"The need obliged us" : culture as capacity during the Hurricane Stan emergency response. A case study from Tectitan, Huehuetenango, Guatemala

A major deficiency in disaster management plans is the assumption that pre-disaster civil-society does not have the capacity to respond effectively during crises. Following from this assumption a dominant emergency management strategy is to replace weak civil-society organizations with specialized disaster organizations that are often either military or Para-military and seek to centralize decision-making. Many criticisms have been made of this approach, but few specifically addresses disasters in the developing world. Disasters in the developing world present unique problems not seen in the developed world because they often occur in the context of compromised governments, and marginalized populations. In this context it is often community members themselves who possess the greatest capacity to respond to disasters. This paper focuses on the capacity of community groups to respond to disaster in a small town in rural Guatemala. Key informant interviews and ethnographic observations are used to reconstruct the community response to the disaster instigated by Hurricane Stan (2005) in the municipality of Tectitán in the Huehuetenango department. The interviews were analyzed using techniques adapted from grounded theory to construct a narrative of the events, and identify themes in the community’s disaster behavior. These themes are used to critique the emergency management plans advocated by the Guatemalan National Coordination for the Reduction of Disasters (CONRED). This paper argues that CONRED uncritically adopts emergency management strategies that do not account for the local realities in communities throughout Guatemala. The response in Tectitán was characterized by the formation of new organizations, whose actions and leadership structure were derived from “normal” or routine life. It was found that pre-existing social networks were resilient and easily re-oriented meet the novel needs of a crisis. New or emergent groups that formed during the disaster utilized social capital accrued by routine collective behavior, and employed organizational strategies derived from “normal” community relations. Based on the effectiveness of this response CONRED could improve its emergency planning on the local-level by utilizing the pre-existing community organizations rather than insisting that new disaster-specific organizations be formed.

The p53 tumor suppressor pathway in culture and {\it in vivo\/}

p53 functions as a tumor suppressor through its ability to initiate either growth arrest or apoptosis in cells which have sustained DNA damage. p53 elicits these cellular phenotypes through its biochemical function as a transcriptional activator. By inducing the expression of a battery of target genes, p53 is able to prevent the propagation of cells with damaged DNA. However, the genes transcriptionally induced by p53 which have been identified to date do not fully explain p53 function. p53 has been demonstrated to activate genes involved in cell cycle inhibition, apoptosis and cell proliferation. The reasons for simultaneous activation of p53 targets with disparate, opposing functions are not clear, but may be due to the use of transformed cell lines in previous experiments. In the studies presented in this thesis, the pathway of p53 tumor suppression has been studied in detail in two systems chosen for their relevance to the natural cell environment. One utilizes a normal, unaltered cultured cell system; the other the whole mouse. In order to better understand the role of the known p53 targets in effecting p53 function in normal cells, early rat embryo fibroblasts were irradiated with ultraviolet light to induce DNA damage. It was discovered that p53 protein levels increased in response to irradiation. The known targets of p53, namely, $p21\sp{WAF1/CIP1},\ mdm2,\ cyclin\ G,$ and bax, were shown for the first time to have a differential temporal induction. The growth suppressor $p21\sp{WAF1/CIP1}$ was induced first, followed by cyclin G then mdm2, which is involved in proliferation through its inactivation of p53, and finally, the apoptosis promoter, bax. These findings indicated that p53 activates its target genes in a manner to allow maximum effectiveness of target function. The rat embryo fibroblasts were shown to undergo apoptosis 24 h after irradiation. Additionally, investigation of these cells for cell cycle alterations demonstrated a brief arrest in G1. In the second study, thymocytes from mice with wild type p53 were shown to undergo apoptosis and activate p53 target genes upon ionizing radiation treatment, while thymocytes from mice deficient in p53 could not. The p53 target genes mdm2 and fas were tested in vivo for their ability to mediate p53-regulated apoptosis, and were found dispensible for that cellular function. Therefore, the p53 targets identified to date do not fully explain the ability of p53 to function as a tumor suppressor. Potentially, functional redundancy between the known targets would account for the data seen in these experiments. Additionally, identification of additional target genes should add further understanding of the p53 pathway of tumor suppression. ^

The interweaving of popular generic modes and modernist techniques in Nostromo : A tale of the seaboard

Nostromo. A Tale of the Seaboard (1904), una de las novelas más experimentales del período modernista temprano, ha sido considerada la obra maestra de Joseph Conrad y su más grande lienzo. Un rasgo particular de la novela es que en ella coexisten la experimentación técnica del modernismo, la altamente difusa ficción del narrador y la situación de narración de historias y el paradigma narrativo de los subgéneros "degradados" de la novela de aventuras, el romance y el melodrama. Este estudio indaga las relaciones que se establecen entre dichos elementos, a todas luces anacrónicos o pertenecientes a espacios culturales diferentes, y se propone mostrar que estas discontinuidades no deben ser consideradas una falla sino índices de una tensión constitutiva de la novela.

The interweaving of popular generic modes and modernist techniques in Nostromo : A tale of the seaboard

Nostromo. A Tale of the Seaboard (1904), una de las novelas más experimentales del período modernista temprano, ha sido considerada la obra maestra de Joseph Conrad y su más grande lienzo. Un rasgo particular de la novela es que en ella coexisten la experimentación técnica del modernismo, la altamente difusa ficción del narrador y la situación de narración de historias y el paradigma narrativo de los subgéneros "degradados" de la novela de aventuras, el romance y el melodrama. Este estudio indaga las relaciones que se establecen entre dichos elementos, a todas luces anacrónicos o pertenecientes a espacios culturales diferentes, y se propone mostrar que estas discontinuidades no deben ser consideradas una falla sino índices de una tensión constitutiva de la novela.

The interweaving of popular generic modes and modernist techniques in Nostromo : A tale of the seaboard

Nostromo. A Tale of the Seaboard (1904), una de las novelas más experimentales del período modernista temprano, ha sido considerada la obra maestra de Joseph Conrad y su más grande lienzo. Un rasgo particular de la novela es que en ella coexisten la experimentación técnica del modernismo, la altamente difusa ficción del narrador y la situación de narración de historias y el paradigma narrativo de los subgéneros "degradados" de la novela de aventuras, el romance y el melodrama. Este estudio indaga las relaciones que se establecen entre dichos elementos, a todas luces anacrónicos o pertenecientes a espacios culturales diferentes, y se propone mostrar que estas discontinuidades no deben ser consideradas una falla sino índices de una tensión constitutiva de la novela.

The interweaving of popular generic modes and modernist techniques in Nostromo : A tale of the seaboard

Nostromo. A Tale of the Seaboard (1904), una de las novelas más experimentales del período modernista temprano, ha sido considerada la obra maestra de Joseph Conrad y su más grande lienzo. Un rasgo particular de la novela es que en ella coexisten la experimentación técnica del modernismo, la altamente difusa ficción del narrador y la situación de narración de historias y el paradigma narrativo de los subgéneros "degradados" de la novela de aventuras, el romance y el melodrama. Este estudio indaga las relaciones que se establecen entre dichos elementos, a todas luces anacrónicos o pertenecientes a espacios culturales diferentes, y se propone mostrar que estas discontinuidades no deben ser consideradas una falla sino índices de una tensión constitutiva de la novela.

Computational methods to create and analyze a digital gene expression atlas of embryo development from microscopy images

Abstract The creation of atlases, or digital models where information from different subjects can be combined, is a field of increasing interest in biomedical imaging. When a single image does not contain enough information to appropriately describe the organism under study, it is then necessary to acquire images of several individuals, each of them containing complementary data with respect to the rest of the components in the cohort. This approach allows creating digital prototypes, ranging from anatomical atlases of human patients and organs, obtained for instance from Magnetic Resonance Imaging, to gene expression cartographies of embryo development, typically achieved from Light Microscopy. Within such context, in this PhD Thesis we propose, develop and validate new dedicated image processing methodologies that, based on image registration techniques, bring information from multiple individuals into alignment within a single digital atlas model. We also elaborate a dedicated software visualization platform to explore the resulting wealth of multi-dimensional data and novel analysis algo-rithms to automatically mine the generated resource in search of bio¬logical insights. In particular, this work focuses on gene expression data from developing zebrafish embryos imaged at the cellular resolution level with Two-Photon Laser Scanning Microscopy. Disposing of quantitative measurements relating multiple gene expressions to cell position and their evolution in time is a fundamental prerequisite to understand embryogenesis multi-scale processes. However, the number of gene expressions that can be simultaneously stained in one acquisition is limited due to optical and labeling constraints. These limitations motivate the implementation of atlasing strategies that can recreate a virtual gene expression multiplex. The developed computational tools have been tested in two different scenarios. The first one is the early zebrafish embryogenesis where the resulting atlas constitutes a link between the phenotype and the genotype at the cellular level. The second one is the late zebrafish brain where the resulting atlas allows studies relating gene expression to brain regionalization and neurogenesis. The proposed computational frameworks have been adapted to the requirements of both scenarios, such as the integration of partial views of the embryo into a whole embryo model with cellular resolution or the registration of anatom¬ical traits with deformable transformation models non-dependent on any specific labeling. The software implementation of the atlas generation tool (Match-IT) and the visualization platform (Atlas-IT) together with the gene expression atlas resources developed in this Thesis are to be made freely available to the scientific community. Lastly, a novel proof-of-concept experiment integrates for the first time 3D gene expression atlas resources with cell lineages extracted from live embryos, opening up the door to correlate genetic and cellular spatio-temporal dynamics. La creación de atlas, o modelos digitales, donde la información de distintos sujetos puede ser combinada, es un campo de creciente interés en imagen biomédica. Cuando una sola imagen no contiene suficientes datos como para describir apropiadamente el organismo objeto de estudio, se hace necesario adquirir imágenes de varios individuos, cada una de las cuales contiene información complementaria respecto al resto de componentes del grupo. De este modo, es posible crear prototipos digitales, que pueden ir desde atlas anatómicos de órganos y pacientes humanos, adquiridos por ejemplo mediante Resonancia Magnética, hasta cartografías de la expresión genética del desarrollo de embrionario, típicamente adquiridas mediante Microscopía Optica. Dentro de este contexto, en esta Tesis Doctoral se introducen, desarrollan y validan nuevos métodos de procesado de imagen que, basándose en técnicas de registro de imagen, son capaces de alinear imágenes y datos provenientes de múltiples individuos en un solo atlas digital. Además, se ha elaborado una plataforma de visualization específicamente diseñada para explorar la gran cantidad de datos, caracterizados por su multi-dimensionalidad, que resulta de estos métodos. Asimismo, se han propuesto novedosos algoritmos de análisis y minería de datos que permiten inspeccionar automáticamente los atlas generados en busca de conclusiones biológicas significativas. En particular, este trabajo se centra en datos de expresión genética del desarrollo embrionario del pez cebra, adquiridos mediante Microscopía dos fotones con resolución celular. Disponer de medidas cuantitativas que relacionen estas expresiones genéticas con las posiciones celulares y su evolución en el tiempo es un prerrequisito fundamental para comprender los procesos multi-escala característicos de la morfogénesis. Sin embargo, el número de expresiones genéticos que pueden ser simultáneamente etiquetados en una sola adquisición es reducido debido a limitaciones tanto ópticas como del etiquetado. Estas limitaciones requieren la implementación de estrategias de creación de atlas que puedan recrear un multiplexado virtual de expresiones genéticas. Las herramientas computacionales desarrolladas han sido validadas en dos escenarios distintos. El primer escenario es el desarrollo embrionario temprano del pez cebra, donde el atlas resultante permite constituir un vínculo, a nivel celular, entre el fenotipo y el genotipo de este organismo modelo. El segundo escenario corresponde a estadios tardíos del desarrollo del cerebro del pez cebra, donde el atlas resultante permite relacionar expresiones genéticas con la regionalización del cerebro y la formación de neuronas. La plataforma computacional desarrollada ha sido adaptada a los requisitos y retos planteados en ambos escenarios, como la integración, a resolución celular, de vistas parciales dentro de un modelo consistente en un embrión completo, o el alineamiento entre estructuras de referencia anatómica equivalentes, logrado mediante el uso de modelos de transformación deformables que no requieren ningún marcador específico. Está previsto poner a disposición de la comunidad científica tanto la herramienta de generación de atlas (Match-IT), como su plataforma de visualización (Atlas-IT), así como las bases de datos de expresión genética creadas a partir de estas herramientas. Por último, dentro de la presente Tesis Doctoral, se ha incluido una prueba conceptual innovadora que permite integrar los mencionados atlas de expresión genética tridimensionales dentro del linaje celular extraído de una adquisición in vivo de un embrión. Esta prueba conceptual abre la puerta a la posibilidad de correlar, por primera vez, las dinámicas espacio-temporales de genes y células.

Self-organizing cultured neural networks: Image analysis techniques for longitudinal tracking and modeling of the underlying network structure

Esta tesis estudia la evolución estructural de conjuntos de neuronas como la capacidad de auto-organización desde conjuntos de neuronas separadas hasta que forman una red (clusterizada) compleja. Esta tesis contribuye con el diseño e implementación de un algoritmo no supervisado de segmentación basado en grafos con un coste computacional muy bajo. Este algoritmo proporciona de forma automática la estructura completa de la red a partir de imágenes de cultivos neuronales tomadas con microscopios de fase con una resolución muy alta. La estructura de la red es representada mediante un objeto matemático (matriz) cuyos nodos representan a las neuronas o grupos de neuronas y los enlaces son las conexiones reconstruidas entre ellos. Este algoritmo extrae también otras medidas morfológicas importantes que caracterizan a las neuronas y a las neuritas. A diferencia de otros algoritmos hasta el momento, que necesitan de fluorescencia y técnicas inmunocitoquímicas, el algoritmo propuesto permite el estudio longitudinal de forma no invasiva posibilitando el estudio durante la formación de un cultivo. Además, esta tesis, estudia de forma sistemática un grupo de variables topológicas que garantizan la posibilidad de cuantificar e investigar la progresión de las características principales durante el proceso de auto-organización del cultivo. Nuestros resultados muestran la existencia de un estado concreto correspondiente a redes con configuracin small-world y la emergencia de propiedades a micro- y meso-escala de la estructura de la red. Finalmente, identificamos los procesos físicos principales que guían las transformaciones morfológicas de los cultivos y proponemos un modelo de crecimiento de red que reproduce el comportamiento cuantitativamente de las observaciones experimentales. ABSTRACT The thesis analyzes the morphological evolution of assemblies of living neurons, as they self-organize from collections of separated cells into elaborated, clustered, networks. In particular, it contributes with the design and implementation of a graph-based unsupervised segmentation algorithm, having an associated very low computational cost. The processing automatically retrieves the whole network structure from large scale phase-contrast images taken at high resolution throughout the entire life of a cultured neuronal network. The network structure is represented by a mathematical object (a matrix) in which nodes are identified neurons or neurons clusters, and links are the reconstructed connections between them. The algorithm is also able to extract any other relevant morphological information characterizing neurons and neurites. More importantly, and at variance with other segmentation methods that require fluorescence imaging from immunocyto- chemistry techniques, our measures are non invasive and entitle us to carry out a fully longitudinal analysis during the maturation of a single culture. In turn, a systematic statistical analysis of a group of topological observables grants us the possibility of quantifying and tracking the progression of the main networks characteristics during the self-organization process of the culture. Our results point to the existence of a particular state corresponding to a small-world network configuration, in which several relevant graphs micro- and meso-scale properties emerge. Finally, we identify the main physical processes taking place during the cultures morphological transformations, and embed them into a simplified growth model that quantitatively reproduces the overall set of experimental observations.

Self-organizing cultured neural networks: Image analysis techniques for longitudinal tracking and modeling of the underlying network structure (versión actualizada)

Esta tesis estudia la evolución estructural de conjuntos de neuronas como la capacidad de auto-organización desde conjuntos de neuronas separadas hasta que forman una red (clusterizada) compleja. Esta tesis contribuye con el diseño e implementación de un algoritmo no supervisado de segmentación basado en grafos con un coste computacional muy bajo. Este algoritmo proporciona de forma automática la estructura completa de la red a partir de imágenes de cultivos neuronales tomadas con microscopios de fase con una resolución muy alta. La estructura de la red es representada mediante un objeto matemático (matriz) cuyos nodos representan a las neuronas o grupos de neuronas y los enlaces son las conexiones reconstruidas entre ellos. Este algoritmo extrae también otras medidas morfológicas importantes que caracterizan a las neuronas y a las neuritas. A diferencia de otros algoritmos hasta el momento, que necesitan de fluorescencia y técnicas inmunocitoquímicas, el algoritmo propuesto permite el estudio longitudinal de forma no invasiva posibilitando el estudio durante la formación de un cultivo. Además, esta tesis, estudia de forma sistemática un grupo de variables topológicas que garantizan la posibilidad de cuantificar e investigar la progresión de las características principales durante el proceso de auto-organización del cultivo. Nuestros resultados muestran la existencia de un estado concreto correspondiente a redes con configuracin small-world y la emergencia de propiedades a micro- y meso-escala de la estructura de la red. Finalmente, identificamos los procesos físicos principales que guían las transformaciones morfológicas de los cultivos y proponemos un modelo de crecimiento de red que reproduce el comportamiento cuantitativamente de las observaciones experimentales. ABSTRACT The thesis analyzes the morphological evolution of assemblies of living neurons, as they self-organize from collections of separated cells into elaborated, clustered, networks. In particular, it contributes with the design and implementation of a graph-based unsupervised segmentation algorithm, having an associated very low computational cost. The processing automatically retrieves the whole network structure from large scale phase-contrast images taken at high resolution throughout the entire life of a cultured neuronal network. The network structure is represented by a mathematical object (a matrix) in which nodes are identified neurons or neurons clusters, and links are the reconstructed connections between them. The algorithm is also able to extract any other relevant morphological information characterizing neurons and neurites. More importantly, and at variance with other segmentation methods that require fluorescence imaging from immunocyto- chemistry techniques, our measures are non invasive and entitle us to carry out a fully longitudinal analysis during the maturation of a single culture. In turn, a systematic statistical analysis of a group of topological observables grants us the possibility of quantifying and tracking the progression of the main networks characteristics during the self-organization process of the culture. Our results point to the existence of a particular state corresponding to a small-world network configuration, in which several relevant graphs micro- and meso-scale properties emerge. Finally, we identify the main physical processes taking place during the cultures morphological transformations, and embed them into a simplified growth model that quantitatively reproduces the overall set of experimental observations.

Prolonged activation of the N-methyl-d-aspartate receptor–Ca2+ transduction pathway causes spontaneous recurrent epileptiform discharges in hippocampal neurons in culture

The molecular basis for developing symptomatic epilepsy (epileptogenesis) remains ill defined. We show here in a well characterized hippocampal culture model of epilepsy that the induction of epileptogenesis is Ca2+-dependent. The concentration of intracellular free Ca2+ ([Ca2+]i) was monitored during the induction of epileptogenesis by prolonged electrographic seizure activity induced through low-Mg2+ treatment by confocal laser-scanning fluorescent microscopy to directly correlate changes in [Ca2+]i with alterations in membrane excitability measured by intracellular recording using whole-cell current–clamp techniques. The induction of long-lasting spontaneous recurrent epileptiform discharges, but not the Mg2+-induced spike discharges, was prevented in low-Ca2+ solutions and was dependent on activation of the N-methyl-d-aspartate (NMDA) receptor. The results provide direct evidence that prolonged activation of the NMDA–Ca2+ transduction pathway causes a long-lasting plasticity change in hippocampal neurons causing increased excitability leading to the occurrence of spontaneous, recurrent epileptiform discharges.

From cell death to embryo arrest: Mathematical models of human preimplantation embryo development

Human preimplantation embryos exhibit high levels of apoptotic cells and high rates of developmental arrest during the first week in vitro. The relation between the two is unclear and difficult to determine by conventional experimental approaches, partly because of limited numbers of embryos. We apply a mixture of experiment and mathematical modeling to show that observed levels of cell death can be reconciled with the high levels of embryo arrest seen in the human only if the developmental competence of embryos is already established at the zygote stage, and environmental factors merely modulate this. This suggests that research on improving in vitro fertilization success rates should move from its current concentration on optimizing culture media to focus more on the generation of a healthy zygote and on understanding the mechanisms that cause chromosomal and other abnormalities during early cleavage stages.

Hybrid vigor, fetal overgrowth, and viability of mice derived by nuclear cloning and tetraploid embryo complementation

To assess whether heterozygosity of the donor cell genome was a general parameter crucial for long-term survival of cloned animals, we tested the ability of embryonic stem (ES) cells with either an inbred or F1 genetic background to generate cloned mice by nuclear transfer. Most clones derived from five F1 ES cell lines survived to adulthood. In contrast, clones from three inbred ES cell lines invariably died shortly after birth due to respiratory failure. Comparison of mice derived from nuclear cloning, in which a complete blastocyst is derived from a single ES cell, and tetraploid blastocyst complementation, in which only the inner cell mass is formed from a few injected ES cells, allows us to determine which phenotypes depend on the technique or on the characteristics of the ES cell line. Neonatal lethality also has been reported in mice entirely derived from inbred ES cells that had been injected into tetraploid blastocysts (ES cell-tetraploids). Like inbred clones, ES cell-tetraploid pups derived from inbred ES cell lines died shortly after delivery with signs of respiratory distress. In contrast, most ES cell-tetraploid neonates, derived from six F1 ES cell lines, developed into fertile adults. Cloned pups obtained from both inbred and F1 ES cell nuclei frequently displayed increased placental and birth weights whereas ES cell-tetraploid pups were of normal weight. The potency of F1 ES cells to generate live, fertile adults was not lost after either long-term in vitro culture or serial gene targeting events. We conclude that genetic heterozygosity is a crucial parameter for postnatal survival of mice that are entirely derived from ES cells by either nuclear cloning or tetraploid embryo complementation. In addition, our results demonstrate that tetraploid embryo complementation using F1 ES cells represents a simple, efficient procedure for deriving animals with complex genetic alterations without the need for a chimeric intermediate.