New insights on the mechanism of polyethylenimine transfection and their implications on gene therapy and DNA vaccines

Polyethylenimine (PEI) has been demonstrated as an efficient DNA delivery vehicle both in vitro and in vivo. There is a consensus that PEI-DNA complexes enter the cells by endocytosis and escape from endosomes by the so-called “proton sponge” effect. However, little is known on how and where the polyplexes are de-complexed for DNA transcription and replication to occur inside the cell nucleus. To better understand this issue, we (i) tracked the cell internalization of PEI upon transfection to human epithelial cells and (ii) studied the interaction of PEI with phospholipidic layers mimicking nuclear membranes. Both the biological and physicochemical experiments provided evidence of a strong binding affinity between PEI and the lipidic bilayer. Firstly, confocal microscopy revealed that PEI alone could not penetrate the cell nucleus; instead, it arranged throughout the cytoplasm and formed a sort of aureole surrounding the nuclei periphery. Secondly, surface tension measurements, fluorescence dye leakage assays, and differential scanning calorimetry demonstrated that a combination of hydrophobic and electrostatic interactions between PEI and the phospholipidic monolayers/bilayers led to the formation of stable defects along the model membranes, allowing the intercalation of PEI through the monolayer/bilayer structure. Results are also supported by molecular dynamics simulation of the pore formation in PEI-lipidic bilayers. As discussed throughout the text, these results might shed light on a the mechanism in which the interaction between PEI and the nucleus membrane might play an active role on the DNA release: on the one hand, the PEI-membrane interaction is anticipated to facilitate the DNA disassembly from the polyplex by establishing a competition with DNA for the PEI binding and on the other hand, the forming defects are expected to serve as channels for the entrance of de-complexed DNA into the cell nucleus. A better understanding of the mechanism of transfection of cationic polymers opens paths to development of more efficiency vectors to improve gene therapy treatment and the new generation of DNA vaccinesThis work was supported by the Spanish "Ministerio de Ciencia, Innovación y Universidades" (Project PID2019–109517RB-I00)S

Determining major factors controlling phosphorus removal by promising adsorbents used for lake restoration: a linear mixed model approach

Phosphorus (P) removal from lake/drainage waters by novel adsorbents may be affected by competitive substances naturally present in the aqueous media. Up to date, the effect of interfering substances has been studied basically on simple matrices (single-factor effects) or by applying unsuitable statistical approaches when using natural lake water. In this study, we determined major factors controlling P removal efficiency in 20 aquatic ecosystems in the southeast Spain by using linear mixed models. Two non-magnetic -CFH-12® and Phoslock®- and two magnetic materials -hydrous lanthanum oxide loaded silica-coated magnetite (Fe-Si-La) and commercial zero-valent iron particles (FeHQ)- were tested to remove P at two adsorbent dosages. Results showed that the type of adsorbent, the adsorbent dosage and color of water (indicative of humic substances) are major factors controlling P removal efficiency. Differences in physico-chemical properties (i.e. surface charge or specific surface), composition and structure explain differences in maximum P adsorption capacity and performance of the adsorbents when competitive ions are present. The highest P removal efficiency, independently on whether the adsorbent dosage was low or high, were 85-100% for Phoslock and CFH-12®, 70-100% for Fe-Si-La and 0-15% for FeHQ. The low dosage of FeHQ, compared to previous studies, explained its low P removal efficiency. Although non-magnetic materials were the most efficient, magnetic adsorbents (especially Fe-Si-La) could be proposed for P removal as they can be recovered along with P and be reused, potentially making them more profitable in a long-term period.This work was supported by Junta de Andalucía project P10-RNM-6630 [Proyectos de Excelencia, Spain]; MINECO CTM 2013-46951-R project; and MAT 2016-78778-R project [Spain]. Authors would also like to thank Fernando Ortega for helping in the sampling procedure

La investigación biomédica en España (II). Evaluación del Fondo de Investigación Sanitaria (FIS) a través de los proyectos de investigación financiados en el período 1988-1995 a centros de investigación, facultades y escuelas

En un trabajo previo, publicado en esta misma revista, los autores han estudiado los resultados generados por los proyectos de investigación financiados por el Fondo de Investigación Sanitaria (FIS) a las Instituciones Sanitarias Asistenciales (hospitales) y los procedentes de los cuestionarios de encuestas remitidos a los investigadores responsables de tales proyectos y a los directores gerentes de los hospitales en los que se realizaron los mismos. En el presente trabajo, se continúa el proceso de evaluación del FIS, centrándose la atención en la distribución de los proyectos de investigación concedidos a los centros de investigación, facultades y escuelas, y en los resultados obtenidos a través de los cuestionarios de encuesta cumplimentados por los investigadores principales pertenecientes a estas instituciones o entidades.La realización de este trabajo ha sido posible gracias a la financiación otorgada por el FIS al proyecto de investigación 96/1803.Peer reviewe

Isotropic Image Analysis for Improving CBR Forecasting

A novel hybrid forecasting Case-Based Reasoning (CBR) system is presented in this interdisciplinary study in which an isotropic buffer operator is applied for case-based creation. Commonly used as an image analysis technique by commercial Geographic Information Systems (GIS), the buffer operator in this particular system calculates the area of an oil slick for prediction and visualization tasks. The use of the buffer operator improves the quality of the data used by the system and in consequence the quality of the results obtained. The system generates predictions by using historical data on oil-slick formation following a spill

(OBIFS) isotropic image analysis for improving a predicting agent based systems

In this interdisciplinary study a novel hybrid forecasting system is presented, in which an isotropic buffer operator is applied for case-based creation within the structure of the organization-based multi-agent system. Commonly used as an image analysis technique by commercial Geographic Information Systems (GIS), the buffer operator in this particular system calculates the area of a forest fire for prediction and visualization tasks. The use of the buffer operator improves the quality of the data used by the system and in consequence the quality of the results obtained. The system has been successfully tested using real historical data on forest fires evolution, by generating accurate predictions

Estado de la oferta técnica de servicios de higiene y seguridad industrial, Colombia, 2010

Introduction. Institutions that supply occupational health services must offer services that are reliable and of high quality across the spectrum of industrial hygiene and safety needs.Objective. Services for occupational health were identified at several institutions, and the technical quality and reliability of these services were compared in different regions of Colombia.Materials and methods. This descriptive study identified the services available for industrial hygiene and safety in 15 cities of Colombia. A survey was conducted in 192 institutions offering such services and a statistical analysis of these results was undertaken. This sample was taken from a nationwide list of institutions purportedly licensed for this activity.Results. Thirty-two percent (61) of the evaluated institutions provided hygiene services, and 48% (93) provided safety services. The range of health services was provided on a subcontract basis both for professional personnel and the equipment. Six institutions in the area of industrial hygiene and 1 in the area of industrial security were supplying services with pending or suspended institutional licenses.Conclusion. Deficiencies in the quality, infrastructure and levels of automation were identified at institutions that supply services of hygiene and industrial security. The resulting recommendatios are that the Ministry of the Social Protection fortifies mechanisms for (1) the evaluation and control of the supplied services, and (2) verify that the institutional activity is in accordance with current and valid licensing.Introducción. Las instituciones prestadoras de servicios de prevención de riesgos laborales tienen la función de ofrecer servicios técnicos y confiables en salud ocupacional, seguridad e higiene industrial.Objetivo. Identificar la oferta de servicios existentes respecto a la prevención de riesgos laborales (parte de higiene y seguridad industrial), considerando sus características técnicas y de calidad, en diferentes regiones del país.Materiales y métodos. Corresponde a un estudio descriptivo de corte transversal (2009-2010) de cobertura nacional (15 ciudades) para identificar la oferta de servicios de higiene y seguridad industrial. Se obtuvo un listado nacional de instituciones con licencia para prestar este tipo de servicios, se seleccionó una muestra de 192 instituciones, se aplicó una encuesta y se hizo el análisis estadístico de la información.Resultados. Sesenta y una (31,77 %) de las instituciones evaluadas prestan servicios de higiene y, 93 (48,44 %), de seguridad industrial. El estudio evidenció una oferta de servicios de higiene basada en la subcontratación. Se encontró que 6 (6,52 %) instituciones en el área de higiene industrial y 1 (0,52 %) en el área de seguridad industrial, no contaban con licencia vigente para la prestación del servicio.Conclusión. Se identificaron deficiencias en la calidad, condiciones de infraestructura y tecnificación de las instituciones que ofrecen servicios de higiene y seguridad industrial. Es necesario que el Ministerio de la Protección Social fortalezca los mecanismos para la evaluación y control de los servicios ofrecidos y revise la forma en que hoy se expiden las licencias que autorizan la prestación de estos servicios. DOI: http://dx.doi.org/10.7705/biomedica.v32i1.58

Ghrelin attenuates hepatocellular injury and liver fibrogenesis in rodents and influences fibrosis progression in humans

El pdf del artículo es la versión pre-print.-- et al-There are no effective antifibrotic therapies for patients with liver diseases. We performed an experimental and translational study to investigate whether ghrelin, an orexigenic hormone with pleiotropic properties, modulates liver fibrogenesis. Recombinant ghrelin was administered to rats with chronic (bile duct ligation) and acute (carbon tetrachloride) liver injury. Hepatic gene expression was analyzed by way of microarray analysis and quantitative polymerase chain reaction. The hepatic response to chronic injury was also evaluated in wild-type and ghrelin-deficient mice. Primary human hepatic stellate cells were used to study the effects of ghrelin in vitro. Ghrelin hepatic gene expression and serum levels were assessed in patients with chronic liver diseases. Ghrelin gene polymorphisms were analyzed in patients with chronic hepatitis C. Recombinant ghrelin treatment reduced the fibrogenic response, decreased liver injury and myofibroblast accumulation, and attenuated the altered gene expression profile in bile duct-ligated rats. Moreover, ghrelin reduced the fibrogenic properties of hepatic stellate cells. Ghrelin also protected rats from acute liver injury and reduced the extent of oxidative stress and inflammation. Ghrelin-deficient mice developed exacerbated hepatic fibrosis and liver damage after chronic injury. In patients with chronic liver diseases, ghrelin serum levels decreased in those with advanced fibrosis, and ghrelin gene hepatic expression correlated with expression of fibrogenic genes. In patients with chronic hepatitis C, polymorphisms of the ghrelin gene (994CT and 604GA) influenced the progression of liver fibrosis. Conclusion: Ghrelin exerts antifibrotic effects in the liver and may represent a novel antifibrotic therapy. Copyright © 2010 by the American Association for the Study of Liver Diseases.Supported by grants from the Ministerio de Ciencia e Investigación (SAF2005-06245), from the Instituto de Salud Carlos III (FIS2005-050567, FIS 2008-PI08/0237 and PI070497), and from the European Community FP6 (LSHB-CT-2007-036644 - DIALOK) and by fellowships from Institut d’Investigacions Biomèdiques August Pi i Sunyer (to M. M. and M. D.), the Fundación Bilbao Vizcaya Argentaria (to M. D.) and the Fundació Clínic (to P. S. B.).Peer Reviewe

BRAF activation by metabolic stress promotes glycolysis sensitizing NRASQ61-mutated melanomas to targeted therapy

Glycolysis; Melanomas; Targeted therapyGlucólisis; Melanomas; Terapia dirigidaGlucòlisi; Melanomes; Teràpia dirigidaNRAS-mutated melanoma lacks a specific line of treatment. Metabolic reprogramming is considered a novel target to control cancer; however, NRAS-oncogene contribution to this cancer hallmark is mostly unknown. Here, we show that NRASQ61-mutated melanomas specific metabolic settings mediate cell sensitivity to sorafenib upon metabolic stress. Mechanistically, these cells are dependent on glucose metabolism, in which glucose deprivation promotes a switch from CRAF to BRAF signaling. This scenario contributes to cell survival and sustains glucose metabolism through BRAF-mediated phosphorylation of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-2/3 (PFKFB2/PFKFB3). In turn, this favors the allosteric activation of phosphofructokinase-1 (PFK1), generating a feedback loop that couples glycolytic flux and the RAS signaling pathway. An in vivo treatment of NRASQ61 mutant melanomas, including patient-derived xenografts, with 2-deoxy-D-glucose (2-DG) and sorafenib effectively inhibits tumor growth. Thus, we provide evidence for NRAS-oncogene contributions to metabolic rewiring and a proof-of-principle for the treatment of NRASQ61-mutated melanoma combining metabolic stress (glycolysis inhibitors) and previously approved drugs, such as sorafenib.This work was funded by Instituto de Salud Carlos III and co-funded by European Union (ERDF/ESF, “A way to make Europe”/“Investing in your future”) PI14/0375-Fondos FEDER J.A.R., PI17/00043-Fondos FEDER; J.A.R., PI20/0384-Fondos FEDER; J.A.R., Euronanomed2-ISCIII (AC16/00019)-Fondos FEDER; J.A.R., Asociación Española Contra el Cancer (AECC-GCB15152978SOEN) (supported P.G.M., K.M.); J.A.R., Ramón Areces Foundation (supported K.M. and research); J.A.R. (PI17/00412)-Fondos FEDER; R.B., A.M., A.N.S. We thank A. Zorzano’s laboratory for technical assistance and performance of Seahorse technology

Daylight luminescence system for silicon solar panels based on a bias switching method

Among the many characterization techniques for solar panel testing, two, electroluminescence (EL) and photoluminescence (PL), can provide useful visual information about the presence of different types of cell defects. EL is performed outdoors by night in commercial solar plants due to the very weak luminescence emission compared to sunlight. PL faces the added difficulty of needing to find a large-area homogeneous light source to excite the modules. Since nighttime work poses many drawbacks and risks, a daylight outdoor EL/PL system would be useful for offering safe inspection of solar plants. We present daylight luminescence techniques based on a bias switching method, in which a pulsed luminescence signal is obtained by alternating the polarization state of the solar panels, synchronizing it with the luminescence image detection by an InGaAs camera. Fast switching and selecting an optimized exposure time are key to achieving high-quality images. The daylight luminescence method described herein allows both EL and PL luminescence images to be obtained, even under high solar irradiance conditions.Proyecto de Investigación ENE2014-56069-C4- 4-R (MCIN)Proyecto de Investigación ENE2017-89561-C4-3-R (MCIN)Proyecto de Investigación RTC-2017-6712-3 (MCIN)Proyecto IDI-20151194 (CDTI)Proyecto de Investigación VA081U16 (Junta de Castilla y León)Proyecto de Investigación VA283P18 (Junta de Castilla y León

Alegacion en hecho, y derecho, por don Ioseph Vicente Beluis y Moncada, Marques de Belgida, & c. Con doña Iuana Manuela Mingot, Condesa viuda de Peñalva, madre, y curadora de don Carlos Iuan Torres, Conde de Peñalva; y con don Luis Iuan Torres, tambien

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