Person: Gil Dones, Félix
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First Name
Félix
Last Name
Gil Dones
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Ciencias Biológicas
Department
Genética, Fisiología y Microbiología
Area
Genética
Identifiers
11 results
Search Results
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Item Proteomic characterization of human coronary thrombus in patients with ST-segment elevation acute myocardial infarction(Journal of Proteomics, 2014) Alonso-Orgaz, Sergio; Moreno-Luna, Rafael; López, Juan A.; Gil Dones, Félix; Padial, Luis R.; Moreu, Jose; Cuesta, Fernando de la; Barderas, Maria G.Abstract Acute myocardial infarction with ST-segment elevation (STEMI) initiates with intraluminal thrombosis and results in total occlusion of the coronary artery. To date, characterization of the coronary thrombus proteome in STEMI patients has not been yet accomplished. Therefore, we aimed to perform an in-depth proteomic characterization of the human coronary thrombus by means of three different approaches: 2-DE followed by mass spectrometry (MALDI MS/MS), 1-DE combined either with liquid chromatography coupled to mass spectrometry in a MALDI TOF/TOF (LC–MALDI-MS/MS), or in a LTQ-Orbitrap (LC–ESI-MS/MS). This approach allowed us to identify a total of 708 proteins in the thrombus. Expression in coronary thrombi (n = 20) of 14 proteins was verified, and the expression of fibrin and 6 cell markers (platelets, monocytes, neutrophils, eosinophils, T-cells and B-cells) quantified by selected reaction monitoring (SRM). A positive correlation of 5 proteins (fermitin homolog 3, thrombospondin-1, myosin-9, beta parvin and ras-related protein Rap-1b) with CD41 was found, pointing out the potential activation of a focal adhesion pathway within thrombus platelets. DIDO1 protein was found to correlate negatively with thrombus fibrin, and was found up-regulated in the plasma of these STEMI patients, which may constitute a starting point for further analyses in the search for biomarkers of thrombosis. Biological significance The proteomic characterization of the human coronary thrombus may contribute to a better understanding of the mechanisms involved in acute coronary syndrome, and thus pave the road for the identification of new therapeutic targets that may help addressing this and other thrombotic diseases. A novel methodology to characterize thrombus composition and expression of a sub-group of proteins is hereby described, which allowed linking protein expression with cellular and ECM matrix composition of the thrombus. Five proteins (fermitin homolog 3, thrombospondin-1, myosin-9, beta parvin and ras-related protein Rap-1b) co-express within the human coronary thrombus with CD41, pointing out the potential activation of a focal adhesion pathway within thrombus platelets during thrombus formation. Besides, the protein death-inducer obliterator 1, found to be expressed within the human coronary thrombus, has been proved to increase in the plasma of STEMI patients, which constitutes an important starting point for further analyses in the search for biomarkers of thrombosis.Item 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.Item 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, FernandoAcute 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.Item 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.Item 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.Item 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.Item Secretome analysis of atherosclerotic and non-atherosclerotic arteries reveals dynamic extracellular remodeling during pathogenesis(Journal of Proteome, 2012) Cuesta, Fernando de la; Barderas, Maria G.; Calvo, Enrique; Zubiri, Irene; Maroto, Aroa S.; Darde, Veronica M.; Martin-Rojas, Tatiana; Gil Dones, Félix; Posada-Ayala, Maria; Tejerina, Teresa; Lopez, Juan A.; Vivanco Martínez, Fernando; Alvarez-Llamas, GloriaAims: Early detection of cardiovascular diseases and knowledge of underlying mechanisms is essential. Tissue secretome studies resemble more closely to the in vivo situation, showing a much narrower protein concentrations dynamic range than plasma. This study was aimed to the analysis of human arterial tissue secretome and to the quantitative comparison of healthy and atherosclerotic secretome to discover proteins with key roles in atherosclerosis development. Methods and results: Secretomes from three biological replicates of human atherosclerotic coronary arteries (APC), preatherosclerotic coronaries (PC) and mammaries (M) were analyzed by LC-MS/MS. The identified proteins were submitted to Ingenuity Pathway Analysis (IPA) tool. Label-free MS/MS based quantification was performed and validated by immunohistochemistry. 64 proteins were identified in the 3 replicates of at least one of the 3 groups and 15 secreted proteins have not been previously reported in plasma. Four proteins were significantly released in higher amounts by mammary tissue: gelsolin, vinculin, lamin A/C and phosphoglucomutase 5. Conclusion: The study of tissue secretome reveals key proteins involved in atherosclerosis which have not been previously reported in plasma. Novel proteins are here highlighted which could be potential therapeutic targets in clinical practice. This article is part of a Special Issue entitled: Proteomics: The clinical link.Item 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.Item A clinical perspective on the utility of alpha 1 antichymotrypsin for the early diagnosis of calcific aortic stenosis(Clinical Proteomics, 2017) Martin-Rojas, Tatiana; Mourino-Alvarez, Laura; Gil Dones, Félix; Cuesta, Fernando de la; Rosello-Lleti, Esther; Laborde, Carlos ; Rivera, Miguel; Lopez-Almodovar, Luis Fernando; Lopez, Juan Antonio; Akerstrom, Finn ; Padial, Luis ; Barderas, MariaBackground: Calcific aortic stenosis (CAS) is the most common heart valve disease in the elderly, representing an important economic and social burden in developed countries. Currently, there is no way to predict either the onset or progression of CAS, emphasizing the need to identify useful biomarkers for this condition. Methods: We performed a multi-proteomic analysis on different kinds of samples from CAS patients and healthy donors: tissue, secretome and plasma. The results were validated in an independent cohort of subjects by immunohistochemistry, western blotting and selected reaction monitoring. Results: Alpha 1 antichymotrypsin (AACT) abundance was altered in the CAS samples, as confirmed in the validation phase. The significant changes observed in the amounts of this protein strongly suggest that it could be involved in the molecular mechanisms underlying CAS. In addition, our results suggest there is enhanced release of AACT into the extracellular fluids when the disease commences. Conclusions: The significant increase of AACT in CAS patients suggests it fulfils an important role in the physiopathology of this disease. These results permit us to propose that AACT may serve as a potential marker for the diagnosis of CAS, with considerable clinical value.Item An Optimal Protocol to Analyze the Rat Spinal Cord Proteome(Biomarker Insights, 2009) Gil Dones, Félix; Alonso-Orgaz, Sergio; Avila, Gerardo; Martin-Rojas, Tatiana; Moral-Darde, Veronica; Barroso, Gemma; Vivanco Martínez, Fernando; Scott-Taylor, Julian; Barderas, MariaSince the function of the spinal cord depends on the proteins found there, better defing the normal Spinal Cord Proteome is an important and challenging task. Although brain and cerebrospinal fluid samples from patients with different central nervous system (CNS) disorders have been studied, a thorough examination of specific spinal cord proteins and the changes induced by injury or associated to conditions such as neurodegeneration, spasticity and neuropathies has yet to be performed. In the present study, we aimed to describe total protein content in the spinal cord of healthy rats, employing different proteomics tools. Accordingly, we have developed a fast, easy, and reproducible sequential protocol for protein extraction from rat spinal cords. We employed conventional two dimensional electrophoresis (2DE) in different pH ranges (eg. 4–7, 3–11 NL) combined with identification by mass spectrometry (MALDI-TOF/TOF), as well as first dimension protein separation combined with Liquid Chromatography Mass Spectrometry/Mass Spectrometry (LC-MS/MS) to maximise the benefits of this technology. The value of these techniques is demonstrated here by the identification of several proteins known to be associated with neuroglial structures, neurotransmission, cell survival and nerve growth in the central nervous system. Furthermore this study identified many spinal proteins that have not previously been described in the literature and which may play an important role as either sensitive biomarkers of dysfunction or of recovery after Spinal Cord Injury.