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1.
BMC Genomics ; 25(1): 484, 2024 May 16.
Artigo em Inglês | MEDLINE | ID: mdl-38755526

RESUMO

Childhood glaucoma (CG) encompasses a heterogeneous group of genetic eye disorders that is responsible for approximately 5% of childhood blindness worldwide. Understanding the molecular aetiology is key to improving diagnosis, prognosis and unlocking the potential for optimising clinical management. In this study, we investigated 86 CG cases from 78 unrelated families of diverse ethnic backgrounds, recruited into the Genomics England 100,000 Genomes Project (GE100KGP) rare disease cohort, to improve the genetic diagnostic yield. Using the Genomics England/Genomic Medicine Centres (GE/GMC) diagnostic pipeline, 13 unrelated families were solved (13/78, 17%). Further interrogation using an expanded gene panel yielded a molecular diagnosis in 7 more unrelated families (7/78, 9%). This analysis effectively raises the total number of solved CG families in the GE100KGP to 26% (20/78 families). Twenty-five percent (5/20) of the solved families had primary congenital glaucoma (PCG), while 75% (15/20) had secondary CG; 53% of this group had non-acquired ocular anomalies (including iris hypoplasia, megalocornea, ectopia pupillae, retinal dystrophy, and refractive errors) and 47% had non-acquired systemic diseases such as cardiac abnormalities, hearing impairment, and developmental delay. CYP1B1 was the most frequently implicated gene, accounting for 55% (11/20) of the solved families. We identified two novel likely pathogenic variants in the TEK gene, in addition to one novel pathogenic copy number variant (CNV) in FOXC1. Variants that passed undetected in the GE100KGP diagnostic pipeline were likely due to limitations of the tiering process, the use of smaller gene panels during analysis, and the prioritisation of coding SNVs and indels over larger structural variants, CNVs, and non-coding variants.


Assuntos
Glaucoma , Humanos , Glaucoma/genética , Glaucoma/diagnóstico , Masculino , Feminino , Criança , Pré-Escolar , Citocromo P-450 CYP1B1/genética , Mutação , Lactente , Genômica/métodos , Linhagem , Adolescente , Fatores de Transcrição Forkhead
2.
Stem Cell Reports ; 19(6): 839-858, 2024 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-38821055

RESUMO

Genetic perturbations influencing early eye development can result in microphthalmia, anophthalmia, and coloboma (MAC). Over 100 genes are associated with MAC, but little is known about common disease mechanisms. In this study, we generated induced pluripotent stem cell (iPSC)-derived optic vesicles (OVs) from two unrelated microphthalmia patients and healthy controls. At day 20, 35, and 50, microphthalmia patient OV diameters were significantly smaller, recapitulating the "small eye" phenotype. RNA sequencing (RNA-seq) analysis revealed upregulation of apoptosis-initiating and extracellular matrix (ECM) genes at day 20 and 35. Western blot and immunohistochemistry revealed increased expression of lumican, nidogen, and collagen type IV, suggesting ECM overproduction. Increased apoptosis was observed in microphthalmia OVs with reduced phospho-histone 3 (pH3+) cells confirming decreased cell proliferation at day 35. Pharmacological inhibition of caspase-8 activity with Z-IETD-FMK decreased apoptosis in one patient model, highlighting a potential therapeutic approach. These data reveal shared pathophysiological mechanisms contributing to a microphthalmia phenotype.


Assuntos
Apoptose , Células-Tronco Pluripotentes Induzidas , Microftalmia , Microftalmia/genética , Microftalmia/patologia , Microftalmia/metabolismo , Humanos , Apoptose/genética , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/citologia , Proliferação de Células , Caspase 8/metabolismo , Caspase 8/genética , Matriz Extracelular/metabolismo , Olho/metabolismo , Olho/patologia , Fenótipo
3.
Am J Biol Anthropol ; 183(2): e24866, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-37929663

RESUMO

OBJECTIVES: Analyses of external bone shape using geometric morphometrics (GM) and cross-sectional geometry (CSG) are frequently employed to investigate bone structural variation and reconstruct activity in the past. However, the association between these methods has not been thoroughly investigated. Here, we analyze whole bone shape and CSG variation of metacarpals 1-5 and test covariation between them. MATERIALS AND METHODS: We analyzed external metacarpal shape using GM and CSG of the diaphysis at three locations in metacarpals 1-5. The study sample includes three modern human groups: crew from the shipwrecked Mary Rose (n = 35 metacarpals), a Pre-industrial group (n = 50), and a Post-industrial group (n = 31). We tested group differences in metacarpal shape and CSG, as well as correlations between these two aspects of metacarpal bone structure. RESULTS: GM analysis demonstrated metacarpus external shape variation is predominately related to changes in diaphyseal width and articular surface size. Differences in external shape were found between the non-pollical metacarpals of the Mary Rose and Pre-industrial groups and between the third metacarpals of the Pre- and Post-industrial groups. CSG results suggest the Mary Rose and Post-industrial groups have stronger metacarpals than the Pre-industrial group. Correlating CSG and external shape showed significant relationships between increasing external robusticity and biomechanical strength across non-pollical metacarpals (r: 0.815-0.535; p ≤ 0.05). DISCUSSION: Differences in metacarpal cortical structure and external shape between human groups suggest differences in the type and frequency of manual activities. Combining these results with studies of entheses and kinematics of the hand will improve reconstructions of manual behavior in the past.


Assuntos
Ossos Metacarpais , Humanos , Metacarpo , Mãos , Diáfises , Extremidade Superior
4.
PLoS Biol ; 21(10): e3002336, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37856539

RESUMO

The transparent corneal epithelium in the eye is maintained through the homeostasis regulated by limbal stem cells (LSCs), while the nontransparent epidermis relies on epidermal keratinocytes for renewal. Despite their cellular similarities, the precise cell fates of these two types of epithelial stem cells, which give rise to functionally distinct epithelia, remain unknown. We performed a multi-omics analysis of human LSCs from the cornea and keratinocytes from the epidermis and characterized their molecular signatures, highlighting their similarities and differences. Through gene regulatory network analyses, we identified shared and cell type-specific transcription factors (TFs) that define specific cell fates and established their regulatory hierarchy. Single-cell RNA-seq (scRNA-seq) analyses of the cornea and the epidermis confirmed these shared and cell type-specific TFs. Notably, the shared and LSC-specific TFs can cooperatively target genes associated with corneal opacity. Importantly, we discovered that FOSL2, a direct PAX6 target gene, is a novel candidate associated with corneal opacity, and it regulates genes implicated in corneal diseases. By characterizing molecular signatures, our study unveils the regulatory circuitry governing the LSC fate and its association with corneal opacity.


Assuntos
Opacidade da Córnea , Epitélio Corneano , Limbo da Córnea , Humanos , Limbo da Córnea/metabolismo , Córnea/metabolismo , Epitélio Corneano/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Diferenciação Celular/genética , Opacidade da Córnea/metabolismo
5.
Biomolecules ; 13(2)2023 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-36830662

RESUMO

Ocular coloboma (OC) is a failure of complete optic fissure closure during embryonic development and presents as a tissue defect along the proximal-distal axis of the ventral eye. It is classed as part of the clinical spectrum of structural eye malformations with microphthalmia and anophthalmia, collectively abbreviated to MAC. Despite deliberate attempts to identify causative variants in MAC, many patients remain without a genetic diagnosis. To reveal potential candidate genes, we utilised transcriptomes experimentally generated from embryonic eye tissues derived from humans, mice, zebrafish, and chicken at stages coincident with optic fissure closure. Our in-silico analyses found 10 genes with optic fissure-specific enriched expression: ALDH1A3, BMPR1B, EMX2, EPHB3, NID1, NTN1, PAX2, SMOC1, TENM3, and VAX1. In situ hybridization revealed that all 10 genes were broadly expressed ventrally in the developing eye but that only PAX2 and NTN1 were expressed in cells at the edges of the optic fissure margin. Of these conserved optic fissure genes, EMX2, NID1, and EPHB3 have not previously been associated with human MAC cases. Targeted genetic manipulation in zebrafish embryos using CRISPR/Cas9 caused the developmental MAC phenotype for emx2 and ephb3. We analysed available whole genome sequencing datasets from MAC patients and identified a range of variants with plausible causality. In combination, our data suggest that expression of genes involved in ventral eye development is conserved across a range of vertebrate species and that EMX2, NID1, and EPHB3 are candidate loci that warrant further functional analysis in the context of MAC and should be considered for sequencing in cohorts of patients with structural eye malformations.


Assuntos
Coloboma , Anormalidades do Olho , Neuropeptídeos , Feminino , Gravidez , Humanos , Animais , Camundongos , Coloboma/genética , Coloboma/metabolismo , Olho/metabolismo , Peixe-Zebra/genética , Perfilação da Expressão Gênica , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/genética , Neuropeptídeos/metabolismo , Proteínas de Homeodomínio/metabolismo
6.
J Pathol ; 259(4): 441-454, 2023 04.
Artigo em Inglês | MEDLINE | ID: mdl-36656098

RESUMO

The crumbs cell polarity complex plays a crucial role in apical-basal epithelial polarity, cellular adhesion, and morphogenesis. Homozygous variants in human CRB1 result in autosomal recessive Leber congenital amaurosis (LCA) and retinitis pigmentosa (RP), with no established genotype-phenotype correlation. The associated protein complexes have key functions in developmental pathways; however, the underlying disease mechanism remains unclear. Using the oko meduzym289/m289 (crb2a-/- ) zebrafish, we performed integrative transcriptomic (RNA-seq data) and methylomic [reduced representation bisulphite sequencing (RRBS)] analysis of whole retina to identify dysregulated genes and pathways. Delayed retinal cell specification was identified in both the crb2a-/- zebrafish and CRB1 patient-derived retinal organoids, highlighting the dysfunction of cell cycle modulation and epigenetic transcriptional control. Differential DNA methylation analysis revealed novel hypermethylated pathways involving biological adhesion, Hippo, and transforming growth factor ß (TGFß) signalling. By integrating gene expression with DNA methylation using functional epigenetic modules (FEM), we identified six key modules involving cell cycle control and disturbance of TGFß, bone morphogenetic protein (BMP), Hippo, and SMAD protein signal transduction pathways, revealing significant interactome hotspots relevant to crb2a function and confirming the epigenetic control of gene regulation in early retinal development, which points to a novel mechanism underlying CRB1-retinopathies. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.


Assuntos
Polaridade Celular , Peixe-Zebra , Animais , Humanos , Peixe-Zebra/genética , Polaridade Celular/genética , Retina/metabolismo , Ciclo Celular , Epigênese Genética , Proteínas do Olho/genética , Proteínas do Olho/metabolismo , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
7.
Genet Med ; 24(5): 1073-1084, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35034853

RESUMO

PURPOSE: Ocular coloboma arises from genetic or environmental perturbations that inhibit optic fissure (OF) fusion during early eye development. Despite high genetic heterogeneity, 70% to 85% of patients remain molecularly undiagnosed. In this study, we have identified new potential causative genes using cross-species comparative meta-analysis. METHODS: Evolutionarily conserved differentially expressed genes were identified through in silico analysis, with in situ hybridization, gene knockdown, and rescue performed to confirm spatiotemporal gene expression and phenotype. Interrogation of the 100,000 Genomes Project for putative pathogenic variants was performed. RESULTS: Nine conserved differentially expressed genes between zebrafish and mouse were identified. Expression of zebrafish ank3a, bmpr1ba/b, cdh4, and pdgfaa was localized to the OF, periocular mesenchyme cells, or ciliary marginal zone, regions traversed by the OF. Knockdown of ank3, bmpr1b, and pdgfaa revealed a coloboma and/or microphthalmia phenotype. Novel pathogenic variants in ANK3, BMPR1B, PDGFRA, and CDH4 were identified in 8 unrelated coloboma families. We showed BMPR1B rescued the knockdown phenotype but variant messenger RNAs failed, providing evidence of pathogenicity. CONCLUSION: We show the utility of cross-species meta-analysis to identify several novel coloboma disease-causing genes. There is a potential to increase the diagnostic yield for new and unsolved patients while adding to our understanding of the genetic basis of OF morphogenesis.


Assuntos
Coloboma , Microftalmia , Animais , Anquirinas/genética , Anquirinas/metabolismo , Receptores de Proteínas Morfogenéticas Ósseas Tipo I/genética , Receptores de Proteínas Morfogenéticas Ósseas Tipo I/metabolismo , Coloboma/genética , Testes Genéticos , Humanos , Camundongos , Microftalmia/genética , Fenótipo , Peixe-Zebra/genética
8.
Commun Biol ; 4(1): 802, 2021 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-34183774

RESUMO

Regeneration of the testis from pluripotent stem cells is a real challenge, reflecting the complexity of the interaction of germ cells and somatic cells. Here we report the generation of testicular somatic cell-like cells (TesLCs) including Sertoli cell-like cells (SCLCs) from mouse embryonic stem cells (ESCs) in xeno-free culture. We find that Nr5a1/SF1 is critical for interaction between SCLCs and PGCLCs. Intriguingly, co-culture of TesLCs with epiblast-like cells (EpiLCs), rather than PGCLCs, results in self-organised aggregates, or testicular organoids. In the organoid, EpiLCs differentiate into PGCLCs or gonocyte-like cells that are enclosed within a seminiferous tubule-like structure composed of SCLCs. Furthermore, conditioned medium prepared from TesLCs has a robust inducible activity to differentiate EpiLCs into PGCLCs. Our results demonstrate conditions for in vitro reconstitution of a testicular environment from ESCs and provide further insights into the generation of sperm entirely in xeno-free culture.


Assuntos
Células-Tronco Embrionárias/citologia , Camadas Germinativas/citologia , Espermatozoides/citologia , Testículo/citologia , Animais , Diferenciação Celular , Técnicas de Cocultura , Masculino , Camundongos , Organoides/citologia , Transcriptoma
9.
JCI Insight ; 6(9)2021 05 10.
Artigo em Inglês | MEDLINE | ID: mdl-33755601

RESUMO

Choroideremia (CHM) is an X-linked recessive chorioretinal dystrophy caused by mutations in CHM, encoding for Rab escort protein 1 (REP1). Loss of functional REP1 leads to the accumulation of unprenylated Rab proteins and defective intracellular protein trafficking, the putative cause for photoreceptor, retinal pigment epithelium (RPE), and choroidal degeneration. CHM is ubiquitously expressed, but adequate prenylation is considered to be achieved, outside the retina, through the isoform REP2. Recently, the possibility of systemic features in CHM has been debated; therefore, in this study, whole metabolomic analysis of plasma samples from 25 CHM patients versus age- and sex-matched controls was performed. Results showed plasma alterations in oxidative stress-related metabolites, coupled with alterations in tryptophan metabolism, leading to significantly raised serotonin levels. Lipid metabolism was disrupted with decreased branched fatty acids and acylcarnitines, suggestive of dysfunctional lipid oxidation, as well as imbalances of several sphingolipids and glycerophospholipids. Targeted lipidomics of the chmru848 zebrafish provided further evidence for dysfunction, with the use of fenofibrate over simvastatin circumventing the prenylation pathway to improve the lipid profile and increase survival. This study provides strong evidence for systemic manifestations of CHM and proposes potentially novel pathomechanisms and targets for therapeutic consideration.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Coroideremia/metabolismo , Metabolismo dos Lipídeos/genética , Estresse Oxidativo/genética , Proteínas de Peixe-Zebra/genética , Adulto , Animais , Estudos de Casos e Controles , Coroideremia/genética , Fenofibrato/farmacologia , Glicerofosfolipídeos/metabolismo , Humanos , Hipolipemiantes/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Lipidômica , Masculino , Metabolômica , Pessoa de Meia-Idade , Prenilação , Serotonina/metabolismo , Sinvastatina/farmacologia , Esfingolipídeos/metabolismo , Triptofano/metabolismo , Adulto Jovem , Peixe-Zebra
10.
Int J Mol Sci ; 22(4)2021 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-33671840

RESUMO

EPHA2 is a transmembrane tyrosine kinase receptor that, when disrupted, causes congenital and age-related cataracts. Cat-Map reports 22 pathogenic EPHA2 variants associated with congenital cataracts, variable microcornea, and lenticonus, but no previous association with microphthalmia (small, underdeveloped eye, ≥2 standard deviations below normal axial length). Microphthalmia arises from ocular maldevelopment with >90 monogenic causes, and can include a complex ocular phenotype. In this paper, we report two pathogenic EPHA2 variants in unrelated families presenting with bilateral microphthalmia and congenital cataracts. Whole genome sequencing through the 100,000 Genomes Project and cataract-related targeted gene panel testing identified autosomal dominant heterozygous mutations segregating with the disease: (i) missense c.1751C>T, p.(Pro584Leu) and (ii) splice site c.2826-9G>A. To functionally validate pathogenicity, morpholino knockdown of epha2a/epha2b in zebrafish resulted in significantly reduced eye size ± cataract formation. Misexpression of N-cadherin and retained fibre cell nuclei were observed in the developing lens of the epha2b knockdown morphant fish by 3 days post-fertilisation, which indicated a putative mechanism for microphthalmia pathogenesis through disruption of cadherin-mediated adherens junctions, preventing lens maturation and the critical signals stimulating eye growth. This study demonstrates a novel association of EPHA2 with microphthalmia, suggesting further analysis of pathogenic variants in unsolved microphthalmia cohorts may increase molecular diagnostic rates.


Assuntos
Catarata/genética , Efrina-A2/genética , Microftalmia/genética , Adolescente , Adulto , Processamento Alternativo , Animais , Catarata/etiologia , Criança , Embrião não Mamífero , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Microftalmia/etiologia , Pessoa de Meia-Idade , Morfolinos/genética , Mutação de Sentido Incorreto , Oligonucleotídeos Antissenso/genética , Linhagem , Receptor EphA2 , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genética
11.
Genes (Basel) ; 12(2)2021 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-33562844

RESUMO

Dual-specificity tyrosine phosphorylation-regulated kinase 1A or DYRK1A, contributes to central nervous system development in a dose-sensitive manner. Triallelic DYRK1A is implicated in the neuropathology of Down syndrome, whereas haploinsufficiency causes the rare DYRK1A-related intellectual disability syndrome (also known as mental retardation 7). It is characterised by intellectual disability, autism spectrum disorder and microcephaly with a typical facial gestalt. Preclinical studies elucidate a role for DYRK1A in eye development and case studies have reported associated ocular pathology. In this study families of the DYRK1A Syndrome International Association were asked to self-report any co-existing ocular abnormalities. Twenty-six patients responded but only 14 had molecular confirmation of a DYRK1A pathogenic variant. A further nineteen patients from the UK Genomics England 100,000 Genomes Project were identified and combined with 112 patients reported in the literature for further analysis. Ninety out of 145 patients (62.1%) with heterozygous DYRK1A variants revealed ocular features, these ranged from optic nerve hypoplasia (13%, 12/90), refractive error (35.6%, 32/90) and strabismus (21.1%, 19/90). Patients with DYRK1A variants should be referred to ophthalmology as part of their management care pathway to prevent amblyopia in children and reduce visual comorbidity, which may further impact on learning, behaviour, and quality of life.


Assuntos
Sistema Nervoso Central/anormalidades , Sistema Nervoso Central/patologia , Deficiência Intelectual/genética , Doenças do Nervo Óptico/genética , Nervo Óptico/anormalidades , Proteínas Serina-Treonina Quinases/genética , Proteínas Tirosina Quinases/genética , Sistema Nervoso Central/crescimento & desenvolvimento , Sistema Nervoso Central/metabolismo , Criança , Pré-Escolar , Síndrome de Down/genética , Síndrome de Down/patologia , Olho/patologia , Anormalidades do Olho/genética , Anormalidades do Olho/patologia , Feminino , Haploinsuficiência/genética , Humanos , Lactente , Deficiência Intelectual/patologia , Masculino , Pessoa de Meia-Idade , Nervo Óptico/patologia , Doenças do Nervo Óptico/patologia , Erros de Refração/genética , Erros de Refração/patologia , Estrabismo/genética , Estrabismo/patologia , Quinases Dyrk
12.
Genes (Basel) ; 12(2)2021 01 22.
Artigo em Inglês | MEDLINE | ID: mdl-33499292

RESUMO

Inherited optic neuropathies, including Leber Hereditary Optic Neuropathy (LHON) and Dominant Optic Atrophy (DOA), are monogenetic diseases with a final common pathway of mitochondrial dysfunction leading to retinal ganglion cell (RGC) death and ultimately loss of vision. They are, therefore, excellent models with which to investigate this ubiquitous disease process-implicated in both common polygenetic ocular diseases (e.g., Glaucoma) and late-onset central nervous system neurodegenerative diseases (e.g., Parkinson disease). In recent years, cellular and animal models of LHON and DOA have matured in parallel with techniques (such as RNA-seq) to determine and analyze the transcriptomes of affected cells. This confluence leaves us at a particularly exciting time with the potential for the identification of novel pathogenic players and therapeutic targets. Here, we present a discussion of the importance of inherited optic neuropathies and how transcriptomic techniques can be exploited in the development of novel mutation-independent, neuroprotective therapies.


Assuntos
Predisposição Genética para Doença , Atrofias Ópticas Hereditárias/genética , Atrofias Ópticas Hereditárias/terapia , Transcriptoma , Alelos , Animais , Gerenciamento Clínico , Perfilação da Expressão Gênica/métodos , Estudos de Associação Genética , Terapia Genética , Genótipo , Humanos , Mutação , Atrofias Ópticas Hereditárias/diagnóstico , Fenótipo
13.
J Hazard Mater ; 412: 125192, 2021 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-33517053

RESUMO

We explore the role of various solution environments - chloride brines, acid mine drainage (sulfate) and groundwater (carbonate), as well as pore pressure in producing secular disequilibrium among the various radionuclides (RN) in the U-decay series upon leaching of uraninite - the most abundant U-ore and a widespread accessory mineral in U-rich rocks. We observed that the end products of the U-decay chain, 206Pb and 207Pb, exist primarily at the surface/edges of grains or within large pores in the uraninite. In contrast, the intermediate daughters 226Ra, 210Pb, 210Po, and 234/230Th, exist primarily within the bulk of uraninite, requiring breakdown by leaching for subsequent mobility to occur. Overall, pore pressure had little effect on RN mobility, with solution environment being the primary factor in creating significant mobility and disequilibrium among the RN, as it drives the initial breakdown of uraninite and influences the subsequent differential solubility of individual RNs. This was particularly the case for carbonate-bearing fluids, leading to significant fractionation of the various daughter RN arising from variable complexation and sorption phenomena. Understanding the geochemical behaviour of the RN in the U-decay series is important for predicting and managing the risks associated with RN in both environmental (acid-mine drainage) and engineered (metallurgical extraction) processes. Effective modelling of long-term RN behaviour should incorporate this strong relative fractionation caused by contrasting geochemical behaviour of individual RN during and after their release into the water from uraninite and subsequent interaction with the surrounding aquifer host rocks.

14.
Eur J Hum Genet ; 29(2): 349-355, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33024313

RESUMO

PAX6 is considered the master regulator of eye development, the majority of variants affecting this gene cause the pan-ocular developmental eye disorder aniridia. Although no genotype-phenotype correlations are clearly established, missense variants affecting the DNA-binding paired domain of PAX6 are usually associated with non-aniridia phenotypes like microphthalmia, coloboma or isolated foveal hypoplasia. In this study, we report two missense heterozygous variants in the paired domain of PAX6 resulting in isolated foveal hypoplasia with nystagmus in two independent families: c.112 C > G; p.(Arg38Gly) and c.214 G > C; p.(Gly72Arg) in exons 5 and 6, respectively. Furthermore, we provide evidence that paternal postzygotic mosaicism is the cause of inheritance, with clinically unaffected fathers and reduced affected allele fraction. This work contributes to increase the phenotypic spectrum caused by PAX6 variants, and to our knowledge, is the first report to describe the presence of postzygotic parental mosaicism causing isolated foveal hypoplasia with nystagmus. These results support the growing evidence that suggest an overestimation of sporadic cases with PAX6 variants, which has strong implications for both genetic counselling and family planning.


Assuntos
Oftalmopatias Hereditárias/genética , Fóvea Central/anormalidades , Mosaicismo , Mutação de Sentido Incorreto , Nistagmo Congênito/genética , Nistagmo Patológico/genética , Fator de Transcrição PAX6/genética , Adolescente , Adulto , Aniridia/genética , Coloboma , Proteínas do Olho/genética , Feminino , Estudos de Associação Genética , Testes Genéticos , Genótipo , Humanos , Masculino , Microftalmia/genética , Pessoa de Meia-Idade , Mutação , Pais , Linhagem , Fenótipo , Adulto Jovem
15.
J Hazard Mater ; 410: 124553, 2021 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-33223312

RESUMO

Knowledge of the behavior of technologically enhanced naturally occurring radioactive materials derived through the decay of U and its daughter products, and their subsequent fractionation, mobilization and retention, is essential to develop effective mitigation strategies and long-term radiological risk prediction. In the present study, multiple state-of-the-art, spatially resolved micro-analytical characterization techniques were combined to systematically track the liberation and migration of radionuclides (RN) from U-bearing phases in an Olympic Dam Cu flotation concentrate following sulfuric-acid-leach processing. The results highlighted the progressive dissolution of U-bearing minerals (mainly uraninite) leading to the release, disequilibrium and ultimately upgrade of daughter RN from the parent U. This occurred in conjunction with primary Cu-Fe-sulfide minerals undergoing coupled-dissolution reprecipitation to the porous secondary Cu-mineral, covellite. The budget of RN remaining in the leached concentrate was split between RN still hosted in the original U-bearing minerals, and RN that were mobilized and subsequently sorbed/precipitated onto porous covellite and auxiliary gangue mineral phases (e.g. barite). Further grinding of the flotation concentrate prior to sulfuric-acid-leach led to dissolution of U-bearing minerals previously encapsulated within Cu-Fe-sulfide minerals, resulting in increased release and disequilibrium of daughter RN, and causing further RN upgrade. The various processes that affect RN (mobility, sorption, precipitation) and sulfide minerals (coupled-dissolution reprecipitation and associated porosity generation) occur continuously within the hydrometallurgical circuit, and their interplay controls the rapid and highly localized enrichment of RN. The innovative combination of tools developed here reveal the heterogeneous distribution and fractionation of the RN in the ores following hydrometallurgical treatment at nm to cm-scales in exquisite detail. This approach provides an effective blueprint for understanding of the mobility and retention of U and its daughter products in complex anthropogenic and natural processes in the mining and energy industries.

17.
Ther Adv Ophthalmol ; 11: 2515841419835460, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30911735

RESUMO

High-throughput, massively parallel sequence analysis has revolutionized the way that researchers design and execute scientific investigations. Vast amounts of sequence data can be generated in short periods of time. Regarding ophthalmology and vision research, extensive interrogation of patient samples for underlying causative DNA mutations has resulted in the discovery of many new genes relevant to eye disease. However, such analysis remains functionally limited. RNA-sequencing accurately snapshots thousands of genes, capturing many subtypes of RNA molecules, and has become the gold standard for transcriptome gene expression quantification. RNA-sequencing has the potential to advance our understanding of eye development and disease; it can reveal new candidates to improve our molecular diagnosis rates and highlight therapeutic targets for intervention. But with a wide range of applications, the design of such experiments can be problematic, no single optimal pipeline exists, and therefore, several considerations must be undertaken for optimal study design. We review the key steps involved in RNA-sequencing experimental design and the downstream bioinformatic pipelines used for differential gene expression. We provide guidance on the application of RNA-sequencing to ophthalmology and sources of open-access eye-related data sets.

18.
Sci Rep ; 7(1): 5644, 2017 07 17.
Artigo em Inglês | MEDLINE | ID: mdl-28717200

RESUMO

Fibrosis-related events play a part in most blinding diseases worldwide. However, little is known about the mechanisms driving this complex multifactorial disease. Here we have carried out the first genome-wide RNA-Sequencing study in human conjunctival fibrosis. We isolated 10 primary fibrotic and 7 non-fibrotic conjunctival fibroblast cell lines from patients with and without previous glaucoma surgery, respectively. The patients were matched for ethnicity and age. We identified 246 genes that were differentially expressed by over two-fold and p < 0.05, of which 46 genes were upregulated and 200 genes were downregulated in the fibrotic cell lines compared to the non-fibrotic cell lines. We also carried out detailed gene ontology, KEGG, disease association, pathway commons, WikiPathways and protein network analyses, and identified distinct pathways linked to smooth muscle contraction, inflammatory cytokines, immune mediators, extracellular matrix proteins and oncogene expression. We further validated 11 genes that were highly upregulated or downregulated using real-time quantitative PCR and found a strong correlation between the RNA-Seq and qPCR results. Our study demonstrates that there is a distinct fibrosis gene signature in the conjunctiva after glaucoma surgery and provides new insights into the mechanistic pathways driving the complex fibrotic process in the eye and other tissues.


Assuntos
Doenças da Túnica Conjuntiva/genética , Redes Reguladoras de Genes , Estudo de Associação Genômica Ampla/métodos , Glaucoma/cirurgia , Análise de Sequência de RNA/métodos , Adulto , Idoso , Linhagem Celular , Doenças da Túnica Conjuntiva/etiologia , Feminino , Fibroblastos/citologia , Fibrose , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica , Ontologia Genética , Predisposição Genética para Doença , Humanos , Masculino , Pessoa de Meia-Idade
19.
Invest Ophthalmol Vis Sci ; 57(3): 1053-62, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26968735

RESUMO

PURPOSE: Mutations in the ciliary transporter gene IFT140, usually associated with a severe syndromic ciliopathy, may also cause isolated retinal dystrophy. A series of patients with nonsyndromic retinitis pigmentosa (RP) due to IFT140 was investigated in this study. METHODS: Five probands and available affected family members underwent detailed phenotyping including retinal imaging and electrophysiology. Whole exome sequencing was performed on two probands, a targeted sequencing panel of 176 retinal genes on a further two, and whole genome sequencing on the fifth. Missense mutations of IFT140 were further investigated in vitro using transient plasmid transfection of hTERT-RPE1 cells. RESULTS: Eight affected patients from five families had preserved visual acuity until at least the second decade; all had normal development without skeletal manifestations or renal failure at age 13 to 67 years (mean, 42 years; median, 44.5 years). Bi-allelic mutations in IFT140 were identified in all families including two novel mutations: c.2815T > C (p.Ser939Pro) and c.1422_23insAA (p.Arg475Asnfs*14). Expression studies demonstrated a significantly reduced number of cells showing localization of mutant IFT140 with the basal body for two nonsyndromic mutations and two syndromic mutations compared with the wild type and a polymorphism. CONCLUSIONS: This study highlights the phenotype of nonsyndromic RP due to mutations in IFT140 with milder retinal dystrophy than that associated with the syndromic disease.


Assuntos
Proteínas de Transporte/genética , Corpo Ciliar/metabolismo , DNA/genética , Mutação , Distrofias Retinianas/genética , Adolescente , Adulto , Idoso , Alelos , Proteínas de Transporte/metabolismo , Corpo Ciliar/patologia , Análise Mutacional de DNA , Exoma , Feminino , Angiofluoresceinografia , Fundo de Olho , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Linhagem , Fenótipo , Distrofias Retinianas/diagnóstico , Distrofias Retinianas/metabolismo , Distrofias Retinianas/patologia , Adulto Jovem
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