A window of opportunity for cooperativity in the T Cell Receptor.

The T-cell antigen receptor (TCR) is pre-organised in oligomers, known as nanoclusters. Nanoclusters could provide a framework for inter-TCR cooperativity upon peptide antigen-major histocompatibility complex (pMHC) binding. Here we have used soluble pMHC oligomers in search for cooperativity effects along the plasma membrane plane. We find that initial binding events favour subsequent pMHC binding to additional TCRs, during a narrow temporal window. This behaviour can be explained by a 3-state model of TCR transition from Resting to Active, to a final Inhibited state. By disrupting nanoclusters and hampering the Active conformation, we show that TCR cooperativity is consistent with TCR nanoclusters adopting the Active state in a coordinated manner. Preferential binding of pMHC to the Active TCR at the immunological synapse suggests that there is a transient time frame for signal amplification in the TCR, allowing the T cells to keep track of antigen quantity and binding time.

Sangrado variceal duodenal secundario a trombosis venosa mesentérica: a propósito de un caso / Duodenal variceal bleeding secondary to mesenteric vein thrombosis

La trombosis venosa mesentérica corresponde a un factor obstructivo del sistema venoso intestinal intraluminal, trayendo consecuencias clínicas dadas por isquemia intestinal y aumento de la circulación colateral debido a una dificultad del drenaje sanguíneo. Raramente, esto podría generar várices ectópicas, pero esta descrito en la literatura la existencia de várices duodenales producto de este mecanismo. En este articulo se presenta un reporte de un caso, que presenta un sangrado variceal de origen duodenal secundaria a una trombosis venosa portal y mesentérica, sin causa aparente.

Mesenteric vein thrombosis is an intraluminal obstruction of the intestinal venous system with clinical consequences due to intestinal ischemia and an increase in collateral circulation caused by compromised venous return. It very rarely generates ectopic varicose veins but duodenal varicose veins caused by this mechanism have been described in the literature. In this article we report the case of duodenal variceal bleeding secondary to a portal and mesenteric venous thrombosis with no apparent cause.

Enfermedad de Still del Adulto, a propósito de un caso / Still's disease, a clinical case

La enfermedad de Still del Adulto es una variante sistémica del universo llamado "artritis idiopática juvenil", cuya diferencia es la edad de aparición, recibiendo este nombre cuando el cuadro se manifiesta en personas mayores de 15 años. Es una enfermedad autoinflamatoria caracterizada por fiebre intermitente, artritis, rash evanescente y linfadenopatías, asociado a manifestaciones de compromiso sistémico. Es una patología de baja prevalencia, pero es importante considerarla como uno de los principales diagnósticos etiológicos de la fiebre de origen desconocido.

Adult-onset Still's disease is a systemic variant of Still's disease distinguished from juvenile idiopathic arthritis by its onset age of over 15. It is an auto-inflammatory disease characterized by intermittent fever, arthritis, short-lived rash and lymphadenopathies. It has low prevalence but is important to consider as one of the principle etiological diagnoses of fever of unknown origin.

Contribución de la temperatura fría y el sabor ácido en la intervención fonoaudiológica de la disfagia orofaríngea / Contribution of cold temperature and sour taste in speech and language intervention of oropharyngeal dysphagia

El fonoaudiólogo es el principal profesional en la rehabilitación no farmacológica y no quirúrgica del usuario con disfagia. Su participación es fundamental tanto para el aminoramiento del riesgo de aspiración o penetración laríngea, como para mejorar o restaurar la función deglutoria. Para este fin, posee opciones terapéuticas directas e indirectas, cuya elección y aplicación dependerá de la patología que curse el usuario, las redes que posea para su recuperación y la motivación intrínseca del mismo. Entre las estrategias de intervención indirecta se encuentra el Tratamiento Sensorio-Motor Oral (OSMT, por sus siglas en inglés), el cual pretende producir una aceleración en el desencadenamiento del proceso deglutorio mediante la ejercitación de los músculos orofaciales en conjunto con diferentes estímulos sensoriales (específicamente la temperatura fría y el sabor ácido). La presente revisión tiene por objetivo dilucidar si la utilización de la temperatura fría y el sabor ácido son útiles como mecanismo de intervención indirecta de la disfagia. Se concluye que las acciones propuestas son efectivas simplemente como mecanismos compensatorios en el proceso deglutorio, puesto que modifican las características del bolo alimenticio e incrementan momentáneamente las sensaciones intraorales.

The speech-language pathologist (SLP) is the main professional in the nonpharmacological and non-surgical rehabilitation of patients with dysphagia. Their role is essential for both reducing the risk of aspiration or laryngeal penetration and improving or restoring the swallowing function. To this end, the SLP has direct and indirect therapeutic options, whose choice and application will depend on the patient’s condition, support networks, and their intrinsic motivation. As part of the indirect intervention strategies, the oral sensorymotor treatment (OSMT) aims to exercise the orofacial muscles, and introduce sensory input by the application of cold temperature and sour taste to increase the triggering speed of the swallowing reflex. This review seeks to determine whether the use of cold temperature and sour taste are effective indirect mechanisms for treating patients with dysphagia. It is concluded that the proposed actions in this review are useful simply as compensatory mechanisms in the swallowing process, as they modify the bolus properties and increase, temporarily, the intra-oral sensations.

PCR-based procedures for detection and quantification of Staphylococcus aureus and their application in food.

AIMS: To evaluate the specificity of nuc targeted primers for PCR detection of Staphylococcus aureus in different food matrices and to establish a RTQ-PCR procedure suitable for the routine detection and quantification of this pathogen in food. METHODS AND RESULTS: Specificity of nuc targeted primers (Pri1-Pri2 and the newly designed RTQ-PCR primers) was tested on a total of 157 strains of genetically confirmed identity, including reference and food isolates. PCR detection on artificially inoculated beef samples by DNA extraction using a DNeasy Tissue Kit (Qiagen GmhH, Hilden, Germany) showed a sensitivity value around 10(3) CFU g(-1). The two RTQ-PCR systems, incorporating SYBR-Green I and TaqMan, respectively, applied in the present work improved the sensitivity of conventional PCR by lowering the detection level to 10 and 100 cells, respectively. Out of 164 naturally contaminated foods tested for the presence of Staph. aureus, 74 were positive by conventional PCR and 69 by the traditional culture method with a high degree of result agreement between both methodologies (93.3%). CONCLUSIONS: PCR approaches, using nuc targeted primers, have proved specific and combined with growth techniques may improve detection of Staph. aureus in different types of food. SIGNIFICANCE AND IMPACT OF THE STUDY: The SYBR-Green I real-time PCR approach established allows the sensitive, automated and quantitative detection of Staph. aureus for routine analysis at a reasonable cost.

[Results of infrainguinal revascularization in patients with end-stage renal disease]. / Résultats des revascularisations sous-inguinales chez les malades en insuffisance rénale terminale.

OBJECTIVES: The aim of this retrospective study was to evaluate the efficacy of bypass in patients with endstage renal disease (ESRD) and to determine predictive factors and precise bypass indications. METHOD: Forty one patients with ESRD underwent 50 bypass, 6 limbs were stage II and 44 stage III or IV according to Leriche and Fontaine classification. Revascularisations procedures were 47 infrainguinal bypass and 3 miscellaneous. Median follow up was 17,0+/-15,7 months. RESULTS: Perioperative mortality rate was 12% (n = 6). Morbidity was as follow : 1 perioperative major imputation and 8 secondary ones. There were 26 secondary death (12 from cardiac events), cumulative survival rate declined to 42,9+/-7,7% and limb salvage rate to 77,2+/-7,5% at 2 years. Primary and secondary potency rates were 53,5+/-10,4% and 70,6+/-10%. Quality of life was good in 28% of revascularised patients. Among risk factors, myocardial events showed a statistical significance in predicting survival, good runoff and bypass occlusion showed a statistical significance in predicting limb salvage. CONCLUSION: Revascularisation can be performed in ESRD patients. However to improve the results full evaluation of myocardial risks, skin lesions and infection of the feet, available autologous vein and nutritional status may be needed in those patients.

PCR detection of Listeria monocytogenes: a study of multiple factors affecting sensitivity.

AIMS: To test, under comparable conditions, several parameters affecting sensitivity of PCR detection in order to establish a PCR procedure suitable for the routine detection of Listeria monocytogenes in food. METHODS AND RESULTS: Beef samples artificially inoculated were used to determine sensitivity of PCR detection under different parameters. As few as 1 CFU g(-1) were detected by DNA extraction using a DNeasy Tissue Kit (Qiagen GmhH, Hilden, Germany) of 1 ml aliquot and PCR amplification with primers directed to the hlyA gene. This PCR protocol was applied in 60 naturally contaminated foods, comparing two enrichment procedures with the traditional culture method. The highest number of positives was recorded by PCR following a 24-h pre-enrichment step at 30 degrees C and a 24-h enrichment step at 37 degrees C. Afterwards, it was applied in 217 naturally contaminated foods and 56 of them tested positive for L. monocytogenes in which only 17 tested positive using the culture method. CONCLUSIONS: The PCR procedure described has proved to be a rapid and sensitive method suitable for the routine analysis of different types of food. SIGNIFICANCE AND IMPACT OF THE STUDY: The method proposed for the detection of L. monocytogenes, has been validated in naturally contaminated food and is suitable to implement in the food industry.

[Percutaneous transluminal angioplasty and severe limb ischaemia]. / Angioplastie transluminale percutanée et ischémie sévère des membres inférieurs.

OBJECTIVES: To determine the outcome and the place of transluminal angioplasty (ATL) in the treatment of severe limb ischaemia. MATERIALS AND METHODS: Seventy two legs at stage III and IV of Fontaine's classification have been exclusively treated by endoluminal procedures. The success was valued both on the haemodynamic post-operative improvement of the run-off flow and on the clinical statut leading to the conservation at mid-term of a functional limb. Patency and survival rate had been valued by actuarial method. RESULTS: Seventy percent of the limbs were haemodynamically improved. For the global population, a 48% clinical success rate was obtained at 6 months but 30.5% of limbs were loss. Primary patency rate was respectively 79, 71 and 68% at 6, 12 and 24 month. The quality of the run-off arteries has been the most influential factor. CONCLUSION: Endoluminal treatment of chronic limb ischaemia had lead to a clinical improvement in 48% of cases. Multi-stages and distal atherosclerotic disease of this patients limits ATL indications which results depend of run-off quality.

Evaluación psicométrica del inventario multifásico de personalidad de Minnesota para adolescentes (MMPI-A) en muestras chilenas / Psychometric evaluation for the Minnesota multiphasic personality inventory for adolescent (MMPI-A) in chilean samples

Este artículo presenta una caracterización psicométrica del Inventario Multifásico de Personalidad de Minnesota para Adolescentes (MMPI-A), instrumento de evaluación clínica que fue traducido y adaptado a los usos del lenguaje y la realidad sociocultural de los adolescentes chilenos a través de un enfoque de comité. El MMPI-A fue aplicado a una muestra de 705 adolescentes, considerando su género y tres ámbitos de procedencia (escolar, clínico y judicial). Se evaluaron las propiedades psicométricas de confiabilidad (estabilidad y consistencia interna) y validez (capacidad de discriminación entre grupos) de sus 38 escalas. Los resultados muestran una prueba estable y consistente, con niveles de confiabilidad semejantes a los obtenidos en estudios foráneos y con adecuada capacidad de discriminación de acuerdo a las variables estudiadas.

On the specificity of PCR detection of Listeria monocytogenes in food: a comparison of published primers.

A total of nine pairs of primers, seven previously published and two newly developed, have been assayed for PCR detection of Listeria monocytogenes in food. They have been tested for specificity on a total of 72 strains including reference and food isolates belonging to L. monocytogenes and other species in the genus. First of all, a polyphasic approach has been carried out in order to establish a reference strain collection. They were biochemically and genetically characterized by API-Lis and randomly amplified polymorphic DNA PCR (RAPD-PCR), respectively. Random amplification of DNA was performed with M13, T7 and T3 universal primers and a data bank was created to compile the RAPD patterns of all the analyzed strains. The UPGMA cluster analysis of RAPD profiles with primer M13 showed eight clusters at 72.3% similarity. Clusters 2 and 7 corresponded to L. monocytogenes. Clusters 1 and 6 grouped L. ivanovii strains. Clusters 3, 4, 5 and 8 corresponded to L. grayi, L. innocua, L. welshimeri and L. seeligeri, respectively. Pattern analysis revealed the existence of miss-identified reference strains which was confirmed by 16S rDNA sequence analysis. RAPD-PCR is a rapid genetic test which helped to confirm strain identity. On the basis of PCR specificity results, primers LM1-LM2 were the best combination for the detection of L. monocytogenes since they only amplified the specific fragment in strains that had been genetically and biochemically assessed as belonging to the species. Specificity of other assayed primers is discussed.

Modulación negativa del TCR por un mecanismo de todo-o-nada / All-or-none downregulation of the T cell antigen receptor

El contacto del TCR con MHC/antígeno resulta en su modulación y desaparición de la superficie celular. Recientemente hemos descrito que este proceso ocurre por al menos dos mecanismos: uno es dependiente de transmisión de señales, predomina a bajas concentraciones de antígeno y resulta en la modulación en trans de moléculas de TCR no contactadas. El otro requiere contacto directo y es independiente de transmisión de señales. En este artículo describimos que el TCR es modulado en una forma discontinua, es decir las células estimuladas oscilan de un estado no-modulado a otro completamente modulado sin transición aparente por estados intermedios, cuando las células T son estimuladas con altas dosis de antígeno o de anticuerpos anti-TCR. El fenómeno se reproduce cuando un receptor quimérico, que contiene la parte extracelular y transmembránica de CD8 y el tallo citoplásmico de CD3 , es entre cruzado con anticuerpos inmovilizados a un substrato. Este proceso de modulación de "todo-o-nada" no requiere de transmisión de señales o de polimerización del citoesqueleto de actina. El análisis por microscopía confocal muestra que el anticuerpo estimulante es tomado del substrato y concentrado junto con la quimera en un polo de la célula donde se constituye un sitio de nucleación para la formación de vesículas endocíticas. El efecto de "todo-o-nada" puede explicarse por la concentración lenta del TCR o de la quimera, seguido de la internalización rápida de los receptores agregados (AU)

Intracellular redistribution of nucleolin upon interaction with the CD3epsilon chain of the T cell receptor complex.

T cell activation through the antigen receptor (TCR) involves the cytoplasmic tails of the CD3 subunits CD3gamma, CD3delta, CD3epsilon, and CD3zeta. Whereas the biological significance of the cytoplasmic tails of these molecules is suggested, in part, by their evolutionarily conserved sequences, their interactions with signal transduction molecules are not completely understood. We used affinity chromatography columns of glutathione S-transferase fused to the CD3epsilon cytoplasmic tail to isolate proteins that specifically interact with this subunit. In this way, we identified the shuttling protein nucleolin as a specific CD3epsilon-interacting molecule. Using competition studies and affinity chromatography on peptide columns, we were able to identify a central proline-rich sequence as the nucleolin-interacting sequence in CD3epsilon. Transfection in COS cells of wild type CD3epsilon, but not of nonbinding mutants of CD3epsilon, resulted in redistribution of nucleolin from the nucleus and nucleoli to the cytoplasm. This property was transferred to a CD8 protein chimera by appending the cytoplasmic tail of CD3epsilon. We also found that nucleolin associated with the TCR complex. This association was increased upon TCR engagement, suggesting that the CD3epsilon/nucleolin interaction may have a role in T cell activation.

[Decompensation of lower limb arteritis after bone and joint surgery]. / Décompensation d'une artérite après chirurgie osseuse et articulaire.

PURPOSE OF THE STUDY: Decompensation of lower limb arteritis after bone and joint surgery is an unusual finding compared with the large number of procedures performed in both emergency and controlled settings. There is however a functional and limb-threatening risk that must not be overlooked. MATERIAL AND METHODS: We report a series of 9 patients followed in our department over the last 3 years. Emergency surgery had been required in 6 patients after trauma and 3 had undergone a planned orthopedic procedure. All the patients had at least one vascular risk factor, and 7 of them had a cardiovascular history. The inaugural sign was a trophic disorder due to a grade IV decompensated arteritis in 8 patients, including 2 with nonunion. Delay to treatment ranged from 1 to 3 months. Acute embolic ischemia required emergency care in 1 patient. RESULTS: A revascularization procedure was performed on 6 limbs and was successful in 3. There were also 6 amputations, three initially, 1 after septic shock and 2 because revascularization was impossible. Three of the amputations were required after failed revascularization. Prosthesis wearing and walking was possible in only two amputated patients. Overall rate of successful salvage was 33% (3 successful revascularizations among 9 limbs). One of the nonunions healed after revascularization; the limb was amputated for the other one. One patient died from septicemia. DISCUSSION: Our series further illustrates the severity of decompensated arteritis after bone and joint surgery, emphasizing the importance of searching for cardiovascular risk factors and functional signs suggestive of a vascular disorder. Arterial duplex Doppler and if necessary arteriography of the lower limbs should be obtained in case of doubt. Two different situations can be distinguished depending on the predictable vascular risk and the localization of the planned bone reconstruction. If the patient has an asymptomatic proximal arteritis and bone and joint surgery is planned above the knee, a revascularization procedure would not appear necessary prior to bone surgery. In other cases, it may be more advisable to treat the arteritis before attempting bone surgery. For trauma victims, the osteosynthesis technique depends greatly on knowledge of the vascular risk.

Internalization and intracellular fate of TCR-CD3 complexes.

The number of surface TCR-CD3 complexes is maintained by an equilibrium between the synthesis and secretion of new polypeptides, their internalization, recycling, and degradation. The different subunits of the TCR-CD3 complex do not display the same intracellular trafficking dynamics. Thus, in the absence of stimuli, TCR and zeta chains may be degraded at a higher rate than CD3 subunits, which are mostly recycled. T-cell activation by antigen, anti-TCR-CD3 antibodies, or pharmacological activators of protein kinase C, results in increased TCR-CD3 internalization, followed by the downmodulation of TCR-CD3 surface levels. Once internalized, TCR-CD3 complexes may either enter a recycling pathway or be sorted to lysosomes and degraded. Protein serine kinases and protein tyrosine kinases may influence the internalization and intracellular sorting of TCR-CD3 complexes. In line with these results TCR-CD3 ligands stimulate both TCR-CD3 internalization and degradation, whereas protein kinase C activators stimulate internalization only. Depending on the stimulus applied, internalization motifs from one or several TCR-CD3 subunits mediate endocytic routing of the complex. The involvement of signaling molecules in the intracellular fate of TCR-CD3, the nature and location of sequences for internalization and intracellular sorting, and the role of downregulation in T-cell activation are still the main open questions.

Rho regulates T cell receptor ITAM-induced lymphocyte spreading in an integrin-independent manner.

T cell receptor (TCR) engagement increases integrin-mediated adhesion to APC, resulting in the stabilization of the T cell : APC interaction and the close apposition of the two cell membranes. Here we show that engagement of either the TCR or CD3 chimeras with immobilized antibodies causes the rapid spreading of T cells in an integrin-independent fashion. This effect concurs with the polymerization of the actin cytoskeleton and is dependent on the integrity of the immunoreceptor tyrosine-based activation motifs of the CD3 subunits. Expression of a dominant negative mutant of RhoA, as well as the Rho-specific inhibitor C3 toxin, abolished TCR-induced spreading. In contrast, constitutively active or dominant negative forms of Rac and Cdc42 did not affect cell spreading. We conclude that signals emanating from the TCR can directly induce T cell spreading, independently of integrins, and via a Rho-dependent reorganization of the actin cytoskeleton.

CD3delta couples T-cell receptor signalling to ERK activation and thymocyte positive selection.

Thymocytes from mice lacking the CD3delta chain of the T-cell receptor (TCR), unlike those of other CD3-deficient mice, progress from a CD4- CD8- double-negative to a CD4+ CD8+ double-positive stage. However, CD3delta-/- double-positive cells fail to undergo positive selection, by which double-positive cells differentiate into more mature thymocytes. Positive selection is also impaired in mice expressing inactive components of the Ras/mitogen activated protein (MAP) kinase signalling pathway. Here we show that CD3delta-/- thymocytes are defective in the induction of extracellular signal-regulated protein kinase (ERK) MAP kinases upon TCR engagement, whereas activation of other MAP kinases is unaffected. The requirement for CD3delta maps to its extracellular or transmembrane domains, or both, as expression of a tail-less CD3delta rescues both ERK activation and positive selection in CD3delta-/- mice. Furthermore, the defect correlates with severely impaired tyrosine phosphorylation of the linker protein LAT, and of the CD3zeta chain that is localized to membrane lipid rafts upon TCR engagement. Our data indicate that the blockade of positive selection of CD3delta-/- thymocytes may derive from defective tyrosine phosphorylation of CD3zeta in lipid rafts, resulting in impaired activation of the LAT/Ras/ERK pathway.

Triggering the TCR complex causes the downregulation of nonengaged receptors by a signal transduction-dependent mechanism.

Downregulation of the TCR complex is believed to be intimately tied to T cell activation, allowing serial triggering of receptors and desensitization of stimulated cells. We studied transfected and transgenic T cells expressing CD3zeta chimeras to demonstrate that ligand engagement of the TCR or chimeras causes comodulation of nonengaged receptors. Comodulation required protein tyrosine kinase activity but not trans-phosphorylation of nonengaged receptors. The TCR appears to be downregulated by at least two mechanisms. One mechanism requires direct engagement, independent of signaling. The second requires signaling and downregulates nontriggered receptors. These results shed new light on the process of TCR downregulation and indicate that the number of downregulated TCRs cannot be assumed to equal the number of engaged receptors.

Receptor engagement transiently diverts the T cell receptor heterodimer from a constitutive degradation pathway.

In the absence of ligand, the T cell receptor (TCR)/CD3 complex is continuously internalized and recycled to the cell surface, whereas receptor engagement results in its down-regulation. The present study shows that the TCR and CD3 components follow different fates accompanying their constitutive internalization. Although the CD3 moiety is recycled to the cell surface, the TCR heterodimer is degraded and replaced by newly synthesized chains. Since the TCR heterodimer cannot reach the cell membrane on its own, we propose a model in which recycling CD3 is transported along a retrograde pathway to the endoplasmic reticulum, where it associates with newly made TCR. Interestingly, engagement of the TCR.CD3 complex by superantigen resulted not only in the down-regulation of the TCR and CD3 components but also caused a transient stabilization of the TCR heterodimer. This suggests that TCR engagement diverts the TCR heterodimer from a degradation to a recycling pathway. Contrary to CD3, the intracellular fate of the TCR heterodimer is thus regulated, providing a mechanism for rapidly replacing nonfunctional TCR during intrathymic development of T cells.

Conformational and biochemical differences in the TCR.CD3 complex of CD8(+) versus CD4(+) mature lymphocytes revealed in the absence of CD3gamma.

Mature CD4(+) and CD8(+) T lymphocytes are believed to build and express essentially identical surface alphabeta T-cell receptor-CD3 (TCR.CD3) complexes. However, TCR.CD3 expression has been shown to be more impaired in CD8(+) cells than in CD4(+) cells when CD3gamma is absent in humans or mice. We have addressed this paradox by performing a detailed phenotypical and biochemical analysis of the TCR.CD3 complex in human CD3gamma-deficient CD8(+) and CD4(+) T cells. The results indicated that the membrane TCR.CD3 complex of CD8(+) T lymphocytes was conformationally different from that of CD4(+) lymphocytes in the absence of CD3gamma. In addition, CD8(+), but not CD4(+), CD3gamma-deficient T lymphocytes were shown to contain abnormally glycosylated TCRbeta proteins, together with a smaller, abnormal TCR chain (probably incompletely processed TCRalpha). These results suggest the existence of hitherto unrecognized biochemical differences between mature CD4(+) and CD8(+) T lymphocytes in the intracellular control of alphabetaTCR. CD3 assembly, maturation, or transport that are revealed when CD3gamma is absent. Such lineage-specific differences may be important in receptor-coreceptor interactions during antigen recognition.