postnatal lymphangiogenesis; disrupted corneal innervation patterns; and persisting yet altered limbal stem cell marker expression with age. The model recapitulates many of the known features of human disease, enabling investigation of underlying disease mechanisms and, importantly, access to a well-defined temporal window for evaluating future therapeutics.

Congenital aniridia is a rare eye disease characterized by loss of PAX6 protein leading to aniridia-associated keratopathy that significantly reduces vision. The miR-204-5p is a possible regulator of PAX6 function and here we evaluate its effect in multiple in vitro and in vivo models. In vitro, miR-204-5p overexpression suppressed vascular factor ANGPT1 in human limbal stem cells (T-LSC) and Pax6-knockdown LSC (mut-LSC), and in primary human limbal epithelial cells (LEC) at the gene and protein levels and following LPS stimulation. However, miR-204-5p inhibited VEGFA expression only in mut-LSCs and LPS-stimulated LEC. Also, miR-204-5p increased PAX6 expression in mut-LSC and differentiated corneal epithelial cells, but not in LEC. Topical miR-204-5p after LPS-induced keratitis in mice failed to suppress Vegfa, Angpt1, Il-1β, and Tnf-α or rescue Pax6 levels in contrast to in vitro results, although it significantly reduced the inflammatory infiltrate in the cornea. In Pax6 Sey/+ aniridia mice, miR-204-5p did not rescue PAX6 levels or suppress Vegfa, Angpt1, or inhibit the ERK1/2 pathway. While short-term miR-204-5p treatment effectively suppresses VEGFA and ANGPT1 and enhances PAX6 expression in multiple corneal epithelia, effects are variable across primary and immortalized cells. Effects of longer-term in vivo treatment, however, require further study., (© 2024. The Author(s).)

Aniridia-associated keratopathy originates from a haploinsufficiency of the transcription factor PAX6 (PAX6 +/- ). In the corneal epithelium of PAX6 +/- mice, a significant increase in oxidized proteins was observed, accompanied by impaired compensation for elevated oxidative stress (OS). The extent to which limbal fibroblast cells (LFCs) are affected by an increased susceptibility to OS in cases of congenital aniridia (AN) has not been determined, yet. Our aim was to examine the impact of OS on antioxidant enzyme expression in normal and AN-LFCs. Following isolation and culture of primary LFCs (n = 8) and AN-LFCs (n = 8), cells were treated with cobalt chloride for 48 h to chemically induce hypoxic conditions and OS. Subsequently, HIF-1α/-2α, PHD1/2, Nrf2, CAT, SOD1, PRDX6, and GPX1 gene expression was examined by qPCR. SOD1, PRDX6, and GPX1 protein levels were assessed from the cell lysate by Western blot. The induction of hypoxia led to reduced HIF-1α gene expression in both fibroblast groups (p≤0.008), while the decrease in PHD1 was limited to AN-LFCs (p = 0.0007). On the other hand, under hypoxic conditions, PHD2 showed higher mRNA expression in AN-LFCs compared to normal LFCs (p = 0.013). As a result of OS, the mRNA levels of Nrf2 (p<0.0001) and the antioxidant enzymes CAT (p = 0.005), SOD1 (p = 0.005), GPX1 (p = 0.002) decreased in AN-LFCs. This was accompanied by an increased protein expression of SOD1 (p = 0.019) and PRDX6 (p=0.0009). In the normal LFC group, the induced extent of OS had no impact on the gene (p≥0.151) and protein expression (p ≥ 0.629) of antioxidant enzymes, except for the GPX1 mRNA level (p = 0.027). AN-LFCs exhibit higher susceptibility to OS than normal LFCs. Therefore, in AN-LFCs, there are sustained alterations in gene and protein expression of antioxidative enzymes even after 48 h of CoCl 2 treatment., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Aniridia is an autosomal dominant congenital malformation associated with mutations in the PAX6 gene. It can be associated with deletion in the contiguous WT1 gene, leading to WAGR syndrome, characterized by Wilm tumor, aniridia, genitourinary anomalies, and mental retardation. Persistent fetal vasculature is a developmental malformation caused by incomplete regression of hyaloid vasculature. Most cases of persistent fetal vasculature occur sporadically; however, some inherited forms are described. We report a case of genetically confirmed WAGR associated with congenital cataract and persistent fetal vasculature., (Copyright © 2023 American Association for Pediatric Ophthalmology and Strabismus. Published by Elsevier Inc. All rights reserved.)

Background: The genotype characteristics and their associated clinical phenotypes in patients with aniridia were analyzed to explore pathogenic variants using whole-exome sequencing., Methods: One patient with aniridia was enrolled at the Beijing Tongren Hospital. Comprehensive ophthalmic and general examinations were performed on the patient. DNA was extracted from the patient, and whole-exome sequencing was performed to identify the causative variant. The pathogenicity of the variant was predicted using in silico analysis and evaluated according to American College of Medical Genetics and Genomics guidelines. Relationships between genetic variants and clinical features were analyzed., Results: In addition to the classical aniridia phenotype showing complete iris aplasia, foveal hypoplasia, and ectopic lentis, the patient also exhibited spontaneous reattachment rhegmatogenous retinal detachment (SRRRD). Whole-exome sequencing identified a novel heterozygous variant, exon8:c.640&#95;646del:p.R214Pfs*28., Conclusions: The present study broadens the range of genetic variants described in aniridia and presents an aniridia patient with SRRRD., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Aniridia, which is a rare congenital defect of the eye, consists of iris hypoplasia or aplasia, and additional ocular abnormalities. It is most commonly caused by autosomal dominant PAX6 gene mutations. However, in about 30% of cases, it is associated with chromosomal rearrangements in the 11p13 region. The aim of this study was to identify the potential PAX6 gene variants, which could cause the isolated aniridia. Eight patients with isolated aniridia were included in this study. MLPA analysis allowed in the past to exclude large structural rearrangements of the PAX6 and adjacent genes like WT1. Blood samples were collected from the patients (and their families in familial cases) and genomic DNA was extracted from peripheral blood leukocytes and buccal cells. The amplification of the 11 exons of the PAX6 gene was performed. Bidirectional Sanger Sequencing was conducted for the identification of the potentially pathogenic variants, and for the segregation analysis of the identified variant in the family. The results were analyzed with the use of CodonCode Aligner software. In three patients, aniridia was sporadic, whereas in another five cases, the eye defect was familial. The potentially pathogenic variants in the PAX6 gene were found in 6 out of 8 patients with aniridia. We identified four known (c.781C > T, c.607C > T, and c.949C > T twice), and two novel variants (c.258&#95;265del and c.495&#95;496insG). Point mutations in the PAX6 gene are the most frequent cause of aniridia. The investigation of the genetic background of the disease is essential for patients to evaluate recurrence risk in the offspring., (© 2023 Japanese Teratology Society.)

We aimed to study aniridia-related keratopathy (ARK) relevant cell signaling pathways [Notch1, Wnt/β-catenin, Sonic hedgehog (SHH) and mTOR] in normal human fetal corneas compared with normal human adult corneas and ARK corneas. We found that fetal corneas at 20 weeks of gestation (wg) and normal adult corneas showed similar staining patterns for Notch1; however 10-11 wg fetal corneas showed increased presence of Notch1. Numb and Dlk1 had an enhanced presence in the fetal corneas compared with the adult corneas. Fetal corneas showed stronger immunolabeling with antibodies against β-catenin, Wnt5a, Wnt7a, Gli1, Hes1, p-rpS6, and mTOR when compared with the adult corneas. Gene expression of Notch1, Wnt5A, Wnt7A, β-catenin, Hes1, mTOR, and rps6 was higher in the 9-12 wg fetal corneas compared with adult corneas. The cell signaling pathway differences found between human fetal and adult corneas were similar to those previously found in ARK corneas with the exception of Notch1. Analogous profiles of cell signaling pathway activation between human fetal corneas and ARK corneas suggests that there is a less differentiated host milieu in ARK., (© 2022. The Author(s).)

Aniridia is a panocular inherited rare eye disease linked to heterozygous mutations on the PAX6 gene, which fail to properly produce sufficient protein essential for normal eye development and function. Most of the patients suffer from aniridia-related keratopathy, a progressive opacification of the cornea. There is no effective treatment for this blinding disease. Here we screen for small compounds and identified Ritanserin, a serotonin 2A receptor antagonist, that can rescue PAX6 haploinsufficiency of mutant limbal cells, defective cell migration and PAX6-target gene expression. We further demonstrated that Ritanserin activates PAX6 production through the selective inactivation of the MEK/ERK signaling pathway. Our data strongly suggest that repurposing this therapeutic molecule could be effective in preventing or treating existing blindness by restoring corneal transparency., Competing Interests: Declaration of competing interest None., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Monogenic syndromic disorders frequently feature ocular manifestations, one of which is glaucoma. In many cases, glaucoma in children may go undetected, especially in those that have other severe systemic conditions that affect other parts of the eye and the body. Similarly, glaucoma may be the first presenting sign of a systemic syndrome. Awareness of syndromes associated with glaucoma is thus critical both for medical geneticists and ophthalmologists. In this review, we highlight six categories of disorders that feature glaucoma and other ocular or systemic manifestations: anterior segment dysgenesis syndromes, aniridia, metabolic disorders, collagen/vascular disorders, immunogenetic disorders, and nanophthalmos. The genetics, ocular and systemic features, and current and future treatment strategies are discussed. Findings from rare diseases also uncover important genes and pathways that may be involved in more common forms of glaucoma, and potential novel therapeutic strategies to target these pathways.

Congenital aniridia (AN) is a severe autosomal dominant panocular disorder associated with pathogenic variants in the PAX6 gene. Previously, we performed a molecular genetic study of a large cohort of Russian patients with AN and revealed four noncoding nucleotide variants in the PAX6 5'UTR. 14 additional PAX6-5'UTR variants were also reported in the literature, but the mechanism of their pathogenicity remained unclear. In the present study, we experimentally analyze five patient-derived PAX6 5'UTR-variants: four variants that we identified in Russian patients (c.-128-2delA, c.-125dupG, c.-122dupG, c.-118&#95;-117del) and one previously reported (c.-52+5G>C). We show that the variants lead to a decrease in the protein translation efficiency, while mRNA expression level is not significantly reduced. Two of these variants also affect splicing. Furthermore, we predict and experimentally validate the presence of an evolutionarily conserved small uORF in the PAX6 5'UTR. All studied variants lead to the frameshift of the uORF, resulting in its extension. This extended out-of-frame uORF overlaps with the downstream CDS and thereby reduces its translation efficiency. We conclude that the uORF frameshift may be the main mechanism of pathogenicity for at least 15 out of 18 known PAX6 5'UTR variants. Moreover, we predict additional uORFs in the PAX6 5'UTR., (© 2021 Wiley Periodicals LLC.)

Background: Congenital aniridia is a complex ocular disorder, usually associated with severe visual impairment, generally caused by mutations on the PAX6 gene. The clinical phenotype of PAX6 mutations is highly variable, making the genotype-phenotype correlations difficult to establish., Methods: we describe the phenotype of eight patients from seven unrelated families with confirmed mutations in PAX6 , and very different clinical manifestations., Results: Only two patients had the classical aniridia phenotype while the other two presented with aniridia-related manifestations, such as aniridia-related keratopathy or partial aniridia. Congenital cataracts were the main manifestation in three of the patients in this series. All the patients had nystagmus and low visual acuity., Conclusions: The diagnosis of mild forms of aniridia is challenging, but these patients have a potentially blinding hereditary disease that might present with a more severe phenotype in future generations. Clinicians should be aware of the mild aniridia phenotype and request genetic testing to perform an accurate diagnosis.

BackgroundAniridia is a severe autosomal dominant panocular disorder associated with pathogenic sequence variants of the PAX6 gene or 11p13 chromosomal aberrations encompassing the coding and/or regulatory regions of the PAX6 gene in a heterozygous state. Patients with aniridia display several ocular anomalies including foveal hypoplasia, cataract, keratopathy, and glaucoma, which can vary in severity and combination.MethodsA cohort of 155 patients from 125 unrelated families with identified point PAX6 pathogenic variants (118 patients) or large chromosomal 11p13 deletions (37 patients) was analyzed. Genetic causes were divided into 6 types. The occurrence of 6 aniridic eye anomalies was analyzed. Fisher's exact test was applied for 2×2 contingency tables assigning numbers of patients with/without each sign and each type of the PAX6 variants or 11p13 deletions with Benjamini-Hochberg correction. The age of patients with different types of mutation did not differ.ResultsPatients with 3'- cis -regulatory region deletions had a milder aniridia phenotype without keratopathy, nystagmus, or foveal hypoplasia. The phenotypes of the patients with other rearrangements involving 11p13 do not significantly differ from those associated with point pathogenic variants in the PAX6 gene. Missense mutations and genetic variants disrupting splicing are associated with a severe aniridia phenotype and resemble loss-of-function mutations. It is particularly important that in all examined patients, PAX6 mutations were found to be associated with multiple eye malformations. The age of patients with keratopathy, cataract, and glaucoma was significantly higher than the age of patients without these signs.ConclusionWe got clear statistically significant genotype-phenotype correlations in congenital aniridia and evident that aniridia severity indeed had worsened with age., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. No commercial re-use. See rights and permissions. Published by BMJ.)

Introduction: Wilms tumor (WT) is the most common renal malignancy of children and can be seen in WAGR syndrome (WT, aniridia, genitourinary anomalies, and intellectual disability). WAGR results from a contiguous gene deletion within the 11p13 region, encompassing the WT1 gene, often responsible for WT development, and the PAX6 gene, responsible for aniridia. Aniridia, a pan-ocular disease resulting from iris hypoplasia, is thought to increase the risk for WT development if their genetic alteration spans both the WT1 and the PAX6 genes on 11p13. Case Description: We describe a unique case of a patient with aniridia secondary to a heterozygous PAX6 nonsense mutation who developed WT despite no additional identifiable germline genetic drivers for this disease. Discussion: Isolated mutations in PAX6 previously have not been associated with increased risk of WT development case raises the question of if surveillance for WT should be continued in patients with aniridia with an isolated PAX6 mutation identified.

Congenital aniridia is caused by heterozygous mutations in the PAX6 gene. In this disease, congenital iris and foveal hypoplasia is associated with juvenile onset cataract, glaucoma, and corneal keratopathy. In rodents, Pax6 mutations result in a congenital reduction in ocular size that is not typically described in human aniridia. Here, the ocular morphometry of aniridia patients is compared with the lens phenotype of Pax6 +/tm1/Pgr mice to reveal whether there are species differences in Pax6 regulation of lens development and homeostasis. Ultrasound biometry (UBM) revealed that eleven percent of aniridia patients exhibited mild microphthalmia while the anterior chamber depth of aniridic eyes was significantly reduced from 6 months of age onward. Although aniridic lens thickness was normal from birth, it was significantly decreased in aniridic lenses older than 30. Notably, 86% of aniridic lenses exhibited cataractous changes in this cohort. In addition, a significant proportion of aniridia patients develop lens subluxation as they age associated with reduced lens diameter as measured by anterior segment optical coherence tomography (AS-OCT). Analysis of young adult Pax6 +/tm1/Pgr mouse lenses by micro-computed tomography (microCT), bright field and dark field imaging revealed that they are reduced in size but did not exhibit overt cataracts at this age. Overall, this study reveals that congenital microphthalmia as assessed by axial length, or microphakia, as assessed by lens thickness, are not typical in human aniridia, although these are primary manifestations of Pax6 mutations in mice, suggesting that PAX6 regulates some aspects of lens development differently between these species., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Purpose : To evaluate the choroidal thickness (CT) in children with congenital aniridia in comparison with age-matched controls. Methods : This was a cross-sectional, observational study that included 64 eyes of 32 children with congenital aniridia (aged 5-12 years) and 80 eyes of 40 healthy subjects who were age-matched. In all subjects, subfoveal choroidal thickness (SFCT) was assessed at 750-μm intervals from the fovea to 1.5 mm in the temporal and nasal directions with spectral-domain optical coherence tomography (SD-OCT). Results : The mean SFCT was 207.67 ± 30.99 µm in the aniridic eyes. This SFCT was significantly thinner than that in control eyes (288.55 ± 30.06 µm) ( P < .001). The SFCTs at 1.5 mm and 0.75 mm intervals in the temporal and nasal directions from the fovea were also significantly thinner in eyes with aniridia than control eyes ( P < .001).There was a significant negative correlation between the SFCT and axial length in eyes with aniridia (B = -10.60, 95%CI = -19.31~-1.89, P = .017). Conclusions : The subfoveal and parafoveal CTs were significantly thinner in eyes with congenital aniridia than in control eyes. These choroidal changes could open up a new way for the research related to the pathophysiology of congenital aniridia.

Background: Because of the significant occurrence of "WAGR-region" deletions among de novo mutations detected in congenital aniridia, DNA diagnosis is critical for all sporadic cases of aniridia due to its help in making an early diagnosis of WAGR syndrome. Standard cytogenetic karyotype study is a necessary step of molecular diagnostics in patients with deletions and in the patients' parents as it reveals complex chromosomal rearrangements and the risk of having another affected child, as well as to provide prenatal and/or preimplantation diagnostics., Case Presentation: DNA samples were obtained from the proband (a 2-year-old boy) and his two healthy parents. Molecular analysis revealed a 977.065 kb deletion that removed loci of the ELP4, PAX6, and RCN1 genes but did not affect the coding sequence of the WT1 gene. The deletion occurred de novo on the paternal allele. The patient had normal karyotype 46,XY and a de novo pericentric inversion of chromosome 11, inv(11)(p13q14)., Conclusions: We confirmed the diagnosis of congenital aniridia at the molecular level. For the patient, the risk of developing Wilms' tumor is similar to that in the general population. The recurrence risk for sibs in the family is low, but considering the possibility of gonadal mosaicism, it is higher than in the general population.

Background: Congenital aniridia is a severe ocular abnormality characterized by incomplete formation of the iris and many other ocular complications. Most cases are caused by the paired box 6 (PAX6) gene mutations generating premature termination codons (PTCs)., Methods: Ophthalmic examination was performed on a Chinese pedigree with congenital aniridia. The mutation was identified by targeted next-generation sequencing. Nonsense suppression therapy was applied on patient-derived lymphocytes. The PAX6 expression was assayed by real-time polymerase chain reaction and Western blot., Results: Complete aniridia was complicated with horizontal nystagmus, contract, foveal hypoplasia, and microphthalmia. A novel heterozygous c.702&#95;703delinsAT (p.Tyr234*) mutation was found in exon 9 of PAX6, generating a PTC at the homeodomain. There were about 50% reductions of both full-length PAX6 protein and PAX6 mRNA in patient-derived lymphocytes, indicating haploinsufficiency due to nonsense-mediated mRNA decay. Ataluren (PTC124) and geneticin (G418) could induce about 30%-40% translational readthrough. Nonsense suppression therapy restored PAX6 protein to about 65%-70% of unaffected family controls., Conclusion: Our data expanded the genetic and phenotypic variations of congenital aniridia, and showed the therapeutic effect of nonsense suppression on this disease using patient-derived cells., (© 2020 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.)

Background: Congenital aniridia involves total or partial hypoplasia of the iris and is due to a deficiency in PAX6 gene expression. WAGR syndrome is comprised of Wilms tumor, aniridia, genitourinary abnormalities, and intellectual disability. Numerous genitourinary pathologies may be associated with WAGR syndrome, necessitating an evaluation of the genitourinary anatomy. The WT1 is vital for the development of kidneys, ovaries in females, and testes in males. WT1 gene mutations result in a WT1 protein with a decreased ability to bind to DNA, leading to uncontrolled growth, and cell division in the kidney which permits the development of Wilms tumor. A congenital ureteral valve is an exceedingly rare cause of obstructive uropathy., Results: A renal and bladder ultrasound demonstrated a renal cyst. A voiding cystourethrogram revealed grade 3 vesicoureteral reflux, and a MAG3 renal scan showed ureteropelvic junction obstruction and hydronephrosis. A ureteral stent was inserted at 3 months of age after which the renal cyst resolved. The patient was urinary tract infection-free at 27 months of age. Genetic testing confirmed a heterozygous alteration in PAX6 (c.495delG, p.Thr166Leufs*41) and no abnormalities of WT1, excluding WAGR syndrome., Conclusion: The genitourinary risks potentially associated with aniridia necessitate prompt genetic analysis to evaluate for WAGR syndrome., (© 2020 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.)

PAX6 encodes a highly conserved transcription factor necessary for normal development of the eyes and central nervous system. Heterozygous loss-of-function mutations in PAX6 cause the disorder aniridia in humans and the Small eye trait in mice. Aniridia is a congenital and progressive disorder known for ocular phenotypes; however, recently, consequences of PAX6 haploinsufficiency in the brains of aniridia patients have been identified. These findings span structural and functional abnormalities, including deficits in cognitive and sensory processing. Furthermore, some of these abnormalities are accelerated as aniridia patients age. Although some functional abnormalities may be explained by structural changes, variability of results remain, and the effects of PAX6 heterozygous loss-of-function mutations on neuroanatomy, particularly with regard to aging, have yet to be resolved. Our study used high-resolution magnetic resonance imaging (MRI) and histology to investigate structural consequences of such mutations in the adult brain of our aniridia mouse model, Small eye Neuherberg allele (Pax6 SeyNeu/+ ), at two adult age groups. Using both MRI and histology enables a direct comparison with human studies, while providing higher resolution for detection of more subtle changes. We show volumetric changes in major brain regions of the the Pax6 SeyNeu/+ mouse compared to wild-type including genotype- and age-related olfactory bulb differences, age-related cerebellum differences, and genotype-related eye differences. We also show alterations in thickness of major interhemispheric commissures, particularly those anteriorly located within the brain including the optic chiasm, corpus callosum, and anterior commissure. Together, these genotype and age related changes to brain volumes and structures suggest a global decrease in adult brain structural plasticity in our Pax6 SeyNeu/+ mice., Competing Interests: Declaration of Competing Interest There are no conflicts of interest for any of the authors of this study. The funders of this research had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript., (Published by Elsevier B.V.)

The transcription factor PAX6 is essential in ocular development in vertebrates, being considered the master regulator of the eye. During eye development, it is essential for the correct patterning and formation of the multi-layered optic cup and it is involved in the developing lens and corneal epithelium. In adulthood, it is mostly expressed in cornea, iris, and lens. PAX6 is a dosage-sensitive gene and it is highly regulated by several elements located upstream, downstream, and within the gene. There are more than 500 different mutations described to affect PAX6 and its regulatory regions, the majority of which lead to PAX6 haploinsufficiency, causing several ocular and systemic abnormalities. Aniridia is an autosomal dominant disorder that is marked by the complete or partial absence of the iris, foveal hypoplasia, and nystagmus, and is caused by heterozygous PAX6 mutations. Other ocular abnormalities have also been associated with PAX6 changes, and genotype-phenotype correlations are emerging. This review will cover recent advancements in PAX6 regulation, particularly the role of several enhancers that are known to regulate PAX6 during eye development and disease. We will also present an updated overview of the mutation spectrum, where an increasing number of mutations in the non-coding regions have been reported. Novel genotype-phenotype correlations will also be discussed.

Purpose: To investigate fundus autofluorescence (FAF) and other fundus manifestations in congenital aniridia., Methods: Fourteen patients with congenital aniridia and 14 age- and sex-matched healthy controls were examined. FAF images were obtained with an ultra-widefield scanning laser ophthalmoscope. FAF intensity was quantified in the macular fovea and in a macular ring surrounding fovea and related to an internal reference within each image. All aniridia patients underwent an ophthalmologic examination, including optical coherence tomography and slit-lamp biomicroscopy., Results: Mean age was 28.4 ± 15.0 years in both the aniridia and control groups. Fovea could be defined by subjective assessment of FAF images in three aniridia patients (21.4%) and in all controls (P = 0.001). Mean ratio between FAF intensity in the macular ring and fovea was 1.01 ± 0.15 in aniridia versus 1.18 ± 0.09 in controls (P = 0.034). In aniridia, presence of foveal hypoplasia evaluated by biomicroscopy correlated with lack of foveal appearance by subjective analyses of FAF images (P = 0.031) and observation of nystagmus (P = 0.009)., Conclusions: Aniridia patients present a lower ratio between FAF intensity in the peripheral and central macula than do healthy individuals. Both subjective and objective analyses of FAF images are useful tools in evaluation of foveal hypoplasia in aniridia.

Introduction: Corneal tattooing is a noninvasive technique which appears relatively well-tolerated in the medium term. We report the cases of 3 patients with a significant change in the color of their tattoos performed over 5 years previously., Patients and Methods: Three patients with a history of intracorneal tattooing several years previously were studied because of a significant change from their initial color. Each patient's file was reviewed with analysis of slit lamp photographs, OCT and specular microscopy., Results: All three patients experienced a significant color change in their tattoos between 5 and 6 years after surgery. The color had changed to golden-brown., Discussion: Retrospective analysis of the components of the tattoo ink found the presence of iron in the black pigment. We believe that pigments composed of iron oxide are transformed into golden-brown ferric iron oxide in the presence of oxygen in the aqueous environment. The presence of moderate corneal edema in these three cases of multioperated patients could explain, in these specific cases, the occurrence of oxidation typically not described., Conclusion: Corneal tattooing remains a simple and very interesting technique when partial or total absence of iris causes significant photophobia. However, the significant changes in color that we report more than 5 years later suggest omitting iron from the dyes used for the cornea and limiting its use in cases of limited endothelial prognosis. A long-term evaluation of corneal tattoos appears necessary., (Copyright © 2019. Published by Elsevier Masson SAS.)

Nucleotide variants that disrupt normal splicing might be the cause of a large number of diseases. Nevertheless, because of the complexity of splicing regulation, it is not always possible to accurately predict the effect of nucleotide sequence changes on splicing events and mRNA structure. Thereby, a number of newly identified nucleotide variants are falsely classified as VUS (a variant of uncertain significance). In the present study we used the minigene assay to analyze the functional consequences of six intronic (c.142-5T>G, c.142-14C>G, c.142-64A>C, c.141+4A>G, c.1032+ 6T>G, c.682+4delA), one missense (c.140A>G) and one synonymous (c.174C>T) variants in the PAX6 gene found in patients with congenital aniridia. We revealed that all except one (c.142-64A>C) variants lead to the disruption of normal splicing patterns resulting in premature termination codon formation followed by mRNA degradation through the nonsense mediated decay pathway. This produces a null allele of the PAX6 gene. That allowed us to reclassify the analyzed variants as loss-of-function and to establish their functional role.

A foetus affected by a congenital rubella infection can develop congenital rubella syndrome (CRS). Aniridia is the absence of iris, rarely been described in literature in association with CRS, can easily be overlooked, leading to complications e.g. glaucoma and blindness later in life. We report a case of a neonate with CRS and aniridia presenting at a tertiary care hospital.

Aniridia is a panocular disorder characterized chiefly by iris hypoplasia. Most cases result from mutations of the PAX6 gene, which is important in both eye and brain development. In addition to ocular alterations, differences in global brain volume and functional connectivity have been reported in humans with aniridia. Understanding neural alterations in aniridia may require examination of possible differences in white matter structure, as few studies have assessed white matter in this population. The current study utilized diffusion-weighted imaging to assess white matter structure in 11 people with aniridia and 11 healthy comparison participants, matched for sex and age. A map of the local connectome was calculated to compare quantitative anisotropy (QA), an index of white matter tract density, in all white matter voxels, revealing subcomponents of white matter tracts with differing QA between people with aniridia and healthy comparisons. The analysis indicated that QA was lower for people with aniridia in portions of bilateral optic tract [t(20)=-4.23, P=0.001, d=-1.80], bilateral optic radiation [t(20)=-4.06, P=0.001, d=-1.73], forceps major [t(20)=-3.65, P=0.002, d=-1.55], bilateral superior longitudinal fasciculus [left: t(20)=-3.15, P=0.005, d=-1.34; right, t(20)=-4.28, P<0.001, d=-1.83], and right posterior corona radiata [t(20)=-3.19, P=0.006, d=-1.36]. These differences demonstrate that white matter structure is altered in people with aniridia in both visual tracts and associated posterior visual pathways.

The small eye (Sey) mouse is a model of PAX6-aniridia syndrome (aniridia). Aniridia, a congenital ocular disorder caused by heterozygous loss-of-function mutations in PAX6, needs new vision saving therapies. However, high phenotypic variability in Sey mice makes development of such therapies challenging. We hypothesize that genetic background is a major source of undesirable variability in Sey mice. Here we performed a systematic quantitative examination of anatomical, histological, and molecular phenotypes on the inbred C57BL/6J, hybrid B6129F1, and inbred 129S1/SvImJ backgrounds. The Sey allele significantly reduced eye weight, corneal thickness, PAX6 mRNA and protein levels, and elevated blood glucose levels. Surprisingly, Pax6 Sey/Sey brains had significantly elevated Pax6 transcripts compared to Pax6 +/+ embryos. Genetic background significantly influenced 12/24 measurements, with inbred strains introducing severe ocular and blood sugar phenotypes not observed in hybrid mice. Additionally, significant interactions (epistasis) between Pax6 genotype and genetic background were detected in measurements of eye weight, cornea epithelial thickness and cell count, retinal mRNA levels, and blood glucose levels. The number of epistatic interactions was reduced in hybrid mice. In conclusion, severe phenotypes in the unnatural inbred strains reinforce the value of more naturalistic F1 hybrid mice for the development of therapies for aniridia and other disorders.

Background: Congenital aniridia is a severe autosomal dominant binocular developmental disorder, the primary feature of which is congenital absence or hypoplasia of the iris. PAX6 is the main disease-causing gene of congenital aniridia; inheritance is autosomal dominant. But the current mutations do not fully explain this disorder., Methods: We investigated the mutation profile of genes related in three Chinese families with congenital aniridia through targeted sequencing technology. And we validated the candidate variants by PCR-based Sanger sequencing. Different degree impairments of islet function were observed in the patients with aniridia by carbohydrate tolerance butter and insulin release tests in our study., Results: We identified four novel mutations of PAX6 from three Chinese families with congenital aniridia, which included heterozygous double mutation c.879&#95;880delCA (p.S294Cfs*46) and c.1124C>G (p.P375R) in Family 1 with three patients, heterozygous frameshift mutation c.308delG (p.P103Qfs*21) in Family 2 with one patient, and c.1192delT (p.S398Pfs*126) in Family 3 with two patients. The three frameshift mutations of PAX6 are co-segregated with the aniridia from controls in the families, but the novel missense mutation is not co-segregated with the phenotype. The frameshift mutations in Family 1 and Family 2 have effects to truncate the protein, but the frameshift mutation in Family 3 will prolong it. We confirmed the phenomenon of male gonadal mosaicism of PAX6 by the sequencing of two linked novel mutations in Family 1. Most of the patients with isolated aniridia have different degrees of islet damage through related clinical tests., Conclusion: It is therefore noteworthy that we found different types of pathogenic mutation, which have effects of truncating or prolonging protein leaded by frameshift mutation. Our results of this study extended the pathogenic mutation spectrum of PAX6 for congenital aniridia and demonstrated the male germline chimerism by molecular experiments., (© 2018 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.)

Congenital aniridia is a rare genetic disorder characterized by a variable degree of hypoplasia or absence of iris. It is frequently associated with keratopathy, cataract, juvenile‑onset glaucoma and foveal and optic nerve hypoplasia. Mutations in the Paired Box 6 (PAX6) gene on chromosome 11p13 have been demonstrated to cause aniridia. The aim of the present study was to investigate the genetic variations of PAX6 in two sporadic patients from southern China with classic congenital aniridia and cataract. Complete ophthalmic and physical examinations were performed, including best‑corrected visual acuity, intraocular pressure, slit‑lamp examination, fundus examination, optical coherence tomography, ultrasound biomicroscopy, and Pentacam scanning. Genomic DNA was extracted from leukocytes of peripheral blood collected from the two patients, their unaffected parents and 200 unrelated control subjects from the same population. Exons 4‑13 of the PAX6 gene were amplified by polymerase chain reaction and sequenced directly. Patient 1 was affected with aniridia accompanied by congenital cataract and nystagmus. A novel heterozygous PAX6 frameshift mutation c.277delG (p.Glu93SerfsX31) in exon 6 was identified in this patient. Patient 2 was presented with aniridia, congenital cataract, lens subluxation and glaucoma. A recurrent nonsense mutation c.718C>T (p.Arg240X) in exon 9 was identified in this patient. The present results expand the mutation spectrum of PAX6 and will be valuable for genetic counseling in the affected families. Additionally, the identification of these mutations reiterates the importance of PAX6 in ocular development and sheds light on the pathogenesis of congenital aniridia.

Heterozygous PAX6 gene mutations leading to haploinsufficiency are the main cause of congenital aniridia, a rare and progressive panocular disease characterized by reduced visual acuity. Up to 90% of patients suffer from aniridia-related keratopathy (ARK), caused by a combination of factors including limbal epithelial stem cell (LSC) deficiency, impaired healing response and abnormal differentiation of the corneal epithelium. It usually begins in the first decade of life, resulting in recurrent corneal erosions, sub-epithelial fibrosis, and corneal opacification. Unfortunately, there are currently no efficient treatments available for these patients and no in vitro model for this pathology. We used CRISPR/Cas9 technology to introduce into the PAX6 gene of LSCs a heterozygous nonsense mutation found in ARK patients. Nine clones carrying a p.E109X mutation on one allele were obtained with no off-target mutations. Compared with the parental LSCs, heterozygous mutant LSCs displayed reduced expression of PAX6 and marked slow-down of cell proliferation, migration and detachment. Moreover, addition to the culture medium of recombinant PAX6 protein fused to a cell penetrating peptide was able to activate the endogenous PAX6 gene and to rescue phenotypic defects of mutant LSCs, suggesting that administration of such recombinant PAX6 protein could be a promising therapeutic approach for aniridia-related keratopathy. More generally, our results demonstrate that introduction of disease mutations into LSCs by CRISPR/Cas9 genome editing allows the creation of relevant cellular models of ocular disease that should greatly facilitate screening of novel therapeutic approaches. Stem Cells 2018;36:1421-1429., (© 2018 The Authors Stem Cells published by Wiley Periodicals, Inc. on behalf of AlphaMed Press 2018.)

The present study investigated the clinical and mutational spectrum of aniridia in a cohort of 17 affected individuals from six families from Cyprus. Each proband was initially evaluated for copy number variants at the PAX6 locus and subsequently underwent PAX6 mutation screening. Sequence analysis of FOXC1 and PITX2 was performed in patients who did not carry a PAX6 mutation. The most common clinical features in the group of aniridia patients associated with aniridia were nystagmus, cataracts and glaucoma. PAX6 pathogenic mutations were identified in five out of six families (a diagnostic yield of 84%). Previously reported pathogenic mutations in PAX6 were identified in four families, which comprise p.R203*, p.R240* and p.R317*. In addition, a novel pathogenic variant (p.E220Gfs*23) was identified in a single family. No pathogenic mutations were detected in PAX6, FOXC1 or PITX2 in the only patient with a sporadic form of aniridia‑like phenotype, confirming the genetic heterogeneity associated with this disease. To the best of our knowledge this is the first report on the mutational spectrum of PAX6 in aniridia patients of Cypriot ancestry. Mutational screening of PAX6 serves a crucial role in distinguishing isolated from syndromic forms of aniridia, and it may therefore eliminate the need for renal ultrasound scan surveillance, delineate the phenotype and improve genetic counseling.

Aniridia is a rare congenital ocular malformation that follows an autosomal dominant mode of inheritance. Most patients carry pathogenic point mutations in the paired box 6 gene ( PAX6 ), but some carry deletions involving the 11p13 region, encompassing partly or completely PAX6 or the region downstream. We identified a novel deletion, ~564 kb in size located about 46.5 kb downstream of PAX6 in a family with bilateral aniridia and foveal hypoplasia using array-CGH and multiplex ligation-dependent probe amplification. We also reviewall of the reported deletions downstream of PAX6 in patients with aniridia and/or other congenital malformations and define the overlapping region that leads to aniridia when deleted.

Purpose: Aniridia is a congenital disorder caused by variants in the PAX6 gene. In this study, we assessed the involvement of PAX6 in patients with aniridia from Australasia and Southeast Asia., Methods: Twenty-nine individuals with aniridia from 18 families originating from Australia, New Caledonia, Cambodia, Sri Lanka, and Bhutan were included. The PAX6 gene was investigated for sequence variants and analyzed for deletions with multiplex ligation-dependent probe amplification., Results: We identified 11 sequence variants and six chromosomal deletions, including one in mosaic. Four deleterious sequence variants were novel: p.(Pro81HisfsTer12), p.(Gln274Ter), p.(Ile29Thr), and p.(Met1?). Ocular complications were associated with a progressive loss of visual function as shown by a visual acuity ≤ 1.00 logMAR reported in 65% of eyes. The prevalence of keratopathy was statistically significantly higher in the Australasian cohort (78.6%) compared with the Southeast Asian cohort (9.1%, p=0.002). Variants resulting in protein truncating codons displayed limited genotype-phenotype correlations compared with other variants., Conclusions: PAX6 variants and deletions were identified in 94% of patients with aniridia from Australasia and Southeast Asia. This study is the first report of aniridia and variations in PAX6 in individuals from Cambodia, Sri Lanka, Bhutan, and New Caledonia, and the largest cohort from Australia.

Gillespie syndrome (GS) [MIM: 206700] is a very rare condition characterized by bilateral iris defect, congenital hypotonia, cerebellar ataxia and intellectual disability. The typical iris anomaly is considered necessary to the diagnosis of GS. Recently, variants in ITPR1 were described causing GS. Non-neurological features were reported in few patients. Here we describe two consanguineous siblings with GS and a novel homozygous ITPR1 pathogenic variant (p.N984fs). They also present a cardiac defect (pulmonary valve stenosis) and one sib had a genitourinary malformation (ureteropelvic junction obstruction). Our report reinforces ITPR1 as the cause of GS and suggests a possible role of ITPR1 in the development of other organs., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)

Purpose: To relate central corneal epithelial phenotype to degree of keratopathy in a limbal stem cell deficient population., Methods: 37 patients (67 eyes) with aniridia-associated keratopathy (AAK) underwent corneal examination including slit lamp biomicroscopy to determine the Grade of AAK, Cochet-Bonnet esthesiometry, and in vivo confocal microscopy (IVCM) to assess morphology of the central corneal epithelium and subepithelial region., Results: AAK Grade ranged from 1 (limbal involvement only) to 4 (total conjunctivalization), with progression from Grade 1 occurring after the age of 20. 30% of subjects had an asymmetric Grade between eyes. In early-stage AAK (Grades 1-2), central epithelial cells had mixed corneal-conjunctival phenotype, touch sensitivity and subbasal nerves diminished, and mature dendritic cells, inflammatory leukocytes, and blood vessels were present despite central transparency in the slit lamp. In later stages (Grades 3-4) of the LSCD, neural deficit and nerve function worsened, immune cell invasion increased, and lymphatic vessels were detected in several cases. Goblet cells and epithelial cysts were observed to varying degrees in all stages, but without clear association to AAK severity. The clinical grade and progression of AAK was strongly associated with the central corneal epithelial phenotype., Conclusions: AAK is associated with degradation of epithelial phenotype, a neural deficit, and immune compromised status even in the clear central cornea in the earliest stages. IVCM can aid in assessing whether the conditions for limbal stem cell maintenance are likely to exist, based on morphology of the central epithelial microenvironment., (Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.)

Intraocular pressure lowering surgery in congenital aniridia glaucoma (CAG) can be complicated by dysgenesis of the limbal region, anterior chamber angle, iris, and lens. The anterior segments of 23 eyes (17 patients) with congenital aniridia were investigated under general anesthesia using ultrasound biomicroscopy (UBM). The structures of the anterior segment were examined: distance of ciliary body processes from the anterior chamber angle and positioning of Schlemm's canal. A surgical plan was created on the basis of these data. Schlemm's canal was detected in 21 of the 23 examined eyes. The mean distance from the anterior chamber angle was 1.3 ± 0.4 mm (range: 0.5-to 2.1 mm). The mean distance between the anterior chamber angle and the ciliary body was 561 ± 301 µm (range: 270-1,300 µm). The mean prominence of the ciliary body towards the lens was 799 ± 352 µm (range: 210-1,660 µm). This resulted in a precise UBM-based trabeculotomy. In addition, the ciliary body was detected and coagulated ab externo with a diode laser probe (810 nm) using diaphanoscopy and UBM. An initial UBM examination of the anterior segment is essential in eyes with CAG scheduled for trabeculotomy or cyclophotocoagulation. Clin. Anat. 31:64-67, 2018. © 2017 Wiley Periodicals, Inc., (© 2017 Wiley Periodicals, Inc.)

Congenital aniridia is a severe autosomal dominant congenital panocular disorder, mainly associated with pathogenic variants in the PAX6 gene. The objective of the study was to investigate the mutational and clinical spectra of congenital aniridia in a cohort of 117 patients from Russia. Each patient underwent detailed ophthalmological examination. From 91 unrelated families, 110 patients were diagnosed with congenital aniridia and 7 with WAGR syndrome (Wilms tumor, Aniridia, Genitourinary anomalies, and mental Retardation syndrome). The clinical presentation in aniridia patients varied from the complete bilateral absence of the iris (75.5%) to partial aniridia or iris hypoplasia (24.5%). Additional ocular abnormalities were consistent with previous reports. In our cohort, we saw a previously not described high percentage of patients (45%) who showed non-ocular phenotypes. Prevalence of deletions coherent with WAGR syndrome appeared to be 19.4% out of sporadic patients. Among the other aniridia cases, PAX6 deletions were identified in 18 probands, and small intragenic changes were detected in 58 probands with 27 of these mutations being novel and 21 previously reported. In 3 families mosaic mutation was transmitted from a subtly affected parent. Therefore, PAX6 mutations explained 96.7% of aniridia phenotypes in this study with only 3 of 91 probands lacking pathogenic variants in the gene., (© 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Aniridia is a rare, congenital ocular disorder caused by the mutations of the paired box gene-6 (PAX6) (OMIM 607108), which encodes a highly conserved transcriptional regulator. In order to investigate the clinical characterizations and genetic defects of two Chinese families affected with aniridia, we recruited the family members and 200 ethnically matched controls. The entire exons and flanking intronic sequences of the PAX6 gene (NG&#95;008679.1) were analyzed and effects of variants on splicing were assessed in silico and in vitro using exon trapping assay with pET01. The donor site (c.1183+1G>A) mutation identified in family 1 would result in a complete skipping of exon 12 and cause a frameshift and run-on translation past the normal termination codon, creating an enlarged PAX6 protein with extended COOH-terminal domain. Novel c.1033-1&#95;1033delinsCT mutation was detected in family 2. This mutation provoked both complete exon 12 skipping and partial skipping of exon 12 deleting 7bp. This would lead to a frameshift translation and the introduction of pre-mature termination code, which resulted in severely truncated PAX6 protein likely to be degraded. Our study further expands the spectrum of genetic pathology underlying PAX6., (Copyright © 2017. Published by Elsevier B.V.)

The Pax6 transcription factor is essential for development of the brain, eye, olfactory and endocrine systems. Haploinsufficiency of PAX6 in humans and mice causes the congenital condition aniridia, with defects in each of these organs and systems. Identification of the PAX6 transcription networks driving normal development is therefore critical in understanding the pathophysiology observed with loss-of-function defects. Here we have focused on identification of the downstream targets for Pax6 in the developing iris and ciliary body, where we used laser capture microdissection in mouse eyes from E12.5-E16.5, followed by chromatin immunoprecipitation, promoter-reporter assays and immunohistochemistry. We identified 6 differentially expressed genes between wildtype and Pax6 heterozygous mouse tissues and demonstrated that Bmp4, Tgfβ2, and Foxc1 were direct downstream targets of Pax6 in developing iris/ciliary body. These results improve our understanding of how mutations in Bmp4, Tgfβ2, and Foxc1 result in phenocopies of the aniridic eye disease and provide possible targets for therapeutic intervention., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Gillespie syndrome (GS) is characterized by bilateral iris hypoplasia, congenital hypotonia, non-progressive ataxia, and progressive cerebellar atrophy. Trio-based exome sequencing identified de novo mutations in ITPR1 in three unrelated individuals with GS recruited to the Deciphering Developmental Disorders study. Whole-exome or targeted sequence analysis identified plausible disease-causing ITPR1 mutations in 10/10 additional GS-affected individuals. These ultra-rare protein-altering variants affected only three residues in ITPR1: Glu2094 missense (one de novo, one co-segregating), Gly2539 missense (five de novo, one inheritance uncertain), and Lys2596 in-frame deletion (four de novo). No clinical or radiological differences were evident between individuals with different mutations. ITPR1 encodes an inositol 1,4,5-triphosphate-responsive calcium channel. The homo-tetrameric structure has been solved by cryoelectron microscopy. Using estimations of the degree of structural change induced by known recessive- and dominant-negative mutations in other disease-associated multimeric channels, we developed a generalizable computational approach to indicate the likely mutational mechanism. This analysis supports a dominant-negative mechanism for GS variants in ITPR1. In GS-derived lymphoblastoid cell lines (LCLs), the proportion of ITPR1-positive cells using immunofluorescence was significantly higher in mutant than control LCLs, consistent with an abnormality of nuclear calcium signaling feedback control. Super-resolution imaging supports the existence of an ITPR1-lined nucleoplasmic reticulum. Mice with Itpr1 heterozygous null mutations showed no major iris defects. Purkinje cells of the cerebellum appear to be the most sensitive to impaired ITPR1 function in humans. Iris hypoplasia is likely to result from either complete loss of ITPR1 activity or structure-specific disruption of multimeric interactions., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Gillespie syndrome (GS) is a rare variant form of aniridia characterized by non-progressive cerebellar ataxia, intellectual disability, and iris hypoplasia. Unlike the more common dominant and sporadic forms of aniridia, there has been no significant association with PAX6 mutations in individuals with GS and the mode of inheritance of the disease had long been regarded as uncertain. Using a combination of trio-based whole-exome sequencing and Sanger sequencing in five simplex GS-affected families, we found homozygous or compound heterozygous truncating mutations (c.4672C>T [p.Gln1558(∗)], c.2182C>T [p.Arg728(∗)], c.6366+3A>T [p.Gly2102Valfs5(∗)], and c.6664+5G>T [p.Ala2221Valfs23(∗)]) and de novo heterozygous mutations (c.7687&#95;7689del [p.Lys2563del] and c.7659T>G [p.Phe2553Leu]) in the inositol 1,4,5-trisphosphate receptor type 1 gene (ITPR1). ITPR1 encodes one of the three members of the IP3-receptors family that form Ca(2+) release channels localized predominantly in membranes of endoplasmic reticulum Ca(2+) stores. The truncation mutants, which encompass the IP3-binding domain and varying lengths of the modulatory domain, did not form functional channels when produced in a heterologous cell system. Furthermore, ITPR1 p.Lys2563del mutant did not form IP3-induced Ca(2+) channels but exerted a negative effect when co-produced with wild-type ITPR1 channel activity. In total, these results demonstrate biallelic and monoallelic ITPR1 mutations as the underlying genetic defects for Gillespie syndrome, further extending the spectrum of ITPR1-related diseases., (Copyright © 2016 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

Purpose: To evaluate the corneal button of primary penetrating keratoplasty of patients diagnosed with congenital aniridia. The study took place at the Instituto Universitario Barraquer and the Centro de Oftalmología Barraquer., Methods: A retrospective analysis of cases diagnosed with congenital aniridia was carried out. We analyzed 13 corneal buttons of 11 eyes with congenital aniridia. We only included those patients who underwent penetrating keratoplasty for the first time. The corneal buttons were analyzed for histological characteristics of the presence of vascularization, the presence or not of Bowman's layer, the thickness of the stroma and Descemet's membrane, and endothelium layer alterations., Results: We found alterations in the epithelium and stroma in all patients, although this loss of architecture was not seen in Descemet's membrane and the endothelial population., Conclusion: Patients with advanced congenital aniridic keratopathy may be good candidates for deep or superficial anterior lamellar keratoplasty for the preservation of normal endothelium and Descemet's membrane, along with limbal stem cell transplantation, to address epithelial and stromal pathology., (© 2016 S. Karger AG, Basel.)

Mutations in the Pax6 gene cause ocular defects in both vertebrate and invertebrate animal species, and the disease aniridia in humans. Despite extensive experimentation on this gene in multiple species, including humans, we still do not understand the earliest effects on development mediated by this gene. This prompted us to develop pax6 mutant lines in Xenopus tropicalis taking advantage of the utility of the Xenopus system for examining early development and in addition to establish a model for studying the human disease aniridia in an accessible lower vertebrate. We have generated mutants in pax6 by using Transcription Activator-Like Effector Nuclease (TALEN) constructs for gene editing in X. tropicalis. Embryos with putative null mutations show severe eye abnormalities and changes in brain development, as assessed by changes in morphology and gene expression. One gene that we found is downregulated very early in development in these pax6 mutants is myc, a gene involved in pluripotency and progenitor cell maintenance and likely a mediator of some key pax6 functions in the embryo. Changes in gene expression in the developing brain and pancreas reflect other important functions of pax6 during development. In mutations with partial loss of pax6 function eye development is initially relatively normal but froglets show an underdeveloped iris, similar to the classic phenotype (aniridia) seen in human patients with PAX6 mutations. Other eye abnormalities observed in these froglets, including cataracts and corneal defects, are also common in human aniridia. The frog model thus allows us to examine the earliest deficits in eye formation as a result of pax6 lesions, and provides a useful model for understanding the developmental basis for the aniridia phenotype seen in humans., (Copyright © 2015 Elsevier Inc. All rights reserved.)

A 33-year-old woman with congenital aniridia presented with decreased vision in her right eye. Slit lamp examination revealed diffuse conjunctivalisation of the ocular surface with mild subepithelial fibrosis consistent with aniridic keratopathy secondary to limbal stem cell deficiency. She underwent limbal stem cell transplantation with cadaver donor tissue (keratolimbal allograft (KLAL) surgery) and received systemic immunosuppression. Despite optimal combination immunosuppressive therapy managed by a renal transplant specialist, 2 weeks after the KLAL, the patient developed intractable eye pain, conjunctival injection, dilation of the KLAL graft blood vessels and limbal haemorrhages. There were no epithelial defects noted. Donor-specific antibody testing was positive, and intravenous immunoglobulin therapy was initiated. There was immediate symptomatic and objective improvement. Fifteen months postoperatively, the patient's vision was 20/400 with a stable corneal epithelium and no evidence of inflammation., (2015 BMJ Publishing Group Ltd.)

Purpose: Aniridia is a rare panocular disorder characterized by iris hypoplasia and other associated eye anomalies. Heterozygous null mutations in paired box gene 6 (PAX6) are the major cause of the classic aniridia phenotype. This study aims to detect the mutational spectrum of PAX6 and associated phenotypes in southern Indian patients with sporadic and familial aniridia., Methods: Genomic DNA was isolated from peripheral blood from all participants. The coding regions and flanking intronic sequences of PAX6 were screened with Sanger sequencing in 30 probands with aniridia. The identified variations were further evaluated in available family members and 150 healthy controls. The pathogenic potential of the mutations were assessed using bioinformatics tools., Results: Thirteen different mutations were detected in eight sporadic and five familial cases. Eleven novel mutations, including five insertions (c.7&#95;10dupAACA, c.567dupC, c.704dupC, c.868dupA and c.753&#95;754insTA), two deletions (c.242delC and c.249delT), and four splicing variants (c.10+1G>A, c.141G>A, c.141+4A>G and c.764A>G) were identified in this study. Clinical findings of the patients revealed phenotypic heterogeneity with the same or different mutations., Conclusions: This study reported 11 novel mutations and thus expanded the spectrum of PAX6 mutations. Interestingly, all mutations reported in this study were truncations, which confirms the hypothesis that haploinsufficiency of PAX6 causes the aniridia phenotype. Our observations revealed inter- and intrafamilial phenotypic variability with PAX6 mutations. The common ocular findings associated with PAX6 mutations were iris hypoplasia, nystagmus, and foveal hypoplasia reported in almost all cases, with cataract, glaucoma, and keratopathy reported in approximately 50% of the patients.

Purpose: Efficacy of cultivated oral mucosa epithelial transplantation (COMET) procedure in corneal epithelium restoration of aniridia patients., Methods: Study subjects were aniridia patients (13 patients; 17 eyes) with irregular, vascular conjunctival pannus involving visual axis who underwent autologous transplantation of cultivated epithelium. For the procedure oral mucosa epithelial cells were obtained from buccal mucosa with further enzymatic treatment. Suspension of single cells was seeded on previously prepared denuded amniotic membrane. Cultures were carried on culture dishes inserts in the presence of the inactivated with Mitomycin C monolayer of 3T3 fibroblasts. Cultures were carried for seven days. Stratified oral mucosa epithelium with its amniotic membrane carrier was transplanted on the surgically denuded corneal surface of aniridia patients with total or subtotal limbal stem cell deficiency. Outcome Measures. Corneal surface, epithelial regularity, and visual acuity improvement were evaluated., Results: At the end of the observation period, 76.4% of the eyes had regular transparent epithelium and 23.5% had developed epithelial defects or central corneal haze; in 88.2% of cases visual acuity had increased. VA range was from HM 0.05 before the surgery to HM up to 0.1 after surgery., Conclusion: Application of cultivated oral mucosa epithelium restores regular epithelium on the corneal surface with moderate improvement in quality of vision.

Congenital aniridia is a rare genetic disorder characterized by varying degrees of iris hypoplasia that are associated with additional ocular abnormalities. More than 90% of the causal mutations identified are found in the PAX6 gene, a transcription factor of critical importance in the process of neurogenesis and ocular development. Here, we investigate clinical, molecular, and craniofacial features of a large Brazilian family with congenital aniridia. Among the 56 eyes evaluated, phenotype variation encompassed bilateral total aniridia to mild iris defects with extensive variation between eyes of the same individual. PAX6 molecular screening indicated a heterozygous splice mutation (c.141 + 1G>A). Thus, we hypothesize that this splicing event may cause variation in the expression of the wild-type transcript, which may lead to the observed variation in phenotype. Affected individuals were more brachycephalic, even though their face height and cephalic circumference were not significantly different when compared to those of non-affected relatives. From this, we infer that the head shape of affected subjects may also be a result of the PAX6 splice-site mutation. Our data summarize the clinical variability associated with the ocular phenotype in a large family with aniridia, and help shed light on the role of PAX6 in neurocranial development., (© 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Purpose: To describe a surgical technique and intraoperative modifications to create a secondary capsulotomy using the femtosecond cataract laser in an eye with traumatic aphakia and aniridia., Methods: The expanded use of the femtosecond cataract laser (Catalys Precision Laser System; Optimedica, Sunnyvale, CA) for creation of a well-centered and circular secondary capsulotomy in the capsular remnants of an eye with traumatic aphakia and aniridia., Results: Using software modifications based on intraoperative anterior segment spectral-domain optical coherence tomography imaging, and customizing the capsulotomy settings, the femtosecond cataract laser was able to penetrate through most of the fused anterior and posterior capsular leaflets and a circular capsulotomy was achieved. The latter permitted optic capture and excellent centration of a three-piece intraocular lens placed in the sulcus with good postoperative visual rehabilitation., Conclusion: The femtosecond cataract laser can be customized to create a well-centered and circular secondary capsulotomy in aniridic and aphakic eyes where optic capture is critical for long-term intraocular lens stability., (Copyright 2014, SLACK Incorporated.)