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International audience; Nemaline myopathies are clinically and genetically heterogeneous muscle diseases characterized by the presence of nemaline bodies (rods) in muscle fibers. Mutations in the KLHL40 (kelch-like family member 40) gene (NEM 8) are common cause of severe/lethal nemaline myopathy. We report an 8-year-old girl born to consanguineous Moroccan parents, who presented with hypotonia and poor sucking at birth, delayed motor development, and further mild difficulties in walking and fatigability. A muscle biopsy revealed the presence of nemaline bodies. KLHL40 gene Sanger sequencing disclosed a never before reported pathogenic homozygous mutation which resulted in absent KLHL40 protein expression in the muscle. This further expands the phenotypical spectrum of KLHL40 related nemaline myopathy.

International audience; OBJECTIVES: Exertional Heat Stroke (EHS) is one of the top three causes of sudden death in athletes. Extrinsic and intrinsic risk factors have been identified but the genetic causes still remain unclear. Our aim was to identify genes responsible for EHS, which is a necessary step to identify patients at risk and prevent crises. DESIGN: Genetic and functional laboratory studies METHODS: Whole Exome Sequencing (WES) was performed to search for candidate genes in a cohort of 15 soldiers who had a documented EHS episode. In silico and in vitro functional studies were performed to evaluate the effect of mutations identified in the candidate gene TRPV1. RESULTS: WES led to the identification of two missense variations in the TRPV1 gene. These variations were very rare or unreported in control databases and located in critical domains of the protein. In vitro functional studies revealed that both variations induce a strong modification of the channel response to one of its natural agonist, the capsaicin. CONCLUSIONS: We evidenced mutations altering channel properties of the TRPV1 gene and demonstrated that TRPV1, which is involved in thermoregulation and nociception, is a new candidate gene for EHS. Our data provide the bases to explore genetic causes and molecular mechanisms governing the pathophysiology of EHS.

Purpose Glioblastoma is one of the most aggressive primary brain cancers. The precise grading of tumors is important to adopt the best follow-up treatment but complementary methods to histopathological diagnosis still lack in achieving an unbiased and reliable classification. Experimental design To progress in the field, a rapid Matrix Assisted Laser Desorption Ionization - Time of Flight Mass spectrometry (MALDI-TOF MS) protocole, devised for the identification and taxonomic classification of microorganisms and based on the analysis of whole cell extracts, was applied to glioma cell lines. Results The analysis of different human glioblastoma cell lines permitted to identify distinct proteomic profiles thus demonstrating the ability of MALDI-TOF to distinguish different malignant cell types. Conclusions and clinical relevance In the study, the authors showed the ability of MALDI-TOF profiling to discriminate glioblastoma cell lines, demonstrating that this technique could be used in complement to histological tumor classification. The proposed procedure is rapid and inexpensive and could be used to improve brain tumors classification and help propose a personalized and more efficient treatment.

Tubulin carboxypeptidase identity revealed Enzymes of the α-tubulin detyrosination/tyrosination cycle create landmarks on microtubules that are essential for their multiple cellular functions and are altered in disease. Tubulin carboxypeptidases (TCPs) responsible for detyrosination have remained elusive for 40 years (see the Perspective by Akhmanova and Maiato). Aillaud et al. identified vasohibins as enzymes that perform the TCP function and found that their small interacting partner SBVP was essential for their activity. Vasohibin/SVBP complexes were involved in neuron polarization and brain cortex development. The authors also developed an inhibitor targeting this family of enzymes. Using a completely different strategy, Nieuwenhuis et al. also showed that vasohibins can remove the C-terminal tyrosine of α-tubulin. Science , this issue p. 1448 , p. 1453 ; see also p. 1381

Synchrotron microbeam radiation therapy (MRT) relies on the spatial fractionation of a synchrotron beam into parallel micron-wide beams allowing deposition of hectogray doses. MRT controls the intracranial tumor growth in rodent models while sparing normal brain tissues. Our aim was to identify the early biological processes underlying the differential effect of MRT on tumor and normal brain tissues. The expression of 28,000 transcripts was tested by microarray 6 hr after unidirectional MRT (400 Gy, 50 µm-wide microbeams, 200 µm spacing). The specific response of tumor tissues to MRT consisted in the significant transcriptomic modulation of 431 probesets (316 genes). Among them, 30 were not detected in normal brain tissues, neither before nor after MRT. Areg, Trib3 and Nppb were down-regulated, whereas all others were up-regulated. Twenty-two had similar expression profiles during the 2 weeks observed after MRT, including Ccnb1, Cdc20, Pttg1 and Plk1 related to the mitotic role of the Polo-like kinase (Plk) pathway. The up-regulation of Areg expression may indicate the emergence of survival processes in tumor cells triggered by the irradiation; while the modulation of the "mitotic role of Plk1" pathway, which relates to cytokinetic features of the tumor observed histologically after MRT, may partially explain the control of tumor growth by MRT. The identification of these tumor-specific responses permit to consider new strategies that might potentiate the antitumoral effect of MRT.

A scalable hardware/software hybrid module--called Ubidule--endowed with bio-inspired ontogenetic and epigenetic features is configured to run a neural networks simulation with developmental and evolvable capabilities. We simulated the activity of hierarchically organized spiking neural networks characterized by an initial developmental phase featuring cell death followed by spike timing dependent synaptic plasticity in presence of background noise. An upstream 'sensory' network received a spatiotemporally organized external input and downstream networks were activated only via the upstream network. Precise firing sequences, formed by recurrent patterns of spikes intervals above chance levels, were observed in all recording conditions, thus suggesting the build-up of a connectivity able to sustain temporal information processing. The activity of a Ubinet--a network of Ubidules--is analyzed by means of virtual electrodes that recorded neural signals similar to EEG. The analysis of these signals was compared with a small set of human recordings and revealed common patterns of shift in quadratic phase coupling. The results suggest some interpretations of changes and plasticity of functional interactions between cortical areas driven by external stimuli and by learning/cognitive

Introduction RNA interference is efficient in in vitro studies, and appears as a therapeutic tool of major clinical interest. Nevertheless, the clinical utilisation of siRNAs is restrained by the poor availability of biodistribution data on this new class of pharmaceutics. This study aimed at defining the biodistribution and pharmacokinetics properties of an siRNA directed to the Casein Kinase-2 beta (CK2β) subunit, a potential target in cancer therapy. Methods Four CK2β siRNAs were chemically modified on each extremity of sense or anti-sense strand and radioiodinated. The biodistribution of each entity was analysed in glioblastoma-bearing mice using nuclear imaging and compared to a control GFP siRNA. Results The labelling process was associated with preservation of interference activity, except when applied to the 5 ' antisense terminus. Radioactivity was predominantly observed in organs of the excretory system after intravenous administration: liver, kidneys and bladder. Tumor/Contralateral muscle ratio showed significant differences depending on the labelling site. Activity associated with CK2β 5 ' s was quite constant over 2hours, while CK2β 3 ' as activity decreased by 40% in tumor. Finally, synchrotron X-ray analysis showed that CK2β 3 ' s is more abundant in tumor than in liver, brain or muscle, and uniformly distributed between intra- and extracellular compartments. Conclusions In this study, we highlighted the large influence of siRNAs radiolabelling position on their biodistribution and pharmacokinetic profiles, and proposed a systematic approach for the imaging of all siRNAs of clinical interest.

International audience; Asymmetrical cell division is a mechanism that gives rise to two daughter cells with different proliferative and differentiative fates. It occurs mainly during development and in adult stem cells. Accumulating evidence suggests that tumour cells arise from the transformation of normal stem cells. Here, we propose that the asymmetrical mitosis potential of stem cells is associated with the generation of migrating tumour progenitors. Application of this speculative model to glioma proposes that the sites where tumour-initiating stem cells reside are indolent and distinct from the tumour mass, and implies that the tumour mass is continuously replenished with new migrating tumour cells from these clinically silent regions. This hypothesis offers explanations for our inability to cure glioblastoma and points to asymmetrical division as a new potential therapeutic target.

Summary. We have previously described a culture system for human bone marrow endothelial cells that organize into capillary tubes associated to pericytes. In the present work, we used this model to assess the angiogenic properties of tachykinins, which have been demonstrated to be involved in neuro–immuno–haematopoietic interactions. The substance P (SP) and neurokinin A (NKA) were similarly potent at increasing in vitro angiogenesis, via NK1 and NK2 receptors respectively. These mediators were not produced by cells in culture, suggesting that in vivo they may be released by nerve fibres in the bone marrow. Therefore, we looked for in situ innervation of the human bone marrow, unknown to date, using immunohistochemistry techniques. As in rodents, arterioles were largely innervated, associated with between one and 10 nerve fibres. Capillary innervation was more restrictive as a unique thin nerve fibre was found in the vicinity of only 6% of these vessels. Finally, no nerve fibres were observed in the vicinity of sinus walls. In conclusion, both in vitro results and the anatomical display of nerve fibres suggest a role in human bone marrow for the vasoactive neuropeptides SP and NKA, which were secreted into a perivascular location. These neural mediators might modulate blood flow in the bone marrow both in the short term by adjusting vascular tone and in the long term by inducing angiogenesis.

International audience; Gliomas such as oligodendrogliomas (ODG) and glioblastomas (GBM) are brain tumours with different clinical outcomes. Histology-based classification of these tumour types is often difficult. Therefore the first aim of this study was to gain microRNA data that can be used as reliable signatures of oligodendrogliomas and glioblastomas. We investigated the levels of 282 microRNAs using membrane-array hybridisation and real-time PCR in ODG, GBM and control brain tissues. In comparison to these control tissues, 26 deregulated microRNAs were identified in tumours and the tissue levels of seven microRNAs (miR-21, miR-128, miR-132, miR-134, miR-155, miR-210 and miR-409-5p) appropriately discriminated oligodendrogliomas from glioblastomas. Genomic, epigenomic and host gene expression studies were conducted to investigate the mechanisms involved in these deregulations. Another aim of this study was to better understand glioma physiopathology looking for targets of deregulated microRNAs. We discovered that some targets of these microRNAs such as STAT3, PTBP1 or SIRT1 are differentially expressed in gliomas consistent with deregulation of microRNA expression. Moreover, MDH1, the target of several deregulated microRNAs, is repressed in glioblastomas, making an intramitochondrial-NAD reduction mediated by the mitochondrial aspartate-malate shuttle unlikely. Understanding the connections between microRNAs and bioenergetic pathways in gliomas may lead to identification of novel therapeutic targets.

International audience; The extremely low incidence of sporadic prion diseases suggests that they may arise as a rare stochastic event in otherwise healthy animals or humans. Current hypotheses for sporadic prion disease include horizontal transmission, spontaneous conversion of Prp C into Prp Sc , and somatic mutation of the Prp gene. Here, we suggest RNA mutation as a possible initial event in the etiology of sporadic prion disease. The proposed model is based on (i) the fact that in Prp-expressing cells, mutations are statistically more likely to occur in the Prp mRNA population than in the corresponding two copies of the Prp gene, and (ii) the absence of RNA repair mechanisms analagous to those found for DNA mismatch correction resulting in a relatively higher rate of RNA mutations. Here, we suggest that translation of mutated Prp mRNA could lead to the synthesis of transient Prp Sc which results in the conversion of Prp C into Prp Sc and the propagation of a disease-associated isoform. This model points to RNA mutation as a possible mechanism for the generation of sporadic prion diseases and other pathological disorders in which infectious proteins other than Prp Sc might be implicated.

We have previously shown that N6-methyldeoxyadenosine (MDA) is an inducer of differentiation in several tumor cells. Here we show that in addition to its ability to induce neurite-outgrowth in PC12 cells, MDA also significantly enhances the nerve-growth factor-mediated neurite outgrowth of these cells. Thus, MDA acts synergistically with NGF to repress cdc2 and cdk2 synthesis and to enhance tyrosine hydroxylase synthesis. To further elucidate the mechanisms of action of MDA, we investigated the effect of this drug on various signaling pathways. The neuritogenesis observed in PC12 following MDA treatment is mediated through activation of adenylyl cyclase in a PKA independent process and through the recruitment of the p44/p42 MAPK pathway. Furthermore, the adenosine A2a receptor antagonist ZM 241385 prevents the MDA-induced neuritogenesis, suggesting that MDA mediates its effect via this adenylyl cyclase-coupled A2a receptor. Collectively, these findings suggest that, in PC12 cells, the MDA-induced neuritogenesis requires the recruitment of adenosine A2a receptor, the stimulation of adenylate cyclase, and the activation of the p44/42MAP kinase cascade.

This study describes a human bone marrow endothelial cell culture in which endothelial cells are organized into capillary tubes. These endothelial cells were positive for von Willebrand Factor, expressed CD34, CD31, and L-fucose residues, took up acetylated low-density lipoproteins, contained Weibel-Palade bodies, and were ensheathed in a basal lamina (which included laminin beta1, EDa+ and EDb+ fibronectin, and collagen type iv). Pericytes expressing alpha-smooth muscle (alpha-SM) actin were spatially associated with the capillary tubes and there was a highly significant correlation between the number of capillary tubes and pericytes. In this model, basal angiogenesis was found to be vascular endothelial growth factor (VEGF)-dependent, because neutralization of endogenous VEGF induced a dramatic regression in the number of tubes. However, the presence of alpha-SM actin-expressing pericytes in the linings of endothelial tubes partially prevented the VEGF-neutralized tube regression. We also observed that nitric oxide production contributed to basal angiogenesis and that upregulation of nitric oxide increased the number of tubes. Tube numbers also decreased when antibodies neutralizing the integrin alphavbeta5 were applied to the cultures. Moreover, addition of any of the hematopoietic cytokines, erythropoietin, stem cell factor, granulocytic colony stimulating factor, or granulomonocytic colony stimulating factor induced a highly significant increase in tube formation. When erythropoietin and granulocytic colony stimulating factor were added, this increase was larger than the maximum increase observed with VEGF. Thus, we have described an in vitro model for human bone marrow angiogenesis in which pericytes and basal lamina matrix were associated with endothelial cells and formed fully organized capillary tubes. In this model, cytokines known to regulate hematopoiesis also seemed to be mediators of angiogenesis. This culture system may therefore prove to be a valuable tool for the study of hematopoietic cytokines on angiogenesis.

BACKGROUND: Vascular endothelial (VE)-cadherin is an endothelial specific adhesion molecule of vital importance for the maintenance of endothelium integrity. Because gliomas are hypervascular tumors, the present study was designed to address the question whether Vascular endothelial-cadherin (VE-cadherin), a protein exclusively expressed in endothelial cells, could be a potential candidate biomarker for clinical use in the setting of gliomas. METHODS: Fifty-six patients newly diagnosed glioma who underwent radiochemotherapy were enrolled in this retrospective study. Serum samples were collected prior treatment. Therapeutic response to Temodal was observed by RMI in 24 patients (25 oligodendroglioma, 2 glioblastoma multiform) median age 42.15 (range 23-65) while 32 patients were resistant (5 anaplastic oligodendroglioma, 9 anaplastic astrocytomas and 17 glioblastomas multiforme), median age 51.9 (range 19-84). sVE-cadherin levels was evaluated by westernblotting and sandwich ELISA. Data were analyzed using logistic regression and receiver operating characteristics (ROC) curve analysis. RESULTS: VE-cadherin 90 kDa extracellular domain was detected in human serum by western blot and its accurate detection was dependent upon a pre-dilution in a detergent-containing buffer (Triton X-100). Of importance, we found that sVE-cadherin was carried by human lipoproteins and that the detergent allowed to separate both entities to increase protein detectability. Interestingly, we found that sVE cadherin was significantly lower in the non responders group as compared to responders. We developped an ELISA that clearly discriminate patients NR (0.669+/-0.104 AU) from patients R (0.975+/-0.098 AU (p=0.0024). Receiver-operating characteristic (ROC) curve analysis revealed that the predictive cut-off value of serum VE-cadherin for chemoresistance was 0.735 AU (area-under-the-curve 0.82; 95% confidence interval 0.71-0.87; maximal accuracy 0.875) Patients lipid profile (total cholesterol, triglycerides, LDL and HDL) were not significantly different between the two groups and could not account for the differences observed in serum VE-cadherin levels. CONCLUSIONS: We show for the first time that the lipoprotein component of serum — which is most often ignored in protein diagnostics — contains proteins that can unexpectedly serve as biomarkers for disease. This finding may open completely new avenues of biomarker research. Second, we demonstrate, one candidate marker (soluble VE-cadherin) to follow glioma therapy. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2724.

International audience; BACKGROUND: Tissue invasion or tissue infiltration are clinical behaviors of a poor-prognosis subset of meningiomas. We carried out proteomic analyses of tissue extracts to discover new markers to accurately distinguish between infiltrative and noninfiltrative meningiomas. METHODOLOGY/PRINCIPAL FINDINGS: Protein lysates of 64 different tissue samples (including two brain-invasive and 32 infiltrative tumors) were submitted to SELDI-TOF mass spectrometric analysis. Mass profiles were used to build up both unsupervised and supervised hierarchical clustering. One marker was found at high levels in noninvasive and noninfiltrative tumors and appeared to be a discriminative marker for clustering infiltrative and/or invasive meningiomas versus noninvasive meningiomas in two distinct subsets. Sensitivity and specificity were 86.7% and 100%, respectively. This marker was purified and identified as a multiphosphorylated form of vimentin, a cytoskeletal protein expressed in meningiomas. CONCLUSIONS/SIGNIFICANCE: Specific forms of vimentin can be surrogate molecular indicators of the invasive/infiltrative phenotype in tumors.