Circulación y excreción, funciones esenciales en el proceso de la nutrición. Una propuesta didáctica para una situación de enseñanza no presencial

El presente Trabajo Fin de Máster recoge una descripción y análisis de la propuesta didáctica llevada a cabo durante el Prácticum II/III del Máster Universitario en Profesorado de Educación Secundaria Obligatoria, Bachillerato, Formación Profesional y Enseñanza de Idiomas, Artísticas y Deportivas. Esta propuesta de unidad didáctica aborda los contenidos relativos a los aparatos circulatorio y excretor y su relación con la función de la nutrición. Dado que el Prácticum II/III se realizó en periodo de cierre de los centros escolares debido a la pandemia mundial de Covid-19, esta propuesta didáctica ha sido diseñada para una situación de educación no presencial, aunque todos los recursos y actividades elaborados son fácilmente adaptables a la enseñanza presencial. Para abordar esta unidad didáctica se ha utilizado una metodología flipped classroom o clase invertida, a través de vídeos breves e interactivos elaborados con la plataforma PlayPosit. Las dudas que surgieron de estos vídeos o de las actividades propuestas, se resolvieron a través de vídeos elaborados con el soporte de una presentación PowerPoint y compartidos a través de Youtube con los alumnos. Para completar esta propuesta, se aprovecharon los recursos TICs para elaborar múltiples actividades interactivas entre las que destacan las actividades de indagación en internet y el uso de modelos en 3D de la web Biodigital para profundizar en la fisiología del corazón y el sistema urinario.<br /

Synergistic combinations and new photosensitizers for the enhancement of photodynamic therapy

Tesis doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 11-10-2019Esta tesis tiene embargado el acceso al texto completo hasta el 11-04-2021La terapia fotodinámica (TFD) es una modalidad terapéutica basada en la acción conjunta de tres elementos: i) un agente activable por luz, denominado fotosensibilizador (FS), ii) luz de una longitud de onda adecuada y iii) el oxígeno molecular. Cada uno de estos tres elementos no induce toxicidad de forma individual, pero su actuación conjunta desencadena la producción de especies reactivas de oxígeno (ROS) que dan lugar a la muerte celular. Desde su aprobación clínica en Canadá, en el año 1993, la TFD ha demostrado su eficacia en el tratamiento de múltiples enfermedades relacionadas con altas tasas de proliferación celular y, en especial, en el tratamiento de enfermedades neoplásicas. Sin embargo, en la actualidad, la aplicación clínica de la TFD como tratamiento de primera opción sigue siendo limitada. Por este motivo, existen actualmente numerosas investigaciones centradas en mejorar la eficacia de los tratamientos fotodinámicos para permitir una aplicación más efectiva tanto en forma de monoterapia como en combinación con otras modalidades terapéuticas. En la presente tesis doctoral se han investigado varias estrategias con este objetivo. En primer lugar, se exponen los resultados de un nuevo protocolo de quimiofototerapia con el agente quimioterapéutico doxorrubicina (Dox) y el FS protoporfirina IX (PpIX). Esta combinación se ha explorado aplicando los fármacos libres o vehiculizados en distintas nanoplataformas como liposomas o nanoclusters de oro (AuNCs). Los resultados obtenidos demuestran el elevado potencial de esta combinación, que induce un efecto citotóxico aditivo y, en algunas ocasiones, claramente sinérgico en células tumorales HeLa. Además, el diseño de estos AuNCs permite el control dirigido de la activación de la toxicidad del fármaco Dox, lo que potencialmente reduciría los graves efectos secundarios asociados a este compuesto (p. ej. elevada cardiotoxicidad). Asimismo, se ha explorado el efecto de la combinación de dos FSs en un mismo tratamiento fotodinámico, obteniendo excelentes resultados. La TFD combinada con los FS ZnPc (ftalocianina de Zinc) y TMPyP (meso‐tetra(4-N-metilpiridil)porfina) tiene un elevado efecto tóxico que induce una elevada muerte celular incluso con concentraciones de fármacos y dosis de luz muy reducidas. Este tratamiento ha demostrado también ser eficaz en la inactivación de células tumorales procedentes de biopsias de pacientes y en cultivos en 3D de estas mismas células, que recrean con más precisión las condiciones de un tumor in vivo. Finalmente, se ha analizado el potencial de varios compuestos de la familia BODIPY (4,4-difluoro-4 bora-3a,4a-diaza-s-indaceno) como nuevos FSs. Ha quedado demostrado que estos compuestos no sólo cuentan con propiedades óptimas para su aplicación en TFD, sino que además son excelentes sondas fluorescentes para distintos orgánulos celulares, lo que los convierten en candidatos ideales para aplicaciones teragnósticas.Photodynamic therapy (PDT) is a minimally invasive and clinically approved procedure based on the synergistic action of three elements: i) a photoactivatable agent, named photosensitizer (PS), ii) light of a specific wavelength, and iii) molecular oxygen. These three elements are not toxic by themselves, but its combination triggers a toxic effect based on the generation of reactive oxygen species (ROS). Since its clinical approval in Canada, in 1993, PDT has proven to be an effective therapy for the treatment of several diseases associated with uncontrolled cell proliferation, especially cancer. However, by now PDT has not been developed enough to become a first-choice treatment except in a very limited number of cases. In this sense, lots of efforts are being focused on the improvement of PDT in order to enhance the clinical application of this therapy (alone or in combination with other therapeutic modalities, such as radiotherapy or chemotherapy). The main objective of this PhD thesis was to evaluate different strategies that potentially would be able to improve PDT effectiveness. First, we have evaluated the effect of a combined protocol involving the chemotherapeutic agent doxorrubicin (Dox) and the PS protoporphyrin IX (PpIX). To explore in depth this combination therapy, we have attached both drugs in two different nanoparticles: liposomes and gold nanoclusters (AuNCs). Our results prove the potential of this combined protocol, which induces an additive or synergistic effect in the inactivation of tumor HeLa cells. Furthermore, attachment of drug to AuNC allows the spatio-temporal release of the chemotherapeutic agent Dox, which could contribute to reduce unwanted side effects usually related with this drug (e.g. high cardiotoxicity). Moreover, this work analyzes the effect of a new protocol that combines two PSs in a unique PDT treatment: Zinc phthalocyanine (ZnPc) and meso-tetrakis(4-N-methylpyridyl)porphyrin (TMPyP) . Our results show that this combined PDT is able to induce an extreme cytotoxic effect on tumor cell lines by using very low drug concentration and light dose. The treatment has been proven highly effective even with patient-derived tumor cells and 3D cultures, which recreates much more precisely in vivo tumor conditions that conventional cell lines or 2D cultures. Finally, several BODIPYs (4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) derived compounds had been analyzed to determine their potential as new PDT agents. We have demonstrated that these compounds have excellent biological and phototoxic properties for its application as new PSs. Moreover, thanks to its strong fluorescence emission, these BODIPYs derivatives are optimal fluorescent probes for several cellular organelles, which could be highly useful for theragnostic applicationsLa realización de la presente Tesis Doctoral ha sido posible gracias a la financiación de los siguientes organismos: MINECO (CTQ2013-48767-C3-3-R). MINECO (CTQ2016-78454-C2-2-R). UNIVERSIDAD AUTÓNOMA DE MADRID. Contratos predoctorales Formación de Personal Investigador (FPI-UAM

AcetylacetonateBODIPY-Biscyclometalated Iridium(III) Complexes: Effective Strategy towards Smarter Fluorescent Photosensitizer Agents

Biscyclometalated IrIII complexes involving boron-dipyrromethene (BODIPY)-based ancillary ligands, where the BODIPY unit is grafted to different chelating cores (acetylacetonate for Ir-1 and Ir-2, and bipyridine for Ir-3) by the BODIPY meso position, have been synthesized and characterized. Complexes with the BODIPY moiety directly grafted to acetylacetonate (Ir-1 and Ir-2) exhibit higher absorption coefficients (ϵ≈4.46×104 m−1 cm−1 and 3.38×104 m−1 cm−1 at 517 nm and 594 nm, respectively), higher moderate fluorescence emission (φfl≈0.08 and 0.22 at 528 nm and 652 nm, respectively) and, in particular, more efficient singlet oxygen generation upon visible-light irradiation (φΔ≈0.86 and 0.59, respectively) than that exhibited by Ir-3 (φΔ≈0.51, but only under UV light). Phosphorescence emission, nanosecond time-resolved transient absorption, and DFT calculations suggest that BODIPY-localized long-lived 3IL states are populated for Ir-1 and Ir-2. In vitro photodynamic therapy (PDT) activity studied for Ir-1 and Ir-2 in HeLa cells shows that such complexes are efficiently internalized into the cells, exhibiting low dark- and high photocytoxicity, even at significantly low complex concentration, making them potentially suitable as theranostic agents. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinhei

In-situ particles reorientation during magnetic hyperthermia application: Shape matters twice

Promising advances in nanomedicine such as magnetic hyperthermia rely on a precise control of the nanoparticle performance in the cellular environment. This constitutes a huge research challenge due to difficulties for achieving a remote control within the human body. Here we report on the significant double role of the shape of ellipsoidal magnetic nanoparticles (nanorods) subjected to an external AC magnetic field: first, the heat release is increased due to the additional shape anisotropy; second, the rods dynamically reorientate in the orthogonal direction to the AC field direction. Importantly, the heating performance and the directional orientation occur in synergy and can be easily controlled by changing the AC field treatment duration, thus opening the pathway to combined hyperthermic/mechanical nanoactuators for biomedicine. Preliminary studies demonstrate the high accumulation of nanorods into HeLa cells whereas viability analysis supports their low toxicity and the absence of apoptotic or necrotic cell death after 24 or 48 h of incubation.Ministerio de Economía, Comercio y Empresa (España)Comunidad de MadridEC FP-7 grant “NanoMag”Xunta de GaliciaUniversidad Autónoma de MadridDepto. de Química en Ciencias FarmacéuticasFac. de FarmaciaTRUEpu

Halogen-free photosensitizers based on meso-enamine-BODIPYs for bioimaging and photodynamic therapy

The search for efficient heavy atom free photosensitizers (PSs) for photodynamic therapy (PDT) is a very active field. We describe herein a simple and easily accessible molecular design based on the attachment of an enamine group as an electron-donor moiety at the meso position of the BODIPY core with different alkylation patterns. The effect of the alkylation degree and solvent polarity on the photophysical properties in terms of splitting absorption bands, fluorescence efficiencies and singlet oxygen production is analyzed in depth experimentally using spectroscopic techniques, including femtosecond and nanosecond transient absorption (fs- and ns-TA) and using computational simulations based on time-dependent density functional theory. The correlation between the theoretical/experimental results permits the rationalization of the observed photophysical behavior exhibited by meso-enamine-BODIPY compounds and the determination of mechanistic details, which rule the population of the triplet state manifold. The potential applicability as a theragnostic agent for the most promising compound is demonstrated through in vitro assays in HeLa cells by analyzing the internalization, localization and phototoxic action

Tailor-made PEG coated iron oxide nanoparticles as contrast agents for long lasting magnetic resonance molecular imaging of solid cancers

Magnetic resonance imaging (MRI) is the most powerful technique for non-invasive diagnosis of human diseases and disorders. Properly designed contrast agents can be accumulated in the damaged zone and be internalized by cells, becoming interesting cellular MRI probes for disease tracking and monitoring. However, this approach is sometimes limited by the relaxation rates of contrast agents currently in clinical use, which show neither optimal pharmacokinetic parameters nor toxicity. In this work, a suitable contrast agent candidate, based on iron oxide nanoparticles (IONPs) coated with polyethyleneglycol, was finely designed, prepared and fully characterized under a physical, chemical and biological point of view. To stand out the real potential of our study, all the experiments were performed in comparison with Ferumoxytol, a FDA approved IONPs. IONPs with a core size of 15 nm and coated with polyethyleneglycol of 5 kDa (OD15-P5) resulted the best ones, being able to be uptaken by both tumoral cells and macrophages and showing no toxicity for in vitro and in vivo experiments. In vitro and in vivo MRI results for OD15-P5 showed r2 relaxivity values higher than Ferumoxitol. Furthermore, the injected OD15-P5 were completely retained at the tumor site for up to 24 h showing high potential as MRI contrast agents for real time long-lasting monitoring of the tumor evolution.European Seventh Framework ProgrammeMinisterio de Economía, Comercio y Empresa (España)Universidad Autónoma de MadridPro-CNIC FoundationCentro Nacional de Investigaciones CardiovascularesCOST Action CA1520Comunidad de MadridUniversidad Complutense de MadridFondo Europeo de Desarrollo RegionalDepto. de Química en Ciencias FarmacéuticasFac. de FarmaciaTRUEpu

A versatile fluorescent molecular probe endowed with singlet oxygen generation under white-light photosensitization

Despite fluorescent photodynamic therapy (fluorescent-PDT) dyes are promising theranostic agents, current approaches unfortunately involve crucial shortcomings (such as, narrow absorption bands, high cost, low bio-compatibility and specificity, low dual efficiency) making difficult their clinical translation. Particularly, efficient fluorescent-PDT agents triggered under white-light, with potential application in topic solar treatments, are scarce. Here, we describe the rational development of a novel fluorescent-PDT molecular biomaterial based on BODIPY building blocks able to sustain, simultaneously, synthetic accessibility, high fluorescence and phototoxicity within a broad spectral window, biocompatibility, including low dark toxicity and high cell permeability with selective accumulation in lysosomes and, what is more important, excellent efficient activity triggered under white light. These all-in-one combined properties make the new dye a valuable ground platform for the development of future smarter theranostic agents