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Hospital de Órbigo, Spain

Zubiri I.,IIS Fundacion Jimenez Diaz | Posada-Ayala M.,IIS Fundacion Jimenez Diaz | Sanz-Maroto A.,IIS Fundacion Jimenez Diaz | Martin-Lorenzo M.,IIS Fundacion Jimenez Diaz | And 7 more authors.
Journal of Proteomics

Diabetic nephropathy (DN) is a major complication of diabetes mellitus (DM), the most frequent cause of end-stage renal disease (ESRD). Exosomes isolated from urine are considered a rich non-invasive source of markers for renal events. Proteinuria associated with DN patients at advanced stages may result in "contamination" of exosomal fraction by co-precipitation of high abundance urine proteins, making it enormously difficult to obtain a reliable comparison of healthy individuals and DN patients and to detect minor proteins. We evaluated different protocols for urinary exosome isolation (ultracentrifugation-based and Exoquick® reagent-based) in combination with an easy and quick depletion procedure of contaminating high abundance proteins (albumin). The optimal methodology was then applied to investigate the proteome of human urinary exosomes in DN and controls using spectral counting LC-MS/MS analysis followed by selected reaction monitoring (SRM) confirmation. A panel of 3 proteins (AMBP, MLL3, and VDAC1) is differentially present in urinary exosomes from DN patients, opening a new field of research focused on improving diagnosis and follow-up of this pathology. Biological significance: Diabetic nephropathy (DN) is a progressive proteinuric kidney disease, a major complication of diabetes mellitus, and the most frequent cause of end-stage renal disease. Current markers of disease (i.e. creatinine and urinary albumin excretion) have proven limitations (i.e. some patients regress to normoalbuminuria, kidney damage may be already present in recently diagnoses microalbuminuric patients and renal function may decrease in the absence of significant albuminuria). We show here the first study on human DN proteome of urinary exosomes. Proteinuria associated to DN patients resulting in contamination of exosomal fraction and the associated difficulty to reliably compare healthy and disease conditions, are here overcome. A combined methodology pointed to increase exosomal proteome recovery and depletion of high-abundance proteome was here set-up. A total of 352 proteins were here identified for the first time associated to human urinary exosomes. Label-free quantitative comparison of DN urinary exosomes vs control group and SRM further validation, resulted in the discovery of a panel of three proteins (AMBP, MLL3 and VDAC1) which changes in DN, opening a new field of research focused to improve diagnosis and follow-up of this pathology. © 2013 Elsevier B.V. Source

Martinez-Maza C.,CNRS Paris Institute of Earth Sciences | Martinez-Maza C.,CSIC - National Museum of Natural Sciences | Rosas A.,CSIC - National Museum of Natural Sciences | Nieto-Diaz M.,Hospital Nacional de Paraplejicos SESCAM
American Journal of Physical Anthropology

Developmental and evolutionary changes in craniofacial morphology are a central issue in paleoanthropology, but the underlying bone growth processes have been scarcely studied. Relevant knowledge on bone growth dynamics can be obtained from the spatial distribution of bone formation and resorption activities. Determining these patterns from the valuable samples typically used in anthropology and palaeoanthropology necessarily implies nondestructive procedures. In this work, we present a methodology based on the analysis of high-resolution replicas by reflected light microscopy, describing how microfeatures related to bone formation and resorption activities are recognized on both recent and fossil bone surfaces. The proposed method yields highly similar images to those obtained with scanning electron microscope and has proven its utility in an analysis of a large sample of extant and extinct hominoids. © 2010 Wiley-Liss, Inc. Source

Calvino-Nunez C.,Pablo De Olavide University | Dominguez-Del-toro E.,Pablo De Olavide University | De Castro F.,Hospital Nacional de Paraplejicos SESCAM

During the development of the nervous system, the perinatal period is particularly sensitive as neuronal connections are still forming in the brain of the neonate. Alpha2-adrenergic receptors are overexpressed temporarily in proliferative zones in the developing brain, reaching a peak during the first postnatal week of life. Both stimulation and blocking of these receptors during this period alter the development of neural circuits, affecting synaptic connectivity and neuronal responses. They even affect motor and cognitive skills later on in the adult. It's especially important to look for the early neurological consequences resulting from such modifications, because they may go unnoticed. The main objective of the present study has been to reaffirm the importance of the maturation of alpha-adrenergic system in mice, by carrying out a comprehensive examination of motor, behavioral and cognitive effects in neonates, during early postnatal development, following chronic administration of the drug Clonidine, an alpha2 adrenergic system agonist. Our study shows that mice treated postnatally with clonidine present a temporal delay in the appearance of developmental markers, a slow execution of vestibular reflexes during first postnatal week of life and a blockade of the short term memory in the novel object recognition task. Shortly after the treatment the startle response is hyperreactive. © 2014 Calvino-Nunez, Dominguezdel-Toro. Source

Lopez-Dolado E.,Hospital Nacional de Paraplejicos SESCAM | Gonzalez-Mayorga A.,Hospital Nacional de Paraplejicos SESCAM | Portoles M.T.,Complutense University of Madrid | Feito M.J.,Complutense University of Madrid | And 4 more authors.
Advanced Healthcare Materials

The increasing prevalence and high sanitary costs of lesions affecting the central nervous system (CNS) at the spinal cord are encouraging experts in different fields to explore new avenues for neural repair. In this context, graphene and its derivatives are attracting significant attention, although their toxicity and performance in the CNS in vivo remains unclear. Here, the subacute tissue response to 3D flexible and porous scaffolds composed of partially reduced graphene oxide is investigated when implanted in the injured rat spinal cord. The interest of these structures as potentially useful platforms for CNS regeneration mainly relies on their mechanical compliance with neural tissues, adequate biocompatibility with neural cells in vitro and versatility to carry topographical and biological guidance cues. Early tissue responses are thoroughly investigated locally (spinal cord at C6 level) and in the major organs (i.e., kidney, liver, lung, and spleen). The absence of local and systemic toxic responses, along with the positive signs found at the lesion site (e.g., filler effect, soft interface for no additional scaring, preservation of cell populations at the perilesional area, presence of M2 macrophages), encourages further investigation of these materials as promising components of more efficient material-based platforms for CNS repair. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. Source

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