Person:
Gil Dones, Félix

First Name

Félix

Last Name

Gil Dones

URI

https://hdl.handle.net/20.500.14352/74303

Affiliation

Universidad Complutense de Madrid

Faculty / Institute

Ciencias Biológicas

Department

Genética, Fisiología y Microbiología

Area

Genética

Identifiers

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Now showing 1 - 9 of 9

Development of an Optimal Protocol for the Proteomic Analysis of Stenotic and Healthy Aortic Valves
(Revista Española de Cardiologia, 2010) Martín-Rojas, Tatiana; López-Almodovar, Luis F.; Juárez-Tosina, Rocío; Gil Dones, Félix; Cuesta, Fernando de La; Álvarez-Llamas, Gloria; Alonso-Orgaz, Sergio; Vivanco, Fernando; Rodríguez-Padial, Luis; Barderas, María G.
Introduction and objectives: For many years, degenerative aortic stenosis was thought to be a passive process secondary to calcium deposition in aortic valves. Although its etiology remains unknown, several authors have pointed out that degenerative aortic stenosis is associated with the same risk factors as coronary artery disease. Furthermore, histological similarities have been found between aortic valve stenosis and atherosclerotic plaque, giving rise to the hypothesis that degenerative aortic stenosis is an inflammatory process similar to atherosclerosis. Nevertheless, some data do not fit with this hypothesis and, consequently, greater understanding of the condition is needed. The main aim of this study was to develop a practical protocol for extracting protein for use in proteomic analysis from both stenotic and healthy aortic valves. Methods: The study was carried out using a number of different proteomic methods: two-dimensional electrophoresis, mass spectrometry, and additional techniques. Results: We developed a simple and reproducible methodology in the laboratory for carrying out the proteomic analysis of human aortic valves and for identifying their component proteins. Conclusions: We developed a simple and reproducible method for extracting protein that can be used with mass spectrometry and that makes it possible to carry out large-scale proteomic analysis of stenotic aortic valves. Furthermore, the methodology will significantly increase our understanding of the valve proteome.
Analysis of the Plasma Proteome Associated with Acute Coronary Syndrome: Does a Permanent Protein Signature Exist in the Plasma of ACS Patients?
(Journal of protemoe research, 2010) Dardé, Veronica M.; Cuesta, Fernando de la; Gil Dones, Félix; Alvarez-Llamas, Gloria; Barderas, Maria G.; Vivanco, Fernando
Acute coronary syndrome (ACS) is triggered by the occlusion of a coronary artery usually due to the thrombosis caused by an atherosclerotic plaque. The identification of proteins directly involved in the pathophysiological events underlying ACS will enable more precise diagnoses and a more accurate prognosis to be determined. Accordingly, we have performed a longitudinal study of the plasma proteome in ACS patients by 2-DE and DIGE. Plasma samples from patients, healthy controls, and stable coronary artery disease (CAD) patients were immunodepleted of the six most abundant proteins, and they were analyzed in parallel at four different times: 0 (on admission) and after 4, 60, and 180 days. From a total of 1400 spot proteins analyzed, 33 proteins were differentially expressed in ACS patients when compared with control subjects/stable patients. A small group of seven proteins that appear to be altered at admission remain affected for 6 months and also in the stable CAD patients. Interestingly, the maximum number of altered proteins was observed in the stable CAD patients. Some of the proteins identified had been previously associated with ACS whereas others (such as Alpha-1-B-glycoprotein, Hakata antigen, Tetranectin, Tropomyosin 4) constitute novel proteins that are altered in this pathology.
Targeting antigens to an invariant epitope of the MHC Class II DR molecule potentiates the immune response to subunit vaccines
(Virus Research, 2011) Gil Dones, Félix; Pérez-Filgueira, Mariano; Barderas, María G.; Pastor Vargas, Carlos; Alonso, Covadonga; Vivanco, Fernando; Escribano, José M.
Recombinant subunit and peptidic vaccines in general present a reduced immunogenicity in vaccinated individuals with respect to the whole pathogen from which they derived. The generation of strong immune responses to these vaccines requires the use of potent adjuvants, high antigen doses and repetitive vaccinations. In this report, we document the enhanced antibody response obtained against two recombinant subunit vaccines by means of targeting to antigen-presenting cells by a recombinant single chain antibody. This antibody, named APCH1, recognizes an epitope of MHC Class II DR molecule preserved in different animal species, including humans. We showed that vaccinal antigens translationally fused to APCH1 antibody and produced by recombinant baculoviruses in insect larvae (Trichoplusia ni), elicited an increased antibody response in comparison with the same antigens alone or fused to a carrier molecule. These results suggest that targeting of antigens to this invariant MHC Class II epitope has immunopotentiating effects that could circumvent the reduced potency of peptidic or subunit vaccines, opening the possibility of widespread application of APCH1 as a new adjuvant antibody of general use.
Modification of the Secretion Pattern of Proteases, Inflammatory Mediators, and Extracellular Matrix Proteins by Human Aortic Valve is Key in Severe Aortic Stenosis
(Molecular and cellular proteomics, 2013) Alvarez-Llamas, Gloria; Martín-Rojas, Tatiana; Cuesta, Fernando de la; Calvo, Enrique; Gil Dones, Félix; Dardé, Veronica M.; Lopez-Almodovar, Luis F; Padial, Luis R.; Lopez, Juan-Antonio; Vivanco, Fernando; Barderas, Maria G.
One of the major challenges in cardiovascular medicine is to identify candidate biomarker proteins. Secretome analysis is particularly relevant in this search as it focuses on a subset of proteins released by a cell or tissue under certain conditions. The sample can be considered as a plasma subproteome and it provides a more direct approximation to the in vivo situation. Degenerative aortic stenosis is the most common worldwide cause of valve replacement. Using a proteomic analysis of the secretome from aortic stenosis valves we could identify candidate markers related to this pathology, which may facilitate early diagnosis and treatment. For this purpose, we have designed a method to validate the origin of secreted proteins, demonstrating their synthesis and release by the tissue and ruling out blood origin. The nLC-MS/MS analysis showed the labeling of 61 proteins, 82% of which incorporated the label in only one group. Western blot and selective reaction monitoring differential analysis, revealed a notable role of the extracellular matrix. Variation in particular proteins such as PEDF, cystatin and clusterin emphasizes the link between aortic stenosis and atherosclerosis. In particular, certain proteins variation in secretome levels correlates well, not only with label incorporation trend (only labeled in aortic stenosis group) but, more importantly, with alterations found in plasma from an independent cohort of samples, pointing to specific candidate markers to follow up in diagnosis, prognosis, and therapeutic intervention.
Inside human aortic stenosis: a proteomic analysis of plasma
(Journal of Proteomics, 2012) Gil Dones, Félix; Darde, Verónica M.; Alonso-Orgaz, Sergio; Lopez-Almodovar, Luis F.; Mourino-Alvarez, Laura; Padial, Luis R.; Vivanco, Fernando; Barderas, Maria G.
Valvular aortic stenosis (AS) produces a slowly progressive obstruction in left ventricular outflow track. For this reason, aortic valve replacement is warranted when the valvular stenosis is hemodinamically significant, becoming the most common worldwide cause of aortic valve surgery. Recent epidemiologic studies have revealed an association between degenerative AS and cardiovascular risk factors for atherosclerosis, althought reducing the exposure to such factors and statin therapies both fail to delay or reverse the pathology. Hence, a deeper understanding of the pathophysiology of this disease is required to identify appropriate preventive measures. A proteomic analysis of plasma will permit to know and identify the changes in protein expression induced by AS in this tissue. Using two-dimensional difference gel electrophoresis (2D-DIGE) followed by mass spectrometry (MS), we compared the crude (not pre-fractioned) and pre-fractioned plasma from AS patients and control subjects. We sought to identify plasma proteins whose expression is modified in AS. In addition we investigated if crude plasma presented some alterations in the more abundant proteins since to date, has never been studied before. We also further investigated the link between this disease and atherosclerosis with a view to identifying new potential markers and therapeutic targets.
Potential blood biomarkers for stroke
(Expert Review of Proteomics, 2012) Laborde, Carlos M.; Mourino–Alvarez, Laura; Akerstrom, Finn; Padial, Luis R.; Vivanco, Fernando; Gil Dones, Félix; Barderas, Maria G.
Stroke is one of the most common causes of death worldwide and a major cause of acquired disability in adults. Despite advances in research during the last decade, prevention and treatment strategies still suffer from significant limitations, and therefore new theoretical and technical approaches are required. Technological advances in the proteomic and metabolomic areas, during recent years, have permitted a more effective search for novel biomarkers and therapeutic targets that may allow for effective risk stratification and early diagnosis with subsequent rapid treatment. This review provides a comprehensive overview of the latest candidate proteins and metabolites proposed as new potential biomarkers in stroke.
Proteomic Profile of Human Aortic Stenosis: Insights into the Degenerative Process
(Journal of Proteome Research, 2012) Gil Dones, Félix; Martín-Rojas, Tatiana; Gil-Dones, Felix; Lopez-Almodovar, Luis F.; Padial, Luis R.; Vivanco, Fernando; Barderas, Maria G.
Degenerative aortic stenosis is the most common worldwide cause of valve replacement. While it shares certain risk factors with coronary artery disease, it is not delayed or reversed by reducing exposure to risk factors (e.g., therapies that lower lipids). Therefore, it is necessary to better understand its pathophysiology for preventive measures to be taken. In this work, aortic valve samples were collected from 20 patients that underwent aortic valve replacement (55% males, mean age of 74 years) and 20 normal control valves were obtained from necropsies (40% males, mean age of 69 years). The proteome of the samples was analyzed by quantitative differential electrophoresis (2D-DIGE) and mass spectrometry, and 35 protein species were clearly increased in aortic valves, including apolipoprotein AI, alpha-1-antitrypsin, serum albumin, lumican, alfa-1-glycoprotein, vimentin, superoxide dismutase Cu–Zn, serum amyloid P-component, glutathione S-transferase-P, fatty acid-binding protein, transthyretin, and fibrinogen gamma. By contrast, 8 protein species were decreased (transgelin, haptoglobin, glutathione peroxidase 3, HSP27, and calreticulin). All of the proteins identified play a significant role in cardiovascular processes, such as fibrosis, homeostasis, and coagulation. The significant changes observed in the abundance of key cardiovascular proteins strongly suggest that they can be involved in the pathogenesis of degenerative aortic stenosis. Further studies are warranted to better understand this process before we can attempt to modulate it.
Valvular Aortic Stenosis: A Proteomic Insight
(Clinical Medicine Insights: Cardiology, 2010) Gil Dones, Félix; Martin-Rojas, Tatiana; Lopez-Almodovar, Luis ; Cuesta, Fernando de La; Darde, Veronica ; Alvarez-Llamas, Gloria; Juarez-Tosina, Rocio; Barroso, Gemma; Vivanco, Fernando; Padial, Luis ; Barderas, Maria
Calcified aortic valve disease is a slowly progressive disorder that ranges from mild valve thickening with no obstruction of blood flow, known as aortic sclerosis, to severe calcification with impaired leaflet motion or aortic stenosis. In the present work we describe a rapid, reproducible and effective method to carry out proteomic analysis of stenotic human valves by conventional 2-DE and 2D-DIGE, minimizing the interference due to high calcium concentrations. Furthermore, the protocol permits the aortic stenosis proteome to be analysed, advancing our knowledge in this area. Until recently, aortic stenosis (AS) was considered a passive process secondary to calcium deposition in the aortic valves. However, it has recently been highlighted that the risk factors associated with the development of calcified AS in the elderly are similar to those of coronary artery disease. Furthermore, degenerative AS shares histological characteristics with atherosclerotic plaques, leading to the suggestion that calcified aortic valve disease is a chronic inflammatory process similar to atherosclerosis. Nevertheless, certain data does not fit with this theory making it necessary to further study this pathology. The aim of this study is to develop an effective protein extraction protocol for aortic stenosis valves such that proteomic analyses can be performed on these structures. In the present work we have defined a rapid, reproducible and effective method to extract proteins and that is compatible with 2-DE, 2D-DIGE and MS techniques. Defining the protein profile of this tissue is an important and challenging task that will help to understand the mechanisms of physiological/pathological processes in aortic stenosis valves.
Proteomics - A Powerful Tool to Deepen the Molecular Mechanisms of Aortic Stenosis Disease
(Aortic Stenosis - Etiology, Pathophysiology and Treatment, 2011) Gil Dones, Félix; Cuesta, Fernando de la; Álvarez Llamas, Gloria; Padial, Luis ; López-Almodovar, Luis ; Martin-Rojas, Tatiana; Vivanco, Fernando; Barderas, Maria