Alpha-Synuclein Expression in the Oligodendrocyte Lineage: an In Vitro and In Vivo Study Using Rodent and Human Models.

In this study, we sought evidence for alpha-synuclein (ASYN) expression in oligodendrocytes, as a possible endogenous source of ASYN to explain its presence in glial inclusions found in multiple system atrophy (MSA) and Parkinson's disease (PD). We identified ASYN in oligodendrocyte lineage progenitors isolated from the rodent brain, in oligodendrocytes generated from embryonic stem cells, and in induced pluripotent stem cells produced from fibroblasts of a healthy individual and patients diagnosed with MSA or PD, in cultures in vitro. Notably, we observed a significant decrease in ΑSYN during oligodendrocyte maturation. Additionally, we show the presence of transcripts in PDGFRΑ/CD140a(+) cells and SOX10(+) oligodendrocyte lineage nuclei isolated by FACS from rodent and human healthy and diseased brains, respectively. Our work identifies ASYN in oligodendrocyte lineage cells, and it offers additional in vitro cellular models that should provide significant insights of the functional implication of ASYN during oligodendrocyte development and disease

Generation of human pluripotent stem cell reporter lines for the isolation of and reporting on astrocytes generated from ventral midbrain and ventral spinal cord neural progenitors

Astrocytes play a critical role during the development and the maintenance of the CNS in health and disease. Yet, their lack of accessibility from fetuses and from the brain of diseased patients has hindered our understanding of their full implication in developmental and pathogenic processes. Human pluripotent stem cells (PSCs) are an alternative source to obtain large quantities of astrocytes in vitro, for mechanistic studies of development and disease. However, these studies often require highly pure populations of astrocytes, which are not always achieved, depending on the PSC lines and protocols used. Here, we describe the generation and characterization of human PSC reporter lines expressing TagRFP driven by the ABC1D region of the human GFAP promoter, as new cellular model for generating homogenous population of astrocytes generated from CNS regionally defined PSC-derived neural progenitors. GFAABC1D::TagRFP-expressing astrocytes can be purified by fluorescent-activated cell sorting and maintain a bright expression for several additional weeks. These express canonical astrocyte markers NF1A, S100β, CX43, GLAST, GS and CD44. These new cellular models, from which highly pure populations of fluorescence-expressing astrocytes can be obtained, provide a new platform for studies where pure or fluorescently labeled astrocyte populations are necessary, for example to assess pro-inflammatory cytokine and chemokine release in response to specific treatment, and uptake and degradation of fluorescently labeled pathogenic proteins, as reported in this study

Creation of a library of induced pluripotent stem cells from Parkinsonian patients.

Induced pluripotent stem cells (iPSCs) are becoming an important source of pre-clinical models for research focusing on neurodegeneration. They offer the possibility for better understanding of common and divergent pathogenic mechanisms of brain diseases. Moreover, iPSCs provide a unique opportunity to develop personalized therapeutic strategies, as well as explore early pathogenic mechanisms, since they rely on the use of patients' own cells that are otherwise accessible only post-mortem, when neuronal death-related cellular pathways and processes are advanced and adaptive. Neurodegenerative diseases are in majority of unknown cause, but mutations in specific genes can lead to familial forms of these diseases. For example, mutations in the superoxide dismutase 1 gene lead to the motor neuron disease amyotrophic lateral sclerosis (ALS), while mutations in the SNCA gene encoding for alpha-synuclein protein lead to familial Parkinson's disease (PD). The generations of libraries of familial human ALS iPSC lines have been described, and the iPSCs rapidly became useful models for studying cell autonomous and non-cell autonomous mechanisms of the disease. Here we report the generation of a comprehensive library of iPSC lines of familial PD and an associated synucleinopathy, multiple system atrophy (MSA). In addition, we provide examples of relevant neural cell types these iPSC can be differentiated into, and which could be used to further explore early disease mechanisms. These human cellular models will be a valuable resource for identifying common and divergent mechanisms leading to neurodegeneration in PD and MSA

Leptin is required for hypothalamic regulation of miRNAs targeting POMC 3 ′ UTR

International audienceThe central nervous system (CNS) monitors modifications in metabolic parameters or hormone levels and elicits adaptive responses such as food intake regulation. Particularly, within the hypothalamus, leptin modulates the activity of pro-opiomelanocortin (POMC) neurons which are critical regulators of energy balance. Consistent with a pivotal role of the melanocortin system in the control of energy homeostasis, disruption of the POMC gene causes hyperphagia and obesity. MicroRNAs (miRNAs) are short noncoding RNA molecules that post-transcriptionally repress the expression of genes by binding to 3 ′-untranslated regions (3 ′ UTR) of the target mRNAs. However, little is known regarding the role of miRNAs that target POMC 3 ′ UTR in the central control energy homeostasis. Particularly, their interaction with the leptin signaling pathway remain unclear. First, we used common prediction programs to search for potential miRNAs target sites on 3 ′ UTR of POMC mRNA. This screening identified a set of conserved miRNAs seed sequences for mir-383, mir-384-3p, and mir-488. We observed that mir-383, mir-384-3p, and mir-488 are up-regulated in the hypothalamus of leptin deficient ob/ob mice. In accordance with these observations, we also showed that mir-383, mir-384-3p, and mir-488 were increased in db/db mice that exhibit a non-functional leptin receptor. The intraperitoneal injection of leptin down-regulated the expression of these miRNAs of interest in the hypothalamus of ob/ob mice showing the involvement of leptin in the expression of mir-383, mir-384-3p, and mir-488. Finally, the evaluation of responsivity to intracerebroventricular administration of leptin exhibited that a chronic treatment with leptin decreased mir-488 expression in hypothalamus of C57BL/6 mice. In summary, these results suggest that leptin modulates the expression of miRNAs that target POMC mRNA in hypothalamus

L'ablation chimique des neurones olfactifs : un modèle d'étude de la molécule RAE-1 induite dans les processus neuroinflammatoires et de neurogénèse

Les pathologies du SNC associent neurodégénérescence, troubles de la transmission et des processus neuroinflammatoires : réaction gliale et recrutement de cellules immunes. Les cellules immunes peuvent participer aux processus de souffrance neuronale et inhiber les processus de réparation comme la neurogenèse, mais peuvent également exercer des rôles neuroprotecteurs. Notre équipe a décrit que la molécule RAE-1, connue pour son rôle de ligand du récepteur activateur NKG2D exprimé par les NK, est exprimée constitutivement dans la SVZ, la zone de neurogenèse adulte qui génère des neuroblastes qui migrent vers le bulbe olfactif. De plus, RAE-1 est exprimée par les neurosphères issues de la SVZ et exerce un rôle non immun sur la prolifération cellulaire.L'objectif était de comprendre le rôle de RAE-1 et des NK en situation pathologique. J'ai étudié, en particulier, le modèle de lésion chimique des neurones olfactifs après injection de dichlobenil qui induit une mort neuronale suivie d'une réparation des réseaux neuronaux dans le bulbe olfactif. Ce modèle, connu des neurobiologistes, n'avait jamais été étudié sur le plan neuroinflammatoire. Nous avons caractérisé dans le bulbe olfactif, en RT-qPCR et cytométrie en flux multicouleur : la cinétique des processus de dé-afférentation, de ré-afférentation, de réaction gliale et le recrutement de cellules immunes. Je décris également l'induction de RAE-1 dans le bulbe olfactif, qui n'est pas liée à la présence d'une neurogenèse locale mais qui corrèle avec la prolifération microgliale et l'expression du M-CSF. Ces processus sont modifiés après un traitement à la minocycline et la déplétion des NK. Avec des modèles de culture in vitro, j'ai identifié RAE-1 comme un marqueur et un acteur de la microglie proliférative qui exerce, contrairement à la microglie non proliférative, des effets neuroprotecteurs.CNS pathologies are characterized by neurodegeneration, neurotransmission impairment and neuroinflammation including glial reactivity and immune cells recruitment. Neuroinflammation induces neuronal loss and impairs neurogenesis but can also exerts neuroprotective effects. Our team has recently demonstrated that RAE-1 is constitutively expressed in the adult subventricular zone (SVZ), the main neurogenic niche where resident neural stem cells generate neuroblasts which migrate in chains to the olfactory bulbs, where they differentiate into interneurons. We also have reported in vitro that RAE-1 is a marker of proliferating neural stem/progenitor cells and plays a non-immune role in cell proliferation. The main objective was to understand the role of RAE-1 and NK cells in pathological condition. We mainly studied a model of chemical ablation of olfactory neurons using dichlobenil which induced neuronal loss followed by an olfactory neuronal network recovery. Neuroinflammatory processes have never been studied in this model, used by neurobiologists. Using RT-qPCR and multicolor flow cytometry, I characterized the processes of deafferentation, reafferentation, astroglia and microglia activation and the recruitment of immune cells. Moreover, RAE-1 expression was induced in the olfactory bulbs. This expression correlated with the increase of microglial cell proliferation markers and M-CSF expression. These neuroinflammatory processes were altered after NK cells depletion and minocyline treatments. Using in vitro culture models, I showed that RAE-1 is a marker and an actor of microglia proliferation. Moreover, proliferative microglia but not non proliferative cells exerts a neuroprotective effect

An Emerging Role of micro-RNA in the Effect of the Endocrine Disruptors

International audienceEndocrine-disrupting chemicals (EDCs) are diverse natural and synthetic ă chemicals that may alter various mechanisms of the endocrine system and ă produce adverse developmental, reproductive, metabolic, and neurological ă effects in both humans and wildlife. Research on EDCs has revealed that ă they use a variety of both nuclear receptor-mediated and ă non-receptor-mediated mechanisms to modulate different components of the ă endocrine system. The molecular mechanisms underlying the effects of ă EDCs are still under investigation. Interestingly, some of the effects ă of EDCs have been observed to pass on to subsequent unexposed ă generations, which can be explained by the gametic transmission of ă deregulated epigenetic marks. Epigenetics is the study of heritable ă changes in gene expression that occur without a change in the DNA ă sequence. Epigenetic mechanisms, including histone modifications, DNA ă methylation, and specific micro-RNAs (miRNAs) expression, have been ă proposed to mediate transgenerational transmission and can be triggered ă by environmental factors. MiRNAs are short non-coding RNA molecules that ă post-transcriptionally repress the expression of genes by binding to ă 3'-untranslated regions of the target mRNAs. Given that there is ă mounting evidence that miRNAs are regulated by hormones, then clearly it ă is important to investigate the potential for environmental EDCs to ă deregulate miRNA expression and action

The Role of MicroRNA in the Modulation of the Melanocortinergic System

International audienceThe central control of energy balance involves a highly regulated neuronal network within the hypothalamus and the dorsal vagal complex. In these structures, pro-opiomelanocortin (POMC) neurons are known to reduce meal size and to increase energy expenditure. In addition, leptin, a peripheral signal that relays information regarding body fat content, modulates the activity of melanocortin pathway neurons including POMC-, Agouti-related peptide (AgRP)/Neuropeptide Y (NPY)-, melanocortin receptors (MC3R and MC4R)-expressing neurons. MicroRNAs (miRNAs) are short non-coding RNAs of 22–26 nucleotides that post-transcriptionally interfere with target gene expression by binding to their mRNAs. Evidence has demonstrated that miRNAs play important roles in the central regulation of energy balance. In this context, different studies identified miRNAs including miR-200 family, miR-103, or miR-488 that could target the genes of melanocortin pathway. More precisely, these different miRNAs can modulate energy homeostasis by affecting leptin transduction pathway in the POMC, or AgRP/NPY neurons. This article reviews the role of identified miRNAs in the modulation of melanocortin pathway in the context of energy homeostasis

Reporting on methods to generate and purify rodent and human oligodendrocytes from different sources

Oligodendrocytes are part of the glial cells located in the central nervous system, capable of providing trophic support to neurons and ensheathing their axons. These cells can become dysfunctional under pathologic condition. Rodent and human pluripotent stem cells are inexhaustible sources for producing oligodendrocytes that can be used for disease modeling and cell replacement therapy studies. They also offer many opportunities to model the contribution of oligodendrocytes in non-genetic disorders such as multiple system atrophy. In this method article, we provide robust and reproducible differentiation protocols to obtain oligodendrocyte progenitor cells and purify them using fluorescence activated cell sorting

RAE-1 expression is induced during experimental autoimmune encephalomyelitis and is correlated with microglia cell proliferation

International audienceRetinoic acid early induced transcript-1 (RAE-1) glycoproteins are ligands of the activating immune receptor NKG2D. They are known as stress molecules induced in pathological conditions. We previously reported that progenitor cells express RAE-1 in physiological conditions and we described a correlation between RAE-1 expression and cell proliferation. In addition, we showed that Raet1 transcripts are induced in the spinal cord of experimental autoimmune encephalomyelitis (EAE) mice. EAE is a model for multiple sclerosis which is accompanied by microglia proliferation and activation, recruitment of immune cells and neurogenesis. We herein studied the time course expression of the two members of the Raet1 gene family present in C57BL/6 mice, namely Raet1d and Raet1e, in the spinal cord during EAE. We report that Raet1d and Raet1e genes are induced early upon EAE onset and reach a maximal expression at the peak of the pathology. We show that myeloid cells, i.e. macrophages as well as microglia, are cellular sources of Raet1 transcripts. We also demonstrate that only Raet1d expression is induced in microglia, whereas macrophages expressed both Raet1d and Raet1e. Furthermore, we investigated the dynamics of RAE-1 expression in microglia cultures. RAE-1 induction correlated with cell proliferation but not with M1/M2 phenotypic orientation. We finally demonstrate that macrophage colony-stimulating factor (M-CSF) is a major factor controlling RAE-1 expression in microglia

Design of a 5G experimental platform based on OpenAirInterface

International audience5G has been designed for providing the appropriate services for a large range of applications requiring high throughput, low latency, a support for the IoT, or for Industry 4.0 business, etc. One of its strong statements is the sofwarization of most of its functions for providing more flexibility, and a support that can easily evolve for providing new services. In that context, OpenSource implementations of 5G functions arise. One of this implementation is the 5G OpenAirInterface (OAI). This paper then describes how a 5G experimental platform taking advantage of the 5G OAI software suite was designed and deployed at LAAS-CNRS. The aim of the platform is to be as generic as possible for being able to experiment and evaluate all 5G new mechanisms and protocols issued from researchers. This paper then specifically addresses how the compatibility issues between 5G OAI and the equipments (USRP, servers, operating systems, etc.) were fixed. The paper also proposes a performance evaluation of the 5G OAI platform and analyzes its limits