Optic cup morphogenesis across species and related inborn human eye defects.

The vertebrate eye is shaped as a cup, a conformation that optimizes vision and is acquired early in development through a process known as optic cup morphogenesis. Imaging living, transparent teleost embryos and mammalian stem cell-derived organoids has provided insights into the rearrangements that eye progenitors undergo to adopt such a shape. Molecular and pharmacological interference with these rearrangements has further identified the underlying molecular machineries and the physical forces involved in this morphogenetic process. In this Review, we summarize the resulting scenarios and proposed models that include common and species-specific events. We further discuss how these studies and those in environmentally adapted blind species may shed light on human inborn eye malformations that result from failures in optic cup morphogenesis, including microphthalmia, anophthalmia and coloboma.

Adenohypophysis placodal precursors exhibit distinctive features within the rostral preplacodal ectoderm.

Placodes are discrete thickenings of the vertebrate cranial ectoderm that generate morpho-functionally distinct structures, such as the adenohypophysis, olfactory epithelium and lens. All placodes arise from a horseshoe-shaped preplacodal ectoderm in which the precursors of individual placodes are intermingled. However, fate-map studies indicated that cells positioned at the preplacodal midline give rise to only the adenohypophyseal placode, suggesting a unique organization of these precursors within the preplacode. To test this possibility, we combined embryological and molecular approaches in chick embryos to show that, at gastrula stage, adenohypophyseal precursors are clustered in the median preplacodal ectoderm, largely segregated from those of the adjacent olfactory placode. Median precursors are elongated, densely packed and, at neurula stage, express a molecular signature that distinguishes them from the remaining preplacodal cells. Olfactory placode precursors and midline neural cells can replace ablated adenohypophyseal precursors up to head-fold stage, although with a more plastic organization. We thus propose that adenohypophyseal placode precursors are unique within the preplacodal ectoderm possibly because they originate the only single placode and the only one with an endocrine character.

Reduce, reuse, recycle - Developmental signals in spinal cord regeneration.

Anamniotes, fishes and amphibians, have the capacity to regenerate spinal cord tissue after injury, generating new neurons that mature and integrate into the spinal circuitry. Elucidating the molecular signals that promote this regeneration is a fundamental question in regeneration research. Model systems, such as salamanders and larval and adult zebrafish are used to analyse successful regeneration. This shows that many developmental signals, such as Notch, Hedgehog (Hh), Bone Morphogenetic Protein (BMP), Wnt, Fibroblast Growth Factor (FGF), Retinoic Acid (RA) and neurotransmitters are redeployed during regeneration and activate resident spinal progenitor cells. Here we compare the roles of these signals in spinal cord development and regeneration of the much larger and fully patterned adult spinal cord. Understanding how developmental signalling systems are reactivated in successfully regenerating species may ultimately lead to ways to reactivate similar systems in mammalian progenitor cells, which do not show neurogenesis after spinal injury.

Secreted frizzled related proteins modulate pathfinding and fasciculation of mouse retina ganglion cell axons by direct and indirect mechanisms.

Retina ganglion cell (RGC) axons grow along a stereotyped pathway undergoing coordinated rounds of fasciculation and defasciculation, which are critical to establishing proper eye-brain connections. How this coordination is achieved is poorly understood, but shedding of guidance cues by metalloproteinases is emerging as a relevant mechanism. Secreted Frizzled Related Proteins (Sfrps) are multifunctional proteins, which, among others, reorient RGC growth cones by regulating intracellular second messengers, and interact with Tolloid and ADAM metalloproteinases, thereby repressing their activity. Here, we show that the combination of these two functions well explain the axon guidance phenotype observed in Sfrp1 and Sfrp2 single and compound mouse mutant embryos, in which RGC axons make subtle but significant mistakes during their intraretinal growth and inappropriately defasciculate along their pathway. The distribution of Sfrp1 and Sfrp2 in the eye is consistent with the idea that Sfrp1/2 normally constrain axon growth into the fiber layer and the optic disc. Disheveled axon growth instead seems linked to Sfrp-mediated modulation of metalloproteinase activity. Indeed, retinal explants from embryos with different Sfrp-null alleles or explants overexpressing ADAM10 extend axons with a disheveled appearance, which is reverted by the addition of Sfrp1 or an ADAM10-specific inhibitor. This mode of growth is associated with an abnormal proteolytic processing of L1 and N-cadherin, two ADAM10 substrates previously implicated in axon guidance. We thus propose that Sfrps contribute to coordinate visual axon growth with a dual mechanism: by directly signaling at the growth cone and by regulating the processing of other relevant cues.

Maternal vgll4a regulates zebrafish epiboly through Yap1 activity.

Gastrulation in zebrafish embryos commences with the morphogenetic rearrangement of blastodermal cells, which undergo a coordinated spreading from the animal pole to wrap around the egg at the vegetal pole. This rearrangement, known as epiboly, relies on the orchestrated activity of maternal transcripts present in the egg, compensating for the gradual activation of the zygotic genome. Epiboly involves the mechano-transducer activity of yap1 but what are the regulators of yap1 activity and whether these are maternally or zygotically derived remain elusive. Our study reveals the crucial role of maternal vgll4a, a proposed Yap1 competitor, during zebrafish epiboly. In embryos lacking maternal/zygotic vgll4a (MZvgll4a), the progression of epiboly and blastopore closure is delayed. This delay is associated with the ruffled appearance of the sliding epithelial cells, decreased expression of yap1-downstream targets and transient impairment of the actomyosin ring at the syncytial layer. Our study also shows that, rather than competing with yap1, vgll4a modulates the levels of the E-cadherin/ß-catenin adhesion complex at the blastomeres' plasma membrane and hence their actin cortex distribution. Taking these results together, we propose that maternal vgll4a acts at epiboly initiation upstream of yap1 and the E-cadherin/ß-catenin adhesion complex, contributing to a proper balance between tissue tension/cohesion and contractility, thereby promoting a timely epiboly progression.

Disruption of chemoreceptor signalling arrays by high levels of CheW, the receptor-kinase coupling protein.

During chemotactic signalling by Escherichia coli, the small cytoplasmic CheW protein couples the histidine kinase CheA to chemoreceptor control. Although essential for assembly and operation of receptor signalling complexes, CheW in stoichiometric excess disrupts chemotactic behaviour. To explore the mechanism of the CheW excess effect, we measured the physiological consequences of high cellular levels of wild-type CheW and of several CheW variants with reduced or enhanced binding affinities for receptor molecules. We found that high levels of CheW interfered with trimer assembly, prevented CheA activation, blocked cluster formation, disrupted chemotactic ability and elevated receptor methylation levels. The severity of these effects paralleled the receptor-binding affinities of the CheW variants. Because trimer formation may be an obligate step in the assembly of ternary signalling complexes and higher-order receptor arrays, we suggest that all CheW excess effects stem from disruption of trimer assembly. We propose that the CheW-binding sites in receptor dimers overlap their trimer contact sites and that high levels of CheW saturate the receptor-binding sites, preventing trimer assembly. The CheW-trapped receptor dimers seem to be improved substrates for methyltransferase reactions, but cannot activate CheA or assemble into clusters, processes that are essential for chemotactic signalling.

Mosmo Is Required for Zebrafish Craniofacial Formation.

Hedgehog (Hh) signaling is a highly regulated molecular pathway implicated in many developmental and homeostatic events. Mutations in genes encoding primary components or regulators of the pathway cause an array of congenital malformations or postnatal pathologies, the extent of which is not yet fully defined. Mosmo (Modulator of Smoothened) is a modulator of the Hh pathway, which encodes a membrane tetraspan protein. Studies in cell lines have shown that Mosmo promotes the internalization and degradation of the Hh signaling transducer Smoothened (Smo), thereby down-modulating pathway activation. Whether this modulation is essential for vertebrate embryonic development remains poorly explored. Here, we have addressed this question and show that in zebrafish embryos, the two mosmo paralogs, mosmoa and mosmob, are expressed in the head mesenchyme and along the entire ventral neural tube. At the cellular level, Mosmoa localizes at the plasma membrane, cytoplasmic vesicles and primary cilium in both zebrafish and chick embryos. CRISPR/Cas9 mediated inactivation of both mosmoa and mosmob in zebrafish causes frontonasal hypoplasia and craniofacial skeleton defects, which become evident in the adult fish. We thus suggest that MOSMO is a candidate to explain uncharacterized forms of human congenital craniofacial malformations, such as those present in the 16p12.1 chromosomal deletion syndrome encompassing the MOSMO locus.

Stretching of the retinal pigment epithelium contributes to zebrafish optic cup morphogenesis.

The vertebrate eye primordium consists of a pseudostratified neuroepithelium, the optic vesicle (OV), in which cells acquire neural retina or retinal pigment epithelium (RPE) fates. As these fates arise, the OV assumes a cup shape, influenced by mechanical forces generated within the neural retina. Whether the RPE passively adapts to retinal changes or actively contributes to OV morphogenesis remains unexplored. We generated a zebrafish Tg(E1-bhlhe40:GFP) line to track RPE morphogenesis and interrogate its participation in OV folding. We show that, in virtual absence of proliferation, RPE cells stretch and flatten, thereby matching the retinal curvature and promoting OV folding. Localized interference with the RPE cytoskeleton disrupts tissue stretching and OV folding. Thus, extreme RPE flattening and accelerated differentiation are efficient solutions adopted by fast-developing species to enable timely optic cup formation. This mechanism differs in amniotes, in which proliferation drives RPE expansion with a much-reduced need of cell flattening.

Rounded eyeballs help to optimize vision ­ but how do they acquire their distinctive shape? In animals with backbones, including humans, the eye begins to form early in development. A single layer of embryonic tissue called the optic vesicle reorganizes itself into a two-layered structure: a thin outer layer of cells, known as the retinal pigmented epithelium (RPE for short), and a thicker inner layer called the neural retina. If this process fails, the animal may be born blind or visually impaired. How this flat two-layered structure becomes round is still being investigated. In fish, studies have shown that the inner cell layer ­ the neural retina ­ generates mechanical forces that cause the developing tissue to curve inwards to form a cup-like shape. But it was unclear whether the outer layer of cells (the RPE) also contributed to this process. Moreno-Marmol et al. were able to investigate this question by genetically modifying zebrafish to make all new RPE cells fluoresce. Following the early development of the zebrafish eye under a microscope revealed that RPE cells flattened themselves into long thin structures that stretched to cover the entire neural retina. This change was made possible by the cell's internal skeleton reorganizing. In fact, preventing this reorganization stopped the RPE cells from flattening, and precluded the optic cup from acquiring its curved shape. The results thus confirmed a direct role for the RPE in generating curvature. The entire process did not require the RPE to produce new cells, allowing the curved shape to emerge in just a few hours. This is a major advantage for fast-developing species such as zebrafish. In species whose embryos develop more slowly, such as mice and humans, the RPE instead grows by producing additional cells ­ a process that takes many days. The development of the eye thus shows how various species use different evolutionary approaches to achieve a common goal.

Patterning the Vertebrate Retina with Morphogenetic Signaling Pathways.

The primordium of the vertebrate eye is composed of a pseudostratified and apparently homogeneous neuroepithelium, which folds inward to generate a bilayered optic cup. During these early morphogenetic events, the optic vesicle is patterned along three different axes-proximo-distal, dorso-ventral, and naso-temporal-and three major domains: the neural retina, the retinal pigment epithelium (RPE), and the optic stalk. These fundamental steps that enable the subsequent development of a functional eye, entail the precise coordination among genetic programs. These programs are driven by the interplay of signaling pathways and transcription factors, which progressively dictate how each tissue should evolve. Here, we discuss the contribution of the Hh, Wnt, FGF, and BMP signaling pathways to the early patterning of the retina. Comparative studies in different vertebrate species have shown that their morphogenetic activity is repetitively used to orchestrate the progressive specification of the eye with evolutionary conserved mechanisms that have been adapted to match the specific need of a given species.

Interaction of Axonal Chondrolectin with Collagen XIXa1 Is Necessary for Precise Neuromuscular Junction Formation.

Chondrolectin (Chodl) is needed for motor axon extension in zebrafish and is dysregulated in mouse models of spinal muscular atrophy (SMA). However, the mechanistic basis of Chodl function is not known. Here, we use Chodl-deficient zebrafish and mouse mutants to show that the absence of Chodl leads to anatomical and functional defects of the neuromuscular synapse. In zebrafish, the growth of an identified motor axon beyond an "en passant" synapse and later axon branching from synaptic points are impaired, leading to functional deficits. Mechanistically, motor-neuron-autonomous Chodl function depends on its intracellular domain and on binding muscle-derived collagen XIXa1 by its extracellular C-type lectin domain. Our data support evolutionarily conserved roles of Chodl in synaptogenesis and provide evidence for a "synapse-first" scenario of motor axon growth in zebrafish.

Mapping the Global Chromatin Connectivity Network for Sox2 Function in Neural Stem Cell Maintenance.

The SOX2 transcription factor is critical for neural stem cell (NSC) maintenance and brain development. Through chromatin immunoprecipitation (ChIP) and chromatin interaction analysis (ChIA-PET), we determined genome-wide SOX2-bound regions and Pol II-mediated long-range chromatin interactions in brain-derived NSCs. SOX2-bound DNA was highly enriched in distal chromatin regions interacting with promoters and carrying epigenetic enhancer marks. Sox2 deletion caused widespread reduction of Pol II-mediated long-range interactions and decreased gene expression. Genes showing reduced expression in Sox2-deleted cells were significantly enriched in interactions between promoters and SOX2-bound distal enhancers. Expression of one such gene, Suppressor of Cytokine Signaling 3 (Socs3), rescued the self-renewal defect of Sox2-ablated NSCs. Our work identifies SOX2 as a major regulator of gene expression through connections to the enhancer network in NSCs. Through the definition of such a connectivity network, our study shows the way to the identification of genes and enhancers involved in NSC maintenance and neurodevelopmental disorders.

Myiasis associated with an invasive ductal carcinoma of the left breast: case study.

Most breast cancers originate in the ductal epithelium and are referred to as invasive ductal carcinoma. In this study we report on the clinical procedures adopted to diagnose myiasis in association with infiltrating metastatic breast carcinoma in a female patient. A 41 years old woman came to the Federal Hospital of Andaraí complaining of intense itching, warmth, redness and hardening of the breast, which had acquired the aspect of an orange peel. A lesion in the left breast was cavitated, dimpled, had fetid odor, and had fibrotic and infected air nodules filled with exudate and Dipteran larvae. The tissue was cleaned and 33 larvae were extracted. The patient was hospitalized and received Ivermectin. Eighteen of the larvae extracted from the patient were placed in 70% alcohol, and twelve were placed in a container with sterile wood shavings under controlled conditions until they metamorphosed into adults. The taxonomic identification of the flies revealed that the culprit was Cochliomyia hominivorax. A histopathological exam conducted three months earlier had revealed infiltrating ductal carcinoma. Two months after the myiasis treatment, the breast tissue had healed. The patient had waited ten days from the onset of the myiasis to seek treatment, and that delay interfered negatively in the prognosis of both the neoplasm and the myiasis. This study is relevant to public health in view of the strong social impact of myiasis.

Cdon acts as a Hedgehog decoy receptor during proximal-distal patterning of the optic vesicle.

Patterning of the vertebrate optic vesicle into proximal/optic stalk and distal/neural retina involves midline-derived Hedgehog (Hh) signalling, which promotes stalk specification. In the absence of Hh signalling, the stalks are not specified, causing cyclopia. Recent studies showed that the cell adhesion molecule Cdon forms a heteromeric complex with the Hh receptor Patched 1 (Ptc1). This receptor complex binds Hh and enhances signalling activation, indicating that Cdon positively regulates the pathway. Here we show that in the developing zebrafish and chick optic vesicle, in which cdon and ptc1 are expressed with a complementary pattern, Cdon acts as a negative Hh signalling regulator. Cdon predominantly localizes to the basolateral side of neuroepithelial cells, promotes the enlargement of the neuroepithelial basal end-foot and traps Hh protein, thereby limiting its dispersion. This Ptc-independent function protects the retinal primordium from Hh activity, defines the stalk/retina boundary and thus the correct proximo-distal patterning of the eye.

Diagnóstico de helmintiasis por videocápsula endoscópica / Diagnosis of helminthiasis by endoscopic videocapsule

Resumen La helmintiasis corresponde a una parasitosis intestinal altamente prevalente en países en desarrollo, y que viene aumentando en los países desarrollados. Teniendo en cuenta el ciclo de vida de los helmintos, su presentación se relaciona con situaciones ambientales y socioeconómicas de alto riesgo. Por tanto, su presencia endémica puede considerarse un problema de salud pública. Para el diagnóstico, se disponen de varios elementos que incluyen el estudio de la materia fecal, la endoscopia y las imágenes diagnósticas como los estudios de rayos X de vías digestivas, la tomografía axial computarizada (TAC) y la resonancia magnética (RM). En la actualidad, existen reportes de diagnóstico de helmintiasis por cápsula endoscópica que también han permitido el estudio de la ecología y de la patología in situ de estos parásitos. Se presenta un caso clínico de helmintiasis compatible con estrongiloidosis, diagnosticado por cápsula endoscópica por el Grupo de Gastroenterología Clínico-Quirúrgica de la Universidad de Caldas, Colombia.

Abstract Helminthiasis is an intestinal parasitosis that is very prevalent in developing countries but is increasing in developed countries. Taking into account the life cycle of helminths, their presentation is related to high-risk environmental and socioeconomic situations. Endemic presence can be considered a public health problem. Several methods are available for diagnosis including the study of fecal matter, endoscopy and diagnostic imaging such as X-ray studies of the digestive tract, CT scans and magnetic resonance. Recently, there have been reports of diagnosis of helminthiasis by capsule endoscopy which has also allowed the study of the ecology and in situ pathology of these parasites. We present a clinical case of helminthiasis compatible with strongyloidiasis that was diagnosed by endoscopic capsule by the surgical clinical gastroenterology group of the University of Caldas in Manizales, Colombia.

Carcinomas escamosos com miíase, uma nova tendência? / Squamous cell carcinomas with myiasis, a new trend?

Importância do problema: Este artigo se propõe a descrever dois casos de miíase em feridas neoplásicas de carcinomas escamosos, uma associação rara que vem se tornando comum. Comentários: Esta associação deve-se as altas prevalências de HPV, baixas condições socioeconômicas e de higiene, além do manejo inadequado de feridas e do descaso dos homens para com sua saúde, os principais afetados quando o assunto é câncer orofaríngeo e de pênis.(AU)

Relevance: This article intends to describe two cases of myiasis on squamous cells neoplasic wounds, a rare association that are becoming more easily seen nowadays. Comments: This association due to high HPV serum prevalences, low social and hygiene conditions, in addition to the precarious wounds care and men´s health seek relapse, the mainly affected when the issue is oropharynx and penis cancers. (AU)

Cdon and Boc: Two transmembrane proteins implicated in cell-cell communication.

Cdon and Boc, and their Drosophila homologues Ihog and Boi, are evolutionary conserved transmembrane glycoproteins belonging to a subgroup of the Immunoglobulin superfamily of cell adhesion molecules (CAMs). Initially isolated in vertebrates as CAMs that link cadherin function with MAPK signaling in myoblast differentiation, they have thereafter been shown to act as essential receptors for the Hedgehog (Hh) family of secreted proteins. They associate with both ligand and other Hh receptor components, including Ptch and Gas1, thus forming homo- and heteromeric complexes. In Drosophila, they are also involved in ligand processing and release from Hh producing cells. Cdon/Boc and Ihog/Boi can substitute one another and play redundant functions is some contexts. In addition, Boc, but not Cdon, mediates axon guidance information provided by Hh in specific neuronal populations, whereas mutations in the CDON cause holoprosencephaly, a human congenital anomaly defined by forebrain midline defects prominently associated with diminished Hh pathway activity.

Pαx6 expression in postmitotic neurons mediates the growth of axons in response to SFRP1.

During development, the mechanisms that specify neuronal subclasses are coupled to those that determine their axonal response to guidance cues. Pax6 is a homedomain transcription factor required for the specification of a variety of neural precursors. After cell cycle exit, Pax6 expression is often shut down in the precursor progeny and most postmitotic neurons no longer express detectable levels of the protein. There are however exceptions and high Pax6 protein levels are found, for example, in postmitotic retinal ganglion cells (RGCs), dopaminergic neurons of the olfactory bulb and the limbic system in the telencephalon. The function of Pax6 in these differentiating neurons remains mostly elusive. Here, we demonstrate that Pax6 mediates the response of growing axons to SFRP1, a secreted molecule expressed in several Pax6-positive forebrain territories. Forced expression of Pax6 in cultured postmitotic cortical neurons, which do not normally express Pax6, was sufficient to increment axonal length. Growth was blocked by the addition of anti-SFRP1 antibodies, whereas exogenously added SFRP1 increased axonal growth of Pax6-transfected neurons but not that of control or untransfected cortical neurons. In the reverse scenario, shRNA-mediated knock-down of Pax6 in mouse retinal explants specifically abolished RGCs axonal growth induced by SFRP1, but had no effect on RGCs differentiation and it did not modify the effect of Shh or Netrin on axon growth. Taken together these results demonstrate that expression of Pax6 is necessary and sufficient to render postmitotic neurons competent to respond to SFRP1. These results reveal a novel and unexpected function of Pax6 in postmitotic neurons and situate Pax6 and SFRP1 as pair regulators of axonal connectivity.

Myiasis associated with an invasive ductal carcinoma of the left breast: case study

ABSTRACT Most breast cancers originate in the ductal epithelium and are referred to as invasive ductal carcinoma. In this study we report on the clinical procedures adopted to diagnose myiasis in association with infiltrating metastatic breast carcinoma in a female patient. A 41 years old woman came to the Federal Hospital of Andaraí complaining of intense itching, warmth, redness and hardening of the breast, which had acquired the aspect of an orange peel. A lesion in the left breast was cavitated, dimpled, had fetid odor, and had fibrotic and infected air nodules filled with exudate and Dipteran larvae. The tissue was cleaned and 33 larvae were extracted. The patient was hospitalized and received Ivermectin. Eighteen of the larvae extracted from the patient were placed in 70% alcohol, and twelve were placed in a container with sterile wood shavings under controlled conditions until they metamorphosed into adults. The taxonomic identification of the flies revealed that the culprit was Cochliomyia hominivorax. A histopathological exam conducted three months earlier had revealed infiltrating ductal carcinoma. Two months after the myiasis treatment, the breast tissue had healed. The patient had waited ten days from the onset of the myiasis to seek treatment, and that delay interfered negatively in the prognosis of both the neoplasm and the myiasis. This study is relevant to public health in view of the strong social impact of myiasis.