Mécanismes andogènes de contrôle des cellules souches du cerveau du mammifère nouveau-né et adulte

Dans le cerveau de mammifères adultes des cellules souches persistent au sein de la zone sous-ventriculaire (SVZ). Ces cellules prolifèrent et génèrent des neurones et des cellules gliales. Les cellules souches neurales contribuent au remplacement constitutif de neurones du bulbe olfactif et sont recrutées après lésion. Toutefois, le remplacement des neurones perdus est limité et les mécanismes sous-jacents restent à élucider. Notre objectif a été d identifier des facteurs diffusibles endogènes capables d influencer la prolifération, l autorenouvellement, la différenciation et la mort des cellules souches neurales. Ces facteurs ont été recherchés au sein, d une part, du cortex cérébral lésé et d autre part, du foie, un organe ayant d impressionnantes capacités de régénération. Nos résultats établissent que les facteurs libérés par le cortex suite à une lésion favorisent la prolifération des cellules de SVZ et la différenciation neuronale des précurseurs in vitro. Nous identifions le FGF-2 (Fibroblast Growth Factor) comme responsable des effets mitogènes du cortex cérébral lésé. Nous montrons aussi que les facteurs libérés par le tissu hépatique adulte augmentent la prolifération des cellules de la SVZ et orientent leur différenciation vers un phénotype glial radiaire. Notre étude identifie le HGF (Hepatocyte Growth Factor) comme responsable majeur de l effet prolifératif. Nous montrons que le récepteur du HGF, c-Met, est exprimé par les cellules immatures et que l administration de HGF in vitro ou in vivo augmente le nombre de cellules aux caractéristiques de souches dans la SVZ. De plus, nous démontrons que les cellules de SVZ libèrent du HGF qui agit de manière autocrine/paracrine sur les cellules souches neurales aussi bien in vitro que in vivo. L identification des régulateurs endogènes des cellules souches neurales est une étape majeure dans la mise en place de stratégies thérapeutiques basées sur l utilisation de ces cellules.Neural stem cells persist in the adult brain, within the subventricular zone (SVZ). These cells proliferate and generate glial and neuronal cells. Neural stem cells provide a constitutive supply of olfactory bulb interneurons and are recruited by brain injury. However, post-injury, neuronal replacement is limited and the mechanisms underlying this process are not fully elucidated. The major aim of our studies is the elucidation of the endogenous mechanisms regulating neural stem cell proliferation, differentiation and self-renewing. Our project was focused on two structures: the cerebral cortex after lesion and the liver, an organ that possesses important regenerative capacities. Our results established that apoptotic cortex explants release factors that promote SVZ cells proliferation and neuronal differentiation. Furthermore, Fibroblast Growth Factor-2 (FGF-2) was identified as responsible for the mitogenic effect. We also demonstrate that factors released by adult liver tissue increase SVZ cells proliferation and direct their phenotype towards a radial glial phenotype. The mitogenic effects of adult liver tissue were mainly due to Hepatocyte Growth Factor (HGF). We showed that HGF and c-Met are expressed in the SVZ and that HGF administration increases neural stem cells population expansion both in vitro and in vivo. Finally, we demonstrated that SVZ cells release HGF that acts on neural stem cells in a autocrine/paracrine way. The identification of endogenous mechanisms that regulate neural stem cells activity is a determinant step in the developpement of therapeutic strategies based on neural stem cells use.POITIERS-BU Sciences (861942102) / SudocSudocFranceF

Role of the calcium toolkit in cancer stem cells

International audienceCancer stem cells are a subpopulation of tumor cells that proliferate, self-renew and produce more differentiated tumoral cells building-up the tumor. Responsible for the sustained growth of malignant tumors, cancer stem cells are proposed to play significant roles in cancer resistance to standard treatment and in tumor recurrence. Among the mechanisms dysregulated in neoplasms, those related to Ca2+ play significant roles in various aspects of cancers. Ca2+ is a ubiquitous second messenger whose fluctuations of its intracellular concentrations are tightly controlled by channels, pumps, exchangers and Ca2+ binding proteins. These components support the genesis of Ca2+ signals with specific spatio-temporal characteristics that define the cell response. Being involved in the coupling of extracellular events with intracellular responses, the Ca2+ toolkit is often hijacked by cancer cells to promote notably their proliferation and invasion. Growing evidence obtained during the last decade pointed to a role of Ca2+ handling and mishandling in cancer stem cells. In this review, after a general overview of the concept of cancer stem cells we analyse and discuss the studies and current knowledge regarding the complex roles of Ca2+ toolkit and signaling in these cells. We highlight that numbers of Ca2+ signaling actors promote cancer stem cell state and are associated with cell resistance to current cancer treatments and thus may represent promising targets for potential clinical applications

Fonctions nouvelles de Gas-6 et de la protéine S

Les protéines vitamine K-dépendantes sont essentiellement connues pour leur implication dans la coagulation du sang. Récemment, deux protéines vitamine K-dépendantes, le facteur anticoagulant protéine S et son homologue structural Gas-6, ont été identifiés comme ligands des récepteurs à activité tyrosine kinase TAM (Tyro-3/Axl/Mer). L’analyse des phénotypes de souris transgéniques invalidées pour les gènes codant pour Gas-6 ou pour ses récepteurs a révélé que Gas-6 et la protéine S sont impliquées dans la régulation de la phagocytose des cellules apoptotiques, processus important dans la réponse immunitaire ainsi que dans les processus de différenciation cellulaire. Les nouvelles fonctions de ces protéines vitamine K-dépendantes ainsi que leur utilisation potentielle dans l’élaboration de traitements de pathologies associées à un déficit de phagocytose font l’objet de cet article

Calcium Channels in Adult Brain Neural Stem Cells and in Glioblastoma Stem Cells

International audienceThe brain of adult mammals, including humans, contains neural stem cells (NSCs) located within specific niches of which the ventricular-subventricular zone (V-SVZ) is the largest one. Under physiological conditions, NSCs proliferate, self-renew and produce new neurons and glial cells. Several recent studies established that oncogenic mutations in adult NSCs of the V-SVZ are responsible for the emergence of malignant primary brain tumors called glioblastoma. These aggressive tumors contain a small subpopulation of cells, the glioblastoma stem cells (GSCs), that are endowed with proliferative and self-renewal abilities like NSCs from which they may arise. GSCs are thus considered as the cells that initiate and sustain tumor growth and, because of their resistance to current treatments, provoke tumor relapse. A growing body of studies supports that Ca2+ signaling controls a variety of processes in NSCs and GSCs. Ca2+ is a ubiquitous second messenger whose fluctuations of its intracellular concentrations are handled by channels, pumps, exchangers, and Ca2+ binding proteins. The concerted action of the Ca2+ toolkit components encodes specific Ca2+ signals with defined spatio-temporal characteristics that determine the cellular responses. In this review, after a general overview of the adult brain NSCs and GSCs, we focus on the multiple roles of the Ca2+ toolkit in NSCs and discuss how GSCs hijack these mechanisms to promote tumor growth. Extensive knowledge of the role of the Ca2+ toolkit in the management of essential functions in healthy and pathological stem cells of the adult brain should help to identify promising targets for clinical applications

The vitamin K-dependent factor, protein S, regulates brain neural stem cell migration and phagocytic activities towards glioma cells

International audienceMalignant gliomas are the most common primary brain tumors. Due to both their invasive nature and resistance to multimodal treatments, these tumors have a very high percentage of recurrence leading in most cases to a rapid fatal outcome. Recent data demonstrated that neural stem/progenitor cells possess an inherent ability to migrate towards glioma cells, track them in the brain and reduce their growth. However, mechanisms involved in these processes have not been explored in-depth. In the present report, we investigated interactions between glioma cells and neural stem/progenitor cells derived from the subventricular zone, the major brain stem cell niche. Our data show that neural stem/pro-genitor cells are attracted by cultured glioma-derived factors. Using multiple approaches, we demonstrate for the first time that the vitamin K-dependent factor protein S produced by glioma cells is involved in tumor tropism through a mechanism involving the tyrosine kinase receptor Tyro3 that, in turn, is expressed by neural stem/ progenitor cells. Neural stem/progenitor cells decrease the growth of both glioma cell cultures and clonogenic population. Cultured neural stem/progenitor cells also engulf, by phagocytosis, apoptotic glioma cell-derived fragments and this mechanism depends on the exposure of phosphatidylserine eat-me signal and is stimulated by protein S. The disclosure of a role of protein S/Tyro3 axis in neural stem/progenitor cell tumor-tropism and the demonstration of a phagocytic activity of neural stem/progenitor cells towards dead glioma cells that is regulated by protein S open up new perspectives for both stem cell biology and brain physiopathology

Aberrant expression and localization of connexin43 and connexin30 in a rat glioma cell line

International audienceGap junctions are cellular structures which permit direct exchanges of small molecules from cytoplasm to cytoplasm in most of the cells of metazoan organisms. For four decades, it has been observed that the inhibition of this type of intercellular communication is often associated with tumorigenesis. The assumption that loss of homeostasis which characterizes tumor growth could be a consequence of a lack of gap junctional intercellular communication (GJIC) has been reinforced by strategies able to reinduce both GJIC and normalization of the phenotype. So far, no molecular data may explain clearly how gap junctions can regulate cell proliferation. It has been argued that the gap-junction tumor suppressive effect may depend specifically on the connexin type which is expressed. For instance, the transfection of connexin30 (Cx30), a gap junction protein, has been previously associated with a slower growth of rat glioma cells (9L cells). Here, we show that these cells do communicate less compared to the Cx43-expressing parental cells even if the Cx30-transfected cells do express more Cx43. This result was related to the cytoplasmic distribution of Cx43 and a nuclear localization of both the Cx30 and a 20-kDa fragment corresponding to a Cx43 signal. According to these data, it seems that cell growth regulation may depend more on the behavior of connexins than the simple establishment of GJIC

Characterization of neural stem cells in the dorsal vagal complex of adult rat by in vivo proliferation labeling and in vitro neurosphere assay

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Leptin-dependent neurotoxicity via induction of apoptosis in adult rat neurogenic cells

International audienceAdipocyte-derived hormone leptin has been recently implicated in the control of neuronal plasticity. To explore whether modulation of adult neurogenesis may contribute to leptin control of neuronal plasticity, we used the neurosphere assay of neural stem cells derived from the adult rat subventricular zone (SVZ). Endogenous expression of specific leptin receptor (ObRb) transcripts, as revealed by RT-PCR, is associated with activation of both ERK and STAT-3 pathways via phosphorylation of the critical ERK/STAT-3 amino acid residues upon addition of leptin to neurospheres. Furthermore, leptin triggered withdrawal of neural stem cells from the cell cycle as monitored by Ki67 labeling. This effect was blocked by pharmacological inhibition of ERK activation thus demonstrating that ERK mediates leptin effects on neural stem cell expansion. Leptin-dependent withdrawal of neural stem cells from the cell cycle was associated with increased apoptosis, as detected by TUNEL, which was preceded by cyclin D1 induction. Cyclin D1 was indeed extensively colocalized with TUNEL-positive, apoptotic nuclei. Cyclin-D1 silencing by specific shRNA prevented leptin-induced decrease of the cell number per neurosphere thus pointing to the causal relationship between leptin actions on apoptosis and cyclin D1 induction. Leptin target cells in SVZ neurospheres were identified by double TUNEL/phenotypic marker immunocytofluorescence as differentiating neurons mostly. The inhibition of neural stem cell expansion via ERK/cyclin D1-triggered apoptosis defines novel biological action of leptin which may be involved in adiposity-dependent neurotoxicity

Store-Operated Calcium Entries Control Neural Stem Cell Self-Renewal in the Adult Brain Subventricular Zone

International audienceThe subventricular zone (SVZ) is the major stem cell niche in the brain of adult mammals. Within this region, neural stem cells (NSC) proliferate, self-renew and give birth to neurons and glial cells. Previous studies underlined enrichment in calcium signaling-related transcripts in adult NSC. Because of their ability to mobilize sustained calcium influxes in response to a wide range of extracellular factors, store-operated channels (SOC) appear to be, among calcium channels, relevant candidates to induce calcium signaling in NSC whose cellular activities are continuously adapted to physiological signals from the microenvironment. By Reverse Transcription Polymerase Chain Reaction (RT-PCR), Western blotting and immunocytochemistry experiments, we demonstrate that SVZ cells express molecular actors known to build up SOC, namely transient receptor potential canonical 1 (TRPC1) and Orai1, as well as their activator stromal interaction molecule 1 (STIM1). Calcium imaging reveals that SVZ cells display store-operated calcium entries. Pharmacological blockade of SOC with SKF-96365 or YM-58483 (also called BTP2) decreases proliferation, impairs self-renewal by shifting the type of SVZ stem cell division from symmetric proliferative to asymmetric, thereby reducing the stem cell population. Brain section immunostainings show that TRPC1, Orai1, and STIM1 are expressed in vivo, in SOX2-positive SVZ NSC. Injection of SKF-96365 in brain lateral ventricle diminishes SVZ cell proliferation and reduces the ability of SVZ cells to form neurospheres in vitro. The present study combining in vitro and in vivo approaches uncovers a major role for SOC in the control of SVZ NSC population and opens new fields of investigation for stem cell biology in health and disease. Stem Cells 2018