RESUMEN
The basic structure of the eye, which is crucial for visual function, is established during the embryonic process of optic cup morphogenesis. Molecular pathways of specification and patterning are integrated with spatially distinct cell and tissue shape changes to generate the eye, with discrete domains and structural features: retina and retinal pigment epithelium enwrap the lens, and the optic fissure occupies the ventral surface of the eye and optic stalk. Interest in the underlying cell biology of eye morphogenesis has led to a growing body of work, combining molecular genetics and imaging to quantify cellular processes such as adhesion and actomyosin activity. These studies reveal that intrinsic machinery and spatiotemporally specific extrinsic inputs collaborate to control dynamics of cell movements and morphologies. Here we consider recent advances in our understanding of eye morphogenesis, with a focus on the mechanics of eye formation throughout vertebrate systems, including insights and potential opportunities using organoids, which may provide a tractable system to test hypotheses from embryonic models.
Asunto(s)
Ojo/embriología , Disco Óptico/embriología , Actomiosina/metabolismo , Animales , Movimiento Celular , Ojo/metabolismo , Ojo/patología , Humanos , Cristalino/embriología , Morfogénesis/genética , Morfogénesis/fisiología , Disco Óptico/metabolismo , Organogénesis/genética , Organogénesis/fisiología , Retina/embriología , Epitelio Pigmentado de la Retina/citología , Transducción de Señal , Vertebrados/fisiologíaRESUMEN
The vertebrate eye anlage grows out of the brain and folds into bilayered optic cups. The eye is patterned along multiple axes, precisely controlled by genetic programs, to delineate neural retina, pigment epithelium, and optic stalk tissues. Pax genes encode developmental regulators of key morphogenetic events, with Pax2 being essential for interpreting inductive signals, including in the eye. PAX2 mutations cause ocular coloboma, when the ventral optic fissure fails to close. Previous studies established that Pax2 is necessary for fissure closure and to maintain the neural retina -- glial optic stalk boundary. Using a Pax2GFP/+ knock-in allele we discovered that the mutant optic nerve head (ONH) lacks molecular boundaries with the retina and RPE, rendering the ONH larger than normal. This was preceded by ventronasal cup mispatterning, a burst of overproliferation and followed by optic cup apoptosis. Our findings support the hypothesis that ONH cells are tripotential, requiring Pax2 to remain committed to glial fates. This work extends current models of ocular development, contributes to broader understanding of tissue boundary formation and informs the underlying mechanisms of human coloboma.
Asunto(s)
Ojo/embriología , Ojo/metabolismo , Disco Óptico/embriología , Factor de Transcripción PAX2/genética , Factor de Transcripción PAX2/metabolismo , Animales , Animales Modificados Genéticamente , Tipificación del Cuerpo/genética , Proliferación Celular/genética , Coloboma/genética , Femenino , Regulación del Desarrollo de la Expresión Génica , Técnicas de Sustitución del Gen , Masculino , Ratones , Ratones Endogámicos C57BL , Disco Óptico/anomalías , Disco Óptico/citología , Retina/embriología , Células Madre/metabolismoRESUMEN
The retinal basal glia (RBG) is a group of glia that migrates from the optic stalk into the third instar larval eye disc while the photoreceptor cells (PR) are differentiating. The RBGs are grouped into three major classes based on molecular and morphological characteristics: surface glia (SG), wrapping glia (WG) and carpet glia (CG). The SGs migrate and divide. The WGs are postmitotic and wraps PR axons. The CGs have giant nucleus and extensive membrane extension that each covers half of the eye disc. In this study, we used lineage tracing methods to determine the lineage relationships among these glia subtypes and the temporal profile of the lineage decisions for RBG development. We found that the CG lineage segregated from the other RBG very early in the embryonic stage. It has been proposed that the SGs migrate under the CG membrane, which prevented SGs from contacting with the PR axons lying above the CG membrane. Upon passing the front of the CG membrane, which is slightly behind the morphogenetic furrow that marks the front of PR differentiation, the migrating SG contact the nascent PR axon, which in turn release FGF to induce SGs' differentiation into WG. Interestingly, we found that SGs are equally distributed apical and basal to the CG membrane, so that the apical SGs are not prevented from contacting PR axons by CG membrane. Clonal analysis reveals that the apical and basal RBG are derived from distinct lineages determined before they enter the eye disc. Moreover, the basal SG lack the competence to respond to FGFR signaling, preventing its differentiation into WG. Our findings suggest that this novel glia-to-glia differentiation is both dependent on early lineage decision and on a yet unidentified regulatory mechanism, which can provide spatiotemporal coordination of WG differentiation with the progressive differentiation of photoreceptor neurons.
Asunto(s)
Diferenciación Celular/fisiología , Neuroglía/fisiología , Disco Óptico/embriología , Animales , Axones/metabolismo , Movimiento Celular , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/embriología , Drosophila melanogaster/metabolismo , Morfogénesis/fisiología , Neurogénesis/fisiología , Neuroglía/metabolismo , Neuronas/metabolismo , Disco Óptico/metabolismo , Células Fotorreceptoras de Invertebrados/fisiología , Retina/embriología , Retina/metabolismo , Transducción de Señal/fisiologíaRESUMEN
The bHLH transcription factor Hes1 is a key downstream effector for the Notch signaling pathway. During embryogenesis neural progenitors express low levels of Hes1 in an oscillating pattern, whereas glial brain boundary regions (e.g., isthmus) have high, sustained Hes1 levels that suppress neuronal fates. Here, we show that in the embryonic mouse retina, the optic nerve head and stalk express high Hes1, with the ONH constituting a boundary between the neural retina and glial cells that ultimately line the optic stalk. Using two Cre drivers with distinct spatiotemporal expression we conditionally inactivated Hes1, to delineate the requirements for this transcriptional repressor during retinal neurogenesis versus patterning of the optic cup and stalk. Throughout retinal neurogenesis, Hes1 maintains proliferation and blocks retinal ganglion cell formation, but surprisingly we found it also promotes cone photoreceptor genesis. In the postnatal eye, Hes1 inactivation with Rax-Cre resulted in increased bipolar neurons and a mispositioning of Müller glia. Our results indicate that Notch pathway regulation of cone genesis is more complex than previously assumed, and reveal a novel role for Hes1 in maintaining the optic cup-stalk boundary.SIGNIFICANCE STATEMENT The bHLH repressor Hes1 regulates the timing of neurogenesis, rate of progenitor cell division, gliogenesis, and maintains tissue compartment boundaries. This study expands current eye development models by showing Notch-independent roles for Hes1 in the developing optic nerve head (ONH). Defects in ONH formation result in optic nerve coloboma; our work now inserts Hes1 into the genetic hierarchy regulating optic fissure closure. Given that Hes1 acts analogously in the ONH as the brain isthmus, it prompts future investigation of the ONH as a signaling factor center, or local organizer. Embryonic development of the ONH region has been poorly studied, which is surprising given it is where the pan-ocular disease glaucoma is widely believed to inflict damage on RGC axons.
Asunto(s)
Ojo/embriología , Neurogénesis/fisiología , Factor de Transcripción HES-1/fisiología , Animales , Coloboma/genética , Coloboma/patología , Células Ependimogliales/citología , Ojo/crecimiento & desarrollo , Gastrulación , Estudios de Asociación Genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Microftalmía/genética , Microftalmía/patología , Disco Óptico/embriología , Disco Óptico/patología , Receptores Notch/fisiología , Retina/anomalías , Retina/embriología , Células Bipolares de la Retina/citología , Células Fotorreceptoras Retinianas Conos/metabolismo , Células Ganglionares de la Retina/citología , Transducción de Señal , Factor de Transcripción HES-1/deficiencia , Factor de Transcripción HES-1/genéticaRESUMEN
Optic cup morphogenesis is an intricate process. Especially, the formation of the optic fissure is not well understood. Persisting optic fissures, termed coloboma, are frequent causes for congenital blindness. Even though the defective fusion of the fissure margins is the most acknowledged reason for coloboma, highly variable morphologies of coloboma phenotypes argue for a diverse set of underlying pathomechanisms. Here, we investigate optic fissure morphogenesis in zebrafish to identify potential morphogenetic defects resulting in coloboma. We show that the formation of the optic fissure depends on tissue flow movements, integrated into the bilateral distal epithelial flow forming the optic cup. On the temporal side, the distal flow translates into a ventral perpendicular flow, shaping the temporal fissure margin. On the nasal side, however, the distal flow is complemented by tissue derived from the optic stalk, shaping the nasal fissure margin. Notably, a distinct population of TGFß-signalling positive cells is translocated from the optic stalk into both fissure margins. Furthermore, we show that induced BMP signalling as well as Wnt-signalling inhibition result in morphogenetic defects of the optic fissure. Our data also indicate that morphogenesis is crucial for a proper positioning of pre-specified dorsal-ventral optic cup domains.
Asunto(s)
Morfogénesis , Disco Óptico/metabolismo , Proteínas Wnt/metabolismo , Proteínas de Pez Cebra/metabolismo , Pez Cebra/metabolismo , Animales , Animales Modificados Genéticamente , Proteínas Morfogenéticas Óseas/genética , Proteínas Morfogenéticas Óseas/metabolismo , Coloboma/embriología , Coloboma/genética , Coloboma/metabolismo , Embrión no Mamífero/embriología , Embrión no Mamífero/metabolismo , Hibridación in Situ/métodos , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Microscopía Confocal , Disco Óptico/embriología , Imagen de Lapso de Tiempo/métodos , Proteínas Wnt/genética , Pez Cebra/embriología , Pez Cebra/genética , Proteínas de Pez Cebra/genéticaRESUMEN
Congenital abnormalities of the optic disc are not uncommon in clinical practice and should be recognized. Size abnormalities of the optic disc include optic disc aplasia, hypoplasia, megalopapilla, and optic disc cupping in prematurity. Among congenital excavations of the optic disc head, morning glory disc anomaly and optic disc pit can be complicated by serous retinal detachment; the papillorenal disc is an association of bilateral optic disc cupping and renal hypoplasia which should be ruled out; optic disc coloboma is caused by an abnormal closure of the embryonic fissure and can be complicated by choroidal neovascularization and retinal detachment. Other abnormalities that will be discussed are congenital tilted disc syndrome, duplicity of the optic disc head, congenital pigmentation of the optic disc head and myelinated retinal nerve fibers. All of these abnormalities can be associated with syndromes and neurological diseases, as well as other potentially blinding ophthalmological defects which can be secondarily complicated by amblyopia, strabismus and nystagmus. Thus, they should be recognized in order to plan for appropriate follow-up.
Asunto(s)
Anomalías del Ojo , Disco Óptico/anomalías , Coloboma/diagnóstico , Coloboma/terapia , Anomalías del Ojo/diagnóstico , Anomalías del Ojo/terapia , Humanos , Disco Óptico/diagnóstico por imagen , Disco Óptico/embriología , Disco Óptico/cirugía , Nervio Óptico/anomalíasRESUMEN
Embryonic development of the vertebrate eye begins with the formation of an optic vesicle which folds inwards to form a double-layered optic cup with a fissure on the ventral surface, known as the optic fissure. Closure of the optic fissure is essential for subsequent growth and development of the eye. A defect in this process can leave a gap in the iris, retina or optic nerve, known as a coloboma, which can lead to severe visual impairment. This review brings together current information about genes and pathways regulating fissure closure from human coloboma patients and animal models. It focuses especially on current understanding of the morphological changes and processes of epithelial remodelling occurring at the fissure margins.
Asunto(s)
Coloboma/embriología , Ojo/embriología , Disco Óptico/embriología , Trastornos de la Visión/embriología , Animales , Coloboma/genética , Ojo/metabolismo , Regulación del Desarrollo de la Expresión Génica , Humanos , Morfogénesis/genética , Disco Óptico/metabolismo , Transducción de Señal/genética , Trastornos de la Visión/genéticaRESUMEN
Purpose: The optic fissure (OF) is a transient opening in the ventral optic cup (OC) that acts as a passage for blood vessels and retinal ganglion cell axons during early eye development. Failure to close the OF is the developmental basis for uveal coloboma, a congenital blinding eye disease that significantly contributes to childhood blindness. Genes specifically expressed in the OF region may play important roles in OF development and function. The aim of this study was to characterize the transcriptome of OC cells in the OF region and investigate the function of OF-specific genes during OF closure. Methods: Laser-assisted microdissection was used to collect different regions of OC tissues. Microarray analysis was used to obtain and compare gene expression profiles of different OC regions. RNA in situ hybridization (ISH) was used to further characterize OF-specific gene expression patterns. Morpholino knockdown in zebrafish was used to study the function of a newly discovered OF-specific gene during OF closure. Results: Microarray comparison revealed that the OC at the OF region exhibited a unique gene expression profile. OC expression patterns of a number of newly discovered OF-specific genes were confirmed by ISH. Morpholino knockdown and downstream target expression and function analysis demonstrated that afap1l2, a newly discovered OF-specific gene, controls OF closure by regulating pax2a expression. Conclusions: Our study characterized the unique transcriptome of the OF region of the OC and demonstrated the essential role of a newly discovered OF-specific gene in OF closure. This study provides a valuable foundation for future mechanism dissection in OF development and physiology, and for human coloboma etiology exploration.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Embrión no Mamífero/metabolismo , Regulación del Desarrollo de la Expresión Génica/fisiología , Proteínas de Microfilamentos/genética , Disco Óptico/embriología , Factor de Transcripción PAX2/genética , Transcriptoma/genética , Proteínas de Pez Cebra/genética , Animales , Femenino , Perfilación de la Expresión Génica , Hibridación in Situ , Masculino , Ratones , Ratones Endogámicos C57BL , Análisis por Micromatrices , Morfolinos/farmacología , Disco Óptico/metabolismo , Plásmidos , ARN/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Retina/embriología , Pez Cebra/embriologíaRESUMEN
Defects in choroid fissure (CF) formation and closure lead to coloboma, a major cause of childhood blindness. Despite genetic advances, the cellular defects underlying coloboma remain poorly elucidated due to our limited understanding of normal CF morphogenesis. We address this deficit by conducting high-resolution spatio-temporal analyses of CF formation and closure in the chick, mouse and fish. We show that a small ventral midline invagination initiates CF formation in the medial-proximal optic cup, subsequently extending it dorsally toward the lens, and proximally into the optic stalk. Unlike previously supposed, the optic disc does not form solely as a result of this invagination. Morphogenetic events that alter the shape of the proximal optic cup also direct clusters of outer layer and optic stalk cells to form dorsal optic disc. A cross-species comparison suggests that CF closure can be accomplished by breaking down basement membranes (BM) along the CF margins, and by establishing BM continuity along the dorsal and ventral surfaces of the CF. CF closure is subsequently accomplished via two distinct mechanisms: tissue fusion or the intercalation of various tissues into the inter-CF space. We identify several novel cell behaviors that underlie CF fusion, many of which involve remodeling of the retinal epithelium. In addition to BM disruption, these include NCAD downregulation along the SOX2+ retinal CF margin, and the protrusion or movement of partially polarized retinal cells into the inter-CF space to mediate fusion. Proximally, the inter-CF space does not fuse or narrow and is instead loosely packed with migrating SOX2+/PAX2+/Vimentin+ astrocytes until it is closed by the outgoing optic nerve. Taken together, our results highlight distinct proximal-distal differences in CF morphogenesis and closure and establish detailed cellular models that can be utilized for understanding the genetic bases of coloboma.
Asunto(s)
Coroides/embriología , Coloboma/embriología , Coloboma/fisiopatología , Animales , Embrión de Pollo , Coroides/fisiología , Coloboma/genética , Ojo/embriología , Ratones/embriología , Morfogénesis/fisiología , Disco Óptico/embriología , Retina/embriología , Análisis Espacio-Temporal , Pez Cebra/embriologíaRESUMEN
Purpose: Coloboma is a sight-threatening congenital eye disease caused by a failure in optic fissure (OF) closure. The aim of this study was to investigate the role of Adamts16, a metalloproteinase, in OF closure. Methods: RNA in situ hybridization was used to examine the expression of Adamts16 in developing mouse and zebrafish eyes. Morpholino knockdowns were performed to study adamts16 function during zebrafish eye development. Additionally, immunofluorescent staining, RNA in situ hybridization, bromodeoxyuridine (BrdU) labeling, TUNEL assays, and high-throughput sequencing were used to examine altered cellular and molecular events in adamts16-morphant optic cups (OCs). Results: Adamts16 is expressed at the edges of the closing OF in both mice and zebrafish eyes. Zebrafish adamts16 knockdown resulted in coloboma formation. In adamts16-morphant eyes, the basement membrane failed to disassemble at the closing OF edges, OC cells exhibited decreased proliferation and increased apoptosis, and fibroblast growth factor 8 (fgf8) was ectopically upregulated in the OC. Conclusions: adamts16 is required for proper OF closure in zebrafish eyes. adamts16 controls OF closure possibly through the combined functions of degrading the basement membrane at the closing OF edges, promoting cell proliferation and survival, and restricting fgf8 expression. Our study linked a metalloproteinase to OF closure, which may facilitate future etiologic studies on human coloboma cases.
Asunto(s)
Proteínas ADAMTS/fisiología , Coloboma/embriología , Disco Óptico/anomalías , Disco Óptico/embriología , Proteínas ADAMTS/metabolismo , Animales , Membrana Basal/patología , Coloboma/metabolismo , Modelos Animales de Enfermedad , Ratones , Ratones Endogámicos C57BL , Disco Óptico/metabolismo , Pez CebraRESUMEN
During vertebrate eye morphogenesis, a transient fissure forms at its inferior part, known as the optic fissure. This will gradually close, giving rise to a healthy, spherical optic cup. Failure of the optic fissure to close gives rise to an ocular disorder known as coloboma. During this developmental process, Foxg1 is expressed in the optic neuroepithelium, with highest levels of expression in the nasal optic stalk. Foxg1-/- mutant mice have microphthalmic eyes with a large ventral coloboma. We found Wnt8b expression upregulated in the Foxg1-/- optic stalk and hypothesized that, similar to what is observed in telencephalic development, Foxg1 directs development of the optic neuroepithelium through transcriptional suppression of Wnt8b To test this, we generated Foxg1-/-;Wnt8b-/- double mutants of either sex and found that the morphology of the optic cup and stalk and the closure of the optic fissure were substantially rescued in these embryos. This rescue correlates with restored Pax2 expression in the anterior tip of the optic fissure. In addition, although we do not find evidence implicating altered proliferation in the rescue, we observe a significant increase in apoptotic cell density in Foxg1-/-;Wnt8b-/- double mutants compared with the Foxg1-/- single mutant. Upregulation of Wnt/ß-catenin target molecules in the optic cup and stalk may underlie the molecular and morphological defects in the Foxg1-/- mutant. Our results show that proper optic fissure closure relies on Wnt8b suppression by Foxg1 in the nasal optic stalk to maintain balanced apoptosis and Pax2 expression in the nasal and temporal edges of the fissure.SIGNIFICANCE STATEMENT Coloboma is an ocular disorder that may result in a loss of visual acuity and accounts for â¼10% of childhood blindness. It results from errors in the sealing of the optic fissure (OF), a transient structure at the bottom of the eye. Here, we investigate the colobomatous phenotype of the Foxg1-/- mutant mouse. We identify upregulated expression of Wnt8b in the optic stalk of Foxg1-/- mutants before OF closure initiates. Foxg1-/-;Wnt8b-/- double mutants show a substantial rescue of the Foxg1-/- coloboma phenotype, which correlates with a rescue in molecular and cellular defects of Foxg1-/- mutants. Our results unravel a new role of Foxg1 in promoting OF closure providing additional knowledge about the molecules and cellular mechanisms underlying coloboma formation.
Asunto(s)
Factores de Transcripción Forkhead/deficiencia , Proteínas del Tejido Nervioso/deficiencia , Disco Óptico/embriología , Disco Óptico/metabolismo , Proteínas Wnt/antagonistas & inhibidores , Proteínas Wnt/biosíntesis , Animales , Femenino , Factores de Transcripción Forkhead/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos CBA , Ratones Noqueados , Proteínas del Tejido Nervioso/genética , Embarazo , Factores de Transcripción/deficiencia , Factores de Transcripción/efectos de los fármacos , Proteínas Wnt/genéticaRESUMEN
BACKGROUND: The conjunctival papillae are epithelial thickenings of the conjunctiva that are required for the induction of underlying bones (the scleral ossicles). These transient papillae develop and become inductively active over an extended temporal period (HH 30-36, 6.5-10 dpf). While their inductive capacity was discovered in the mid-1900s, little is known about their development. RESULTS: Through a series of timed surgical ablations followed by in situ hybridization for Bmp2, we show that the ring of conjunctival papillae is not altered if the conjunctival epithelium is ablated either prior to or shortly after papillae induction (i.e., HH 29-30, 6.5-7 dpf). A conjunctival papilla ablated at or prior to HH 34 (8 dpf), when the complete ring is present, regenerates and quickly becomes inductively active, inducing an underlying scleral condensation with only a slight delay. This regenerative capacity extends until HH 35.5, a full 36 hours beyond the normal timeline of papillae induction. As such, the period of epithelial competency for papilla induction is longer than previously identified. CONCLUSIONS: Papilla regeneration is a mechanism that ensures the formation of a complete sclerotic ring and provides another level of redundancy for the induction of a complete sclerotic ring during the normal inductive period. Developmental Dynamics 246:381-391, 2017. © 2017 Wiley Periodicals, Inc.
Asunto(s)
Conjuntiva/crecimiento & desarrollo , Disco Óptico/crecimiento & desarrollo , Esclerótica/crecimiento & desarrollo , Animales , Embrión de Pollo , Conjuntiva/citología , Conjuntiva/embriología , Epitelio , Disco Óptico/embriología , Regeneración , Factores de TiempoRESUMEN
In the early embryo, the eyes form initially as relatively spherical optic vesicles (OVs) that protrude from both sides of the brain tube. Each OV grows until it contacts and adheres to the overlying surface ectoderm (SE) via an extracellular matrix (ECM) that is secreted by the SE and OV. The OV and SE then thicken and bend inward (invaginate) to create the optic cup (OC) and lens vesicle, respectively. While constriction of cell apices likely plays a role in SE invagination, the mechanisms that drive OV invagination are poorly understood. Here, we used experiments and computational modeling to explore the hypothesis that the ECM locally constrains the growing OV, forcing it to invaginate. In chick embryos, we examined the need for the ECM by (1) removing SE at different developmental stages and (2) exposing the embryo to collagenase. At relatively early stages of invagination (Hamburger-Hamilton stage HH14[Formula: see text]), removing the SE caused the curvature of the OV to reverse as it 'popped out' and became convex, but the OV remained concave at later stages (HH15) and invaginated further during subsequent culture. Disrupting the ECM had a similar effect, with the OV popping out at early to mid-stages of invagination (HH14[Formula: see text] to HH14[Formula: see text]). These results suggest that the ECM is required for the early stages but not the late stages of OV invagination. Microindentation tests indicate that the matrix is considerably stiffer than the cellular OV, and a finite-element model consisting of a growing spherical OV attached to a relatively stiff layer of ECM reproduced the observed behavior, as well as measured temporal changes in OV curvature, wall thickness, and invagination depth reasonably well. Results from our study also suggest that the OV grows relatively uniformly, while the ECM is stiffer toward the center of the optic vesicle. These results are consistent with our matrix-constraint hypothesis, providing new insight into the mechanics of OC (early retina) morphogenesis.
Asunto(s)
Matriz Extracelular/metabolismo , Morfogénesis , Disco Óptico/crecimiento & desarrollo , Actinas/metabolismo , Animales , Proliferación Celular , Embrión de Pollo , Simulación por Computador , Ectodermo/metabolismo , Ratones , Modelos Biológicos , Disco Óptico/anatomía & histología , Disco Óptico/embriología , Coloración y Etiquetado , Tomografía de Coherencia ÓpticaRESUMEN
The development of complex organs such as the eye requires a delicate and coordinated balance of cell division and cell death. Although apoptosis is prevalent in the proximoventral optic cup, the precise role it plays in eye development needs to be investigated further. In this study, we show that reduced apoptosis in the proximoventral optic cup prevents closure of the optic fissure. We also show that expression of ephrin A5 (Efna5) partially overlaps with Eph receptor B2 (Ephb2) expression in the proximoventral optic cup and that binding of EphB2 to ephrin A5 induces a sustained activation of JNK. This prolonged JNK signal promotes apoptosis and prevents cell proliferation. Thus, we propose that the unique cross-subclass interaction of EphB2 with ephrin A5 has evolved to function upstream of JNK signaling for the purpose of maintaining an adequate pool of progenitor cells to ensure proper closure of the optic fissure.
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Efrina-A5/metabolismo , Sistema de Señalización de MAP Quinasas , Disco Óptico/embriología , Disco Óptico/metabolismo , Receptor EphB2/metabolismo , Animales , Apoptosis/efectos de los fármacos , Membrana Celular/metabolismo , Proliferación Celular/efectos de los fármacos , Embrión de Mamíferos/metabolismo , Activación Enzimática , Efrina-A5/deficiencia , Células HEK293 , Humanos , Ratones Transgénicos , Modelos Biológicos , Morfogénesis , Receptor EphB2/deficiencia , Transducción de SeñalRESUMEN
Intramembranous ossification is a complex multi-step process which relies on extensive interactions among bone cells and surrounding tissues. The embryonic vasculature is essential in regulating endochondral ossification; however, its role during intramembranous ossification remains poorly understood, and in vivo studies are lacking. Previous research from our lab on the development of the intramembranous scleral ossicles has demonstrated an intriguing pattern of vascular development in which the areas of future osteogenesis remain avascular until after bone induction has occurred. Such avascular zones are located directly beneath each of the conjunctival papillae, epithelial structures which provide osteogenic signals to the underlying mesenchyme. Here we provide a high-resolution map of the developing vasculature from the time of ossicle induction to mineralization using a novel technique. We show that vegfa is expressed by the papillae and nearby mesenchymal tissue throughout HH 34-37, when vascular growth is taking place, and is down-regulated thereafter. Localized inhibition of Vegf results in expansion of the avascular zone surrounding the implanted papilla and mispatterning of the scleral ossicles. These results demonstrate that Vegf signaling could provide important insights into the complex relationship between bone and vasculature during intramembranous bone development.
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Osículos del Oído/embriología , Neovascularización Fisiológica/fisiología , Osteogénesis/fisiología , Esclerótica/embriología , Factor A de Crecimiento Endotelial Vascular/metabolismo , Animales , Huesos , Calcificación Fisiológica/fisiología , Embrión de Pollo , Osículos del Oído/irrigación sanguínea , Endotelio Vascular/embriología , Disco Óptico/irrigación sanguínea , Disco Óptico/embriología , Esclerótica/irrigación sanguínea , Transducción de SeñalRESUMEN
Iris epithelium is a double-layered pigmented cuboidal epithelium. According to the current model, the neural retina and the posterior iris pigment epithelium (IPE) are derived from the inner wall of the optic cup, while the retinal pigment epithelium (RPE) and the anterior IPE are derived from the outer wall of the optic cup during development. Our current study shows evidence, contradicting this model of fetal iris development. We demonstrate that human fetal iris expression patterns of Otx2 and Mitf transcription factors are similar, while the expressions of Otx2 and Sox2 are complementary. Furthermore, IPE and RPE exhibit identical morphologic development during the early embryonic period. Our results suggest that the outer layer of the optic cup forms two layers of the iris epithelium, and the posterior IPE is the inward-curling anterior rim of the outer layer of the optic cup. These findings provide a reasonable explanation of how IPE cells can be used as an appropriate substitute for RPE cells.
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Iris/embriología , Pigmentación , Animales , Diferenciación Celular , Epitelio/embriología , Epitelio/inervación , Epitelio/metabolismo , Femenino , Regulación del Desarrollo de la Expresión Génica , Humanos , Iris/citología , Iris/inervación , Iris/metabolismo , Ratones , Factor de Transcripción Asociado a Microftalmía/metabolismo , Disco Óptico/embriología , Disco Óptico/metabolismo , Factores de Transcripción Otx/metabolismo , Embarazo , Transporte de Proteínas , Factores de Transcripción SOXB1/metabolismoRESUMEN
Mutations in Peroxidasin (PXDN) cause severe inherited eye disorders in humans, such as congenital cataract, corneal opacity and developmental glaucoma. The role of peroxidasin during eye development is poorly understood. Here, we describe the first Pxdn mouse mutant which was induced by ENU (N-ethyl-N-nitrosourea) and led to a recessive phenotype. Sequence analysis of cDNA revealed a T3816A mutation resulting in a premature stop codon (Cys1272X) in the peroxidase domain. This mutation causes severe anterior segment dysgenesis and microphthalmia resembling the manifestations in patients with PXDN mutations. The proliferation and differentiation of the lens is disrupted in association with aberrant expression of transcription factor genes (Pax6 and Foxe3) in mutant eyes. Additionally, Pxdn is involved in the consolidation of the basement membrane and lens epithelium adhesion in the ocular lens. Lens material including γ-crystallin is extruded into the anterior and posterior chamber due to local loss of structural integrity of the lens capsule as a secondary damage to the anterior segment development leading to congenital ocular inflammation. Moreover, Pxdn mutants exhibited an early-onset glaucoma and progressive retinal dysgenesis. Transcriptome profiling revealed that peroxidasin affects the transcription of developmental and eye disease-related genes at early eye development. These findings suggest that peroxidasin is necessary for cell proliferation and differentiation and for basement membrane consolidation during eye development. Our studies provide pathogenic mechanisms of PXDN mutation-induced congenital eye diseases.
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Proteínas de la Matriz Extracelular/genética , Ojo/embriología , Ojo/metabolismo , Organogénesis/genética , Peroxidasa/genética , Animales , Adhesión Celular , Proliferación Celular , Análisis Mutacional de ADN , Matriz Extracelular/metabolismo , Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Estudios de Asociación Genética , Ligamiento Genético , Genotipo , Cristalino/embriología , Cristalino/metabolismo , Masculino , Ratones , Mutación , Disco Óptico/embriología , Disco Óptico/metabolismo , Fenotipo , Polimorfismo de Nucleótido Simple , Retina/embriología , Retina/metabolismo , Retina/patología , PeroxidasinaRESUMEN
BACKGROUND: Heparan sulfate proteoglycans (HSPG) are important for embryonic development by means of the regulation of gradient formation and signaling of multiple growth factors and morphogens. Previous studies have shown that Bmp/Shh/Fgf signaling are required for the regionalization of the optic vesicle (OV) and for the closure of the optic fissure (OF), the disturbance of which underlie ocular anomalies such as microphthalmia, coloboma, and optic nerve hypoplasia. RESULTS: To study HSPG-dependent coordination of these signaling pathways during mammalian visual system development, we have generated a series of OV-specific mutations in the heparan sulfate (HS) N-sulfotransferase genes (Ndst1 and Ndst2) and HS O-sulfotransferase genes (Hs2st, Hs6st1, and Hs6st2) in mice. Of interest, the resulting HS undersulfation still allowed for normal retinal neurogenesis and optic fissure closure, but led to defective optic disc and stalk development. The adult mutant animals further developed optic nerve aplasia/hypoplasia and displayed retinal degeneration. We observed that MAPK/ERK signaling was down-regulated in Ndst mutants, and consistent with this, HS-related optic nerve morphogenesis defects in mutant mice could partially be rescued by constitutive Kras activation. CONCLUSIONS: These results suggest that HSPGs, depending on their HS sulfation pattern, regulate multiple signaling pathways in optic disc and stalk morphogenesis.
Asunto(s)
Proteoglicanos de Heparán Sulfato/fisiología , Morfogénesis , Disco Óptico/embriología , Tracto Óptico/embriología , Amidohidrolasas/genética , Animales , Embrión de Mamíferos , Ratones , Ratones Transgénicos , Morfogénesis/genética , Disco Óptico/metabolismo , Enfermedades del Nervio Óptico/genética , Tracto Óptico/metabolismo , Degeneración Retiniana/genética , Transducción de Señal/genética , Sulfotransferasas/genéticaRESUMEN
Nineteen Wnt ligands and 10 Frizzled (Fz) receptors mediate multiple distinct cellular events during neuronal development. However, their precise roles in cell-type specification and organogenesis are poorly delineated because of overlapping functions and expression profiles. Here, we have explored the role of two closely related Frizzled receptors, Fz5 and Fz8, in mouse retinal development. We previously showed that Fz5(-/-) mice exhibit mild coloboma and microphthalmia at ~50% penetrance. Fz8 expression overlaps with Fz5 in the neural retina and optic fissure/disc. Mice lacking Fz8 show minimal eye and retinal defects. The embryos lacking both Fz5 and Fz8 die early in development, but a majority of triallelic Fz5(-/-);Fz8(+/-) mutants survive until birth. The triallelic mutant develops severe retinal coloboma and microphthalmia with full penetrance. At the cellular level, impaired neurogenesis is indicated by increased early-born retinal neurons that result from accelerated cell cycle exit of progenitors. Deficiency of apical retinal neuroepithelium is indicated by altered localization of apical junction markers, such as atypical protein kinase C, RhoA and ß-catenin. Hes1 expression, which is critical for retinal progenitor expansion, is down-regulated in the triallelic mutant mouse. Furthermore, blocking Frizzled receptors in cultured retinal explants led to basally shifted divisions of retinal progenitors. Together, our studies suggest a dose-dependent regulation of signaling by Fz5 and Fz8 in optic fissure/disc formation and progenitor expansion.
Asunto(s)
Receptores Frizzled/genética , Receptores Frizzled/fisiología , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/fisiología , Retina/embriología , Neuronas Retinianas/fisiología , Animales , Axones/fisiología , Coloboma/genética , Coloboma/metabolismo , Regulación hacia Abajo , Regulación del Desarrollo de la Expresión Génica , Gliosis/genética , Gliosis/metabolismo , Ratones , Ratones Noqueados , Microftalmía/genética , Microftalmía/metabolismo , Mitosis , Mutación , Células-Madre Neurales/citología , Células-Madre Neurales/fisiología , Neurogénesis , Disco Óptico/embriología , Disco Óptico/metabolismo , Retina/metabolismo , Neuronas Retinianas/citología , Transducción de Señal , beta Catenina/genética , beta Catenina/metabolismo , Proteínas de Unión al GTP rho/genética , Proteínas de Unión al GTP rho/metabolismo , Proteína de Unión al GTP rhoARESUMEN
Fatty Liver Shionogi (FLS) mice have been shown to develop a hereditary disorder characterized by localized retinochoroidal defects of the ventral fundus very similar to human typical ocular coloboma without microphthalmia. The objective of this study was to determine when and how the failure of the optic fissure closure occurs, and to clarify the disturbed mechanism of basement membrane disintegration during embryonal stage in FLS mice. Fetuses at day 11.5-15.5 of gestation were obtained from dams of FLS and BALB/c strain of mice. Coronal serial sections through the eye were examined by light and electron microscopy. The sections were followed by observation of the basement membrane using reaction with periodic acid-Schiff (PAS) reagent and immunohistochemical staining with anti-Laminin and anti-Type IV collagen antibodies. Both optic fissure margins closely approached each other up to GD 11.5 in all FLS and BALB/c embryos. The inner and outer layers of the optic cup did not normally fuse at midlenticular levels of the optic fissure in almost 70% of FLS fetuses by GD 15.5, whereas both margins were completely fused in all BALB/c fetuses of the same gestational day. In the FLS fetuses at GD 12.5, rolling on one side of fissure margins and consequent asymmetry were observed at the ventral optic fissure. The basement membrane persisted after the close contact of both sides of the fissure margins during GD 11.5 and 15.5. Ultrastructurally, the basal lamina was not disintegrated and mesenchymal cells intervened between the two neuroepithelial layers, resulting in complete separation of both fissure margins at GD 13.0. It is highly probable that the disturbed basement membrane disintegration right before optic fissure closure causes mild ocular coloboma without microphthalmia in FLS mice.