Littérature scientifique sur le sujet « Meninges, neural stem cells, postnatal neurogenesis »

Neural stem cells are capable of generating new neurons during development as well as in the adulthood and represent one of the most promising tools to replace lost or damaged neurons after injury or neurodegenerative disease. Unlike the brain, neurogenesis in the adult spinal cord is poorly explored and the comprehensive characterization of the cells that constitute stem cell neurogenic niche is still missing. Moreover, the terminology used to specify developmental and/or anatomical CNS regions, where neurogenesis in the spinal cord occurs, is not consensual and the analogy with the brain is often unclear. In this review, we will try to describe the heterogeneity of the stem cell types in the spinal cord ependymal zone, based on their origin and stem cell potential. We will also consider specific animal in vitro models that could be useful to identify “the right” stem cell candidate for cell replacement therapies.

Adult neurons are believed to be in a state of growth arrest. The generation of neurons is complete at the time of birth in most of the brain regions. However neurogenesis is present through life in the dentate gyrus of hippocampus and the lateral ventricles due to the presence of neural stem cells (NSC). This postnatal neurogenesis in hippocampus plays a critical role in cognitive development mainly in learning and memory functions. NSC are self-renewing, multipotent cells that generate the neurons and glia of the nervous system. Due to their high proliferation, NSC are highly sensitive to ionizing radiation. This review describes the current knowledge on impact of ionizing radiation on neural stem cells biology. Widening the knowledge of mechanisms involved in radiation-induced neurotoxicity at the level of NSC may help to overcome in the future the side effects occurring after anti-cancer therapies of the brain and help to protect and maintain neurogenesis.

Stem cells of the central nervous system have received a great deal of attention in neurobiology in the last decade. It has been shown that neurogenesis occurs in the postnatal period in specialized niches of the adult mammalian brain. The niche is a key regulator of stem cell behavior. Recent data underscore the complexity and heterogeneity of the different components of the niche, and the presence of local signaling microdomain. The review is devoted to recent views on the structural organization of neurogenic niches and regulatory factors involved at different stages of neurogenesis in the postnatal period. Understanding of stem cells behavior in the niches can serve as a basis for determination of these cells function in the adult brain.

Neural stem cells continuously generate newborn neurons that integrate into and modify neural circuitry in the adult hippocampus. The molecular mechanisms that regulate or perturb neural stem cell proliferation and differentiation, however, remain poorly understood. Here, we have found that mouse hippocampal radial glia-like (RGL) neural stem cells express the synaptic cochaperone cysteine string protein-α (CSP-α). Remarkably, in CSP-α knockout mice, RGL stem cells lose quiescence postnatally and enter into a high-proliferation regime that increases the production of neural intermediate progenitor cells, thereby exhausting the hippocampal neural stem cell pool. In cell culture, stem cells in hippocampal neurospheres display alterations in proliferation for which hyperactivation of the mechanistic target of rapamycin (mTOR) signaling pathway is the primary cause of neurogenesis deregulation in the absence of CSP-α. In addition, RGL cells lose quiescence upon specific conditional targeting of CSP-α in adult neural stem cells. Our findings demonstrate an unanticipated cell-autonomic and circuit-independent disruption of postnatal neurogenesis in the absence of CSP-α and highlight a direct or indirect CSP-α/mTOR signaling interaction that may underlie molecular mechanisms of brain dysfunction and neurodegeneration.

Endogenous DNA double-strand breaks (DSBs) formation and repair in neural stem/progenitor cells (NSPCs) play fundamental roles in neurogenesis and neurodevelopmental disorders. NSPCs exhibit heterogeneity in terms of lineage fates and neurogenesis activity. Whether NSPCs also have heterogeneous regulations on DSB formation and repair to accommodate region-specific neurogenesis has not been explored. Here, we identified a regional regulator Filia, which is predominantly expressed in mouse hippocampal NSPCs after birth and regulates DNA DSB formation and repair. On one hand, Filia protects stalling replication forks and prevents the replication stress-associated DNA DSB formation. On the other hand, Filia facilitates the homologous recombination–mediated DNA DSB repair. Consequently, Filia−/− mice had impaired hippocampal NSPC proliferation and neurogenesis and were deficient in learning, memory, and mood regulations. Thus, our study provided the first proof of concept demonstrating the region-specific regulations of DSB formation and repair in subtypes of NSPCs.

Two main stem cell pools exist in the postnatal mammalian brain that, although they share some “stemness” properties, also exhibit significant differences. Multipotent neural stem cells survive within specialized microenvironments, called niches, and they are vulnerable to ageing. Oligodendroglial lineage-restricted progenitor cells are widely distributed in the brain parenchyma and are more resistant to the effects of ageing. Here, we create polymorphic neural stem cell cultures and allow cells to progress towards the neuronal and the oligodendroglial lineage. We show that the divergence of cell fate is accompanied by a divergence in the properties of progenitors, which reflects their adaptation to life in the niche or the parenchyma. Neurogenesis shows significant spatial restrictions and a dependence on laminin, a major niche component, while oligodendrogenesis shows none of these constraints. Furthermore, the blocking of integrin-β1 leads to opposing effects, reducing neurogenesis and enhancing oligodendrogenesis. Therefore, polymorphic neural stem cell assays can be used to investigate the divergence of postnatal brain stem cells and also to predict the in vivo effects of potential therapeutic molecules targeting stem and progenitor cells, as we do for the microneurotrophin BNN-20.

While various modalities of chronic nicotine use have been associated with numerous negative consequences to human health, one possible benefit of nicotine exposure has been uncovered. The discovery of an inverse correlation between smoking and Parkinson’s disease, and later Alzheimer’s disease as well, motivated investigation of nicotine as a neuroprotective agent. Some studies have demonstrated that nicotine elicits improvements in cognitive function. The hippocampus, along with the subventricular zone (SVZ), is a distinct brain region that allow for ongoing postnatal neurogenesis throughout adulthood and plays a major role in certain cognitive behaviors like learning and memory. Therefore, one hypothesis underlying nicotine-induced neuroprotection is possible effects on neural stem cells and neural precursor cells. On the other hand, nicotine withdrawal frequently leads to cognitive impairments, particularly in hippocampal-dependent behaviors, possibly suggesting an impairment of hippocampal neurogenesis with nicotine exposure. This review discusses the current body of evidence on nicotine’s effects on neural stem cells and neural progenitors. Changes in neural stem cell proliferation, survival, intracellular dynamics, and differentiation following acute and chronic nicotine exposure are examined.

During the last two decades basic research in neuroscience has remarkably expanded due to the discovery of neural stem cells (NSCs) and adult neurogenesis in the mammalian central nervous system (CNS). The existence of such unexpected plasticity triggered hopes for alternative approaches to brain repair, yet deeper investigation showed that constitutive mammalian neurogenesis is restricted to two small “neurogenic sites” hosting NSCs as remnants of embryonic germinal layers and subserving homeostatic roles in specific neural systems. The fact that in other classes of vertebrates adult neurogenesis is widespread in the CNS and useful for brain repair sometimes creates misunderstandings about the real reparative potential in mammals. Nevertheless, in the mammalian CNS parenchyma, which is commonly considered as “nonneurogenic,” some processes of gliogenesis and, to a lesser extent, neurogenesis also occur. This “parenchymal” cell genesis is highly heterogeneous as to the position, identity, and fate of the progenitors. In addition, even the regional outcomes are different. In this paper the heterogeneity of mammalian parenchymal neurogliogenesis will be addressed, also discussing the most common pitfalls and misunderstandings of this growing and promising research field.

Abstract It remains unknown whether H3K4 methylation, an epigenetic modification associated with gene activation, regulates fate determination of the postnatal neural stem and progenitor cells (NSPCs). By inactivating the Dpy30 subunit of the major H3K4 methyltransferase complexes in specific regions of mouse brain, we demonstrate a crucial role of efficient H3K4 methylation in maintaining both the self-renewal and differentiation capacity of postnatal NSPCs. Dpy30 deficiency disrupts development of hippocampus and especially the dentate gyrus and subventricular zone, the major regions for postnatal NSC activities. Dpy30 is indispensable for sustaining the self-renewal and proliferation of NSPCs in a cell-intrinsic manner and also enables the differentiation of mouse and human neural progenitor cells to neuronal and glial lineages. Dpy30 directly regulates H3K4 methylation and the induction of several genes critical in neurogenesis. These findings link a prominent epigenetic mechanism of gene expression to the fundamental properties of NSPCs and may have implications in neurodevelopmental disorders.

MicroRNAs are short RNA molecules composed of 20-22 nucleotides. They highly accurately indicate cell identity and hence they are useful in labeling cells and tacking lineage commitment. However, this requires accurate microRNA profiling of cells in individual developmental stages. Since microRNAs are important negative regulators of eukaryotic gene expression, microRNA profiling allows better understanding of molecular regulatory networks of important cellular events, such as adult neurogenesis. Adult neurogenesis is the process in which neurons, as well as glia, are generated from neural stem cells. It was found to be responsible for brain regeneration, olfactory discrimination, memory formation and learning. Depression was suggested to be related to dysregulation of neurogenesis. Thus, knowledge in cellular and molecular mechanisms of adult neurogenesis will lay solid foundation to develop therapies to regenerate neural cells after injuries or onsets of neurodegenerative diseases and to understand the cognitive ability, memory formation and learning of the brain. In spite of its importance, investigation into the miRNA profiles and functions in neurogenesis is still infant. This project aimed to establish a preliminary microRNA profile on neurogenesis. Although this was not completed, the project could be extended to a large-scale microRNA profiling in neurogenesis. This would enable future workers to track the lineage commitment, the migration, and the distribution of NSCs and their derived cells accurately by in situ hybridization. Also, the future workers may construct a 2D representation of the changes in miRNA profiles and this may lead to discovery of previously unknown molecular and cellular differences among cells of same cell identity.
published_or_final_version
Anatomy
Master
Master of Medical Sciences

Les malformations du cortex cérébral (MDC) représentent une cause importante de handicap et d'épilepsie pharmaco-résistante. Le séquençage à haut débit a permis une amélioration considérable de l'identification des bases moléculaires des MDC non syndromiques. Toutefois, certaines formes, notamment les MDC complexes, demeurent inexpliquées. Mon projet de thèse a pour objectif de progresser dans la compréhension des MDC complexes en utilisant deux modèles : les microlissencéphalies (MLIS) et le syndrome d'Aicardi (AIC), une forme syndromique particulière associant des malformations de l'oeil et du cerveau uniquement rapporté chez les filles. L'étude par séquençage d'exome en trios de 16 familles MLIS m'a permis d'identifier et de caractériser un nouveau gène, WDR81, impliqué dans le cycle cellulaire. Par la même stratégie, j'ai pu identifier un variant homozygote pathogène dans TLE1, un partenaire majeur de FOXG1 dans la balance prolifération/différenciation de progéniteurs neuronaux, dans une famille consanguine de microcéphalie postnatale dont le phénotype est proche du syndrome FOXG1. En parallèle, mes travaux ont permis de préciser les spectres phénotypiques associés à RTTN, EPG5, COL4A1, COL4A2, TBR1, KIF5C, KIF2A et FOXG1. La deuxième partie de mon projet avait pour objet l'identification des bases moléculaires du syndrome d'Aicardi à partir d'une cohorte internationale de 19 patientes. Après avoir exclu un biais d'inactivation du chromosome X et la présence de microremaniements chromosomiques, j'ai réalisé un séquençage d'exome en trio. Aucun variant récurrent n'a été retrouvé dans les séquences codantes. Dans un second temps, j'ai testé une approche combinant les données du séquençage de génome et l'analyse du transcriptome (RNA-Seq) sur fibroblastes, me permettant d'identifier des transcrits dérégulés qui étaient impliqués dans le développement du cerveau et de l'oeil. J'ai comparé les résultats de cette analyse avec ceux de l'analyse du génome dans le but d'identifier des variants dans ces gènes candidats. En conclusion, mon travail de thèse a permis d'améliorer la connaissance des bases moléculaires des MDC complexes et d'ouvrir des perspectives de nouveaux mécanismes tels que ceux engageant les gènes WDR81 et EPG5, et le rôle des endosomes et de l'autophagie dans les MDC, et aussi TLE1 comme nouvelle cause de microcéphalies postnatales. Mes travaux ont également permis de générer une collection de données de séquençage haut débit (WES, WGS et RNA-Seq) qui seront mises en commun dans le cadre d'un consortium international afin de développer des nouvelles stratégies d'analyse en particulier pour les séquences non codantes. Cette approche permettra également d'ouvrir la voie vers la compréhension des mécanismes cellulaires impliqués dans la formation du cerveau et de l' œil
Malformations of cortical development (MCD) are a major cause of intellectual disability and drug-resistant epilepsy. Next Generation Sequencing (NGS) has considerably improved the identification of the molecular basis of non-syndromic MCD. However, certain forms, including complex MCD, remain unexplained. My PhD project aimed to improve the understanding of complex MCD using two disorders: Microlissencephaly (MLIS) and Aicardi Syndrome (AIC), the latter associating brain and eye malformations and only reported in girls. Trio Whole Exome Sequencing (WES) performed in 16 MLIS families allowed me to identify and functionally characterize a new MLIS gene, WDR81, in which mutations lead to cell cycle alteration. Moreover, using the same strategy, I was able to identify a pathogenic homozygous variant in TLE1 in a patient from consanguineous family with a postnatal microcephaly, suggestive of a FOXG1-like presentation. Interestingly, TLE1 is a major partner of FOXG1, a gene involved in maintaining the balance between progenitor proliferation and differentiation. In parallel, my work allowed me to redefine the phenotypic spectrum associated with RTTN, EPG5, COL4A1 and COL4A2, TBR1, KIF5C, KIF2A and FOXG1. The second part of my PhD program was aimed at identifying the genetic basis of AIC in an international cohort of 19 patients. After excluding a skewed X chromosome inactivation and the presence of chromosomal rearrangements, I performed WES in trios. The analysis of the data from WES did not allow me to identify any recurrent variants. I therefore tested a new approach combining Whole Genome Sequencing (WGS) and RNA-Sequencing (RNA-Seq) on fibroblast cells. I identified a number of deregulated transcripts implicated in brain and eye development. I compared the results of this analysis with the WGS analysis in order to find variants in these candidate genes. In conclusion, these studies have improved the knowledge of the molecular basis of complex MCD, such as TLE1 in postnatal microcephaly, and revealed the pathogenic mechanisms such as WDR81 in cell cycle progression and EPG5 in endosomes and autophagy. My work has also generated a collection of NGS data (WES, WGS and RNA-Seq) that will be shared in an international consortium to develop new analytical strategies, in particular for the non-coding DNA regions. This novel strategy provides opportunities to improve understanding of the cellular mechanisms involved in brain and eye development

La neurogenesi nel cervello dei mammiferi prosegue durante il corso di tutta la vita (Eriksson et al., 1998; Gage, 2000) in due nicchie germinative: la zona sottoventricolare e il giro dentato dell’ippocampo (Gage and Temple, 2013). Le cellule della glia radiale (Kriegstein and Alvarez-Buylla, 2009) sono le cellule staminali che, durante lo sviluppo embrionale e postnatale, danno origine a diversi tipi cellulari tra cui neuroblasti, neuroni, oligodendrociti, astrociti e cellule ependimali (Kriegstein and Alvarez-Buylla, 2009). Oltre a queste nicchie di cellule staminali ampiamente caratterizzate, è stata descritta l’esistenza di nicchie neurali ectopiche in seguito a condizioni traumatiche (Pluchino et al., 2010) e in condizioni fisiologiche in regioni specifiche, come la retina, il cervelletto e i bulbi olfattivi (Menezes et al., 1995; Ponti et al., 2008; Tropepe et al., 2000). Diversi gruppi hanno recentemente identificato una nuova funzione per le meningi, da semplice struttura protettiva del sistema nervoso centrale a nicchia contenente cellule staminali endogene. E’ stato infatti dimostrato che le meningi contengono cellule che presentano un potenziale differenziativo in senso neurale nel sistema nervoso centrale dell’adulto (Bifari et al., 2009, 2015; Decimo et al., 2011; Nakagomi et al., 2011, 2012; Petricevic et al., 2011). Infatti, i precursori identificati nelle meningi sono in grado di differenziare in neuroni con elevata efficienza, sia in vitro che dopo trapianto in vivo (Bifari et al., 2009; Decimo et al., 2011). Inoltre, è stata osservata un’attivazione di questa popolazione in seguito a lesioni ischemiche cerebrali, contribuendo ad una notevole espansione del pool di cellule staminali e progenitori presenti (Nakagomi et al., 2012). I precursori neurali delle meningi contribuiscono alla reazione parenchimale che si verifica dopo una lesione al midollo spinale, migrando verso l’area lesionata ed esprimendo i marcatori (come nestina e DCX) che sono espressi anche dai precursori neurali all’interno delle nicchie staminali classiche (Decimo et al., 2011). L’identificazione di questa nuova popolazione nelle meningi, che presenta caratteristiche tipiche delle cellule staminali neurali, suggerisce un potenziale ruolo per i precursori delle meningi nel mantenimento dell’omeostasi cerebrale. Tuttavia, il possibile contributo delle cellule delle meningi alla neurogenesi in condizioni fisiologiche non è ancora stato investigato. Durante il corso dei miei studi, ho studiato il contributo delle cellule delle meningi alla neurogenesi in vivo. Abbiamo sviluppato una tecnica che ci permettesse di marcare le cellule delle meningi, in modo da poterle visualizzare e seguire nel tempo, combinando iniezioni di coloranti vitali o marcatori genetici (lentivirus o plasmidi) con l’utilizzo di modelli transgenici. Abbiamo osservato che le cellule delle meningi migrano nel periodo postnatale, dalla loro sede esterna al parenchima cerebrale fino alle cortecce restrospleniale e visuo-motoria. In seguito, queste cellule differenziano in neuroni che sono funzionali dal punto di vista elettrofisiologico, integrati nel network neuronale esistente e responsivi a stimoli farmacologici. Inoltre, abbiamo dimostrato che queste cellule neurogeniche appartengono alla popolazione perivascolare delle meningi che esprime PDGFRß. Nonostante non sia stata ancora chiarita l’origine embrionale di queste cellule, i nostri dati preliminari suggeriscono una possibile provenienza dalla popolazione della cresta neurale. In conclusione, una riserva di progenitori residente nelle meningi, di origine embrionale, contribuisce alla neurogenesi corticale nel periodo postnatale. Queste cellule rappresentano una riserva di cellule staminali endogene e potrebbero quindi essere utilizzate in medicina rigenerativa nel trattamento dei disordini neurodegenerativi.
Neurogenesis continues throughout life in mammalian brain (Eriksson et al., 1998; Gage, 2000) in two germinal niches: the subventricular zone lining the lateral ventricle and subgranular zone in the dentate gyrus of the hippocampus (Gage and Temple, 2013). Radial glial cells (Kriegstein and Alvarez-Buylla, 2009) are the neural stem cells that, during embryonic and postnatal development, give rise to various cell types including neuroblasts, neurons, oligodendrocytes, astrocytes and ependymal cells (Kriegstein and Alvarez-Buylla, 2009). In adult mice, newly formed neuroblasts migrate through the rostral migratory stream to the olfactory bulb, where they continually replace local interneurons (Imayoshi et al., 2008). Apart from these well-established neural stem niches, the existence of ectopic neural stem cell niches has been reported following injury (Pluchino et al., 2010), as well as in selected physiological conditions in the retina, cerebellum and olfactory bulb (Menezes et al., 1995; Ponti et al., 2008; Tropepe et al., 2000). Interestingly, several independent groups have recently identified a novel role for meninges as a potential niche harbouring endogenous stem cells with neural differentiation potential in the adult central nervous system (Bifari et al., 2009, 2015; Decimo et al., 2011; Nakagomi et al., 2011, 2012; Petricevic et al., 2011). Surprisingly, meningeal neural precursors are able to differentiate both in vitro and, after transplantation in vivo, into neurons with extremely high efficiency (Bifari et al., 2009; Decimo et al., 2011). Moreover, these cells can be activated by central nervous system parenchymal injuries, undergoing an extensive expansion of stem cells and progenitors (Nakagomi et al., 2012). Meningeal neural precursors contribute to neural parenchymal reaction after spinal cord injury, migrating to the perilesioned area, while expressing the same markers (nestin and DCX) that are transiently expressed by neural precursors within classic neurogenic niches (Decimo et al., 2011). The finding of this new cell population in the meninges, with stem cell features, provides new insights into the complexity of the parenchymal reaction to a traumatic injury and suggests a potential role for meningeal progenitor cells in the maintainance of brain homeostasis. However, the possible contribution of meningeal neural precursors to neurogenesis in physiological conditions has not previously been investigated. During the course of my studies, I explored the hypothesis that meningeal cells may contribute to neurogenesis in vivo. We were able to specifically tag meningeal cells in P0 pups and track them during time, combining injection of cell tracers in the meningeal subarachnoid space and transgenic mouse lines. We found that neurogenic meningeal cells migrate from their location outside the brain parenchyma, along the meningeal substructures, to the retrosplenial and visual motor cortices during the neonatal period. Subsequently, meningeal-derived cells differentiate into cortical neurons that are electrophysiologically functional, integrated in the existing network and responsive to pharmacological stimuli. In addition we found that these meningeal neurogenic cells belongs to the perivascular PDGFRß+ lineage and are mainly additive to the well-characterized neurogenic parenchymal radial glia. Although the developmental origin of these cells still has to be elucidated, our preliminary data indicate a possible neural crest-derivation. Thus, a reservoir of embryonic derived progenitors residing in the meninges contributes to postnatal cortical neurogenesis. These cells may have a role as endogenous stem cell pool that can be exploited in regenerative medicine for neurodegenerative diseases.

博士
長庚大學
臨床醫學研究所
97
Stem cells are defined to have the capability of self-renewal, proliferation and differentiation. In mammals, subventricular zone (SVZ) is a neural stem niche where thousands of neuroblasts are born everyday and migrate to the olfactory bulb (OB). There are a few signals pathways reported to be involved in this neurogenic niche; epigenetic control is a one of the major mechanisms remained to be elucidate. Chromatin remodelling, a key process to activate or suppress gene function, is important for stem cell maintenance. However, for neural stem cells, the relationship with chromatin remodelling is still poorly understood. Trithorax (Trx) and polycomb (Pcb) groups are both important chromatin remodelling factors which modulating embryo development. From published expression profiles of SVZ and OB, several chromatin modifiers were identified. Mll, a trithorax member, is first identified as a leukemic oncogene. With the utilization of reversed-transcription polymerase reaction (RT-PCR), in situ hybridization and conditional knock-in mice, we are able to demonstrate and confirm the persistent expression of Mll in the SVZ, starting from embryo stage. With an unique Mll1 conditional knockout mice, when crossed to an hGFAP-Cre strain, we observed a significant defective neurogenesis. The DCX-expressing neuroblasts accumulated in the SVZ without migration to the OB. The defect is limited to neurogenesis but not to gliogenesis; oligodrencytes and astrocytes are normally differentiated from the SVZ. This phenotype is re-confirmed by injecting the Cre-carrying adenovirus to the SVZ of conditional knockout mice. To further demonstrate this ex vivo, with monolayer neural stem cell culture, we compared the neurogenesis from SVZ of conditional knock-out mice. In vitro, the neurogenesis was decreased by near 40 folds; while oligodendrocytes and astrocytes were compensatory increased. Since in a conditional knockout model, the Mll1 is deleted since embryo stage day 11.5-12.5, it is possible that Mll1 may not affect neurogenesis directly. We utilized shRNAi and Cre-carrying virus to infect the monolayer SVZ stem cells and knockdown Mll1 immediately. In both methods, the neurogenesis decreased in vitro. Neuroblasts transcription signals were not decreased symmetrically; Dlx2 was decreased while MASH1 was not. We designed a Dlx2-carrying plasmid to infect the Mll1-depleted cells and the neurogenesis was partially rescued. To further explore the downstream targets, chromatin immunoprecipitation (CHIP) with MLL antibody was performed. Mll1 is directly binding on the Dlx2 promoter regions, also abundant at 1Kb upstream at the initiation site. To investigate which histone methylation manipulates the activation of Dlx2, we performed another two CHIP experiments with H3K4me3 and H3K27me3 antibodies. We found that in Mll1-deleted SVZ, H3K4me3 is not different on Dlx2, MASH1 and Olig2 loci; whiles Dlx2 locus is strongly methylated on H3K27. In conclusions, Mll1 is expressed in the mammals SVZ. Lacking Mll1 may lead to defective neurogenesis and failure to migration; it does not affect gliogenesis. Mll1 in the CNS does not act as H3K4 methyl-transferase; more possibly, it may recruit a demethylase to specifically remove the methylation on H3K27 of Dlx2 locus.