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1.
Mo Med ; 111(3): 202-6, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25011341

RESUMO

Tourette Syndrome (TS) is recognized as a more common neurodevelopmental disorder than once thought. In this article we present an update on TS including the DSM-5 revised criteria, new findings in the genetics of TS, treatment advances such as new medications for tics and the use of new tools including Cognitive Behavioral Intervention for Tics (CBIT). We also explore supportive services for the ongoing care of patients using nursing education and family therapy.


Assuntos
Síndrome de Tourette/epidemiologia , Síndrome de Tourette/terapia , Terapia Cognitivo-Comportamental , Comorbidade , Exposição Ambiental , Família , Predisposição Genética para Doença , Humanos , Síndrome de Tourette/genética
2.
Pathol Biol (Paris) ; 58(5): 381-6, 2010 Oct.
Artigo em Francês | MEDLINE | ID: mdl-20181440

RESUMO

Autism is a neurodevelopmental disorder characterized by a deficit of language and communication both associated with a restricted repertoire of activities and interests. The current prevalence of autistic disorder stricto sensu is estimated at 1/500 whereas autism spectrum disorders (ASD) increases up to 1/150 to 1/200. Mental deficiency (MD) and epilepsy are present in numerous autistic individuals. Consequently, autism is as a major public health issue. Autism was first considered as a non biological disease; however various rational approaches for analysing epidemiological data suggested the possibility of the influence of genetic factors. In 2003, this hypothesis was clearly illustrated by the characterization of genetic mutations transmitted through a mendelian manner. Subsequently, the glutamate synapse appeared as a preferential causal target in autism because the identified genes encoded proteins present in this structure. Strikingly, the findings that an identical genetic dysfunction of the synapse might also explain some MD suggested the possibility of a genetic comorbidity between these neurodevelopmental conditions. To date, various identified genes are considered indifferently as "autism" or "MD" genes. The characterization of mutations in the NLGN4X gene in patients with Asperger syndrome, autism without MD, or MD without autism, was the first example. It appears that a genetic continuum between ASD on one hand, and between autism and MD on the other hand, is present. Consequently, it is likely that genes already involved in MD will be found mutated in autistic patients and will represent future target for finding new factors in autism.


Assuntos
Transtorno Autístico/genética , Transmissão Sináptica , Síndrome de Asperger/genética , Síndrome de Asperger/fisiopatologia , Astrócitos/patologia , Astrócitos/fisiologia , Transtorno Autístico/epidemiologia , Transtorno Autístico/fisiopatologia , Proteínas de Transporte/genética , Moléculas de Adesão Celular Neuronais , Criança , Transtornos Globais do Desenvolvimento Infantil/epidemiologia , Transtornos Globais do Desenvolvimento Infantil/genética , Comorbidade , Epilepsia/epidemiologia , Epilepsia/genética , Ácido Glutâmico/fisiologia , Humanos , Incidência , Deficiência Intelectual/epidemiologia , Deficiência Intelectual/genética , Proteínas de Membrana/deficiência , Proteínas de Membrana/genética , Receptores de Glutamato/genética , Receptores de Glutamato/fisiologia , Transmissão Sináptica/genética , Transmissão Sináptica/fisiologia
3.
Sci Rep ; 10(1): 6790, 2020 04 22.
Artigo em Inglês | MEDLINE | ID: mdl-32321933

RESUMO

Faba bean (Vicia faba L.) is a pulse crop of high nutritional value and high importance for sustainable agriculture and soil protection. With the objective of identifying gene-based SNPs, transcriptome sequencing was performed in order to reduce faba bean genome complexity. A set of 1,819 gene-based SNP markers polymorphic in three recombinant line populations was selected to enable the construction of a high-density consensus genetic map encompassing 1,728 markers well distributed in six linkage groups and spanning 1,547.71 cM with an average inter-marker distance of 0.89 cM. Orthology-based comparison of the faba bean consensus map with legume genome assemblies highlighted synteny patterns that partly reflected the phylogenetic relationships among species. Solid blocks of macrosynteny were observed between faba bean and the most closely-related sequenced legume species such as pea, barrel medic or chickpea. Numerous blocks could also be identified in more divergent species such as common bean or cowpea. The genetic tools developed in this work can be used in association mapping, genetic diversity, linkage disequilibrium or comparative genomics and provide a backbone for map-based cloning. This will make the identification of candidate genes of interest more efficient and will accelerate marker-assisted selection (MAS) and genomic-assisted breeding (GAB) in faba bean.


Assuntos
Mapeamento Cromossômico/métodos , Cromossomos de Plantas/genética , Melhoramento Vegetal/métodos , Polimorfismo de Nucleotídeo Único , Vicia faba/genética , Agricultura/métodos , Conservação dos Recursos Naturais/métodos , Fabaceae/classificação , Fabaceae/genética , Perfilação da Expressão Gênica/métodos , Marcadores Genéticos/genética , Genômica/métodos , Sintenia , Vicia faba/metabolismo
4.
Rev Mal Respir ; 36(3): 396-404, 2019 Mar.
Artigo em Francês | MEDLINE | ID: mdl-30902444

RESUMO

Airborne isolation is the main confinement measure used to limit human-to-human transmission of tuberculosis. If implemented early, precisely as soon as the patient is clinically diagnosed with tuberculosis, this measure will protect the population, particularly the health workers who are exposed. A patient suspected of being infected with tuberculosis can create a difficult situation if microbiological examination of his respiratory secretions is negative. This is a complex laboratory technique and sensitivity varies from one test to another. Thus, a false negative result is possible; meaning that a patient can have positive results on a microbiological culture performed later. This patient would still have low, but not no, contagiousness as long as a treatment has not been initiated. This situation can extend the period of respiratory isolation while further diagnostic investigations are carried out. This extended isolation can reduce the quality of health care delivered and patients can show signs of depression and anxiety. The use in routine clinical investigation of gene amplification tools should allow a rethinking of respiratory isolation rules. These tools, which are very sensitive and with a short reporting time, could drastically reduce the duration of respiratory isolation for patients suspected of being infected with tuberculosis.


Assuntos
Controle de Infecções/métodos , Respiração , Escarro/microbiologia , Tuberculose/diagnóstico , Tuberculose/terapia , Microbiologia do Ar , Ambiente Controlado , Reações Falso-Negativas , Humanos , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/isolamento & purificação , Isolamento de Pacientes/métodos , Saúde Pública/métodos
5.
Seizure ; 69: 41-43, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30959424

RESUMO

PURPOSE: The ketogenic diet has been found to be safe and effective in the treatment of drug resistant epilepsy in childhood. The age range of children undergoing this treatment has steadily been going down. There is strong evidence that it is a safe alternative in infants with drug resistant seizures. The American Academy of Pediatrics strongly supports continuing a breast milk diet until infants are at least six months of age. The purpose of this study is to evaluate the safety and efficacy of the ketogenic diet in infants while maintaining a breast milk diet. METHOD: This is a cohort study of 9 infants between the ages of 1 and 13 months with drug resistant epilepsy treated with the ketogenic diet while maintained on breast milk. The data from the first two patients was gathered retrospectively while the other seven were studied prospectively. RESULTS: We show that all nine infants achieved and maintained ketosis effectively. While one infant had no change in seizure frequency, three were seizure free at the first follow-up visit and four had a burden of seizure reduction greater than 50%. The diet was overall well tolerated, although one child required a hospital stay for dehydration and metabolic acidosis. CONCLUSION: The ketogenic diet can be safely and effectively initiated in infants while continuing human breast milk feedings.


Assuntos
Dieta Cetogênica , Epilepsia Resistente a Medicamentos/dietoterapia , Cetose/dietoterapia , Leite Humano , Convulsões/dietoterapia , Estudos de Coortes , Dieta Cetogênica/efeitos adversos , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Estudos Retrospectivos , Resultado do Tratamento
6.
Curr Biol ; 11(7): 524-8, 2001 Apr 03.
Artigo em Inglês | MEDLINE | ID: mdl-11413004

RESUMO

Left-hemiparetic patients show predominant postural imbalance as compared to right-hemiparetic patients. The right hemisphere is crucial for generating internal maps used for perceptual and premotor processing of spatial information. Predominant postural imbalance with right-brain damage could thus result from a distortion of an internal postural map. Well-known manifestations of distorted internal maps due to right-hemisphere lesions, such as hemineglect, may show improvement following prism adaptation shifting the visual field to the right. We therefore investigated the effect of prism adaptation on postural imbalance in left-hemiparetic patients. Three groups of five patients were either adapted to prisms deviating the visual field to the right or left or exposed to neutral prisms while performing reaching movements of the right arm. Postural imbalance was reduced only following prism adaptation to the right. Thus, brief adaptation (i.e., 3 min) to rightward-shifting prisms can dramatically improve postural imbalance. This result shows that the effect of exposure to prisms that horizontally shift the visual field to the right in a reaching task generalizes to the postural system, and it suggests an interaction between horizontal and vertical reference frames. This also supports the theory that predominant postural imbalance in patients with right-brain damage may be partly related to a distortion of an internal postural map.


Assuntos
Óculos , Hemiplegia/reabilitação , Postura , Adulto , Idoso , Dano Encefálico Crônico/diagnóstico , Feminino , Lateralidade Funcional , Hemiplegia/fisiopatologia , Humanos , Masculino , Pessoa de Meia-Idade , Óptica e Fotônica , Equilíbrio Postural , Comportamento Espacial , Campos Visuais/fisiologia
7.
Phys Rev E Stat Nonlin Soft Matter Phys ; 75(1 Pt 1): 011125, 2007 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17358128

RESUMO

Systems whose potential energies consists of pieces that scale as r;{-2} together with pieces that scale as r;{2} , show no violent relaxation to Virial equilibrium but may pulsate at considerable amplitude forever. Despite this pulsation these systems form lattices when the nonpulsational "energy" is low, and these disintegrate as that energy is increased. The "specific heats" show the expected halving as the "solid" is gradually replaced by the "fluid" of independent particles. The forms of the lattices are described here for N18 and they become hexagonal close packed for large N . In the larger N limit, a shell structure is formed. Their large N behavior is analogous to a gamma=53 polytropic fluid with a quasigravity such that every element of fluid attracts every other in proportion to their separation. For such a fluid, we study the "rotating pulsating equilibria" and their relaxation back to uniform but pulsating rotation. We also compare the rotating pulsating fluid to its discrete counterpart, and study the rate at which the rotating crystal redistributes angular momentum and mixes as a function of extra heat content.

8.
Prog Urol ; 17(3): 393-8, 2007 May.
Artigo em Francês | MEDLINE | ID: mdl-17622065

RESUMO

Lower urinary tract dysfunction is frequent in Parkinson's disease and other Parkinsonian syndromes and can cause urinary incontinence complicating a urgency-frequency syndrome or on the contrary, dysuria. These disorders are a frequent urological presenting complaint due to their impact on the patient's quality of life. Urologists must be aware of the different natural histories of diseases such as Parkinson's disease and Parkinsonian syndromes such as multisystem atrophy, which often have a severe course and are marked by resistance to neuropharmacological treatments. These various diseases can also directly induce urinary symptoms, independently of urological complications. Inversely, the development of urinary disorders, especially obstructive symptoms, in a patient with Parkinsonian syndrome may require review of the neurological diagnosis. Finally, therapeutic management is complex due to the difficulty of using pharmacological treatments, and the risk of deterioration after surgical treatment of obstructive uropathy.


Assuntos
Transtornos Parkinsonianos/fisiopatologia , Doenças Urológicas/etiologia , Humanos , Doença de Parkinson/complicações , Doença de Parkinson/fisiopatologia , Transtornos Parkinsonianos/complicações
9.
Cancer Res ; 54(8): 2102-7, 1994 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-7513610

RESUMO

Using whole viable human colon carcinoma HT29 cells as immunogen, we produced a monoclonal antibody (mAb) termed 69-6-5. The antibody was functionally selected on its anti-cell-spreading activity. By immunoprecipitation of surface radiolabeled cell lysates from HT29-D4 cells (an HT29 cell clone), mAb 69-6-5 recognized a molecular complex resembling integrin heterodimers. Sequential immunodepletions with mAb to the integrin alpha v subunit demonstrated that this complex was composed of alpha v-containing integrins. Accordingly, mAb 69-6-5 reacted with integrin alpha v beta 3 immunopurified from melanoma cells and integrins alpha v beta 5 and alpha v beta 6 immunopurified from pancreatic carcinoma cells. In cell adhesion assays, the 69-6-5 mAb was able to inhibit strongly in a dose-dependent manner arginine-glycine-aspartic acid-mediated adhesion of HT29-D4 cells to vitronectin, fibronectin, or ProNectin F but not to laminin or collagen. Immunoprecipitations with beta chain-specific antisera indicated that these cells express integrins alpha v beta 5 (receptor for vitronectin) and alpha v beta 6 (receptor for fibronectin) but neither alpha v beta 1 nor alpha v beta 3. In summary, these results indicated that mAb 69-6-5 reacts with several alpha v integrins and that it can effectively interfere with the adhesive functions of at least alpha v beta 5 and alpha v beta 6, which represent the major receptors on HT29-D4 cells responsible for their adhesion on vitronectin and fibronectin.


Assuntos
Anticorpos Monoclonais/farmacologia , Antígenos de Neoplasias , Adesão Celular/fisiologia , Fibronectinas , Glicoproteínas , Integrinas/fisiologia , Sequência de Aminoácidos , Adesão Celular/efeitos dos fármacos , Linhagem Celular , Movimento Celular , Neoplasias do Colo , Reações Cruzadas , Eletroforese em Gel de Poliacrilamida , Epitopos/química , Epitopos/imunologia , Humanos , Integrinas/análise , Integrinas/imunologia , Dados de Sequência Molecular , Peso Molecular , Receptores de Citoadesina/imunologia , Receptores de Citoadesina/fisiologia , Receptores de Vitronectina , Células Tumorais Cultivadas , Vitronectina
10.
Biochim Biophys Acta ; 475(2): 217-27, 1977 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-321026

RESUMO

The rate of individual ribosomal protein synthesis relative to total protein synthesis has been determined in Escherichia coli rel+ and rel- cells, under valyltRNA deprivation. These strains have a temperature-sensitive valyl-tRNA synthetase. Starvation was obtained following transfer to the cells to non-permissive temperature. Ribosomal proteins were obtained by treatment of either total lysates of freeze-thawed lysozyme spheroplasts or ammonium sulphate precipitate of ribosomes, with acetic acid. Differential labelling of the ribosomal proteins was observed in both strains: proteins from the rel+ strain appear more labelled than those from the rel- strain, the rate of labelling of individual proteins being about the same in both strains. Moreover ribosomal proteins were found as stable during starvation as total protein. It is thus concluded that in starving cells individual ribosomal proteins are not synthesized at equal rates. This indicates that the synthesis of ribosomal proteins is not only under the control of the rel gene.


Assuntos
Aminoácidos/metabolismo , Escherichia coli/metabolismo , Biossíntese de Proteínas , Proteínas Ribossômicas/biossíntese , Peso Molecular , Mutação , RNA de Transferência/metabolismo , Ribossomos/metabolismo , Temperatura , Valina/metabolismo
11.
Biochim Biophys Acta ; 475(2): 228-40, 1977 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-321027

RESUMO

The rate of synthesis of ribosomal proteins relative to that of total protein was measured at various times during recovery from arginine starvation in isogenic re+ and rel- strains of Escherichia coli K 12. Total ribosomal proteins are preferentially synthesized early during recovery. Higher rates of synthesis are obtained in the rel+ strain than in the rel- strain. Differential rates of synthesis of individual ribosomal proteins are observed at the various times studied. The rate of synthesis of individual proteins increases with time up to maximum values then the rates come down to values similar to those found in exponentially growing cells. The time of restart of synthesis of each protein has been estimated (1) by the time at which the maximum value is reached, and (2) by measuring the rate of synthesis at early time (3 min). Most ribosomal proteins behave similarlly in rel- and rel+ strains. Proteins have been listed from highly labelled (early proteins) to poorly labelled (late proteins). The significance of the order of restart is considered.


Assuntos
Aminoácidos/metabolismo , Escherichia coli/metabolismo , Biossíntese de Proteínas , Proteínas Ribossômicas/biossíntese , Arginina/metabolismo , Replicação do DNA , Cinética , Peso Molecular , Mutação , Ribossomos/metabolismo
12.
Eur J Cell Biol ; 64(1): 71-7, 1994 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-7957315

RESUMO

Nitric oxide (NO) and the NO generating agent nitroprusside (SNP), inhibit the binding of [125I] vasoactive intestinal peptide (VIP) to its receptor at the surface of IGR39 human melanoma cells. Cysteine (10 mM) increases the sensitivity of the system to SNP while N-acetylcysteine (10 mM) decreases it. The NO gas as well as SNP inhibits the [125I]VIP binding capacity. These observations sustain an effect of SNP-generated NO rather than an effect of the SNP molecule per se or the cyanoferrate portion of the molecule. The inhibitory effect of NO is time and concentration dependent and is fully reversible. Affinity constants of high and low affinity VIP receptors of SNP-treated IGR39 cells are not modified while maximal binding capacity (Bmax) of both receptor types are decreased to the same extent. Production of cGMP by SNP-treated cells is time and concentration dependent and the maximum amount of cGMP obtained reaches 13 times the basal level. The cAMP production is not affected by SNP. However, the SNP effects on the [125I]VIP binding are not mimicked by the membrane permeant cGMP analogs dibutyryl cGMP and 8-bromo cGMP even at concentrations as high as 0.5 mM. Taken altogether, these data demonstrate a regulatory action of NO on VIP binding capacity of IGR39 melanoma cells which is not cGMP mediated. They also evidence a new step which could be involved in the NO-VIP interaction.


Assuntos
GMP Cíclico/fisiologia , Melanoma/patologia , Óxido Nítrico/farmacologia , Nitroprussiato/farmacologia , Receptores de Peptídeo Intestinal Vasoativo/antagonistas & inibidores , Neoplasias Cutâneas/patologia , Peptídeo Intestinal Vasoativo/metabolismo , AMP Cíclico/fisiologia , GMP Cíclico/análogos & derivados , GMP Cíclico/farmacologia , Dibutiril GMP Cíclico/farmacologia , Humanos , Proteínas de Neoplasias/metabolismo , Ligação Proteica/efeitos dos fármacos , Receptores de Peptídeo Intestinal Vasoativo/metabolismo , Células Tumorais Cultivadas
13.
Eur J Cell Biol ; 59(2): 382-8, 1992 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-1337321

RESUMO

The effects of specific sulfhydryl reagents, N-ethylmaleimide (NEM), p-chloromercuribenzoic acid (PCMB) and 5-5'-dithiobis(2-nitrobenzoic acid) (DTNB), were tested on the vasoactive intestinal peptide (VIP) receptor binding capacity of the human superficial melanoma-derived IGR39 cells. On intact cell monolayers NEM and PCMB inhibit the specific [125I]VIP binding in a time and dose-dependent manner while DTNB has no effect at any concentration tested. Inhibitory effects of NEM and PCMB on high and low affinity VIP receptor are not identical. With NEM-treated cells, only low affinity sites remained accessible to the ligand. Their affinity constant is not modified. With PCMB-treated cells, the binding capacity of high affinity sites is reduced by 56% while the binding capacity of low affinity sites is not significantly affected. For both types of binding sites, the affinity constants remain in the same range of that of untreated cells. On cells made permeable by lysophosphatidylcholine, DTNB is able to inhibit the specific [125I]VIP binding in a time and dose-dependent manner. The three sulfhydryl reagents stabilize the preformed [125I]VIP receptor complex whose dissociation in the presence of native VIP is significantly reduced. Labeling of free SH groups with tritiated NEM after preincubation of cells with DTNB and VIP made possible the characterization of reacting SH groups which probably belong to the receptor. Taken together, these data allow us to define three classes of sulfhydryl groups. In addition, it is shown that high and low affinity sites have different sensibility to sulfhydryl reagents.


Assuntos
Receptores dos Hormônios Gastrointestinais/efeitos dos fármacos , Reagentes de Sulfidrila/farmacologia , Peptídeo Intestinal Vasoativo/metabolismo , Alquilação , Autorradiografia , Permeabilidade da Membrana Celular/fisiologia , Ácido Ditionitrobenzoico , Humanos , Radioisótopos do Iodo , Proteínas de Membrana/análise , Ensaio Radioligante , Receptores dos Hormônios Gastrointestinais/metabolismo , Receptores de Peptídeo Intestinal Vasoativo , Células Tumorais Cultivadas
14.
Eur J Cell Biol ; 46(3): 458-65, 1988 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-2846304

RESUMO

HT29-D4, a clone of the human colonic adenocarcinoma cell line (HT29), possesses at its cell surface specific binding sites for the vasoactive intestinal peptide (VIP) (KD = 0.5 nM). Their molecular weight was previously estimated to 117 kDa and 64 kDa. This clone underwent functional and structural differentiation when grown in a glucose-free galactose-containing medium. The [125I]VIP binding capacity of cells grown in this medium gradually declined while the cell density increased and reached a value close to zero when cell monolayer was able to form hemicysts. At this time, cells presented numerous tight junctions and desmosomes and a well organized brush border. Binding capacity could be recovered when the post-confluent monolayers were previously disaggregated with EDTA. Neither the affinity for VIP nor the molecular weight of the [125I]VIP cross-linked polypeptides were modified in these cells compared to cells grown in glucose-containing medium. However, surface receptor number of differentiated cells was twice that of undifferentiated cells. Leakproof differentiated cell monolayers grown on permeable substratum produced cAMP in response to VIP only when the peptide was present in the lower chamber of the culture wells. Taking these data altogether, we conclude that the localization of functional VIP receptors is restricted to the basolateral domain in differentiated post-confluent HT29-D4 cells.


Assuntos
Receptores dos Hormônios Gastrointestinais/metabolismo , Células Tumorais Cultivadas/metabolismo , Adenocarcinoma/metabolismo , Adenilil Ciclases/metabolismo , Diferenciação Celular , Linhagem Celular , Neoplasias do Colo/metabolismo , Humanos , Microscopia Eletrônica , Peso Molecular , Receptores de Peptídeo Intestinal Vasoativo , Células Tumorais Cultivadas/citologia , Peptídeo Intestinal Vasoativo/metabolismo
15.
J Invest Dermatol ; 111(4): 650-5, 1998 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-9764848

RESUMO

Data from the literature indicate that ICAM-1 molecules play an important role in keratinocyte interactions with lymphocytes via the lymphocyte function-associated-1 lymphocyte-adhesion molecule. We examined the role of beta1 integrins in keratinocyte-lymphocyte adhesion under different activation conditions. Among the beta1 integrins expressed on keratinocytes and lymphocytes detected by indirect immunofluorescence microscopy and flow cytofluorometry, primarily the alpha2 and the alpha3 subunits on both cell types were involved in keratinocyte-lymphocyte adhesion. Moreover, the highest adhesion level was observed when both cell types were activated by IFN-gamma for keratinocytes and phorbol 12-myristate 13-acetate for lymphocytes, suggesting that the former involved the protein kinase C pathway. Keratinocyte activation, characterized by the expression of ICAM-1, a decrease of beta1 integrins, and the absence of alpha5beta1 integrin, was required for optimal lymphocyte adhesion. Thus, beta1 integrins remaining at the surface of IFN-gamma-treated keratinocytes could be activated by this cytokine, and could synergize with ICAM-1 and lymphocyte function-associated-1 molecules to consolidate keratinocyte-lymphocyte adhesion.


Assuntos
Integrina beta1/farmacologia , Queratinócitos/citologia , Linfócitos/citologia , Adesão Celular/efeitos dos fármacos , Comunicação Celular/efeitos dos fármacos , Humanos , Interferon gama/farmacologia
16.
Matrix Biol ; 15(7): 503-7, 1997 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-9106161

RESUMO

Human fibroblasts with mutated type I collagen have marked defective adhesive capacities on exogenous type I collagen and exogenous fibronectin in comparison to normal fibroblasts. This defective cell adhesion could be partly explained by the decreased level of cell surface receptors of the beta 1-integrin family, i.e., the alpha 2 integrin subunit for type I collagen and the alpha 5 integrin subunit for fibronectin, observed in pathological fibroblasts. However, it appeared that the presence of altered collagen interfered both with fibronectin biosynthesis and with its surface expression. Using a binding assay on immobilized fibronectin, we demonstrated that the mutated collagen had a weaker binding to fibronectin. In addition, the pathological fibroblasts plated on a mixture of normal exogenous type I collagen and fibronectin exhibited the same maximal level of adhesion as control fibroblasts. These results indicate that fibroblasts with the mutated collagen exhibit a decreased binding to normal fibronectin, a modification of synthesis and surface expression of fibronectin, and, finally, altered adhesive capacities.


Assuntos
Colágeno/fisiologia , Fibroblastos/patologia , Fibronectinas/biossíntese , Adesão Celular/fisiologia , Colágeno/genética , Feminino , Feto , Fibronectinas/fisiologia , Humanos , Mutação
17.
Biochimie ; 70(10): 1311-22, 1988 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-2852963

RESUMO

Vasoactive intestinal peptide (VIP) is a neuropeptide with a broad range of biological activities in various tissues. After interaction with its membrane receptor, VIP generally induces a very large increase in the intracellular cyclic AMP level. Receptors for VIP have been described in numerous tissues and cell lines. The first results on VIP receptor structure have been obtained by covalent cross-linking using bifunctional reagents. The molecular mass of the different components characterized in this way differs greatly according to the species and the tissue used. This heterogeneity may reflect either a difference in the length of the cross-linked polypeptide backbone or differently glycosylated forms of the same polypeptide. The VIP binding site of intact human adenocarcinoma cells (HT29 cells) is an Mr 64,000 glycoprotein with 20kDa of N-linked oligosaccharide side chains containing sialic acid. The structure of the VIP binding site from HT29 cell is compared, first to the structure of the VIP receptor from other tissues, particularly that from rat liver, and second to the structure of the hepatic glucagon binding site. Recently, solubilization of the VIP receptor in an active form has provided a new way of studying this receptor. The HT29 cell line is an appropriate model to study the dynamics of the VIP receptor. After binding to its receptor, VIP is rapidly internalized, probably by receptor-mediated endocytosis. This internalization leads to a decrease in the cell surface receptor number and simultaneously to a homologous desensitization of adenylate cyclase. VIP is then degraded in the lysosomes, while most of the receptors are recycled back to the cell surface. The presence of an intracellular pool of unoccupied VIP receptors has been demonstrated after inactivation of the cell surface receptors by chymotrypsin. The kinetics of the receptor reappearance at the cell surface, after inactivation by chymotrypsin or after receptor-mediated endocytosis, indicate 2 possible intracellular pathways for occupied and unoccupied VIP receptors.


Assuntos
Receptores dos Hormônios Gastrointestinais/análise , Peptídeo Intestinal Vasoativo/metabolismo , Acetilglucosaminidase/metabolismo , Animais , Invaginações Revestidas da Membrana Celular , Endocitose , Glucagon/metabolismo , Humanos , Lisossomos/metabolismo , Manosil-Glicoproteína Endo-beta-N-Acetilglucosaminidase , Peso Molecular , Receptores dos Hormônios Gastrointestinais/fisiologia , Receptores de Peptídeo Intestinal Vasoativo , Sialoglicoproteínas/análise
18.
Biochimie ; 70(5): 663-71, 1988 May.
Artigo em Inglês | MEDLINE | ID: mdl-2844304

RESUMO

HT 29, a cell line derived from a human colonic adenocarcinoma, is highly responsive to the vasoactive intestinal peptide (VIP) as shown by a more than 100-fold intracellular cAMP increase (Ka = 0.3 nM), the stimulations of protein kinase A (Ka = 0.1 nM) and the low-Km cAMP phosphodiesterase (Ka = 40 nM). Remarkably, adenylate cyclase, cAMP-dependent kinase and cAMP-specific phosphodiesterase are activated in a sequential manner. Binding studies with [125I]-labeled VIP indicate a high affinity site with a Kd value (0.5 nM) close to the activation constant value (Ka) of the three enzymes. The molecular structure of the VIP receptor was studied by immunological and chemical approaches. A monoclonal antibody (mAb 109-10-16) which partially decreased the binding of VIP to its receptor allowed the characterization of Mr = 53,000 and Mr = 48-49,000 polypeptides. More precise identification of protein components of the VIP receptor resulted from covalent cross-linking on intact HT 29 cells by four bifunctional reagents: dithiobis-(succinimidyl propionate) and its non-cleavable analog disuccinimidyl suberate, the photoactivable azido phenyl glyoxal and dimethylpimelimidate. Analysis by SDS-polyacrylamide gel electrophoresis demonstrated a major band of Mr = 67,000 regardless of which cross-linker was used. The same band and an Mr = 49,000 species were found in experiments using a crude membrane fraction of HT 29 cells. Assuming one molecule of VIP (Mr = 3326) linked per polypeptide, these observations suggest that an Mr = 64,000 species belongs to the VIP specific plasma membrane receptor. This protein contains an Mr = 20,000 N-linked sialic acid rich oligosaccharidic moiety.(ABSTRACT TRUNCATED AT 250 WORDS)


Assuntos
Adenocarcinoma/metabolismo , Neoplasias do Colo/metabolismo , Receptores dos Hormônios Gastrointestinais/metabolismo , Células Tumorais Cultivadas/metabolismo , Peptídeo Intestinal Vasoativo/farmacologia , 3',5'-AMP Cíclico Fosfodiesterases/metabolismo , Adenilil Ciclases/metabolismo , AMP Cíclico/biossíntese , AMP Cíclico/farmacologia , Eletroforese em Gel de Poliacrilamida , Ativação Enzimática/efeitos dos fármacos , Humanos , Imunoensaio , Lipídeos de Membrana/fisiologia , Proteínas Quinases/metabolismo , Receptores de Peptídeo Intestinal Vasoativo , Peptídeo Intestinal Vasoativo/metabolismo
19.
J Immunol Methods ; 178(1): 41-51, 1995 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-7829864

RESUMO

A simple and sensitive cell-cell adhesion microplate assay was established using the cytoplasmic fluorescent dye, calcein AM. The procedure involves three steps: the labeling of lymphocytes with an adequate concentration of calcein AM (20 microM) during a short incubation period (30 min); the adhesion of 2 x 10(5) labeled lymphocytes per well to confluent keratinocyte or fibroblast monolayers grown in microtiter plates for 90 min; and, finally, measurement of the fluorescent signal utilizing a new system of cold-light microfluorimetry (Rat, 1993). During the adhesion assay, the release of calcein from labeled lymphocytes is low and the method permits the detection of as few as 1000 adherent cells. This non-radioactive procedure takes less than 4 h to perform and has proven to be as accurate and reliable as the common method using radioactive isotopes. In addition to its simplicity, the use of a fluorescent molecular probe in conjunction with cold-light microfluorimetry (CLF) offers many advantages of safety and economy, and can readily be adapted to the different cell types that participate in cell-cell adhesion.


Assuntos
Adesão Celular , Fluorometria/métodos , Células Cultivadas , Técnicas Citológicas , Fibroblastos/fisiologia , Fluoresceínas , Corantes Fluorescentes , Humanos , Marcação por Isótopo , Queratinócitos/fisiologia , Linfócitos/fisiologia
20.
Thromb Haemost ; 54(3): 635-8, 1985 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-3841420

RESUMO

Heparin cofactor II (HC II) is a heparin-dependent inhibitor of thrombin, distinct from antithrombin III (AT III). This study was designed to evaluate its metabolism in healthy subjects. Purified HC II was labelled with 125I by the lactoperoxidase-glucose oxidase technique. The biological activity of the HC II was unchanged after labelling as was its migratory pattern by crossed immunoelectrophoresis in the presence of heparin or dermatan sulfate. Three healthy volunteers were injected with 10 microCi and the plasma radioactivity was measured daily. The data were approximated by a sum of two exponential terms and the metabolism of HC II was described by a two compartment mamillary system. The mean values of fractional catabolic rate, intravascular fraction and half-life of the elimination phase were respectively: 0.44 d-1, 0.60 and 2.53 d. These parameters are of the same order of magnitude as those reported in the literature for AT III. The plasma HC II concentration in the 3 subjects ranged from 61 to 82 micrograms/ml as estimated using our purified preparation. Accordingly, the absolute catabolic rate ranged from 1.17 to 1.36 mg X kg-1 X d-1.


Assuntos
Antitrombinas/metabolismo , Glicoproteínas/metabolismo , Adulto , Cofator II da Heparina , Humanos , Masculino , Taxa de Depuração Metabólica
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