Ocular Anterior Segment Dysgenesis Research Articles

TGFβ Signaling Dysregulation May Contribute to COL4A1-Related Glaucomatous Optic Nerve Damage.

Mutations in the genes encoding type IV collagen alpha 1 (COL4A1) and alpha 2 (COL4A2) cause a multisystem disorder that includes ocular anterior segment dysgenesis (ASD) and glaucoma. We previously showed that transforming growth factor beta (TGFβ) signaling was elevated in developing anterior segments from Col4a1 mutant mice and that reducing TGFβ signaling ameliorated ASD, supporting a role for the TGFβ pathway in disease pathogenesis. Here, we tested whether altered TGFβ signaling also contributes to glaucoma-related phenotypes in Col4a1 mutant mice. To test the role of TGFβ signaling in glaucoma-relevant phenotypes, we genetically reduced TGFβ signaling using mice with mutated Tgfbr2, which encodes the common receptor for all TGFβ ligands in Col4a1+/G1344D mice. We performed slit-lamp biomicroscopy and optical coherence tomography for qualitative and quantitative analyses of anterior and posterior ocular segments, histological analyses of ocular tissues and optic nerves, and intraocular pressure assessments using rebound tonometry. Col4a1+/G1344D mice showed defects of the ocular drainage structures, including iridocorneal adhesions, and phenotypes consistent with glaucomatous neurodegeneration, including thinning of the nerve fiber layer, retinal ganglion cell loss, optic nerve head excavation, and optic nerve degeneration. We found that reducing TGFβ receptor 2 (TGFBR2) was protective for ASD, ameliorated ocular drainage structure defects, and protected against glaucomatous neurodegeneration in Col4a1+/G1344D mice. Our results suggest that elevated TGFβ signaling contributes to glaucomatous neurodegeneration in Col4a1 mutant mice.

Evaluating neural crest cell migration in a Col4a1 mutant mouse model of ocular anterior segment dysgenesis

The periocular mesenchyme (POM) is a transient migratory embryonic tissue derived from neural crest cells (NCCs) and paraxial mesoderm that gives rise to most of the structures in front of the eye. Morphogenetic defects of these structures can impair aqueous humor outflow, leading to elevated intraocular pressure and glaucoma. Mutations in collagen type IV alpha 1 (COL4A1) and alpha 2 (COL4A2) cause Gould syndrome – a multisystem disorder often characterized by variable cerebrovascular, ocular, renal, and neuromuscular manifestations. Approximately one-third of individuals with COL4A1 and COL4A2 mutations have ocular anterior segment dysgenesis (ASD), including congenital glaucoma resulting from abnormalities of POM-derived structures. POM differentiation has been a major focus of ASD research, but the underlying cellular mechanisms are still unclear. Moreover, earlier events including NCC migration and survival defects have been implicated in ASD; however, their roles are not as well understood. Vascular defects are among the most common consequences of COL4A1 and COL4A2 mutations and can influence NCC survival and migration. We therefore hypothesized that NCC migration might be impaired by COL4A1 and COL4A2 mutations. In this study, we used 3D confocal microscopy, gross morphology, and quantitative analyses to test NCC migration in Col4a1 mutant mice. We show that homozygous Col4a1 mutant embryos have severe embryonic growth retardation and lethality, and we identified a potential maternal effect on embryo development. Cerebrovascular defects in heterozygous Col4a1 mutant embryos were present as early as E9.0, showing abnormal cerebral vasculature plexus remodeling compared to controls. We detected abnormal NCC migration within the diencephalic stream and the POM in heterozygous Col4a1 mutants whereby mutant NCCs formed smaller diencephalic migratory streams and POMs. In these settings, migratory NCCs within the diencephalic stream and POM localize farther away from the developing vasculature. Our results show for the first time that Col4a1 mutations lead to cranial NCCs migratory defects in the context of early onset defective angiogenesis without affecting cell numbers, possibly impacting the relation between NCCs and the blood vessels during ASD development.

The phenotypic spectrum of terminal and subterminal 6p deletions based on a social media-derived cohort and literature review

BackgroundTerminal 6p deletions are rare, and information on their clinical consequences is scarce, which impedes optimal management and follow-up by clinicians. The parent-driven Chromosome 6 Project collaborates with families of affected children worldwide to better understand the clinical effects of chromosome 6 aberrations and to support clinical guidance. A microarray report is required for participation, and detailed phenotype information is collected directly from parents through a multilingual web-based questionnaire. Information collected from parents is then combined with case data from literature reports. Here, we present our findings on 13 newly identified patients and 46 literature cases with genotypically well-characterised terminal and subterminal 6p deletions. We provide phenotype descriptions for both the whole group and for subgroups based on deletion size and HI gene content.ResultsThe total group shared a common phenotype characterised by ocular anterior segment dysgenesis, vision problems, brain malformations, congenital defects of the cardiac septa and valves, mild to moderate hearing impairment, eye movement abnormalities, hypotonia, mild developmental delay and dysmorphic features. These characteristics were observed in all subgroups where FOXC1 was included in the deletion, confirming a dominant role for this gene. Additional characteristics were seen in individuals with terminal deletions exceeding 4.02 Mb, namely complex heart defects, corpus callosum abnormalities, kidney abnormalities and orofacial clefting. Some of these additional features may be related to the loss of other genes in the terminal 6p region, such as RREB1 for the cardiac phenotypes and TUBB2A and TUBB2B for the cerebral phenotypes. In the newly identified patients, we observed previously unreported features including gastrointestinal problems, neurological abnormalities, balance problems and sleep disturbances.ConclusionsWe present an overview of the phenotypic characteristics observed in terminal and subterminal 6p deletions. This reveals a common phenotype that can be highly attributable to haploinsufficiency of FOXC1, with a possible additional effect of other genes in the 6p25 region. We also delineate the developmental abilities of affected individuals and report on previously unrecognised features, showing the added benefit of collecting information directly from parents. Based on our overview, we provide recommendations for clinical surveillance to support clinicians, patients and families.

Enlargement of the Eye Socket Early after Birth with an Ocular Prosthesis for Clinical Congenital Anophthalmia

Clinical congenital anophthalmia causes facial asymmetry and difficulty in wearing ocular prostheses due to underdevelopment of the orbits. Orbital expansion must be started early after birth for treatment, but consensus regarding the treatment for clinical congenital anophthalmia has not been established. We report a successful orbital expansion using a thermoplastic splint. A one-month-old boy had left clinical congenital anophthalmia (ophthalmatrophia), right congenital glaucoma, and right anterior segment ocular dysgenesis. Two months after birth, left orbital expansion was initiated using a custom-made thermoplastic splint. The splint was replaced with larger ones in steps as the infant grew, and the orbit was successfully expanded. There is no appreciable size difference between the left and right orbit after 5 years of follow up. Treatment using a splint for the conjunctival sac is a simple and effective procedure that can be performed on an outpatient basis. Safer and simpler material should be widely used for the treatment.

A rare genotype of biallelic mosaic variants in BCOR gene causing a bilateral ocular anterior segment dysgenesis and cataracts.

Oculofaciocardiodental (OFCD) syndrome is a rare X-linked dominant syndrome characterized by the involvement of the eyes, face, teeth, and heart with variable expressivity. The syndrome is caused by loss-of-function variants in the BCOR gene located on the X chromosome. OFCD affects only females with presumed embryonic lethality among males. We report a first case of a female with biallelic mosaic variants in BCOR gene, leading to a severe ocular phenotype including anterior segment dysgenesis, cataracts, and retinal involvement. The unique condition of biallelic mosaic loss-of-function mutations leads to a variable expression of an allele with the pathogenic variant, independent of the X-Inactivation pattern. This novel mechanism of co-existent biallelic mosaicism should be suspected in unexplained severe cases of OFCD.

Elevated TGFβ signaling contributes to ocular anterior segment dysgenesis in Col4a1 mutant mice

Ocular anterior segment dysgenesis (ASD) refers to a collection of developmental disorders affecting the anterior structures of the eye. Although a number of genes have been implicated in the etiology of ASD, the underlying pathogenetic mechanisms remain unclear. Mutations in genes encoding collagen type IV alpha 1 (COL4A1) and alpha 2 (COL4A2) cause Gould syndrome, a multi-system disorder that often includes ocular manifestations such as ASD and glaucoma. COL4A1 and COL4A2 are abundant basement membrane proteins that provide structural support to tissues and modulate signaling through interactions with other extracellular matrix proteins, growth factors, and cell surface receptors. In this study, we used a combination of histological, molecular, genetic and pharmacological approaches to demonstrate that altered TGFβ signaling contributes to ASD in mouse models of Gould syndrome. We show that TGFβ signaling was elevated in anterior segments from Col4a1 mutant mice and that genetically reducing TGFβ signaling partially prevented ASD. Notably, we identified distinct roles for TGFβ1 and TGFβ2 in ocular defects observed in Col4a1 mutant mice. Importantly, we show that pharmacologically promoting type IV collagen secretion or reducing TGFβ signaling ameliorated ocular pathology in Col4a1 mutant mice. Overall, our findings demonstrate that altered TGFβ signaling contributes to COL4A1-related ocular dysgenesis and implicate this pathway as a potential therapeutic target for the treatment of Gould syndrome.

Axenfeld-Rieger Syndrome: A Case Report with Literature Review

Axenfeld–Rieger syndrome (ARS) is a rare autosomal dominant disorder that has both systemic and ocular anterior segment dysgenesis. The ocular manifestations include posterior embryotoxon, iris and anterior angle abnomalies with a high risk of glaucoma and blindness. The systemic manifestations can include craniofacial abnomalies such as maxillary hypoplasia, hypodontia, oligodontia and microdont.

Identification of fibronectin 1 as a candidate genetic modifier in a Col4a1 mutant mouse model of Gould syndrome.

ABSTRACTCollagen type IV alpha 1 and alpha 2 (COL4A1 and COL4A2) are major components of almost all basement membranes. COL4A1 and COL4A2 mutations cause a multisystem disorder that can affect any organ but typically involves the cerebral vasculature, eyes, kidneys and skeletal muscles. In recent years, patient advocacy and family support groups have united under the name of Gould syndrome. The manifestations of Gould syndrome are highly variable, and animal studies suggest that allelic heterogeneity and genetic context contribute to the clinical variability. We previously characterized a mouse model of Gould syndrome caused by a Col4a1 mutation in which the severities of ocular anterior segment dysgenesis (ASD), myopathy and intracerebral hemorrhage (ICH) were dependent on genetic background. Here, we performed a genetic modifier screen to provide insight into the mechanisms contributing to Gould syndrome pathogenesis and identified a single locus [modifier of Gould syndrome 1 (MoGS1)] on Chromosome 1 that suppressed ASD. A separate screen showed that the same locus ameliorated myopathy. Interestingly, MoGS1 had no effect on ICH, suggesting that this phenotype could be mechanistically distinct. We refined the MoGS1 locus to a 4.3 Mb interval containing 18 protein-coding genes, including Fn1, which encodes the extracellular matrix component fibronectin 1. Molecular analysis showed that the MoGS1 locus increased Fn1 expression, raising the possibility that suppression is achieved through a compensatory extracellular mechanism. Furthermore, we found evidence of increased integrin-linked kinase levels and focal adhesion kinase phosphorylation in Col4a1 mutant mice that is partially restored by the MoGS1 locus, implicating the involvement of integrin signaling. Taken together, our results suggest that tissue-specific mechanistic heterogeneity contributes to the variable expressivity of Gould syndrome and that perturbations in integrin signaling may play a role in ocular and muscular manifestations.

First observation of secondary childhood glaucoma in Coffin-Siris syndrome: a case report and literature review

BackgroundSevere congenital ophthalmological malformations and glaucoma might be an important occasional feature in patients with Coffin-Siris syndrome (CSS), especially Coffin-Siris syndrome 9 (CSS9, OMIM #615866) caused by SOX11 mutation. Recently, primary (open-angle) glaucoma was described in two children with the most common form of Coffin-Siris syndrome, CSS1 (OMIM #135900) by ARID1B (AT-rich interaction domain-containing protein 1B) gene mutation. In this article, we present the first report of glaucoma with Coffin-Siris syndrome 9 as well as the first report of secondary glaucoma with any form of Coffin-Siris syndrome. These findings indicate that secondary glaucoma is an occasional finding in patients with Coffin-Siris syndrome.Case presentationA child with secondary childhood glaucoma and additional ocular manifestations was evaluated and treated at the childhood glaucoma centre in Mainz, Germany. Examination under general anaesthesia revealed ocular anterior segment dysgenesis (ASD) (Peters type iridocorneal dysgenesis) in combination with congenital limbal stem cell deficiency (LSCD), aniridia, and cataract. The patient also had multiple other congenital anomalies and severe developmental delay. To explain his combination of anomalies, molecular genetic analysis from peripheral blood was performed in late 2018 and early 2019. Following normal findings with a panel diagnostic of 18 genes associated with congenital glaucoma, whole exome sequencing was performed and revealed a novel likely pathogenic heterozygous variant c.251G>T, p.(Gly84Val) in the SOX11 gene (SRY-related HMG-box gene 11). The variant had occurred de novo. Thus, the multiple congenital anomalies and developmental delay of the patient represented Coffin-Siris syndrome 9 (CSS9, OMIM #615866).ConclusionsWhen eye diseases occur in combination with other systemic features, genetic analysis can be seminal. Results indicate that glaucoma is an occasional feature of patients with Coffin-Siris syndrome. As early treatment may improve the visual outcome of patients with glaucoma, we suggest that patients with Coffin-Siris syndrome should receive specific ophthalmological screening.

A donor twin discordant with Peters anomaly in a twin\u2013twin transfusion syndrome case: a case report

BackgroundPeters anomaly is a rare form of anterior segment ocular dysgenesis, the antenatal image of Peters anomaly had not been reported. We herein showcased a discordant finding of Peters anomaly in a monozygotic twin complicated with twin-twin transfusion syndrome (TTTS) and exhibited its antenatal sonographic images,Case presentationA 38-year-old gravida 2 para 1 pregnant woman visited our clinic at the gestational age of 18 weeks where TTTS stage III was diagnosed and the following laser therapy was done successfully. Ten days after the surgery, the follow-up ultrasound detected the opacity of both fetal eyeballs in the donor twin and thus congenital cataract was suspected initially. Then magnetic resonance imaging (MRI) examination was arranged at the gestational age of 23 weeks, and no central nervous system or other anomaly was found. At the 29 weeks of gestation, the opacity of both fetal eyeballs of the donor twin did not clear. The pregnancy resulted in cesarean section at the gestational age of 37 weeks indicated by malpresentation where two male live births were born. Examination under anesthesia was arranged for donor twin after delivery and Peters anomaly was diagnosed based on central corneal opacity with iridocorneal and corneolenticular adhesions.ConclusionsThe prenatal image of Peters anomaly may present as the opacity of the fetal eyeballs similar to congenital cataract. Some cases of the Peters anomaly had been reported with a genetic abnormality, but since our case presented discordant presentation in monozygotic twin pregnancy where both twins are supposed to share the same genetic make-up, therefore other factors that are epigenetic may be held accountable. Nevertheless, a genetic origin of the anomaly in our case cannot be excluded.

Axenfeld-Rieger Syndrome–A Rare Phenomenon

Purpose: Axenfeld-Rieger syndrome is an ocular anterior segment dysgenesis, autosomal dominantly inherited, commonly associated with glaucoma and systemic anomalies. This study presents various clinical manifestations of Axenfeld-Rieger syndrome within one family. Material and Method: Three members of the family: patient 1--father (54 years old), patient 2--son (12 years old) and patient 3--daughter (8 years old), underwent complete ophthalmic examination, including standard glaucoma diagnostics. It was impossible to perform complete eye examination in one case (patient 1) because of severity of ocular changes. Results: All family members described had iris abnormalities (hypoplastic iris stroma) and advanced glaucoma, however severity of symptoms were different in each case. The most advanced disease was recognized in patient 1. Other findings included: posterior embryotoxon (patients 2 and 3), iridocorneal angle abnormalities (patients 2 and 3), stretch holes and extraocular features (patient 2): dental anomalies (microdontia and hypodontia), maxillary hypoplasia and periumbilical skin fold. All of these symptoms supported the diagnosis of Axenfeld-Rieger syndrome. In addition, we also diagnosed high myopia & strabismus (patient 2) and corneal scar in (patient 1). Conclusion: Reported cases of Axenfeld-Rieger syndrome demonstrate phenotypic variability of the disease among family members, which is characteristic for this disorder and can cause diagnostic problem.

Biallelic CPAMD8 Variants Are a Frequent Cause of Childhood and Juvenile Open-Angle Glaucoma

Developmental abnormalities of the ocular anterior segment in some cases can lead to ocular hypertension and glaucoma. CPAMD8 is a gene of unknown function recently associated with ocular anterior segment dysgenesis, myopia, and ectopia lentis. We sought to assess the contribution of biallelic CPAMD8 variants to childhood and juvenile open-angle glaucoma. Retrospective, multicenter case series. A total of 268 probands and their relatives with a diagnosis of childhood or juvenile open-angle glaucoma. Developmental abnormalities of the ocular anterior segment in some cases can lead to ocular hypertension and glaucoma. CPAMD8 is a gene of unknown function recently associated with ocular anterior segment dysgenesis, myopia, and ectopia lentis. We sought to assess the contribution of biallelic CPAMD8 variants to childhood and juvenile open-angle glaucoma. Patients underwent a comprehensive ophthalmic assessment, with DNA from patients and their relatives subjected to genome, exome, or capillary sequencing. CPAMD8 RNA expression analysis was performed on tissues dissected from cadaveric human eyes. Diagnostic yield within a cohort of childhood and juvenile open-angle glaucoma, prevalence and risk of ophthalmic phenotypes, and relative expression of CPAMD8 in the human eye. We identified rare (allele frequency < 4×10-5) biallelic CPAMD8 variants in 5.7% (5/88) of probands with childhood glaucoma and 2.1% (2/96) of probands with juvenile open-angle glaucoma. When including family members, we identified 11 individuals with biallelic variants in CPAMD8 from 7 unrelated families. Nine of these individuals were diagnosed with glaucoma (9/11, 81.8%), with a mean age at diagnosis of 9.22±14.89 years, and all individuals with glaucoma required 1 or more incisional procedures to control high intraocular pressure. Iris abnormalities were observed in 9 of 11 individuals, cataract was observed in 8 of 11 individuals (72.7%), and retinal detachment was observed in 3 of 11 individuals (27.3%). CPAMD8 expression was highest in neural crest-derived tissues of the adult anterior segment, suggesting that CPAMD8 variation may cause malformation or obstruction of key drainage structures. Biallelic CPAMD8 variation was associated with a highly heterogeneous phenotype and in our cohorts was the second most common inherited cause of childhood glaucoma after CYP1B1 and juvenile open-angle glaucoma after MYOC. CPAMD8 sequencing should be considered in the investigation of both childhood and juvenile open-angle glaucoma, particularly when associated with iris abnormalities, cataract, or retinal detachment.

Lens-specific conditional knockout of Msx2 in mice leads to ocular anterior segment dysgenesis via activation of a calcium signaling pathway

Ocular anterior segment dysgenesis (ASD) is a failure of normal development of anterior structures of the eye, leading to lens opacification. The underlying mechanisms relating to ASD are still unclear. Previous studies have implicated transcriptional factor muscle segment homeobox 2 (Msx2) in ASD. In this study, we used Msx2 conditional knockout (CKO) mice as a model and found that Msx2 deficiency in surface ectoderm induced ASD. Loss of Msx2 function specifically affected lens development, while other eye structures were not significantly affected. Multiple lines of evidence show that calcium signaling pathways are involved in this pathogenesis. Our study demonstrates that Msx2 plays an essential role in lens development by activating a yet undetermined calcium signaling pathway.

Ciliary margin-derived BMP4 does not have a major role in ocular development.

Heterozygous Bmp4 mutations in humans and mice cause severe ocular anterior segment dysgenesis (ASD). Abnormalities include pupil displacement, corneal opacity, iridocorneal adhesions, and variable intraocular pressure, as well as some retinal and vascular defects. It is presently not known what source of BMP4 is responsible for these defects, as BMP4 is expressed in several developing ocular and surrounding tissues. In particular, BMP4 is expressed in the ciliary margins of the optic cup which give rise to anterior segment structures such as the ciliary body and iris, making it a good candidate for the required source of BMP4 for anterior segment development. Here, we test whether ciliary margin-derived BMP4 is required for ocular development using two different conditional knockout approaches. In addition, we compared the conditional deletion phenotypes with Bmp4 heterozygous null mice. Morphological, molecular, and functional assays were performed on adult mutant mice, including histology, immunohistochemistry, in vivo imaging, and intraocular pressure measurements. Surprisingly, in contrast to Bmp4 heterozygous mutants, our analyses revealed that the anterior and posterior segments of Bmp4 conditional knockouts developed normally. These results indicate that ciliary margin-derived BMP4 does not have a major role in ocular development, although subtle alterations could not be ruled out. Furthermore, we demonstrated that the anterior and posterior phenotypes observed in Bmp4 heterozygous animals showed a strong propensity to co-occur, suggesting a common, non-cell autonomous source for these defects.

Mutations of conserved non-coding elements of PITX2 in patients with ocular dysgenesis and developmental glaucoma.

Mutations in FOXC1 and PITX2 constitute the most common causes of ocular anterior segment dysgenesis (ASD), and confer a high risk for secondary glaucoma. The genetic causes underlying ASD in approximately half of patients remain unknown, despite many of them being screened by whole exome sequencing. Here, we performed whole genome sequencing on DNA from two affected individuals from a family with dominantly inherited ASD and glaucoma to identify a 748-kb deletion in a gene desert that contains conserved putative PITX2 regulatory elements. We used CRISPR/Cas9 to delete the orthologous region in zebrafish in order to test the pathogenicity of this structural variant. Deletion in zebrafish reduced pitx2 expression during development and resulted in shallow anterior chambers. We screened additional patients for copy number variation of the putative regulatory elements and found an overlapping deletion in a second family and in a potentially-ancestrally-related index patient with ASD and glaucoma. These data suggest that mutations affecting conserved non-coding elements of PITX2 may constitute an important class of mutations in patients with ASD for whom the molecular cause of their disease have not yet been identified. Improved functional annotation of the human genome and transition to sequencing of patient genomes instead of exomes will be required before the magnitude of this class of mutations is fully understood.

Genetic dissection of anterior segment dysgenesis caused by a Col4a1 mutation in mouse.

ABSTRACTOcular anterior segment dysgenesis (ASD) describes a spectrum of clinically and genetically heterogeneous congenital disorders affecting anterior structures that often lead to impaired vision. More importantly, 50-75% of patients with ASD develop early onset and aggressive glaucoma. Although several genes have been implicated in the etiology of ASD, the underlying mechanisms remain elusive. Type IV collagen alpha 1 (COL4A1) is an extracellular matrix protein and a critical component of nearly all basement membranes. COL4A1 mutations cause multi-system disorders in patients, including ASD (congenital cataracts, Axenfeld-Rieger's anomaly, Peter's anomaly and microphthalmia) and congenital or juvenile glaucoma. Here, we use a conditional Col4a1 mutation in mice to determine the location and timing of pathogenic events underlying COL4A1-related ocular dysgenesis. Our results suggest that selective expression of the Col4a1 mutation in neural crest cells and their derivatives is not sufficient to cause ocular dysgenesis and that selective expression of the Col4a1 mutation in vascular endothelial cells can lead to mild ASD and optic nerve hypoplasia but only on a sensitized background. In contrast, lens-specific expression of the conditional Col4a1 mutant allele led to cataracts, mild ASD and optic nerve hypoplasia, and age-related intraocular pressure dysregulation and optic nerve damage. Finally, ubiquitous expression of the conditional Col4a1 mutation at distinct developmental stages suggests that pathogenesis takes place before E12.5. Our results show that the lens and possibly vasculature play important roles in Col4a1-related ASD and that the pathogenic events occur at mid-embryogenesis in mice, during early stages of ocular development.

FOXE3 contributes to Peters anomaly through transcriptional regulation of an autophagy-associated protein termed DNAJB1.

FOXE3 is a lens-specific transcription factor that has been associated with anterior segment ocular dysgenesis. To determine the transcriptional target(s) of FOXE3 that are indispensable for the anterior segment development, we examined the transcriptome and the proteome of cells expressing truncated FOXE3 responsible for Peters anomaly identified through linkage-coupled next-generation whole-exome sequencing. We found that DNAJB1, an autophagy-associated protein, was the only candidate exhibiting differential expression in both screens. We confirmed the candidacy of DNAJB1 through chromatin immunoprecipitation and luciferase assays while knockdown of DNAJB1 in human lens epithelial cells resulted in a mitotic arrest. Subsequently, we targeted dnajb1a in zebrafish through injection of a splice-blocking morpholino. The dnajb1a morphants exhibited underdeveloped cataractous lenses with persistent apoptotic nuclei. In conclusion, here we report DNAJB1 is a transcriptional target of FOXE3 in a novel pathway that is crucial for the development of the anterior segment of the eye.

Targeted deletion of the murine Lgr4 gene decreases lens epithelial cell resistance to oxidative stress and induces age-related cataract formation.

Oxidative stress contributes to the formation of cataracts. The leucine rich repeat containing G protein-coupled receptor 4 (LGR4, also known as GPR48), is important in many developmental processes. Since deletion of Lgr4 has previously been shown to lead to cataract formation in mice, we sought to determine the specific role that Lgr4 plays in the formation of cataracts. Initially, the lens opacities of Lgr4−/− mice at different ages without ocular anterior segment dysgenesis (ASD) were evaluated with slit-lamp biomicroscopy. Lenses from both Lgr4−/− and wild-type mice were subjected to oxidation induced protein denaturation to assess the ability of the lens to withstand oxidation. The expression of antioxidant enzymes was evaluated with real-time quantitative PCR. Phenotypically, Lgr4−/− mice showed earlier onset of lens opacification and higher incidence of cataract formation compared with wild-type mice of similar age. In addition, Lgr4−/− mice demonstrated increased sensitivity to environmental oxidative damage, as evidenced by altered protein expression. Real-time quantitative PCR showed that two prominent antioxidant defense enzymes, catalase (CAT) and superoxidase dismutase-1 (SOD1), were significantly decreased in the lens epithelial cells of Lgr4−/− mice. Our results suggest that the deletion of Lgr4 can lead to premature cataract formation, as well as progressive deterioration with aging. Oxidative stress and altered expression of several antioxidant defense enzymes contribute to the formation of cataracts.

Whole exome sequence analysis of Peters anomaly.

Peters anomaly is a rare form of anterior segment ocular dysgenesis, which can also be associated with additional systemic defects. At this time, the majority of cases of Peters anomaly lack a genetic diagnosis. We performed whole exome sequencing of 27 patients with syndromic or isolated Peters anomaly to search for pathogenic mutations in currently known ocular genes. Among the eight previously recognized Peters anomaly genes, we identified a de novo missense mutation in PAX6, c.155G>A, p.(Cys52Tyr), in one patient. Analysis of 691 additional genes currently associated with a different ocular phenotype identified a heterozygous splicing mutation c.1025+2T>A in TFAP2A, a de novo heterozygous nonsense mutation c.715C>T, p.(Gln239*) in HCCS, a hemizygous mutation c.385G>A, p.(Glu129Lys) in NDP, a hemizygous mutation c.3446C>T, p.(Pro1149Leu) in FLNA, and compound heterozygous mutations c.1422T>A, p.(Tyr474*) and c.2544G>A, p.(Met848Ile) in SLC4A11; all mutations, except for the FLNA and SLC4A11 c.2544G>A alleles, are novel. This is the first study to use whole exome sequencing to discern the genetic etiology of a large cohort of patients with syndromic or isolated Peters anomaly. We report five new genes associated with this condition and suggest screening of TFAP2A and FLNA in patients with Peters anomaly and relevant syndromic features and HCCS, NDP and SLC4A11 in patients with isolated Peters anomaly.

First functional analysis of a novel splicing mutation in the B3GALTL gene by an ex vivo approach in Tunisian patients with typical Peters plus syndrome

Peters plus syndrome is a rare recessive autosomal disorder comprising ocular anterior segment dysgenesis, short stature, hand abnormalities and distinctive facial features. It was related only to mutations in the B3GALTL gene in the 13q12.3 region. In this study, we undertook the first functional analysis of a novel c.597-2 A>G splicing mutation within the B3GALTL gene using an ex-vivo approach. The results showed a complete skipping of exon 8 in the B3GALTL cDNA, which altered the open reading frame of the mutant transcript and generated a PTC within exon 9. This finding potentially elicits the nonsense mRNA to degradation by NMD (nonsense-mediated mRNA decay). The theoretical consequences of splice site mutations, predicted with the bioinformatics tool Human Splice Finder, were investigated and evaluated in relation to ex-vivo results. The findings confirmed the key role played by the B3GALTL gene in typical Peters-plus syndromes and the utility of mRNA analysis to understand the primary impacts of this mutation and the phenotype of the disease.