Attenuation of Reactive Gliosis Does Not Affect Infarct Volume in Neonatal Hypoxic-Ischemic Brain Injury in Mice

Astroglial cells are activated following injury and up-regulate the expression of the intermediate filament proteins glial fibrillary acidic protein (GFAP) and vimentin. Adult mice lacking the intermediate filament proteins GFAP and vimentin (GFAP(-/-)Vim(-/-)) show attenuated reactive gliosis, reduced glial scar formation and improved regeneration of neuronal synapses after neurotrauma. GFAP(-/-)Vim(-/-) mice exhibit larger brain infarcts after middle cerebral artery occlusion suggesting protective role of reactive gliosis after adult focal brain ischemia. However, the role of astrocyte activation and reactive gliosis in the injured developing brain is unknown.We subjected GFAP(-/-)Vim(-/-) and wild-type mice to unilateral hypoxia-ischemia (HI) at postnatal day 9 (P9). Bromodeoxyuridine (BrdU; 25 mg/kg) was injected intraperitoneally twice daily from P9 to P12. On P12 and P31, the animals were perfused intracardially. Immunohistochemistry with MAP-2, BrdU, NeuN, and S100 antibodies was performed on coronal sections. We found no difference in the hemisphere or infarct volume between GFAP(-/-)Vim(-/-) and wild-type mice at P12 and P31, i.e. 3 and 22 days after HI. At P31, the number of NeuN(+) neurons in the ischemic and contralateral hemisphere was comparable between GFAP(-/-)Vim(-/-) and wild-type mice. In wild-type mice, the number of S100(+) astrocytes was lower in the ipsilateral compared to contralateral hemisphere (65.0+/-50.1 vs. 85.6+/-34.0, p<0.05). In the GFAP(-/-)Vim(-/-) mice, the number of S100(+) astrocytes did not differ between the ischemic and contralateral hemisphere at P31. At P31, GFAP(-/-)Vim(-/-) mice showed an increase in NeuN(+)BrdU(+) (surviving newly born) neurons in the ischemic cortex compared to wild-type mice (6.7+/-7.7; n = 29 versus 2.9+/-3.6; n = 28, respectively, p<0.05), but a comparable number of S100(+)BrdU(+) (surviving newly born) astrocytes.Our results suggest that attenuation of reactive gliosis in the developing brain does not affect the hemisphere or infarct volume after HI, but increases the number of surviving newborn neurons

Astrocyte activation and reactive gliosis : A new target in stroke?

Stroke is an acute insult to the central nervous system (CNS) that triggers a sequence of responses in the acute, subacute as well as later stages, with prominent involvement of astrocytes. Astrocyte activation and reactive gliosis in the acute stage of stroke limit the tissue damage and contribute to the restoration of homeostasis. Astrocytes also control many aspects of neural plasticity that is the basis for functional recovery. Here, we discuss the concept of intermediate filaments (nanofilaments) and the complement system as two handles on the astrocyte responses to injury that both present attractive opportunities for novel treatment strategies modulating astrocyte functions and reactive gliosis.Peer reviewe

Astrocytes, reactive gliosis and CNS regeneration

The regenerative capacity in the adult central nervous system (CNS) is very limited, and the glial cells have been implicated in the inhibition of CNS regeneration. In essentially all CNS pathologies, astrocytes are activated and this process is known as reactive gliosis. Upregulation of intermediate filament proteins GFAP and vimentin and hypertrophy of cellular processes are the hallmarks of reactive gliosis. The focus of this thesis were astrocytes, in particular reactive astrocytes, and their role in regeneration in the adult CNS.Loss of GFAP, seen in many high grade astrocytomas, has been suspected to constitute a step in tumor development. By induction of astrocytomas in GFAP-/- mice in vivo, no difference in tumor development or progression was seen, suggesting that loss of GFAP seen in tumor cells rather reflects the undifferentiated state of these cells. Response to electrically induced injury of brain cortex and the hippocampus was assessed in knock-out mice depleted of neuron-derived PDGF-B, a possible modulator of reactive gliosis. We concluded that neuron-derived PDGF-B is not required for injury-triggered reactive gliosis, activation of microglia or neovascularization.GFAP-/-Vim-/- mice were previously reported to display less prominent post-traumatic glial scars. This animal model was used to study the integration of retinal grafts in the adult mouse retina. In contrast to wild-type, the GFAP-/-Vim-/- retinas provided a highly permissive environment for retinal transplants. Grafted cells migrated extensively into the host retina, differentiated into neurons and stayed integrated in the GFAP-/-Vim-/- retinas by six months after transplantation.To assess the role of reactive astrocytes in neuroprotection and regeneration after CNS trauma, GFAP-/-Vim-/- mice were subjected to a lesion partially denervating the dentate gyrus of the hippocampus. At an early stage after the lesion, GFAP-/-Vim-/- astrocytes provided less neuroprotection, but at a later time-point the same mice showed vastly improved regeneration of neuronal synapses as well as increased number of newborn neurons. Cell proliferation and neurogenesis in the hippocampus was also studied in healthy old GFAP-/-Vim-/- and wild-type mice, in which aging triggers mild and progressing reactive gliosis. Compared to wild-type, GFAP-/-Vim-/- mice showed increased cell proliferation and neurogenesis in the dentate gyrus of the hippocampus. This suggests that modulation of astrocyte reactivity increases neurogenesis even in the absence of a trauma or other challenge

Synemin is expressed in reactive astrocytes in neurotrauma and interacts differentially with vimentin and GFAP intermediate filament networks

Immature astrocytes and astrocytoma cells contain synemin and three other intermediate filament (IF) proteins: glial fibrillary acidic protein (GFAP), vimentin and nestin. Here, we show that, after neurotrauma, reactive astrocytes produce synemin and thus propose synemin as a new marker of reactive astrocytes. Comparison of synemin mRNA and protein levels in brain tissues and astrocyte cultures from wild-type, Vim-/- and Gfap-/-Vim-/- mice showed that in the absence of vimentin, synemin protein was undetectable although synemin mRNA was present at wild-type levels. By contrast, in Gfap-/- astrocytes, synemin protein and mRNA levels, as well as synemin incorporation into vimentin IFs, were unaltered. Biochemical assays with purified proteins suggested that synemin interacts with GFAP IFs like an IF-associated protein rather than like a polymerization partner, whereas the opposite was true for synemin interaction with vimentin. In transfection experiments, synemin did not incorporate into normal, filamentous GFAP networks, but integrated into vimentin and GFAP heteropolymeric networks. Thus, alongside GFAP, vimentin and nestin, reactive astrocytes contain synemin, whose accumulation is suppressed post-transcriptionally in the absence of a polymerization partner. In astrocytes, this partner is vimentin and not GFAP, which implies a functional difference between these two type III IF proteins

Astrocytoma grade IV (glioblastoma multiforme) displays 3 subtypes with unique expression profiles of intermediate filament proteins

Summary Astrocytoma grade IV (glioblastoma multiforme) is the most common and most malignant tumor of the central nervous system and is currently noncurable. Here, we have examined a population-based cohort of 47 patients with grade IV astrocytoma, who underwent tumor surgery at Sahlgrenska University Hospital in Sweden and who survived after surgery for less than 200 days (short survivors, 28 patients) and more than 500 days (long survivors, 19 patients). For each tumor, we ascertained information on patient age, sex, tumor location, oncological treatment, and survival after surgery. The analysis of the tumor volume and the extent of tumor resection (incomplete versus complete resection of the macroscopic tumor) was made retrospectively from the preoperative radiological investigations and, when available, also from postoperative radiology. We performed semiquantitative immunohistochemical evaluation of the presence of intermediate filament (nanofilament) proteins glial fibrillary acidic protein, vimentin, nestin, and synemin in tumor cells. The intermediate filament system helps cells and tissues to cope with various types of stress, and thus, it might affect the malignant potential of grade IV astrocytoma. We propose a subclassification of astrocytomas grade IV with respect to the expression of the intermediate filament proteins glial fibrillary acidic protein, vimentin, nestin, and synemin, namely, type A, B, and C. Our results suggest that the expression of the intermediate filament proteins glial fibrillary acidic protein, vimentin, nestin, and synemin is coregulated in grade IV astrocytomas. The expression patterns of the intermediate filament proteins in astrocytoma type A, B, and C might have biological and clinical significance. © 2013 Elsevier Inc

Synemin is expressed in reactive astrocytes and Rosenthal fibers in Alexander disease

Alexander disease (AxD) is a neurodegenerative disorder with prominent white matter degeneration and the presence of Rosenthal fibers containing aggregates of glial fibrillary acidic protein (GFAP), and small stress proteins HSP27 and αB-crystallin, and widespread reactive gliosis. AxD is caused by mutations in GFAP, the main astrocyte intermediate filament protein. We previously showed that intermediate filament protein synemin is upregulated in reactive astrocytes after neurotrauma. Here, we examined immunohistochemically the presence of synemin in reactive astrocytes and Rosenthal fibers in two patients with AxD. There was an abundance of GFAP-positive Rosenthal fibers and widespread reactive gliosis in the white matter and subpial regions. Many of the GFAP-positive reactive astrocytes were positive for synemin, and synemin was also present in Rosenthal fibers. We show that synemin is expressed in reactive astrocytes in AxD, and is also present in Rosenthal fibers. The potential role of synemin in AxD pathogenesis remains to be investigated. © 2013 APMIS Published by Blackwell Publishing Ltd

Etude des propriétés électroniques du graphène et des systèmes à base de graphène sous champs magnétiques intenses

Cette thèse présente des mesures de transport électronique dans des systèmes bi-dimensionels et uni-dimensionels à base de graphène sous champ magnétique pulsé (60T). L'objectif de ces travaux consiste à sonder la dynamique des porteurs de charge en modifiant la densité d'états du système par l'application d'un champ magnétique. Une première partie est consacrée à l'étude de l'influence des îlots électrons-trous sur les propriétés de transport du graphène au voisinage du point de neutralité de charge. Nous avons constaté l'apparition de fluctuations de la magnéto-résistance liée à la transition progressive des îlots de taille finie dans le régime quantique lorsque le champ magnétique augmente. Nous avons aussi montré que la variation de l'énergie de Fermi, liée à l'augmentation de la dégénérescence orbitale des niveaux de Landau, est directement responsable d'une modification du ratio entre électrons et trous. Dans une deuxième partie consacrée à l'étude des nanorubans de graphène, nous avons exploré deux gammes de largeur différentes. Dans les rubans larges (W>60nm), la quantification de la résistance a été observée révélant ainsi une signature évidente de la quantification du spectre énergétique en niveaux de Landau. Le confinement magnétique des porteurs de charge sur les bords des nanorubans a permis de mettre en évidence, pour la première fois, la levée de dégénérescence de vallée liée à la configuration armchair du ruban. Pour des rubans plus étroits (W<30nm), en présence de défauts de bord et d'impuretés chargées, la formation progressive des états de bords chiraux donne lieu à une magnéto-conductance positive quelque soit la densité de porteurs. Enfin, la dernière partie traite du magnéto-transport dans le graphene multi-feuillet. En particulier, nous avons observé l'effet Hall quantique dans les systèmes tri-couche de graphène. Une étude comparative des résultats expérimentaux avec des simulations numériques a permis de déterminer l'empilement rhombohedral des trois couches de graphene constituant l'échantillon.------------------------------------------------------------------------------------------------------------------------------------------This thesis presents transport measurements on two-dimensional and one-dimensional graphene-based systems under pulsed magnetic field (60T). The objective of this work is to probe the dynamics of charge carriers by changing the density of states of the system by applying a strong magnetic field. The first part is devoted to the study of the influence of electron-hole pockets on the transport properties of graphene near the charge neutrality point. We found the appearance of fluctuations in the magneto-resistance due to the progressive transition of the electron/hole puddles of finite size in the quantum regime as the magnetic field increases. We have also shown that the variation of the Fermi energy, due to the increase of orbital Landau level degeneracy, is directly responsible of a change in the electron and hole ratio. The second part is devoted to the study of graphene nano-ribbons, we explored two different ranges of width. In the broad nano-ribbons of width W larger than 60 nm, the quantification of the resistance is observed, revealing a clear signature of the quantization of the energy spectrum into Landau levels. We show for the first time the effect of valley degeneracy lifting induced by the magnetic confinement of charge carriers at the edges of the armchair nano-ribbons. For narrower nano-ribbons (W <30 nm) in presence of edge defects and charged impurities, the progressive formation of chiral edge states leads to a positive magneto-conductance whatever the carrier density. Finally, the last part of this thesis deals with magneto-transport fingerprints in multi-layer graphene as we observed the quantum Hall effect in tri-layer graphene. A comparative study of the experimental results with numerical simulations was used to determine the rhombohedral stacking of three layers of graphene in the sample

Neurofilament Light Chain (NfL) in Blood—A Biomarker Predicting Unfavourable Outcome in the Acute Phase and Improvement in the Late Phase after Stroke

Increased sensitivity of methods assessing the levels of neurofilament light chain (NfL), a neuron-specific intermediate filament protein, in human plasma or serum, has in recent years led to a number of studies addressing the utility of monitoring NfL in the blood of stroke patients. In this review, we discuss that elevated blood NfL levels after stroke may reflect several different neurobiological processes. In the acute and post-acute phase after stroke, high blood levels of NfL are associated with poor clinical outcome, and later on, the blood levels of NfL positively correlate with secondary neurodegeneration as assessed by MRI. Interestingly, increased blood levels of NfL in individuals who survived stroke for more than 10 months were shown to predict functional improvement in the late phase after stroke. Whereas in the acute phase after stroke the injured axons are assumed to be the main source of blood NfL, synaptic turnover and secondary neurodegeneration could be major contributors to blood NfL levels in the late phase after stroke. Elevated blood NfL levels after stroke should therefore be interpreted with caution. More studies addressing the clinical utility of blood NfL assessment in stroke patients are needed before the inclusion of NfL in the clinical workout as a useful biomarker in both the acute and the chronic phase after stroke