RESUMEN
Epithelial cells lining the intestinal mucosa constitute a selective-semipermeable barrier acting as first line of defense in the organism. The number of those cells remains constant during physiological conditions, but disruption of epithelial cell homeostasis has been observed in several pathologies. During colitis, epithelial cell proliferation decreases and cell death augments. The mechanism responsible for these changes remains unknown. Here, we show that the pro-inflammatory cytokine IFNγ contributes to the inhibition of epithelial cell proliferation in intestinal epithelial cells (IECs) by inducing the activation of mTORC1. Activation of mTORC1 in response to IFNγ was detected in IECs present along the crypt axis and in colonic macrophages. mTORC1 inhibition enhances cell proliferation, increases DNA damage in IEC. In macrophages, mTORC1 inhibition strongly reduces the expression of pro-inflammatory markers. As a consequence, mTORC1 inhibition exacerbated disease activity, increased mucosal damage, enhanced ulceration, augmented cell infiltration, decreased survival and stimulated tumor formation in a model of colorectal cancer CRC associated to colitis. Thus, our findings suggest that mTORC1 signaling downstream of IFNγ prevents epithelial DNA damage and cancer development during colitis.
RESUMEN
The intestinal epithelium constitutes a first line of defense of the innate immune system. Epithelial dysfunction is a hallmark of intestinal disorders such as inflammatory bowel diseases (IBDs). The actin cytoskeleton controls epithelial barrier integrity but the function of actin regulators such as cortactin is poorly understood. Given that cortactin controls endothelial permeability, we hypothesized that cortactin is also important for epithelial barrier regulation. We found increased permeability in the colon of cortactin-KO mice that was accompanied by reduced levels of ZO-1, claudin-1, and E-cadherin. By contrast, claudin-2 was upregulated. Cortactin deficiency increased RhoA/ROCK1-dependent actomyosin contractility, and inhibition of ROCK1 rescued the barrier defect. Interestingly, cortactin deficiency caused increased epithelial proliferation without affecting apoptosis. KO mice did not develop spontaneous colitis, but were more susceptible to dextran sulfate sodium colitis and showed severe colon tissue damage and edema formation. KO mice with colitis displayed strong mucus deposition and goblet cell depletion. In healthy human colon tissues, cortactin co-localized with ZO-1 at epithelial cell contacts. In IBDs patients, we observed decreased cortactin levels and loss of co-localization with ZO-1. Thus, cortactin is a master regulator of intestinal epithelial barrier integrity in vivo and could serve as a suitable target for pharmacological intervention in IBDs.
Asunto(s)
Actomiosina/metabolismo , Colitis/inmunología , Cortactina/metabolismo , Enfermedades Inflamatorias del Intestino/inmunología , Mucosa Intestinal/patología , Quinasas Asociadas a rho/metabolismo , Proteína de Unión al GTP rhoA/metabolismo , Animales , Apoptosis , Proliferación Celular , Colitis/inducido químicamente , Cortactina/genética , Citoesqueleto/metabolismo , Sulfato de Dextran , Modelos Animales de Enfermedad , Humanos , Inmunidad Innata , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteína de la Zonula Occludens-1/metabolismo , Proteína de Unión al GTP rhoA/genéticaRESUMEN
BACKGROUND AND OBJECTIVE: Understanding the causes of disagreement among experts in clinical decision making has been a challenge for decades. In particular, a high amount of variability exists in diagnosis of retinopathy of prematurity (ROP), which is a disease affecting low birth weight infants and a major cause of childhood blindness. A possible cause of variability, that has been mostly neglected in the literature, is related to discrepancies in the sets of important features considered by different experts. In this paper we propose a methodology which makes use of machine learning techniques to understand the underlying causes of inter-expert variability. METHODS: The experiments are carried out on a dataset consisting of 34 retinal images, each with diagnoses provided by 22 independent experts. Feature selection techniques are applied to discover the most important features considered by a given expert. Those features selected by each expert are then compared to the features selected by other experts by applying similarity measures. Finally, an automated diagnosis system is built in order to check if this approach can be helpful in solving the problem of understanding high inter-rater variability. RESULTS: The experimental results reveal that some features are mostly selected by the feature selection methods regardless the considered expert. Moreover, for pairs of experts with high percentage agreement among them, the feature selection algorithms also select similar features. By using the relevant selected features, the classification performance of the automatic system was improved or maintained. CONCLUSIONS: The proposed methodology provides a handy framework to identify important features for experts and check whether the selected features reflect the pairwise agreements/disagreements. These findings may lead to improved diagnostic accuracy and standardization among clinicians, and pave the way for the application of this methodology to other problems which present inter-expert variability.
Asunto(s)
Aprendizaje Automático , Retinopatía de la Prematuridad/patología , Humanos , LactanteRESUMEN
Guanylate-binding protein-1 (GBP-1) is an interferon inducible large GTPase involved in endothelial cell proliferation and invasion. In this report, expression and function of GBP-1 were investigated in vitro in intestinal epithelia after exposure to interferon-gamma and in human colonic mucosa from individuals with inflammatory bowel disease (IBD). Interestingly, in contrast to other epithelia, GBP-1 distributed to the plasma membrane in intestinal epithelial cells where it colocalized with the tight junction protein coxsackie- and adenovirus receptor. In addition, expression of GBP-1 was upregulated in colonic epithelia of individuals with IBD. Downregulation of GBP-1 by siRNA resulted in enhanced permeability that correlated with increased apoptosis. Indeed, inhibition of caspase activity prevented the inhibition of barrier formation induced by the loss of GBP-1. These data suggest that GBP-1 is a novel marker of intestinal mucosal inflammation that may protect against epithelial apoptosis induced by inflammatory cytokines and subsequent loss of barrier function.
Asunto(s)
Apoptosis/efectos de los fármacos , Células Epiteliales/metabolismo , Proteínas de Unión al GTP/metabolismo , Interferón gamma/farmacología , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Uniones Estrechas/metabolismo , Apoptosis/inmunología , Línea Celular , Colon/inmunología , Colon/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Células Epiteliales/inmunología , Proteínas de Unión al GTP/genética , Humanos , Enfermedades Inflamatorias del Intestino/genética , Enfermedades Inflamatorias del Intestino/inmunología , Enfermedades Inflamatorias del Intestino/metabolismo , ARN Interferente Pequeño/genéticaRESUMEN
A fundamental function of epithelia and endothelia is to separate different compartments within the organism and to regulate the exchange of substances between them. The tight junction (TJ) constitutes the barrier both to the passage of ions and molecules through the paracellular pathway and to the movement of proteins and lipids between the apical and the basolateral domains of the plasma membrane. In recent years more than 40 different proteins have been discovered to be located at the TJs of epithelia, endothelia and myelinated cells. This unprecedented expansion of information has changed our view of TJs from merely a paracellular barrier to a complex structure involved in signaling cascades that control cell growth and differentiation. Both cortical and transmembrane proteins integrate TJs. Among the former are scaffolding proteins containing PDZ domains, tumor suppressors, transcription factors and proteins involved in vesicle transport. To date two components of the TJ filaments have been identified: occludin and claudin. The latter is a protein family with more than 20 members. Both occludin and claudins are integral proteins capable of interacting adhesively with complementary molecules on adjacent cells and of co-polymerizing laterally. These advancements in the knowledge of the molecular structure of TJ support previous physiological models that exhibited TJ as dynamic structures that present distinct permeability and morphological characteristics in different tissues and in response to changing natural, pathological or experimental conditions.
Asunto(s)
Células Epiteliales/fisiología , Uniones Estrechas , Animales , Humanos , Inmunoglobulinas/química , Lípidos , Vaina de Mielina/fisiología , Estructura Terciaria de Proteína , Xenopus/embriologíaRESUMEN
Zonula occludens 2 (ZO-2) protein is a tight-junction phos phorylated protein that belongs to the membrane-associated guanylate kinase ('MAGUK') family. Here we study the interaction between ZO-2 and protein kinase C (PKC). We have constructed two ZO-2 fusion proteins of the middle (3PSG) and C-terminal (AP) regions of the molecule and demonstrate that they are phosphorylated by PKC isoenzymes beta, epsilon, lambda and zeta. To understand the physiological significance of the interaction between ZO-2 and PKC, we analysed the phosphorylation state of ZO-2 immunoprecipitated from monolayers with mature tight junctions or from cells that either lack them or have them disassembled through Ca(2+) chelation. We found that in the latter condition the phosphorylation level of ZO-2 is significantly higher and is due to the action of both PKC and cAMP-dependent protein kinase. These results therefore suggest that the phosphorylated state of ZO-2 restrains its capacity to operate at the junctional complex.
Asunto(s)
Proteínas de la Membrana/metabolismo , Proteína Quinasa C/metabolismo , Uniones Estrechas/metabolismo , Animales , Calcio/metabolismo , Línea Celular , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Perros , Glutatión Transferasa/genética , Isoenzimas/metabolismo , Proteínas de la Membrana/genética , Fosforilación , Pruebas de Precipitina , Proteína Quinasa C beta , Proteína Quinasa C-epsilon , Proteínas Serina-Treonina Quinasas/metabolismo , Estructura Terciaria de Proteína/genética , Ratas , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Serina/metabolismo , Treonina/metabolismo , Uniones Estrechas/enzimología , Proteína de la Zonula Occludens-2RESUMEN
The recognition of accelerative and decelerative patterns in the fetal heart rate (FHR) is one of the tasks carried out manually by obstetricians when they analyze cardiotocograms for information respecting the fetal state. An approach based on artificial neural networks formed by a multilayer perceptron (MLP) is developed. However, since the system utilizes the FHR signal as direct input, an anterior stage must be incorporated that applies a principal component analysis (PCA) so as to make the system independent of the signal baseline. Furthermore, the introduction of multiresolution into the PCA has resolved other problems that were detected in the application of the system. Presented in this paper are the results of validation of these systems designated the PCA-MLP and multiresolutlon principal component analysis (MR-PCA) systems against three clinical experts.
RESUMEN
ZO-1, ZO-2 and ZO-3 are tight junction (TJ)-associated proteins that belong to the MAGUK family. In addition to the presence of the characteristic MAGUK modules (PDZ, SH3 and GK), ZOs have a distinctive carboxyl terminal with splicing domains, acidic- and proline-rich regions. The modular organization of these proteins allows them to function as scaffolds, which associate to transmembrane TJ proteins, the cytoskeleton and signal transduction molecules. ZOs shuttle between the TJ and the nucleus, where they may regulate gene expression.
Asunto(s)
Nucleósido-Fosfato Quinasa/química , Nucleósido-Fosfato Quinasa/fisiología , Uniones Estrechas/fisiología , Animales , Proteínas Portadoras/química , Proteínas Portadoras/fisiología , Conexina 43/fisiología , Citoesqueleto/fisiología , Guanilato-Quinasas , Humanos , Sustancias Macromoleculares , Proteínas de la Membrana/química , Proteínas de la Membrana/fisiología , Ocludina , Fosfoproteínas/química , Fosfoproteínas/fisiología , Fosforilación , Estructura Terciaria de Proteína , Proteínas de la Zonula Occludens , Proteína de la Zonula Occludens-1 , Proteína de la Zonula Occludens-2 , Dominios Homologos srcRESUMEN
A common situation in the field of medicine is the availability of a huge quantity of data and knowledge relevant to a problem which is nevertheless, to a greater or lesser degree, incomplete or imprecise. This kind of problem occurs, for example, with respect to the information available for the prognostic tasks required for pregnancy monitoring, where decision-making by physicians calls for the incorporation of predictive skills. There are available however, several knowledge discovery methods that can be applied to data resulting from the performance of one or several of the non-stress tests (NSTs) that are used to evaluate a pregnant patient's antenatal status. This paper presents, discusses and compares the results obtained as a consequence of the application of different prediction methods, namely the Bayes' model, discriminant analysis, artificial neural networks (ANNs) and the Shortliffe and Buchanan uncertainty-based model.
Asunto(s)
Redes Neurales de la Computación , Resultado del Embarazo , Simulación por Computador , Sistemas Especialistas , Femenino , Humanos , Modelos Estadísticos , Valor Predictivo de las Pruebas , EmbarazoRESUMEN
Most of the information on the structure and function of the tight junction (TJ) has been obtained in MDCK cells. Accordingly, we have sequenced ZO-1 in this cell type, because this protein is involved in the response of the TJ to changes in Ca2+, phosphorylation, and the cytoskeleton. ZO-1 of MDCK cells comprises 6805 bp with a predicted open reading frame of 1769 amino acids. This sequence is 92 and 87% homologous to human and mouse ZO-1, respectively. Two nuclear sorting signals located at the PDZ1 and GK domains and 17 SH3 putative binding sites at the proline-rich domain were detected. We found two new splicing regions at the proline-rich region: beta had not been reported in human and mouse counterparts, and gamma, which was previously sequenced in human and mouse ZO-1, is now identified as a splicing region. The expression of different beta and gamma isoforms varies according to the tissue tested. With the information provided by the sequence, Southern blot, and PCR experiments we can predict a single genomic copy of MDCK-ZO-1 that is at least 13.16 kb long. MDCK-ZO-1 mRNA is 7.4 kb long. Its expression is regulated by calcium, while the expression of MDCK-ZO-1 protein is not.
Asunto(s)
Proteínas de la Membrana/genética , Fosfoproteínas/genética , Uniones Estrechas , Empalme Alternativo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Sitios de Unión , Calcio/metabolismo , Línea Celular , Núcleo Celular , ADN Complementario , Perros , Regulación de la Expresión Génica , Humanos , Proteínas de la Membrana/clasificación , Proteínas de la Membrana/metabolismo , Ratones , Datos de Secuencia Molecular , Fosfoproteínas/clasificación , Fosfoproteínas/metabolismo , Filogenia , Isoformas de Proteínas , ARN Mensajero , Homología de Secuencia de Aminoácido , Transducción de Señal , Proteína de la Zonula Occludens-1 , Dominios Homologos srcRESUMEN
Validation of intelligent systems is an important task to perform. Typically the results of the validation analysis are used to verify whether or not the system satisfies the initial design requirements, and to acquire new knowledge and/or refine the knowledge already acquired. In practice, the validation of intelligent systems usually requires the application of several different techniques (e.g., retrospective, prospective, quantitative). In this work the authors present the methodology devised to validate PATRICIA: an intelligent monitoring system designed to advise clinicians on the management of patients dependent on mechanical ventilation. The application of this methodology requires that appropriate validation paradigms are selected, depending on both the application domain and the characteristics of the intelligent system. The article also presents and discusses validation results.
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Unidades de Cuidados Intensivos , Monitoreo Fisiológico , Sistemas Especialistas , Humanos , Respiración Artificial , Programas InformáticosRESUMEN
NST-EXPERT is a medical expert system designed to evaluate fetal condition (especially in 'high risk' pregnancies). The system reasons based on data from the Non-Stress Test (NST), a monitoring test widely used in obstetrics. From specific limited maternal-fetal context information together with the analysis of the NST results the system infers a diagnosis for each case, elaborates a therapeutic plan, and suggests a prognosis of an early neonatal outcome. This article describes a new version based on a previous research prototype. Such systems can aid clinicians of varied experience in assessing and managing complicated pregnancies.
Asunto(s)
Sistemas Especialistas , Embarazo de Alto Riesgo , Diagnóstico Prenatal/métodos , Femenino , Humanos , Embarazo , Diagnóstico Prenatal/instrumentación , PronósticoRESUMEN
The system described, VISC, has as its aim the acquisition and subsequent digitization of monodimensional cardiotocographic signals. These signals can originate in any type of hospital cardiotocographic register, as well as in a book of uterine activity (UA) or fetal heart rate (FHR) signals. VISC consists of a set of algorithms to identify the fetal heart rate signal, although it can be used with minor corrections to work on any continuous curve. Some of the problems that may arise, such as signal loss and presence of noise, are discussed to illustrate the proposed method. The program is integrated into a wider environment, whose main nucleus is the NST-EXPERT expert system, for the diagnosis of fetal well-being, the recommendation of therapeutical plans and the prognosis of immediate neonatal states.
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Cardiotocografía/métodos , Sistemas Especialistas , Enfermedades Fetales/diagnóstico , Algoritmos , Computadores , Computadores Analógicos , Femenino , Enfermedades Fetales/fisiopatología , Monitoreo Fetal/métodos , Frecuencia Cardíaca Fetal/fisiología , Humanos , Embarazo , Diseño de Software , EspañaRESUMEN
Assessment of the fetus in a high-risk pregnancy uses a variety of tests for screening and continued detection of in utero compromise. This paper describes Foetos, an expert system designed to help clinical personnel to interpret several fetal assessment tests: fetal biophysical profile, contraction stress test, and nonstress tests. Foetos has been built using the knowledge engineering tool Genie, which adopts a mixed frame- and rule-based approach to represent the clinical knowledge in the field. Foetos includes diagnostic, prognostic, and therapeutic structures, based on heuristic interpretation of such tests and contextual structures which relate interpretation to the overall clinical picture. The results of initial retrospective and prospective program validation are included in the report. These results show a substantial level of agreement between Foetos' recommendations and clinical management. Interpretation of the results indicates that the obstetrical field could be an area of interest for the application of AI techniques.
Asunto(s)
Diagnóstico por Computador , Sistemas Especialistas , Monitoreo Fetal/métodos , Tamizaje Masivo , Femenino , Humanos , Sistemas en Línea , Embarazo , Programas Informáticos , Interfaz Usuario-ComputadorRESUMEN
In intensive care units (ICUs), certain parameters must be interpreted taking into account the intrinsic characteristics of each patient or the peculiarities of the clinical case under consideration. Similar temporal evolutions of some parameters in different patients could have different interpretations. Artificial intelligence techniques can aid in resolving this problem through the construction of expert systems (ES). These systems are capable of performing contextual evaluations of the parameters typically monitored in ICUs. This contextual evaluation is usually carried out using symbolic elements. Thus, the symbolic processing of numeric data is an important task to perform. In any event, the assignment of semantic labels to numeric values is always an uncertain and arbitrary process. This suggests the convenience of defining and implementing representation schemes capable of dealing with uncertain knowledge. This paper presents a model for the symbolic processing of numeric variables in which the uncertainty associated with the assignment of literals appears spontaneously. The categorical approach for the symbolic classification of numeric values is a particular case of the proposed model.
Asunto(s)
Sistemas Especialistas , Unidades de Cuidados Intensivos , Monitoreo Fisiológico , Interpretación Estadística de Datos , Modelos Teóricos , Estados UnidosRESUMEN
The withdrawal of mechanical ventilation generally presents no problems if carried out when the values of certain predictive physiological variables suggest that the patient will be able to breathe for him or herself. In certain cases, however, a more gradual weaning procedure is necessary, in which the amount of mechanical assistance provided is in some way reduced little by little. One of the usual means of achieving this gradual reduction is by intermittent mandatory ventilation (IMV), in which short bursts of mechanical ventilation are separated by progressively longer periods of "ventilator silence" which stimulate the patient's endogenous respiration. ESTER is an expert system in which the most widely accepted criteria are systematically called upon to supply advice to the clinician who has to prescribe a respiratory therapy regime suited to the needs of the patient. After effecting a preliminary prognosis of the patient's condition, ESTER asks for certain physiological parameters to be keyed in. Analysis of these parameters reveals the patient's condition and allows a recommended respiratory therapy to be designed.