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1.
Pediatr Transplant ; 14(2): 261-7, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20470360

RESUMO

Despite significant interest by pediatric transplant patients in meeting others who have undergone transplantation, geographic distances combined with their daily routines make this difficult. This mixed-method study describes the use of Zora, a Web-based virtual community designed to create a support system for these patients. The Zora software allows participants to create a graphical online virtual city with houses expressing their individuality and objects conveying their concerns and personal stories. Zora allows real-time chat between participants further facilitating communication. Twenty-two post-transplant patients used Zora over nine months. The median number of log ons per participant was 19.50 times (q1 = 5.25, q3 = 41.50), and each participant spent a median of 12.48 h (q1 = 2.13, q3 = 25.55) logged into the program. This represented a median of 18.27 min/wk (q1 = 6.88, q3 = 37.40) per participant. Users created a total of 3736 objects (median/participant = 12.5, q1 = 2.25, q3 = 30) and created 66 virtual houses (median/participant = 2.00, q1 = 1.00, q3 = 3.00). In addition, a total of 14,444 lines of chat were recorded (median/participant = 228.5, q1 = 30.00, q3 = 663.25), and a total 278 messages were sent between users (median/participant = 3.50, q1 = 0.25, q3 = 15.5). Qualitative data show the preliminary success of the project, as three major themes emerged: (i) increased sense of normalcy for the patients, (ii) enhanced sense of self and contribution to the community, and (iii) increased social network. There were no instances of harmful interactions in the virtual world. This study demonstrates the feasibility and safety of a virtual community as a potential psychosocial intervention for post-transplant adolescents.


Assuntos
Internet , Transplante de Órgãos/psicologia , Apoio Social , Adolescente , Criança , Estudos de Viabilidade , Feminino , Humanos , Relações Interpessoais , Masculino , Software
2.
Paediatr Anaesth ; 19(10): 1031-3, 2009 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-19754495

RESUMO

A fiberoptic-assisted laryngoscope (FLS) (Acoma Medical Industry Co. Ltd., Tokyo, Japan) is a modified Macintosh laryngoscope with a tubular holder into which a fiberoptic bronchoscope can be inserted. We present three cases of Treacher Collins syndrome whose tracheas were successfully intubated with the aid of the FLS. These cases suggest that the FLS may be a useful alternative in the case of difficult pediatric intubation.


Assuntos
Intubação Intratraqueal , Laringoscopia/métodos , Disostose Mandibulofacial/diagnóstico , Anestesia por Inalação , Criança , Orelha Externa/cirurgia , Feminino , Fístula/congênito , Fístula/cirurgia , Humanos , Laringoscópios , Boca/cirurgia , Fibras Ópticas
3.
Neurosci Res ; 52(3): 250-62, 2005 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15927725

RESUMO

Postmitotic neural precursors are generated in the ventricular zone (VZ) of the developing neural tube and immediately migrate to the mantle layer (ML) where they differentiate into mature neurons. Although the regulation of neuronal differentiation and migration has extensively been studied, the behavior of the early postmitotic precursors migrating toward the ML is largely unknown. In this study, we have identified Neph3 as a specific marker for early postmitotic neural precursors in the VZ of the developing spinal cord. Analysis of Neph3 localization by immunofluorescence and immunoelectron microscopy revealed that early neural precursors in the VZ possessed not only pia-connected processes but also ones that reached the ventricle. This apical extension of processes was confirmed by analyzing another early postmitotic marker, Dll1 mRNA, which was actively transported toward the ventricle and accumulated at the termini of the processes. Furthermore, adherens junctions (AJs) were formed around the apical end of processes extending from Neph3- and Dll1 mRNA-positive postmitotic precursors. Taken together, these observations suggest that migrating early postmitotic neural precursors in the VZ of the developing spinal cord form a neuroepithelial cell-like bipolar morphology and communicate with their neighboring cells through AJs.


Assuntos
Junções Aderentes/fisiologia , Movimento Celular/fisiologia , Ventrículos Cerebrais/citologia , Neurônios/fisiologia , Medula Espinal/citologia , Células-Tronco/fisiologia , Junções Aderentes/ultraestrutura , Animais , Northern Blotting , Caderinas/metabolismo , Agregação Celular/fisiologia , Células Cultivadas , Ventrículos Cerebrais/embriologia , Ventrículos Cerebrais/ultraestrutura , Clonagem Molecular/métodos , Proteínas de Ligação a DNA/metabolismo , Proteínas de Drosophila/metabolismo , Embrião de Mamíferos , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Vetores Genéticos/fisiologia , Proteínas de Homeodomínio/metabolismo , Imuno-Histoquímica/métodos , Hibridização In Situ/métodos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Microscopia Imunoeletrônica , Proteínas de Neurofilamentos/metabolismo , Neurônios/ultraestrutura , Fosfoproteínas/metabolismo , RNA Mensageiro/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Medula Espinal/embriologia , Tubulina (Proteína)/metabolismo , Proteína da Zônula de Oclusão-1
4.
J Biochem Biophys Methods ; 60(1): 61-7, 2004 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-15236911

RESUMO

The single nucleotide polymorphism (SNP) of aldehyde dehydrogenase-2 (ALDH2) codon 487, GAA (Glu) or AAA (Lys), was examined using green fluorescent protein (GFP)-display, an electrophoretic detection method for single amino acid changes. Although no shift in migration between the GFP-ALDH (Glu487) and GFP-ALDH (Lys487) fusion proteins was observed on SDS/urea gel, the two migrated to different positions when tagged with Asp. The SNP analysis was performed with GFP-ALDH-Asp3, and GFP-ALDH-Asp3 constructed from donors having the codon GAA/GAA, GAA/AAA or AAA/AAA was detected as different patterns as expected. GFP-display is potentially a unique method in SNP analysis, which does not require any special equipment or chemicals.


Assuntos
Técnicas Genéticas , Proteínas de Fluorescência Verde/química , Polimorfismo de Nucleotídeo Único , Aldeído Desidrogenase/genética , Aldeído-Desidrogenase Mitocondrial , Ácido Aspártico/química , Códon , DNA/química , Eletroforese em Gel de Poliacrilamida/métodos , Escherichia coli/metabolismo , Humanos , Peptídeos/química , Plasmídeos/metabolismo
5.
J Biol Chem ; 279(17): 18015-25, 2004 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-14871893

RESUMO

Cell migration plays roles in invasion of transformed cells and scattering of embryonic mesenchymal cells into surrounding tissues. We have found that Ig-like Necl-5/Tage4 is up-regulated in NIH3T3 cells transformed by an oncogenic Ras (V12Ras-NIH3T3 cells) and heterophilically trans-interacts with a Ca(2+)-independent Ig-like cell adhesion molecule nectin-3, eventually enhancing their intercellular motility. We show here that Necl-5 furthermore enhances cell migration in a nectin-3-independent manner. Studies using L fibroblasts expressing various mutants of Necl-5, NIH3T3 cells, and V12Ras-NIH3T3 cells have revealed that Necl-5 enhances serum- and platelet-derived growth factor-induced cell migration. The extracellular region of Necl-5 is necessary for directional cell migration, but not for random cell motility. The cytoplasmic region of Necl-5 is necessary for both directional and random cell movement. Necl-5 colocalizes with integrin alpha(V)beta(3) at leading edges of migrating cells. Analyses using an inhibitor or an activator of integrin alpha(V)beta(3) or a dominant negative mutant of Necl-5 have shown the functional association of Necl-5 with integrin alpha(V)beta(3) in cell motility. Cdc42 and Rac small G proteins are activated by the action of Necl-5 and required for the serum-induced, Necl-5-enhanced cell motility. These results indicate that Necl-5 regulates serum- and platelet-derived growth factor-induced cell migration in an integrin-dependent, nectin-3-independent manner, when cells do not contact other cells. We furthermore show here that enhanced motility and metastasis of V12Ras-NIH3T3 cells are at least partly the result of up-regulated Necl-5.


Assuntos
Antígenos de Neoplasias/fisiologia , Moléculas de Adesão Celular/fisiologia , Integrinas/metabolismo , Proteínas de Neoplasias/fisiologia , Animais , Antígenos de Neoplasias/metabolismo , Cálcio/metabolismo , Adesão Celular , Moléculas de Adesão Celular/metabolismo , Divisão Celular , Linhagem Celular Transformada , Movimento Celular , Transformação Celular Neoplásica , Citoplasma/metabolismo , Inibidores Enzimáticos/farmacologia , Fibroblastos/metabolismo , Transferência Ressonante de Energia de Fluorescência , Integrina alfaVbeta3/metabolismo , Camundongos , Camundongos Nus , Microscopia de Fluorescência , Células NIH 3T3 , Nectinas , Metástase Neoplásica , Proteínas de Neoplasias/metabolismo , Fator de Crescimento Derivado de Plaquetas/metabolismo , Ligação Proteica , Transfecção , Regulação para Cima
6.
J Biol Chem ; 278(37): 35421-7, 2003 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-12826663

RESUMO

Nectins are Ca2+-independent immunoglobulin-like cell-cell adhesion molecules that play roles in organization of a variety of cell-cell junctions in cooperation with or independently of cadherins. Four nectins have been identified. Five nectin-like molecules, which have domain structures similar to those of nectins, have been identified, and we characterized here nectin-like molecule-2 (Necl-2)/IGSF4/RA175/SgIGSF/TSLC1/SynCAM1. Necl-2 showed Ca2+-independent homophilic cell-cell adhesion activity. It furthermore showed Ca2+-independent heterophilic cell-cell adhesion activity with Necl-1/TSLL1/SynCAM3 and nectin-3. Necl-2 was widely expressed in rat tissues examined. Necl-2 localized at the basolateral plasma membrane in epithelial cells of the mouse gall bladder, but not at specialized cell-cell junctions, such as tight junctions, adherens junctions, and desmosomes. Nectins bind afadin, whereas Necl-2 did not bind afadin but bound Pals2, a membrane-associated guanylate kinase family member known to bind Lin-7, implicated in the proper localization of the Let-23 protein in Caenorhabditis elegans, the homologue of mammalian epidermal growth factor receptor. These results indicate the unique localization of Necl-2 and its possible involvement in localization of a transmembrane protein(s) through Pals2.


Assuntos
Moléculas de Adesão Celular/fisiologia , Adesão Celular/fisiologia , Células Epiteliais/fisiologia , Imunoglobulinas/fisiologia , Proteínas de Membrana/fisiologia , Animais , Molécula 1 de Adesão Celular , Moléculas de Adesão Celular/genética , Membrana Celular/metabolismo , Clonagem Molecular , Fibroblastos/fisiologia , Vetores Genéticos , Imunoglobulinas/genética , Células L , Proteínas de Membrana/genética , Camundongos , Dados de Sequência Molecular , Especificidade de Órgãos , Plasmídeos , Ligação Proteica , Transporte Proteico , Ratos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/fisiologia , Transfecção
7.
Anal Biochem ; 317(1): 107-15, 2003 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-12729607

RESUMO

The migrating position of green fluorescent protein (GFP)-fused polypeptide varied on an SDS/urea gel by a single amino acid change in the fused polypeptide segment. An easy detection method for a single amino acid change based on this observation was called "GFP-display." Using various target polypeptides, staphylococcal protein A (SpA), Ras, p53, and human beta3 adrenergic receptor (AR), and their mobility-shift patterns resulting from the single amino acid changes, several important properties of GFP-display were revealed as follows: (i). since the binding of dodecyl sulfate ions to acidic or hydrophilic amino acids is weaker than that to basic or hydrophobic amino acids, the ions bound weakly to the fused polypeptide segment are forced to come off by high concentrations of urea prior to the ions bound strongly, resulting in the mobility shift, (ii). the mobility shift is estimated to a certain extent using a new parameter called the "GD value" calculated from the isoelectric point, hydrophilicity, and number of fused amino acids, and (iii). the fluorescence intensity of GFP-fused polypeptide tends to increase with the average hydrophilicity of the fused polypeptide segment. GFP-display will be a helpful technique for many kinds of gene or protein studies related to amino acid substitutions such as the random mutagenesis in a gene of interest.


Assuntos
Proteínas Luminescentes/química , Peptídeos/análise , Peptídeos/genética , Substituição de Aminoácidos , Bases de Dados Genéticas , Ensaio de Desvio de Mobilidade Eletroforética , Escherichia coli/química , Escherichia coli/metabolismo , Fluorescência , Proteínas de Fluorescência Verde , Humanos , Proteínas Luminescentes/genética , Mutagênese , Peptídeos/química , Receptores Adrenérgicos beta 3/análise , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteína Estafilocócica A/análise , Proteína Supressora de Tumor p53/análise , Ureia/química , Proteínas ras/análise
8.
J Biol Chem ; 278(30): 28167-72, 2003 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-12740392

RESUMO

Malignant transformation of cells causes disruption of cell-cell adhesion, enhancement of cell motility, and invasion into surrounding tissues. Nectins have both homophilic and heterophilic cell-cell adhesion activities and organize adherens junctions in cooperation with cadherins. We examined here whether Tage4, which was originally identified to be a gene overexpressed in colon carcinoma and has a domain structure similar to those of nectins, is involved in cell adhesion and/or migration. Tage4 heterophilically trans-interacted with nectin-3, but not homophilically with Tage4. Expression of Tage4 was markedly elevated in NIH3T3 cells transformed by an oncogenic Ki-Ras (V12Ras-NIH3T3 cells) as compared with that of wild-type NIH3T3 cells. trans-Interaction of Tage4 with nectin-3 enhanced motility of V12Ras-NIH3T3 cells. Tage4 did not bind afadin, a nectin- and actin filament-binding protein that connects nectins to the actin cytoskeleton and cadherins through catenins. Thus, Tage4 heterophilically trans-interacts with nectin-3 and regulates cell migration. Tage4 is tentatively re-named here nectin-like molecule-5 (necl-5) on the basis of its function and domain structure similar to those of nectins.


Assuntos
Antígenos de Neoplasias , Moléculas de Adesão Celular/metabolismo , Proteínas de Neoplasias , Células 3T3 , Animais , Adesão Celular , Movimento Celular , Clonagem Molecular , Citoesqueleto/metabolismo , DNA Complementar/metabolismo , Dimerização , Humanos , Cinesinas , Cinética , Camundongos , Proteínas dos Microfilamentos/química , Proteínas dos Microfilamentos/metabolismo , Miosinas , Nectinas , Filogenia , Plasmídeos/metabolismo , Ligação Proteica , Estrutura Terciária de Proteína , Proteínas Recombinantes/metabolismo , Ressonância de Plasmônio de Superfície , Fatores de Tempo , Transfecção
9.
Rapid Commun Mass Spectrom ; 17(10): 1071-8, 2003.
Artigo em Inglês | MEDLINE | ID: mdl-12720288

RESUMO

Improvement of in-gel digestion efficiency is highly desirable for one- or two-dimensional gel electrophoretic separation and mass spectrometric (MS) analysis in proteomics, because the resultant increases in sequence coverage and MS signal intensity lead to higher confidence in protein identification. Here an optimized in-gel digestion system, in combination with thin-gel separation and negative staining in a high-throughput format using 96-well plates, is described. The combination of negative staining and protein separation on a 0.9 mm thick gel showed a clear improvement in in-gel digestion efficiency in comparison with the more typical protocols such as the combination of silver staining and a 1.0 mm gel. In addition, the use of 96-well plates to increase throughput did not decrease the efficiency of this strategy when the stirring of the gel pieces in processes such as destaining, washing, gel-shrinking and peptide extraction was performed by sonication instead of shaking the plates. This procedure was optimized and applied to identify proteins of the postsynaptic density fraction; 105 proteins were identified after SDS-PAGE separation.


Assuntos
Proteínas/química , Animais , Química Encefálica , Eletroforese em Gel de Poliacrilamida , Géis , Hidrólise , Camundongos , Análise de Sequência de Proteína , Coloração pela Prata , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , beta-Galactosidase/química
10.
J Gastroenterol ; 38(1): 92-6, 2003.
Artigo em Inglês | MEDLINE | ID: mdl-12560929

RESUMO

Radical surgery for fulminant amoebic colitis leads to extremely high mortality; however, resective surgery is mandatory if a patient develops massive fecal peritonitis. We herein report an extremely rare case of fulminant amoebic colitis with multiple perforations, which was successfully treated by staged surgical procedures. A 48-year-old man who had been treated with predonisolone under a diagnosis of ulcerative colitis was admitted. Biopsy specimens from the colonic mucosa revealed Entamoeba histolytica. On the day of diagnosis, he developed severe abdominal pain and underwent emergency laparoptomy, showing total colonic gangrene with multiple perforations associated with massive fecal peritonitis. Subtotal colectomy, mucous fistula of the rectosigmoid, and ileostomy were performed. He recovered well although disseminated intravascular coagulopathy developed postoperatively. As the middle and upper part of rectum was found to be severely stenotic 4 months after surgery, we performed proctectomy, ileal pouch anal canal anastomosis, and diverting ileostomy, which was reversed 6 months later. The patient has been well with satisfactory anal function 37 months after the initial surgery. This case suggests that (1). early and accurate diagnosis of amoebiasis is important to avoid surgical intervention, and (2). staged surgery including total colectomy should be considered as one of the treatment choices even in patients with total necrotizing amoebic colitis.


Assuntos
Disenteria Amebiana/cirurgia , Perfuração Intestinal/cirurgia , Colectomia , Bolsas Cólicas , Disenteria Amebiana/complicações , Humanos , Ileostomia , Perfuração Intestinal/etiologia , Masculino , Pessoa de Meia-Idade , Reoperação
11.
J Cell Biol ; 158(3): 577-90, 2002 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-12163476

RESUMO

The cytomatrix at the active zone (CAZ) has been implicated in defining the site of Ca2+-dependent exocytosis of neurotransmitter. We have identified here a novel CAZ protein of approximately 120 kD from rat brain and named it CAST (CAZ-associated structural protein). CAST had no transmembrane segment, but had four coiled-coil domains and a putative COOH-terminal consensus motif for binding to PDZ domains. CAST was localized at the CAZ of conventional synapses of mouse brain. CAST bound directly RIM1 and indirectly Munc13-1, presumably through RIM1, forming a ternary complex. RIM1 and Munc13-1 are CAZ proteins implicated in Ca2+-dependent exocytosis of neurotansmitters. Bassoon, another CAZ protein, was also associated with this ternary complex. These results suggest that a network of protein-protein interactions among the CAZ proteins exists at the CAZ. At the early stages of synapse formation, CAST was expressed and partly colocalized with bassoon in the axon shaft and the growth cone. The vesicles immunoisolated by antibassoon antibody-coupled beads contained not only bassoon but also CAST and RIM1. These results suggest that these CAZ proteins are at least partly transported on the same vesicles during synapse formation.


Assuntos
Encéfalo/metabolismo , Citoplasma/metabolismo , Proteínas de Ligação ao GTP , Proteínas do Tecido Nervoso/isolamento & purificação , Proteínas do Tecido Nervoso/metabolismo , Neurônios/metabolismo , Membranas Sinápticas/metabolismo , Envelhecimento/metabolismo , Sequência de Aminoácidos/genética , Animais , Sequência de Bases/genética , Encéfalo/crescimento & desenvolvimento , Encéfalo/ultraestrutura , Compartimento Celular/fisiologia , Diferenciação Celular , Células Cultivadas , Clonagem Molecular , Citoplasma/ultraestrutura , DNA Complementar/análise , DNA Complementar/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Substâncias Macromoleculares , Proteínas de Membrana , Camundongos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/ultraestrutura , Neurônios/ultraestrutura , Ligação Proteica/fisiologia , Ratos , Membranas Sinápticas/ultraestrutura , Transmissão Sináptica/fisiologia , Sinaptofisina/metabolismo
12.
Genes Cells ; 7(2): 187-97, 2002 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-11895482

RESUMO

BACKGROUND: The postsynaptic density (PSD) at synapses is a specialized submembranous structure where neurotransmitter receptors are linked to cytoskeleton and signalling molecules. Activity-dependent dynamic change in the components of the PSD is a mechanism of synaptic plasticity. Identification of the PSD proteins and examination of their modulations dependent on synaptic activity will be valuable for an understanding of the molecular basis of learning and memory. RESULT: We attempted here to identify proteins in the PSD fraction by two-dimensional (2D) gel electrophoresis and mass spectrometry. About 1.7 x 103 protein spots were detected on 2D gels. A total of 90 spots were identified, containing 47 different protein species. In addition to previously identified PSD proteins such as PSD-95/SAP90, several new proteins were identified in the PSD fraction. They included stomatin-like protein 2 and NIPSNAP1. We also examined activity-dependent modulations of PSD proteins by 2D gel electrophoresis. The spot concentration of G protein beta subunit 5 and NIPSNAP1 increased 2 h after kainate treatment that caused generalized seizures. CONCLUSION: These results indicate that the combination of 2D gel electrophoresis and mass spectrometry is an excellent tool for the identification of activity-regulated PSD proteins.


Assuntos
Proteínas Sanguíneas/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Núcleosídeo-Difosfato Quinase , Proteínas/metabolismo , Sinapses/metabolismo , Fatores de Transcrição/metabolismo , Animais , Eletroforese em Gel Bidimensional , Peptídeos e Proteínas de Sinalização Intercelular , Espectrometria de Massas , Camundongos , Nucleosídeo NM23 Difosfato Quinases , Proibitinas , Prosencéfalo/metabolismo , Proteínas Repressoras , Frações Subcelulares/metabolismo
13.
Nihon Kokyuki Gakkai Zasshi ; 40(9): 771-6, 2002 Sep.
Artigo em Japonês | MEDLINE | ID: mdl-12607304

RESUMO

A 55-year-old woman visited our hospital for further examination of abnormal shadows on chest radiographs. Her routine chest radiograph showed two nodular shadows in the right lower lung field. A chest CT scan revealed other nodules, small patchy shadows in both lung fields, and enlargement of the mediastinal lymphnodes (#2, 3). Laboratory data showed polyclonal hyperimmunoglobulinemia. This case was initially considered on the basis of a transbronchial lung biopsy to be a plasma cell granuloma. However, serum gammaglobulin levels gradually increased, and video-assisted thoracoscopic surgery was performed to aid in making a definite diagnosis. Biopsy specimens revealed lymphoid follicles with plasma cells which were stained with both anti-kappa chain and anti-lambda chain antibodies. The patient was treated with prednisolone (50 mg/day), and the serum gammaglobulin level and the shadows on the chest CT were temporarily slightly improved. During the clinical course, her laboratory data and histological specimens were re-examined, and the final diagnosis was multicentric Castleman's disease.


Assuntos
Hiperplasia do Linfonodo Gigante/diagnóstico por imagem , Pulmão/diagnóstico por imagem , Hiperplasia do Linfonodo Gigante/diagnóstico , Feminino , Humanos , Pessoa de Meia-Idade , Tomografia Computadorizada por Raios X
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