Synergies and trade-offs between insect and pathogen resistance in maize leaves and roots

Determining links between plant defence strategies is important to understand plant evolution and to optimize crop breeding strategies. Although several examples of synergies and trade-offs between defence traits are known for plants that are under attack by multiple organisms, few studies have attempted to measure correlations of defensive strategies using specific single attackers. Such links are hard to detect in natural populations because they are inherently confounded by the evolutionary history of different ecotypes. We therefore used a range of 20 maize inbred lines with considerable differences in resistance traits to determine if correlations exist between leaf and root resistance against pathogens and insects. Aboveground resistance against insects was positively correlated with the plant's capacity to produce volatiles in response to insect attack. Resistance to herbivores and resistance to a pathogen, on the other hand, were negatively correlated. Our results also give first insights into the intraspecific variability of root volatiles release in maize and its positive correlation with leaf volatile production. We show that the breeding history of the different genotypes (dent versus flint) has influenced several defensive parameters. Taken together, our study demonstrates the importance of genetically determined synergies and trade-offs for plant resistance against insects and pathogens.

Minimally invasive approach to lumbosacral decompression in a cadaveric canine model

AIMS To determine efficacy of a minimally invasive (MI) surgical approach using a human MI lumbar retractor for canine lumbosacral dorsal laminectomy and partial discectomy and to compare this technique to the standard open surgical (OS) approach. METHODS Lumbosacral dorsal laminectomy and partial discectomy was performed on 16 large-breed canine cadavers using either a standard OS (n=8) or MI (n=8) approach. Skin and fascial incision length, procedure time, and intraoperative complications were recorded. Postoperatively specimens were evaluated for laminectomy and discectomy dimensions, and visible damage to the cauda equina and exiting nerve roots. RESULTS Median length of skin and fascial incisions in the OS group were longer than in the MI group (p<0.001). Median laminectomy length was similar between both approaches (p=0.234) but width was greater for the MI than OS approach (p=0.002). Both approaches achieved similar partial discectomy width (p=0.279). Overall surgical time was longer for MI approaches compared to OS, with a median of 18.5 (min 15.5, max 21.8) minutes for MI compared to 14.6 (min 13.1, max 16.9) minutes for OS (p=0.001). CONCLUSIONS The MI approach reduced incision lengths while retaining comparable laminectomy and discectomy dimensions. For this in vitro model the MI approach required more time to complete, but this difference may not be relevant in clinical cases. CLINICAL RELEVANCE Dogs undergoing lumbosacral dorsal laminectomy are commonly large-breed dogs. The traditional open approach requires a large skin incision and soft tissue dissection, especially in overweight animals. A MI approach accomplishing the same surgical result while minimising soft tissue trauma could reduce post-operative pain and recovery time, and may lower wound-related complications. Clinical studies are needed to confirm postoperative benefit and assess operating times in vivo.

Geochemistry of samples from Valtournenche, Western Italian Alps

Slices of polycyclic metasediments (marbles and meta-cherts) are tectonically amalgamated with the polydeformed basement of the Dent Blanche tectonic system along a major Alpine shear zone in the Western Alps (Becca di Salé area, Valtournenche Valley). A combination of techniques (structural analysis at various scales, metamorphic petrology, geochronology and trace element geochemistry) was applied to determine the age and composition of accessory phases (titanite, allanite and zircon) and their relation to major minerals. The results are used to reconstruct the polyphase structural and metamorphic history, comprising both pre-Alpine and Alpine cycles. The pre-Alpine evolution is associated with low-pressure high-temperature metamorphism related to Permo-Triassic lithospheric thinning. In meta-cherts, microtextural relations indicate coeval growth of allanite and garnet during this stage, at ~ 300 Ma. Textures of zircon also indicate crystallization at HT conditions; ages scatter from 263-294 Ma, with a major cluster of data at ~ 276 Ma. In impure marble, U-Pb analyses of titanite domains (with variable Al and F contents) yield apparent 206Pb/238U dates range from Permian to Jurassic. Chemical and isotopic data suggest that titanite formed at Permian times and was then affected by (extension-related?) fluid circulation during the Triassic and Jurassic, which redistributed major elements (Al and F) and partially opened the U-Pb system. The Alpine cycle lead to early blueschist facies assemblages, which were partly overprinted under greenschist facies conditions. The strong Alpine compressional overprint disrupted the pre-Alpine structural imprint and/or reactivated earlier structures. The pre-Alpine metamorphic record, preserved in these slices of metasediments, reflects the onset of the Permo-Triassic lithospheric extension to Jurassic rifting.

The Lagrangian description of aperiodic flows: a case study of the Kuroshio Current

This article reviews several recently developed Lagrangian tools and shows how their com- bined use succeeds in obtaining a detailed description of purely advective transport events in general aperiodic flows. In particular, because of the climate impact of ocean transport processes, we illustrate a 2D application on altimeter data sets over the area of the Kuroshio Current, although the proposed techniques are general and applicable to arbitrary time depen- dent aperiodic flows. The first challenge for describing transport in aperiodical time dependent flows is obtaining a representation of the phase portrait where the most relevant dynamical features may be identified. This representation is accomplished by using global Lagrangian descriptors that when applied for instance to the altimeter data sets retrieve over the ocean surface a phase portrait where the geometry of interconnected dynamical systems is visible. The phase portrait picture is essential because it evinces which transport routes are acting on the whole flow. Once these routes are roughly recognised it is possible to complete a detailed description by the direct computation of the finite time stable and unstable manifolds of special hyperbolic trajectories that act as organising centres of the flow.

Detection and pattern recognition applied to leaves and chromosomes

The Project you are about to see it is based on the technologies used on object detection and recognition, especially on leaves and chromosomes. To do so, this document contains the typical parts of a scientific paper, as it is what it is. It is composed by an Abstract, an Introduction, points that have to do with the investigation area, future work, conclusions and references used for the elaboration of the document. The Abstract talks about what are we going to find in this paper, which is technologies employed on pattern detection and recognition for leaves and chromosomes and the jobs that are already made for cataloguing these objects. In the introduction detection and recognition meanings are explained. This is necessary as many papers get confused with these terms, specially the ones talking about chromosomes. Detecting an object is gathering the parts of the image that are useful and eliminating the useless parts. Summarizing, detection would be recognizing the objects borders. When talking about recognition, we are talking about the computers or the machines process, which says what kind of object we are handling. Afterwards we face a compilation of the most used technologies in object detection in general. There are two main groups on this category: Based on derivatives of images and based on ASIFT points. The ones that are based on derivatives of images have in common that convolving them with a previously created matrix does the treatment of them. This is done for detecting borders on the images, which are changes on the intensity of the pixels. Within these technologies we face two groups: Gradian based, which search for maximums and minimums on the pixels intensity as they only use the first derivative. The Laplacian based methods search for zeros on the pixels intensity as they use the second derivative. Depending on the level of details that we want to use on the final result, we will choose one option or the other, because, as its logic, if we used Gradian based methods, the computer will consume less resources and less time as there are less operations, but the quality will be worse. On the other hand, if we use the Laplacian based methods we will need more time and resources as they require more operations, but we will have a much better quality result. After explaining all the derivative based methods, we take a look on the different algorithms that are available for both groups. The other big group of technologies for object recognition is the one based on ASIFT points, which are based on 6 image parameters and compare them with another image taking under consideration these parameters. These methods disadvantage, for our future purposes, is that it is only valid for one single object. So if we are going to recognize two different leaves, even though if they refer to the same specie, we are not going to be able to recognize them with this method. It is important to mention these types of technologies as we are talking about recognition methods in general. At the end of the chapter we can see a comparison with pros and cons of all technologies that are employed. Firstly comparing them separately and then comparing them all together, based on our purposes. Recognition techniques, which are the next chapter, are not really vast as, even though there are general steps for doing object recognition, every single object that has to be recognized has its own method as the are different. This is why there is not a general method that we can specify on this chapter. We now move on into leaf detection techniques on computers. Now we will use the technique explained above based on the image derivatives. Next step will be to turn the leaf into several parameters. Depending on the document that you are referring to, there will be more or less parameters. Some papers recommend to divide the leaf into 3 main features (shape, dent and vein] and doing mathematical operations with them we can get up to 16 secondary features. Next proposition is dividing the leaf into 5 main features (Diameter, physiological length, physiological width, area and perimeter] and from those, extract 12 secondary features. This second alternative is the most used so it is the one that is going to be the reference. Following in to leaf recognition, we are based on a paper that provides a source code that, clicking on both leaf ends, it automatically tells to which specie belongs the leaf that we are trying to recognize. To do so, it only requires having a database. On the tests that have been made by the document, they assure us a 90.312% of accuracy over 320 total tests (32 plants on the database and 10 tests per specie]. Next chapter talks about chromosome detection, where we shall pass the metaphasis plate, where the chromosomes are disorganized, into the karyotype plate, which is the usual view of the 23 chromosomes ordered by number. There are two types of techniques to do this step: the skeletonization process and swiping angles. Skeletonization progress consists on suppressing the inside pixels of the chromosome to just stay with the silhouette. This method is really similar to the ones based on the derivatives of the image but the difference is that it doesnt detect the borders but the interior of the chromosome. Second technique consists of swiping angles from the beginning of the chromosome and, taking under consideration, that on a single chromosome we cannot have more than an X angle, it detects the various regions of the chromosomes. Once the karyotype plate is defined, we continue with chromosome recognition. To do so, there is a technique based on the banding that chromosomes have (grey scale bands] that make them unique. The program then detects the longitudinal axis of the chromosome and reconstructs the band profiles. Then the computer is able to recognize this chromosome. Concerning the future work, we generally have to independent techniques that dont reunite detection and recognition, so our main focus would be to prepare a program that gathers both techniques. On the leaf matter we have seen that, detection and recognition, have a link as both share the option of dividing the leaf into 5 main features. The work that would have to be done is to create an algorithm that linked both methods, as in the program, which recognizes leaves, it has to be clicked both leaf ends so it is not an automatic algorithm. On the chromosome side, we should create an algorithm that searches for the beginning of the chromosome and then start to swipe angles, to later give the parameters to the program that searches for the band profiles. Finally, on the summary, we explain why this type of investigation is needed, and that is because with global warming, lots of species (animals and plants] are beginning to extinguish. That is the reason why a big database, which gathers all the possible species, is needed. For recognizing animal species, we just only have to have the 23 chromosomes. While recognizing a plant, there are several ways of doing it, but the easiest way to input a computer is to scan the leaf of the plant. RESUMEN. El proyecto que se puede ver a continuación trata sobre las tecnologías empleadas en la detección y reconocimiento de objetos, especialmente de hojas y cromosomas. Para ello, este documento contiene las partes típicas de un paper de investigación, puesto que es de lo que se trata. Así, estará compuesto de Abstract, Introducción, diversos puntos que tengan que ver con el área a investigar, trabajo futuro, conclusiones y biografía utilizada para la realización del documento. Así, el Abstract nos cuenta qué vamos a poder encontrar en este paper, que no es ni más ni menos que las tecnologías empleadas en el reconocimiento y detección de patrones en hojas y cromosomas y qué trabajos hay existentes para catalogar a estos objetos. En la introducción se explican los conceptos de qué es la detección y qué es el reconocimiento. Esto es necesario ya que muchos papers científicos, especialmente los que hablan de cromosomas, confunden estos dos términos que no podían ser más sencillos. Por un lado tendríamos la detección del objeto, que sería simplemente coger las partes que nos interesasen de la imagen y eliminar aquellas partes que no nos fueran útiles para un futuro. Resumiendo, sería reconocer los bordes del objeto de estudio. Cuando hablamos de reconocimiento, estamos refiriéndonos al proceso que tiene el ordenador, o la máquina, para decir qué clase de objeto estamos tratando. Seguidamente nos encontramos con un recopilatorio de las tecnologías más utilizadas para la detección de objetos, en general. Aquí nos encontraríamos con dos grandes grupos de tecnologías: Las basadas en las derivadas de imágenes y las basadas en los puntos ASIFT. El grupo de tecnologías basadas en derivadas de imágenes tienen en común que hay que tratar a las imágenes mediante una convolución con una matriz creada previamente. Esto se hace para detectar bordes en las imágenes que son básicamente cambios en la intensidad de los píxeles. Dentro de estas tecnologías nos encontramos con dos grupos: Los basados en gradientes, los cuales buscan máximos y mínimos de intensidad en la imagen puesto que sólo utilizan la primera derivada; y los Laplacianos, los cuales buscan ceros en la intensidad de los píxeles puesto que estos utilizan la segunda derivada de la imagen. Dependiendo del nivel de detalles que queramos utilizar en el resultado final nos decantaremos por un método u otro puesto que, como es lógico, si utilizamos los basados en el gradiente habrá menos operaciones por lo que consumirá más tiempo y recursos pero por la contra tendremos menos calidad de imagen. Y al revés pasa con los Laplacianos, puesto que necesitan más operaciones y recursos pero tendrán un resultado final con mejor calidad. Después de explicar los tipos de operadores que hay, se hace un recorrido explicando los distintos tipos de algoritmos que hay en cada uno de los grupos. El otro gran grupo de tecnologías para el reconocimiento de objetos son los basados en puntos ASIFT, los cuales se basan en 6 parámetros de la imagen y la comparan con otra imagen teniendo en cuenta dichos parámetros. La desventaja de este método, para nuestros propósitos futuros, es que sólo es valido para un objeto en concreto. Por lo que si vamos a reconocer dos hojas diferentes, aunque sean de la misma especie, no vamos a poder reconocerlas mediante este método. Aún así es importante explicar este tipo de tecnologías puesto que estamos hablando de técnicas de reconocimiento en general. Al final del capítulo podremos ver una comparación con los pros y las contras de todas las tecnologías empleadas. Primeramente comparándolas de forma separada y, finalmente, compararemos todos los métodos existentes en base a nuestros propósitos. Las técnicas de reconocimiento, el siguiente apartado, no es muy extenso puesto que, aunque haya pasos generales para el reconocimiento de objetos, cada objeto a reconocer es distinto por lo que no hay un método específico que se pueda generalizar. Pasamos ahora a las técnicas de detección de hojas mediante ordenador. Aquí usaremos la técnica explicada previamente explicada basada en las derivadas de las imágenes. La continuación de este paso sería diseccionar la hoja en diversos parámetros. Dependiendo de la fuente a la que se consulte pueden haber más o menos parámetros. Unos documentos aconsejan dividir la morfología de la hoja en 3 parámetros principales (Forma, Dentina y ramificación] y derivando de dichos parámetros convertirlos a 16 parámetros secundarios. La otra propuesta es dividir la morfología de la hoja en 5 parámetros principales (Diámetro, longitud fisiológica, anchura fisiológica, área y perímetro] y de ahí extraer 12 parámetros secundarios. Esta segunda propuesta es la más utilizada de todas por lo que es la que se utilizará. Pasamos al reconocimiento de hojas, en la cual nos hemos basado en un documento que provee un código fuente que cucando en los dos extremos de la hoja automáticamente nos dice a qué especie pertenece la hoja que estamos intentando reconocer. Para ello sólo hay que formar una base de datos. En los test realizados por el citado documento, nos aseguran que tiene un índice de acierto del 90.312% en 320 test en total (32 plantas insertadas en la base de datos por 10 test que se han realizado por cada una de las especies]. El siguiente apartado trata de la detección de cromosomas, en el cual se debe de pasar de la célula metafásica, donde los cromosomas están desorganizados, al cariotipo, que es como solemos ver los 23 cromosomas de forma ordenada. Hay dos tipos de técnicas para realizar este paso: Por el proceso de esquelotonización y barriendo ángulos. El proceso de esqueletonización consiste en eliminar los píxeles del interior del cromosoma para quedarse con su silueta; Este proceso es similar a los métodos de derivación de los píxeles pero se diferencia en que no detecta bordes si no que detecta el interior de los cromosomas. La segunda técnica consiste en ir barriendo ángulos desde el principio del cromosoma y teniendo en cuenta que un cromosoma no puede doblarse más de X grados detecta las diversas regiones de los cromosomas. Una vez tengamos el cariotipo, se continua con el reconocimiento de cromosomas. Para ello existe una técnica basada en las bandas de blancos y negros que tienen los cromosomas y que son las que los hacen únicos. Para ello el programa detecta los ejes longitudinales del cromosoma y reconstruye los perfiles de las bandas que posee el cromosoma y que lo identifican como único. En cuanto al trabajo que se podría desempeñar en el futuro, tenemos por lo general dos técnicas independientes que no unen la detección con el reconocimiento por lo que se habría de preparar un programa que uniese estas dos técnicas. Respecto a las hojas hemos visto que ambos métodos, detección y reconocimiento, están vinculados debido a que ambos comparten la opinión de dividir las hojas en 5 parámetros principales. El trabajo que habría que realizar sería el de crear un algoritmo que conectase a ambos ya que en el programa de reconocimiento se debe clicar a los dos extremos de la hoja por lo que no es una tarea automática. En cuanto a los cromosomas, se debería de crear un algoritmo que busque el inicio del cromosoma y entonces empiece a barrer ángulos para después poder dárselo al programa que busca los perfiles de bandas de los cromosomas. Finalmente, en el resumen se explica el por qué hace falta este tipo de investigación, esto es que con el calentamiento global, muchas de las especies (tanto animales como plantas] se están empezando a extinguir. Es por ello que se necesitará una base de datos que contemple todas las posibles especies tanto del reino animal como del reino vegetal. Para reconocer a una especie animal, simplemente bastará con tener sus 23 cromosomas; mientras que para reconocer a una especie vegetal, existen diversas formas. Aunque la más sencilla de todas es contar con la hoja de la especie puesto que es el elemento más fácil de escanear e introducir en el ordenador.

Determinación de la higroscopicidad y comportamiento termodinámico de la madera juvenil y madura a través de sus isotermas de sorción

La respuesta higroscópica de la madera varía a lo largo de la dirección radial del árbol. El aumento de corta de ejemplares jóvenes y el uso de troncos de pequeños diámetros en la industria de los productos forestales, hacen preciso estudiar el comportamiento higroscópico tanto de la madera juvenil como de la madura. Su determinación proporciona información para comprender los mecanismos de sorción. Asimismo, la obtención de las propiedades termodinámicas de ambos tipos de madera facilita la modelización de procesos industriales como el secado o el encolado. En el presente trabajo, se ha comparado el comportamiento higroscópico y las propiedades termodinámicas de la madera juvenil y madura de Abies pinsapo Boissier, Abies alba Mill., Pinus canariensis C. Sm. ex DC., Pinus nigra Arnold, Pinus uncinata Mill. ex Mirb. y Pinus pinea L. Para este propósito se han utilizado las isotermas de sorción obtenidas mediante el método tradicional de sales saturadas descrito por COST Action E8 a 15, 35 y 50ºC en Abies pinsapo, Abies alba, Pinus nigra, Pinus uncinata y Pinus pinea, y a 35 y 50ºC en Pinus canariensis. Igualmente, se ha empleado el equipo dynamic vapor sorption (DVS) en la obtención de las isotermas de Pinus pinea a 35 y 50ºC. El ajuste de las curvas se ha realizado mediante el modelo Guggenheim, Anderson y de Boer-Dent (GAB), cumpliendo todas las muestras los criterios de aceptación establecidos. En el estudio de las isotermas se ha calculado el coeficiente y área de histéresis entre el proceso de adsorción y desorción para cada una de las muestras. Con el fin de comprender el comportamiento higroscópico experimentado por cada madera se ha determinado la composición química, espectros de infrarrojos (FTIR) y difractogramas de rayos X de cada una. Los parámetros termodinámicos - calor isostérico neto y total heat of wetting - se han obtenido a partir de las isotermas de sorción mediante el método de integración de la ecuación de Clausius-Clapeyron. Finalmente, se han comparado los datos obtenidos con el método tradicional de sales saturadas y con dynamic vapor sorption con el propósito de conocer la existencia de similitudes entre ambas metodologías. Los resultados mostraron que los puntos de equilibrio son, en la mayor parte de los casos, superiores en la madera madura frente a la juvenil, y por tanto las isotermas de la madera madura se encuentran siempre por encima de las de la juvenil, debido principalmente a la composición química. Respecto a las propiedades termodinámicas, se ha determinado que la energía involucrada en los procesos de sorción es superior en la madera madura que en la madera juvenil, siendo mayor en el proceso de desorción frente al de adsorción. En la comparación de las metodologías de sales saturadas y dynamic vapor sorption no se han detectado casi diferencias significativas en el proceso de adsorción, mientras que sí se han obtenido en el de desorción. ABSTRACT The hygroscopic response of wood varies throughout the radial direction of the tree. The longer cut of young trees and the use of small-diameter trunks in the forest product industry make it necessary to study the hygroscopic behaviour of both juvenile and mature wood. Determining this behaviour in both types of wood provides information for understanding the sorption mechanisms. Similarly, obtaining the thermodynamic properties of juvenile and mature wood facilitates modelling of industrial processes such as drying and bonding. In this study a comparison was made of the hygroscopic behaviour and thermodynamic properties of juvenile and mature wood of Abies pinsapo Boissier, Abies alba Mill., Pinus canariensis C. Sm. ex DC., Pinus nigra Arnold, Pinus uncinata Mill. ex Mirb. and Pinus pinea L. This was done by obtaining the sorption isotherms using the traditional saturated salt method described by COST Action E8 at 15, 35 and 50ºC in Abies pinsapo, Abies alba, Pinus nigra, Pinus uncinata and Pinus pinea, and at 35 and 50ºC in Pinus canariensis. In addition, dynamic vapour sorption (DVS) was used to obtain the isotherms of Pinus pinea at 35 and 50ºC. The curves were fitted using the Guggenheim, Anderson and de Boer- Dent (GAB) model and all samples met the established acceptance criteria. In the study of the isotherms, the hysteresis coefficient and area of the hysteresis loop between adsorption and desorption were calculated for each sample. To understand the hygroscopic behaviour of juvenile and mature wood, the chemical composition, infrared spectra (FTIR) and X-ray diffractograms of each type of wood were determined. The thermodynamic parameters - net isosteric heat and total heat of wetting - were obtained from the sorption isotherms by applying the integration method of the Clausius-Clapeyron equation. The data obtained using the traditional saturated salt method and with dynamic vapour sorption were compared to determine the similarities between the two methods. The results showed that the equilibrium points are greater in the mature wood than in the juvenile wood in most cases, and therefore the mature wood isotherms are always above the juvenile wood isotherms, mainly because of the chemical composition. As regards the thermodynamic properties, it was determined that the energy involved in the sorption processes is greater in the mature wood than in the juvenile wood, and is greater in the desorption process than in the adsorption process. On comparing the saturated salt and dynamic vapour sorption methods, almost no significant differences were detected in the adsorption process, but significant differences were obtained in the desorption process.

T helper type 2 inflammatory disease in the absence of interleukin 4 and transcription factor STAT6

An important signaling pathway for the differentiation of T helper type 2 (TH2) cells from uncommitted CD4 T cell precursors is activation of the STAT6 transcription factor by interleukin 4 (IL-4). The protooncogene BCL-6 is also involved in TH2 differentiation, as BCL-6 −/− mice develop an inflammation of the heart and lungs associated with an overproduction of TH2 cells. Surprisingly, IL-4 −/− BCL-6 −/− and STAT6 −/− BCL-6 −/− double-mutant mice developed the same TH2-type inflammation of the heart and lungs as is characteristic of BCL-6 −/− mice. Furthermore, a TH2 cytokine response developed in STAT6 −/− BCL-6 −/− and IL-4 −/− BCL-6 −/− mice after immunization with a conventional antigen in adjuvant. In contrast to these in vivo findings, STAT6 was required for the in vitro differentiation of BCL-6 −/− T cells into TH2 cells. BCL-6, a transcriptional repressor that can bind to the same DNA binding motifs as STAT transcription factors, seems to regulate TH2 responses in vivo by a pathway independent of IL-4 and STAT6.

The Ras/Rac1/Cdc42/SEK/JNK/c-Jun Cascade Is a Key Pathway by Which Agonists Stimulate DNA Synthesis in Primary Cultures of Rat Hepatocytes

The ability of signaling via the JNK (c-Jun NH2-terminal kinase)/stress-activated protein kinase cascade to stimulate or inhibit DNA synthesis in primary cultures of adult rat hepatocytes was examined. Treatment of hepatocytes with media containing hyperosmotic glucose (75 mM final), tumor necrosis factor α (TNFα, 1 ng/ml final), and hepatocyte growth factor (HGF, 1 ng/ml final) caused activation of JNK1. Glucose, TNFα, or HGF treatments increased phosphorylation of c-Jun at serine 63 in the transactivation domain and stimulated hepatocyte DNA synthesis. Infection of hepatocytes with poly-l-lysine–coated adenoviruses coupled to constructs to express either dominant negatives Ras N17, Rac1 N17, Cdc42 N17, SEK1−, or JNK1− blunted the abilities of glucose, TNFα, or HGF to increase JNK1 activity, to increase phosphorylation of c-Jun at serine 63, and to stimulate DNA synthesis. Furthermore, infection of hepatocytes by a recombinant adenovirus expressing a dominant-negative c-Jun mutant (TAM67) also blunted the abilities of glucose, TNFα, and HGF to stimulate DNA synthesis. These data demonstrate that multiple agonists stimulate DNA synthesis in primary cultures of hepatocytes via a Ras/Rac1/Cdc42/SEK/JNK/c-Jun pathway. Glucose and HGF treatments reduced glycogen synthase kinase 3 (GSK3) activity and increased c-Jun DNA binding. Co-infection of hepatocytes with recombinant adenoviruses to express dominant- negative forms of PI3 kinase (p110α/p110γ) increased basal GSK3 activity, blocked the abilities of glucose and HGF treatments to inhibit GSK3 activity, and reduced basal c-Jun DNA binding. However, expression of dominant-negative PI3 kinase (p110α/p110γ) neither significantly blunted the abilities of glucose and HGF treatments to increase c-Jun DNA binding, nor inhibited the ability of these agonists to stimulate DNA synthesis. These data suggest that signaling by the JNK/stress-activated protein kinase cascade, rather than by the PI3 kinase cascade, plays the pivotal role in the ability of agonists to stimulate DNA synthesis in primary cultures of rat hepatocytes.

Radiation-induced Release of Transforming Growth Factor α Activates the Epidermal Growth Factor Receptor and Mitogen-activated Protein Kinase Pathway in Carcinoma Cells, Leading to Increased Proliferation and Protection from Radiation-induced Cell Death

Exposure of A431 squamous and MDA-MB-231 mammary carcinoma cells to ionizing radiation has been associated with short transient increases in epidermal growth factor receptor (EGFR) tyrosine phosphorylation and activation of the mitogen-activated protein kinase (MAPK) and c-Jun NH2-terminal kinase (JNK) pathways. Irradiation (2 Gy) of A431 and MDA-MB-231 cells caused immediate primary activations (0–10 min) of the EGFR and the MAPK and JNK pathways, which were surprisingly followed by later prolonged secondary activations (90–240 min). Primary and secondary activation of the EGFR was abolished by molecular inhibition of EGFR function. The primary and secondary activation of the MAPK pathway was abolished by molecular inhibition of either EGFR or Ras function. In contrast, molecular inhibition of EGFR function abolished the secondary but not the primary activation of the JNK pathway. Inhibition of tumor necrosis factor α receptor function by use of neutralizing monoclonal antibodies blunted primary activation of the JNK pathway. Addition of a neutralizing monoclonal antibody versus transforming growth factor α (TGFα) had no effect on the primary activation of either the EGFR or the MAPK and JNK pathways after irradiation but abolished the secondary activation of EGFR, MAPK, and JNK. Irradiation of cells increased pro-TGFα cleavage 120–180 min after exposure. In agreement with radiation-induced release of a soluble factor, activation of the EGFR and the MAPK and JNK pathways could be induced in nonirradiated cells by the transfer of media from irradiated cells 120 min after irradiation. The ability of the transferred media to cause MAPK and JNK activation was blocked when media were incubated with a neutralizing antibody to TGFα. Thus radiation causes primary and secondary activation of the EGFR and the MAPK and JNK pathways in autocrine-regulated carcinoma cells. Secondary activation of the EGFR and the MAPK and JNK pathways is dependent on radiation-induced cleavage and autocrine action of TGFα. Neutralization of TGFα function by an anti-TGFα antibody or inhibition of MAPK function by MEK1/2 inhibitors (PD98059 and U0126) radiosensitized A431 and MDA-MB-231 cells after irradiation in apoptosis, 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT), and clonogenic assays. These data demonstrate that disruption of the TGFα–EGFR–MAPK signaling module represents a strategy to decrease carcinoma cell growth and survival after irradiation.

Roles for Basal and Stimulated p21Cip-1/WAF1/MDA6 Expression and Mitogen-activated Protein Kinase Signaling in Radiation-induced Cell Cycle Checkpoint Control in Carcinoma Cells

We investigated the role of the cdk inhibitor protein p21Cip-1/WAF1/MDA6 (p21) in the ability of MAPK pathway inhibition to enhance radiation-induced apoptosis in A431 squamous carcinoma cells. In carcinoma cells, ionizing radiation (2 Gy) caused both primary (0–10 min) and secondary (90–240 min) activations of the MAPK pathway. Radiation induced p21 protein expression in A431 cells within 6 h via secondary activation of the MAPK pathway. Within 6 h, radiation weakly enhanced the proportion of cells in G1 that were p21 and MAPK dependent, whereas the elevation of cells present in G2/M at this time was independent of either p21 expression or MAPK inhibition. Inhibition of the MAPK pathway increased the proportion of irradiated cells in G2/M phase 24–48 h after irradiation and enhanced radiation-induced apoptosis. This correlated with elevated Cdc2 tyrosine 15 phosphorylation, decreased Cdc2 activity, and decreased Cdc25C protein levels. Caffeine treatment or removal of MEK1/2 inhibitors from cells 6 h after irradiation reduced the proportion of cells present in G2/M phase at 24 h and abolished the ability of MAPK inhibition to potentiate radiation-induced apoptosis. These data argue that MAPK signaling plays an important role in the progression/release of cells through G2/M phase after radiation exposure and that an impairment of this progression/release enhances radiation-induced apoptosis. Surprisingly, the ability of irradiation/MAPK inhibition to increase the proportion of cells in G2/M at 24 h was found to be dependent on basal p21 expression. Transient inhibition of basal p21 expression increased the control level of apoptosis as well as the abilities of both radiation and MEK1/2 inhibitors to cause apoptosis. In addition, loss of basal p21 expression significantly reduced the capacity of MAPK inhibition to potentiate radiation-induced apoptosis. Collectively, our data argue that MAPK signaling and p21 can regulate cell cycle checkpoint control in carcinoma cells at the G1/S transition shortly after exposure to radiation. In contrast, inhibition of MAPK increases the proportion of irradiated cells in G2/M, and basal expression of p21 is required to maintain this effect. Our data suggest that basal and radiation-stimulated p21 may play different roles in regulating cell cycle progression that affect cell survival after radiation exposure.

PEA3 sites within the progression elevated gene-3 (PEG-3) promoter and mitogen-activated protein kinase contribute to differential PEG-3 expression in Ha-ras and v-raf oncogene transformed rat embryo cells

Transformation of normal cloned rat embryo fibroblast (CREF) cells with cellular oncogenes results in acquisition of anchorage-independent growth and oncogenic potential in nude mice. These cellular changes correlate with an induction in the expression of a cancer progression-promoting gene, progression elevated gene-3 (PEG-3). To define the mechanism of activation of PEG-3 as a function of transformation by the Ha-ras and v-raf oncogenes, evaluations of the signaling and transcriptional regulation of the ~2.0 kb promoter region of the PEG-3 gene, PEG-Prom, was undertaken. The full-length and various mutated regions of the PEG-Prom were linked to a luciferase reporter construct and tested for promoter activity in CREF and oncogene-transformed CREF cells. An analysis was also performed using CREF cells doubly transformed with Ha-ras and the Ha-ras specific suppressor gene Krev-1, which inhibits the transformed phenotype in vitro. These assays document an association between expression of the transcription regulator PEA3 and PEG-3. The levels of PEA3 and PEG-3 RNA and proteins are elevated in the oncogenically transformed CREF cells, and reduced in transformation and tumorigenic suppressed Ha-ras/Krev-1 doubly transformed CREF cells. Enhanced tumorigenic behavior, PEG-3 promoter function and PEG-3 expression in Ha-ras transformed cells were all dependent upon increased activity within the mitogen-activated protein kinase (MAPK) pathway. Electrophoretic mobility shift assays and DNase I footprinting experiments indicate that PEA3 binds to sites within the PEG-Prom in transformed rodent cells in an area adjacent to the TATA box in a MAPK-dependent fashion. These findings demonstrate an association between Ha-ras and v-raf transformation of CREF cells with elevated PEA3 and PEG-3 expression, and they implicate MAPK signaling via PEA3 as a signaling cascade involved in activation of the PEG-Prom.

Oral soft tissue biopsies in Oporto, Portugal: An eight year retrospective analysis

The diseases that affect the oral cavity are wide and diverse, comprising a broad spectrum of either benign or malignant lesions. However, few histological-based studies were performed for the evaluation of oral cavity lesions, and very few directed to oral soft tissue pathology. The aim of this study was to carry out pioneering research, within a Portuguese population, to determine the frequency and characteristics of oral malignancies, potential malignant disorders, and soft benign tissues pathologies submitted for biopsy in a north Portugal (Oporto) hospital population.

Unemployment is the scourge, not youth unemployment per se. The misguided policy preoccupation with youth. CEPS Policy Brief No. 294, 26 June 2013

This Policy Brief argues that too much effort and political capital is being spent by the Commission and member states on being seen to be doing something quickly about youth unemployment when, in fact, the structural measures proposed will only have long-term effects. Expectations of immediate relief are running well above what can be delivered, especially at the EU level. Given the macroeconomic situation, no policy option will deliver a significant dent in either youth unemployment or unemployment in general. The EU policies on the table that are supposed to have an immediate effect, such as increased lending from the European Investment Bank to SMEs for the hiring of young people, will only have a very marginal impact on youth unemployment. Moreover, this impact will come mostly to the detriment of older unemployed persons excluded from such a scheme. Given the perceived need to ‘be seen to be doing something’, we fear that policies subsidising young workers de facto at the expense of older workers or, even worse, policies that subsidise older workers for not taking young people’s jobs, will proliferate. In fact, it is not at all clear that young people suffer more from being unemployed than older people, or even disproportionately more than older unemployed individuals. In particular, it is not clear that the much-publicised notion of a ‘lost generation’ with permanent ‘scars’ is relevant only to the young generation. The paper ends by highlighting the much-neglected policy option of encouraging labour mobility within the internal market. Although the Commission is ‘upgrading and modernising’ its tools, much more could be done in this area – to the benefit of the individuals concerned, the member states, and European integration in general.

Metabolomics analysis in rats with thiamine deficiency identifies key metabolites in vulnerable brain regions and suggests neural stem progenitor cells play a role in ameliorating metabolic dysfunction

La documentation scientifique fait état de la présence, chez l’adulte, de cellules souches et progénitrices neurales (CSPN) endogènes dans les zones sous-ventriculaire et sous-granulaire du cerveau ainsi que dans le gyrus denté de l’hippocampe. De plus, un postulat selon lequel il serait également possible de retrouver ce type de cellules dans la moelle épinière et le néocortex des mammifères adultes a été énoncé. L’encéphalopathie de Wernicke, un trouble neurologique grave toutefois réversible qui entraîne un dysfonctionnement, voire une défaillance du cerveau, est causée principalement par une carence importante en thiamine (CT). Des observations récentes laissent envisager que les facteurs en cause dans la prolifération et la différenciation des CSPN pourraient également jouer un rôle important lors d’un épisode de CT. L’hypothèse, selon laquelle l’identification de nouveaux métabolites entrant dans le mécanisme ou la séquence de réactions se soldant en une CT pourraient en faciliter la compréhension, a été émise au moyen d'une démarche en cours permettant d’établir le profil des modifications métaboliques qui surviennent en de telles situations. Cette approche a été utilisée pour constater les changements métaboliques survenus au niveau du foyer cérébral dans un modèle de rats déficients en thiamine (rats DT), particulièrement au niveau du thalamus et du colliculus inférieur (CI). La greffe de CSPN a quant à elle été envisagée afin d’apporter de nouvelles informations sur la participation des CSPN lors d’un épisode de CT et de déterminer les bénéfices thérapeutiques potentiels offerts par cette intervention. Les sujets de l’étude étaient répartis en quatre groupes expérimentaux : un premier groupe constitué de rats dont la CT était induite par la pyrithiamine (rats DTiP), un deuxième groupe constitué de rats-contrôles nourris ensemble (« pair-fed control rats » ou rats PFC) ainsi que deux groupes de rats ayant subi une greffe de CSPN, soit un groupe de rats DTiP greffés et un dernier groupe constitué de rats-contrôles (rats PFC) greffés. Les échantillons de foyers cérébraux (thalamus et CI) des quatre groupes de rats ont été prélevés et soumis à des analyses métabolomiques non ciblées ainsi qu’à une analyse visuelle par microscopie à balayage électronique (SEM). Une variété de métabolites-clés a été observée chez les groupes de rats déficients en thiamine (rats DTiP) en plus de plusieurs métabolites dont la documentation ne faisait pas mention. On a notamment constaté la présence d’acides biliaires, d’acide cynurénique et d’acide 1,9— diméthylurique dans le thalamus, alors que la présence de taurine et de carnosine a été observée dans le colliculus inférieur. L’étude a de plus démontré une possible implication des CSPN endogènes dans les foyers cérébraux du thalamus et du colliculus inférieur en identifiant les métabolites-clés ciblant les CSPN. Enfin, les analyses par SEM ont montré une amélioration notable des tissus à la suite de la greffe de CSPN. Ces constatations suggèrent que l’utilisation de CSPN pourrait s’avérer une avenue thérapeutique intéressante pour soulager la dégénérescence symptomatique liée à une grave carence en thiamine chez l’humain.