Free-water diffusion tensor imaging detects occult periependymal abnormality in the AQP4-IgG-seropositive neuromyelitis optica spectrum disorder.

To compare free-water corrected diffusion tensor imaging (DTI) measures in the normal-appearing periependymal area between AQP4-IgG-seropositive NMOSD and multiple sclerosis (MS) to investigate occult pathophysiology. This prospective study included 44 patients (mean age, 39.52 ± 11.90 years; 14 men) with AQP4-IgG-seropositive NMOSD (n = 20) and MS (n = 24) who underwent DTI between April 2014 and April 2020. Based on free-water corrected DTI measures obtained from normal-appearing periependymal voxels of (1) lateral ventricles and (2) the 3rd and 4th ventricles as dependent variables, MANCOVA was conducted to compare the two groups, using clinical variables as covariates. A significant difference was found between AQP4-IgG-seropositive NMOSD and MS in the 3rd and 4th periependymal voxels (λ = 0.462, P = 0.001). Fractional anisotropy, axial diffusivity was significantly decreased and radial diffusivity was increased in AQP4-IgG-seropositive NMOSD in post-hoc analysis, compared with MS (F = 27.616, P < 0.001, F = 7.336, P = 0.011, and F = 5.800, P = 0.022, respectively). Free-water corrected DTI measures differ in the periependymal area surrounding the diencephalon and brain stem/cerebellum between MS and NMOSD, which may suggest occult white matter injury in areas with distribution of AQP-4 in NMOSD.

Tomografía computada y resonancia magnética de variantes normales/congénitas de apariencia quística y presentación frecuente en el encéfalo / CT and MRI of Normal/Congenital Variables with Cystic Appearance and Frequent Presence in the Brain

La presentación de una imagen de apariencia quística durante el estudio del encéfalo constituye un hallazgo incidental cada vez más frecuente, pudiendo encontrarse en el espacio extra o intraaxial. Las mismas pueden ser de naturaleza congénita o adquirida, benigna o maligna, ocupantes de espacio con desplazamiento de la línea media o simplemente presentarse sin efecto compresivo alguno. De localización supra o infratentorial, esas imágenes constituyen un desafío diagnóstico, siendo imprescindible su reconocimiento para no solicitar estudios o tratamientos innecesarios. Valoraremos las imágenes de apariencia quística más frecuentes empleando tomografía computada o imágenes de resonancia magnética.

Presence of a cystic formation in brain examination is frequently an incidental finding. They can be intra or extra-axial in location, congenital or acquired, benign or malignant with or without mass effect. Intracranial cysts can be a diagnostic challenge and we should know them to avoid unnecessary exams or treatment. We will analyze the most common cystic formations seen in computed tomography and magnetic resonance.

Mutations in Kinesin family member 6 reveal specific role in ependymal cell ciliogenesis and human neurological development.

Cerebrospinal fluid flow is crucial for neurodevelopment and homeostasis of the ventricular system of the brain, with localized flow being established by the polarized beating of the ependymal cell (EC) cilia. Here, we report a homozygous one base-pair deletion, c.1193delT (p.Leu398Glnfs*2), in the Kinesin Family Member 6 (KIF6) gene in a child displaying neurodevelopmental defects and intellectual disability. To test the pathogenicity of this novel human KIF6 mutation we engineered an analogous C-terminal truncating mutation in mouse. These mutant mice display severe, postnatal-onset hydrocephalus. We generated a Kif6-LacZ transgenic mouse strain and report expression specifically and uniquely within the ependymal cells (ECs) of the brain, without labeling other multiciliated mouse tissues. Analysis of Kif6 mutant mice with scanning electron microscopy (SEM) and immunofluorescence (IF) revealed specific defects in the formation of EC cilia, without obvious effect of cilia of other multiciliated tissues. Dilation of the ventricular system and defects in the formation of EC cilia were also observed in adult kif6 mutant zebrafish. Finally, we report Kif6-GFP localization at the axoneme and basal bodies of multi-ciliated cells (MCCs) of the mucociliary Xenopus epidermis. Overall, this work describes the first clinically-defined KIF6 homozygous null mutation in human and defines KIF6 as a conserved mediator of neurological development with a specific role for EC ciliogenesis in vertebrates.

Hydrocephalus due to multiple ependymal malformations is caused by mutations in the MPDZ gene.

Congenital hydrocephalus is considered as either acquired due to haemorrhage, infection or neoplasia or as of developmental nature and is divided into two subgroups, communicating and obstructive. Congenital hydrocephalus is either syndromic or non-syndromic, and in the latter no cause is found in more than half of the patients. In patients with isolated hydrocephalus, L1CAM mutations represent the most common aetiology. More recently, a founder mutation has also been reported in the MPDZ gene in foetuses presenting massive hydrocephalus, but the neuropathology remains unknown. We describe here three novel homozygous null mutations in the MPDZ gene in foetuses whose post-mortem examination has revealed a homogeneous phenotype characterized by multiple ependymal malformations along the aqueduct of Sylvius, the third and fourth ventricles as well as the central canal of the medulla, consisting in multifocal rosettes with immature cell accumulation in the vicinity of ependymal lining early detached from the ventricular zone. MPDZ also named MUPP1 is an essential component of tight junctions which are expressed from early brain development in the choroid plexuses and ependyma. Alterations in the formation of tight junctions within the ependyma very likely account for the lesions observed and highlight for the first time that primary multifocal ependymal malformations of the ventricular system is genetically determined in humans. Therefore, MPDZ sequencing should be performed when neuropathological examination reveals multifocal ependymal rosette formation within the aqueduct of Sylvius, of the third and fourth ventricles and of the central canal of the medulla.

Zebrafish models of idiopathic scoliosis link cerebrospinal fluid flow defects to spine curvature.

Idiopathic scoliosis (IS) affects 3% of children worldwide, yet the mechanisms underlying this spinal deformity remain unknown. Here we show that ptk7 mutant zebrafish, a faithful developmental model of IS, exhibit defects in ependymal cell cilia development and cerebrospinal fluid (CSF) flow. Transgenic reintroduction of Ptk7 in motile ciliated lineages prevents scoliosis in ptk7 mutants, and mutation of multiple independent cilia motility genes yields IS phenotypes. We define a finite developmental window for motile cilia in zebrafish spine morphogenesis. Notably, restoration of cilia motility after the onset of scoliosis blocks spinal curve progression. Together, our results indicate a critical role for cilia-driven CSF flow in spine development, implicate irregularities in CSF flow as an underlying biological cause of IS, and suggest that noninvasive therapeutic intervention may prevent severe scoliosis.

Cortical hypoplasia and ventriculomegaly of p73-deficient mice: Developmental and adult analysis.

Trp73, a member of the p53 gene family, plays a crucial role in neural development. We describe two main phenotypic variants of p73 deficiency in the brain, a severe one characterized by massive apoptosis in the cortex leading to early postnatal death and a milder, non-/low-apoptosis one in which 50% of pups may reach adulthood using an intensive-care breeding protocol. Both variants display the core triad of p73 deficiency: cortical hypoplasia, hippocampal malformations, and ventriculomegaly. We studied the development of the neocortex in p73 KO mice from early embryonic life into advanced age (25 months). Already at E14.5, the incipient cortical plate of the p73 KO brains showed a reduced width. Examination of adult neocortex revealed a generalized, nonprogressive reduction by 10-20%. Area-specific architectonic landmarks and lamination were preserved in all cortical areas. The surviving adult animals had moderate ventricular distension, whereas pups of the early lethal phenotypic variant showed severe ventriculomegaly. Ependymal cells of wild-type ventricles strongly express p73 and are particularly vulnerable to p73 deficiency. Ependymal denudation by apoptosis and reduction of ependymal cilia were already evident in young mice, with complete absence of cilia in older animals. Loss of p73 function in the ependyma may thus be one determining factor for chronic hydrocephalus, which leads to atrophy of subcortical structures (striatum, septum, amygdala). p73 Is thus involved in a variety of CNS activities ranging from embryonic regulation of brain size to the control of cerebrospinal fluid homeostasis in the adult brain via maintenance of the ependyma.

Conditional N-WASP knockout in mouse brain implicates actin cytoskeleton regulation in hydrocephalus pathology.

Cerebrospinal fluid (CSF) is produced by the choroid plexus and moved by multi-ciliated ependymal cells through the ventricular system of the vertebrate brain. Defects in the ependymal layer functionality are a common cause of hydrocephalus. N-WASP (Neural-Wiskott Aldrich Syndrome Protein) is a brain-enriched regulator of actin cytoskeleton and N-WASP knockout caused embryonic lethality in mice with neural tube and cardiac abnormalities. To shed light on the role of N-WASP in mouse brain development, we generated N-WASP conditional knockout mouse model N-WASP(fl/fl); Nestin-Cre (NKO-Nes). NKO-Nes mice were born with Mendelian ratios but exhibited reduced growth characteristics compared to their littermates containing functional N-WASP alleles. Importantly, all NKO-Nes mice developed cranial deformities due to excessive CSF accumulation and did not survive past weaning. Coronal brain sections of these animals revealed dilated lateral ventricles, defects in ciliogenesis, loss of ependymal layer integrity, reduced thickness of cerebral cortex and aqueductal stenosis. Immunostaining for N-cadherin suggests that ependymal integrity in NKO-Nes mice is lost as compared to normal morphology in the wild-type controls. Moreover, scanning electron microscopy and immunofluorescence analyses of coronal brain sections with anti-acetylated tubulin antibodies revealed the absence of cilia in ventricular walls of NKO-Nes mice indicative of ciliogenesis defects. N-WASP deficiency does not lead to altered expression of N-WASP regulatory proteins, Fyn and Cdc42, which have been previously implicated in hydrocephalus pathology. Taken together, our results suggest that N-WASP plays a critical role in normal brain development and implicate actin cytoskeleton regulation as a vulnerable axis frequently deregulated in hydrocephalus.

Platelet-derived growth factor C deficiency in C57BL/6 mice leads to abnormal cerebral vascularization, loss of neuroependymal integrity, and ventricular abnormalities.

Platelet-derived growth factors (PDGFs) and their tyrosine kinase receptors (PDGFRs) are known to play important roles during development of the lungs, central nervous system (CNS), and skeleton and in several diseases. PDGF-C is a ligand for the tyrosine kinase receptor PDGFRα. Mutations in the gene encoding PDGF-C have been linked to clefts of the lip and/or palate in humans, and ablation of PDGF-C in 129/Sv background mice results in death during the perinatal period. In this study, we report that ablation of PDGF-C in C57BL/6 mice results in a milder phenotype than in 129/Sv mice, and we present a phenotypic characterization of PDGF-C deficiency in the adult murine CNS. Multiple congenital defects were observed in the CNS of PDGF-C-null C57BL/6 mice, including cerebral vascular abnormalities with abnormal vascular smooth muscle cell coverage. In vivo imaging of mice deficient in PDGF-C also revealed cerebral ventricular abnormalities, such as asymmetry of the lateral ventricles and hypoplasia of the septum, reminiscent of cavum septum pellucidum in humans. We further noted that PDGF-C-deficient mice displayed a distorted ependymal lining of the lateral ventricles, and we found evidence of misplaced neurons in the ventricular lining. We conclude that PDGF-C plays a critical role in the development of normal cerebral ventricles and neuroependymal integrity as well as in normal cerebral vascularization.

Ependymal alterations in sudden intrauterine unexplained death and sudden infant death syndrome: possible primary consequence of prenatal exposure to cigarette smoking.

BACKGROUND: The ependyma, the lining providing a protective barrier and filtration system separating brain parenchyma from cerebrospinal fluid, is still inadequately understood in humans. In this study we aimed to define, by morphological and immunohistochemical methods, the sequence of developmental steps of the human ependyma in the brainstem (ventricular ependyma) and thoracic spinal cord (central canal ependyma) of a large sample of fetal and infant death victims, aged from 17 gestational weeks to 8 postnatal months. Additionally, we investigated a possible link between alterations of this structure, sudden unexplained fetal and infant death and maternal smoking. RESULTS: Our results demonstrate that in early fetal life the human ependyma shows a pseudostratified cytoarchitecture including many tanycytes and ciliated cells together with numerous apoptotic and reactive astrocytes in the subependymal layer. The ependyma is fully differentiated, with a monolayer of uniform cells, after 32 to 34 gestational weeks. We observed a wide spectrum of ependymal pathological changes in sudden death victims, such as desquamation, clusters of ependymal cells in the subventricular zone, radial glial cells, and the unusual presence of neurons within and over the ependymal lining. These alterations were significantly related to maternal smoking in pregnancy. CONCLUSIONS: We conclude that in smoking mothers, nicotine and its derivatives easily reach the cerebrospinal fluid in the fetus, immediately causing ependymal damage. Consequently, we suggest that the ependyma should be examined in-depth first in victims of sudden fetal or infant death with mothers who smoke.

Mutations in Hydin impair ciliary motility in mice.

Chlamydomonas reinhardtii hydin is a central pair protein required for flagellar motility, and mice with Hydin defects develop lethal hydrocephalus. To determine if defects in Hydin cause hydrocephalus through a mechanism involving cilia, we compared the morphology, ultrastructure, and activity of cilia in wild-type and hydin mutant mice strains. The length and density of cilia in the brains of mutant animals is normal. The ciliary axoneme is normal with respect to the 9 + 2 microtubules, dynein arms, and radial spokes but one of the two central microtubules lacks a specific projection. The hydin mutant cilia are unable to bend normally, ciliary beat frequency is reduced, and the cilia tend to stall. As a result, these cilia are incapable of generating fluid flow. Similar defects are observed for cilia in trachea. We conclude that hydrocephalus in hydin mutants is caused by a central pair defect impairing ciliary motility and fluid transport in the brain.

Patterned neuropathologic events occurring in hyh congenital hydrocephalic mutant mice.

Hyh mutant mice develop long-lasting hydrocephalus and represent a good model for investigating neuropathologic events associated with hydrocephalus. The study of their brains by use of lectin binding, bromodeoxyuridine labeling, immunochemistry, and scanning electron microscopy revealed that certain events related to hydrocephalus followed a well-defined pattern. A program of neuroepithelium/ependyma denudation was initiated at embryonic day 12 and terminated at the end of the second postnatal week. After the third postnatal week the denuded areas remained permanently devoid of ependyma. In contrast, a selective group of ependymal areas resisted denudation throughout the lifespan. Ependymal denudation triggered neighboring astrocytes to proliferate. These astrocytes expressed particular glial markers and formed a superficial cell layer replacing the lost ependyma. The loss of the neuroepithelium/ependyma layer at specific regions of the ventricular walls and at specific stages of brain development would explain the fact that only certain brain structures had abnormal development. Therefore, commissural axons forming the corpus callosum and the hippocampal commissure displayed abnormalities, whereas those forming the anterior and posterior commissures did not; and the brain cortex was not homogenously affected, with the cingular and frontal cortices being the most altered regions. All of these telencephalic alterations developed at stages when hydrocephalus was not yet patent at the lateral ventricles, indicating that abnormal neural development and hydrocephalus are linked at the etiologic level, rather than the former being a consequence of the latter. All evidence collected on hydrocephalic hyh mutant mice indicates that a primary alteration in the neuroepithelium/ependyma cell lineage triggers both hydrocephalus and abnormalities in telencephalic development.

Biosynthesis of Wdr16, a marker protein for kinocilia-bearing cells, starts at the time of kinocilia formation in rat, and wdr16 gene knockdown causes hydrocephalus in zebrafish.

The rat ortholog of the WD40 repeat protein Wdr16 is abundantly expressed in testis and cultured ependymal cells. Low levels are found in lung and brain, respectively, while it is absent from kinocilia-free tissues. In testis and ependymal primary cultures, Wdr16 messenger RNA appears concomitantly with the messages for sperm-associated antigen 6, a kinocilia marker, and for hydin, a protein linked to ciliary function and hydrocephalus. In testis, ependyma and respiratory epithelium, the Wdr16 protein is up-regulated together with kinocilia formation. The wdr16 gene is restricted to genera in possession of kinocilia, and it is strongly conserved during evolution. The human and zebrafish proteins are identical in 62% of their aligned amino acids. On the message level, the zebrafish Wdr16 ortholog was found exclusively in kinocilia-bearing tissues by in situ hybridisation. Gene knockdown in zebrafish embryos by antisense morpholino injection resulted in severe hydrocephalus formation with unaltered ependymal morphology or ciliary beat. Wdr16 can be considered a differentiation marker of kinocilia-bearing cells. In the brain, it appears to be functionally related to water homeostasis or osmoregulation.

High levels of Cre expression in neuronal progenitors cause defects in brain development leading to microencephaly and hydrocephaly.

Hydrocephalus is a common and variegated pathology often emerging in newborn children after genotoxic insults during pregnancy (Hicks and D'Amato, 1980). Cre recombinase is known to have possible toxic effects that can compromise normal cell cycle and survival. Here we show, by using three independent nestin Cre transgenic lines, that high levels of Cre recombinase expression into the nucleus of neuronal progenitors can compromise normal brain development. The transgenics analyzed are the nestin Cre Balancer (Bal1) line, expressing the Cre recombinase with a nuclear localization signal, and two nestin CreER(T2) (Cre recombinase fused with a truncated estrogen receptor) mice lines with different levels of expression of a hybrid CreER(T2) recombinase that translocates into the nucleus after tamoxifen treatment. All homozygous Bal1 nestin Cre embryos displayed reduced neuronal proliferation, increased aneuploidy and cell death, as well as defects in ependymal lining and lamination of the cortex, leading to microencephaly and to a form of communicating hydrocephalus. An essentially overlapping phenotype was observed in the two nestin CreER(T2) transgenic lines after tamoxifen mediated-CreER(T2) translocation into the nucleus. Neither tamoxifen-treated wild-type nor nestin CreER(T2) oil-treated control mice displayed these defects. These results indicate that some forms of hydrocephalus may derive from a defect in neuronal precursors proliferation. Furthermore, they underscore the potential risks for developmental studies of high levels of nuclear Cre in neurogenic cells.

A deficiency in RFX3 causes hydrocephalus associated with abnormal differentiation of ependymal cells.

Ciliated ependymal cells play central functions in the control of cerebrospinal fluid homeostasis in the mammalian brain, and defects in their differentiation or ciliated properties can lead to hydrocephalus. Regulatory factor X (RFX) transcription factors regulate genes required for ciliogenesis in the nematode, drosophila and mammals. We show here that Rfx3-deficient mice suffer from hydrocephalus without stenosis of the aqueduct of Sylvius. RFX3 is expressed strongly in the ciliated ependymal cells of the subcommissural organ (SCO), choroid plexuses (CP) and ventricular walls during embryonic and postnatal development. Ultrastructural analysis revealed that the hydrocephalus is associated with a general defect in CP differentiation and with severe agenesis of the SCO. The specialized ependymal cells of the CP show an altered epithelial organization, and the SCO cells lose their characteristic ultrastructural features and adopt aspects more typical of classical ependymal cells. These differentiation defects are associated with changes in the number of cilia, although no obvious ultrastructural defects of these cilia can be observed in adult mice. Moreover, agenesis of the SCO is associated with downregulation of SCO-spondin expression as early as E14.5 of embryonic development. These results demonstrate that RFX3 is necessary for ciliated ependymal cell differentiation in the mouse.

Malformations of cortical development: high-resolution MR and diffusion tensor imaging of fiber tracts at 3T.

Patients with malformations of cortical development and epilepsy may have a variety of abnormal brain findings, including abnormal gyral patterns, cortical thickening, decreased volume of white matter, and increased diffusion of white matter. The status of individual white matter fiber tracts, however, is unknown. We present a case of bilateral frontal schizencephaly and subcortical heterotopia and illustrate alterations of white matter fascicles by combined structural and functional diffusion tensor imaging at 3 T.

RNA-binding protein Musashi family: roles for CNS stem cells and a subpopulation of ependymal cells revealed by targeted disruption and antisense ablation.

Homologues of the Musashi family of RNA-binding proteins are evolutionarily conserved across species. In mammals, two members of this family, Musashi1 (Msi1) and Musashi2 (Msi2), are strongly coexpressed in neural precursor cells, including CNS stem cells. To address the in vivo roles of msi in neural development, we generated mice with a targeted disruption of the gene encoding Msi1. Homozygous newborn mice frequently developed obstructive hydrocephalus with aberrant proliferation of ependymal cells in a restricted area surrounding the Sylvius aqueduct. These observations indicate a vital role for msi1 in the normal development of this subpopulation of ependymal cells, which has been speculated to be a source of postnatal CNS stem cells. On the other hand, histological examination and an in vitro neurosphere assay showed that neither the embryonic CNS development nor the self-renewal activity of CNS stem cells in embryonic forebrains appeared to be affected by the disruption of msi1, but the diversity of the cell types produced by the stem cells was moderately reduced by the msi1 deficiency. Therefore, we performed antisense ablation experiments to target both msi1 and msi2 in embryonic neural precursor cells. Administration of the antisense peptide-nucleotides, which were designed to specifically down-regulate msi2 expression, to msi1(-/-) CNS stem cell cultures drastically suppressed the formation of neurospheres in a dose-dependent manner. Antisense-treated msi1(-/-) CNS stem cells showed a reduced proliferative activity. These data suggest that msi1 and msi2 are cooperatively involved in the proliferation and maintenance of CNS stem cell populations.

Analysis of ependymal abnormalities in subjects with schizophrenia, bipolar disorder, and depression.

Abnormalities of the ependyma can serve as a marker of early brain insults. The presence of ependymal abnormalities was determined in sections containing ependyma of the temporal horn obtained from the Stanley Neuropathology Consortium: 15 subjects with schizophrenia, 15 with bipolar illness, 15 with major depression and 15 normal controls. There were no significant differences in numbers of subjects with ependymal discontinuities or subventricular rosettes. Subjects with schizophrenia had significantly less nodular gliosis than normal subjects (p=0.02); there was a trend for subjects with depression to have less nodular gliosis than normal subjects (p=0.06). Control subjects had unexpectedly high rates of ependymal abnormalities, indicating that ependymal abnormalities may not be a useful marker of pre- or perinatal events associated with schizophrenia and other psychiatric disorders in adult postmortem tissue.

Gelastic seizure with tectal tumor, lobar holoprosencephaly, and subependymal nodules: clinical report.

Gelastic seizures are characterized by inappropriate, stereotyped laughter and are often first recognized when other epileptic manifestations occur. They are frequently associated with hypothalamic hamartomas. Central nervous system developmental abnormalities are rarely reported with gelastic seizures. There is only one case report of gelastic seizure caused by holoprosencephaly. We report a 2-year-old girl with multiple brain structural abnormalities including tectal tumor (possibly hamartoma), multiple subependymal nodules, and holoprosencephaly. She developed seizures during the newborn period and presented with gelastic seizure and simple partial seizure at 3 months of age.