Sirtuin 1 activation as a new therapeutic target for age-related macular degeneration

Motivation: Age-related macular degeneration (AMD) is a retinal degenerative disease that affects the macula which is the central part of the retina. The macula is enriched in cone-photoreceptors and is responsible for color and fine detail vision. AMD is the major cause of blindness in adults in the developed world. In the search for new treatments, numerous lines of evidence suggest that dietary polyphenols such as resveratrol have the capacity to mitigate age-associated diseases due to their antioxidant capacity, their ability to activate the antioxidant defenses, and their anti-inflammatory capacity. These effects are partly due to activation of deacetylase enzymes like sirtuin 1 (SIRT1). The problem is that bioavailability of resveratrol is low considering its poorly solubility and high metabolic rate. In addition, resveratrol is toxic at high doses. Because of this, previous work of our group had synthetized resveratrol derivatives that improve its characteristics and maintain or improve its therapeutic effect. These compounds have been tested in retinal degenerative animal models where they have shown neuroprotective effect. In the present work, we aim to study the mechanism of action of these compounds in a cellular model and the implication of SIRT1 in order these derivatives serve as future treatment for AMD.Methods: All the analysis has been carried out on a human retinal pigment epithelial cell line (ARPE-19). Cells have been treated with different concentrations of resveratrol and its derivatives. The treated cells have been subjected to toxicity assays and have also been analyzed using molecular biology techniques such as quantitative PCR, immunofluorescence and western blot. In particular, it is studied whether the mechanism of action is due to an increase in expression or activity of SIRT1 at different concentrations.Results and conclusions: Up to now, it has been observed that the different derivatives are not toxic at low concentrations but at high concentrations. In addition, they are capable of increase the activity of SIRT1 dependent on concentration, as well as the mechanism of action may be mediated by the activation of HSP70, a heat shock activated chaperone whose transcription factor is deacetylated and activated by SIRT1. Later, these derivatives will be tested in a cellular model of protein aggregation, mimicking what occurs in the retinal epithelium with a mutation that causes macular degeneration

Resveratrol derivates for age-related macular degeneration treatment

Motivation: Age-related macular degeneration (AMD) is a condition characterized by the acumulation of deposits called drusen between the retinal pigment epitheliun (RPE) and Bruch's membrane. As the disease progresses, occurs the degeneration of RPE and photoreceptors cells (dry AMD) or the infiltration of blood vessels from the choroidal vasculature into the retina (wet AMD). To date AMD has no effective treatment. This condition is the leading cause of blindness among elderly individuals worldwide and the total number of patients is expected to increase to 288 million affected individuals in 2040. Resveratrol has shown to have protective effects in others ophthalmologic disorders as diabetic retinopathy, likely through upregulation of Sirt1. The aim of this work is to test some small molecules derivated from resveratrol to see if there is a protective effect against the progression of AMD. Methods: The compounds were tested in vitro using the RPE cell line ARPE-19. Posible toxic effects were studied using RealTime-Glo™ MT Cell Viability Assay and RealTime-Glo™ Annexin V Apoptosis and Necrosis Assay. The efects in the cell cycle were annalised by flow cytometry. Changes in Sirt1 enzymatic activity were messured using SIRT-Glo™ Assay System and Sirt1 expression has been annalised by inmunofluorescence (IF) and western blot (WB) Results: The compounds did not show significant toxic effects or effects for the cell cycle in the concentration range annalised except resveratrol, that seems to cause cell cycle arrest at S and G2/M phase and cell apoptosis. Sirt1 enzymatic activity and Sirt1 expression in the cells do not seem to be increased in response to resveratrol derivates treatment at high doses compared to vehicle control. Conclusions: Resveratrol derivates seem to be safer and less toxic than resveratrol, at least for in vitro use. Further studies are needed to determinate their possible therapeutic applications

Neuromuscular disease classification system

Diagnosis of neuromuscular diseases is based on subjective visual assessment of biopsies from patients by the pathologist specialist. A system for objective analysis and classification of muscular dystrophies and neurogenic atrophies through muscle biopsy images of fluorescence microscopy is presented. The procedure starts with an accurate segmentation of the muscle fibers using mathematical morphology and a watershed transform. A feature extraction step is carried out in two parts: 24 features that pathologists take into account to diagnose the diseases and 58 structural features that the human eye cannot see, based on the assumption that the biopsy is considered as a graph, where the nodes are represented by each fiber, and two nodes are connected if two fibers are adjacent. A feature selection using sequential forward selection and sequential backward selection methods, a classification using a Fuzzy ARTMAP neural network, and a study of grading the severity are performed on these two sets of features. A database consisting of 91 images was used: 71 images for the training step and 20 as the test. A classification error of 0% was obtained. It is concluded that the addition of features undetectable by the human visual inspection improves the categorization of atrophic pattern

Retinal pigment epithelium degeneration caused by aggregation of PRPF31 and the role of HSP70 family of proteins

Background Mutations in pre-mRNA splicing factor PRPF31 can lead to retinitis pigmentosa (RP). Although the exact disease mechanism remains unknown, it has been hypothesized that haploinsufficiency might be involved in the pathophysiology of the disease. Methods In this study, we have analyzed a mouse model containing the p.A216P mutation in Prpf31 gene. Results We found that mutant Prpf31 protein produces cytoplasmic aggregates in the retinal pigment epithelium and decreasing the protein levels of this splicing factor in the nucleus. Additionally, normal protein was recruited in insoluble aggregates when the mutant protein was overexpressed in vitro. In response to protein aggregation, Hspa4l is overexpressed. This member of the HSP70 family of chaperones might contribute to the correct folding and solubilization of the mutant protein, allowing its translocation to the nucleus. Conclusions Our data suggests that a mechanism haploinsufficiency and dominant-negative is involved in retinal degeneration due to mutations in PRPF31. HSP70 over-expression might be a new therapeutic target for the treatment of retinal degeneration due to PRPF31 mutations.This project has been financed through a) The ISCIII (Miguel Servet-I, 2015), co-financed by the European Regional Development Fund (ERDF), No CP15/00071. b) The European Union’s Horizon 2020 research and innovation program, under grant agreement No 634479. c) Regional Ministry of Economy, Innovation and Science of the Junta de Andalucía, No P09-CTS-04967.info:eu-repo/semantics/publishedVersio

Quantifiable diagnosis of muscular dystrophies and neurogenic atrophies through network analysis

[Background] The diagnosis of neuromuscular diseases is strongly based on the histological characterization of muscle biopsies. However, this morphological analysis is mostly a subjective process and difficult to quantify. We have tested if network science can provide a novel framework to extract useful information from muscle biopsies, developing a novel method that analyzes muscle samples in an objective, automated, fast and precise manner.[Methods] Our database consisted of 102 muscle biopsy images from 70 individuals (including controls, patients with neurogenic atrophies and patients with muscular dystrophies). We used this to develop a new method, Neuromuscular DIseases Computerized Image Analysis (NDICIA), that uses network science analysis to capture the defining signature of muscle biopsy images. NDICIA characterizes muscle tissues by representing each image as a network, with fibers serving as nodes and fiber contacts as links.[Results] After a ‘training’ phase with control and pathological biopsies, NDICIA was able to quantify the degree of pathology of each sample. We validated our method by comparing NDICIA quantification of the severity of muscular dystrophies with a pathologist’s evaluation of the degree of pathology, resulting in a strong correlation (R = 0.900, P <0.00001). Importantly, our approach can be used to quantify new images without the need for prior ‘training’. Therefore, we show that network science analysis captures the useful information contained in muscle biopsies, helping the diagnosis of muscular dystrophies and neurogenic atrophies.[Conclusions] Our novel network analysis approach will serve as a valuable tool for assessing the etiology of muscular dystrophies or neurogenic atrophies, and has the potential to quantify treatment outcomes in preclinical and clinical trials.LME and AM were supported by the Miguel Servet (Instituto Carlos III) program that also funded the work. LME is funded by the Spanish Ministry of Science (BFU2011-25734), and AP is funded by the Spanish Ministry of Science and the Andalusian Government.LME and AM were supported by the Miguel Servet (Instituto Carlos III) program that also funded the work. LME is funded by the Spanish Ministry of Science (BFU2011-25734), and AP is funded by the Spanish Ministry of Science and the Andalusian Government. AS is funded by the Consejería de Innovación, Ciencia y Empresa of the Junta de Andalucía.Peer Reviewe

Método para obtener información útil para el diagnóstico de enfermedades neuromusculares

Procedimiento para obtener, a partir de la biopsia de un paciente, nuevos parámetros que permiten diagnosticar de manera objetiva diferentes enfermedades neuromusculares y su grado de afectación al paciente, que comprende: - realizar una tinción de la biopsia para resaltar las fibras musculares tipo I, las fibras musculares tipo II y el endomisio; - obtener una imagen de la biopsia tras la tinción; - segmentar la imagen para identificar los contornos de las fibras musculares; - formar una red donde las fibras musculares constituyen nodos y los contactos entre fibras musculares constituyen las uniones entre los nodos; - formar un vector característico de la biopsia cuyos elementos se eligen entre parámetros geométricos de las fibras y parámetros de la red construida; y - comparar biopsias control y afectas por medio de ACP utilizando en cada caso el vector característico seleccionadoPeer reviewedServicio Andaluz de Salud, Universidad de Sevilla, Consejo Superior de Investigaciones CientíficasB1 Patente sin examen previ

Método para obtener información útil para el diagnóstico de enfermedades neuromusculares

Método para obtener información útil para el diagnóstico de enfermedades neuromusculares. Procedimiento para obtener, a partir de la biopsia de un paciente, nuevos parámetros que permiten diagnosticar de manera objetiva diferentes enfermedades neuromusculares y su grado de afectación al paciente, que comprende: realizar una tinción de la biopsia para resaltar las fibras musculares tipo I, las fibras musculares tipo II y el endomisio; obtener una imagen de la biopsia tras la tinción; segmentar la imagen para identificar los contornos de las fibras musculares; formar una red donde las fibras musculares constituyen nodos y los contactos entre fibras musculares constituyen las uniones entre los nodos; formar un vector característico de la biopsia cuyos elementos se eligen entre parámetros geométricos de las fibras y parámetros de la red construida; y comparar biopsias control y afectas por medio de ACP utilizando en cada caso el vector característico seleccionado.Peer reviewedServicio Andaluz de Salud, Universidad de Sevilla, Consejo Superior de Investigaciones Científicas (España)A1 Solicitud de patente con informe sobre el estado de la técnic

Compuestos acilados para el tratamiento de patologías oculares

La presente invención se refiere al uso terapéutico de compuestos derivados piceido acilados en patologías oculares, en particular retinitis pigmentosa y en degeneración macular asociada a la edad, entre otras.Peer reviewedConsejo Superior de Investigaciones Científicas (España), Fundación Pública Andaluza Progreso y SaludB1 Patente sin examen previ

Compuestos acilados para el tratamiento de patologías oculares

The present invention relates to the therapeutic use of acylated piceid derivative compounds in ocular pathologies, in particular retinitis pigmentosa and in age-related macular degeneration, ínter afiaPeer reviewedConsejo Superior de Investigaciones Científicas (España), Fundación Pública Andaluza Progreso y SaludA1 Solicitud de patente con informe sobre el estado de la técnic