Joel Eyer

Review on intermediate filaments of the nervous system and their pathological alterations

AbstractIntermediate filaments (IFs) of the nervous system, including neurofilaments, α-internexin, glial fibrillary acidic protein, synemin, nestin, peripherin and vimentin, are finely expressed following elaborated cell, tissue and developmental specific patterns. A common characteristic of several neurodegenerative diseases is the abnormal accumulation of neuronal IFs in cell bodies or along the axon, often associated with impairment of the axonal transport and degeneration of neurons. In this review, we also present several perturbations of IF metabolism and organization associated with neurodegenerative disorders. Such modifications could represent strong markers of neuronal damages. Moreover, recent data suggest that IFs represent potential biomarkers to determine the disease progression or the differential stages of a neuronal disorder. Finally, recent investigations on IF expression and function in cancer provide evidence that they may be useful as markers, or targets of brain tumours, especially high-grade glioma. A better knowledge of the molecular mechanisms of IF alterations, combined to neuroimaging, is essential to improve diagnosis and therapeutic strategies of such neurodegenerative diseases and glioma.

Photo-responsive polymer with erasable and reconfigurable micro- and nano-patterns: An in vitro study for neuron guidance

The interaction of cells with nanoscale topography has proven to be an important modality in controlling cell responses. Topographic parameters on material surfaces play a role in cell growth. We have synthesized a new bio compatible polymer containing photoswitching molecules. Stripepatterned (groove/ridge pattern) were patterned and erased with ease on this bio azopolymer with two different set-ups: one with the projection of an optical interference pattern and the other one by molecular self-organization with one single laser beam. These two set-ups allow the re-writing of pattern after erasing and its inscription in vitro. PC12 cells were cultured on the bio-photoswitching patterned polymer and compared with PC12 cells growing on a well know substrate: poly-L-lysine. This result is of interest for facilitating contact guidance and designing reconfigurable scaffold for neural network formation in vitro.

The NFL-TBS.40-63 anti-glioblastoma peptide enters selectively in glioma cells by endocytosis

Glioblastoma are the most frequent and aggressive tumour of the nervous system despite surgical resection associated with chemotherapy and radiotherapy. Recently, we showed that the NFL-TBS.40-63 peptide corresponding to the sequence of a tubulin-binding site of neurofilaments, enters selectively in glioblastoma cells where it blocks microtubule polymerization, inhibits their proliferation, and reduces tumour development in rats bearing glioblastoma (Bocquet et al., 2009; Berges et al., 2012a). Here, we characterized the molecular mechanism responsible for the uptake of NFL-TBS.40-63 peptide by glioblastoma cells. Unlike other cell penetrating peptides (CPPs), which use a balance between endocytosis and direct translocation, the NFL-TBS.40-63 peptide is unable to translocate directly through the membrane when incubated with giant plasma membrane vesicles. Then, using a panel of markers and inhibitors, flow cytometry and confocal microscopy investigations showed that the uptake occurs mainly through endocytosis. Moreover, glycosaminoglycans and αVβ3 integrins are not involved in the NFL-TBS.40-63 peptide recognition and internalization by glioblastoma cells. Finally, the signalling of tyrosine kinase receptors is involved in the peptide uptake, especially via EGFR overexpressed in tumour cells, indicating that the uptake of NFL-TBS.40-63 peptide by glioblastoma cells is related to their abnormally high proliferative activity.

The vimentin-tubulin binding site peptide (Vim-TBS.58-81) crosses the plasma membrane and enters the nuclei of human glioma cells

Cell-penetrating peptides (CPPs) can translocate through the plasma membrane and localize in different cell compartments providing a promising delivery system for peptides, proteins, nucleic acids, and other products. Here we describe features of a novel cell-penetrating peptide derived from the intermediate filament protein vimentin, called Vim-TBS.58-81. We show that it enters cells from a glioblastoma line via endocytosis where it distributes throughout the cytoplasm and nucleus. Moreover, when coupled to the pro-apoptogenic peptide P10, it localizes to the nucleus inhibiting cell proliferation. Thus, the Vim-TBS.58-81 peptide represents an effective vector for delivery of peptides and potentially a broad range of cargos to the nucleus.

NFL-lipid nanocapsules for brain neural stem cell targeting in vitro and in vivo.

The replacement of injured neurons by the selective stimulation of neural stem cells in situ represents a potential therapeutic strategy for the treatment of neurodegenerative diseases. The peptide NFL-TBS.40-63 showed specific interactions towards neural stem cells of the subventricular zone. The aim of our work was to produce a NFL-based drug delivery system able to target neural stem cells through the selective affinity between the peptide and these cells. NFL-TBS.40-63 (NFL) was adsorbed on lipid nanocapsules (LNC) whom targeting efficiency was evaluated on neural stem cells from the subventricular zone (brain) and from the central canal (spinal cord). NFL-LNC were incubated with primary neural stem cells in vitro or injected in vivo in adult rat brain (right lateral ventricle) or spinal cord (T10). NFL-LNC interactions with neural stem cells were different depending on the origin of the cells. NFL-LNC showed a preferential uptake by neural stem cells from the brain, while they did not interact with neural stem cells from the spinal cord. The results obtained in vivo correlate with the results observed in vitro, demonstrating that NFL-LNC represent a promising therapeutic strategy to selectively deliver bioactive molecules to brain neural stem cells.

The therapeutic contribution of nanomedicine to treat neurodegenerative diseases via neural stem cell differentiation.

The discovery of adult neurogenesis drastically changed the therapeutic approaches of central nervous system regenerative medicine. The stimulation of this physiologic process can increase memory and motor performances in patients affected by neurodegenerative diseases. Neural stem cells contribute to the neurogenesis process through their differentiation into specialized neuronal cells. In this review, we describe the most important methods developed to restore neurological functions via neural stem cell differentiation. In particular, we focused on the role of nanomedicine. The application of nanostructured scaffolds, nanoparticulate drug delivery systems, and nanotechnology-based real-time imaging has significantly improved the safety and the efficacy of neural stem cell-based treatments. This review provides a comprehensive background on the contribution of nanomedicine to the modulation of neurogenesis via neural stem cell differentiation.

The NFL-TBS.40-63 Anti-Glioblastoma Peptide Disrupts Microtubule and Mitochondrial Networks in the T98G Glioma Cell Line

Despite aggressive therapies, including combinations of surgery, radiotherapy and chemotherapy, glioblastoma remains a highly aggressive brain cancer with the worst prognosis of any central nervous system disease. We have previously identified a neurofilament-derived cell-penetrating peptide, NFL-TBS.40-63, that specifically enters by endocytosis in glioblastoma cells, where it induces microtubule destruction and inhibits cell proliferation. Here, we explore the impact of NFL-TBS.40-63 peptide on the mitochondrial network and its functions by using global cell respiration, quantitative PCR analysis of the main actors directing mitochondrial biogenesis, western blot analysis of the oxidative phosphorylation (OXPHOS) subunits and confocal microscopy. We show that the internalized peptide disturbs mitochondrial and microtubule networks, interferes with mitochondrial dynamics and induces a rapid depletion of global cell respiration. This effect may be related to reduced expression of the NRF-1 transcription factor and of specific miRNAs, which may impact mitochondrial biogenesis, in regard to default mitochondrial mobility.

Mycolacton: ein Bakterientoxin als zukünftiges Schmerzmittel?

Buruli Ulcer ist eine durch das Bakterium Mycobakterium ulcerans hervorgerufene Krankheit der Haut und Weichteile. Durch das Toxin Mycolacton entstehen dabei grose Gewebeschaden, die zu Beginn der Krankheit keine Schmerzen hervorrufen. Betroffene nehmen daher die Gewebelasionen erst ernst, wenn die Krankheit schon weit fortgeschritten ist. Wissenschaftler um Priscilla Brodin und Laurent Marsollier vom franzosischen Zentrum fur wissenschaftliche Forschung (CNRS) haben nun den molekularen Mechanismus dieser Analgesie aufgedeckt.

V - 2 Propriétés électrophysiologies des souris NFHGFP : intérêt pour l’étude de la régénération axonale

Introduction Pour l’etude du metabolisme des neurofilaments, une lignee de souris transgeniques (NFHGFP), exprimant une proteine de fusion entre la sous-unite NFH et une proteine fluorescente (GFP) a ete creee (Letournel et al. , 2006). Objectifs L’etude veut demontrer que l’expression de la proteine modifiee n’altere pas les vitesses de conduction nerveuse (VCN) et que la queue peut etre utilisee pour une telle etude. Methodes Nous avons enregistre les VCN motrice des nerfs sciatiques des souris NFHGFP (n = 15) et de souris controles (n = 13), âgees de 3 a 4 mois. Dans le meme temps ont ete mesurees les vitesses de conduction motrice sur la queue en adaptant une technique decrite chez le rat. Un bloc de conduction a ensuite ete cree par ecrasement (« crush ») de la queue et du nerf sciatique, avec suivi de son evolution clinique et electrophysiologique. Resultats Aucune difference significative des VCN n’a ete constatee entre les souris NFHGFP et controles au niveau du nerf sciatique (40 m/s SEM ± 0,78 ; 42,7 m/s SEM ± 1,07) ou de la queue (30,6 m/s SEM ± 0,64 ; 32 m/s SEM ± 0,46). De plus le rapport entre vitesse de conduction sur la queue et sur le nerf sciatique est reste stable (0,76). Finalement l’aspect du bloc de conduction etait comparable entre les deux groupes. Discussion Cette etude montre que les VCN des souris NFHGFP et controles, et leur evolution lors de la regeneration nerveuse, sont similaires. Ces resultats montrent l’absence d’anomalie fonctionnelle des nerfs en accord avec l’absence d’anomalie morphologique deja rapportee. Les resultats obtenus par la methode de mesure des vitesses de conduction sur la queue des souris apparaissent plus reproductibles que ceux obtenus sur le nerf sciatique. Conclusion Les souris NFHGFP peuvent etre utilisees pour l’etude des Neurofilaments lors de la regeneration axonale. L’analyse des VCN sur la queue donne des resultats plus fiables que sur le nerf sciatique.