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DIFFERENZIAMENTO E CREAZIONE DI UN MICROAMBIENTE RIGENERATIVO NELLA LESIONE MIDOLLARE DA PARTE DEI PRECURSORI NEURALI POST MORTEM (PM-NPCS). ANALISI DI TIPO BIOCHIMICO, MORFOLOGICO, ED IN VIVO ATTRAVERSO LA RISONANZA MAGENTICA ED IL RECUPERO FUNZIONALE DEGLI ARTI POSTERIORI
Full text linkTraumatic injuries in central nervous system lead to severe and permanent neurological deficit. Particularly, traumatic spinal cord injury often results in a devastating loss of neurological function below the injury site. Since the loss of CNS neurons may not be replaced by the proliferation of the surviving ones, intraspinal transplantation of exogenous neuronal cells or tissue has been accepted for a long time as a way to obtain a partial reconstruction of the lost cord tissue and to promote recovery of neurological function.
Cell-based therapies in the injured spinal cord are intended to fill lesion cavities, which typically develop at an injury site, and to provide a cellular growth-conducive substrate for re-growing axons. Various cell types such as fibroblasts, olfactory ensheathing cells, Schwann cells and neural stem ⁄precursor cells have been used to regenerate or replace injured spinal cord parenchyma, and to elicit axonal regen- eration which is the primary goal of regenerative medicine and one of the prime motivations for the study of stem cells (NCSs).
Unfortunately when NSCs are administrated in a spinal cord injury model they modulate the inflammatory response but do not differentiate into mature cells and are quickly engulfed by macrophages present at lesion site. Recently we isolated a new class of neural stem cell from the subventricular zone of mice forebrain named Post-Mortem Neural Precursor Cells (PM-NPCs), that are capable of surviving after a prolonged ischemic insult. PM-NPCs for their potentiality in terms of proliferation and differentiation capabilities, are a good tool for tissue replacement therapies. In this study we focused on transplantation of PM-NPCs in a murine model of spinal cord injury by endovenous injection within 2 hours after injury. After administration, cells migrate specifically to the site of injury, as demonstrated both by ex-vivo immunoistochemistry and in-vivo MRI after PM-NPCs labelling with Superparamagnetic Iron Oxide Particles (SPIOs). Interestingly, these cells survive in such an unfavorable enviroment wich is the injury site and differentiate predominantly into cholinergic neurons, reconstituting a rich axonal and dendritic network and promoting a marked axonal regeneration across the injury site of monoaminergic fibers within 30 days from their administration.
Fluororuby injection in the dorsal funiculi rostral to the lesion site shows
that some spared and/or regenereting corticospinal (CS) labelled fibres bypass the lesion site and are in the caudal portion of PM-NSCs mice treated spinal cord, where few or none CS labelled fibers are found in saline trated mice. Moreover, beavhioral assesment evaluated by means of Basso Mouse Scale (BMS), shows a significantly improve of hind limb function in PM-NPCs treated mice compared with animals treated with placebo.
Eventually, the molecular analysis of the lesion site shows that PM-NPCs induce a remodulation of inflammatory response through the reduction of astrocytes activation and glial scar formation, and the reduction of immunitary cells infiltration (neutrophils and macrophages) leading to a better tissue preservation.
Moreover, the expression of proinflammatory cytokines and release of neurotrophic factors are reguated by PM-NPCs treatement: some proinflammatory cytokines (IL-6, MIP-2 and TNF alpha) levels significantly decrease after 48 hours from spinal cord injury and PM-NPCs transplantation, while after 7 days we observe an increase of IL-6 and TNF alpha probably because at longer time those cytokines are necessary to support the regenerative process according to the literature. These data suggest that PM-NPCs may represent a good source for cellular therapy in neurodegenerative disorders, specially on spinal cord injury
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Dispelling the Myths Behind First-author Citation Counts
Full text linkWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
Nucleotide receptors in trigeminal satellite glial cells as new targets for the pharmacological control of migraine pain: in vitro and in vivo studies
No full textThe main aim of my PhD research project is to understand what occurs to pain transduction mechanisms of trigeminal sensory neurons in migraine, why they become hyperactive, and how we may stop them. We focused our studies on the role of the purinergic system in the neuron-to-glial cells communication within the trigeminal ganglion, and its cross-talk with known pro-algogenic systems (such as bradikynin, BK, calcitonin gene-related peptide, CGRP, and prostaglandins). The final goal is the identification of new cellular and molecular players in the onset and maintenance of trigeminal-associated pain, for the development of new effective therapeutic strategies for migraine.
To this purpose, we set up and in vitro model of primary mixed neuron-glia or purified satellite glial cells (SGCs) cultures from trigeminal ganglion (TG) from C57BL6 mice. G protein-coupled P2Y receptor function was evaluated by single cell calcium imaging, and the extracellular concentrations of CGRP were measured by an ELISA assay. Western blot experiments on P2Y1 and P2Y2 receptors subtypes were also performed. Concerning the role of metabotropic purinergic receptors, our data show that exposure of mixed-neuron glia cultures to BK induces neuronal release of CGRP, which in turn significantly potentiates the ADP responsive P2Y1 and the UTP-sensitive P2Y2 receptor subtypes on surrounding SGCs. The increased activity of P2 receptors is not only due to increased receptor protein expression, but also, and especially for the P2Y1 subtype, to modulation of the receptor localization to membrane lipid rafts. Interestingly, the anti-migraine drug sumatriptan fully inhibits both CGRP release and glial P2Y-receptor potentiation. Moreover, exposure to BK leads to increased production of PGE2, an effect completely abolished by the COX-1 inhibitor acetylsalicylic acid. The latter also blocks neuronal CGRP release. Taken together, these results suggest that a complex cross-talk between neuronal and glial cells takes place in the TG, involving pain mediators and extracellular nucleotides. Modulation of this network by known anti-migraine drugs, such as triptans and COX inhibitors, suggests that it might play an important role in the development of migraine pain.
The second part of the current project is aimed at evaluating the pro- or anti-algogenic role of P2Y receptors through their selective inhibition in vivo. To this purpose, we set up a sub-chronic inflammatory model characterized by inflammatory pain and trigeminal hypersensitivity, by injecting complete Freund adjuvant (CFA) into the temporomandibular joint (TMJ) of rats. CFA-injected animals showed ipsilateral mechanical allodynia and TMJ edema. Glial cell activation was then evaluated in the spinal-trigeminal system by immunohistochemistry, confirming that our model leads to trigeminal inflammation and sensitization. Western blot experiments on GFAP protein expression, a typical marker of glial cells, confirmed the presence of glial activation within the trigeminal ganglion starting from 24 hours up to 11 days after CFA injection. Moreover, P2Y1 receptor subtype protein expression is upregulated in the ipsilateral trigeminal ganglia of CFA-injected rats starting from a week after injection, while this increase is observed starting from 72 h post-injection for P2Y2 receptor subtype. Interestingly, the non-selective P2Y antagonist PPADS shows a strong analgesic effect on CFA-induced trigeminal inflammation, which is comparable to acetylsalicylic acid (ASA)-mediated analgesia.
These results suggest a possible the pro-algogenic role for P2Y receptors in the development of trigeminal sensitization and migraine pain, opening the future perspective of identifying innovative and more selective pharmacological approaches for the sake of those migraineurs who are insensitive to currently available drugs
Nucleotide receptors in trigeminal satellite glial cells as new targets for the pharmacological control of migraine pain: in vitro and in vivo studies
No full textBackground and Purpose - The main aim of my PhD research project is to study pain transduction mechanisms in trigeminal ganglia (TG) in migraine, to understand why sensory trigeminal neurons become hyperactive in this pathological status, and how their activity can be pharmacologically modulated. We focused our studies on the role of the purinergic system in the neuron-to-glial cells communication within the TG, and its cross-talk with known pro-algogenic systems (such as bradykinin, BK, calcitonin gene-related peptide, CGRP, and prostaglandins). The final goal is the identification of new cellular and molecular players in the onset and maintenance of trigeminal-associated pain, for the development of new effective therapeutic strategies for migraine.
Methods and Results – For the in vitro studies, we set up primary mixed neuron-glia or purified satellite glial cell (SGCs) cultures from theTG of C57BL6 mice. G protein-coupled P2Y receptor function was evaluated by single cell calcium imaging, and the extracellular concentrations of CGRP and PGE2 were measured by ELISA assays. Western blot experiments on P2Y1 and P2Y2 receptor subtypes were also performed. Concerning the role of metabotropic purinergic receptors, our data show that exposure of mixed-neuron glia cultures to BK induced the neuronal release of CGRP, which in turn significantly potentiated the ADP-responsive P2Y1 and the UTP-sensitive P2Y2 receptor subtypes on surrounding SGCs. The increased activity of P2 receptors was related to increased receptor protein expression. Interestingly, the anti-migraine drug sumatriptan fully inhibited both CGRP release and glial P2Y-receptor potentiation. Moreover, exposure to BK led to increased production of PGE2, an effect completely abolished by the COX-1 inhibitor acetylsalicylic acid (ASA), whichalso blocked neuronal CGRP release. Taken together, these results suggest that a complex cross-talk between neuronal and glial cells takes place in the TG, involving pain mediators and extracellular nucleotides. Modulation of this network by known anti-migraine drugs, such as triptans and COX inhibitors, suggests that it might play an important role in the development of migraine pain.
In vivo studies were aimed at evaluating the pro- or anti-algogenic role of P2Y receptors through their selective inhibition. To this purpose, we set up a sub-chronic model of inflammatory trigeminal hypersensitivity, by injecting complete Freund adjuvant (CFA) into the temporomandibular joint (TMJ) of rats. CFA-injected animals showed ipsilateral mechanical allodynia and TMJ edema. Glial cell activation was evaluated in the spinal-trigeminal system by immunohistochemistry and western blotting analysis of GFAP protein expression, a typical marker of activated SGCs. A significant glial activation within the TG was observed starting from 24 hours up to 11 days after CFA injection, thus confirming that our model leads to TG sensitization. Moreover, upregulation of P2Y1 and P2Y2 protein expression was also observed, thus corroborating our in vitro data. Interestingly, the non-selective P2Y antagonist PPADS showed a strong analgesic effect on CFA-induced TG inflammation, which is comparable to ASA-mediated analgesia.
Conclusions - These results suggest a possible the pro-algogenic role for P2Y receptors in the development of trigeminal sensitization and migraine pain, opening the future perspective of identifying innovative and more selective pharmacological approaches for the sake of those migraineurs who are insensitive to currently available drugs.
References
[1] G. Magni, S. Ceruti. P2Y purinergic receptors: new targets for analgesic and antimigraine drugs. Biochem Pharmacol, 85(4):466-77, 2013.
[2] S. Ceruti, G. Villa, M. Fumagalli, L. Colombo, G. Magni, M. Zanardelli, E. Fabbretti, C. Verderio, A. M. J. M. van den Maagdenberg, A. Nistri, M. P. Abbracchio. CGRP−mediated enhancement of purinergic neuron/glia communication by the algogenic factor bradykinin in mouse trigeminal ganglia from wild type and R192Q Cav2.1 knock-in mice: implications for basic mechanisms of migraine pain. J Neuroscience, 31(10):3638 –3649, 2011
[3] G. Villa, S. Ceruti, M. Zanardelli, G. Magni, L. Jasmin, P. T. Ohara, M. P. Abbracchio.. Temporomandibular join inflammation activates glial and immune cells in both the trigeminal ganglia and the spinal trigeminal nucleus. Mol Pain, 6:89, 2010
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