Ependymal Cell Differentiation Research Articles

Neurogenesis and growth factors expression after complete spinal cord transection in Pleurodeles waltlii.

Following spinal lesion, connections between the supra-spinal centers and spinal neuronal networks can be disturbed, which causes the deterioration or even the complete absence of sublesional locomotor activity. In mammals, possibilities of locomotion restoration are much reduced since descending tracts either have very poor regenerative ability or do not regenerate at all. However, in lower vertebrates, there is spontaneous locomotion recuperation after complete spinal cord transection at the mid-trunk level. This phenomenon depends on a translesional descending axon re-growth originating from the brainstem. On the other hand, cellular and molecular mechanisms underlying spinal cord regeneration and in parallel, locomotion restoration of the animal, are not well known. Fibroblast growth factor 2 (FGF-2) plays an important role in different processes such as neural induction, neuronal progenitor proliferation and their differentiation. Studies had shown an over expression of this growth factor after tail amputation. Nestin, a protein specific for intermediate filaments, is considered an early marker for neuronal precursors. It has been recently shown that its expression increases after tail transection in urodeles. Using this marker and western blots, our results show that the number of FGF-2 and FGFR2 mRNAs increases and is correlated with an increase in neurogenesis especially in the central canal lining cells immediately after lesion. This study also confirms that spinal cord re-growth through the lesion site initially follows a rostrocaudal direction. In addition to its role known in neuronal differentiation, FGF-2 could be implicated in the differentiation of ependymal cells into neuronal progenitors.

Ependymal cell differentiation, from monociliated to multiciliated cells.

Primary and motile cilia differ in their structure, composition, and function. In the brain, primary cilia are immotile signalling organelles present on neural stem cells and neurons. Multiple motile cilia are found on the surface of ependymal cells in all brain ventricles, where they contribute to the flow of cerebrospinal fluid. During development, monociliated ependymal progenitor cells differentiate into multiciliated ependymal cells, thus providing a simple system for studying the transition between these two stages. In this chapter, we provide protocols for immunofluorescence staining of developing ependymal cells in vivo, on whole mounts of lateral ventricle walls, and in vitro, on cultured ependymal cells. We also provide a list of markers we currently use to stain both types of cilia, including proteins at the ciliary membrane and tubulin posttranslational modifications of the axoneme.

Rax regulates hypothalamic tanycyte differentiation and barrier function in mice

The wall of the ventral third ventricle is composed of two distinct cell populations: tanycytes and ependymal cells. Tanycytes regulate many aspects of hypothalamic physiology, but little is known about the transcriptional network that regulates their development and function. We observed that the retina and anterior neural fold homeobox transcription factor (Rax) is selectively expressed in hypothalamic tanycytes, and showed a complementary pattern of expression to markers of hypothalamic ependymal cells, such as Rarres2 (retinoic acid receptor responder [tazarotene induced] 2). To determine whether Rax controls tanycyte differentiation and function, we generated Rax haploinsufficient mice and examined their cellular and molecular phenotype in adulthood. These mice appeared grossly normal, but careful examination revealed a thinning of the third ventricular wall and reduction of both tanycyte and ependymal markers. These experiments show that Rax is required for hypothalamic tanycyte and ependymal cell differentiation. Rax haploinsufficiency also resulted in the ectopic presence of ependymal cells in the α2 tanycytic zone, where few ependymal cells are normally found, suggesting that Rax is selectively required for α2 tanycyte differentiation. These changes in the ventricular wall were associated with reduced diffusion of Evans Blue tracer from the ventricle to the hypothalamic parenchyma, with no apparent repercussion on the gross anatomical or behavioral phenotype of these mice. In conclusion, we have provided evidence that Rax is required for the normal differentiation and patterning of hypothalamic tanycytes and ependymal cells, as well as for maintenance of the cerebrospinal fluid-hypothalamus barrier.

Inactivation of Cdc42 in embryonic brain results in hydrocephalus with ependymal cell defects in mice

The establishment of a polarized cellular morphology is essential for a variety of processes including neural tube morphogenesis and the development of the brain. Cdc42 is a Ras-related GTPase that plays an essential role in controlling cell polarity through the regulation of the actin and microtubule cytoskeleton architecture. Previous studies have shown that Cdc42 plays an indispensable role in telencephalon development in earlier embryo developmental stage (before E12.5). However, the functions of Cdc42 in other parts of brain in later embryo developmental stage or in adult brain remain unclear. Thus, in order to address the role of Cdc42 in the whole brain in later embryo developmental stage or in adulthood, we used Cre/loxP technology to generate two lines of tissue-specific Cdc42-knock-out mice. Inactivation of Cdc42 was achieved in neuroepithelial cells by crossing Cdc42/ flox mice with Nestin-Cre mice and resulted in hydrocephalus, causing death to occur within the postnatal stage. Histological analyses of the brains from these mice showed that ependymal cell differentiation was disrupted, resulting in aqueductal stenosis. Deletion of Cdc42 in the cerebral cortex also induced obvious defects in interkinetic nuclear migration and hypoplasia. To further explore the role of Cdc42 in adult mice brain, we examined the effects of knocking-out Cdc42 in radial glial cells by crossing Cdc42/flox mice with human glial fibrillary acidic protein (GFAP)-Cre mice. Inactivation of Cdc42 in radial glial cells resulted in hydrocephalus and ependymal cell denudation. Taken together, these results highlight the importance of Cdc42 for ependymal cell differentiation and maintaining, and suggest that these functions likely contribute to the essential roles played by Cdc42 in the development of the brain.

FM19G11 Favors Spinal Cord Injury Regeneration and Stem Cell Self-Renewal by Mitochondrial Uncoupling and Glucose Metabolism Induction

Spinal cord injury is a major cause of paralysis with no currently effective therapies. Induction of self-renewal and proliferation of endogenous regenerative machinery with noninvasive and nontoxic therapies could constitute a real hope and an alternative to cell transplantation for spinal cord injury patients. We previously showed that FM19G11 promotes differentiation of adult spinal cord-derived ependymal stem cells under hypoxia. Interestingly, FM19G11 induces self-renewal of these ependymal stem cells grown under normoxia. The analysis of the mechanism of action revealed an early increment of mitochondrial uncoupling protein 1 and 2 with an early drop of ATP, followed by a subsequent compensatory recovery with activated mitochondrial metabolism and the induction of glucose uptake by upregulation of the glucose transporter GLUT-4. Here we show that phosphorylation of AKT and AMP-activated kinase (AMPK) is involved in FM19G11-dependent activation of GLUT-4, glucose influx, and consequently in stem cell self-renewal. Small interfering RNA of uncoupling protein 1/2, GLUT-4 and pharmacological inhibitors of AKT, mTOR and AMPK signaling blocked the FM19G11-dependent induction of the self-renewal-related markers Sox2, Oct4, and Notch1. Importantly, FM19G11-treated animals showed accelerated locomotor recovery. In vivo intrathecal sustained administration of FM19G11 in rats after spinal cord injury showed more neurofilament TUJ1-positive fibers crossing the injured area surrounded by an increase of neural precursor Vimentin-positive cells. Overall, FM19G11 exerts an important influence on the self-renewal of ependymal stem progenitor cells with a plausible neuroprotective role, providing functional benefits for spinal cord injury treatment.

Chordoid glioma of the third ventricle: Four cases including one case with papillary features

Chordoid glioma is a rare, slowly growing tumor of the CNS, which is always located in the third ventricle of adults. Chordoid glioma has classic histological features consisting of clusters and cords of epithelioid tumor cells embedded within a mucinous stroma with rich lymphoplasmacytic infiltrate. The important distinctive immunohistochemical feature of this neoplasm is strong and diffuse reactivity for GFAP. Here, we report four cases of chordoid glioma that occupied the anterior portion of the third ventricle or suprasellar region. These four cases were all adult females with almost typical clinical, radiological, histologic and immunohistochemical characteristics of chordoid glioma. However, in one case there was an unusual histologic finding with regard to the papillary region. In this region, elongated tumor cells were observed radiating toward a central vessel to form characteristic papillary structures. Immunohistochemically, three cases showed strong reactivity for GFAP, and one exhibited weak reactivity. All cases were focally positive for epithelial membrane antigen, CD34 and D2-40, but negative for neurofilament protein (NFP). Several ultrastructural investigations have supported the ependymal origin of chordoid glioma. In some cases of immunoreactivity for NFP, some authors have supposed that chordoid glioma originates from a multipotential stem cell with glial and neuronal cell differentiation. With regard to the present four cases with immunoreactivity for D2-40 (an ependymal marker) and CD34 (undifferentiated neural precursors) and based on previously published data, we considered that the majority of chordoid gliomas had an ependymal origin, and that a small minority might have originated from a multipotential stem cell having ependymal and neuronal cell differentiation.

Increased Expression of Foxj1 after Traumatic Brain Injury

Foxj1 is a member of the Forkhead/winged-helix (Fox) family of transcription factors, which is required for postnatal differentiation of ependymal cells and a subset of astrocytes in the subventricular zone. The subpopulation of astrocytes has the ability of self-renew and neurogenic potential differentiated into astrocytes, oligodendrocytes, and neurons. However, its expression and function in the central nervous system lesion are not well understood. In this study, we performed a traumatic brain injury (TBI) model in adult rats and investigated the changed expression of Foxj1 in the brain cortex. Western blot and immunohistochemistry analysis showed that the expression of Foxj1 gradually increased, reached a peak at day 3 after TBI, and declined during the following days. Double immunofluorescence staining revealed that Foxj1 was co-expressed with MAP-2 and GFAP. In addition, we detected that Ki67 had the co-localization with NeuN, GFAP, and Foxj1. All our findings suggested that Foxj1 may be involved in the pathophysiology of brain after TBI.

Proliferation, migration and differentiation of ependymal region neural progenitor cells in the brainstem after hypoglossal nerve avulsion

Cells in the ependymal region in the adult central nervous system (CNS) have been found to possess neural progenitor cell (NPC) like features including capacity for generating new neurons and glia in response to injury and inflammatory disease. Whether these cells are activated after a peripheral nerve injury has not previously been extensively evaluated. We investigate the possible activation and effect of NPCs in the ependymal region in the immediate vicinity to the hypoglossal nucleus in the brainstem using two models of injuries, hypoglossal nerve transection and nerve avulsion after which the proliferation, migration and differentiation of ependymal regional NPCs were evaluated. We showed that: (i) immunoreactivity for Sox2 was detected in cells in the ependymal region of the brainstem and that BrdU/Sox2-positive cells were observed after avulsion, but not after transection injury; (ii) avulsion induces re-expression of nestin in the ependymal layer as well as induced NPC migration from the ependymal layer; (iii) the chemokine SDF-1α (a marker for migrating cells) was upregulated ipsilateral to the nerve injury; (iiii) the NPCs migrating differentiated only into GFAP-positive astrocytes in the hypoglossal nucleus. These results suggest that nerve avulsion injury induces in parallel with the retrograde "axon reaction" activation of endogenous NPCs in the ependymal region and further suggest that these cells could be involved in repair and neuroregeneration after injury within the brainstem.

Neurogenesis after complete spinal cord transection in Pleurodeles waltlii

Event Abstract Back to Event Neurogenesis after complete spinal cord transection in Pleurodeles waltlii Jonathan Chetrit1, Amira Zaky2 and Marie Moftah2* 1 Bordeaux 2 University, INSERM U862, France 2 Alexandria University, Zoology Department, Faculty of Science, Egypt Following spinal lesion, connections between the supraspinal centers and spinal neuronal networks can be disturbed, which causes the deterioration or even the complete absence of sublesional locomotor activity. In Mammals, possibilities of locomotion restoration are much reduced since descending tracts either have very poor regenerative ability or do not regenerate at all. However, in certain lower Vertebrates such as Urodeles, there is spontaneous locomotion recuperation after complete spinal cord transection. This phenomenon depends on a translesional descending axon re-growth originating from the brainstem. On the other hand, cellular and molecular mechanisms underlying spinal cord regeneration resulting in parallel in locomotion restoration of the animal are not well known. Fibroblast Growth Factor-2 (FGF-2) plays an important role in different processes such as neural induction, neuronal progenitor proliferation and differentiation. Recent studies showed an up-regulation of this growth factor after complete trunk spinal cord transection in Pleurodeles. Nestin, a protein specific for intermediate filaments, is considered as a neuronal precursor early marker. It has been recently shown that Nestin expression increases after tail transection in Urodeles. Using this marker, our results show that the increase in FGF-2 mRNAs (grain count following in situ hybridization) and that of its protein during spinal cord regeneration (western blots) are correlated with an increase in neurogenesis (immunohistochemistry) in the Urodele Amphibian Pleurodeles waltlii. This study also confirms that axonal re-growth through the lesion site initially follows a rostrocaudal direction. In addition to its known role in neuronal differentiation, FGF-2 could be implicated in the differentiation of ependymal cells into neuronal progenitors; thus promoting body spinal cord gap-replacement. Acknowledgment: We cordially thank Pr Jean-Marie Cabelguen and Dr Frédéric Nagy for their assistance, revisions and support. Conference: 2nd NEUROMED Workshop, Fez, Morocco, 10 Jun - 12 Jun, 2010. Presentation Type: Oral Presentation Topic: Oral Session 3: The plastic brain: implications for learning and education Citation: Chetrit J, Zaky A and Moftah M (2010). Neurogenesis after complete spinal cord transection in Pleurodeles waltlii. Front. Neurosci. Conference Abstract: 2nd NEUROMED Workshop. doi: 10.3389/conf.fnins.2010.12.00034 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 04 Jun 2010; Published Online: 04 Jun 2010. * Correspondence: Marie Moftah, Alexandria University, Zoology Department, Faculty of Science, Alexandria, Egypt, marie.moftah@alexu.edu.eg Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Jonathan Chetrit Amira Zaky Marie Moftah Google Jonathan Chetrit Amira Zaky Marie Moftah Google Scholar Jonathan Chetrit Amira Zaky Marie Moftah PubMed Jonathan Chetrit Amira Zaky Marie Moftah Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

FoxJ1-dependent gene expression is required for differentiation of radial glia into ependymal cells and a subset of astrocytes in the postnatal brain

Neuronal specification occurs at the periventricular surface of the embryonic central nervous system. During early postnatal periods, radial glial cells in various ventricular zones of the brain differentiate into ependymal cells and astrocytes. However, mechanisms that drive this time- and cell-specific differentiation remain largely unknown. Here, we show that expression of the forkhead transcription factor FoxJ1 in mice is required for differentiation into ependymal cells and a small subset of FoxJ1(+) astrocytes in the lateral ventricles, where these cells form a postnatal neural stem cell niche. Moreover, we show that a subset of FoxJ1(+) cells harvested from the stem cell niche can self-renew and possess neurogenic potential. Using a transcriptome comparison of FoxJ1-null and wild-type microdissected tissue, we identified candidate genes regulated by FoxJ1 during early postnatal development. The list includes a significant number of microtubule-associated proteins, some of which form a protein complex that could regulate the transport of basal bodies to the ventricular surface of differentiating ependymal cells during FoxJ1-dependent ciliogenesis. Our results suggest that time- and cell-specific expression of FoxJ1 in the brain acts on an array of target genes to regulate the differentiation of ependymal cells and a small subset of astrocytes in the adult stem cell niche.

Spinal Cord Injury Reveals Multilineage Differentiation of Ependymal Cells

Spinal cord injury often results in permanent functional impairment. Neural stem cells present in the adult spinal cord can be expanded in vitro and improve recovery when transplanted to the injured spinal cord, demonstrating the presence of cells that can promote regeneration but that normally fail to do so efficiently. Using genetic fate mapping, we show that close to all in vitro neural stem cell potential in the adult spinal cord resides within the population of ependymal cells lining the central canal. These cells are recruited by spinal cord injury and produce not only scar-forming glial cells, but also, to a lesser degree, oligodendrocytes. Modulating the fate of ependymal progeny after spinal cord injury may offer an alternative to cell transplantation for cell replacement therapies in spinal cord injury.

RETRACTED ARTICLE: Cyclophosphamide-induced agenesis of cerebral aqueduct resulting in hydrocephalus in mice

The present work was undertaken to reveal the mechanism of cerebral aqueduct agenesis found to result in hydrocephalus following intrauterine exposure to model teratogen, cyclophosphamide, in murine fetuses. A single dose of 10-mg/kg body weight cyclophosphamide was injected intaperitoneally to pregnant mice on day 10, 11 or 12 of gestation. Fetuses were collected through abdominal incision on day 18 and studied for various malformations of brain and cranium including hydrocephalus. Incomplete development and failure of canalization of the cerebral aqueduct were detected when serial sections of brain in coronal and transverse planes were studied under the microscope. Biotechnological investigations such as % DNA fragmentation, % viable cell count and cell proliferation assay were carried out on brain cells for further studies. Agenesis and non-canalization of the cerebral aqueduct resulted in increased pressure of CSF, which led to rupture of the aqueduct complicated by leakage and accumulation of CSF in brain substance forming a cavity containing CSF parallel and lateral to the unopened part of the cerebral aqueduct. Incomplete development along with non-canalization of the cerebral aqueduct resulted in blockage of CSF flow through the ventricles that manifest as internal hydrocephalus. External hydrocephalus on the other hand was detected where the CSF accumulated in the cavity formed inside the brain substance and established communication with the CSF in the subarachnoid space. Cyclophosphamide induced inhibition of mitosis and cell differentiation of ependymal cells reflecting a decreased % viable cell count and cell proliferation assay along with augmentation of apoptosis of brain cells quantified as increased % DNA fragmentation count, which were identified as the contributing factors underlying the agenesis and incomplete development of the cerebral aqueduct. The study also suggests that cell survival, proliferation, migration or differentiation of ependymal cells might have been affected, and we speculate that CSF may have an inducing role in the development and canalization of the cerebral aqueduct.

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.

Ependymal Cell Differentiation and GLUT1 Expression is a Synchronous Process in the Ventricular Wall

Ependymal cells appear to be totally differentiated during the first 3 weeks in the mouse brain. Early during postnatal development ependymal cells differentiate and undergo metabolic activation, which is accompanied by increased glucose uptake. We propose that ependymal cells induce an overexpression of the glucose transporter, GLUT1, during the first 2 weeks after delivery in order to maintain the early metabolic activation. During the first postnatal day, GLUT1 is strongly induced in the upper region of the third ventricle and in the ventral area of the rostral cerebral aqueduct. During the next 4 days, GLUT1 is expressed in all differentiated ependymal cells of the third ventricle and in hypothalamic tanycytes. At the end of the first week, ependymal cell differentiation and GLUT1 overexpression is concentrated in the latero-ventral area of the aqueduct. We propose that ependymal cell differentiation and GLUT1 overexpression is a synchronous process in the ventricular wall.

Upregulated and prolonged differentiation potential of the ependymal cells lining the ventriculus terminalis in human fetuses

The ventriculus terminalis (VT) is a dilated cavity within the conus medullaris of the spinal cord. Although the VT was discovered in the mid-nineteenth century, little is known about its characteristics during development in human fetuses. Ependymal cells lining the cavities within the CNS retain high differentiation potential, and are believed to be responsible for the postnatal neurogenesis. To evaluate the differentiation capacity of the ependymal cells lining the VT during development, we examined glial fibrillary acidic protein (GFAP) and proliferating cell nuclear antigen (PCNA) expression in the spinal cord of 18–24-week-old human fetuses. GFAP is a marker for the degree of ependymal cell differentiation in the human fetus, and PCNA is a well-known marker for cell division. Morphological characteristics of the VT were also examined. At the lower portion of the conus medullaris, the central canal abruptly expands dorsally to become the VT. Then the VT widens bilaterally while its anteroposterior diameter reduces gradually in a caudal direction. Finally, the VT becomes a narrow, transverse slit at the level of the lowermost conus medullaris. Compared with those lining the central canal, more numerous ependymal cells lining the VT showed more intensive GFAP and PCNA expression throughout all gestational ages examined. This suggests that, in the developing human spinal cord, ependymal cells lining the VT retain their differentiation potential, including a higher proliferative capacity, until a later stage of development than those lining the central canal.

Ependymal cell reactions in spinal cord segments after compression injury in adult rat.

Recently, it has been suggested that neural stem cells and neural progenitor cells exist in the ependyma that forms the central canal of the spinal cord. In this study, we produced various degrees of thoracic cord injury in adult rats using an NYU-weight-drop device, assessed the degree of recovery of lower limb motor function based on a locomotor rating scale, and analyzed the kinetics of ependymal cell proliferation and differentiation by proliferating cell nuclear antigen (PCNA), nestin, glial fibrillary acidic protein (GFAP), or GAP-43 immunostaining. The results showed that the time course of the ependymal cell proliferation and differentiation reactions differed according to the severity of injury, and that the responses occurred not only in the neighborhood of the injury but in the entire spinal cord. An increase in the locomotor rating score was related to an increase in the number of PCNA-positive cells, and the differentiation of ependymal cells into reactive astrocytes was involved in injury repair. No apoptotic cells in the ependyma were detectable by the TUNEL method. These results indicate that the ependymal cells of the spinal central canal are themselves multipotent, can divide and proliferate according to the severity of injury, and differentiate into reactive astrocytes within the ependyma without undergoing apoptosis or cell death.

L-Carnitine accelerates the in vitro regeneration of neural network from adult murine brain cells

The development, growth and regeneration of nerve cells remain an unresolved issue. The up-to-date reported brain repair mechanisms are numerous and evidence suggests that, apart from the required trophism, tropism, microenvironment and specificity of the brain, a plethora of chemical, physiological and immunological compounds can contribute to such events. Among these compounds, we concentrated our interest on l-carnitine ( l-Cn), which regulates the β-oxidation of long chain fatty acids necessary for brain development, myelinization and growth. In contrast to fetal brain cells that grow easily in culture, adult brain cells show limited neurogenesis. Here, using adult brain cells from experimental mice, we show that although l-Cn does not improve their proliferative activity in short-term cultures, it accelerates the growth and differentiation of neurons, astrocytes, oligodendrocytes and ependymal cells from neurospheres in long-term cultures. Thus, the formation of a confluent neural network requires a 2-month period in culture. These observations provide new insights for in vivo use of l-Cn to support brain cell development in cases of injury or brain degenerative diseases.

Differentiation of choroid plexus ependymal cells into astrocytes after grafting into the pre-lesioned spinal cord in mice.

Choroid plexus epithelial cells represent a continuation of, and have the same origin as, ventricular ependymal cells, and are regarded as modified ependymal cells. To extend previous studies of the use of choroid plexus ependymal cell (CPEC) grafting for nerve regeneration in the spinal cord, we investigated the capacity of cultured choroid plexus ependymal cells to differentiate into other types of glial cells in the spinal cord tissue. The choroid plexuses were excised from the fourth ventricle of green fluorescent protein (GFP)-transgenic mice and the cells were dissociated and cultured for 4-6 weeks. CPECs were harvested from the monolayer cultures and injected into the pre-lesioned spinal cords of wild-type mice of the same strain using a Hamilton syringe. One week after injection, some GFP-positive transplanted cells became immunohistochemically positive for glial fibrillary acidic protein (GFAP) but negative for neurofilament and myelin basic protein. All the GFAP-positive transplanted cells were negative for vimentin. Two weeks after grafting, immunoelectron microscopy showed that the GFP-positive transplanted cells that had gained GFAP immunoreactivity contained numerous bundles of intermediate filaments, a morphological characteristic similar to that of astrocytes, and were in close contact with adjacent host tissue. These results indicate that, when grafted into the spinal cord, at least some cultured choroid plexus ependymal cells have the capacity to differentiate into astrocytes.

Proliferation and Differentiation of Ependymal Cells after Transection of the Carp Spinal Cord

Proliferation and differentiation of ependymal cells in the injured carp spinal cord were studied by immunohistochemistry using proliferating cell nuclear antigen (PCNA) and 3H-thymidine (3H-TdR) autoradiography. A surge of proliferation of ependymal cells occurred in the lesion with a peak around 6th day after surgical operation (complete transection) of the spinal cord. The proportion of PCNA-positive ependymal cells at 6th post-operative day (6 POD) was over 14 times that in the normal state. Electron microscopic autoradiography revealed that most of the ependymal cells incorporating 3H-TdR at 5 POD, especially those located in the caudal side of the transection site, contained numerous free ribosomes in their apical portion, but the other organellae, such as rough endoplasmic reticulum and Golgi apparatus, were poorly developed in this portion. In three weeks thereafter, the ependymal cell layer was reconstructed through the lesion, and the apical part of the 3H-TdR-labeled ependymal cells became elongated and morphologically differentiated: that is, it had free ribosomes decreased and glial filaments increased in number. Many bundles of regenerating axons were observed to course within the reconstructed ependymal cell layer. These results may suggest that proliferation, differentiation and reconstruction of the ependymal cell layer following injury of the carp spinal cord are requisite to make the permissive milieu for elongation of the regenerating axons.

Ependymal Reactions to Injury. A Review

The ependyma reacts to injury with a few stereotypical responses and does not regenerate at any age. Non-neoplastic ependymal cells do not undergo mitotic proliferation and do not re-express fetal cytoskeletal or secretory proteins. Atrophy of ependymal cells accompanies generalized cerebral atrophy. The ependyma may be damaged by stretching during ventricular dilatation, by infarcts of the ventricular wall or by infection and inflammation. Tearing of the epithelium leaves discontinuities that become filled with processes of subventricular astrocytes. In some cases reactive gliosis is minimal, but in most it is extensive and gliotic nodules form beneath intact ependyma and within gaps between ependymal islands. Ependymal rosettes may form in several ways: sequestration of diverticuli from the surface; curling of a torn edge or penetration of an edge into the parenchyma; reactive gliosis overgrowing an ependymal edge; in situ differentiation of ependymal cells from deep neuroepithelial cells. Migration and metaplasia are unlikely mechanisms. Bacterial and fungal ependymitis are highly destructive. Several viruses, especially mumps, selectively infect ependymal cells and are an important cause of acquired aqueductal stenosis without inflammation. Damaged ependyma may not be able to perform its function in the regulation of transport of fluid, ions and small molecules between cerebral parenchyma and ventricular fluid and thus may contribute to hydrocephalus. Damage to the fetal ependyma may result in secondary focal dysplasias of the developing brain.