RESUMO
Self-assembled quantum dots (QDs) based on III-V semiconductors have excellent properties for applications in quantum optics. However, the presence of a 2D wetting layer (WL) which forms during the Stranski-Krastanov growth of QDs can limit their performance. Here, we investigate WL formation during QD growth by the droplet epitaxy technique. We use a combination of photoluminescence excitation spectroscopy, lifetime measurements, and transmission electron microscopy to identify the presence of an InGaAs WL in these droplet epitaxy QDs, even in the absence of distinguishable WL luminescence. We observe that increasing the amount of Ga deposited on a GaAs (100) surface prior to the growth of InGaAs QDs leads to a significant reduction in the emission wavelength of the WL to the point where it can no longer be distinguished from the GaAs acceptor peak emission in photoluminescence measurements. However increasing the amount of Ga deposited does not suppress the formation of a WL under the growth conditions used here.
Assuntos
Arsenicais , Gálio , Pontos Quânticos , Arsenicais/química , Luminescência , Gálio/químicaRESUMO
Human lymphocyte antigen (HLA) class I proteins of the major histocompatibility complex are largely dependent for expression on small peptides supplied to them by transporter associated with antigen processing (TAP) protein. An inherited human deficiency in the TAP transporter was identified in two siblings suffering from recurrent respiratory bacterial infections. The expression on the cell surface of class I proteins was very low, whereas that of CD1a was normal, and the cytotoxicity of natural killer cells was affected. In addition, CD8+ alpha beta T cells were present in low but significant numbers and were cytotoxic in the most severely affected sibling, who also showed an increase in CD4+CD8+ T cells and gamma delta T cells.
Assuntos
Transportadores de Cassetes de Ligação de ATP , Proteínas de Transporte/genética , Antígenos de Histocompatibilidade Classe I/análise , Síndromes de Imunodeficiência/genética , Linfócitos/imunologia , Membro 2 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Membro 3 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Adolescente , Sequência de Aminoácidos , Antígenos CD/análise , Antígenos CD1 , Sequência de Bases , Proteínas de Transporte/análise , Criança , Feminino , Antígenos de Histocompatibilidade Classe I/metabolismo , Homozigoto , Humanos , Síndromes de Imunodeficiência/imunologia , Células Matadoras Naturais/imunologia , Células de Langerhans/imunologia , Contagem de Leucócitos , Masculino , Dados de Sequência Molecular , Mutação , Subpopulações de Linfócitos T/imunologia , Linfócitos T Citotóxicos/imunologiaRESUMO
The temporal precision with which EPSPs initiate action potentials in postsynaptic cells determines how activity spreads in neuronal networks. We found that small EPSPs evoked from just subthreshold potentials initiated firing with short latencies in most CA1 hippocampal inhibitory cells, while action potential timing in pyramidal cells was more variable due to plateau potentials that amplified and prolonged EPSPs. Action potential timing apparently depends on the balance of subthreshold intrinsic currents. In interneurons, outward currents dominate responses to somatically injected EPSP waveforms, while inward currents are larger than outward currents close to threshold in pyramidal cells. Suppressing outward potassium currents increases the variability in latency of synaptically induced firing in interneurons. These differences in precision of EPSP-spike coupling in inhibitory and pyramidal cells will enhance inhibitory control of the spread of excitation in the hippocampus.
Assuntos
Potenciais de Ação/fisiologia , Potenciais Pós-Sinápticos Excitadores/fisiologia , Hipocampo/fisiologia , Interneurônios/fisiologia , Células Piramidais/metabolismo , Animais , Estimulação Elétrica , Hipocampo/citologia , Técnicas In Vitro , Interneurônios/citologia , Inibição Neural/fisiologia , Potássio/metabolismo , Células Piramidais/citologia , Ratos , Ratos Sprague-Dawley , Tempo de Reação/fisiologia , Sódio/metabolismoRESUMO
Rhythmic gamma oscillations at 30-70 Hz in cortical and hippocampal slices depend on a maintained excitation and on interactions between interneurons and pyramidal cells. These interactions include gap-junctional connections between inhibitory cells and fast excitatory and inhibitory chemical synapses. Spike timing with precision in the range of several ms may be assured by biphasic signaling mechanisms operating at these different connections. Such temporal precision may be important in cognitive processing.
Assuntos
Potenciais de Ação/fisiologia , Interneurônios/fisiologia , Percepção/fisiologia , Animais , HumanosRESUMO
The transporter associated with antigen processing (TAP), which is composed of two subunits (TAP1 and TAP2) that have different biochemical and functional properties, plays a key role in peptide loading and the cell surface expression of HLA class I molecules. Three cases of HLA class I deficiency have previously been shown to result from the absence of a functional TAP2 subunit. In the present study, we analyzed two cases displaying not only the typical lung syndrome of HLA class I deficiency but also skin lesions, and found these patients to be TAP1-deficient. This defect leads to unstable HLA class I molecules and their retention in the endoplasmic reticulum. However, the absence of TAP1 is compatible with life and does not seem to result in higher susceptibility to viral infections than TAP2 deficiency. This work also reveals that vasculitis is often observed in HLA class I-deficient patients.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Antígenos de Histocompatibilidade Classe I/genética , Mutação , Membro 2 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Antígenos de Histocompatibilidade Classe I/imunologia , HumanosRESUMO
Behavioural analysis of mice carrying engineered mutations is widely used to identify roles of specific genes in components of the mammalian behavioural repertoire. The reproducibility and robustness of phenotypic measures has become a concern that undermines the use of mouse genetic models for translational studies. Contributing factors include low individual study power, non-standardized behavioural testing, failure to address confounds and differences in genetic background of mutant mice. We have examined the importance of these factors using a statistically robust approach applied to behavioural data obtained from three mouse mutations on 129S5 and C57BL/6J backgrounds generated in a standardized battery of five behavioural assays. The largest confounding effect was sampling variation, which partially masked the genetic background effect. Our observations suggest that strong interaction of mutation with genetic background in mice in innate and learned behaviours is not necessarily to be expected. We found composite measures of innate and learned behaviour were similarly impacted by mutations across backgrounds. We determined that, for frequently used group sizes, a single retest of a significant result conforming to the commonly used P < 0.05 threshold results in a reproducibility of 60% between identical experiments. Reproducibility was reduced in the presence of strain differences. We also identified a P-value threshold that maximized reproducibility of mutant phenotypes across strains. This study illustrates the value of standardized approaches for quantitative assessment of behavioural phenotypes and highlights approaches that may improve the translational value of mouse behavioural studies.
Assuntos
Comportamento Animal/fisiologia , Mutação , Animais , Modelos Animais de Doenças , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Atividade Motora/genética , Fenótipo , Reprodutibilidade dos Testes , Comportamento Social , SoftwareRESUMO
Conventional major histocompatibility complex class I molecules are highly polymorphic and present peptides to cytotoxic T cells. These peptides derive from the proteolytic degradation of endogenous proteins in the cytosol and are translocated into the endoplasmic reticulum by a peptide transporter consisting of two transporter associated with antigen processing (TAP) molecules. Absence of this transporter leads to the synthesis of unstable peptide free class I molecules that are weakly expressed on the cell surface. Mouse nonconventional class I molecules (class Ib) may also present TAP-dependent peptides. In humans, CD1 antigens are nonconventional class I molecules. Recently, we characterized a human HLA class I deficiency resulting from a homozygous TAP deficiency. We show here that CD1a and -c are normally expressed on epidermal Langerhans cells of the TAP-deficient patients, as are CD1a, -b, and -c on dendritic cells differentiated in vitro from monocytes. Moreover, the CD1a antigens present on the surface of the dendritic cells are functional, since they internalize by receptor-mediated endocytosis gold-labeled F(ab')2 fragments of an anti-CD1a mAb. This suggests either that CD1 molecules are empty molecules, that they are more stable than empty conventional class I proteins, or that CD1 molecules present TAP-independent peptides.
Assuntos
Transportadores de Cassetes de Ligação de ATP/fisiologia , Antígenos CD/biossíntese , Membro 3 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/genética , Adolescente , Apresentação de Antígeno/imunologia , Antígenos CD1 , Células Cultivadas , Feminino , Citometria de Fluxo , Antígenos HLA/fisiologia , Antígenos de Histocompatibilidade Classe I/fisiologia , Humanos , Imuno-Histoquímica , Monócitos/citologia , Monócitos/ultraestruturaRESUMO
Short-duration, cyclically loaded, axial, fretting corrosion tests were designed and performed to compare the fretting behaviour of different metal Howmedica universal heads connected to coated and uncoated metal cone taper spigots. Concurrent axial push-on and pull-off taper friction tests were also performed. There was no measurable fretting debris found in any test and SEM studies of the contact surfaces showed no evidence of fretting. It is concluded that no significant, long-term fretting corrosion of the Vitalium Co-Cr-Mo heads on the Vitallium or uncoated Ti-6AI-4V spigots, nor of the Orthinox stainless steel heads on the Orthinox spigots, is likely to occur in vivo.
Assuntos
Prótese de Quadril , Teste de Materiais , Metais , Corrosão , Desenho de Prótese , Falha de Prótese , Aço Inoxidável , Propriedades de Superfície , VitálioRESUMO
Static axial push-on and lift-off, and push-on and twist-off experiments were designed and performed to measure the effective, room-temperature coefficient of friction mu for different design femoral prosthesis cone taper joints comprising a universal head on a stem spigot. Alumina and metal heads were tested on metal spigots using either distilled water, Ringer's solution, blood or no lubricant. Complementary push-on and lift-off friction tests of Araldite model joints for subsequent frozen-stress, photoelastic stress analysis were performed at 130 degrees C. It was found that lubricant caused little decrease in the values of the coefficient of friction in prosthesis tapers. The values measured were typically mu = 0.2 for an alumina head on a Co-Cr-Mo or Ti-6Al-4V spigot, mu = 0.15 for a Co-Cr-Mo head on a Co-Cr-Mo or Ti-6Al-4V spigot and mu = 0.13 for a stainless steel head on a stainless steel spigot. For Araldite photoelastic models of an alumina head on a Vitallium spigot, as-cast taper surfaces lubricated with silicone grease gave consistent friction of typically mu = 0.14. The axial displacement of model heads on their spigots were compared with predicted values and previously measured values for prosthesis heads.
Assuntos
Prótese de Quadril , Resinas Epóxi , Cabeça do Fêmur , Lubrificação , Teste de Materiais , Anidridos Ftálicos , Desenho de Prótese , Estresse MecânicoRESUMO
Three-dimensional, frozen-stress photoelasticity was used to study the best shape for a proposed alumina universal head loaded on to a Vitallium cone taper spigot with a 30 degrees inclined force, as in vivo. Typical cone taper friction values were reproduced in the photoelastic models. The location of the highest tensile stresses in the Mark I shape with a flat crown was found to be on the inner surface of the crown. Changing to a torispherical surface in the Mark II shape reduced this magnitude. However, the Mark III shape with a hemispherical inner crown surface gave even lower stress there, equal to the maximum value of the hoop stress at the taper, which was measured to be fairly uniform both around and along the taper except at the ends where contact pressure concentrations were found to occur and it became reduced. Lamé axisymmetric cylinder stress predictions were found to be useful approximations to measured values and were generally overestimates of the tensile hoop stress at the head taper surface.
Assuntos
Óxido de Alumínio , Alumínio , Prótese de Quadril , Elasticidade , Cabeça do Fêmur , Desenho de Prótese , Estresse MecânicoRESUMO
A 3-year-old boy with Proteus syndrome has a novel germline p.Y68D mutation of the PTEN gene inherited from his mother who has Cowden syndrome. In addition, DNA extracted from curettings of his widespread epidermal naevus shows loss of heterozygosity for this mutation. To our knowledge, this has not been described before.
Assuntos
Perda de Heterozigosidade , Nevo Pigmentado/genética , PTEN Fosfo-Hidrolase/genética , Síndrome de Proteu/genética , Neoplasias Cutâneas/genética , Pré-Escolar , Mutação em Linhagem Germinativa , Humanos , Masculino , Nevo Pigmentado/patologia , Linhagem , Neoplasias Cutâneas/patologiaRESUMO
1. The patterns of discharge of spontaneous GABAA-mediated inhibitory postsynaptic currents (sIPSCs), originating from the nucleus reticularis thalami (NRT), and their modulation by GABAB autoreceptors, were studied in rat thalamocortical (TC) neurones using whole-cell voltage-clamp recordings in brain slices. 2. sIPSCs were recorded in all ventro-basal (VB) and dorsal lateral geniculate (LGN) neurones. In VB neurones, in the presence of tetraethylammonium (TEA, 5 mM), these sIPSCs can occur in bursts at frequencies of either 0.1 or 1-2 Hz. In the presence of tetrodotoxin (TTX), these bursting activities are replaced by the continuous discharge of miniature IPSCs (mIPSCs), recorded in the absence of TEA, at a frequency of 4 Hz. The kinetic properties of mIPSCs were similar in VB and LGN TC neurones. 3. In VB TC neurones the GABAB receptor agonist (+/-)-baclofen, at a concentration of 0.05 microM, decreased the mIPSC frequency by 22% without affecting their amplitude distribution. Increasing the (+/-)-baclofen concentration to 1 and 10 microM caused similar reductions (41 and 47%, respectively) in the mIPSCs frequency: these values were significantly different from the one observed with 0.05 microM (+/-)-baclofen. In LGN TC neurones, where mIPSCs originate from both NRT and local interneurone terminals, 1 microM (+/-)-baclofen produced a 66% reduction in the mIPSC frequency. 4. The GABAB receptor antagonist CGP55845A (50 nM) not only blocked the baclofen-mediated decrease in mIPSC frequency, but also produced a 52% increase in the mIPSC frequency compared with control in three out of seven neurones. Application of CGP55845A (50-500 nM) alone produced a 77% increase in the mIPSC frequency in three out of nine VB neurones, and in the LGN, CGP55845A (100 nM) produced a 53% increase in four out of nine neurones. CGP55845A (100 nM) also reversibly increased the amplitude of evoked GABAA IPSCs by 74 and 57% in three out of three VB and three out of five LGN neurones, respectively. 5. Application of GABA (1.5-5 microM) decreased the mIPSC frequency in VB TC neurones by a similar extent (48%) as 1-10 microM (+/-)-baclofen. 6. In the presence of 100 microM Cd2+, (+/-)-baclofen still decreased the mIPSC frequency by about 40%, indicating that the effect of presynaptic GABAB receptor activation on spontaneous GABA release did not occur through a reduction of voltage-dependent Ca2+ currents. 7. Cd2+ (100 microM) decreased the amplitude of both mIPSCs and isoguvacine-induced current by 30 and 19%, respectively, indicating an effect of this divalent cation on postsynaptic GABAA receptors. 8. We conclude that GABAB autoreceptors are present on the GABAergic terminals within the thalamic sensory nuclei and that these receptors can be tonically activated by the ambient GABA.
Assuntos
Autorreceptores/fisiologia , Receptores de GABA-A/fisiologia , Receptores de GABA-B/fisiologia , Núcleos Talâmicos/química , Animais , Baclofeno/farmacologia , Cádmio/farmacologia , Agonistas GABAérgicos/farmacologia , Antagonistas GABAérgicos/farmacologia , Corpos Geniculados/química , Corpos Geniculados/citologia , Corpos Geniculados/fisiologia , Interneurônios/química , Interneurônios/fisiologia , Ácidos Isonicotínicos/farmacologia , Masculino , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Técnicas de Cultura de Órgãos , Técnicas de Patch-Clamp , Ácidos Fosfínicos/farmacologia , Terminações Pré-Sinápticas/química , Terminações Pré-Sinápticas/efeitos dos fármacos , Terminações Pré-Sinápticas/fisiologia , Propanolaminas/farmacologia , Ratos , Ratos Wistar , Tetrodotoxina/farmacologia , Núcleos Talâmicos/citologia , Núcleos Talâmicos/fisiologia , Ácido gama-Aminobutírico/farmacologiaRESUMO
Neurotransmitters affect the membrane potential (Vm) of target cells by modulating the activity of receptor-linked ion channels. The direction and amplitude of the resulting transmembrane current depend on the resting level of Vm and the gradient across the membrane of permeant ion species. Vm, in addition, governs the activation state of voltage-gated channels. Knowledge of the exact level of Vm is therefore crucial to evaluate the nature of the neurotransmitter effect. However, the traditional methods to measure Vm, with microelectrodes or the whole-cell current-clamp technique, have the drawback that the recording pipette is in contact with the cytoplasm, and dialysis with the pipette solution alters the ionic composition of the interior of the cell. Here we describe a novel technique to determine the Vm of an intact cell from the reversal potential of K+ currents through a cell-attached patch. Applying the method to interneurons in hippocampal brain slices yielded more negative values for Vm than subsequent whole-cell current-clamp measurements from the same cell, presumably reflecting the development of a Donnan potential between cytoplasm and pipette solution in the whole-cell mode. Cell-attached Vm measurements were used to study GABAergic actions in intact CA1 interneurons. In 1- to 3-week-old rats, bath-applied GABA inhibited these cells by stabilizing Vm at a level depending on contributions from both GABAA and GABAB components. In contrast, in 1- to 4-d-old animals, only GABAA receptors were activated resulting in a depolarizing GABA response.
Assuntos
Hipocampo/fisiologia , Interneurônios/fisiologia , Ácido gama-Aminobutírico/fisiologia , Animais , Hipocampo/citologia , Técnicas In Vitro , Potenciais da Membrana/fisiologia , Técnicas de Patch-Clamp , RatosRESUMO
1. The cell-attached configuration of the patch-clamp technique was used to assess resting membrane potential and firing threshold of CA1 pyramidal cells and interneurones of rat hippocampal slices. 2. Resting potential was inferred from the reversal potential of voltage-gated K+ currents with symmetrical intracellular and pipette K+ concentrations. Its mean value was -74 +/- 9 mV for silent interneurones (mean +/- s.d.; n = 17) and -84 +/- 7 mV for silent pyramidal cells (n = 8). Spontaneous action currents occurred in thirteen out of thirty-two interneurones and two out of ten pyramidal cells. In active cells, membrane potential values fluctuated by up to 20 mV, due in part to the large hyperpolarizations that followed an action current. 3. Membrane potential values determined from K+ current reversal were 13 +/- 6 mV more hyperpolarized than those measured in whole-cell recordings from the same neurones (n = 8), probably due to a Donnan equilibrium potential between pipette and cytoplasm. 4. Firing threshold of silent cells was determined by elevating external K+ until action currents were generated, while membrane potential was monitored from the cell-attached K+ current reversal. Spike threshold was attained at -49 +/- 8 mV for interneurones (n = 17) and at -60 +/- 8 mV for pyramidal cells (n = 8). Increasing external Ca2+ from 2 to 4 mM shifted the neuronal voltage threshold by +5 mV, without affecting resting potential. 5. For comparison with these values, we examined how the rate of membrane polarization influenced firing threshold in whole-cell records. Ramp current injections, of duration 15-1500 ms, revealed that current threshold followed a classical strength-duration relationship. In contrast voltage threshold, determined from current injection or by elevating extracellular K+, varied little with the rate of membrane polarization. 6. The state of activation and inactivation of Na+ and K+ currents might contribute to the stability of the voltage threshold. Cell-attached records showed that 79 +/- 10 % of Na+ channels and 64 +/- 10 % of K+ channels were available for activation at resting potential in silent cells (n = 8). As cells were depolarized to threshold, Na+ current availability was reduced to 23 +/- 10 %, and K+ current availability to 31 +/- 12 %. 7. The speed of transition into the inactivated states also appears to contribute to the invariance of threshold for all but the fastest depolarizations. At potentials close to threshold, the rate of inactivation of Na+ and K+ followed a double exponential time course, such that Na+ currents were 62 % inactivated and K+ currents were 63 % inactivated within 15 ms.
Assuntos
Adesão Celular/fisiologia , Hipocampo/fisiologia , Interneurônios/fisiologia , Canais de Potássio/fisiologia , Células Piramidais/fisiologia , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/fisiologia , Animais , Cálcio/farmacologia , Cálcio/fisiologia , Técnicas In Vitro , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Técnicas de Patch-Clamp , Ratos , Ratos Sprague-Dawley , Canais de Sódio/fisiologiaRESUMO
Dendritic cells express several alternatively spliced CD1e mRNAs. These molecules encode proteins characterized by the presence of either one, two, or three alpha domains and either a 51- or 63-amino acid cytoplasmic domain. Moreover, mRNAs encoding isoforms lacking the transmembrane domain are observed. Several of these CD1e isoforms were expressed in transfected cells, and two of them, with three alpha domains, displayed a particular processing pathway. These latter isoforms slowly leave the endoplasmic reticulum due to the presence of atypical dilysine motifs in the cytoplasmic tail. These molecules are associated with the beta(2)-microglobulin and accumulate in late Golgi and late endosomal compartments. In the latter compartments, they are cleaved into soluble forms that appear to be stable. In dendritic cells, these isoforms are mainly located in the Golgi apparatus, and upon maturation they are redistributed to late endosomal compartments. This work demonstrates the existence of CD1e molecules. As compared with other CD1 molecules, CD1e displays fundamentally different properties and therefore may represent a third type of CD1 molecules.
Assuntos
Antígenos CD1/imunologia , Células Dendríticas/imunologia , Processamento Alternativo , Antígenos CD1/química , Antígenos CD1/genética , Sequência de Bases , Primers do DNA , Endossomos/imunologia , Complexo de Golgi/imunologia , Dados de Sequência Molecular , Precursores de RNA/genética , RNA Mensageiro/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologiaRESUMO
Changes in gene expression occurring during differentiation of human monocytes into dendritic cells were studied at the RNA and protein levels. These studies showed the induction of several gene classes corresponding to various biological functions. These functions encompass antigen processing and presentation, cytoskeleton, cell signalling and signal transduction, but also an increase in mitochondrial function and in the protein synthesis machinery, including some, but not all, chaperones. These changes put in perspective the events occurring during this differentiation process. On a more technical point, it appears that the studies carried out at the RNA and protein levels are highly complementary.
Assuntos
Diferenciação Celular/genética , Células Dendríticas/fisiologia , Monócitos/fisiologia , Proteínas/metabolismo , RNA/metabolismo , Primers do DNA/química , Perfilação da Expressão Gênica , Humanos , Proteoma/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Human HLA class I deficiency is a rare disease which, in most of the patients described to date, results from a defect in subunit 1 or 2 of the peptide transporter associated with antigen processing (TAP). The clinical features of TAP deficiency include a chronic inflammation of the respiratory tract and/or granulomatous skin lesions. In this report, we describe two adult siblings with an HLA class I deficiency. One individual had only spontaneously-healing skin granulomatous lesions, while the second did not display any of the symptoms associated with HLA class I deficiency and could be considered to be healthy. We show that the patients display a homozygous TAP2 mutation which blocks the maturation of HLA class I molecules. Cell surface expression of these molecules is strongly reduced, but three times higher than on cells from other previously described TAP-deficient individuals. This higher expression results, at least in part, from the presence of HLA-B7 molecules which are probably empty of peptide. The numbers of CD8+ alphabeta T cells are almost normal in these patients. The anti-EBV T-cell response of one patient is mediated by HLA-B7 restricted CD8+ alphabeta T lymphocytes recognizing the BMRF1 nuclear EBV antigen, demonstrating that CD8+ alphabeta T cells can participate in anti-viral responses. This study shows that TAP deficiency can remain totally asymptomatic for several decades, and suggests that in some cases, TAP-independent immune responses provide efficient protection from most of the common intracellular pathogens.