Regulation of fenestra formation via actin-dynamin2 interaction in rat pituitary endothelial cells.

Endothelial fenestrae are transcellular pores divided by a diaphragm consisting of plasmalemma vesicle-associated protein (PLVAP). They function as a channel for peptide hormones and other substances. Invagination of the plasma membrane is necessary for the fenestra formation. The actin cytoskeleton is essential for scission of endocytic vesicles from the invaginated plasma membrane. Therefore, we examined the involvement of the actin cytoskeleton in fenestra formation in cultured endothelial cells isolated from the anterior lobe (AL) of the rat pituitary, using immunofluorescence and scanning electron microscopy. Inhibition of polymerization and depolymerization of the actin cytoskeleton by latrunculin A and jasplakinolide, respectively, remarkably increased the PLVAP-positive sieve plate area and number of fenestrae. Jasplakinolide significantly affected the arrangement of the fenestra on the cell surface, resulting in parallel serpentine furrows of the fenestra. These results suggest that the actin cytoskeleton not only induces fenestra formation but also regulates cell arrangement. Dynamin is a scission protein of the invaginated plasma membrane and interacts with the actin cytoskeleton. We found that dynamin2 is mainly expressed in the endothelial cells of the rat AL. We then investigated the function of dynamin2 by the treatment with dyngo-4a, a potent inhibitor of dynamin1 and dynamin2, on the fenestra formation. As a result, the PLVAP-positive area is significantly increased by the treatment. These results show that the actin-dynamin2 interaction is essential for the control of the fenestra formation in endothelial cells of rat AL. In conclusion, the actin cytoskeleton and dynamin2 function as regulators of endothelial fenestra formation.

Defining the critical period of hedgehog pathway inhibitor-induced cranial base dysplasia in mice.

BACKGROUND: The hedgehog signaling pathway is critical for developmental patterning of the limb, craniofacial and axial skeleton. Disruption of this pathway in mice leads to a series of structural malformations, but the exact role and critical period of the Hh pathway in the early development of the cranial base have been rarely described. RESULTS: Embryos exposed to vismodegib from E7.5, E9.5, and E10.5 had a higher percentage of cranial base fenestra. The peak incidence of hypoplasia in sphenoid winglets and severe craniosynostosis in cranial base synchondroses was observed when vismodegib was administered between E9.5 and E10.5. Cranial base craniosynostosis results from accelerating terminal differentiation of chondrocytes and premature osteogenesis. CONCLUSIONS: We define the critical periods for the induction of cranial base deformity by vismodegib administration at a meticulous temporal resolution. Our findings suggest that the Hh pathway may play a vital role in the early development of the cranial base. This research also establishes a novel and easy-to-establish mouse model of synostosis in the cranial base using a commercially available pathway-selective inhibitor.

Fibronectin-integrin signaling regulates PLVAP localization at endothelial fenestrae by microtubule stabilization.

Endothelial fenestrae are the transcellular pores existing on the capillary walls which are organized in clusters referred to as sieve plates. They are also divided by a diaphragm consisting of plasmalemma vesicle-associated protein (PLVAP). In this study, we examined the involvement of fibronectin signaling in the formation of fenestra and diaphragm in endothelial cells. Results showed that Itga5 and Itgb1 were expressed in PECAM1-positive endothelial cells isolated from the anterior lobe (AL) of the rat pituitary, and integrin α5 was localized at the fenestrated capillaries of the rat pituitary and cultured PECAM1-positive endothelial cells isolated from AL (CECAL). Inhibition of both integrin α5ß1 and FAK, a key molecule for integrin-microtubule signaling, respectively, by ATN-161 and FAK inhibitor 14, caused the delocalization of PLVAP at the sieve plates and depolymerization of microtubules in CECAL. Paclitaxel prevented the delocalization of PLVAP by the inhibition of integrin α5ß1. Microtubule depolymerization induced by colcemid also caused the delocalization of PLVAP. Treatment of CECAL with ATN-161 and colcemid caused PLVAP localization at the Golgi apparatus. The localization of PLVAP at the sieve plates was inhibited by BFA treatment in a time-dependent manner and spread diffusely to the cytoplasm. These results indicate that a constant supply of PLVAP proteins by the endomembrane system via the Golgi apparatus is essential for the localization of PLVAP at sieve plates. In conclusion, the endomembrane transport pathway from the Golgi apparatus to sieve plates requires microtubule cytoskeletons, which are regulated by fibronectin-integrin α5ß1 signaling.

Fibronectin is essential for formation of fenestrae in endothelial cells of the fenestrated capillary.

Endothelial fenestrae are transcellular pores that pierce the capillary walls in endocrine glands such as the pituitary. The fenestrae are covered with a thin fibrous diaphragm consisting of the plasmalemma vesicle-associated protein (PLVAP) that clusters to form sieve plates. The basal surface of the vascular wall is lined by basement membrane (BM) composed of various extracellular matrices (ECMs). However, the relationship between the ECMs and the endothelial fenestrae is still unknown. In this study, we isolated fenestrated endothelial cells from the anterior lobe of the rat pituitary, using a dynabeads-labeled antibody against platelet endothelial cell adhesion molecule 1 (PECAM1). We then analyzed the gene expression levels of several endothelial marker genes and genes for integrin α subunits, which function as the receptors for ECMs, by real-time polymerase chain reaction (PCR). The results showed that the genes for the integrin α subunit, which binds to collagen IV, fibronectin, laminin-411, or laminin-511, were highly expressed. When the PECAM1-positive cells were cultured for 7 days on collagen IV-, fibronectin-, laminins-411-, or laminins-511-coated coverslips, the sieve plate structures equipped with probably functional fenestrae were maintained only when the cells were cultured on fibronectin. Additionally, real-time PCR analysis showed that the fibronectin coating was effective in maintaining the expression pattern of several endothelial marker genes that were preferentially expressed in the endothelial cells of the fenestrated capillaries. These results indicate that fibronectin functions as the principal factor in the maintenance of the sieve plate structures in the endothelial cells of the fenestrated capillary.

Why did we encounter gusher in a stapes surgery case? Was it enlarged medial aperture of the cochlear aqueduct?

BACKGROUND AND PURPOSE: Preoperative prediction of cerebrospinal fluid (CSF) gusher is important for stapes surgery. According to the current opinion settled among otologists and radiologists, the issues of whether enlarged cochlear aqueduct might be a cause of CSF gusher in stapes surgery and which segment of the aqueduct should be taken into account to diagnose enlarged cochlear aqueduct in computerized tomography (CT) are controversial. The case we encountered led us to hypothesize that enlarged cochlear aqueduct might cause CSF gusher in stapes surgery and that shape and diameter of medial aperture of the cochlear aqueduct are important in this prediction. METHODS AND RESULTS: Enlarged medial aperture of the cochlear aqueduct with a shape differed from that of the other side was retrospectively diagnosed in thin-slice CT in a patient who had been undergone middle ear and stapes surgery for conductive hearing loss. This finding went unnoticed in preoperative CT. In the small fenestra stapedotomy operation, CSF gusher occurred through opening in the ill-defined, fixed and thickened stapes footplate. A piece of temporalis fascia and reshaped incus were appropriately placed which stopped the gusher. Re-evaluation of preoperatively taken CT showed that anterior-posterior and superior-inferior diameters of the medial aperture were 11.7 mm and 2.87 mm in CSF gusher side versus 2.95 mm and 1.88 mm on the other side, respectively. Its shape in gusher side differed from that of the other side. CONCLUSION: This report is the first to show video-documented CSF gusher in a patient with enlarged medial aperture of the cochlear aqueduct. It appears to be plausible to propose that these findings have to change the otologists' and radiologists' perspective to the cochlear aqueduct. It can be deduced that difference in shapes of the medial aperture in both sides might be an indicator of potential CSF gusher.

Dolichocebus gaimanensis Is Not a Stem Platyrrhine.

The lower molars of the early Miocene New World monkey Dolichocebus gaimanensis from Patagonia, considered by some to be a stem platyrrhine, are for all intents and purposes indistinguishable from the same teeth preserved in the type specimen of the middle Miocene Laventiana annectens from Colombia. Canine morphology further indicates that Dolichocebus is a cebine, closely related to living Saimiri. The phenetic and phyletic continuity of these extinct species, here classified as congeners, means they are the first fossil platyrrhines that link South America's southern and northern Miocene monkey faunas, possibly as vicariant communities or with ancestral-descendant species that evolved by phyletic evolution. These findings further refute the hypothesis that early Miocene platyrrhines are (1) monophyletic and (2) outside the so-called crown platyrrhine radiation. The connection between these species is additional evidence supporting the hypothesis that the pattern of platyrrhine evolution is notably distinct for the predominance of low-level taxonomic clades (genera, tribes, subfamilies) of prolonged ecophylogenetic stability, which is the essence of the Long-Lineage Hypothesis.

Glomerular endothelial cell maturation depends on ADAM10, a key regulator of Notch signaling.

The principal function of glomeruli is to filter blood through a highly specialized filtration barrier consisting of a fenestrated endothelium, the glomerular basement membrane and podocyte foot processes. Previous studies have uncovered a crucial role of endothelial a disintegrin and metalloprotease 10 (ADAM10) and Notch signaling in the development of glomeruli, yet the resulting defects have not been further characterized nor understood in the context of kidney development. Here, we used several different experimental approaches to analyze the kidneys and glomeruli from mice lacking ADAM10 in endothelial cells (A10ΔEC mice). Scanning electron microscopy of glomerular casts demonstrated enlarged vascular diameter and increased intussusceptive events in A10ΔEC glomeruli compared to controls. Consistent with these findings, genes known to regulate vessel caliber (Apln, AplnR and Vegfr3) are significantly upregulated in A10ΔEC glomeruli. Moreover, transmission electron microscopy revealed the persistence of diaphragms in the fenestrae of A10ΔEC glomerular endothelial cells, which was corroborated by the elevated expression of the protein PLVAP/PV-1, an integral component of fenestral diaphragms. Analysis of gross renal vasculature by light sheet microscopy showed no major alteration of the branching pattern, indicating a localized importance of ADAM10 in the glomerular endothelium. Since intussusceptions and fenestrae with diaphragms are normally found in developing, but not mature glomeruli, our results provide the first evidence for a crucial role of endothelial ADAM10, a key regulator of Notch signaling, in promoting the development and maturation of the glomerular vasculature.

The Crista Fenestra and Its Impact on the Surgical Approach to the Scala Tympani during Cochlear Implantation.

OBJECTIVE: The aim of this work was to describe the dimensions of the crista fenestra and determine its presence by means of high-resolution computed tomography (CT) for the purpose of cochlear implantation via the round window approach. METHODS: A series of 10 adult human temporal bones underwent high-resolution CT scanning and were further dissected for microscopic study of the round window niche. RESULTS: In all of the specimens, the round window membrane was fully visualized after the complete removal of bony overhangs. The crista fenestra was identified as a sharp bony crest located in the anterior and inferior borders of the niche; its area ranged from 0.28 to 0.80 mm2 (mean 0.51 ± 0.18). The proportion of the area occupied by the crista fenestra in the whole circumference of the round window ranged from 23 to 50% (mean 36%). We found a moderate positive correlation between the area of the niche and the dimensions of the crista fenestra (Spearman rho: 0.491). In every case, high-resolution CT scanning was unable to determine the presence of the crista fenestra. CONCLUSION: The crista fenestra occupies a variable but expressive area within the bony round window niche. Narrower round window niches tended to house smaller crests. The presence of the crista fenestra is an important obstacle to adequate access to the scala tympani. Nevertheless, a high-resolution CT scan provides no additional preoperative information with regard to its presence for the purpose of surgical access to the scala tympani via the round window niche.

A Study of Round Window and its Adjacent Anatomy to Guide the Cochlear Implant Electrode Insertion.

Introduction: Residual hearing preservation has gained attention now which has brought round window membrane into the light, as a port for cochlear implantation. Atraumatic insertion of electrodes can be achieved by study of anatomical variations of round window and its forms which can guide the surgeon. Objective: This study was undertaken to examine the anatomical variations of round window and its adjacent structures and their impact on surgical approach during cochlear implantation. Methods: A series of 40 adult human temporal bones underwent high-resolution CT scanning and were further dissected for microscopic study of the round window. Results: The antero posterior dimensions of RW ranged from 1.22 to 2.51 mm on radiology and on dissection 1.76 mm +/- 0.3 mm. Shape of round window in 72.5% of bones was oval, and in 27.5% bones it was round shaped. As per Saint Thomas hospital classification for Round window visualization we found 82.5% bones had type I RW visualization and 17.5% had type IIa RW visualization. Area of crista fenestra on dissection was ranging from 0.41 to 0.69 mm2. Conclusion: Residual hearing preservation has become a new motto for surgeons. Therefore thorough anatomic knowledge of round window is must for careful insertion, as round window is closely related to the sensitive inner ear structures.

Cyclosporine Induces Fenestra-Associated Injury in Human Renal Microvessels In Vitro.

The use of cyclosporine A (CsA) in transplantation is frequently associated with nephrotoxicity, characterized by renal vascular injury, thrombotic microangiopathy, and striped interstitial fibrosis. Here, using human kidney-specific microvascular endothelial cells (HKMECs), we showed that CsA inhibited NFAT1 activation and impaired VEGF signaling in these ECs in a dose- and time-dependent manner. Integrated genome regulatory analyses identified key distinctions in the landscapes of HKMECs compared to human umbilical vein endothelial cells, particularly around genes related to the formation and maintenance of fenestrae. Using a bioengineered flow-directed 3D kidney microphysiological system, we revealed that CsA-induced kidney microvascular injury was associated with fenestrae and cell adhesion impairment, membrane swelling, and erythrocyte adhesion and extravasation into the interstitial space. Our data provide novel insights into kidney-specific molecular and structural mechanisms of CsA-induced microvascular injury. Our results also suggest VEGF-related pathways as potential targets for therapy during CsA treatment and emphasize the importance of leveraging species and organ-specific cells to better reflect human pathophysiology and the response to injury.

The wHole Story About Fenestrations in LSEC.

The porosity of liver sinusoidal endothelial cells (LSEC) ensures bidirectional passive transport of lipoproteins, drugs and solutes between the liver capillaries and the liver parenchyma. This porosity is realized via fenestrations - transcellular pores with diameters in the range of 50-300 nm - typically grouped together in sieve plates. Aging and several liver disorders severely reduce LSEC porosity, decreasing their filtration properties. Over the years, a variety of drugs, stimulants, and toxins have been investigated in the context of altered diameter or frequency of fenestrations. In fact, any change in the porosity, connected with the change in number and/or size of fenestrations is reflected in the overall liver-vascular system crosstalk. Recently, several commonly used medicines have been proposed to have a beneficial effect on LSEC re-fenestration in aging. These findings may be important for the aging populations of the world. In this review we collate the literature on medicines, recreational drugs, hormones and laboratory tools (including toxins) where the effect LSEC morphology was quantitatively analyzed. Moreover, different experimental models of liver pathology are discussed in the context of fenestrations. The second part of this review covers the cellular mechanisms of action to enable physicians and researchers to predict the effect of newly developed drugs on LSEC porosity. To achieve this, we discuss four existing hypotheses of regulation of fenestrations. Finally, we provide a summary of the cellular mechanisms which are demonstrated to tune the porosity of LSEC.

Notes on the cheek region of the Late Jurassic theropod dinosaur Allosaurus.

Allosaurus, from the Late Jurassic of North America and Europe, is a model taxon for Jurassic basal tetanuran theropod dinosaurs. It has achieved an almost iconic status due to its early discovery in the late, 19th century, and due to the abundance of material from the Morrison Formation of the western U.S.A., making Allosaurus one of the best-known theropod taxa. Despite this, various aspects of the cranial anatomy of Allosaurus are surprisingly poorly understood. Here, we discuss the osteology of the cheek region, comprised by the jugal, maxilla, and lacrimal. This region of the skull is of importance for Allosaurus taxonomy and phylogeny, particularly because Allosaurus has traditionally been reconstructed with an unusual cheek configuration, and because the European species Allosaurus europaeus has been said to be different from North American material in the configuration of these bones. Based on re-examination of articulated and disarticulated material from a number of repositories, we show that the jugal participates in the antorbital fenestra, contradicting the common interpretation. The jugal laterally overlies the lacrimal, and forms an extended antorbital fossa with this bone. Furthermore, we document previously unrecorded pneumatic features of the jugal of Allosaurus.

Intraperitoneal injections as an alternative method for micro-CT contrast enhanced detection of murine liver tumors.

Micro-computed tomography (micro-CT) coupled with tissue, or vascular, specific contrast agent has emerged as a powerful tool for detecting and monitoring tumor growth in the liver of murine animals. Intravenous injections of contrast agents can be technically challenging and lead to errors that can considerably influence the outcome of a preclinical study, prompting an alternative method. Here we assessed the effectiveness of intraperitoneal injections of polyiodinated triglycerides emulsions (Fenestra LC) in micro-CT imaging of young SCID (8 weeks) and old BALB/c (48 weeks) mice with xenograft or carcinogen-induced liver tumors, respectively, and determined an optimal acquisition time. Utilizing an intraperitoneal injection is a viable alternative administration route for using Fenestra in detection and quantification of murine liver tumor burden.

Two new species in the genus Andronymus Holland, 1896 (Lepidoptera, Hesperiidae).

During extensive field work in West Africa (Cameroon, Ivory Coast, Liberia, Sierra Leone), the authors collected two skipper species in the genus Andronymus (Lepidoptera, Hesperiidae), which would not fit the descriptions of any existing taxa. Both are described as new, A. magma sp. nov. is known only from Cameroon, while A. fenestra sp. nov. was found in a few localities in the Liberian sub-region of West Africa.

New insights into chasmosaurine (Dinosauria: Ceratopsidae) skulls from the Upper Cretaceous (Campanian) of Alberta, and an update on the distribution of accessory frill fenestrae in Chasmosaurinae.

Chasmosaurine ceratopsids are well documented from the Upper Cretaceous (Campanian) Dinosaur Park Formation (DPF) of southern Alberta and Saskatchewan, and include Chasmosaurus belli, Chasmosaurus russelli, Mercuriceratops gemini, Vagaceratops irvinensis, and material possibly referable to Spiclypeus shipporum. In this study, we describe three recently prepared chasmosaurine skulls (CMN 8802, CMN 34829, and TMP 2011.053.0046) from the DPF, and age-equivalent sediments, of Alberta. CMN 8802 and CMN 34829 are both referred to Chasmosaurus sp. based on the size and shape of the preserved parietal fenestrae. TMP 2011.053.0046 is referred to Vagaceratops sp. based on the position and orientation of its preserved epiparietals. Each skull is characterized by the presence of an accessory fenestra in either the squamosal (CMN 8802 and TMP 2011.053.0046) or parietal (CMN 34829). Such fenestrae are common occurrences in chasmosaurine squamosals, but are rare in the parietal portion of the frill. The origin of the fenestrae in these three specimens is unknown, but they do not appear to exhibit evidence of pathology, as has been previously interpreted for the accessory fenestrae in most other chasmosaurine frills. These three skulls contribute to a better understanding of the morphological variation, and geographic and stratigraphic distribution, of chasmosaurines within the DPF and age-equivalent sediments in Western Canada.

Redescrição e osteologia de Bryconamericus exodon Eigenmann, 1907 (Ostariophysi, Characiformes, Characidae) / Redescription and osteology of Bryconamericus exodon Eigenmann, 1907 (Ostariophysi, Characiformes, Characidae)

Apresenta-se a redescrição de Bryconamericus exodon Eigenmann (1907), espécie-tipo do gênero, uma descrição osteológica da espécie, uma comparação com outras espécies de Characiformes e comentários sobre seu relacionamento com outros caracídeos. Bryconamericus exodon é facilmente distinta de todas as demais espécies do gênero por possuir os lobos superior e inferior da nadadeira caudal castanho-escuros; aproxima-se das espécies de Bryconamericus com corpo alongado e baixo, menos de 30 por cento do comprimento padrão, boca terminal e dentes da série externa do pré-maxilar desalinhados. Compartilha com outros Characidae um palatino fenestrado, perfurado ou adelgaçado, em sua porção médio-anterior, e um pós-cleitro 3 alongado e com aproximadamente a mesma largura em toda sua extensão, sinapomorfias adicionais para os Characidae com ii, 8 raios na nadadeira dorsal e quatro dentes na série interna do pré-maxilar.

The redescription of B. exodon Eigenmann (1907), type species of Bryconamericus, an osteological description for the species, a comparison with other characiform fishes, and comments about its relationships among other characids are presented. Bryconamericus exodon is easily distinguished from other species in the genus by possessing upper and lower caudal-fin lobes dark brown; it is proximate to other Bryconamericus species that possess: an elongated and low body, its depth less than 30 percent of standard length, mouth terminal, and teeth of external premaxillary series not aligned. Furthermore it shares with other Characidae a palatine fenestrate, perforated or shallowed in its medial, anterior, portion, and a third postcleithrum elongated and equally wide along its entire length, character states proposed as additional synapomorphies for the characids with ii, 8 dorsal-fin rays and four teeth in the internal premaxillary series.