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1.
Nat Commun ; 8(1): 1719, 2017 11 23.
Artigo em Inglês | MEDLINE | ID: mdl-29170512

RESUMO

Animal fetuses and embryos may have applications in the generation of human organs. Progenitor cells may be an appropriate cell source for regenerative organs because of their safety and availability. However, regenerative organs derived from exogenous lineage progenitors in developing animal fetuses have not yet been obtained. Here, we established a combination system through which donor cells could be precisely injected into the nephrogenic zone and native nephron progenitor cells (NPCs) could be eliminated in a time- and tissue-specific manner. We successfully achieved removal of Six2+ NPCs within the nephrogenic niche and complete replacement of transplanted NPCs with donor cells. These NPCs developed into mature glomeruli and renal tubules, and blood flow was observed following transplantation in vivo. Furthermore, this artificial nephron could be obtained using NPCs from different species. Thus, this technique enables in vivo differentiation from progenitor cells into nephrons, providing insights into nephrogenesis and organ regeneration.


Assuntos
Néfrons/metabolismo , Transplante de Células-Tronco/métodos , Células-Tronco/metabolismo , Quimeras de Transplante , Animais , Diferenciação Celular , Feminino , Masculino , Mesoderma/citologia , Mesoderma/embriologia , Mesoderma/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Néfrons/citologia , Néfrons/embriologia , Organogênese , Ratos Sprague-Dawley , Ratos Transgênicos , Especificidade da Espécie , Células-Tronco/citologia
2.
Neuroscience ; 300: 585-92, 2015 Aug 06.
Artigo em Inglês | MEDLINE | ID: mdl-26012491

RESUMO

Movement dysfunction in Parkinson's disease (PD) is caused by the degeneration of dopaminergic (DA) neurons in the substantia nigra (SN). Here, we established a method for voxel-based morphometry (VBM) and automatic tissue segmentation of the marmoset monkey brain using a 7-T animal scanner and applied the method to assess DA degeneration in a PD model, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated animals, with tyrosine-hydroxylase staining. The most significant decreases of local tissue volume were detected in the bilateral SN of MPTP-treated marmoset brains (-53.0% in right and -46.5% in left) and corresponded with the location of DA neurodegeneration found in histology (-65.4% in right). In addition to the SN, the decreases were also confirmed in the locus coeruleus, and lateral hypothalamus. VBM using 7-T MRI was effective in detecting volume loss in the SN of the PD-model marmoset. This study provides a potential basis for the application of VBM with ultra-high field MRI in the clinical diagnosis of PD. The developed method may also offer value in automatic whole-brain evaluation of structural changes for the marmoset monkey.


Assuntos
Callithrix/anatomia & histologia , Intoxicação por MPTP/patologia , Imageamento por Ressonância Magnética/métodos , Substância Negra/patologia , Animais , Callithrix/metabolismo , Intoxicação por MPTP/metabolismo , Imageamento por Ressonância Magnética/instrumentação , Masculino , Tamanho do Órgão , Reconhecimento Automatizado de Padrão/métodos , Substância Negra/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo
3.
Neuroscience ; 257: 158-74, 2014 Jan 17.
Artigo em Inglês | MEDLINE | ID: mdl-24220690

RESUMO

The present study characterized fetal sulcation patterns and gyrification in the cerebrum of the New World monkey group, common marmosets, using a 3D T2-weighted high-resolution anatomical magnetic resonance imaging (MRI) sequence from the fixed brain at 7-tesla ex vivo. Fetal sulcation in the marmoset cerebrum began to indent the lateral fissure and hippocampal sulcus in gestational week (GW) 12, and then the following sulci emerged: the callosal and calcarine sulci on GW 15; the superior temporal sulcus on GW 17; and the circular and occipitotemporal sulci on GW 18. The degree of cortical convolution was evaluated quantitatively based on 2D MRI slices by the gyrification index (GI) and based on 3D MRI data by sulcation index (SI). Both the mean GI and SI increased from GW 16, and were closely correlated with the cortical volume and the cortical surface area during fetal periods (their correlation coefficients marked more than 0.95). After birth, both the mean GI and SI decreased slightly by 2years of age, whereas the cortical volume and surface area continuously increased. Notably, histological analysis showed that the outer subventricular zone (oSVZ) in non-sulcal regions was thicker than that in the presumptive calcarine sulcal region on GW 13, preceding the infolding of the calcarine sulcus. The present results showed definite sulcal infolding on the cerebral cortical surface of the marmosets, with similar pattern and sequence of their emergences to other higher-order primates such as macaques and humans. Differential expansion of the oSVZ may be involved in gyral convolution and sulcal infolding in the developing cerebrum.


Assuntos
Mapeamento Encefálico , Córtex Cerebral , Imageamento por Ressonância Magnética , Fatores Etários , Animais , Animais Recém-Nascidos , Callithrix , Córtex Cerebral/anatomia & histologia , Córtex Cerebral/embriologia , Córtex Cerebral/crescimento & desenvolvimento , Embrião de Mamíferos , Proteínas do Olho/metabolismo , Feminino , Lateralidade Funcional , Idade Gestacional , Proteínas de Homeodomínio/metabolismo , Processamento de Imagem Assistida por Computador , Masculino , Fator de Transcrição PAX6 , Fatores de Transcrição Box Pareados/metabolismo , Proteínas Repressoras/metabolismo , Proteínas com Domínio T/metabolismo , Tubulina (Proteína)/metabolismo
4.
Neuroscience ; 230: 102-13, 2013 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-23047019

RESUMO

The developmental anatomy of the brain is largely directed by neural-based cues. Despite this knowledge, the developmental trajectory of the primate brain has not yet been fully characterized. To realize this goal, the advance in noninvasive imaging methods and new brain atlases are essential. The common marmoset (Callithrix jacchus), a small New World primate, is widely used in neuroscience research. The recent introduction of transgenic techniques has enabled the marmoset to be used as a genetically modifiable primate model for brain development. Here, a magnetic resonance histology technique involving the use of ultra-high-resolution ex vivo magnetic resonance imaging (MRI) was performed to identify the developmental anatomy of the marmoset brain at different time points from gestational week 8 through to birth. The data allowed the generation of a multidimensional atlas of brain structures at different developmental stages. Furthermore, in utero MRI techniques were developed to noninvasively monitor brain development during the embryonic and fetal stages. The multidimensional atlas and the MRI tools developed herein are anticipated to further our understanding of the developing primate brain.


Assuntos
Mapeamento Encefálico , Encéfalo , Callithrix/anatomia & histologia , Imageamento por Ressonância Magnética , Animais , Animais Recém-Nascidos , Encéfalo/anatomia & histologia , Encéfalo/embriologia , Encéfalo/crescimento & desenvolvimento , Embrião de Mamíferos , Imageamento Tridimensional , Fatores de Tempo
5.
Br J Cancer ; 106(6): 1148-52, 2012 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-22343623

RESUMO

BACKGROUND: It is becoming increasingly recognised that opioids are responsible for tumour growth. However, the effects of opioids on tumour growth have been controversial. METHODS: The effects of κ-opioid receptor (KOR) agonist on the growth of non-small cell lung cancer (NSCLC) cells were assessed by a cell proliferation assay. Western blotting was performed to ascertain the mechanism by which treatment with KOR agonist suppresses tumour growth. RESULTS: Addition of the selective KOR agonist U50,488H to gefitinib-sensitive (HCC827) and gefitinib-resistant (H1975) NSCLC cells produced a concentration-dependent decrease in their growth. These effects were abolished by co-treatment with the selective KOR antagonist nor-BNI. Furthermore, the growth-inhibitory effect of gefitinib in HCC827 cells was further enhanced by co-treatment with U50,488H. With regard to the inhibition of tumour growth, the addition of U50, 488H to H1975 cells produced a concentration-dependent decrease in phosphorylated-glycogen synthase kinase 3ß (p-GSK3ß). CONCLUSION: The present results showed that stimulation of KOR reduces the growth of gefitinib-resistant NSCLC cells through the activation of GSK3ß.


Assuntos
(trans)-Isômero de 3,4-dicloro-N-metil-N-(2-(1-pirrolidinil)-ciclo-hexil)-benzenoacetamida/farmacologia , Proliferação de Células/efeitos dos fármacos , Receptores Opioides kappa/agonistas , Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas , Linhagem Celular Tumoral , Sobrevivência Celular , Resistencia a Medicamentos Antineoplásicos , Receptores ErbB/genética , Gefitinibe , Expressão Gênica , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta , Humanos , Mutação de Sentido Incorreto , Naltrexona/análogos & derivados , Naltrexona/farmacologia , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Quinazolinas/farmacologia , Receptores Opioides kappa/antagonistas & inibidores , Receptores Opioides kappa/genética , Receptores Opioides kappa/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais
6.
Br J Cancer ; 105(10): 1615-24, 2011 Nov 08.
Artigo em Inglês | MEDLINE | ID: mdl-22015555

RESUMO

BACKGROUND: Although matrix metalloproteinases (MMPs) are implicated in tumourigenesis and cancer progression, the role of MMP-13 in melanoma cell metastases is poorly understood. METHODS: Lung metastases of mouse melanoma B16BL6 cells were analysed in MMP-13 knockout (KO) and wild-type (WT) mice after intravenous injection. The mRNA and protein expression of MMP-13 in lung tissues was analysed by RT-PCR, real-time PCR, immunoblotting and immunohistochemistry. The expression of SDF-1α, CXCR4 and endostatin, and effects of endostatin to cultured melanoma cells and lung metastases were also studied. RESULTS: Lung metastases of B16BL6 cells were significantly higher by 2.5-5.7-fold in MMP-13 KO mice than in WT mice. The expression of MMP-13 in WT mouse lung tissue was stimulated on day 1 after intravenous injection of the melanoma cells and MMP-13 was immunolocalised to vascular endothelial cells in the lungs. Endostatin formation, but not degradation of SDF-1α, in the lung tissue was associated with reduced lung metastasis in WT mice. Endostatin significantly inhibited migration of B16BL6 cells in monolayer wounding assay and remarkably suppressed Matrigel invasion and transendothelial invasion of the cells. In addition, lung metastases of melanoma cells in MMP-13 KO mice were reduced by intraperitoneal administration of endostatin. CONCLUSION: Our results suggest that MMP-13 is overproduced by endothelial cells in the lungs with melanoma cells and has a protective role in lung metastasis by local generation of endostatin.


Assuntos
Endostatinas/biossíntese , Neoplasias Pulmonares/prevenção & controle , Metaloproteinase 13 da Matriz/metabolismo , Melanoma Experimental/patologia , Animais , Sequência de Bases , Primers do DNA , Ensaio de Imunoadsorção Enzimática , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/secundário , Melanoma Experimental/enzimologia , Camundongos , Camundongos Knockout , Microscopia de Fluorescência , Reação em Cadeia da Polimerase Via Transcriptase Reversa
7.
Neuroimage ; 54(4): 2741-9, 2011 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-21044887

RESUMO

Advanced magnetic resonance (MR) neuroimaging analysis techniques based on voxel-wise statistics, such as voxel-based morphometry (VBM) and functional MRI, are widely applied to cognitive brain research in both human subjects and in non-human primates. Recent developments in imaging have enabled the evaluation of smaller animal models with sufficient spatial resolution. The common marmoset (Callithrix jacchus), a small New World primate species, has been widely used in neuroscience research, to which voxel-wise statistics could be extended with a species-specific brain template. Here, we report, for the first time, a tissue-segmented, population-averaged standard template of the common marmoset brain. This template was created by using anatomical T(1)-weighted images from 22 adult marmosets with a high-resolution isotropic voxel size of (0.2 mm)(3) at 7-Tesla and DARTEL algorithm in SPM8. Whole brain templates are available at International Neuroinformatics Japan Node website, http://brainatlas.brain.riken.jp/marmoset/.


Assuntos
Anatomia Artística , Atlas como Assunto , Encéfalo/anatomia & histologia , Callithrix/anatomia & histologia , Animais , Feminino , Processamento de Imagem Assistida por Computador , Imageamento por Ressonância Magnética , Masculino
8.
Curr Top Microbiol Immunol ; 324: 179-96, 2008.
Artigo em Inglês | MEDLINE | ID: mdl-18481461

RESUMO

The radiological modalities that are currently utilized as critical components in clinical medicine have also been adapted to small-animal imaging, among which are ultrasound imaging, X-ray computerized tomography (CT), magnetic resonance imaging (MRI), positron emission tomography (PET), and single-photon emission computed tomography (SPECT). Optical imaging techniques such as bioluminescence imaging (BLI) and fluorescence imaging (FLI) are approaches that are commonly used in small animals. Longitudinal surveys of living (i.e., nonsacrificed) animal models with these modalities provide some clues for the development of clinical applications. The techniques are absolutely essential for translational research. However, there are currently few tools available with sufficient spatial or temporal resolution ideal for all experimental studies. In this chapter, we provide a rationale and techniques for visualizing target cells in living small animals and an overview of the advantages and limitations of current imaging technology. Finally, we introduce a humanized mouse and a novel in vivo imaging system that we have developed. We also discuss real-time observations of reconstructs and clinical manifestations.


Assuntos
Diagnóstico por Imagem/métodos , Modelos Animais , Animais , Camundongos , Camundongos SCID
9.
Mol Cell ; 5(4): 683-93, 2000 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-10882104

RESUMO

The TRAP220 component of the TRAP/SMCC complex, a mammalian homologof the yeast Mediator that shows diverse coactivation functions, interacts directly with nuclear receptors. Ablation of the murine Trap220 gene revealed that null mutants die during an early gestational stage with heart failure and exhibit impaired neuronal development with extensive apoptosis. Primary embryonic fibroblasts derived from null mutants show an impaired cell cycle regulation and a prominent decrease of thyroid hormone receptor function that is restored by ectopic TRAP220 but no defect in activation by Gal4-RARalpha/RXRalpha, p53, or VP16. Moreover, haploinsufficient animals show growth retardation, pituitary hypothyroidism, and widely impaired transcription in certain organs. These results indicate that TRAP220 is essential for a wide range of physiological processes but also that it has gene- and activator-selective functions.


Assuntos
Proteínas de Transporte/metabolismo , Hipófise/embriologia , Receptores dos Hormônios Tireóideos/metabolismo , Glândula Tireoide/embriologia , Hormônios Tireóideos/metabolismo , Fatores de Transcrição , Animais , Proteínas de Transporte/genética , Desenvolvimento Embrionário e Fetal , Fibroblastos/fisiologia , Genes Letais , Cardiopatias Congênitas , Fígado/embriologia , Pulmão/embriologia , Subunidade 1 do Complexo Mediador , Camundongos , Camundongos Knockout , Malformações do Sistema Nervoso , Hipófise/metabolismo , Placenta/embriologia , Glândula Tireoide/metabolismo , Transativadores/metabolismo , Transcrição Gênica , Proteína Supressora de Tumor p53/metabolismo
10.
Proc Natl Acad Sci U S A ; 97(12): 6350-5, 2000 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-10829067

RESUMO

The Nova paraneoplastic antigens are neuron-specific RNA binding proteins that participate in the control of alternative splicing. We have used the yeast two-hybrid system to isolate Nova interacting proteins and identify an RNA binding protein that is closely related to the polypyrimidine tract-binding protein (PTB). The expression of this protein, brPTB, is enriched in the brain, where it is expressed in glia and neurons. brPTB interacts with Nova proteins in cell lines and colocalizes with Nova within neuronal nuclei. We previously found that Nova binds to a pyrimidine-rich RNA element present upstream of an alternatively spliced exon, E3A, in glycine receptor alpha2 (GlyRalpha2) pre-mRNA, and this binding is implicated in Nova-dependent regulation of splicing. Cotransfection assays with a GlyRalpha2 minigene demonstrate that brPTB antagonizes the action of Nova to increase utilization of GlyRalpha2 E3A. brPTB binds to a 90-nt GlyRalpha2 RNA adjacent to the Nova binding site, but with an affinity that is more than 10-fold lower than Nova. When a putative binding site for brPTB on the GlyRalpha2 RNA is mutated, binding is abolished and the inhibitory effect on Nova-dependent exon selection disappears. These results suggest that brPTB is a tissue-restricted RNA binding protein that interacts with and inhibits the ability of Nova to activate exon selection in neurons.


Assuntos
Processamento Alternativo , Antígenos de Neoplasias/fisiologia , Encéfalo/metabolismo , Proteínas do Tecido Nervoso/fisiologia , Proteínas de Ligação a RNA/fisiologia , Ribonucleoproteínas/fisiologia , Animais , Sequência de Bases , Masculino , Dados de Sequência Molecular , Antígeno Neuro-Oncológico Ventral , Proteína de Ligação a Regiões Ricas em Polipirimidinas , Ratos , Ratos Sprague-Dawley , Receptores de Glicina/genética
11.
Neuron ; 25(2): 359-71, 2000 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-10719891

RESUMO

We have combined genetic and biochemical approaches to analyze the function of the RNA-binding protein Nova-1, the paraneoplastic opsoclonus-myoclonus ataxia (POMA) antigen. Nova-1 null mice die postnatally from a motor deficit associated with apoptotic death of spinal and brainstem neurons. Nova-1 null mice show specific splicing defects in two inhibitory receptor pre-mRNAs, glycine alpha2 exon 3A (GlyRalpha2 E3A) and GABA(A) exon gamma2L. Nova protein in brain extracts specifically bound to a previously identified GlyRalpha2 intronic (UCAUY)3 Nova target sequence, and Nova-1 acted directly on this element to increase E3A splicing in cotransfection assays. We conclude that Nova-1 binds RNA in a sequence-specific manner to regulate neuronal pre-mRNA alternative splicing; the defect in splicing in Nova-1 null mice provides a model for understanding the motor dysfunction in POMA.


Assuntos
Processamento Alternativo/fisiologia , Antígenos de Neoplasias , Neurônios Motores/citologia , Neurônios Motores/fisiologia , Proteínas do Tecido Nervoso , Proteínas de Ligação a RNA/genética , Ribonucleoproteínas/genética , Animais , Apoptose/genética , Química Encefálica/genética , Tronco Encefálico/citologia , Tronco Encefálico/embriologia , Sobrevivência Celular/genética , Éxons/genética , Deleção de Genes , Regulação da Expressão Gênica no Desenvolvimento , Genótipo , Marcação In Situ das Extremidades Cortadas , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neurônios Motores/química , Antígeno Neuro-Oncológico Ventral , Ligação Proteica/genética , Precursores de RNA/genética , Proteínas de Ligação a RNA/metabolismo , Receptores de GABA-A/genética , Receptores de GABA-A/metabolismo , Receptores de Glicina/genética , Receptores de Glicina/metabolismo , Ribonucleoproteínas/metabolismo , Medula Espinal/citologia , Medula Espinal/embriologia
12.
Genes Dev ; 13(16): 2087-97, 1999 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-10465786

RESUMO

Paraneoplastic cerebellar degeneration (PCD) is a disorder in which breast or ovarian tumors express an onconeural antigen termed cdr2, which normally is expressed in cerebellar Purkinje neurons. This leads to an immune response to cdr2 that is associated with tumor immunity and autoimmune cerebellar degeneration. We have found that cdr2, a cytoplasmic protein harboring a helix-leucine zipper (HLZ) motif, interacts specifically with the HLZ motif of c-Myc. Both proteins colocalize in the cytoplasm of adult cerebellar Purkinje neurons, and coimmunoprecipitate from tumor cell lines and cerebellar extracts. cdr2 down-regulates c-Myc-dependent transcription in cotransfection assays, and redistributes Myc protein in the cytoplasm. Disease antisera from six of six PCD patients specifically blocked the interaction between cdr2 and c-Myc in vitro. These data indicate that cdr2 normally sequesters c-Myc in the neuronal cytoplasm, thereby down-regulating c-Myc activity, and suggest a mechanism whereby inhibition of cdr2 function by autoantibodies in PCD may contribute to Purkinje neuronal death.


Assuntos
Autoantígenos/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Células de Purkinje/metabolismo , Animais , Autoanticorpos/imunologia , Autoantígenos/genética , Autoantígenos/imunologia , Sobrevivência Celular , Citoplasma/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/imunologia , Regulação para Baixo , Células HeLa , Humanos , Zíper de Leucina , Masculino , Camundongos , Síndromes Paraneoplásicas/imunologia , Células de Purkinje/citologia , Ratos , Ratos Sprague-Dawley , Transcrição Gênica , Células Tumorais Cultivadas
13.
Proc Natl Acad Sci U S A ; 96(17): 9885-90, 1999 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-10449789

RESUMO

Hu proteins are mammalian embryonic lethal abnormal visual system (ELAV)-like neuronal RNA-binding proteins that contain three RNA recognition motifs. Although Drosophila ELAV is required for the correct differentiation and survival of neurons, the roles played by the Hu genes in the mammalian nervous system remain largely unknown. To explore the in vivo functions of mouse Hu proteins, we overexpressed them in rat pheochromocytoma PC12 cells, where they induced neuronal phenotype in the absence of nerve growth factor. We have characterized the functions of various forms of mHuB and mHuC bearing point mutations or deletions. Mutants of mHuC that had amino acid exchanges in the RNP1 domain of the first or second RNA recognition motifs (RRMs) lost biologic activity as well as RNA-binding activity. In addition, the mutants containing only the third RRM failed to induce the neuronal phenotype in PC12 cells and inhibited the biologic activity of cotransfected wild-type mHuB and mHuC, thus acting as a dominant-negative form. However, these mutants could not suppress the nerve growth factor-induced differentiation of PC12 cells. Further, we misexpressed wild-type and dominant-negative Hu in E9.5 mouse embryos, by using electroporation into the neural tube at the level of the rhombencephalon. mHuB and mHuC induced the ectopic expression of neuronal markers, whereas the dominant-negative forms of mHuB and mHuC suppressed the differentiation of central nervous system motor neurons. From these results, we suggest that Hu proteins are required for neuronal differentiation in the mammalian nervous system.


Assuntos
Antígenos de Superfície/fisiologia , Sistema Nervoso Central/crescimento & desenvolvimento , Proteínas do Tecido Nervoso/fisiologia , Neurônios/fisiologia , Sistema Nervoso Periférico/crescimento & desenvolvimento , Proteínas de Ligação a RNA/fisiologia , Animais , Antígenos de Superfície/genética , Análise Mutacional de DNA , Proteínas ELAV , Proteína Semelhante a ELAV 2 , Proteína Semelhante a ELAV 3 , Camundongos , Proteínas do Tecido Nervoso/genética , Células PC12 , Fenótipo , Proteínas de Ligação a RNA/genética , Ratos , Ribonucleoproteínas/fisiologia
14.
Genomics ; 45(2): 313-9, 1997 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-9344654

RESUMO

The paraneoplastic neurologic disorders (PND) are a rare group of neurologic syndromes that arise when an immune response to systemic tumors expressing neuronal proteins ("onconeural antigens") develops into an autoimmune neuronal degeneration. The use of patient antisera to clone the genes encoding PND antigens has led to new insight into the mechanism of these autoimmune disorders. The tumor antigens can now be grouped into three classes: (1) neuron-specific RNA-binding proteins, (2) nerve terminal vesicle-associated proteins, and (3) cytoplasmic signaling proteins. To understand better the evolutionary relatedness of these genes and to evaluate them as candidates for inherited neurological disorders, we have determined the mouse chromosomal locations of nine of these genes-Hua, Hub, Huc, Hud, Nova1, Nova2, Natpb, Cdr2, and Cdr3. These data suggest that the Hua-Hud genes arose from gene duplication and dispersion, while the other genes are dispersed in the genome. We also predict the chromosomal locations of these genes in human and discuss the potential of these genes as candidates for uncloned mouse and human mutations.


Assuntos
Antígenos de Neoplasias/genética , Mapeamento Cromossômico , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/imunologia , Doenças do Sistema Nervoso/genética , Doenças do Sistema Nervoso/imunologia , Síndromes Paraneoplásicas/genética , Síndromes Paraneoplásicas/imunologia , Animais , Autoimunidade , Cruzamentos Genéticos , DNA Complementar/genética , Feminino , Ligação Genética , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Muridae , Mutação , Degeneração Neural/genética , Degeneração Neural/imunologia
15.
Proc Natl Acad Sci U S A ; 94(13): 7001-5, 1997 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-9192681

RESUMO

Leptin's effects are mediated by interactions with a receptor that is alternatively spliced, resulting in at least five different murine forms: Ob-Ra, Ob-Rb, Ob-Rc, Ob-Rd, and Ob-Re. A mutation in one splice form, Ob-Rb, results in obesity in mice. Northern blots, RNase protection assays, and PCR indicate that Ob-Rb is expressed at a relatively high level in hypothalamus and low level in several other tissues. Ob-Ra is expressed ubiquitously, whereas Ob-Rc, -Rd, and -Re RNAs are only detectable using PCR. In hypothalamus, Ob-Rb is present in the arcuate, ventromedial, dorsomedial, and lateral hypothalamic nuclei but is not detectable in other brain regions. These nuclei are known to regulate food intake and body weight. The level of Ob-Rb in hypothalamus is reduced in mice rendered obese by gold thioglucose (GTG), which causes hypothalamic lesions. The obesity in GTG-treated mice is likely to be caused by ablation of Ob-Rb-expressing neurons, which results in leptin resistance.


Assuntos
Encéfalo/metabolismo , Proteínas de Transporte/metabolismo , Receptores de Superfície Celular , Processamento Alternativo , Animais , Northern Blotting , Proteínas de Transporte/genética , Hibridização In Situ , Camundongos , Dados de Sequência Molecular , Mutação , Especificidade de Órgãos , Receptores para Leptina
16.
J Neurosci ; 17(9): 3024-37, 1997 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-9096138

RESUMO

The Hu proteins are a group of antigens targeted in an immune-mediated neurodegenerative disorder associated with cancer. We have cloned and characterized four members of the Hu gene family from mouse. We find that the Hu genes encode a large number of alternatively spliced transcripts to produce a series of related neuron-specific RNA binding proteins. Despite this complexity, we have discerned several ordered features of Hu expression. In the embryo, specific Hu genes are expressed in a hierarchy during early neurogenesis. In the E16 developing cortex, mHuB is induced in very early postmitotic neurons exiting the ventricular zone, mHuD is expressed in migrating neurons of the intermediate zone, and mHuC is expressed in mature cortical plate neurons. Such a hierarchy suggests distinct functional roles for each gene in developing neurons. In the adult, all neurons express some set of Hu mRNA and protein. However, specific patterns are evident such that individual neuronal types in the hippocampus, cerebellum, olfactory cortex, neocortex, and elsewhere express from one to several Hu genes. The complexity of potential protein variants within a gene family and of different Hu family members within a neuron suggests a diverse array of function. Given the strong homologies among the Hu proteins, the Drosophila neurogenic gene elav, and the Drosophila splicing factor sxl, we predict that different combinations of Hu proteins determine different neuron-specific aspects of post-transcriptional RNA regulation. Our findings of specific developmental patterns of expression and the correlation between immune targeting of the Hu proteins and adult neurodegenerative disease suggest that the Hu proteins are critical in both the proper development and function of mature neurons.


Assuntos
Proteínas de Bactérias/genética , Proteínas de Transporte/metabolismo , Proteínas de Ligação a DNA/genética , Neurônios/metabolismo , RNA/metabolismo , Animais , Proteínas de Bactérias/metabolismo , Clonagem Molecular , Proteínas de Ligação a DNA/metabolismo , Humanos , Imuno-Histoquímica , Hibridização In Situ , Camundongos
17.
Proc Natl Acad Sci U S A ; 93(13): 6275-9, 1996 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-8692805

RESUMO

MacMARCKS is a member of the MARCKS family of protein kinase C (PKC) substrates. Biochemical evidence demonstrates that these proteins integrate calcium and PKC-dependent signals to regulate actin structure at the membrane. We report here that deletion of the MacMARCKS gene prevents cranial neural tube closure in the developing brain, resulting in anencephaly. This suggests a central role for MacMARCKS and the PKC signal transduction pathway in the folding of the anterior neural plate during the early phases of brain formation, and supports the hypothesis that actin-based motility directs cranial neural tube closure.


Assuntos
Anencefalia/genética , Proteínas de Membrana , Defeitos do Tubo Neural/genética , Proteínas/genética , Animais , Sequência de Bases , Encéfalo/anormalidades , Encéfalo/embriologia , Proteínas de Ligação a Calmodulina , Linhagem Celular , Quimera , Primers do DNA , Desenvolvimento Embrionário e Fetal/genética , Peptídeos e Proteínas de Sinalização Intracelular , Camundongos , Camundongos Endogâmicos C57BL , Proteínas dos Microfilamentos , Dados de Sequência Molecular , Proteína Quinase C/metabolismo , Proteínas/metabolismo , Transdução de Sinais
18.
Dev Neurosci ; 18(3): 199-209, 1996.
Artigo em Inglês | MEDLINE | ID: mdl-8894448

RESUMO

Immunocytochemical and autoradiographic techniques were used to examine proliferating cells in the adult rat brain along with the expression of specific growth factor receptors and cell cycle proteins. Two hours following an injection of [3H]thymidine ([3H]Thy), dividing cells were detected in the subgranular region of the dentate gyrus and in the subependymal region (SER) extending into the olfactory bulb. Many cells continued to divide over the next 24 h as demonstrated by the ability for thymidine-labeled cells to incorporate bromodeoxyuridine (BrdU); however, the results of BrdU, PSTAIR, and vimentin staining suggest that the majority of the progeny cells detectable by [3H]Thy autoradiography at 3 days and 1 week after injection are postmitotic and at least partially differentiated. Significant numbers of thymidine-labeled cells detected 2 h following thymidine injection in the subgranular region of the dentate gyrus and in the SER of the lateral ventricle stained positively for epidermal growth factor receptor-, vimentin-, and PSTAIR-like immunoreactivity. Significant numbers of thymidine-labeled cells in the SER also stained positively for the low-affinity neurotrophin receptor p75NGFR. No [3H]Thy/p75NGFR-labeled cells were detected in the dentate gyrus. In addition, very few [3H]Thy/PSTAIR- or [3H]Thy/ vimentin-labeled cells were detected in region CA4. These data suggest that proliferating cells located in different regions of the adult brain may not be homogeneous and may be subject to different growth factor regulation.


Assuntos
Proteína Quinase CDC2/biossíntese , Fator de Crescimento Epidérmico/biossíntese , Hipocampo/metabolismo , Proteínas do Tecido Nervoso/biossíntese , Fragmentos de Peptídeos/biossíntese , Prosencéfalo/metabolismo , Receptores de Fator de Crescimento Neural/biossíntese , Animais , Proteína Quinase CDC2/genética , Ciclo Celular , Diferenciação Celular , Divisão Celular , Giro Denteado/citologia , Giro Denteado/metabolismo , Fator de Crescimento Epidérmico/genética , Regulação da Expressão Gênica , Hipocampo/citologia , Masculino , Proteínas do Tecido Nervoso/genética , Bulbo Olfatório/citologia , Bulbo Olfatório/metabolismo , Fragmentos de Peptídeos/genética , Prosencéfalo/citologia , Ratos , Ratos Sprague-Dawley , Receptor de Fator de Crescimento Neural , Receptores de Fator de Crescimento Neural/genética , Células-Tronco/citologia , Células-Tronco/metabolismo , Vimentina/biossíntese , Vimentina/genética
19.
Regul Pept ; 59(2): 143-9, 1995 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-8584749

RESUMO

Intracerebral diffusion, cellular uptake and intracellular localization of oligodeoxynucleotides (ODN) after their microinjection in mouse brains were examined. Using either tetramethylrhodamine-5-(and -6)-isothiocyanate (TRITC)- or gamma-33P ATP-labeled ODNs, it was found that both phosphodiester ODNs (D-ODN) and phosphorothioate ODNs (S-ODN) quickly diffused (up to about 500 microns) and were taken up by many cells around the injection site as early as 15 min after administration. Fluorescence labeling intensity and silver grain accumulation of D-ODNs were greatly reduced by 4 h after injection, whereas those of S-ODNs were stable beyond at least 8 h after injection. Most of labeled ODNs were found in neuronal cells as identified by immunocytochemistry for neurofilament, NF 200, and to a much lesser extent in astrocytic cells as identified by immunocytochemistry for glial fibrillary acidic protein.


Assuntos
Encéfalo/metabolismo , Oligonucleotídeos Antissenso/metabolismo , Animais , Sequência de Bases , Encéfalo/citologia , Difusão , Feminino , Proteína Glial Fibrilar Ácida/metabolismo , Imuno-Histoquímica , Injeções Intraventriculares , Camundongos , Dados de Sequência Molecular , Oligonucleotídeos Antissenso/farmacocinética
20.
Cell ; 82(5): 773-83, 1995 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-7671305

RESUMO

We have identified a target antigen in autoimmune cerebellar degeneration, beta-NAP, that is closely related to the beta-adaptin and beta-COP coat proteins. Beta-NAP is a nonclathrin-associated phosphoprotein expressed exclusively in neurons, from E12 through adulthood. Beta-NAP is present in the neuronal soma and nerve terminal as soluble and membrane-bound pools and is associated with a discrete set of nerve-terminal vesicles. These results establish beta-NAP as a neuron-specific vesicle coat protein. We propose a model in which beta-NAP mediates vesicle transport between the soma and the axon terminus and suggest that beta-NAP may represent a general class of coat proteins that mediates apical transport in polarized cells.


Assuntos
Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Doenças Cerebelares/imunologia , Proteínas de Membrana/genética , Proteínas do Tecido Nervoso/genética , Neurônios/química , Fosfoproteínas/genética , Complexo 3 de Proteínas Adaptadoras/análise , Subunidades beta do Complexo de Proteínas Adaptadoras/análise , Animais , Antígenos/fisiologia , Capsídeo , Cerebelo/química , Cerebelo/embriologia , Cerebelo/patologia , Feto/fisiologia , Expressão Gênica/fisiologia , Imuno-Histoquímica , Proteínas de Membrana/análise , Camundongos , Microscopia Eletrônica , Dados de Sequência Molecular , Proteínas do Tecido Nervoso/análise , Fosfoproteínas/análise , Terminações Pré-Sinápticas/química , RNA Mensageiro/análise , Homologia de Sequência de Aminoácidos , Vesículas Sinápticas/química
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