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Gomez-Choco M.,Functional Unit of Cerebrovascular Diseases | Doucerain C.,Institute for Biomedical Research of Barcelona IIBB | Doucerain C.,August Pi i Sunyer Biomedical Research Institute IDIBAPS | Urra X.,Functional Unit of Cerebrovascular Diseases | And 6 more authors.
Journal of Neuroimmunology | Year: 2014

Heat shock protein 70 (Hsp-70) can act as a danger signal and activate immune responses. We studied the presence of Hsp-70 in lymphoid tissue and plasma of acute stroke patients and asymptomatic controls free of neurological disease. Immunofluorescence, Western blotting, qRT-PCR and flow cytometry studies were performed. Plasma Hsp-70 concentration at day 7 was similar in patients and controls, whereas patients disclosed stronger immunoreactivity to Hsp-70 in lymphoid tissue than controls. Most Hsp-70. + cells were antigen presenting cells located in T cell zones. Stronger immunoreactivity to Hsp-70 was associated with smaller infarctions and better functional outcome. © 2014 Elsevier B.V. Source


De La Rosa X.,Institute for Biomedical Research of Barcelona IIBB | Santalucia T.,Institute for Biomedical Research of Barcelona IIBB | Fortin P.-Y.,University of Bordeaux Segalen | Purroy J.,Institute for Biomedical Research of Barcelona IIBB | And 5 more authors.
European Journal of Nuclear Medicine and Molecular Imaging | Year: 2013

Purpose: Stroke induces strong expression of the 72-kDa heat-shock protein (HSP-70) in the ischaemic brain, and neuronal expression of HSP-70 is associated with the ischaemic penumbra. The aim of this study was to image induction of Hsp-70 gene expression in vivo after brain ischaemia using reporter mice. Methods: A genomic DNA sequence of the Hspa1b promoter was used to generate an Hsp70-mPlum far-red fluorescence reporter vector. The construct was tested in cellular systems (NIH3T3 mouse fibroblast cell line) by transient transfection and examining mPlum and Hsp-70 induction under a challenge. After construct validation, mPlum transgenic mice were generated. Focal brain ischaemia was induced by transient intraluminal occlusion of the middle cerebral artery and the mice were imaged in vivo with fluorescence reflectance imaging (FRI) with an intact skull, and with confocal microscopy after opening a cranial window. Results: Cells transfected with the Hsp70-mPlum construct showed mPlum fluorescence after stimulation. One day after induction of ischaemia, reporter mice showed a FRI signal located in the HSP-70-positive zone within the ipsilateral hemisphere, as validated by immunohistochemistry. Live confocal microscopy allowed brain tissue to be visualized at the cellular level. mPlum fluorescence was observed in vivo in the ipsilateral cortex 1 day after induction of ischaemia in neurons, where it is compatible with penumbra and neuronal viability, and in blood vessels in the core of the infarction. Conclusion: This study showed in vivo induction of Hsp-70 gene expression in ischaemic brain using reporter mice. The fluorescence signal showed in vivo the induction of Hsp-70 in penumbra neurons and in the vasculature within the ischaemic core. © 2012 Springer-Verlag Berlin Heidelberg. Source


Penas C.,Cell Medica | Font-Nieves M.,Institute for Biomedical Research of Barcelona IIBB | Fores J.,University of Barcelona | Petegnief V.,Institute for Biomedical Research of Barcelona IIBB | And 3 more authors.
Cell Death and Differentiation | Year: 2011

Disconnection of the axon from the soma of spinal motoneurons (MNs) leads either to a retrograde degenerative process or to a regenerative reaction, depending on the severity and the proximity to the soma of the axonal lesion. The endoplasmic reticulum (ER) is a continuous membranous network that extends from the nucleus to the entire cytoplasm of the neuronal soma, axon and dendrites. We investigated whether axonal injury is sensed by the ER and triggers the activation of protective mechanisms, such as the unfolded protein response (UPR) and autophagy. We found early (at 3 days) accumulation of beclin1, LC3II and Lamp-1, hallmarks of autophagy, in both degenerating MNs after spinal root avulsion and in non-degenerating MNs after distal nerve section, although Lamp-1 disappeared by 5 days only in the former. In contrast, only degenerating MNs presented early activation of IRE1α, revealed by an increase of the spliced isoform of Xbp1 and accumulation of ATF4 in their nucleus, two branches of the UPR, and late BiP downregulation in association with cytoskeletal and organelle disorganization. We conclude that BiP decrease is a signature of the degenerating process, as its overexpression led to an increase in MN survival after root avulsion. Besides, Bcl2 is strongly implicated in the survival pathway activated by BiP overexpression. © 2011 Macmillan Publishers Limited All rights reserved. Source


Psilodimitrakopoulos S.,ICFO - Institute of Photonic Sciences | Petegnief V.,Institute for Biomedical Research of Barcelona IIBB | Soria G.,Institute for Biomedical Research of Barcelona IIBB | Amat-Roldan I.,ICFO - Institute of Photonic Sciences | And 4 more authors.
Progress in Biomedical Optics and Imaging - Proceedings of SPIE | Year: 2010

We use high resolution polarization second harmonic generation (PSHG) imaging microscopy in cultured neurons, and we provide estimation on the effective orientation of the SHG source in neuronal processes in vitro. We performed pixel by pixel analysis and we found a picked distribution of angles with maximum frequency at θe = 34.54°, with Δθe=11.44°. This angle value is very close to the inclination of the tubulin heterodimmer with respect to the long axis of the microtubule. © 2010 Copyright SPIE - The International Society for Optical Engineering. Source


Psilodimitrakopoulos S.,ICFO - Institute of Photonic Sciences | Petegnief V.,Institute for Biomedical Research of Barcelona IIBB | Soria G.,Institute for Biomedical Research of Barcelona IIBB | De Vera N.,Institute for Biomedical Research of Barcelona IIBB | And 5 more authors.
Progress in Biomedical Optics and Imaging - Proceedings of SPIE | Year: 2010

Polarization sensitive second harmonic generation (PSHG) imaging can provide useful information which is unreachable by intensity SHG imaging. Specifically, it can provide geometrical characteristics of the SHG source molecular architecture. The information is obtained by rotating the excitation linear polarization and by fitting the SHG intensity variation to a cylindrical symmetry biophysical model. As a result, the ratios of the non-vanishing χ2 tensor elements, responsible for the SHG conversion, are retrieved. In the end, by assuming a SHG source with dominant hyperpolarizability, its molecular orientation can be estimated. Here, we developed and used this approach to retrieve submicron structural information from cultured neurons and to provide estimation on the effective orientation of the molecular SHG source in axons. For that purpose, the PSHG images of axons were fitted pixel by pixel using an algorithm based on the above mentioned model. A coefficient of determination of r2>90% was chosen as a filtering mechanism. For a selected region of interest we then retrieved the pixels' values histogram of the harmonophores' effective orientations, θe. The distribution was centred at θe=34.93°, with σ=7.62°. These angle values correspond to the geometrical characteristics of the tubulin heterodimmers forming the microtubules. Modifications on tubulin dimers may alter θe, σ thus the PSHG optical technique suggests novel quantitative biomarkers able to characterize neurons' plasticity as well as disease progression. © 2010 SPIE. Source

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