Malignant infarction of the middle cerebral artery in a porcine model. A pilot study

Animal models; Central nervous system; InfarctionModels animals; Sistema nerviós central; InfartModelos animales; Sistema nervioso central; InfartoBackground and purpose Interspecies variability and poor clinical translation from rodent studies indicate that large gyrencephalic animal stroke models are urgently needed. We present a proof-of-principle study describing an alternative animal model of malignant infarction of the middle cerebral artery (MCA) in the common pig and illustrate some of its potential applications. We report on metabolic patterns, ionic profile, brain partial pressure of oxygen (PtiO2), expression of sulfonylurea receptor 1 (SUR1), and the transient receptor potential melastatin 4 (TRPM4). Methods A 5-hour ischemic infarct of the MCA territory was performed in 5 2.5-to-3-month-old female hybrid pigs (Large White x Landrace) using a frontotemporal approach. The core and penumbra areas were intraoperatively monitored to determine the metabolic and ionic profiles. To determine the infarct volume, 2,3,5-triphenyltetrazolium chloride staining and immunohistochemistry analysis was performed to determine SUR1 and TRPM4 expression. Results PtiO2 monitoring showed an abrupt reduction in values close to 0 mmHg after MCA occlusion in the core area. Hourly cerebral microdialysis showed that the infarcted tissue was characterized by reduced concentrations of glucose (0.03 mM) and pyruvate (0.003 mM) and increases in lactate levels (8.87mM), lactate-pyruvate ratio (4202), glycerol levels (588 μM), and potassium concentration (27.9 mmol/L). Immunohistochemical analysis showed increased expression of SUR1-TRPM4 channels. Conclusions The aim of the present proof-of-principle study was to document the feasibility of a large animal model of malignant MCA infarction by performing transcranial occlusion of the MCA in the common pig, as an alternative to lisencephalic animals. This model may be useful for detailed studies of cerebral ischemia mechanisms and the development of neuroprotective strategies.The Neurotraumatology and Neurosurgery Research Unit is supported by a grant from the Departament d'Universitats, Recerca i Societat de la Informació de la Generalitat de Catalunya (SGR 2014-844). This work has been supported in part by the Fondo de Investigación Sanitaria (Instituto de Salud Carlos III) with grant FIS PI11/00700, which was co-financed by the European Regional Development Fund (ERDF) and awarded to Dr. J. Sahuquillo. A. Sánchez-Guerrero is the recipient of a personal pre-doctoral grant from the Instituto de Salud Carlos III (grant number grant number FI12/00074)

La grabación sonora: un recurso pedagógico multidisciplinar para la reinterpretación de la Historia

Mediante el presente proyecto se ha pretendido abordar un nuevo acercamiento a la historia de la música, en tanto manifestación cultural, basado en una metodología de reciente creación que entronca directamente con los estudios sobre la praxis interpretativa. Para ello hemos profundizado en el correcto manejo de los software informáticos que permiten el análisis de estas fuentes sonoras (fundamentalmente visualizadores de ondas y editores de sonido). Como resultado, hemos constituido un grupo de trabajo abierto a profesores y alumnos, ubicado físicamente en la Facultad de Geografía e Historia de la UCM

La influencia de la tecnología en los procesos de producción musical: análisis de la grabación sonora y sus aplicaciones en el ámbito docente

Memoria de las actividades del proyecto de Innova-Docencia nº256, coordinado por el Dr. Marco Antonio Juan de Dios Cuartas, profesor del Departamento de Musicología de la Universidad Complutense de Madrid

Impact of interstitial lung disease on the survival of systemic sclerosis with pulmonary arterial hypertension

To assess severity markers and outcomes of patients with systemic sclerosis (SSc) with or without pulmonary arterial hypertension (PAH-SSc/non-PAH-SSc), and the impact of interstitial lung disease (ILD) on PAH-SSc. Non-PAH-SSc patients from the Spanish SSc registry and PAH-SSc patients from the Spanish PAH registry were included. A total of 364 PAH-SSc and 1589 non-PAH-SSc patients were included. PAH-SSc patients had worse NYHA-functional class (NYHA-FC), worse forced vital capacity (FVC) (81.2 +/- 20.6% vs 93.6 +/- 20.6%, P < 0.001), worse tricuspid annular plane systolic excursion (TAPSE) (17.4 +/- 5.2 mm vs 19.9 +/- 6.7 mm, P < 0.001), higher incidence of pericardial effusion (30% vs 5.2%, P < 0.001) and similar prevalence of ILD (41.8% vs. 44.9%). In individuals with PAH-SSc, ILD was associated with worse hemodynamics and pulmonary function tests (PFT). Up-front combination therapy was used in 59.8% and 61.7% of patients with and without ILD, respectively. Five-year transplant-free survival rate was 41.1% in PAH-SSc patients and 93.9% in non-PAH-SSc patients (P < 0.001). Global survival of PAH-SSc patients was not affected by ILD regardless its severity. The multivariate survival analysis in PAH-SSc patients confirmed age at diagnosis, worse NYHA-FC, increased PVR, reduced DLCO, and lower management with up-front combination therapy as major risk factors. In conclusion, in PAH-SSc cohort risk of death was greatly increased by clinical, PFT, and hemodynamic factors, whereas it was decreased by up-front combination therapy. Concomitant ILD worsened hemodynamics and PFT in PAH-SSc but not survival regardless of FVC impairment

Impact of interstitial lung disease on the survival of systemic sclerosis with pulmonary arterial hypertension

To assess severity markers and outcomes of patients with systemic sclerosis (SSc) with or without pulmonary arterial hypertension (PAH-SSc/non-PAH-SSc), and the impact of interstitial lung disease (ILD) on PAH-SSc. Non-PAH-SSc patients from the Spanish SSc registry and PAH-SSc patients from the Spanish PAH registry were included. A total of 364 PAH-SSc and 1589 non-PAH-SSc patients were included. PAH-SSc patients had worse NYHA-functional class (NYHA-FC), worse forced vital capacity (FVC) (81.2 ± 20.6% vs 93.6 ± 20.6%, P &lt; 0.001), worse tricuspid annular plane systolic excursion (TAPSE) (17.4 ± 5.2 mm vs 19.9 ± 6.7 mm, P &lt; 0.001), higher incidence of pericardial effusion (30% vs 5.2%, P &lt; 0.001) and similar prevalence of ILD (41.8% vs. 44.9%). In individuals with PAH-SSc, ILD was associated with worse hemodynamics and pulmonary function tests (PFT). Up-front combination therapy was used in 59.8% and 61.7% of patients with and without ILD, respectively. Five-year transplant-free survival rate was 41.1% in PAH-SSc patients and 93.9% in non-PAH-SSc patients (P &lt; 0.001). Global survival of PAH-SSc patients was not affected by ILD regardless its severity. The multivariate survival analysis in PAH-SSc patients confirmed age at diagnosis, worse NYHA-FC, increased PVR, reduced DLCO, and lower management with up-front combination therapy as major risk factors. In conclusion, in PAH-SSc cohort risk of death was greatly increased by clinical, PFT, and hemodynamic factors, whereas it was decreased by up-front combination therapy. Concomitant ILD worsened hemodynamics and PFT in PAH-SSc but not survival regardless of FVC impairment

El laboratorio sonoro como herramienta docente para el análisis de la creación e interpretación musical a través de las grabaciones

Depto. de MusicologíaFac. de Geografía e HistoriaFALSEsubmitte

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. For example, a key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process versus those that measure fl ux through the autophagy pathway (i.e., the complete process including the amount and rate of cargo sequestered and degraded). In particular, a block in macroautophagy that results in autophagosome accumulation must be differentiated from stimuli that increase autophagic activity, defi ned as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (inmost higher eukaryotes and some protists such as Dictyostelium ) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the fi eld understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. It is worth emphasizing here that lysosomal digestion is a stage of autophagy and evaluating its competence is a crucial part of the evaluation of autophagic flux, or complete autophagy. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation it is imperative to delete or knock down more than one autophagy-related gene. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways so not all Atg proteins can be used as a specific marker for an autophagic process. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field

Educafarma 10.0

Memoria ID-030. Ayudas de la Universidad de Salamanca para la innovación docente, curso 2021-2022

Aplicación de la microdiálisis en la definición del perfil iónico del edema cerebral postraumático e isquémico. estudio de la contribución del canal SUR1-TRPM4 en su formación

Los traumatismos craneoenfálicos (TCE) y los infartos malignos de la arteria cerebral media (IMACM) provocan importantes desequilibrios iónicos, generando una entrada masiva de iones en las células, con el consecuente arrastre osmótico de agua y la formación de edema cerebral. La alteración de las concentraciones iónicas induce la acumulación de agua en el espacio intra y extracelular (EEC), causando hinchazón del tejido cerebral lesionado y deterioro clínico de estos pacientes. Estos trastornos iónicos están directamente relacionados con la sobreexpresión de distintos canales iónicos, constitutivos o de nueva expresión como el canal formado por el receptor de la sulfonilurea 1 (SUR1) y el receptor de potencial transitorio de la melastatina 4 (TRPM4) conocido como SUR1-TRPM4. Las técnicas actuales de neuromonitorización permiten el estudio de los procesos que tienen lugar tras una lesión cerebral aguda. Entre ellos, la microdiálisis (MD) cerebral es una técnica de neuromonitorización avanzada que permite el muestreo continuo del EEC. Los objetivos principales de esta tesis fueron: 1) estudiar la expresión del SUR1 en distintos tipos celulares (neuronas, células endoteliales, astrocitos, microglía reactiva/macrófagos y neutrófilos) en el tejido cerebral pericontusional determinado, además, si su grado de expresión estaba relacionado con el tiempo postraumatismo, 2) determinar la recuperación relativa (RR) in vitro de Na+, K+, Cl-, albúmina y cistatina C con membranas de MD de 100 kDa (CMA71) a una velocidad de perfusión de 0,30 µL/min, 3) estimar el perfil iónico y molecular del EEC cerebral en las distintas áreas cerebrales en las que se insertó el catéter de MD en pacientes con lesiones cerebrales agudas y 4) definir la aparición de edema vasogénico a partir de la detección de albúmina y cistatina C en el EEC del encéfalo. En primer lugar, para el estudio de la expresión del SUR1 en el tejido pericontusional, se analizó un grupo de 26 muestras de contusiones cerebrales de pacientes tratados quirúrgicamente y 3 controles de tejido cerebral normal mediante inmunohistoquímica de fluorescencia. Los resultados objetivaron que el SUR1 se sobreexpresaba en todos los tipos celulares cerebrales estudiados. Sin embargo, el patrón temporal de expresión del SUR1 era distinto para cada tipo celular. En segundo lugar, para el cálculo de la RR de iones se llevaron a cabo 9 experimentos in vitro con catéteres de MD CMA-71 aplicando las condiciones estándar utilizadas en un entorno clínico, determinándose las concentraciones iónicas, tanto en el dializado y como de la matriz. Un modelo de regresión lineal permitió calcular las concentraciones iónicas reales en la matriz. Por último, para el análisis de perfil iónico del EEC cerebral se analizaron los microdializados de 34 pacientes con un TCE o IMACM que recibieron monitorización multimodal durante las primeras 48 horas tras el TCE o del inicio de los síntomas del IMACM. Se determinó la concentración iónica en 12 muestras por paciente mediante espectrometría de masas con plasma acoplado inductivamente, considerando que el perfil iónico de los pacientes depende de la zona de inserción del catéter de MD. Los resultados mostraron que los niveles de K+ en el EEC guardan relación con la viabilidad del tejido, mientras que los Na+ con la permeabilidad de barrera hematoencefálica. La MD ofrece la posibilidad única de observar las transformaciones dinámicas de los iones en el cerebro a través del tiempo y abre una nueva vía para explorar el perfil iónico del cerebro, sus cambios en el edema cerebral y cómo este perfil se puede modificar con diferentes terapias. Definir el perfil molecular del edema puede servir como índice cuantitativo de progresión del edema, permitiendo la modulación farmacológica de sus distintas etapas y potenciales tratamientos dirigidos al bloqueo selectivo de SUR1.Traumatic brain injury (TBI) and malignant middle cerebral artery infarction (MMCAI) cause significant ionic imbalances, generating massive ionic fluxes with the consequent osmotic water movement across the cells and the cerebral edema formation. Changes in ionic concentrations induce water accumulation in the intracellular and extracellular space and cause swelling of the injured brain tissue and neurological worsening in patients. These ionic disorders are directly related to the overexpression of different ion channels, that can be constitutive or newly synthesized as the channel formed by the sulfonylurea receptor 1 (SUR1) and transient receptor potential of melastatin 4 (TRPM4) known as SUR1-TRPM4. Current neuromonitorization techniques allow the study of the processes occurring after acute brain injury. Cerebral microdialysis (MD) is an advanced neuromonitorization technique that allows continuous sampling of the cerebral parenchyma. The main objectives of this thesis were: 1) to study SUR1 expression in different cell types (neurons, endothelial cells, astrocytes, reactive microglia/macrophages and neutrophils) in pericontusional brain tissue and whether SUR1 up-regulation was related to time post injury, 2) to calculate the in vitro relative recovery (RR) of Na+, K+, Cl-, albumin and cystatin C with membranes of 100 kDa MD (CMA71) at an infusion rate of 0.30 µL/min, 3) to determine the ionic and molecular profile of brain extracellular space in different brain areas in which MD catheter was inserted in patients with acute brain injury and 4) to define the development of vasogenic edema by the albumin and cystatin C detection in the brain extracellular space. First, to study SUR1 expression in pericontusional tissue, 26 samples obtained from brain-injured patients who underwent surgical treatment and 3 normal brain tissue samples as a control were analyzed using inmunofluorescence. The results showed that SUR1 is overexpressed in all studied cell types. However, the temporal pattern of SUR1 expression was found to be different in each cell type. Secondly, to calculate the RR of ions, 9 experiments were performed in vitro with MD CMA-71 catheters under standard conditions for clinical settings. Ionic concentrations were determined in both the dialysate and the matrix. A linear regression model was used to calculate the true ionic concentrations in the matrix. Finally, to analyze the ionic profile of brain extracellular space, microdialysate samples of 34 patients were analyzed. The patient group had presented with moderate or severe TBI or MMCAI, and received multimodal monitoring during the first 48 hours after injury or start of symptoms. Ionic concentrations were determined in 12 samples for each patient using inductively coupled plasma mass spectrometry. Ionic profile was studied according to where the MD catheter was inserted. The results showed that K+ levels in the extracellular space were related to tissue viability, while the Na+ concentration was related to blood-brain barrier permeability. MD offers a unique opportunity to observe the dynamic transformations of ions in the brain over time and opens up a new path to explore the ionic profile of the brain in terms of how it changes during brain edema and how it may be modified through different therapies. A definition of the molecular profile in brain edema can be used as a quantitative index of edema progression, which may allow different stages to be managed pharmacologically and potential treatments directed at selectively blocking SUR1 to be developed

Open-Label Study to Evaluate the Efficacy of a Topical Anhydrous Formulation with 15% Pure Ascorbic Acid and Ginger as a Potent Antioxidant

Vitamin C is one of the naturally occurring antioxidants capable of reducing or preventing skin photoaging. Achieving a stable formulation with the optimal dose of ascorbic acid to ensure a biologically significant antioxidant effect is a challenge when developing cosmetic formulations. The objective of this study was to develop a stable formula in a non-aqueous media with 15% pure vitamin C supplemented with ginger and to study its efficacy, skin tolerance, and cosmetic assessment in 33 women. Vitamin C stability over time was determined via a high-performance liquid chromatography (HPLC) technique versus an aqueous option. Reactive oxygen species (ROS) determination was quantified to provide antioxidant effect. A 56-day in vivo study was performed to evaluate skin luminosity and hyperpigmentation reduction. Skin acceptability was verified by a dermatologist. The HPLC studies demonstrated a high stability of the anhydrous formula compared to an aqueous option. The in vitro studies showed a reduction in ROS of 93% (p-value p-value p-value < 0.007). Moreover, very good skin tolerance was determined as the dermatologist did not determine any clinical signs, and the subjects did not report any feelings of discomfort. We were able to develop an anhydrous formula of pure vitamin C that combines very good stability, consumer acceptance, and skin tolerance with a high level of efficacy