Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 25
Filtrar
Mais filtros










Intervalo de ano de publicação
1.
BMC Med Res Methodol ; 22(1): 229, 2022 08 16.
Artigo em Inglês | MEDLINE | ID: mdl-35971088

RESUMO

An increasing number of large-scale multi-modal research initiatives has been conducted in the typically developing population, e.g. Dev. Cogn. Neur. 32:43-54, 2018; PLoS Med. 12(3):e1001779, 2015; Elam and Van Essen, Enc. Comp. Neur., 2013, as well as in psychiatric cohorts, e.g. Trans. Psych. 10(1):100, 2020; Mol. Psych. 19:659-667, 2014; Mol. Aut. 8:24, 2017; Eur. Child and Adol. Psych. 24(3):265-281, 2015. Missing data is a common problem in such datasets due to the difficulty of assessing multiple measures on a large number of participants. The consequences of missing data accumulate when researchers aim to integrate relationships across multiple measures. Here we aim to evaluate different imputation strategies to fill in missing values in clinical data from a large (total N = 764) and deeply phenotyped (i.e. range of clinical and cognitive instruments administered) sample of N = 453 autistic individuals and N = 311 control individuals recruited as part of the EU-AIMS Longitudinal European Autism Project (LEAP) consortium. In particular, we consider a total of 160 clinical measures divided in 15 overlapping subsets of participants. We use two simple but common univariate strategies-mean and median imputation-as well as a Round Robin regression approach involving four independent multivariate regression models including Bayesian Ridge regression, as well as several non-linear models: Decision Trees (Extra Trees., and Nearest Neighbours regression. We evaluate the models using the traditional mean square error towards removed available data, and also consider the Kullback-Leibler divergence between the observed and the imputed distributions. We show that all of the multivariate approaches tested provide a substantial improvement compared to typical univariate approaches. Further, our analyses reveal that across all 15 data-subsets tested, an Extra Trees regression approach provided the best global results. This not only allows the selection of a unique model to impute missing data for the LEAP project and delivers a fixed set of imputed clinical data to be used by researchers working with the LEAP dataset in the future, but provides more general guidelines for data imputation in large scale epidemiological studies.


Assuntos
Transtorno Autístico , Transtorno Autístico/genética , Teorema de Bayes , Criança , Coleta de Dados/métodos , Humanos
2.
Neuroimage ; 229: 117713, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33421594

RESUMO

How spontaneously fluctuating functional magnetic resonance imaging (fMRI) signals in different brain regions relate to behaviour has been an open question for decades. Correlations in these signals, known as functional connectivity, can be averaged over several minutes of data to provide a stable representation of the functional network architecture for an individual. However, associations between these stable features and behavioural traits have been shown to be dominated by individual differences in anatomy. Here, using kernel learning tools, we propose methods to assess and compare the relation between time-varying functional connectivity, time-averaged functional connectivity, structural brain data, and non-imaging subject behavioural traits. We applied these methods to Human Connectome Project resting-state fMRI data to show that time-varying fMRI functional connectivity, detected at time-scales of a few seconds, has associations with some behavioural traits that are not dominated by anatomy. Despite time-averaged functional connectivity accounting for the largest proportion of variability in the fMRI signal between individuals, we found that some aspects of intelligence could only be explained by time-varying functional connectivity. The finding that time-varying fMRI functional connectivity has a unique relationship to population behavioural variability suggests that it might reflect transient neuronal communication fluctuating around a stable neural architecture.


Assuntos
Comportamento/fisiologia , Encéfalo/fisiologia , Conectoma/métodos , Individualidade , Imageamento por Ressonância Magnética/métodos , Rede Nervosa/fisiologia , Encéfalo/diagnóstico por imagem , Humanos , Rede Nervosa/diagnóstico por imagem
3.
Reprod Domest Anim ; 53(3): 733-741, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29602187

RESUMO

Metformin is clinically used to treat diabetes. Given its role-impacting metabolism, metformin has been also added to semen cryopreservation media showing specie-dependent effects. We aimed to investigate metformin effects in both fresh (38.5°C for 2, 24 hr) and refrigerated (17°C for 10 days) boar spermatozoa. Metformin (2 hr) does not affect fresh sperm viability, membrane lipid organization nor acrosome integrity. However, metformin (24 hr) blocks sperm ΔΨm and significantly reduces % motile spermatozoa (65%), % progressive spermatozoa (50%), % rapid (100%), velocities VCL (69%), VSL (86%), VAP (78%) and motility coefficients. Metformin-including extender does not modify sperm viability, membrane lipid organization or acrosome integrity. Furthermore, it significantly reduces high ΔΨ-population spermatozoa at refrigeration day 4. Metformin also significantly reduces sperm motility during refrigeration. Summarizing, metformin inhibits both boar sperm ΔΨ and motility in any sperm condition studied: fresh and refrigerated. These findings dissuade metformin as an additive to improve boar sperm quality.


Assuntos
Metformina/farmacologia , Preservação do Sêmen/veterinária , Motilidade dos Espermatozoides/efeitos dos fármacos , Sus scrofa/fisiologia , Acrossomo/efeitos dos fármacos , Animais , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Refrigeração/veterinária , Análise do Sêmen/veterinária , Preservação do Sêmen/métodos , Espermatozoides/efeitos dos fármacos
4.
Andrology ; 5(6): 1131-1140, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28980456

RESUMO

AMP-activated kinase (AMPK) plays a key function in maintaining cellular energy homeostasis. We recently identified and localized AMPK protein in human spermatozoa and showed that inhibition of AMPK activity significantly modified human sperm motility. Recently, AMPK has gained great relevance as prime target for pharmacological approaches in several energy-related pathologies and therefore pharmacological research is aimed to develop direct AMPK-activating compounds such as A769662. Our aim was to investigate the effect of A769662 in essential functional processes of human spermatozoa. Human spermatozoa were incubated in the presence or absence of the AMPK activator A769662 for different incubation times (0-20 h) and motility was evaluated by CASA system whereas other functional parameters were evaluated by flow cytometry. A769662 treatment significantly reduces the percentages of motile, progressive, and rapid spermatozoa starting at 2 h. Moreover, AMPK activator in human spermatozoa causes a significant reduction in any velocity measured, which is concomitant to a significant decrease in the percentage of rapid spermatozoa, both at short- (2-3 h) and long-time treatment (20 h). Treatment of human spermatozoa with A769662 does not significantly alter any of the following functional parameters: sperm viability, mitochondrial membrane potential, phosphatidylserine translocation to the outer leaf of plasma membrane, acrosome membrane integrity, or mitochondrial superoxide anion production. In summary, our results suggest that AMPK in human spermatozoa contributes to the regulation of sperm motility, without affecting basic physiological parameters of human spermatozoa (viability, mitochondrial membrane potential or reactive oxygen species production, acrosome membrane integrity, phosphatidylserine exposure at plasma membrane). As sperm motility is required in the female reproductive tract to achieve fertilization, we conclude that AMPK is an essential regulatory kinase of the human spermatozoa function. This conclusion needs to be taken into account when AMPK is elected as prime target in pharmacological approaches for several energy-related pathologies.


Assuntos
Pironas/farmacologia , Motilidade dos Espermatozoides/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , Tiofenos/farmacologia , Proteínas Quinases Ativadas por AMP/efeitos dos fármacos , Compostos de Bifenilo , Regulação para Baixo , Humanos , Masculino , Espermatozoides/enzimologia
5.
Theriogenology ; 85(1): 12-20, 2016 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-26074068

RESUMO

Detailed molecular mechanisms mediating signal transduction cascades that regulate boar sperm function involving Ser/Thr and tyrosine phosphorylation of proteins have been reviewed previously. Therefore, this review will focus in those kinase pathways identified recently (<10 years) in boar spermatozoa that regulate different functional spermatozoa processes. AMP-activated protein kinase (AMPK) is a cell energy sensor kinase that was first identified in mammalian spermatozoa in 2012, and since then it has emerged as an essential regulator of boar sperm function. Signaling pathways leading to AMPK activation in boar sperm are highlighted in this review (PKA, CaMKKα/ß, and PKC as well as Ca(2+) and cAMP messengers as upstream regulators). Interestingly, stimuli considered as cell stress (hyperosmotic stress, inhibition of mitochondrial activity, absence of intracellular Ca(2+)) markedly activate AMPK in boar spermatozoa. Moreover, AMPK plays a remarkable and necessary regulatory role in mammalian sperm function, controlling essential boar sperm functional processes such as motility, viability, mitochondrial membrane potential, organization and fluidity of plasma membrane, and outer acrosome membrane integrity. These mentioned processes are all required under fluctuating environment of spermatozoa when transiting through the female reproductive tract to achieve fertilization. An applied role of AMPK in artificial insemination techniques is also suggested as during boar seminal doses preservation at 17 °C, physiological levels of AMPK activity markedly increase (maximum on Day 7) and result essential to maintain the aforementioned fundamental sperm processes. Moreover, regulation of sperm function exerted by the glycogen synthase kinase 3 and Src family kinase pathways is summarized.


Assuntos
Regulação da Expressão Gênica/fisiologia , Espermatozoides/fisiologia , Suínos/fisiologia , Animais , Masculino , Motilidade dos Espermatozoides/fisiologia
6.
Mol Hum Reprod ; 21(1): 31-45, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25281642

RESUMO

We recently demonstrated that AMPK inhibition in spermatozoa regulates motility, plasma membrane organization, acrosome integrity and mitochondrial membrane potential. As AMPK activity varies in different energy conditions induced by sperm environment, this work investigates the functional effects of AMPK activation in boar spermatozoa. Spermatozoa were incubated under non-stimulating (TBM) or Ca(2+) and [Formula: see text]-stimulating (TCM) media in the presence/absence of AMPK activator, A769662, for different times. AMPK activity, evaluated as Thr(172) phosphorylation by western blot, is effectively increased by A769662 in spermatozoa. AMPK activation significantly reduces the percentage of motile spermatozoa under Ca(2+) and/or [Formula: see text]-stimulating conditions. Moreover, AMPK activation in spermatozoa incubated in TBM or TCM significantly reduces curvilinear VCL, straight-line VSL and average VAP velocities, which subsequently lead to a significant decrease in the percentage of rapid spermatozoa (VAP > 80 µm/s). The effect of AMPK activation on motility is intensified by the absence of BSA in the incubation medium. AMPK activation for a short time prevents the decline in cell viability and in the sperm population displaying high mitochondrial membrane potential which is induced by Ca(2+) and [Formula: see text]. Sustained (24 h) AMPK activation under TBM or TCM significantly increases both lipid disorganization and phosphatidylserine externalization in the sperm plasma membrane, and diminishes the acrosome membrane integrity. In summary, AMPK activation modifies essential sperm processes such as motility, viability, mitochondrial membrane potential, acrosome membrane integrity, and organization and fluidity of plasma membrane. As these spermatozoa processes are required under different environmental conditions when transiting through the female reproductive tract to achieve fertilization, we conclude that balanced levels of AMPK activity are essential for regulating sperm function.


Assuntos
Adenilato Quinase/metabolismo , Potencial da Membrana Mitocondrial/fisiologia , Motilidade dos Espermatozoides/fisiologia , Espermatozoides/metabolismo , Animais , Compostos de Bifenilo , Movimento Celular/efeitos dos fármacos , Movimento Celular/fisiologia , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Fosforilação , Pironas/farmacologia , Motilidade dos Espermatozoides/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , Sus scrofa , Tiofenos/farmacologia
7.
Neural Comput ; 26(6): 1108-27, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24684452

RESUMO

We consider the problem of multiclass adaptive classification for brain-computer interfaces and propose the use of multiclass pooled mean linear discriminant analysis (MPMLDA), a multiclass generalization of the adaptation rule introduced by Vidaurre, Kawanabe, von Bünau, Blankertz, and Müller (2010) for the binary class setting. Using publicly available EEG data sets and tangent space mapping (Barachant, Bonnet, Congedo, & Jutten, 2012) as a feature extractor, we demonstrate that MPMLDA can significantly outperform state-of-the-art multiclass static and adaptive methods. Furthermore, efficient learning rates can be achieved using data from different subjects.


Assuntos
Mapeamento Encefálico , Interfaces Cérebro-Computador , Encéfalo/fisiologia , Processamento de Sinais Assistido por Computador , Algoritmos , Bases de Dados Factuais , Análise Discriminante , Eletroencefalografia , Humanos
8.
Anim Reprod Sci ; 139(1-4): 109-14, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23660365

RESUMO

During boar semen liquid preservation, extender is one of the factors that influence storage tolerance of spermatozoa. However, there are few studies about intra-breed variation in the preservation of semen quality during storage in different extenders. Similarly, boar breed is generally not considered a possible factor influencing variation in the semen storage tolerance in a particular extender. The aim of this study was to compare boar semen storage potential, in terms of the ability to maintain sperm viability and motility, of two currently used long-term extenders, MR-A and XCell. Extended semen from two breeds, Iberian and Duroc that had been stored at 17°C for up to 7 days was used. Intra- and inter-breed effect was studied. On Days 1, 4 and 7 (Day 0=day of semen collection), motility parameters and the percentage of total motile sperm and progressively motile sperm using a CASA system was evaluated. Viability (SYBR-14/PI) was evaluated by flow cytometry. Within each breed and for each storage day, there were differences between extenders, although semen tolerance to preservation was more influenced by the extender in the Iberian than in the Duroc breed. Neither breed nor extender influenced the percentage of viable spermatozoa during the storage time. Moreover, differences in motility parameters were observed between breeds, although the differences were greater when the XCell extender was used. In conclusion, both extender and breed influence motility characteristics of liquid-stored boar semen, so both aspects have to be considered in the design of comparative studies about stored boar semen quality from different breeds or with different extenders. Further studies are needed to corroborate these findings.


Assuntos
Análise do Sêmen/veterinária , Preservação do Sêmen/veterinária , Espermatozoides/fisiologia , Suínos/fisiologia , Animais , Citometria de Fluxo/veterinária , Masculino , Preservação do Sêmen/métodos , Estatísticas não Paramétricas
9.
Neural Comput ; 24(11): 2900-23, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22845827

RESUMO

We introduce a probabilistic model that combines a classifier with an extra reinforcement signal (RS) encoding the probability of an erroneous feedback being delivered by the classifier. This representation computes the class probabilities given the task related features and the reinforcement signal. Using expectation maximization (EM) to estimate the parameter values under such a model shows that some existing adaptive classifiers are particular cases of such an EM algorithm. Further, we present a new algorithm for adaptive classification, which we call constrained means adaptive classifier, and show using EEG data and simulated RS that this classifier is able to significantly outperform state-of-the-art adaptive classifiers.


Assuntos
Interfaces Cérebro-Computador , Modelos Neurológicos , Modelos Estatísticos , Algoritmos , Inteligência Artificial , Humanos , Reconhecimento Automatizado de Padrão , Reforço Psicológico , Processamento de Sinais Assistido por Computador
10.
Neural Netw ; 24(10): 1120-7, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-21696919

RESUMO

We propose an adaptive classification method for the Brain Computer Interfaces (BCI) which uses Interaction Error Potentials (IErrPs) as a reinforcement signal and adapts the classifier parameters when an error is detected. We analyze the quality of the proposed approach in relation to the misclassification of the IErrPs. In addition we compare static versus adaptive classification performance using artificial and MEG data. We show that the proposed adaptive framework significantly improves the static classification methods.


Assuntos
Inteligência Artificial , Encéfalo/fisiologia , Potenciais Evocados/fisiologia , Neurorretroalimentação/fisiologia , Interface Usuário-Computador , Adaptação Fisiológica/fisiologia , Algoritmos , Auxiliares de Comunicação para Pessoas com Deficiência , Eletroencefalografia/métodos , Humanos , Magnetoencefalografia/métodos , Neurorretroalimentação/métodos , Reconhecimento Automatizado de Padrão/métodos , Estimulação Luminosa/métodos , Desempenho Psicomotor/fisiologia , Tempo de Reação/fisiologia , Processamento de Sinais Assistido por Computador , Software
11.
Clin Vaccine Immunol ; 16(9): 1352-9, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19641100

RESUMO

Culture filtrate and cell extracts from Mycobacterium bovis cultures contain molecules which could promote protective immunity to tuberculosis in animals. Different protein fractions of M. bovis cultures were obtained by elution electrophoresis and were tested in experimentally infected cattle. The fractions that elicited gamma interferon (IFN-gamma) responses were resolved by two-dimensional gel electrophoresis, and individual proteins were identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry. The open reading frames were cloned, expressed as their recombinant forms, and retested with naturally and experimentally infected animals. Eleven protein fractions were highly reactive, from which the Rv1636, HspX, Rv0138, Rv2524, EsxI, and Rv3740 recombinant proteins were obtained. EsxI and HspX were the antigens most recognized by the IFN-gamma release assay. In summary, a proteomic approach allowed the identification of novel antigens useful for the diagnosis of bovine tuberculosis.


Assuntos
Antígenos de Bactérias/análise , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/análise , Proteínas de Bactérias/imunologia , Mycobacterium bovis/química , Mycobacterium bovis/imunologia , Animais , Bovinos , Eletroforese em Gel Bidimensional , Imunoensaio , Interferon gama/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Tuberculose Bovina/diagnóstico
12.
J Biol Chem ; 276(10): 7312-9, 2001 Mar 09.
Artigo em Inglês | MEDLINE | ID: mdl-11036086

RESUMO

CD69, one of the earliest specific antigens acquired during lymphoid activation, acts as a signal-transducing receptor involved in cellular activation events, including proliferation and induction of specific genes. CD69 belongs to a family of receptors that modulate the immune response and whose genes are clustered in the natural killer (NK) gene complex. The extracellular portion of these receptors represent a subfamily of C-type lectin-like domains (CTLDs), which are divergent from true C-type lectins and are referred to as NK-cell domains (NKDs). We have determined the three-dimensional structure of human CD69 NKD in two different crystal forms. CD69 NKD adopts the canonical CTLD fold but lacks the features involved in Ca(2+) and carbohydrate binding by C-type lectins. CD69 NKD dimerizes noncovalently, both in solution and in crystalline state. The dimer interface consists of a hydrophobic, loosely packed core, surrounded by polar interactions, including an interdomain beta sheet. The intersubunit core shows certain structural plasticity that may facilitate conformational rearrangements for binding to ligands. The surface equivalent to the binding site of other members of the CTLD superfamily reveals a hydrophobic patch surrounded by conserved charged residues that probably constitutes the CD69 ligand-binding site.


Assuntos
Antígenos CD/química , Antígenos de Diferenciação de Linfócitos T/química , Lectinas/química , Sequência de Aminoácidos , Animais , Sítios de Ligação , Divisão Celular , Cristalografia por Raios X , Dimerização , Elétrons , Humanos , Células Matadoras Naturais/metabolismo , Lectinas Tipo C , Ligantes , Modelos Moleculares , Dados de Sequência Molecular , Plasmídeos/metabolismo , Ligação Proteica , Dobramento de Proteína , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Ratos , Proteínas Recombinantes/metabolismo , Homologia de Sequência de Aminoácidos , Transdução de Sinais
13.
Annu Rev Immunol ; 17: 435-66, 1999.
Artigo em Inglês | MEDLINE | ID: mdl-10358765

RESUMO

Superantigens (SAGs) are a class of immunostimulatory and disease-causing proteins of bacterial or viral origin with the ability to activate large fractions (5-20%) of the T cell population. Activation requires simultaneous interaction of the SAG with the V beta domain of the T cell receptor (TCR) and with major histocompatibility complex (MHC) class II molecules on the surface of an antigen-presenting cell. Recent advances in knowledge of the three-dimensional structure of bacterial SAGs, and of their complexes with MHC class II molecules and the TCR beta chain, provide a framework for understanding the molecular basis of T cell activation by these potent mitogens. These structures along with those of TCR-peptide/MHC complexes reveal how SAGs circumvent the normal mechanism for T cell activation by peptide/MHC and how they stimulate T cells expressing TCR beta chains from a number of different families, resulting in polyclonal T cell activation. The crystal structures also provide insights into the basis for the specificity of different SAGs for particular TCR beta chains, and for the observed influence of the TCR alpha chain on SAG reactivity. These studies open the way to the design of SAG variants with altered binding properties for TCR and MHC for use as tools in dissecting structure-activity relationships in this system.


Assuntos
Ativação Linfocitária/fisiologia , Superantígenos/administração & dosagem , Linfócitos T/imunologia , Sequência de Aminoácidos , Animais , Doenças Autoimunes/imunologia , Doenças Transmitidas por Alimentos/imunologia , Antígenos de Histocompatibilidade Classe II/química , Antígenos de Histocompatibilidade Classe II/metabolismo , Humanos , Imunoterapia , Cinética , Substâncias Macromoleculares , Modelos Moleculares , Dados de Sequência Molecular , Conformação Proteica , Receptores de Antígenos de Linfócitos T alfa-beta/química , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Choque Séptico/imunologia , Superantígenos/química , Superantígenos/metabolismo
14.
J Immunol ; 162(10): 6040-5, 1999 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-10229844

RESUMO

The affinities (Ka) and association rate constants (kon) of 23 mouse (BALB/c) anti-lysozyme mAbs obtained after short and prolonged immunizations have been measured by plasmon resonance techniques. The affinities for the 23 Abs, measured using their Fab, range from Ka = 1.1 x 10(7) to 1.4 x 10(10) M-1. There is no significant correlation between time or dose of immunization and affinity or association rates, indicating no time- or dose-dependent maturation of the response within the doses and times that were explored. IgMs are produced early and late in the response, with intrinsic affinities <10(5) M-1. Two independently derived mAbs, D44.1 (short term) and F10.6.6 (from a longer term response), result from identical or nearly identical somatic recombination events of germline gene segments. F10.6.6 has more mutations and a higher affinity constant (Ka = 1.4 x 10(10) M-1) than D44.1 (Ka = 1.1 x 10(7) M-1). Although higher affinities may result from an accumulation of mutations, they do not correlate with the length and dose of immunogenic challenge.


Assuntos
Anticorpos Monoclonais/imunologia , Afinidade de Anticorpos , Muramidase/imunologia , Animais , Galinhas , Fragmentos Fab das Imunoglobulinas/imunologia , Imunoglobulina G/imunologia , Imunoglobulina M/imunologia , Cinética , Camundongos , Camundongos Endogâmicos BALB C , Ressonância de Plasmônio de Superfície , Fatores de Tempo
15.
Immunol Rev ; 163: 177-86, 1998 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-9700510

RESUMO

Superantigens (SAGs) are a class of disease-causing and immunostimulatory proteins of bacterial or viral origin that activate T cells by binding to the V beta domain of the T-cell antigen receptor (TCR). The three-dimensional structure of the complex between a TCR beta chain (mouse V beta 8.2-J beta 2.1-C beta 1) and the SAG staphylococcal enterotoxin C3 (SEC3) has been recently determined. The complementarity-determining region 2 (CDR2) of the beta chain and, to lesser extents, CDR1 and hypervariable region 4 (HV4) bind in a cleft between the small and large domains of the SAG. A model of the TCR-SAG-peptide/MHC complex constructed from available crystal structures reveals how the SAG acts as a wedge between the TCR and MHC, thereby displacing the antigenic peptide away from the TCR and circumventing the normal mechanism for T-cell activation by peptide/MHC. To evaluate the actual contribution of individual SAG residues to stabilizing the V beta C beta-SEC3 complex, as well as to investigate the relationship between the affinity of SAGs for TCB and MHC and their ability to activate T cells, we measured the binding of a set of SEC3 mutants to a soluble recombinant TCR beta chain and to the human MHC class II molecule HLA-DR1. We show that there is direct correlation between affinity and ability to stimulate T cells, with SAGs having the highest affinity for the TCR being the most biologically active. We also find that there is an interplay between TCR-SAG and SAG-MHC interactions in determining mitogenic potency, such that a small increase in the affinity of a SAG for MHC can overcome a large decrease in the SAG's affinity for the TCR. Finally, we observe that those SEC3 residues that make the greatest energetic contribution to stabilizing the V beta C beta-SEC3 complex are strictly conserved among enterotoxins reactive with mouse V beta 8.2, thereby explaining why SAGs having other residues at these positions show different V beta-binding specificities.


Assuntos
Conformação Proteica , Receptores de Antígenos de Linfócitos T alfa-beta/química , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Superantígenos/imunologia , Sequência de Aminoácidos , Animais , Apresentação de Antígeno/imunologia , Antígenos de Bactérias/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Humanos , Cinética , Camundongos , Dados de Sequência Molecular , Mutação , Peptídeos/imunologia , Relação Estrutura-Atividade , Linfócitos T/imunologia
16.
J Exp Med ; 187(6): 823-33, 1998 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-9500785

RESUMO

The three-dimensional structure of the complex between a T cell receptor (TCR) beta chain (mouse Vbeta8.2Jbeta2.1Cbeta1) and the superantigen (SAG) staphylococcal enterotoxin C3 (SEC3) has been recently determined to 3.5 resolution. To evaluate the actual contribution of individual SAG residues to stabilizing the beta-SEC3 complex, as well as to investigate the relationship between the affinity of SAGs for TCR and MHC and their ability to activate T cells, we measured the binding of a set of SEC3 and staphylococcal enterotoxin B (SEB) mutants to soluble recombinant TCR beta chain and to the human MHC class II molecule HLA-DR1. Affinities were determined by sedimentation equilibrium and/or surface plasmon detection, while mitogenic potency was assessed using T cells from rearrangement-deficient TCR transgenic mice. We show that there is a clear and simple relationship between the affinity of SAGs for the TCR and their biological activity: the tighter the binding of a particular mutant of SEC3 or SEB to the TCR beta chain, the greater its ability to stimulate T cells. We also find that there is an interplay between TCR-SAG and SAG-MHC interactions in determining mitogenic potency, such that a small increase in the affinity of a SAG for MHC can overcome a large decrease in the SAG's affinity for the TCR. Finally, we observe that those SEC3 residues that make the greatest energetic contribution to stabilizing the beta-SEC3 complex ("hot spot" residues) are strictly conserved among enterotoxins reactive with mouse Vbeta8.2, thereby providing a basis for understanding why SAGs having other residues at these positions show different Vbeta-binding specificities.


Assuntos
Enterotoxinas/metabolismo , Antígeno HLA-DR1/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Superantígenos/metabolismo , Sequência de Aminoácidos , Animais , Sítios de Ligação , Ativação Linfocitária , Camundongos , Camundongos Transgênicos , Dados de Sequência Molecular , Mutação , Linfócitos T/imunologia
17.
Nature ; 391(6666): 502-6, 1998 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-9461220

RESUMO

Antigen recognition by T lymphocytes is mediated by cell-surface glycoproteins known as T-cell antigen receptors (TCRs). These are composed of alpha and beta, or gamma and delta, polypeptide chains with variable (V) and constant (C) regions. In contrast to alphabeta TCRs, which recognize antigen only as peptide fragments bound to molecules of the major histocompatibility complex (MHC), gammadelta TCRs appear to recognize proteins directly, without antigen processing, and to recognize MHC molecules independently of the bound peptide. Moreover, small phosphate-containing non-peptide compounds have also been identified as ligands for certain gammadelta T cells. These studies indicate that antigen recognition by gammadelta TCRs may be fundamentally different from that by alphabeta TCRs. The three-dimensional structures of several alphabeta TCRs and TCR fragments, and their complexes with peptide-MHC or superantigens, have been determined. Here we report the crystal structure of the Vdelta domain of a human gammadelta TCR at 1.9 A resolution. A comparison with antibody and alphabeta TCR V domains reveals that the framework structure of Vdelta more closely resembles that of VH than of Valpha, Vbeta or VL (where H and L refer to heavy and light chains), whereas the relative positions and conformations of its complementarity-determining regions (CDRs) share features of both Valpha and VH. These results provide the first direct evidence that gammadelta TCRs are structurally distinct from alphabeta TCRs and, together with the observation that the CDR3 length distribution of TCR delta chains is similar to that of immunoglobulin heavy chains, are consistent with functional studies suggesting that recognition of certain antigens by gammadelta TCRs may resemble antigen recognition by antibodies.


Assuntos
Receptores de Antígenos de Linfócitos T gama-delta/química , Sequência de Aminoácidos , Cristalografia por Raios X , Escherichia coli , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Conformação Proteica , Proteínas Recombinantes/química
18.
Immunity ; 9(6): 807-16, 1998 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-9881971

RESUMO

Superantigens (SAGs) are a class of immunostimulatory proteins of bacterial or viral origin that activate T cells by binding to the V beta domain of the T cell antigen receptor (TCR). The three-dimensional structure of the complex between a TCR beta chain (mouse V beta8.2) and the SAG staphylococcal enterotoxin B (SEB) at 2.4 A resolution reveals why SEB recognizes only certain V beta families, as well as why only certain SAGs bind mouse V beta8.2. Models of the TCR-SEB-peptide/MHC class II complex indicate that V alpha interacts with the MHC beta chain in the TCR-SAG-MHC complex. The extent of the interaction is variable and is largely determined by the geometry of V alpha/V beta domain association. This variability can account for the preferential expression of certain V alpha regions among T cells reactive with SEB.


Assuntos
Enterotoxinas/química , Fragmentos de Peptídeos/química , Conformação Proteica , Receptores de Antígenos de Linfócitos T alfa-beta/química , Staphylococcus aureus/imunologia , Superantígenos/química , Animais , Cristalografia por Raios X , Enterotoxinas/imunologia , Antígenos de Histocompatibilidade Classe II/química , Antígenos de Histocompatibilidade Classe II/imunologia , Camundongos , Modelos Moleculares , Fragmentos de Peptídeos/imunologia , Peptídeos/química , Peptídeos/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/fisiologia , Superantígenos/imunologia
19.
Artigo em Inglês | MEDLINE | ID: mdl-7735521

RESUMO

A 37-year-old woman with a previous history of allergic rhinitis and bronchial asthma presented local and generalized allergic reactions to bovine, porcine and human insulins. She developed ketoacidosis, high insulin requirements and transient hypoglycemic episodes. Several desensitizing schedules were applied in order to induce tolerance to human insulin therapy. In addition, the following parameters of the humoral immune response were measured in different serum samples taken within 13 days after one of the anaphylactic episodes: insulin antibodies (binding range = 29.4-47.8%; cutoff = 2.6%), total IgE (range = 500-850 IU/ml; normal values = 3.7-269 IU/ml), specific IgE (range = 0.42-0.83 PRU/ml; class 2) and subclasses of specific IgG (IgG1 = 97.2 SDS; IgG2 = 41.1 SDS; IgG3 = 9.9 SDS; IgG4 = 0.3 SDS, on day 1). A binding capacity of 31.8 IU insulin/l obtained by Scatchard analysis was in agreement with episodes of elevated insulin requirements and hypoglycemia. A high anti-insulin IgG/IgE ratio, along with high levels of specific IgG1 antibodies, suggested that the latter antibodies could be involved in the development of anaphylactic episodes.


Assuntos
Hipersensibilidade a Drogas/imunologia , Imunoglobulina E/sangue , Imunoglobulina G/sangue , Insulina/imunologia , Adulto , Feminino , Humanos , Imunoglobulina G/classificação
20.
J Immunol Methods ; 169(2): 241-9, 1994 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-8133083

RESUMO

On type 1 newly diagnosed and on insulin treated diabetic patients, anti-insulin autoantibodies (IAA) and antibodies (IA) having the same specificity are respectively induced. Such immune response may be evaluated either by radiobinding assay (RBA) or enzyme-linked immunosorbent assay (ELISA). Both methodologies have been compared at previous International Workshops, which pointed out discrepancies in results. In this work, IAA/IA prevalence was assessed by displacement RBA and ELISA, in normal subjects, type 2 (treated with hypoglycaemic agents), insulin treated and newly diagnosed type 1 diabetic patients. Results showed a lack of RBA-ELISA agreement. An attempt was then made to determine whether such results were, at least in part, attributable to iodination site in Tyr-A14. For this purpose parallel RBA assays were carried out by using radiolabelled insulin at A14 and A19 Tyr residues. Control sera and samples from insulin treated and type 1 newly diagnosed diabetic patients were tested. Our results suggest that labelling position is not involved in artifactual binding of tracers, at least as a systematic phenomenon. In the majority of cases the variability in RBA-ELISA signal ratios are best explained in terms of differences in the basic principles operating in both methods instead of artifacts due to tracer preparation.


Assuntos
Ensaio de Imunoadsorção Enzimática/métodos , Anticorpos Anti-Insulina/imunologia , Insulina/imunologia , Radioisótopos do Iodo , Ensaio Radioligante/métodos , Tirosina , Adolescente , Adulto , Idoso , Autoanticorpos/análise , Autoanticorpos/imunologia , Sítios de Ligação de Anticorpos/imunologia , Criança , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 2/imunologia , Feminino , Humanos , Anticorpos Anti-Insulina/análise , Masculino , Pessoa de Meia-Idade
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...