Profundizando en la estenósis aórtica valvular: análisis proteómico del plasma

Comunicaciones a congreso

An Optimal Protocol to Analyze the Rat Spinal Cord Proteome

Since 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

Estudio de la Estenosis Aórtica Valvular desde un punto de vista proteómico

Comunicaciones a congreso

Proteomics - A Powerful Tool to Deepen the Molecular Mechanisms of Aortic Stenosis Disease

We thank the grants from the Instituto de Salud Carlos III (FIS PI070537), Fondo de Investigación Sanitaria de Castilla la Mancha (FISCAM, PI2008/08), Fondo de Investigación Sanitaria de Castilla la Mancha (FISCAM PI2008/28) and Redes Temáticas de Investigación Cooperativa, FONDOS FEDER (RD06/0014/1015)Depto. de Genética, Fisiología y MicrobiologíaFac. de Ciencias BiológicasTRUEpu

Valvular Aortic Stenosis: A Proteomic Insight

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

iTRAQ proteomic analysis of extracellular matrix remodeling in aortic valve disease

Degenerative aortic stenosis (AS) is the most common worldwide cause of valve replacement. The aortic valve is a thin, complex, layered connective tissue with compartmentalized extracellular matrix (ECM) produced by specialized cell types, which directs blood flow in one direction through the heart. There is evidence suggesting remodeling of such ECM during aortic stenosis development. Thus, a better characterization of the role of ECM proteins in this disease would increase our understanding of the underlying molecular mechanisms. Aortic valve samples were collected from 18 patients which underwent aortic valve replacement (50% males, mean age of 74 years) and 18 normal control valves were obtained from necropsies (40% males, mean age of 69 years). The proteome of the samples was analyzed by 2D-LC MS/MS iTRAQ methodology. The results showed an altered expression of 13 ECM proteins of which 3 (biglycan, periostin, prolargin) were validated by Western blotting and/or SRM analyses. These findings are substantiated by our previous results demonstrating differential ECM protein expression. The present study has demonstrated a differential ECM protein pattern in individuals with AS, therefore supporting previous evidence of a dynamic ECM remodeling in human aortic valves during AS development

Proteomic Profile of Human Aortic Stenosis: Insights into the Degenerative Process

This work was supported by grants from the Instituto de Salud Carlos III (FISPI070537,PI080970),Fondo de Investigación Sanitaria de Castilla la Mancha (FISCAM,PI2008/08), Fondo de Investigación Sanitaria de Castilla la Mancha (FISCAM PI2008/28) and Fondos Feder-Redes Telemáticas de Investigación Cooperativa (RD06/0014/1015)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.Depto. de Genética, Fisiología y MicrobiologíaFac. de Ciencias BiológicasTRUEpu

Proteomics - A Powerful Tool to Deepen the Molecular Mechanisms of Aortic Stenosis Disease

We thank the grants from the Instituto de Salud Carlos III (FIS PI070537), Fondo de Investigación Sanitaria de Castilla la Mancha (FISCAM, PI2008/08), Fondo de Investigación Sanitaria de Castilla la Mancha (FISCAM PI2008/28) and Redes Temáticas de Investigación Cooperativa, FONDOS FEDER (RD06/0014/1015)Depto. de Genética, Fisiología y MicrobiologíaFac. de Ciencias BiológicasTRUEpu

Inside human aortic stenosis: a proteomic analysis of plasma

This work was supported by grants from the Instituto de Salud Carlos III (FIS PI070537, CP09/00229), Fondo de Investigación Sanitaria de Castilla la Mancha (FISCAM, PI2008/28 and PI2008/08) and Redes Temáticas de Investigación Cooperativa (FONDOS FEDER, RD06/0014/1015), CNIC.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.Depto. de Genética, Fisiología y MicrobiologíaFac. de Ciencias BiológicasTRUEpu