Establishment of a transgenic system for in vivo detection of apoptosis in the developing heart

All living organisms face death since the beginning of their lives. Cells commit suicide to sculpt correctly all organs during animal development. Currently, there exist at least 13 cell death modalities through which a cell is programmed to die. From these, the most studied is apoptosis. Despite the huge progress made in the field of apoptosis, little is known about its role in mouse heart development and it is still not clear if its upregulation or downregulation can lead to heart disease. One of the main reasons for this uncertainty is the lack of techniques that mark apoptotic cells at different time points in vivo and in fixed tissue. With a specific and broad detection method that overcomes the rapidity in which apoptotic cells die and disappear from tissues, it should be possible to confirm if apoptosis takes place during mouse heart development, in which areas and at which specific rates occurs during different embryonic stages and which are the cells that undergo apoptosis during heart morphogenesis. In this work, I addressed these issues by establishing a genetically encoded reporter tool used to visualize and quantify apoptosis during mouse heart development in vivo. This system is based on the detection of phosphatidylserine (PS) residues—exposed in the outer membrane of apoptotic cells—by Annexin V (A5). Commercial probes of A5 linked to fluorescent probes have been developed. These probes provide excellent results in cells, but have some limitations such as high cost and not-specific for fixed tissue, when used in fixed tissue or in living organisms by systemic injection. Therefore, this system was aimed to overcome these drawbacks by ubiquitously expressing a secreted human Annexin V (sA5) fused to the yellow fluorescent protein (YFP). The ubiquitous expression was ensured by the chicken β-actin promoter with CMV enhancer element (CAG). As cells undergo apoptosis, sA5-YFP binds with high affinity to its PS residues resulting in bright fluorescent signals. In this way, it was possible to better characterize, quantify and monitor apoptotic signals in the developing mouse heart over time. In summary, this study consisted of two major goals. The first one was to validate the sA5-YFP system, in vitro and in vivo, to specifically label apoptotic cells. The second goal was to use this system to describe the apoptotic patterns and rates during all stages of embryonic heart development in more detail. For the first part, I could show that the labeled sA5-YFP signals in transgenic mESCs, in EBs, and in sA5-YFP mice were apoptotic cells after inducing apoptosis ectopically or in entities (EBs and embryonic tissue such as neural tube, otic placode and yolk sac) in which apoptosis is a physiological process. These cells presented the typical morphological and biochemical changes of apoptotic cells. They rounded up, detached from the culture plate or tissue compound, formed membrane blebs or fragmented, and displayed condensed chromatin. Moreover, the sA5-YFP protein co-localized with active forms of pro-caspases (casp-8), effector caspases (active casp-3 and 7) and with TUNEL staining. Thus, I proved that the sA5-YFP system allows the visualization of cells and blebs (fragments of apoptotic cells) in different stages (early-intermediate-late) of apoptosis. For the second goal, I used the transgenic CAG-sA5-YFP mouse line to provide detailed information about apoptosis rates, distribution of apoptotic events and cells undergoing apoptosis during mouse heart development. I could show that apoptosis takes place at all stages of the mouse heart development starting from E9.5. I observed than less than 1.5% of the cells in the embryonic heart undergo physiological apoptosis and the apoptosis rates decrease as the heart ages. In contrast to earlier studies, apoptotic cells were observed very clearly in the primitive ventricle at E9.5 and in the lining of the trabeculae of both ventricles from E10.5 - E14.5 in sA5-YFP hearts. These events coincide with episodes of trabeculae formation, differentiation, remodeling, and compaction, suggesting that apoptosis might be necessary for chamber maturation. In addition, I noticed apoptotic events in the outflow tract, the interventricular septum, and atria, coinciding with previous reports. Lastly, I demonstrated that the labeled sA5-YFP apoptotic cells, found in the trabeculae, co-localize mainly with cardiomyocytes around days E13.5 - E14.5 of mouse heart development. These findings imply a role of cardiomyocyte apoptosis in ventricular differentiation. However, more experiments are needed to undercover the biological role of apoptosis in trabeculae formation. Last but not least, I provided evidence that the CAG-sA5-YFP mouse model enables the real-time imaging of induced apoptosis and physiological occurring apoptosis in the developing mouse embryo at E8.75 and in the embryonic mouse heart at E9. Apoptotic signals can be identified by their roundish morphology and bright sA5-YFP+ signals. Fluorescence intensity of sA5-YFP can be used as quantitative parameter to evaluate changes in apoptosis in vivo. This means that this reporter line can be used to monitor the distribution and morphological changes of apoptotic cells within specific tissues over time in living organisms. In conclusion, the sA5-YFP mouse has the potential to unravel the physiological role of apoptosis in the developing heart, its role in the formation of other mammalian organs, and in diseases displaying aberrant patterns of apoptosis

Disseny d’un mètode per avaluar la qualitat del servei d’una empresa de manteniment d’enllumenat públic

El objectiu principal d’aquest projecte és desenvolupar un mètode per avaluar la qualitat del servei d’una empresa de manteniment d’enllumenat públic. El present document s’estructura en diferents blocs, començant per els temes d’introducció, on s’explica la teoria sobre l’enllumenat públic i es defineixen els elements que el formen, coneixement necessari per la comprensió del projecte. A continuació, es procedeix a explicar les característiques dels serveis dels quals s’encarrega una Empresa de Serveis Energètics (en endavant ESE), les quals basen el negoci en aconseguir un estalvi energètic en les empreses dels seus clients. Aquests serveis són; el servei lumínic, centrat en la quantitat i qualitat de la llum; el servei de manteniment, el qual s’ha d’encarregar de definir i gestionar de manera adequada un programa de manteniment de tots els equips que formen el sistema; i per últim, el sistema energètic, el qual controla que el consum energètic que genera l’enllumenat públic sigui el correcte. Posterior a aquest punt s’exemplificarà el mètode a partir de les dades reals de l’enllumenat públic del municipi de Deltebre, municipi el qual té tot el sistema d’enllumenat públic gestionat per una Empresa de Serveis Energètics. Finalment s’han plantejat tres escenaris en funció dels diners que l’Ajuntament en qüestió vulgui dedicar a l’aplicació del mètode, i en funció d’aquest s’ha explicat el procediment a realitzar i s’han llistat els costos que suposaria, per finalment, acabar arribant a un pressupost final per cadascun d’ells. L’aplicació d’un mètode com el desenvolupat en aquest projecte garanteix un estudi complet de tots els serveis que formen part un sistema d’enllumenat públic. L’estudi exhaustiu de cadascun dels serveix conclou en un llistat de propostes d’accions que hauran de portar-se a termes si es vol garantir el correcte funcionament de tot el sistema d’enllumenat públic

Surface-Bound Molecular Gradients for the High-Throughput Screening of Cell Responses

Chemical gradient surfaces are described as surfaces with a gradually varying composition along their length. Continuous chemical gradients have recently been proposed as an alternative to discrete microarrays for the high-throughput screening of the effects of ligand concentration in cells. Here, we review some of the most recent examples in which gradients have been used to evaluate the effect of a varying ligand concentration in cell adhesion, morphology, growth, and differentiation of cells, including some of our recent findings. They show the importance of the organization of ligands at the nanoscale, which is highlighted by abrupt changes in cell behavior at critical concentration thresholds

Las mujeres víctimas de violencia política en Aragón durante la Guerra Civil y la posguerra (1936-1945)

Las múltiples formas de represión que empleó el Régimen franquista afectaron a hombres y mujeres. Sin embargo, éstas han quedado relegadas en el relato historiográfico y, con él, en la memoria colectiva. Este trabajo pretende ahondar en las violencias de carácter político que sufrieron, y analizar si también el género jugó su papel en el ejercicio de la venganza y la construcción del Nuevo Estado, analizando violencias sufridas por ambos sexos como los asesinatos, la represión económica, la depuración o los consejos de guerra, utilizando Aragón como marco de desarrollo

¿Ángel del hogar o mujer trabajadora?: Presencia femenina en los mercados laborales españoles desde la Revolución Industrial.

RESUMEN: La Revolución Industrial supuso enormes cambios que, dentro del ámbito laboral, afectaron de manera diferente a mujeres y hombres. Con el siguiente trabajo se pretende exponer una visión general de la situación de la mujer trabajadora en los distintos sectores laborales, su discriminación con respecto al trabajador varón y la fuerte carga doméstica que condicionaba su vida. Además, se cuestiona la masiva salida femenina de los mercados de trabajo con el avance del capitalismo industrial, así como el verdadero cumplimiento del modelo de ganador de pan, prácticamente imposible para la mayoría de las familias obreras. Palabras clave: Trabajo, obreras, mercados laborales, hogar, salarios, condiciones laborales, industrialización. ABSTRACT: Inside the labour sphere, the Industrial Revolution caused great changes that affected women and men in a different way. The following proyect pretends to show a general vision about the situation of working woman in the differents labour sectors, her discrimination in relation to working man and the heavy domestic duty that conditioned her life. Besides, it is questioned the masive female exit from the labour market in the industrial capitalism advance, as well as the real fullfilment of the male breadwinner model, practically imposible for most workers families. Key Words: Job, workers, labour market, household, wages, labour conditions, industrialization

Delitos, justicia ordinaria y control social en Zaragoza (1931-1945): de la II República a la Dictadura de Franco

La justicia ha sido empleada por el poder como un mecanismo de control y regulación social. A través de la fiscalización de ciertos comportamientos entendidos como ilegales, los grupos de poder han pretendido preservar el orden y el statu quo ante cualquier alteración o amenaza interna. Su pretensión apuntaba a la conservación de su situación de poder, de sus intereses personales o, en última instancia, del propio Estado. Desde arriba, cualquier comportamiento que sea entendido como un potencial foco de peligro será, inmediatamente, situado fuera de la legalidad; el poder ¿crea¿ ilegalidades. Sin embargo, la aparición de la delincuencia responde a una variada casuística. Las acciones delictuales encuentran su propia lógica en el contexto económico, social, político, cultural o ético en donde se producen. No son simples actos marginales; detrás, encontramos razones que los explican. Desde abajo, la existencia del delito supone la no aceptación por una parte de la población de los dogmas económicos, políticos, éticos, etc., que el poder determina como idóneos y protege mediante el castigo penal. Los comportamientos ilegales en Zaragoza durante la II República, la Guerra Civil y el franquismo fueron constantes, variados y alcanzaron a todas las esferas de la vida. El análisis de los delincuentes nos indica que la delincuencia no se circunscribía únicamente a las clases sociales más bajas, si bien estas eran las que infringían la ley en un mayor número. Elementos pertenecientes a las clases altas, a las estructuras de poder o con buena situación económica también delinquían. Sin embargo, a ambos grupos les movían incentivos bien distintos. Tanto la justicia republicana como la franquista otorgaron mayoritariamente veredictos condenatorios para estas acciones ilegales. No obstante, existió una diferencia: la justicia ordinaria franquista tuvo como objetivo la ejemplaridad y la punición, antes que la reinserción o recuperación del infractor. Finalmente, a través del análisis cualitativo de las sentencias criminales observaremos la existencia de continuidades delictivas entre ambos períodos, pero, también, de algunas nuevas infracciones durante la dictadura. En efecto, pues como fenómeno social, la delincuencia permanece constante, se modifica o se adapta a los cambios políticos, económicos, culturales, etc., acaecidos en el entorno donde surge

Biosensors Integration in blood-brain barrier-on-a-chip: emerging platform for monitoring neurodegenerative diseases

Over the most recent decades, the development of new biological platforms to study disease progression and drug efficacy has been of great interest due to the high increase in the rate of neurodegenerative diseases (NDDs). Therefore, blood–brain barrier (BBB) as an organ-on-a-chip (OoC) platform to mimic brain-barrier performance could offer a deeper understanding of NDDs as well as a very valuable tool for drug permeability testing for new treatments. A very attractive improvement of BBB-oC technology is the integration of detection systems to provide continuous monitoring of biomarkers in real time and a fully automated analysis of drug permeably, rendering more efficient platforms for commercialization. In this Perspective, an overview of the main BBB-oC configurations is introduced and a critical vision of the BBB-oC platforms integrating electronic read out systems is detailed, indicating the strengths and weaknesses of current devices, proposing the great potential for biosensors integration in BBB-oC. In this direction, we name potential biomarkers to monitor the evolution of NDDs related to the BBB and/or drug cytotoxicity using biosensor technology in BBB-oC

Resultados de la recaudación del impuesto a las nóminas en el caribe mexicano: gestión presidencial 2018 - 2019

Analysis has been made on whether the presidential administration of Mexico, which began in December 2018 with anti-neoliberal economic policies, has had an impact and to what extent whit respect to the collection of the payroll tax in the state of Quintana Roo and, as a contrast variable, the international passengers.  The adjustment variable, identify the distribution in order to represent the magnitude of the changes. To find out if there is some independence from the government management, it´s been realized independence test in different presidential terms. It is note that there were significant changes in the collection of the payroll test, however, the Independence test suggest that despite them where unrelated to the presidential management.Se analiza si la administración presidencial de México, iniciada en diciembre 2018, con políticas económicas antineoliberales, ha repercutido y en qué magnitud respecto a la recaudación del impuesto sobre nóminas en el estado de Quintana Roo y, como variable de contraste, en los pasajeros internacionales. Mediante pruebas de ajuste, se identifica la distribución para representar la magnitud de los cambios. Para conocer si existe independencia de la gestión gubernamental, se realizan pruebas de independencia en períodos presidenciales distintos. Se advierte que en la recaudación sí se presentaron cambios significativos, no obstante, la prueba de independencia sugiere que a pesar de ello fueron ajenos a la gestión presidencial