RESUMO
We report on PAX6 alleles associated with a clinical diagnosis of classical aniridia in 81 affected individuals representing 66 families. Allelic variants expected to affect PAX6 function were identified in 61 families (76 individuals). Ten cases of sporadic aniridia (10 families) had complete (8 cases) or partial (2 cases) deletion of the PAX6 gene. Sequence changes that introduced a premature termination codon into the open reading frame of PAX6 occurred in 47 families (62 individuals). Three individuals with sporadic aniridia (three families) had sequence changes (one deletion, two run-on mutations) expected to result in a C-terminal extension. An intronic deletion of unknown functional significance was detected in one case of sporadic aniridia (one family), but not in unaffected relatives. Within these 61 families, single nucleotide substitutions accounted for 30/61 (49%), indels for 23/61 (38%), and complete deletion of the PAX6 locus for 8/61 (13%). In five cases of sporadic aniridia (five families), no disease-causing mutation in the coding region was detected. In total, 23 unique variants were identified that have not been reported in the Leiden Open Variation Database (LOVD) database. Within the group assessed, 92% had sequence changes expected to reduce PAX6 function, confirming the primacy of PAX6 haploinsufficiency as causal for aniridia.
Assuntos
Aniridia/genética , Predisposição Genética para Doença/genética , Mutação , Fator de Transcrição PAX6/genética , Alelos , Análise Mutacional de DNA , Feminino , Haploinsuficiência/genética , Humanos , Mutação INDEL , Masculino , Modelos Moleculares , Mutagênese Insercional , Fator de Transcrição PAX6/química , Mutação Puntual , Domínios Proteicos , Deleção de SequênciaRESUMO
Leukocyte infiltration is a key step in the development of demyelinating lesions in multiple sclerosis (MS), and molecules mediating leukocyte-endothelial interactions represent prime candidates for the development of therapeutic strategies. Here we studied the effects of blocking the integrin-associated tetraspanin CD81 in in vitro and in vivo models for MS. In an in vitro setting mAb against CD81 significantly reduced monocyte transmigration across brain endothelial cell monolayers, both in rodent and human models. Interestingly, leukocyte as well as endothelial CD81 was involved in this inhibitory effect. To assess their therapeutic potential, CD81 mAb were administered to mice suffering from experimental autoimmune encephalomyelitis (EAE). We found that Eat2, but not 2F7 mAb directed against mouse CD81 significantly reduced the development of neurological symptoms of EAE when using a preventive approach. Concomitantly, Eat2 treated animals showed reduced inflammation in the spinal cord. We conclude that CD81 represents a potential therapeutic target to interfere with leukocyte infiltration and ameliorate inflammatory neurological damage in MS.
Assuntos
Anticorpos Monoclonais/farmacologia , Antígenos CD/efeitos dos fármacos , Quimiotaxia de Leucócito/efeitos dos fármacos , Encefalomielite Autoimune Experimental/tratamento farmacológico , Imunossupressores/farmacologia , Monócitos/efeitos dos fármacos , Animais , Anticorpos Monoclonais/uso terapêutico , Antígenos CD/imunologia , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/imunologia , Linhagem Celular Transformada , Artérias Cerebrais/citologia , Artérias Cerebrais/efeitos dos fármacos , Artérias Cerebrais/imunologia , Quimiotaxia de Leucócito/imunologia , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/fisiopatologia , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/imunologia , Feminino , Humanos , Terapia de Imunossupressão/métodos , Imunossupressores/uso terapêutico , Camundongos , Monócitos/imunologia , Esclerose Múltipla/tratamento farmacológico , Esclerose Múltipla/imunologia , Esclerose Múltipla/fisiopatologia , Ratos , Tetraspanina 28 , Resultado do TratamentoRESUMO
The present study is concerned with the projection of the lateral geniculate nucleus onto cortical area 18. Horseradish peroxidase (HRP) was injected into area 18 of 15 cats. Drawings were made to determine the location of the injection site and the distribution of labeled neurons in the lateral geniculate nuclei of each cat. The local retinotopic maps constructed prior to the injections and the reconstructions of the lateral geniculate nucleus were used to determine the location and the extent of each of the HRP injections. In 15 of the 25 hemispheres studied, the ratio of the number of HRP-labeled neurons in lamina A relative to the number of labeled neurons in lamina A1 was calculated. This ratio varied from 1.06 to 0.28, indicating that at least some regions of area 18 are dominated by inputs from lamina A1. However, if the HRP-labeled neurons in lamina C are included in the counts for lamina A, then the ratio A+C/A1 has a mean of 1.11, suggesting that area 18 receives a balanced input, with inputs from the contralateral eye being relayed through laminae A and C, and inputs from the ipsilateral eye being relayed through lamina A1. When the distribution of HRP-labeled neurons in lamina A was plotted onto a dorsal view of the lateral geniculate nucleus, the labeled neurons formed an ellipse with the long axis of the ellipse oriented parallel to the isoelevation lines. The representation of azimuth is compressed in area 18 relative to the lateral geniculate nucleus. In six hemispheres the injections were restricted to a few layers of the area 18. Following small injections into layer IV of area 18, the HRP-labeled neurons occupied an extensive region of the lateral geniculate nucleus, indicating a considerable amount of convergence of the inputs to area 18. In hemispheres where the injections were restricted to layers I and II, labeled neurons were only seen in the medial interlaminar nucleus and the C laminae.
Assuntos
Corpos Geniculados/anatomia & histologia , Córtex Visual/anatomia & histologia , Animais , Gatos , Histocitoquímica , Peroxidase do Rábano Silvestre , Neurônios Aferentes/anatomia & histologiaRESUMO
When a kitten is reared with one eyelid sutured closed, there are profound changes in the developing visual system. In the lateral geniculate nucleus, the neurons in the laminae innervated by the deprived eye are smaller than normal, and some of these neurons may lose connections with the visual cortex. In the present study a variety of double label retrograde transport methods were used to define the effects of monocular deprivation on cortical projections of geniculate neurons. One marker was injected into area 17 and the other was injected into area 18. Neurons projecting to area 17 are on average 16.4% smaller than those in the nondeprived laminae. The neurons that normally would project to both areas 17 and 18 by an axon that branches are the most severely affected by monocular deprivation. These cells are nearly 40% smaller than their counterparts in the nondeprived laminae, and many of the neurons appear to lose their projection to one of the cortical areas. These neurons may be at a distinct disadvantage, since they must compete with neurons from the nondeprived laminae for a considerable amount of cortical territory in two different cortical areas. This competition may be so severe that some of the neurons are no longer capable of maintaining connections with both cortical areas.
Assuntos
Corpos Geniculados/crescimento & desenvolvimento , Privação Sensorial/fisiologia , Visão Ocular/fisiologia , Córtex Visual/fisiologia , Animais , Gatos , Corpos Geniculados/citologia , Vias Visuais/fisiologiaRESUMO
ABAKAN is a keratin sulfate proteoglycan that was identified in rat brain by monoclonal antibody TED15 (Geisert et al. [1992] Brain Res. 571:165-168). It blocks neuronal attachment and neurite outgrowth in culture, is associated with astrocytes, and marks the boundaries of areas in the developing rat brain (Geisert and Bidanset [1993] Dev. Brain Res., 75:163-173). In the present study TED15 was used to examine the distribution of ABAKAN during laminar development of the dorsal lateral geniculate nucleus in ferrets. This distribution was also compared with that of astrocytes as displayed with antibodies to GFAP. In the adult, TED15 and anti-glial fibrillary acidic protein (GFAP) labeling are similar. Both are fairly uniform in the nucleus although somewhat elevated near the optic tract and in the interlaminar zone between laminae A and A1. During development the pattern is quite different. At postnatal day 1 (P1), before lamination is evident, TED15 and anti-GFAP labeling are light in the nucleus. By P10, when laminae are emerging, both are elevated in the A-A1 interlaminar zone and in the C laminae. At P18, when laminae are distinct, TED15 labels the A-A1 interlaminar zone, and it marks the borders between the ON and OFF leaflets within A and A1 (Stryker and Zahs [1983] J. Neurosci. 3:1943-1951). In comparison, anti-GFAP marks the interlaminar zone but not the ON/OFF leaflets. By 6 weeks the nucleus resembles the adult nucleus. These results show that ABAKAN marks the boundaries of the major functional subdivisions of the lateral geniculate nucleus in the developing ferret and suggest that it plays a role in lamination.
Assuntos
Furões/metabolismo , Corpos Geniculados/metabolismo , Sulfato de Queratano/biossíntese , Proteoglicanas/biossíntese , Animais , Animais Recém-Nascidos , Anticorpos Monoclonais/imunologia , Eletroforese em Gel de Poliacrilamida , Corpos Geniculados/crescimento & desenvolvimento , Proteína Glial Fibrilar Ácida/imunologia , Immunoblotting , Imuno-Histoquímica , Sulfato de Queratano/análise , Proteoglicanas/análiseRESUMO
The present study defines the expression pattern of TAPA (target of the antiproliferative antibody, also known as CD81) in the developing rat brain. TAPA is a member of the tetramembrane spanning family of proteins, and like other members of this family it appears to be associated with the stabilization of cellular contacts (Geisert et al. [1996] J. Neurosci. 16:5478-5487). On immunoblots of the brain, TAPA is present in higher levels than any other tissue examined: muscle, tendon, peripheral nerve, cartilage, liver, kidney, skin, and testicle. Immunohistochemical methods were used to define the distribution of TAPA in the brain. This protein is expressed by ependyma, choroid plexus, astrocytes, and oligodendrocytes. TAPA is dramatically upregulated during early postnatal development, at the time of glial birth and maturation. At embryonic day 18, the levels of TAPA are low, with most of the immunoreaction product being associated with the ependyma, choroid plexus, and the glia limitans. As development continues, the amount of TAPA expressed in the brain increases, and at postnatal day 14 the levels approach those of the adult. This increase in the levels of TAPA at postnatal day 14 is due to upregulation in the gray matter and white matter. Thus, TAPA is found in all glial cells, and the level of this protein correlates with their maturation.
Assuntos
Envelhecimento/fisiologia , Antígenos CD/biossíntese , Encéfalo/metabolismo , Córtex Cerebral/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Membrana/biossíntese , Neuroglia/metabolismo , Neuropeptídeos/biossíntese , Sequência de Aminoácidos , Animais , Animais Recém-Nascidos , Antígenos CD/química , Astrócitos/metabolismo , Encéfalo/embriologia , Encéfalo/crescimento & desenvolvimento , Células Cultivadas , Córtex Cerebral/crescimento & desenvolvimento , Hibridização In Situ , Masculino , Proteínas de Membrana/química , Modelos Moleculares , Dados de Sequência Molecular , Neuropeptídeos/química , Oligodendroglia/metabolismo , Especificidade de Órgãos , Conformação Proteica , Ratos , Ratos Sprague-Dawley , Tetraspanina 28RESUMO
The retinal afferents to the medial interlaminar nucleus and to its rostro-dorsal extension at the edge of the pulvinar have been studied in cats by fiber degeneration and autoradiographic methods. Fiber degeneration following section of one optic nerve shows two distinct retinal inputs: One is coarse-fibered and goes to the medial interlaminar nucleus itself; the second, which is fine-fibered, goes to the rostral and medial borders of the medial interlaminar nucleus and continues into the pulvinar. The regions in receipt of these fine fibers have been called the "geniculate wing". The topography of retinal representations and the degree of binocular overlap within the medial interlaminar nucleus and the wing have been studied by combining intraocular injections of 3H proline with local lesions of the injected eye, or with removal of the non-injected eye. In the medial interlaminar nucleus three distinct laminae are recognizable and are particularly clearly shown in horizontal sections. Rostrally and medially, lamina 1 maps the contralateral nasal retina as a mirror reversal of lamina A. Posterior and lateral to this, lamina 2 maps the ipsilateral temporal retina as a mirror reversal of lamina A1. Lamina 3 lies closest to the optic tract and receives crossed afferents from the temporal retina. In this lamina, which is the smallest, vertical retinal dimensions map as in the other layers, but we were unable to determine the mapping of the horizontal dimensions. In the geniculate wing, as in the other geniculate layers, vertical lines of the visual field are mapped as corresponding vertical diencephalic dimensions; horizontal retinal dimensions are mapped as horizontal lines in the wing, with central retinal areas represented furthest from the optic tract. In the geniculate wing the contralateral nasal and ipsilateral temporal retina are mapped with considerable binocular overlap. The crossed temporal retina has no demonstrable representation in the wing.
Assuntos
Retina/anatomia & histologia , Núcleos Talâmicos/anatomia & histologia , Animais , Axônios/ultraestrutura , Mapeamento Encefálico , Gatos , Corpos Geniculados/anatomia & histologia , Campos Visuais , Vias Visuais/anatomia & histologiaRESUMO
CD81 (TAPA), a member of the tetraspanin family of proteins, is upregulated by astrocytes and microglia after traumatic injury to the rat central nervous system (CNS). To further understand the role of CD81 in the microglial response to injury, we analysed the functional effects of a CD81 antibody, AMP1, on cultured rat microglia. We found that AMP1 suppressed microglial proliferation in a dose-dependent manner. Furthermore, AMP1 stimulated myelin phagocytosis, probably by opsonizing the myelin. The phagocytosis of latex beads, as well as the production of nitric oxide, were not significantly influenced by AMP1. These data indicate that CD81 is involved in an important subset of microglial effector functions after CNS injury.
Assuntos
Antígenos CD/imunologia , Proteínas de Membrana , Microglia/citologia , Microglia/imunologia , Óxido Nítrico/biossíntese , Fagocitose/imunologia , Animais , Anticorpos/farmacologia , Antígenos CD/biossíntese , Divisão Celular/imunologia , Células Cultivadas , Técnicas In Vitro , Microglia/metabolismo , Microesferas , Bainha de Mielina/imunologia , Bainha de Mielina/metabolismo , Ratos , Ratos Wistar , Traumatismos da Medula Espinal/imunologia , Tetraspanina 28RESUMO
PURPOSE: The target of the antiproliferative antibody (TAPA, CD81) is a member of the tetramembrane spanning superfamily of proteins and appears to be involved in the regulation of mitotic activity and the stabilization of cellular contacts [J Neurosci 1996; 16:5478-5487]. The present study examines the distribution of this protein in the normal rat retina and its role in reactive gliosis occurring after retinal injury. METHODS: An immunoblot was used to define the relative level of TAPA in the normal rat retina. The distribution of the protein was examined using indirect immunohistochemical methods. Both of these methods were used to define the upregulation of TAPA in the rat retina injured with a needle scrape. RESULTS: The immunohistochemical analysis of the retina shows that TAPA is found in all layers of the normal retina with a distinct lack of labeling in the inner and outer segments of the photoreceptors. After retinal injury, a dramatic upregulation of TAPA was observed. The immunohistochemistry also revealed a pattern of expression similar to that observed in the normal retina with two notable exceptions: (1) small finger-like projections extending down into the outer segments are immunopositive, and (2) the elevated levels of TAPA can be seen outlining cell bodies in the outer nuclear layer and the ganglion cell layer. CONCLUSIONS: TAPA is found in the normal rat retina and there is a dramatic upregulation of this protein following injury. The distribution of the protein within the retina is consistent with its expression in retinal glia, the Muller cells which span the thickness of the retina, and astrocytes found in the ganglion cell layer. These data suggest that TAPA may play a role in the proliferative response of non-neuronal cells that occurs following a mechanical injury to the retina.
Assuntos
Antígenos CD/metabolismo , Proteínas de Membrana/metabolismo , Retina/metabolismo , Animais , Antígenos CD/imunologia , Astrócitos/metabolismo , Proteína Glial Fibrilar Ácida/análise , Gliose/metabolismo , Immunoblotting , Imuno-Histoquímica , Neuroglia/metabolismo , Células Fotorreceptoras/metabolismo , Ratos , Ratos Sprague-Dawley , Retina/química , Retina/lesões , Células Ganglionares da Retina/química , Tetraspanina 28 , Regulação para Cima , Vimentina/análiseRESUMO
In the present study, a novel combination of techniques was used to identify the genes that may be involved in the lack of axonal regeneration in the mammalian adult central nervous system (CNS). The key features of this approach are: (1) a functional assay that can be affected by antibody perturbation; (2) increased specificity of the polyclonal antiserum by adsorption; (3) the expression cloning of the genes from a lambdagt11 library; (4) amplification of the insert cDNA by PCR; and (5) the direct cycle sequencing of PCR products. In this culture assay system, neurons were plated directly on sections of the rat CNS. This assay system could be used to demonstrate the lack of neuronal attachment to or neurite extension over myelinated regions of the CNS (white matter). This prohibitive nature of the CNS sections could be masked by a rabbit polyclonal antiserum directed against rat CNS white matter. This data indicates that the anti-white matter antiserum recognizes and neutralizes inhibitory molecules on the surface of the sections. Making the assumption that the prohibitive antigen is associated with the cell membrane, the antiserum was adsorbed against a soluble protein fraction of the adult rat brain. This adsorption significantly increased the specificity of the antiserum as demonstrated by immunoblot methods. The adsorbed antiserum was then used to screen the cDNA library of the adult rat brain. The present report describes this novel combination of techniques allowing one to go from a functional tissue culture assay system to defining the molecular basis for the cellular interactions.
Assuntos
Inibidores do Crescimento/fisiologia , Proteínas do Tecido Nervoso/fisiologia , Inibição Neural , Neuritos/fisiologia , Adsorção , Animais , Clonagem Molecular , Corpo Caloso/fisiologia , Feminino , Expressão Gênica/imunologia , Hipocampo/fisiologia , Soros Imunes/fisiologia , Masculino , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/imunologia , Inibição Neural/efeitos dos fármacos , Inibição Neural/genética , Neuritos/efeitos dos fármacos , Nervo Óptico/fisiologia , Ratos , Ratos Sprague-DawleyRESUMO
Following injury to the central nervous system (CNS), astrocytes become reactive and in many cases form a glial scar. Very little is known about the adhesive interactions between astrocytes at the glial scar, even though reactive gliosis and scar formation are a central issue in CNS wound healing. In the present study, we examine the role of cadherin in the process of scar formation using immunohistochemistry and immunoblot methods. When a stab wound was made in the cerebral cortex of the rat, cadherins were consistently upregulated by the reactive astrocytes at the glial scar. Our immunoblot analysis demonstrates that the increase in cadherin immunoreactivity was due to a threefold upregulation of a single protein with a molecular weight of 135 kDa. The size (135 kDa) and location of the immunoreactive protein at regions of cell-cell contact in cultured astrocytes indicates that the immunoreactive protein is N-cadherin. These data are the first to demonstrate that N-cadherin plays a prominent role in the response of astrocytes to injury, including the formation and maintenance of the glial scar.
Assuntos
Caderinas/fisiologia , Cicatriz/fisiopatologia , Gliose/fisiopatologia , Proteínas do Tecido Nervoso/fisiologia , Animais , Adesão Celular/fisiologia , Cicatriz/patologia , Immunoblotting , Imuno-Histoquímica , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
The influence of the cortico-geniculate pathway on identified X and Y lateral geniculate cells was studied by reversibly cooling visual cortical areas 17 and 18. The majority (86.5%) of cells changed their response to visual stimulation when cortex was inactivated, and both X- and Y-cells were modulated by the cortical input. The influence of the visual cortex was complex, with both excitatory and inhibitory actions. Furthermore, the mechanism underlying the basic center-surround receptive field organization was influenced.
Assuntos
Corpos Geniculados/fisiologia , Córtex Visual/fisiologia , Animais , Gatos , Temperatura Baixa , Potenciais Evocados , Inibição Neural , Neurônios/fisiologia , Transmissão Sináptica , Vias Visuais/fisiologia , Percepção Visual/fisiologiaRESUMO
The distribution of a 90 kDa protein, termed KG-CAM, was examined in the developing and adult rat central nervous system (CNS) using the monoclonal antibody 11-59. The amino acid sequence of this protein revealed a sequence homology with a group of chick cell adhesion molecules from the immunoglobulin superfamily: DM-GRASP; SC1; and BEN. Immunolabeling of cells cultured from the embryonic and neonatal rat brain demonstrates that the protein recognized by 11-59 is on the external surface of a subpopulation of neurons and a limited population of glial cells. When the 11-59 antibody was used to stain sections of the adult brain and spinal cord, a number of different structures were labeled. The most intense immunoreactivity was found in the somatosensory system, the basal ganglia, the cortex, the olfactory system, and the circumventricular organs. One of the more interesting aspects of KG-CAM is the spatially and temporally regulated patterns of expression observed during the development of the CNS. For example, the dendrites of layer II pyramidal cells in the granular retrosplenial cortex are immunopositive for 11-59 while the dendrites are in the process of bundling in layer I, but not before bundling begins or after the process is completed. These findings reveal the varied roles of this adhesion molecule in the developing brain and spinal cord, as well as its potential role in the maintenance of the structural integrity of the adult CNS.
Assuntos
Moléculas de Adesão Celular/metabolismo , Sistema Nervoso Central/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Molécula de Adesão de Leucócito Ativado , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais , Moléculas de Adesão Celular/análise , Moléculas de Adesão Celular/imunologia , Sistema Nervoso Central/anatomia & histologia , Sistema Nervoso Central/crescimento & desenvolvimento , Endopeptidases , Hidrólise , Immunoblotting , Camundongos/imunologia , Dados de Sequência Molecular , Peso Molecular , Proteínas do Tecido Nervoso/análise , Proteínas do Tecido Nervoso/imunologia , RatosRESUMO
A novel CNS specific keratan sulfate proteoglycan is identified by a monoclonal antibody (mAb) TED15. The antibody recognizes a carbohydrate epitope that is removed from the core protein following keratanase digestion. The relative molecular weight of the glycoprotein is not altered following digestion with heparinase, heparitinase or chondroitinase ABC. By immunoblot analysis, the TED15 antigen is differentially distributed in diencephalic derivatives, being present in the optic nerve but absent from the retina. Within the optic nerve, the TED15 antigen is associated with astrocytes, while in the retina no labeling of astrocytes of Müller cells is observed.
Assuntos
Astrócitos/metabolismo , Sistema Nervoso Central/metabolismo , Sulfato de Queratano/metabolismo , Nervo Óptico/metabolismo , Proteoglicanas/metabolismo , Animais , Anticorpos Monoclonais , Imuno-Histoquímica , Nervo Óptico/citologia , Ratos , Retina/citologia , Retina/metabolismo , Distribuição TecidualRESUMO
The up-regulation of the keratan sulfate proteoglycan (ABAKAN) was examined using indirect immunohistochemical methods. Previous studies indicate that the keratan sulfate proteoglycan is associated with astrocytes in the optic nerve and in the developing rat brain. In model culture systems, this proteoglycan is capable of inhibiting the growth of neurites over laminin. To determine whether the proteoglycan is up-regulated specifically during reactive gliosis, stab wounds were made in the cerebral cortex of early postnatal rats, and the up-regulation of the proteoglycan was related to the developmentally regulated gliotic response to injury. Following a stab wound in the cortex of the late postnatal rat, reactive gliosis was consistently observed along with an up-regulation of ABAKAN. When the cortex was injured on postnatal day 2, there was a variable gliotic response and considerable variation in the regulation of proteoglycan expression. Biochemical analysis revealed that ABAKAN is a large proteoglycan with multiple keratan sulfate side-chains, at least one chondroitin sulfate side-chain and at least one additional carbohydrate chain with a terminal 3-sulfoglucuronic acid. Taken together, these data demonstrate that the boundary proteoglycan ABAKAN is also associated with reactive gliosis during early postnatal development.
Assuntos
Animais Recém-Nascidos/fisiologia , Córtex Cerebral/lesões , Córtex Cerebral/metabolismo , Proteoglicanas de Sulfatos de Condroitina/biossíntese , Sulfato de Queratano/biossíntese , Regulação para Cima/fisiologia , Animais , Anticorpos Monoclonais , Astrócitos/metabolismo , Córtex Cerebral/citologia , Proteoglicanas de Sulfatos de Condroitina/isolamento & purificação , Eletroforese em Gel de Poliacrilamida , Feminino , Hidrólise , Immunoblotting , Imuno-Histoquímica , Sulfato de Queratano/isolamento & purificação , Lumicana , Neuritos/metabolismo , Gravidez , Ratos , Ratos Sprague-DawleyRESUMO
During the development of the central nervous system (CNS), adhesive molecules promote the formation of axonal pathways and appropriate neuronal connections by facilitating cellular interactions. In addition to the interactions that bring neurons together, recent evidence suggests inhibition of neuronal interactions also plays a role by restricting axons to their appropriate pathways and forming boundaries between functional units of the developing CNS. The present study describes the distribution of a recently identified large keratan sulfate proteoglycan, ABAKAN, in the postnatal day 14 (P14) and adult rat brain. In the adult brain ABAKAN appears to be relatively evenly distributed throughout the CNS, while at P14 this proteoglycan is found at high concentrations between different functional units of the neonatal brain. For example, ABAKAN appears to separate different cortical areas and mark the boundaries between thalamic nuclei. In vitro assays demonstrate that this keratan sulfate proteoglycan is a potent inhibitor of neurite growth. The distribution of ABAKAN at P14 and the effects of this keratan sulfate proteoglycan on neurite growth suggest that ABAKAN functions as a molecular barrier to axonal growth in the developing rat brain.
Assuntos
Animais Recém-Nascidos/metabolismo , Astrócitos/metabolismo , Química Encefálica/fisiologia , Encéfalo/crescimento & desenvolvimento , Proteoglicanas de Sulfatos de Condroitina/metabolismo , Sulfato de Queratano/metabolismo , Envelhecimento/metabolismo , Animais , Anticorpos Monoclonais/imunologia , Encéfalo/anatomia & histologia , Encéfalo/citologia , Córtex Cerebral/crescimento & desenvolvimento , Córtex Cerebral/metabolismo , Proteoglicanas de Sulfatos de Condroitina/imunologia , Proteoglicanas de Sulfatos de Condroitina/farmacologia , Immunoblotting , Imuno-Histoquímica , Sulfato de Queratano/imunologia , Sulfato de Queratano/farmacologia , Laminina/metabolismo , Lumicana , Camundongos , Neuritos/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Núcleos Talâmicos/crescimento & desenvolvimento , Núcleos Talâmicos/metabolismoRESUMO
The neuronal response to trauma of the brain and spinal cord was examined by staining sections of injured central nervous system (CNS) with a monoclonal antibody (TuJ1) that recognizes class III beta-tubulin exclusively. Because class III beta-tubulin is expressed by neurons and not by glia, this monoclonal antibody stains neuronal cell bodies, dendrites, axons and axonal terminations darkly with a pale staining background. Thus, the TuJ1 antibody is extremely useful, revealing the fine details of axons and their terminations, as well as significant injury-related alterations in the composition of the somatic cytoskeleton.
Assuntos
Axônios/análise , Encéfalo/ultraestrutura , Neurônios/análise , Tubulina (Proteína)/análise , Animais , Anticorpos Monoclonais , Axônios/ultraestrutura , Química Encefálica , Técnicas Imunoenzimáticas , Degeneração Neural , Neurônios/ultraestrutura , Ratos , Ratos EndogâmicosRESUMO
The upregulation of a 106-kDa glial protein was examined using indirect immunohistochemical methods. The protein is recognized by the monoclonal antibody AMP1. Previous studies demonstrated that the AMP1 antigen is found on the external surface of cultured astrocytes and is involved in stabilizing adhesive interactions between these cells. In sections of injured adult rat brain, the spatial distribution of the AMP1 antigen correlated with the region of reactive gliosis. The relative intensity of immunofluorescence indicates that the AMP1 antigen is dramatically upregulated at the gliotic scar. The correlation between AMP1 antigen upregulation and reactive gliosis suggests that this molecule is critical to the process of CNS scar formation.
Assuntos
Antígenos de Superfície/imunologia , Astrócitos/imunologia , Neuroglia/imunologia , Regulação para Cima/imunologia , Animais , Anticorpos Monoclonais/imunologia , Moléculas de Adesão Celular Neuronais/imunologia , Moléculas de Adesão Celular Neuronais/metabolismo , Proteína Glial Fibrilar Ácida/imunologia , Proteína Glial Fibrilar Ácida/metabolismo , Imuno-Histoquímica , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
The present study uses the monoclonal antibody TED 15 to examine the distribution of a keratan sulfate proteoglycan, ABAKAN, in the barrel field of the rat somatosensory cortex. At birth there is very little ABAKAN present in the somatosensory cortex. The levels of this proteoglycan increase during development until the highest levels are reached in the adult rat. Immunohistochemistry reveals that the TED 15 immunoreaction product marks the boundaries between cortical barrels. At postnatal day 7 (P7) the proteoglycan is localized specifically to the boundary regions. As the brain matures, the levels of ABAKAN increase in the barrel hollows and in the surrounding cortex; however the boundary regions maintain a higher level of expression even in the adult animal. The high levels of ABAKAN observed in the adult barrel fields indicate that unlike other boundary molecules this proteoglycan may be involved in maintaining the structure of the adult somatosensory cortex.
Assuntos
Proteoglicanas de Sulfatos de Condroitina/metabolismo , Sulfato de Queratano/metabolismo , Córtex Somatossensorial/metabolismo , Animais , Animais Recém-Nascidos , Immunoblotting , Imuno-Histoquímica , Lumicana , Regeneração Nervosa/fisiologia , Neurônios Aferentes/fisiologia , Ratos , Ratos Sprague-Dawley , Córtex Somatossensorial/citologia , Córtex Somatossensorial/crescimento & desenvolvimentoRESUMO
Using a monoclonal antibody (mAb), designated AMP1, a novel cell adhesion molecule was identified on rat astrocytes and oligodendrocytes. When confluent monolayers of cultured rat astrocytes were labeled with AMP1, the antigen was discretely localized to the cell surface in regions of cell-cell contacts. The antibody did not label embryonic rat cortical neurons plated on monolayers of neonatal astrocytes, indicating that the antigen is neither present on cultured neurons nor does it reorganize on the glial surface under the neurons. On immunoblots of astrocytic or brain proteins, mAb AMP1 recognized a 106 kDa protein. In the present paper, data are presented demonstrating that the AMP1 antigen is a cell adhesion molecule and is distinct from all the known cell adhesion molecules present on astrocytes: N-CAM, N-cadherin, or members of the beta 1 integrin family. We have tentatively termed this molecule 'glial cell adhesion molecule' (G-CAM).