Mammalian pre-mRNA 3' end processing factor CF I m 68 functions in mRNA export

Export of mRNA from the nucleus is linked to proper processing and packaging into ribonucleoprotein complexes. Although several observations indicate a coupling between mRNA 3' end formation and export, it is not known how these two processes are mechanistically connected. Here, we show that a subunit of the mammalian pre-mRNA 3' end processing complex, CF I(m)68, stimulates mRNA export. CF I(m)68 shuttles between the nucleus and the cytoplasm in a transcription-dependent manner and interacts with the mRNA export receptor NXF1/TAP. Consistent with the idea that CF I(m)68 may act as a novel adaptor for NXF1/TAP, we show that CF I(m)68 promotes the export of a reporter mRNA as well as of endogenous mRNAs, whereas silencing by RNAi results in the accumulation of mRNAs in the nucleus. Moreover, CF I(m)68 associates with 80S ribosomes but not polysomes, suggesting that it is part of the mRNP that is remodeled in the cytoplasm during the initial stages of translation. These results reveal a novel function for the pre-mRNA 3' end processing factor CF I(m)68 in mRNA export.

Trastuzumab treatment beyond progression in advanced breast cancer: patterns of care in six Swiss breast cancer centers.

BACKGROUND Trastuzumab is an established treatment for HER2-positive breast cancer (BC). We analyzed Swiss patterns of care in patients with HER2-positive BC after disease progression on trastuzumab-containing therapy for metastatic BC (MBC). PATIENTS AND METHODS A retrospective analysis was performed in six Swiss BC centers. Patients with HER2-positive MBC treated with at least one infusion of trastuzumab for advanced disease between January 2006 and December 2007 were identified. Treatment patterns in first and further lines were analyzed. RESULTS All of the 72 identified patients received trastuzumab as their first palliative anti-HER2 therapy, either as monotherapy (n = 23) or in combination with chemotherapy (typically taxane or vinorelbine; n = 49). Median time to progression was 8.1, 8.0 and 7.9 months in the monotherapy, trastuzumab-taxane and trastuzumab-vinorelbine cohorts, respectively. After progression on first-line anti-HER2 therapy, trastuzumab was continued in 67 of 68 patients who received further therapy. One patient received second-line lapatinib plus capecitabine. The median duration of anti-HER2 therapy was 20 months. Patients received a median of 4 lines of anti-HER2 therapy. CONCLUSIONS Durable responses were achieved with repeated exposure to anti-HER2 therapy. In a selected patient population, trastuzumab monotherapy appears to be a reasonable first-line treatment option.

In-vitro activity of taurolidine on single species and a multispecies population associated with periodontitis.

The antimicrobial activity of taurolidine was compared with minocycline against microbial species associated with periodontitis (four single strains and a 12-species mixture). Minimal inhibitory concentrations (MICs) and minimal bactericidal concentrations (MBCs), killing as well as activities on established and forming single-species biofilms and a 12-species biofilm were determined. The MICs of taurolidine against single species were always 0.31 mg/ml, the MBCs were 0.64 mg/ml. The used mixed microbiota was less sensitive to taurolidine, MIC and the MBC was 2.5 mg/ml. The strains and the mixture were completely killed by 2.5 mg/ml taurolidine, whereas 256 μg/ml minocycline reduced the bacterial counts of the mixture by 5 log10 colony forming units (cfu). Coating the surface with 10 mg/ml taurolidine or 256 μg/ml minocycline prevented completely biofilm formation of Porphyromonas gingivalis ATCC 33277 but not of Aggregatibacter actinomycetemcomitans Y4 and the mixture. On 4.5 d old biofilms, taurolidine acted concentration dependent with a reduction by 5 log10 cfu (P. gingivalis ATCC 33277) and 7 log10 cfu (A. actinomycetemcomitans Y4) when applying 10 mg/ml. Minocycline decreased the cfu counts by 1-2 log10 cfu independent of the used concentration. The reduction of the cfu counts in the 4.5 d old multi-species biofilms was about 3 log10 cfu after application of any minocycline concentration and after using 10 mg/ml taurolidine. Taurolidine is active against species associated with periodontitis, even within biofilms. Nevertheless a complete elimination of complex biofilms by taurolidine seems to be impossible and underlines the importance of a mechanical removal of biofilms prior to application of taurolidine.

Selective resistance to the PARP inhibitor olaparib in a mouse model for BRCA1-deficient metaplastic breast cancer

Metaplastic breast carcinoma (MBC) is a rare histological breast cancer subtype characterized by mesenchymal elements and poor clinical outcome. A large fraction of MBCs harbor defects in breast cancer 1 (BRCA1). As BRCA1 deficiency sensitizes tumors to DNA cross-linking agents and poly(ADP-ribose) polymerase (PARP) inhibitors, we sought to investigate the response of BRCA1-deficient MBCs to the PARP inhibitor olaparib. To this end, we established a genetically engineered mouse model (GEMM) for BRCA1-deficient MBC by introducing the MET proto-oncogene into a BRCA1-associated breast cancer model, using our novel female GEMM ES cell (ESC) pipeline. In contrast to carcinomas, BRCA1-deficient mouse carcinosarcomas resembling MBC show intrinsic resistance to olaparib caused by increased P-glycoprotein (Pgp) drug efflux transporter expression. Indeed, resistance could be circumvented by using another PARP inhibitor, AZD2461, which is a poor Pgp substrate. These preclinical findings suggest that patients with BRCA1-associated MBC may show poor response to olaparib and illustrate the value of GEMM-ESC models of human cancer for evaluation of novel therapeutics.

Evaluation of memory B cells among perinatally HIV infected children from Houston Texas

Background: HIV associated B cell exhaustion is a notable characteristic of HIV viremic adults. However, it is not known if such alterations are present in perinatal HIV infected children, whose viral dynamics differs from those seen in adults. In the present study we perform an analysis of B cells subsets and measure antigen-specific memory B cells (MBC) in a pediatric HIV infected cohort. ^ Methods: Peripheral mononuclear cells (PBMC) of perinatal HIV infected individuals are characterized into naïve (CD21hi/CD27−), classic (CD27+), tissue like (CD21lo/CD27 −) and activated MBC (CD27+CD21− ) by FACS. A memory ELISPOT assay is used to detect antibody secreting cells. We measure total IgG and antibodies specific for influenza, HBV, mumps, measles, rubella and VZV. Memory was expressed as spot forming cells (SPC) /million of PBMC. Wilcoxon rank-sum was used to compare unpaired groups and linear regression analysis was used to determine predictors of B cell dysfunction ^ Results: 41 HIV perinatal infected children are included (51.2% females and 65.9% Black). Age at study is median (range) 8.78 years (4.39-11.57). At the time of testing they have a CD4% of 30.9 (23.2-39.4), a viral load (VL) of 1.95 log10 copies/ml (1.68-3.29) and a cumulative VL of 3.4 log10 copy × days (2.7-4.0). Ninety two percent of the children are on cARV for > 6 months. Overall, HIV+ children compared with controls have a significant lower number of IgG and antigen specific SFC. In addition, they have a lower proportion of classical MBC 12.9 (8.09-19.85) vs 29.4 (18.7-39.05); 0.01, but a significant higher proportion of tissue like memory MBC 6.01 (2.79-12.7) vs 0.99 (0.87-1.38); 0.003, compared with controls. Patients are parsed on VL (<400 and ≥ 400 copies/ml) with the objective to evaluate the effect of VL on B cell status. Patients with a VL ≥ 400 copies/ml have a significantly lower IgG, HBV, measles, rubella and VZV SPC compared with those with a VL < 400 copies/ml. There are no significant differences in B cell subpopulations between the groups. A moderate negative correlation was observed between the time of cARV initiation and the frequency of IgG memory B cells, suggesting that early initiation of cARV appears to lead to a better functionality of the IgG memory B cells (P=0.05). A statistically significant positive correlation was observed between the total number of IgG memory cells and the number of antigen-specific memory B cells/SPCs. Suggesting that the progressive recovery of the IgG memory B cell pull goes along with a progressive increase in the number of antigen-specific SPCs. ^ Conclusion: A pediatric cohort in overall good status with respect to HIV infection and on ART has defects in B cell function and numbers (reduced total and antigen specific MBC and increased tissue like and reduced classical MBC).^

Análisis de los factores determinantes de la repetición del proceso de compra en el comercio electrónico entre empresas y consumidores finales. Aplicación al mercado español

Las Tecnologías de la Información y la Comunicación en general e Internet en particular han supuesto una revolución en nuestra forma de comunicarnos, relacionarnos, producir, comprar y vender acortando tiempo y distancias entre proveedores y consumidores. A la paulatina penetración del ordenador, los teléfonos inteligentes y la banda ancha fija y/o móvil ha seguido un mayor uso de estas tecnologías entre ciudadanos y empresas. El comercio electrónico empresa–consumidor (B2C) alcanzó en 2010 en España un volumen de 9.114 millones de euros, con un incremento del 17,4% respecto al dato registrado en 2009. Este crecimiento se ha producido por distintos hechos: un incremento en el porcentaje de internautas hasta el 65,1% en 2010 de los cuales han adquirido productos o servicios a través de la Red un 43,1% –1,6 puntos porcentuales más respecto a 2010–. Por otra parte, el gasto medio por comprador ha ascendido a 831€ en 2010, lo que supone un incremento del 10,9% respecto al año anterior. Si segmentamos a los compradores según por su experiencia anterior de compra podemos encontrar dos categorías: el comprador novel –que adquirió por primera vez productos o servicios en 2010– y el comprador constante –aquel que había adquirido productos o servicios en 2010 y al menos una vez en años anteriores–. El 85,8% de los compradores se pueden considerar como compradores constantes: habían comprado en la Red en 2010, pero también lo habían hecho anteriormente. El comprador novel tiene un perfil sociodemográfico de persona joven de entre 15–24 años, con estudios secundarios, de clase social media y media–baja, estudiante no universitario, residente en poblaciones pequeñas y sigue utilizando fórmulas de pago como el contra–reembolso (23,9%). Su gasto medio anual ascendió en 2010 a 449€. El comprador constante, o comprador que ya había comprado en Internet anteriormente, tiene un perfil demográfico distinto: estudios superiores, clase alta, trabajador y residente en grandes ciudades, con un comportamiento maduro en la compra electrónica dada su mayor experiencia –utiliza con mayor intensidad canales exclusivos en Internet que no disponen de tienda presencial–. Su gasto medio duplica al observado en compradores noveles (con una media de 930€ anuales). Por tanto, los compradores constantes suponen una mayoría de los compradores con un gasto medio que dobla al comprador que ha adoptado el medio recientemente. Por consiguiente es de interés estudiar los factores que predicen que un internauta vuelva a adquirir un producto o servicio en la Red. La respuesta a esta pregunta no se ha revelado sencilla. En España, la mayoría de productos y servicios aún se adquieren de manera presencial, con una baja incidencia de las ventas a distancia como la teletienda, la venta por catálogo o la venta a través de Internet. Para dar respuesta a las preguntas planteadas se ha investigado desde distintos puntos de vista: se comenzará con un estudio descriptivo desde el punto de vista de la demanda que trata de caracterizar la situación del comercio electrónico B2C en España, poniendo el foco en las diferencias entre los compradores constantes y los nuevos compradores. Posteriormente, la investigación de modelos de adopción y continuidad en el uso de las tecnologías y de los factores que inciden en dicha continuidad –con especial interés en el comercio electrónico B2C–, permiten afrontar el problema desde la perspectiva de las ecuaciones estructurales pudiendo también extraer conclusiones de tipo práctico. Este trabajo sigue una estructura clásica de investigación científica: en el capítulo 1 se introduce el tema de investigación, continuando con una descripción del estado de situación del comercio electrónico B2C en España utilizando fuentes oficiales (capítulo 2). Posteriormente se desarrolla el marco teórico y el estado del arte de modelos de adopción y de utilización de las tecnologías (capítulo 3) y de los factores principales que inciden en la adopción y continuidad en el uso de las tecnologías (capítulo 4). El capítulo 5 desarrolla las hipótesis de la investigación y plantea los modelos teóricos. Las técnicas estadísticas a utilizar se describen en el capítulo 6, donde también se analizan los resultados empíricos sobre los modelos desarrollados en el capítulo 5. El capítulo 7 expone las principales conclusiones de la investigación, sus limitaciones y propone nuevas líneas de investigación. La primera parte corresponde al capítulo 1, que introduce la investigación justificándola desde un punto de vista teórico y práctico. También se realiza una breve introducción a la teoría del comportamiento del consumidor desde una perspectiva clásica. Se presentan los principales modelos de adopción y se introducen los modelos de continuidad de utilización que se estudiarán más detalladamente en el capítulo 3. En este capítulo se desarrollan los objetivos principales y los objetivos secundarios, se propone el mapa mental de la investigación y se planifican en un cronograma los principales hitos del trabajo. La segunda parte corresponde a los capítulos dos, tres y cuatro. En el capítulo 2 se describe el comercio electrónico B2C en España utilizando fuentes secundarias. Se aborda un diagnóstico del sector de comercio electrónico y su estado de madurez en España. Posteriormente, se analizan las diferencias entre los compradores constantes, principal interés de este trabajo, frente a los compradores noveles, destacando las diferencias de perfiles y usos. Para los dos segmentos se estudian aspectos como el lugar de acceso a la compra, la frecuencia de compra, los medios de pago utilizados o las actitudes hacia la compra. El capítulo 3 comienza desarrollando los principales conceptos sobre la teoría del comportamiento del consumidor, para continuar estudiando los principales modelos de adopción de tecnología existentes, analizando con especial atención su aplicación en comercio electrónico. Posteriormente se analizan los modelos de continuidad en el uso de tecnologías (Teoría de la Confirmación de Expectativas; Teoría de la Justicia), con especial atención de nuevo a su aplicación en el comercio electrónico. Una vez estudiados los principales modelos de adopción y continuidad en el uso de tecnologías, el capítulo 4 analiza los principales factores que se utilizan en los modelos: calidad, valor, factores basados en la confirmación de expectativas –satisfacción, utilidad percibida– y factores específicos en situaciones especiales –por ejemplo, tras una queja– como pueden ser la justicia, las emociones o la confianza. La tercera parte –que corresponde al capítulo 5– desarrolla el diseño de la investigación y la selección muestral de los modelos. En la primera parte del capítulo se enuncian las hipótesis –que van desde lo general a lo particular, utilizando los factores específicos analizados en el capítulo 4– para su posterior estudio y validación en el capítulo 6 utilizando las técnicas estadísticas apropiadas. A partir de las hipótesis, y de los modelos y factores estudiados en los capítulos 3 y 4, se definen y vertebran dos modelos teóricos originales que den respuesta a los retos de investigación planteados en el capítulo 1. En la segunda parte del capítulo se diseña el trabajo empírico de investigación definiendo los siguientes aspectos: alcance geográfico–temporal, tipología de la investigación, carácter y ambiente de la investigación, fuentes primarias y secundarias utilizadas, técnicas de recolección de datos, instrumentos de medida utilizados y características de la muestra utilizada. Los resultados del trabajo de investigación constituyen la cuarta parte de la investigación y se desarrollan en el capítulo 6, que comienza analizando las técnicas estadísticas basadas en Modelos de Ecuaciones Estructurales. Se plantean dos alternativas, modelos confirmatorios correspondientes a Métodos Basados en Covarianzas (MBC) y modelos predictivos. De forma razonada se eligen las técnicas predictivas dada la naturaleza exploratoria de la investigación planteada. La segunda parte del capítulo 6 desarrolla el análisis de los resultados de los modelos de medida y modelos estructurales construidos con indicadores formativos y reflectivos y definidos en el capítulo 4. Para ello se validan, sucesivamente, los modelos de medida y los modelos estructurales teniendo en cuenta los valores umbrales de los parámetros estadísticos necesarios para la validación. La quinta parte corresponde al capítulo 7, que desarrolla las conclusiones basándose en los resultados del capítulo 6, analizando los resultados desde el punto de vista de las aportaciones teóricas y prácticas, obteniendo conclusiones para la gestión de las empresas. A continuación, se describen las limitaciones de la investigación y se proponen nuevas líneas de estudio sobre distintos temas que han ido surgiendo a lo largo del trabajo. Finalmente, la bibliografía recoge todas las referencias utilizadas a lo largo de este trabajo. Palabras clave: comprador constante, modelos de continuidad de uso, continuidad en el uso de tecnologías, comercio electrónico, B2C, adopción de tecnologías, modelos de adopción tecnológica, TAM, TPB, IDT, UTAUT, ECT, intención de continuidad, satisfacción, confianza percibida, justicia, emociones, confirmación de expectativas, calidad, valor, PLS. ABSTRACT Information and Communication Technologies in general, but more specifically those related to the Internet in particular, have changed the way in which we communicate, relate to one another, produce, and buy and sell products, reducing the time and shortening the distance between suppliers and consumers. The steady breakthrough of computers, Smartphones and landline and/or wireless broadband has been greatly reflected in its large scale use by both individuals and businesses. Business–to–consumer (B2C) e–commerce reached a volume of 9,114 million Euros in Spain in 2010, representing a 17.4% increase with respect to the figure in 2009. This growth is due in part to two different facts: an increase in the percentage of web users to 65.1% en 2010, 43.1% of whom have acquired products or services through the Internet– which constitutes 1.6 percentage points higher than 2010. On the other hand, the average spending by individual buyers rose to 831€ en 2010, constituting a 10.9% increase with respect to the previous year. If we select buyers according to whether or not they have previously made some type of purchase, we can divide them into two categories: the novice buyer–who first made online purchases in 2010– and the experienced buyer: who also made purchases in 2010, but had done so previously as well. The socio–demographic profile of the novice buyer is that of a young person between 15–24 years of age, with secondary studies, middle to lower–middle class, and a non–university educated student who resides in smaller towns and continues to use payment methods such as cash on delivery (23.9%). In 2010, their average purchase grew to 449€. The more experienced buyer, or someone who has previously made purchases online, has a different demographic profile: highly educated, upper class, resident and worker in larger cities, who exercises a mature behavior when making online purchases due to their experience– this type of buyer frequently uses exclusive channels on the Internet that don’t have an actual store. His or her average purchase doubles that of the novice buyer (with an average purchase of 930€ annually.) That said, the experienced buyers constitute the majority of buyers with an average purchase that doubles that of novice buyers. It is therefore of interest to study the factors that help to predict whether or not a web user will buy another product or use another service on the Internet. The answer to this question has proven not to be so simple. In Spain, the majority of goods and services are still bought in person, with a low amount of purchases being made through means such as the Home Shopping Network, through catalogues or Internet sales. To answer the questions that have been posed here, an investigation has been conducted which takes into consideration various viewpoints: it will begin with a descriptive study from the perspective of the supply and demand that characterizes the B2C e–commerce situation in Spain, focusing on the differences between experienced buyers and novice buyers. Subsequently, there will be an investigation concerning the technology acceptance and continuity of use of models as well as the factors that have an effect on their continuity of use –with a special focus on B2C electronic commerce–, which allows for a theoretic approach to the problem from the perspective of the structural equations being able to reach practical conclusions. This investigation follows the classic structure for a scientific investigation: the subject of the investigation is introduced (Chapter 1), then the state of the B2C e–commerce in Spain is described citing official sources of information (Chapter 2), the theoretical framework and state of the art of technology acceptance and continuity models are developed further (Chapter 3) and the main factors that affect their acceptance and continuity (Chapter 4). Chapter 5 explains the hypothesis behind the investigation and poses the theoretical models that will be confirmed or rejected partially or completely. In Chapter 6, the technical statistics that will be used are described briefly as well as an analysis of the empirical results of the models put forth in Chapter 5. Chapter 7 explains the main conclusions of the investigation, its limitations and proposes new projects. First part of the project, chapter 1, introduces the investigation, justifying it from a theoretical and practical point of view. It is also a brief introduction to the theory of consumer behavior from a standard perspective. Technology acceptance models are presented and then continuity and repurchase models are introduced, which are studied more in depth in Chapter 3. In this chapter, both the main and the secondary objectives are developed through a mind map and a timetable which highlights the milestones of the project. The second part of the project corresponds to Chapters Two, Three and Four. Chapter 2 describes the B2C e–commerce in Spain from the perspective of its demand, citing secondary official sources. A diagnosis concerning the e–commerce sector and the status of its maturity in Spain is taken on, as well as the barriers and alternative methods of e–commerce. Subsequently, the differences between experienced buyers, which are of particular interest to this project, and novice buyers are analyzed, highlighting the differences between their profiles and their main transactions. In order to study both groups, aspects such as the place of purchase, frequency with which online purchases are made, payment methods used and the attitudes of the purchasers concerning making online purchases are taken into consideration. Chapter 3 begins by developing the main concepts concerning consumer behavior theory in order to continue the study of the main existing acceptance models (among others, TPB, TAM, IDT, UTAUT and other models derived from them) – paying special attention to their application in e–commerce–. Subsequently, the models of technology reuse are analyzed (CDT, ECT; Theory of Justice), focusing again specifically on their application in e–commerce. Once the main technology acceptance and reuse models have been studied, Chapter 4 analyzes the main factors that are used in these models: quality, value, factors based on the contradiction of expectations/failure to meet expectations– satisfaction, perceived usefulness– and specific factors pertaining to special situations– for example, after receiving a complaint justice, emotions or confidence. The third part– which appears in Chapter 5– develops the plan for the investigation and the sample selection for the models that have been designed. In the first section of the Chapter, the hypothesis is presented– beginning with general ideas and then becoming more specific, using the detailed factors that were analyzed in Chapter 4– for its later study and validation in Chapter 6– as well as the corresponding statistical factors. Based on the hypothesis and the models and factors that were studied in Chapters 3 and 4, two original theoretical models are defined and organized in order to answer the questions posed in Chapter 1. In the second part of the Chapter, the empirical investigation is designed, defining the following aspects: geographic–temporal scope, type of investigation, nature and setting of the investigation, primary and secondary sources used, data gathering methods, instruments according to the extent of their use and characteristics of the sample used. The results of the project constitute the fourth part of the investigation and are developed in Chapter 6, which begins analyzing the statistical techniques that are based on the Models of Structural Equations. Two alternatives are put forth: confirmatory models which correspond to Methods Based on Covariance (MBC) and predictive models– Methods Based on Components–. In a well–reasoned manner, the predictive techniques are chosen given the explorative nature of the investigation. The second part of Chapter 6 explains the results of the analysis of the measurement models and structural models built by the formative and reflective indicators defined in Chapter 4. In order to do so, the measurement models and the structural models are validated one by one, while keeping in mind the threshold values of the necessary statistic parameters for their validation. The fifth part corresponds to Chapter 7 which explains the conclusions of the study, basing them on the results found in Chapter 6 and analyzing them from the perspective of the theoretical and practical contributions, and consequently obtaining conclusions for business management. The limitations of the investigation are then described and new research lines about various topics that came up during the project are proposed. Lastly, all of the references that were used during the project are listed in a final bibliography. Key Words: constant buyer, repurchase models, continuity of use of technology, e–commerce, B2C, technology acceptance, technology acceptance models, TAM, TPB, IDT, UTAUT, ECT, intention of repurchase, satisfaction, perceived trust/confidence, justice, feelings, the contradiction of expectations, quality, value, PLS.

Markov blanket-based approach for learning multi-dimensional Bayesian network classifiers: An application to predict the European Quality of Life-5 Dimensions (EQ-5D) from the 39-item Parkinson's Disease Questionnaire (PDQ-39)

Multi-dimensional Bayesian network classifiers (MBCs) are probabilistic graphical models recently proposed to deal with multi-dimensional classification problems, where each instance in the data set has to be assigned to more than one class variable. In this paper, we propose a Markov blanket-based approach for learning MBCs from data. Basically, it consists of determining the Markov blanket around each class variable using the HITON algorithm, then specifying the directionality over the MBC subgraphs. Our approach is applied to the prediction problem of the European Quality of Life-5 Dimensions (EQ-5D) from the 39-item Parkinson’s Disease Questionnaire (PDQ-39) in order to estimate the health-related quality of life of Parkinson’s patients. Fivefold cross-validation experiments were carried out on randomly generated synthetic data sets, Yeast data set, as well as on a real-world Parkinson’s disease data set containing 488 patients. The experimental study, including comparison with additional Bayesian network-based approaches, back propagation for multi-label learning, multi-label k-nearest neighbor, multinomial logistic regression, ordinary least squares, and censored least absolute deviations, shows encouraging results in terms of predictive accuracy as well as the identification of dependence relationships among class and feature variables.

Multi-dimensional classification using Bayesian networks for stationary and evolving streaming data

Hoy en día, con la evolución continua y rápida de las tecnologías de la información y los dispositivos de computación, se recogen y almacenan continuamente grandes volúmenes de datos en distintos dominios y a través de diversas aplicaciones del mundo real. La extracción de conocimiento útil de una cantidad tan enorme de datos no se puede realizar habitualmente de forma manual, y requiere el uso de técnicas adecuadas de aprendizaje automático y de minería de datos. La clasificación es una de las técnicas más importantes que ha sido aplicada con éxito a varias áreas. En general, la clasificación se compone de dos pasos principales: en primer lugar, aprender un modelo de clasificación o clasificador a partir de un conjunto de datos de entrenamiento, y en segundo lugar, clasificar las nuevas instancias de datos utilizando el clasificador aprendido. La clasificación es supervisada cuando todas las etiquetas están presentes en los datos de entrenamiento (es decir, datos completamente etiquetados), semi-supervisada cuando sólo algunas etiquetas son conocidas (es decir, datos parcialmente etiquetados), y no supervisada cuando todas las etiquetas están ausentes en los datos de entrenamiento (es decir, datos no etiquetados). Además, aparte de esta taxonomía, el problema de clasificación se puede categorizar en unidimensional o multidimensional en función del número de variables clase, una o más, respectivamente; o también puede ser categorizado en estacionario o cambiante con el tiempo en función de las características de los datos y de la tasa de cambio subyacente. A lo largo de esta tesis, tratamos el problema de clasificación desde tres perspectivas diferentes, a saber, clasificación supervisada multidimensional estacionaria, clasificación semisupervisada unidimensional cambiante con el tiempo, y clasificación supervisada multidimensional cambiante con el tiempo. Para llevar a cabo esta tarea, hemos usado básicamente los clasificadores Bayesianos como modelos. La primera contribución, dirigiéndose al problema de clasificación supervisada multidimensional estacionaria, se compone de dos nuevos métodos de aprendizaje de clasificadores Bayesianos multidimensionales a partir de datos estacionarios. Los métodos se proponen desde dos puntos de vista diferentes. El primer método, denominado CB-MBC, se basa en una estrategia de envoltura de selección de variables que es voraz y hacia delante, mientras que el segundo, denominado MB-MBC, es una estrategia de filtrado de variables con una aproximación basada en restricciones y en el manto de Markov. Ambos métodos han sido aplicados a dos problemas reales importantes, a saber, la predicción de los inhibidores de la transcriptasa inversa y de la proteasa para el problema de infección por el virus de la inmunodeficiencia humana tipo 1 (HIV-1), y la predicción del European Quality of Life-5 Dimensions (EQ-5D) a partir de los cuestionarios de la enfermedad de Parkinson con 39 ítems (PDQ-39). El estudio experimental incluye comparaciones de CB-MBC y MB-MBC con los métodos del estado del arte de la clasificación multidimensional, así como con métodos comúnmente utilizados para resolver el problema de predicción de la enfermedad de Parkinson, a saber, la regresión logística multinomial, mínimos cuadrados ordinarios, y mínimas desviaciones absolutas censuradas. En ambas aplicaciones, los resultados han sido prometedores con respecto a la precisión de la clasificación, así como en relación al análisis de las estructuras gráficas que identifican interacciones conocidas y novedosas entre las variables. La segunda contribución, referida al problema de clasificación semi-supervisada unidimensional cambiante con el tiempo, consiste en un método nuevo (CPL-DS) para clasificar flujos de datos parcialmente etiquetados. Los flujos de datos difieren de los conjuntos de datos estacionarios en su proceso de generación muy rápido y en su aspecto de cambio de concepto. Es decir, los conceptos aprendidos y/o la distribución subyacente están probablemente cambiando y evolucionando en el tiempo, lo que hace que el modelo de clasificación actual sea obsoleto y deba ser actualizado. CPL-DS utiliza la divergencia de Kullback-Leibler y el método de bootstrapping para cuantificar y detectar tres tipos posibles de cambio: en las predictoras, en la a posteriori de la clase o en ambas. Después, si se detecta cualquier cambio, un nuevo modelo de clasificación se aprende usando el algoritmo EM; si no, el modelo de clasificación actual se mantiene sin modificaciones. CPL-DS es general, ya que puede ser aplicado a varios modelos de clasificación. Usando dos modelos diferentes, el clasificador naive Bayes y la regresión logística, CPL-DS se ha probado con flujos de datos sintéticos y también se ha aplicado al problema real de la detección de código malware, en el cual los nuevos ficheros recibidos deben ser continuamente clasificados en malware o goodware. Los resultados experimentales muestran que nuestro método es efectivo para la detección de diferentes tipos de cambio a partir de los flujos de datos parcialmente etiquetados y también tiene una buena precisión de la clasificación. Finalmente, la tercera contribución, sobre el problema de clasificación supervisada multidimensional cambiante con el tiempo, consiste en dos métodos adaptativos, a saber, Locally Adpative-MB-MBC (LA-MB-MBC) y Globally Adpative-MB-MBC (GA-MB-MBC). Ambos métodos monitorizan el cambio de concepto a lo largo del tiempo utilizando la log-verosimilitud media como métrica y el test de Page-Hinkley. Luego, si se detecta un cambio de concepto, LA-MB-MBC adapta el actual clasificador Bayesiano multidimensional localmente alrededor de cada nodo cambiado, mientras que GA-MB-MBC aprende un nuevo clasificador Bayesiano multidimensional. El estudio experimental realizado usando flujos de datos sintéticos multidimensionales indica los méritos de los métodos adaptativos propuestos. ABSTRACT Nowadays, with the ongoing and rapid evolution of information technology and computing devices, large volumes of data are continuously collected and stored in different domains and through various real-world applications. Extracting useful knowledge from such a huge amount of data usually cannot be performed manually, and requires the use of adequate machine learning and data mining techniques. Classification is one of the most important techniques that has been successfully applied to several areas. Roughly speaking, classification consists of two main steps: first, learn a classification model or classifier from an available training data, and secondly, classify the new incoming unseen data instances using the learned classifier. Classification is supervised when the whole class values are present in the training data (i.e., fully labeled data), semi-supervised when only some class values are known (i.e., partially labeled data), and unsupervised when the whole class values are missing in the training data (i.e., unlabeled data). In addition, besides this taxonomy, the classification problem can be categorized into uni-dimensional or multi-dimensional depending on the number of class variables, one or more, respectively; or can be also categorized into stationary or streaming depending on the characteristics of the data and the rate of change underlying it. Through this thesis, we deal with the classification problem under three different settings, namely, supervised multi-dimensional stationary classification, semi-supervised unidimensional streaming classification, and supervised multi-dimensional streaming classification. To accomplish this task, we basically used Bayesian network classifiers as models. The first contribution, addressing the supervised multi-dimensional stationary classification problem, consists of two new methods for learning multi-dimensional Bayesian network classifiers from stationary data. They are proposed from two different points of view. The first method, named CB-MBC, is based on a wrapper greedy forward selection approach, while the second one, named MB-MBC, is a filter constraint-based approach based on Markov blankets. Both methods are applied to two important real-world problems, namely, the prediction of the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase and protease inhibitors, and the prediction of the European Quality of Life-5 Dimensions (EQ-5D) from 39-item Parkinson’s Disease Questionnaire (PDQ-39). The experimental study includes comparisons of CB-MBC and MB-MBC against state-of-the-art multi-dimensional classification methods, as well as against commonly used methods for solving the Parkinson’s disease prediction problem, namely, multinomial logistic regression, ordinary least squares, and censored least absolute deviations. For both considered case studies, results are promising in terms of classification accuracy as well as regarding the analysis of the learned MBC graphical structures identifying known and novel interactions among variables. The second contribution, addressing the semi-supervised uni-dimensional streaming classification problem, consists of a novel method (CPL-DS) for classifying partially labeled data streams. Data streams differ from the stationary data sets by their highly rapid generation process and their concept-drifting aspect. That is, the learned concepts and/or the underlying distribution are likely changing and evolving over time, which makes the current classification model out-of-date requiring to be updated. CPL-DS uses the Kullback-Leibler divergence and bootstrapping method to quantify and detect three possible kinds of drift: feature, conditional or dual. Then, if any occurs, a new classification model is learned using the expectation-maximization algorithm; otherwise, the current classification model is kept unchanged. CPL-DS is general as it can be applied to several classification models. Using two different models, namely, naive Bayes classifier and logistic regression, CPL-DS is tested with synthetic data streams and applied to the real-world problem of malware detection, where the new received files should be continuously classified into malware or goodware. Experimental results show that our approach is effective for detecting different kinds of drift from partially labeled data streams, as well as having a good classification performance. Finally, the third contribution, addressing the supervised multi-dimensional streaming classification problem, consists of two adaptive methods, namely, Locally Adaptive-MB-MBC (LA-MB-MBC) and Globally Adaptive-MB-MBC (GA-MB-MBC). Both methods monitor the concept drift over time using the average log-likelihood score and the Page-Hinkley test. Then, if a drift is detected, LA-MB-MBC adapts the current multi-dimensional Bayesian network classifier locally around each changed node, whereas GA-MB-MBC learns a new multi-dimensional Bayesian network classifier from scratch. Experimental study carried out using synthetic multi-dimensional data streams shows the merits of both proposed adaptive methods.

Isolation of an ovine pulmonary surfactant-associated anionic peptide bactericidal for Pasteurella haemolytica.

Ovine pulmonary surfactant is bactericidal for Pasteurella haemolytica when surfactant and bacteria mixtures are incubated with normal ovine serum. To isolate this component, surfactant (1 mg/ml) was centrifuged at 100,000 x gav, and the supernatant was fractionated by HPLC. Fractions were eluted with acetonitrile (10-100%)/0.1% trifluoracetic acid and tested for bactericidal activity. Amino acid and sequence analysis of three bactericidal fractions showed that fraction 2 contained H-GDDDDDD-OH, fraction 3 contained H-DDDDDDD-OH, and fraction 6 contained H-GADDDDD-OH. Peptides in 0.14 M NaCl/10 microM ZnCl2 (zinc saline solution) induced killing of P. haemolytica and other bacteria comparable to defensins and beta-defensins [minimal bactericidal concentration (MBC)50 range, 0.01-0.06 mM] but not in 0.14 M NaCl/10 mM sodium phosphate buffer, pH 7.2/0.5 mM CaCl2/0.15 mM MgCl2 (MBC50 range, 2.8-11.5 mM). Bactericidal activity resided in the core aspartate hexapeptide homopolymeric region, and MBC50 values of aspartate dipeptide-to-heptapeptide homopolymers were inversely proportional to the number of aspartate residues in the peptide. P. haemolytica incubated with H-DDDDDD-OH in zinc saline solution was killed within 30 min. Ultrastructurally, cells contained flocculated intracellular constituents. In contrast to cationic defensins and beta-defensins, surfactant-associated anionic peptides are smaller in size, opposite in charge, and are bactericidal in zinc saline solution. They are members of another class of peptide antibiotics containing aspartate, which when present in pulmonary secretions may help clear bacteria as a part of the innate pulmonary defense system.

Novel microbicidal agents derived from Naja atra cardiotoxin 1 (CTX-1)

During my PhD course, I focused my research on antimicrobial peptides (AMPs), in particular on the aspects of their computational design and development. This work led to the development of a new family of AMPs that I designed, starting from the amino acid sequence of a snake venom toxin, the cardiotoxin 1 (CTX-1) of Naja atra. Naja atra atra cardiotoxin 1, produced by Chinese cobra snakes belonging to Elapidae family, is included in the three-finger toxin family and exerts high cytotoxicity and antimicrobial activity too. This toxin family is characterized by specific folding of three beta-sheet loops (“fingers”) extending from the central core and by four conserved disulfide bridges. Using as template the first loop of this toxin, different sequences of 20 amino acids linear cationic peptides have been designed in order to avoid toxic effects but to maintain and strengthen the antimicrobial activity. As a result, the sequence NCP-0 (Naja Cardiotoxin Peptide-0) was designed as ancestor and subsequently other 4 variant sequences of NCP0 were developed. These variant sequences have shown microbicidal activity towards a panel of reference strains of Gram-positive and Gram-negative bacteria, fungi and an enveloped virus. In particular, the sequence designed as NCP-3 (Naja Cardiotoxin Peptide-3) and its variants NCP-3a and NCP-3b have shown the best antimicrobial activity together with low cytotoxicity against eukaryotic cells and low hemolytic activity. Bactericidal activity has been demonstrated by minimum bactericidal concentration (MBC) assay at values below 10 μg/ml for Pseudomonas aeruginosa ATCC 27853, Acinetobacter baumannii ( clinical isolates), Moraxella catharralis ATCC 25238, MRSA ATCC 43400, while towards Staphylococcus aureus ATCC 25923, Enterococcus hirae ATCC 10541 and Streptococcus agalactiae ATCC 13813 the bactericidal activity was demonstrated even below 1.6 μg/ml concentration. This potent antimicrobial activity was confirmed even for unicellular fungi Candida albicans, Candida glabrata and Malassezia pachydermatis (MBC 32.26-6.4 μg/ml), and also against the fast-growing mycobacteria Mycobacterium smegmatis DSMZ 43756 and Mycobacterium fortuitum DSMZ 46621 (MBC 100 μg/ml). Moreover, NCP-3 has shown a virucidal activity on the enveloped virus Bovine Herpesvirus 1 (BoHV1) belonging to herpesviridae family. The bactericidal activity is maintained in a high salt concentration (125 and 250 mM NaCl) medium and PB +20% Mueller Hinton Medium for E. coli, MRSA and Pseudomonas aeruginosa reference strains. Considering these in vitro obtained data, we propose NCP-3 and its variants NCP-3a and NCP-3b as promising antimicrobial candidates. For this reason, the whole novel AMPs family has been protected by a national patent (n°102015000015951).

Análise química e atividades biológicas de Guatteria elliptica R. E. Fries (Annonaceae)

A espécie endêmica G. elliptica R. E. Fries não apresentava estudos fitoquímicos e biológicos detalhados na literatura. Assim, o objetivo desse trabalho foi avaliar a composição química e as propriedades biológicas dos óleos essenciais, extratos brutos, alcaloides totais, tortas, frações das tortas, amostras isoladas dessa espécie. O material vegetal foi coletado em Paranapiacaba (Santo André, SP, Brasil). O óleo essencial extraído das folhas por destilação à vapor apresentou um rendimento de 0,2%. A análise histológica das folhas encontrou óleo em células oleíferas localizadas no parênquima esponjoso. A composição do óleo (CG-EM) indicou espatulenol e óxido de cariofileno como compostos majoritários. Os alcaloides totais foram obtidos dos extratos brutos das folhas e dos galhos e analisados por CG-EM, identificando quatro aporfinas (nornuciferina, estefarina, corituberina e asimilobina) e duas protoberberinas (discretamina e caseadina). Os alcaloides totais foram fracionados em coluna cromatográfica ou por Extração em Fase Sólida e purificados por cromatografia em camada preparativa, originando duas amostras (Amostra 9 e 10). Na Amostra 9, foram identificados dois alcaloides aporfinicos nornuciferina e asimilobina (CG-EM e RMN-1H). Na Amostra 10, foram identificados (LC-EM/EM) cinco alcaloides aporfínicos (desidronantenina, glaunidina, liriodenina, N-óxido de oliverina e telikovina) e um alcaloide protoberberínico (caseadina). Caseadina, glaunidina, N-óxido de oliverina e telikovina não foram previamente identificados em Guatteria. Os resíduos dos extratos brutos livres de alcaloides foram fracionados pelo método de partição com solventes de polaridade crescente. Os extratos brutos e as frações acetato de etila e butanólicas de folhas e galhos apresentaram flavonoides (NP-PEG). Nos ensaios biológicos, a melhor atividade antioxidante (sequestro do radical DPPH) foi encontrada para a fração clorofórmica dos galhos (EC50=24,25±1,14 µg/mL) e a torta dos galhos (EC50=26,23±4,20 µg/mL). No ensaio antimicrobiano pelo método turbidimétrico a atividade mais importante foi obtida contra Staphylococcus aureus (ATCC 6538) para os alcaloides totais dos galhos (CIM/CBM=0,12±0,01/0,26 mg/mL) e das folhas (CIM/CBM=0,21±0,01/0,28 mg/mL), e fração hexânica das folhas (CIM/CBM=0,24±0,02/>1 mg/mL). Uma alta atividade antitumoral foi observada frente a células humanas de mama (MCF-7) para Amostra 10 (IC50=2,28±0,18 µg/mL), fração de acetato de etila das folhas (IC50=4,47±0,40 µg/mL), óleo essencial (IC50=7,01±0,23 µg/mL) e os alcaloides totais das folhas (IC50=9,32±0,36 µg/mL). Para as células de próstata (PC-3), foi encontrada atividade para a Amostra 10 (IC50=1,37±0,36 µg/mL) e o óleo essencial (IC50=5,32±0,35 µg/mL). A futura aplicação dos extratos e frações de G. elliptica como um agente antitumoral parece ser segura, pois mantiveram uma viabilidade celular maior do que 90% no ensaio de citotoxicidade com culturas de fibroblasto murino (BALB/c 3T3, ATCC CCL-163) nas concentrações onde a atividade antitumoral foi promissora (<30 µg/mL) contra MCF-7 e/ou PC-3.

Efeito da aplicação de composto orgânico sobre a qualidade do solo cultivado com cana-de-açúcar (Saccharum spp.)

A dinâmica ambiental possui capacidade limitada de reciclagem e a crescente utilização resíduos agroindustriais, especialmente na agricultura, pode levar a situações de poluição do solo e demais componentes ambientais. A manutenção da produtividade de ecossistemas agrícolas e naturais depende do processo de transformação da matéria orgânica e, por conseguinte, da biomassa microbiana do solo, e que é responsável pela decomposição e mineralização de resíduos no mesmo. A dinâmica natural dos microrganismos do solo, em constante mudança e adaptação, os torna um indicador sensível às mudanças resultantes de diferentes práticas de manejo agrícola. Sendo assim, conhecer essas alterações e suas interferências é fundamental para identificar estratégias adequadas de manejo, apontando técnicas de utilização adequadas. O objetivo deste trabalho foi avaliar a qualidade de um solo agrícola, cultivado com três variedades de cana-de-açúcar (Saccharum spp.), comparando a utilização de adubação mineral frente à utilização de fertilizante orgânico composto no período final de formação dos perfilhos (120 dias após o plantio). Foi implantado, em condições de campo, o cultivo de cana-de-açúcar (cana planta), utilizando as variedades RB 867515, RB 962869 e RB 855453, onde cada variedade foi cultivada de três formas distintas, sendo elas: plantio controle (CT) sem aplicação de insumos para adubação; plantio orgânico (OG) com aplicação de fertilizante orgânico; e plantio convencional (CV) com aplicação de adubação mineral, seguindo recomendações de adubação após análise química inicial do solo local. Cada parcela possuía 37 m2, com 5 sulcos de 5,0 m de comprimento e espaçamento de 1,5 m entrelinhas, sendo os três sulcos centrais formando a área útil. De acordo com a variedade e o tipo de adubação, foram formados nove tratamentos: T1 86CT, T2 96CT, T3 85CT, T4 6OG, T5 96OG, T6 85OG, T7 86CV, T8 96CV e T9 85CV, com delineamento estatístico de blocos ao acaso e quatro repetições. Os parâmetros químicos do solo analisados foram macronutrientes e micronutrientes; os parâmetros microbiológicos foram carbono da biomassa microbiana (CBM), respiração basal do solo (RBS), quociente metabólico (qCO2), número mais provável de fungos e bactérias do solo (NMP); e, por fim, a produtividade agrícola (t/ha). Os resultados foram submetidos a análise de variância (ANOVA) e à comparação das médias através do teste de Tukey (10%). Também foi realizada a análise de variância dos dados e correlação cofenética de Pearson para formação de dendogramas. Com base no período estudado, considerado como fase crítica da formação do canavial, concluiu-se que os parâmetros químicos que evidenciaram alterações no solo foram pH e os macronutrientes Mg, Al e SB, sendo os tratamentos orgânicos equivalentes e/ou melhores que os tratamentos convencionais. Para os parâmetros microbiológicos, o NMP de fungos apresentou os maiores valores nos tratamentos convencionais e controle. A produtividade agrícola não foi influenciada pelos diferentes tratamentos e insumos utilizados, independente da variedade de cana-de-açúcar utilizada. Por fim, foram observadas correlações positivas entre as variáveis CTC e quociente metabólico (qCO2) apontando potencial melhoria da qualidade do solo, com o emprego de insumos orgânicos

Polo-like kinase 4 controls centriole duplication but does not directly regulate cytokinesis

Centrioles organize the centrosome, and accurate control of their number is critical for the maintenance of genomic integrity. Centrioles duplicate once per cell cycle, and duplication is coordinated by Polo-like kinase 4 (Plk4). We previously demonstrated that Plk4 accumulation is autoregulated by its own kinase activity. However, loss of heterozygosity of Plk4 in mouse embryonic fibroblasts has been proposed to cause cytokinesis failure as a primary event, leading to centrosome amplification and gross chromosomal abnormalities. Using targeted gene disruption, we show that human epithelial cells with one inactivated Plk4 allele undergo neither cytokinesis failure nor increase in centrosome amplification. Plk4 is shown to localize exclusively at the centrosome, with none in the spindle midbody. Substantial depletion of Plk4 by small interfering RNA leads to loss of centrioles and subsequent spindle defects that lead to a modest increase in the rate of cytokinesis failure. Therefore, Plk4 is a centriole-localized kinase that does not directly regulate cytokinesis.

Typical medullary breast carcinomas have a basal/myoepithelial phenotype

Medullary breast cancer (MBC) is a rare, diagnostically difficult, pathological subtype. Despite being high grade, it has a good prognosis. MBC patients have an excess of BRCA1 germ-fine mutation and reliable identification of MBC could help to identify patients at risk of carrying germline BRCA1 mutations or in whom chemotherapy could be avoided. The aim of this study was therefore to improve diagnosis by establishing an MBC protein expression profile using immunohistochemistry (IHC) on tissue-microarrays (TMA). Using a series of 779 breast carcinomas ('EC' set), diagnosed initially as MBC, a double-reading session was carried out by several pathologists on all of the histological material to establish the diagnosis as firmly as possible using a 'medullary score'. Only MBCs with high scores, i.e. typical MBC (TMBC) (n = 44) and non-TMBC grade III with no or low scores (n = 160), were included in the IHC study. To validate the results obtained on this first set, a control series of TMBC (n = 17) and non-MBC grade III cases (n = 140) ('IPC' set) was studied. The expression of 18 proteins was studied in the 61 TMBCs and 300 grade III cases from the two sets. The global intra-observer concordance of the first reading for the diagnosis of TMBC was 94%, with almost perfect kappa (kappa) of 0.815. TMBC was characterized by a high degree of basal/myoepithelial differentiation. In multivariate analysis with logistic regression, TMBC was defined by the association of P-cadherin (R = 2.29), MIB1 > 50 (R = 3.80), ERBB2 negativity (R = 2.24) and p53 positivity (RR = 1.45). Copyright (c) 2005 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.