Regulación de NOS-3 durante la muerte hepatocelular inducida por ácidos biliares

1. Introducción o motivación de la tesis. La enfermedad hepática colestásica se caracteriza por una alteración en la formación de la bilis, que viene acompañada o es causada por un defecto en la excreción de la misma hacia el canalículo biliar. Como resultado, se produce una alteración en la circulación enterohepática de los ácidos biliares, aumentando su concentración a nivel sérico y hepático. Las consecuencias de la colestasis no tratadas son severas y se asocian con el desarrollo de fibrosis, cirrosis, hipertensión portal, fallo hepático y la aparición de carcinoma hepatocelular y colangiocelular. En la actualidad, las opciones de tratamiento farmacológico para el manejo de las enfermedades colestásicas son muy limitadas. Así, la terapia con el ácido ursodesoxicólico UDCA) resulta útil en el tratamiento de la colangitis biliar primaria, la colangitis esclerosante primaria o la colestasis intrahepática del ambarazo, aunque su eficacia puede ser limitada en algunas circunstancias (Bessho and Bezerra, 2011; Maillette de Buy Wenniger and Beuers, 2010; Zein and Lindor, 2010). De ahí la importancia de disponer de un mayor conocimiento de los mecanismos a través de los cuales los ácidos biliares ejercen su efecto citotóxico, para poder así diseñar nuevas estrategias terapéuticas eficaces y seguras u optimizar las ya existentes. El ácido glicoquenodesoxicólico (GCDCA) es una de las sales biliares más representativas en el suero de pacientes colestásicos (Woolbright et al., 2015) y su acumulación en el hígado durante la colestasis ha sido directamente asociada con el daño hepatocelular (Spivey et al., 1993). Por tanto, GCDCA es considerado un excelente candidato para valorar los mecanismos de toxicidad por acumulación de sales biliares en modelos experimentales in vitro e in vivo. Numerosas pruebas experimentales han demostrado que el desarrollo de la enfermedad hepática colestásica se relaciona con diversos mecanismos celulares íntimamente relacionados, incluyendo la disfunción mitocondrial, el estrés oxidativo y la muerte celular (Rodrigues et al., 1998a; Rodrigues et al., 1998b). En este sentido, el uso terapéutico de los antioxidantes ha suscitado interés para el tratamiento de las enfermedades colestásicas (Galicia-Moreno et al., 2012; Gane et al., 2010; Spirli et al., 2003). El óxido nítrico (ON) se sintetiza a partir del aminoácido L-arginina mediante la participación de la óxido nítrico sintasa (NOS), cuya activación depende de la concentración intracelular de cálcio ([Ca2+]i. En el hígado, el ON desempeña un papel citoprotector o citotóxico en función de factores tales como su concentración, la fuente que lo genera y la presencia de especies reactivas de oxígeno (EROs) (Coskun et al., 2001; Fiorucci et al., 2001; Iwakiri and Kim, 2015; Moussa et al., 2000). Diversos estudios han demostrado que la desregulación de la expresión de la óxido nítrico sintasa endotelial (NOS-3) actúa como un factor clave durante la progresión del daño hepático (Palmer et al., 1987). Sin embargo, el papel de la NOS-3 no ha sido claramente establecido durante la enfermedad hepática colestásica. El presente trabajo parte de la hipótesis de que la acumulación de sales biliares que ocurre durante la colestasis provoca una respuesta hepatocelular prooxidante y proapoptótica, que podría estar regulada por la biodisponibilidad de ON sintetizado por la NOS-3. Por ello, el objetivo general de esta tesis doctoral fue clarificar los mecanismos moleculares que subyacen a la citotoxicidad hepatocelular durante la enfermedad hepática colestásica y la participación de NOS-3 en este proceso. 2. Contenido de la investigación. Los resultados obtenidos indican que la respuesta citotóxica a GCDCA se caracterizó por la alteración del sistema redox celular, la disminución de la actividad combinada de los complejos respiratorios II+III, la acumulación de especies reactivas de oxígeno (EROs), una mayor expresión y activación de los factores de transcripción (FTs) Sp1, cJun y cFos, así como una mayor capacidad de unión de los mismos a la región promotora de NOS-3 (pNOS-3), en la posiciones -1386, -632 y -104 para Sp1, y -666 para cJun y cFos. Esto se asoció con una disminución de la actividad del pNOS-3 y, en consecuencia, con una menor expresión/actividad de NOS-3. La disminución de la expresión de NOS-3 durante la toxicidad inducida por GCDCA coincidió con el descenso de la concetración de cácio citosólico ([Ca2+]i). El tratamiento antioxidante causó una menor activación de Sp1, cJun y cFos, previno la reducción de la [Ca2+]i, recuperó la actividad del pNOS-3 y la expresión/actividad de NOS-3 y redujo la muerte celular provocada por la acumulación de la sal biliar. De manera similar, la inhibición específica de Sp1 y AP-1 recuperó la expresión de NOS-3 y suprimió los efectos citotóxicos ejercidos por GCDCA. De manera complementaria, el bloqueo farmacológico de AP-1 con SR11302 redujo la expresión de ciclina D1, inducida por GCDCA y relacionada con su efecto deletéreo. La inhibición específica de la actividad NOS por L-NAME inhibió el efecto protector de SR11302. Cabe destacar que, en este modelo de colestasis, no fueron detectadas las isoformas neuronal e inducible de la NOS. La ligadura del conducto biliar en ratas confirmó los resultados del modelo in vitro relativos a la activación de los FTs, la reducción de la expresión de NOS-3 y el daño hepatocelular inducido por la obstrucción biliar. 3. Conclusión. • El daño hepatocelular provocado por GCDCA se asocia con una menor expresión de NOS-3 y una menor acumulación de los productos finales del ON. • La desregulación de la expresión de NOS-3 durante la citotoxicidad por GCDCA coincide con la disminución de la [Ca2+] en el citosol, asociada a su acumulación en el RE. Este efecto es dependiente de la acumulación de estrés oxidativo celular. • Sp1 y AP-1 actúan como represores transcripcionales de la expresión de NOS-3 y favorecen la muerte celular durante la colestasis experimental. • NOS-3 ejerce un papel crítico en la regulación de la expresión de ciclina D1 durante el daño celular inducido por GCDCA. • Se requieren investigaciones adicionales para determinar el valor terapéutico de los inhibidores de Sp1 y de AP-1 para el tratamiento de las enfermedades hepáticas colestásicas

Meta-analytic study on the frequency and treatment of oral antral communications

Objective: To determine the optimum surgical treatment for oral antral communications (OAC) and to understand the main post-operative complications.Study Design: Meta-analytical, observational and retrospective study of 1,072 cases of OAC obtained from a literature review of 15 articles. Results: OAC occur slightly more often in men and during the fourth decade of life. Its primary etiological factor is dental extraction, most often affecting the third molar. The most common treatment has been the use of Bichat's fat pad grafts, whereas the technique with the highest percentage of complications has been the use of the palatal rotation flap. The most frequent complication has been the fistulization of the OAC. Conclusions: Early diagnosis of OAC and its treatment within 48 hours of evolution are fundamental in order to properly resolve this pathology. The use of Bichat's fat pad grafts is a simple technique that offers excellent vascularization and results. © Medicina Oral

Sculpture Making Process of Elba, the Mesolithic Woman of Chan do Lindeiro (Lugo)

[Abstract] This article addresses the process of making the hyper-realistic sculpture of Elba, the Mesolithic shepherd of O Courel, whose remains were found in the cave of Chan do Lindeiro (Pedrafita do Cebreiro, Lugo). It outlines the technical processes that were carried out during the sculpture´s creation, following the same order in which each of the steps of this artistic project were followed based on the scientific knowledge that is available up to now.[Resumen] En este artículo se aborda el proceso de elaboración de la escultura hiperrealista de Elba, la pastora mesolítica de O Courel, cuyos restos fueron encontrados en la cueva de Chan do Lindeiro (Pedrafita do Cebreiro, Lugo). En él se definen los procesos técnicos que se han realizado durante la ejecución de la pieza escultórica, siguiendo el mismo orden en el que se han llevado a cabo cada una de las operaciones de este proyecto artístico basado en los conocimientos científicos que se tienen hasta la fecha

143 PRINCIPAL COMPONENT CLUSTERING OF FRONTAL PLANE KNEE KINEMATICS

Replication forks stall at different DNA obstacles such as those originated by transcription. Fork stalling can lead to DNA double-strand breaks (DSBs) that will be preferentially repaired by homologous recombination when the sister chromatid is available. The Rrm3 helicase is a replisome component that promotes replication upon fork stalling, accumulates at highly transcribed regions and prevents not only transcription-induced replication fork stalling but also transcription-associated hyper-recombination. This led us to explore the possible role of Rrm3 in the repair of DSBs when originating at the passage of the replication fork. Using a mini-HO system that induces mainly single-stranded DNA breaks, we show that rrm3Δ cells are defective in DSB repair. The defect is clearly seen in sister chromatid recombination, the major repair pathway of replication-born DSBs. Our results indicate that Rrm3 recruitment to replication-born DSBs is crucial for viability, uncovering a new role for Rrm3 in the repair of broken replication forks.This work was supported by grants from the Spanish Ministry of Economy and Innovation (BFU2013-42918), the European Union (FEDER), the European Research Council (ERC2014 AdG669898 TARLOOP), and the Junta de Andalucía (BIO1238). SMG was funded by a predoctoral training fellowship from the Spanish National Research Council (CSIC) and BGG by a postdoctoral grant from the Scientific Foundation of the Spanish Association Against Cancer (AECC). Funding for open access charge: Grants from the Spanish Ministry of Economy and Innovation (BFU2013-42918).Peer Reviewe

El concepto de salario emocional

El presente trabajo tiene como objetivo enlistar las diferentes definiciones que se han redactado sobre Salario Emocional (SE) y establecer una definición propia sobre el concepto, a través de la búsqueda bibliográfica entre los años 2014 y 2019, en diferentes bases de datos científicas, utilizando los descriptores: salario, retribución emocional, salario emocional, motivación laboral y emociones en el trabajo; se identificaron diversas acepciones que aluden al termino SE, proponiendo como definición propia del concepto Salario Emocional, como los elementos extrínsecos e intrínsecos del trabajo que generan emociones positivas. Concluyendo que el SE, contribuye en la generación de entornos organizacionales favorables y saludables.This paper aims to list the different variables that have been written about Emotional Salary (ES) and establish a definition of the concept itself, through a bibliographic search between 2014 and 2019, in different scientific databases, using as searching words: salary, emotional compensation, emotional salary, work motivation and emotions at work; several meanings that refer to the term ES were identified, proposing our definition for the concept  Emotional Salary, as the outward and within elements of work that generates positive emotions. Concluding that the ES, contributes to the generation of favorable and healthy organizational environments

Advances in the control of phytopathogenic fungi that infect crops through their root system

[EN] Productivity and economic sustainability of many herbaceous and woody crops are seriously threatened by numerous phytopathogenic fungi. While symptoms associated with phytopathogenic fungal infections of aerial parts (leaves, stems and fruits) are easily observable and therefore recognizable, allowing rapid or preventive action to control this type of infection, the effects produced by soil-borne fungi that infect plants through their root system are more difficult to detect. The fact that these fungi initiate infection and damage underground implies that the first symptoms are not as easily noticeable, and therefore both crop yield and plant survival are frequently severely compromised by the time the infection is found. In this paper we will review and discuss recent insights into plant-microbiota interactions in the root system crucial to understanding the beginning of the infectious process. We will also review different methods for diminishing and controlling the infection rate by phytopathogenic fungi penetrating through the root system including both the traditional use of biocontrol agents such as antifungal compounds as well as some new strategies that could be used because of their effective application, such as nanoparticles, virus-based nanopesticides, or inoculation of plant material with selected endophytes. We will also review the possibility of modeling and influencing the composition of the microbial population in the rhizosphere environment as a strategy for nudging the plant-microbiome interactions toward enhanced beneficial outcomes for the plant, such as controlling the infectious processSIS.G.-G. was supported by a FPU fellowship (Grant number FPU15/03475) from the Ministerio de Educacio´n, Cultura y Deporte (Madrid, Spain). A.M.I. and A.D.G. were supported by a predoctoral fellowship from the Junta de Castilla y Leo´n. C.C.-P. was supported by a technician contract co-financed by the Iniciativa de Empleo Juvenil (Junta de Castilla y León) and the European Social Fun

In-depth in vitro Evaluation of the Activity and Mechanisms of Action of Organic Acids and Essential Oils Against Swine Enteropathogenic Bacteria

P. 1-13Alternative antimicrobials require a deep understanding of their action mechanisms by in vitro assays which support science-based field use. This study focuses on the characterization of bactericidal mechanisms of potential antimicrobial compounds, two organic acids and three single essential oil (EO) compounds against swine enteropathogenic bacteria Escherichia coli, Salmonella enterica subsp. enterica serovar Typhimurium, and Clostridium perfringens. Target concentrations of the compounds were evaluated using the inhibitory potential of the vapor phase and bacterial viability after short-term exposure, while cell targets were disclosed using flow cytometry (FC), Fourier-transform infrared (FTIR) spectroscopy, and scanning electron microscopy (SEM). All tested compounds exhibited vapor phase activity against the three bacterial species, except sodium salt of coconut fatty acid distillates against C. perfringens. Survival test results evidenced that effects on bacterial viability were concentration dependent and higher in single EO compounds than in organic acids. In detail, thymol and its isomer carvacrol were the most effective compounds. Further characterization of thymol and cinnamaldehyde activity revealed that thymol main target was the cell membrane, since it caused striking damages in the membrane permeability, integrity and composition evidenced by FC and FTIR in the three enteric pathogens. In contrast, cinnamaldehyde was more effective against enterobacteria than against C. perfringens and only caused slightly damages at the highest concentration tested. Its target at the molecular level differed between enterobacteria and C. perfringens isolates. The SEM micrographs allowed us to confirm the results previously obtained for both EO compounds by other techniques. Altogether, the study showed the straight effect of these antimicrobials, which could constitute relevant information to optimize their feed inclusion rates in field studies or field use.S

Viticultural and Biotechnological Strategies to Reduce Alcohol Content in Red Wines

Viticultural and biotechnological strategies are two approaches to deal with higher must sugar levels at harvest time. A wide range of factors could significantly affect sugar accumulation in the grape such as choice of vineyard site, soil composition, irrigation strategy, rootstock, and grape cultivar selection as well as grape yield. In this sense, approaches to canopy management are continually evolving in response to changes in other vineyard management practices; some of these could contribute to reduce soluble sugars on grape berries at harvest time. On the other hand, among possible biotechnological strategies, one of the most relevant is the control of the fermentative process by using selected yeast strains. In this chapter, we will show how some viticultural practices have influenced the accumulation of soluble sugars and other enological parameters in grape berries at harvest time. We will also report how a careful yeast selection and the implementation of different fermentation strategies can also contribute to reduce ethanol content in wines

Circulating Tumor Cells in Hepatocellular Carcinoma: A Comprehensive Review and Critical Appraisal

Hepatocellular carcinoma (HCC) is the fifth most common neoplasm and a major cause of cancer-related death worldwide. There is no ideal biomarker allowing early diagnosis of HCC and tumor surveillance in patients receiving therapy. Liquid biopsy, and particularly circulating tumor cells (CTCs), have emerged as a useful tool for diagnosis and monitoring therapeutic responses in different tumors. In the present manuscript, we evaluate the current evidence supporting the quantitative and qualitative assessment of CTCs as potential biomarkers of HCC, as well as technical aspects related to isolation, identification, and classification of CTCs. Although the dynamic assessment of CTCs in patients with HCC may aid the decision-making process, there are still many uncertainties and technical caveats to be solved before this methodology has a true impact on clinical practice guidelines. More studies are needed to identify the optimal combination of surface markers, to increase the efficiency of ex-vivo expansion of CTCs, or even to target CTCs as a potential therapeutic strategy to prevent HCC recurrence after surgery or to hamper tumor progression and extrahepatic spreading

Use of Endophytic and Rhizosphere Actinobacteria from Grapevine Plants To Reduce Nursery Fungal Graft Infections That Lead to Young Grapevine Decline

[EN] Endophytic and rhizosphere actinobacteria isolated from the root system of 1-year-old grafted Vitis vinifera plants were evaluated for their activities against fungi that cause grapevine trunk diseases. A total of 58 endophytic and 94 rhizosphere isolates were tested. Based on an in vitro bioassay, 15.5% of the endophytic isolates and 30.8% of the rhizosphere isolates exhibited antifungal activity against the fungal pathogen Diplodia seriata, whereas 13.8% of the endophytic isolates and 16.0% of the rhizosphere isolates showed antifungal activity against Dactylonectria macrodidyma (formerly Ilyonectria macrodidyma). The strains which showed the greatest in vitro efficacy against both pathogens were further analyzed for their ability to inhibit the growth of Phaeomoniella chlamydospora and Phaeoacremonium minimum (formerly Phaeoacremonium aleophilum). Based on their antifungal activity, three rhizosphere isolates and three endophytic isolates were applied on grafts in an open-root field nursery in a 3-year trial. The field trial led to the identification of one endophytic strain, Streptomyces sp. VV/E1, and two rhizosphere isolates, Streptomyces sp. VV/R1 and Streptomyces sp. VV/R4, which significantly reduced the infection rates produced by the fungal pathogens Dactylonectria sp., Ilyonectria sp., P. chlamydospora, and P. minimum, all of which cause young grapevine decline. The VV/R1 and VV/R4 isolates also significantly reduced the mortality level of grafted plants in the nursery. This study shows that certain actinobacteria could represent a promising new tool for controlling fungal trunk pathogens that infect grapevine plants through the root system in nurseries.SIThis work was supported by Viveros Villanueva Vides S.L. (Larraga, Spain), which was financed by the Centro para el Desarrollo Tecnológico Industrial (CDTI; Madrid, Spain) and the Government of Comunidad Foral de Navarra (Spain). S. González-García was supported by a FPU fellowship of the Ministerio de Educación, Cultura y Deporte (Madrid, Spain