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Neural stem cells (NSCs), crucial for memory in the adult brain, are also pivotal to buffer depressive behavior. However, the mechanisms underlying the boost in NSC activity throughout life are still largely undiscovered. Here, we aimed to explore the role of deacetylase Sirtuin 3 (SIRT3), a central player in mitochondrial metabolism and oxidative protection, in the fate of NSC under aging and depression-like contexts. We showed that chronic treatment with tert-butyl hydroperoxide induces NSC aging, markedly reducing SIRT3 protein. SIRT3 overexpression, in turn, restored mitochondrial oxidative stress and the differentiation potential of aged NSCs. Notably, SIRT3 was also shown to physically interact with the long chain acyl-CoA dehydrogenase (LCAD) in NSCs and to require its activation to prevent age-impaired neurogenesis. Finally, the SIRT3 regulatory network was investigated in vivo using the unpredictable chronic mild stress (uCMS) paradigm to mimic depressive-like behavior in mice. Interestingly, uCMS mice presented lower levels of neurogenesis and LCAD expression in the same neurogenic niches, being significantly rescued by physical exercise, a well-known upregulator of SIRT3 and lipid metabolism. Our results suggest that targeting NSC metabolism, namely through SIRT3, might be a suitable promising strategy to delay NSC aging and confer stress resilience.

Neurogenesis decreases in Alzheimer’s disease (AD) patients, suggesting that restoring the normal neurogenic response could be a disease modifying intervention. To study the mechanisms of pathology-induced neuro-regeneration in vertebrate brains, zebrafish is an excellent model due to its extensive neural regeneration capacity. Here, we report that Kynurenic acid (KYNA), a metabolite of the amino acid tryptophan, negatively regulates neural stem cell (NSC) plasticity in adult zebrafish brain through its receptor, aryl hydrocarbon receptor 2 (Ahr2). The production of KYNA is suppressed after amyloid-toxicity through reduction of the levels of Kynurenine amino transferase 2 (KAT2), the key enzyme producing KYNA. NSC proliferation is enhanced by an antagonist for Ahr2 and is reduced with Ahr2 agonists or KYNA. A subset of Ahr2-expressing zebrafish NSCs do not express other regulatory receptors such as il4r or ngfra, indicating that ahr2-positive NSCs constitute a new subset of neural progenitors that are responsive to amyloid-toxicity. By performing transcriptome-wide association studies (TWAS) in three late onset Alzheimer disease (LOAD) brain autopsy cohorts, we also found that several genes that are components of KYNA metabolism or AHR signaling are differentially expressed in LOAD, suggesting a strong link between KYNA/Ahr2 signaling axis to neurogenesis in LOAD.

Parkinson’s disease (PD) as a progressive neurodegenerative disorder arises from multiple genetic and environmental factors. However, underlying pathological mechanisms remain poorly understood. Using multiplexed single-cell transcriptomics, we analyze human neural precursor cells (hNPCs) from sporadic PD (sPD) patients. Alterations in gene expression appear in pathways related to primary cilia (PC). Accordingly, in these hiPSC-derived hNPCs and neurons, we observe a shortening of PC. Additionally, we detect a shortening of PC in PINK1-deficient human cellular and mouse models of familial PD. Furthermore, in sPD models, the shortening of PC is accompanied by increased Sonic Hedgehog (SHH) signal transduction. Inhibition of this pathway rescues the alterations in PC morphology and mitochondrial dysfunction. Thus, increased SHH activity due to ciliary dysfunction may be required for the development of pathoetiological phenotypes observed in sPD like mitochondrial dysfunction. Inhibiting overactive SHH signaling may be a potential neuroprotective therapy for sPD.

Summary Physical activity is a strong positive physiological modulator of adult neurogenesis in the hippocampal dentate gyrus. Although the underlying regulatory mechanisms are still unknown, systemic processes must be involved. Here we show that platelets are activated after acute periods of running, and that activated platelets promote neurogenesis, an effect that is likely mediated by platelet factor 4. Ex vivo, the beneficial effects of activated platelets and platelet factor 4 on neural precursor cells were dentate gyrus specific and not observed in the subventricular zone. Moreover, the depletion of circulating platelets in mice abolished the running-induced increase in precursor cell proliferation in the dentate gyrus following exercise. These findings demonstrate that platelets and their released factors can modulate adult neural precursor cells under physiological conditions and provide an intriguing link between running-induced platelet activation and the modulation of neurogenesis after exercise., Graphical Abstract, Highlights • Activated platelets increase hippocampal neurogenesis in physiological conditions • Platelet-depleted mice lack the running-induced increase in precursor proliferation • PF4-treated mice have more immature neurons • Pro-neurogenic effects of activated platelets and PF4 are dentate gyrus-specific, Using the neurogenesis-promoting stimulus of physical activity, Walker and colleagues show that platelets are activated after acute running periods and that activated platelets and their released protein PF4 following exercise, increase neurogenesis. They show that the pro-neurogenic effects of platelets are dentate gyrus specific and that platelet depletion in mice abolishes the running-induced increase in precursor proliferation in the dentate gyrus.

The development of the cerebral cortex relies on the controlled division of neural stem and progenitor cells. The requirement for precise spatiotemporal control of proliferation and cell fate places a high demand on the cell division machinery, and defective cell division can cause microcephaly and other brain malformations. Cell-extrinsic and -intrinsic factors govern the capacity of cortical progenitors to produce large numbers of neurons and glia within a short developmental time window. In particular, ion channels shape the intrinsic biophysical properties of precursor cells and neurons and control their membrane potential throughout the cell cycle. We found that hyperpolarization-activated cyclic nucleotide-gated cation (HCN) channel subunits are expressed in mouse, rat, and human neural progenitors. Loss of HCN channel function in rat neural stem cells impaired their proliferation by affecting the cell-cycle progression, causing G1 accumulation and dysregulation of genes associated with human microcephaly. Transgene-mediated, dominant-negative loss of HCN channel function in the embryonic mouse telencephalon resulted in pronounced microcephaly. Together, our findings suggest a role for HCN channel subunits as a part of a general mechanism influencing cortical development in mammals.

Neurotherapeutics : official journal of the American Society for Experimental NeuroTherapeutics (ASENT) 18(3), 1963-1979 (2021). doi:10.1007/s13311-021-01045-2, Published by Springer, New York, NY

International audience; For some time, it has been accepted that the β-site APP cleaving enzyme 1 (BACE1) and the γ-secretase are two main players in the amyloidogenic processing of the β-amyloid precursor protein (APP). Recently, the membrane-type 5 matrix metalloproteinase (MT5-MMP/MMP-24), mainly expressed in the nervous system, has been highlighted as a new key player in APP-processing, able to stimulate amyloidogenesis and also to generate a neurotoxic APP derivative. In addition, the loss of MT5-MMP has been demonstrated to abrogate pathological hallmarks in a mouse model of Alzheimer’s disease (AD), thus shedding light on MT5-MMP as an attractive new therapeutic target. However, a more comprehensive analysis of the role of MT5-MMP is necessary to evaluate how its targeting affects neurons and glia in pathological and physiological situations. In this study, leveraging on CRISPR-Cas9 genome editing strategy, we established cultures of human-induced pluripotent stem cells (hiPSC)-derived neurons and astrocytes to investigate the impact of MT5-MMP deficiency on their phenotypes. We found that MT5-MMP-deficient neurons exhibited an increased number of primary and secondary neurites, as compared to isogenic hiPSC-derived neurons. Moreover, MT5-MMP-deficient astrocytes displayed higher surface area and volume compared to control astrocytes. The MT5-MMP-deficient astrocytes also exhibited decreased GLAST and S100β expression. These findings provide novel insights into the physiological role of MT5-MMP in human neurons and astrocytes, suggesting that therapeutic strategies targeting MT5-MMP should be controlled for potential side effects on astrocytic physiology and neuronal morphology

Physical exercise stimulates adult neurogenesis, yet the underlying mechanisms remain poorly understood. A fundamental component of the innate neuroregenerative capacity of zebrafish is the proliferative and neurogenic ability of the neural stem/progenitor cells. Here, we show that in the intact spinal cord, this plasticity response can be activated by physical exercise by demonstrating that the cholinergic neurotransmission from spinal locomotor neurons activates spinal neural stem/progenitor cells, leading to neurogenesis in the adult zebrafish. We also show that GABA acts in a non-synaptic fashion to maintain neural stem/progenitor cell quiescence in the spinal cord and that training-induced activation of neurogenesis requires a reduction of GABAA receptors. Furthermore, both pharmacological stimulation of cholinergic receptors, as well as interference with GABAergic signaling, promote functional recovery after spinal cord injury. Our findings provide a model for locomotor networks’ activity-dependent neurogenesis during homeostasis and regeneration in the adult zebrafish spinal cord., The mechanisms stimulating adult neurogenesis are unclear. Here, the authors show the contribution of cholinergic and GABAergic signalling within the locomotor network to spinal cord neurogenesis during homeostasis and regeneration, showing neurogenesis depends on circuit activity in the adult zebrafish.

Although the neurogenesis-enhancing effects of exercise have been extensively studied, the molecular mechanisms underlying this response remain unclear. Here, we propose that this is mediated by the exercise-induced systemic release of the antioxidant selenium transport protein, selenoprotein P (SEPP1). Using knockout mouse models, we confirmed that SEPP1 and its receptor low-density lipoprotein receptor-related protein 8 (LRP8) are required for the exercise-induced increase in adult hippocampal neurogenesis. In vivo selenium infusion increased hippocampal neural precursor cell (NPC) proliferation and adult neurogenesis. Mimicking the effect of exercise through dietary selenium supplementation restored neurogenesis and reversed the cognitive decline associated with aging and hippocampal injury, suggesting potential therapeutic relevance. These results provide a molecular mechanism linking exercise-induced changes in the systemic environment to the activation of quiescent hippocampal NPCs and their subsequent recruitment into the neurogenic trajectory.

Summary Intermediate progenitor cells (IPCs) are neocortical neuronal precursors. Although IPCs play crucial roles in corticogenesis, their molecular features remain largely unknown. In this study, we aimed to characterize the molecular profile of IPCs. We isolated TBR2-positive (+) IPCs and TBR2-negative (−) cell populations in the developing mouse cortex. Comparative genome-wide gene expression analysis of TBR2+ IPCs versus TBR2− cells revealed differences in key factors involved in chromatid segregation, cell-cycle regulation, transcriptional regulation, and cell signaling. Notably, mutation of many IPC genes in human has led to intellectual disability and caused a wide range of cortical malformations, including microcephaly and agenesis of corpus callosum. Loss-of-function experiments in cortex-specific mutants of Esco2, one of the novel IPC genes, demonstrate its critical role in IPC maintenance, and substantiate the identification of a central genetic determinant of IPC biogenesis. Our data provide novel molecular characteristics of IPCs in the developing mouse cortex., Graphical abstract, Highlights • Purification and transcriptome profiling of IPCs in developing mouse cortex • Identified 1,119 IPC-enriched genes with over 350 novel IPC genes in SVZ confirmed • IPC gene mutation is linked to intellectual disability and cortical malformation • Esco2, a novel IPC-enriched gene, is needed for IPC viability during corticogenesis, In this article, Tuoc and colleagues show that the expression of many genes, including ESCO2, which encode for key factors involved in chromatid segregation, cell-cycle regulation, transcriptional regulation, and cell signaling, is enriched in intermediate progenitor cells, and their mutation has implications for a wide range of cortical malformations and functional disabilities in the mouse and human brain.

CREB (cyclic AMP response element binding protein) binding protein (CBP, CREBBP) is a ubiquitously expressed transcription coactivator with intrinsic histone acetyltransferase (KAT) activity. Germline mutations within theCBPgene are known to cause Rubinstein-Taybi syndrome (RSTS), a developmental disorder characterized by intellectual disability, specific facial features and physical anomalies. Here, we investigate mechanisms of CBP function during brain development in order to elucidate morphological and functional mechanisms underlying the development of RSTS. Due to the embryonic lethality of conventional CBP knockout mice, we employed a tissue specific knockout mouse model (hGFAP-cre::CBPFl/Fl, mutant mouse) to achieve a homozygous deletion of CBP in neural precursor cells of the central nervous system.Our findings suggest that CBP plays a central role in brain size regulation, correct neural cell differentiation and neural precursor cell migration. We provide evidence that CBP is both important for stem cell viability within the ventricular germinal zone during embryonic development and for unhindered establishment of adult neurogenesis. Prominent histological findings in adult animals include a significantly smaller hippocampus with fewer neural stem cells. In the subventricular zone, we observe large cell aggregations at the beginning of the rostral migratory stream due to a migration deficit caused by impaired attraction from the CBP-deficient olfactory bulb. The cerebral cortex of mutant mice is characterized by a shorter dendrite length, a diminished spine number, and a relatively decreased number of mature spines as well as a reduced number of synapses.In conclusion, we provide evidence that CBP is important for neurogenesis, shaping neuronal morphology, neural connectivity and that it is involved in neuronal cell migration. These findings may help to understand the molecular basis of intellectual disability in RSTS patients and may be employed to establish treatment options to improve patients’ quality of life.

Summary During early cortical development, neural stem cells (NSCs) divide symmetrically to expand the progenitor pool, whereas, in later stages, NSCs divide asymmetrically to self-renew and produce other cell types. The timely switch from such proliferative to differentiative division critically determines progenitor and neuron numbers. However, the mechanisms that limit proliferative division in late cortical development are not fully understood. Here, we show that the BAF (mSWI/SNF) complexes restrict proliferative competence and promote neuronal differentiation in late corticogenesis. Inactivation of BAF complexes leads to H3K27me3-linked silencing of neuronal differentiation-related genes, with concurrent H3K4me2-mediated activation of proliferation-associated genes via de-repression of Wnt signaling. Notably, the deletion of BAF complexes increased proliferation of neuroepithelial cell-like NSCs, impaired neuronal differentiation, and exerted a Wnt-dependent effect on neocortical and hippocampal development. Thus, these results demonstrate that BAF complexes act as both activators and repressors to control global epigenetic and gene expression programs in late corticogenesis., Graphical Abstract, Highlights • Loss of BAF complexes increases H3K27me3 and H3K4me2 marks in late corticogenesis • BAF complexes epigenetically regulate neural proliferation and differentiation • BAF complexes suppress neuroepithelial cell fate and Wnt signaling • BAF complexes control cortical development in a Wnt signaling-dependent manner, In this article, Tuoc and colleagues show that inactivation of BAF complexes in late cortical development leads to H3K27me3-linked silencing of neuronal differentiation-related genes, with concurrent H3K4me2-mediated activation of proliferation-associated genes via de-repression of Wnt signaling. The deletion of BAF complexes increased proliferation of neuroepithelial-like progenitors, impaired neuronal differentiation, and exerted a Wnt-dependent effect on neocortical and hippocampal development.

Objective In familial hypercholesterolemia (FH), mutations in the low-density lipoprotein (LDL) receptor (LDLr) gene result in increased plasma LDL cholesterol. Clinical and preclinical studies have revealed an association between FH and hippocampus-related memory and mood impairment. We here asked whether hippocampal pathology in FH might be a consequence of compromised adult hippocampal neurogenesis. Methods We evaluated hippocampus-dependent behavior and neurogenesis in adult C57BL/6JRj and LDLr−/− mice. We investigated the effects of elevated cholesterol and the function of LDLr in neural precursor cells (NPC) isolated from adult C57BL/6JRj mice in vitro. Results Behavioral tests revealed that adult LDLr−/− mice showed reduced performance in a dentate gyrus (DG)-dependent metric change task. This phenotype was accompanied by a reduction in cell proliferation and adult neurogenesis in the DG of LDLr−/− mice, suggesting a potential direct impact of LDLr mutation on NPC. Exposure of NPC to LDL as well as LDLr gene knockdown reduced proliferation and disrupted transcriptional activity of genes involved in endogenous cholesterol synthesis and metabolism. The LDL treatment also induced an increase in intracellular lipid storage. Functional analysis of differentially expressed genes revealed parallel modulation of distinct regulatory networks upon LDL treatment and LDLr knockdown. Conclusions Together, these results suggest that high LDL levels and a loss of LDLr function, which are characteristic to individuals with FH, might contribute to a disease-related impairment in adult hippocampal neurogenesis and, consequently, cognitive functions., Graphical abstract LDL receptor and cholesterol metabolism regulate adult hippocampal neurogenesis. The LDLr −/− mice show impaired hippocampal related behavior and adult neurogenesis. In vitro exposure of neural precursor cells (NPC) to LDL and LDLr knock-down reduces cell proliferation modulating distinct regulatory networks. Additionally, LDL exposure induces increase in lipid storage and impaired neuronal differentiation.Image 1, Highlights • The LDLr −/− mice show impaired hippocampal related behaviour and adult neurogenesis. • In vitro exposure of NPC to LDL and LDLr knock-down reduces cell proliferation. • LDL exposure induces lipid storage in NPC. • In vitro LDL and LDLr knock-down in NPC modulates distinct regulatory networks.

Axonal degeneration is a key pathology of neurodegenerative diseases, including hereditary spastic paraplegia (HSP), a disorder characterized by spasticity in the lower limbs. Treatments for HSP and other neurodegenerative diseases are mainly symptomatic. While iPSC-derived neurons are valuable for drug discovery and target identification, these applications require robust differentiation paradigms and rapid phenotypic read-outs ranging between hours and a few days. Using spastic paraplegia type 4 (SPG4, the most frequent HSP subtype) as an exemplar, we here present three rapid phenotypic assays for uncovering neuronal process pathologies in iPSC-derived glutamatergic cortical neurons. Specifically, these assays detected a 51% reduction in neurite outgrowth and a 60% increase in growth cone area already 24 hours after plating; axonal swellings, a hallmark of HSP pathology, was discernible after only 5 days. Remarkably, the identified phenotypes were neuron subtype-specific and not detectable in SPG4-derived GABAergic forebrain neurons. We transferred all three phenotypic assays to a 96-well setup, applied small molecules and found that a liver X receptor (LXR) agonist rescued all three phenotypes in HSP neurons, providing a potential drug target for HSP treatment. We expect this multiparametric and rapid phenotyping approach to accelerate development of therapeutic compounds for HSP and other neurodegenerative diseases.

FUS (fused in sarcoma) plays a key role in several steps of RNA metabolism, and dominant mutations in this protein are associated with neurodegenerative diseases. Here, we show that FUS is a component of the cellular response to topoisomerase I (TOP1)–induced DNA breakage; relocalising to the nucleolus in response to RNA polymerase II (Pol II) stalling at sites of TOP1-induced DNA breaks. This relocalisation is rapid and dynamic, reversing following the removal of TOP1-induced breaks and coinciding with the recovery of global transcription. Importantly, FUS relocalisation following TOP1-induced DNA breakage is associated with increased FUS binding at sites of RNA polymerase I transcription in ribosomal DNA and reduced FUS binding at sites of RNA Pol II transcription, suggesting that FUS relocates from sites of stalled RNA Pol II either to regulate pre-mRNA processing during transcriptional stress or to modulate ribosomal RNA biogenesis. Importantly, FUS-mutant patient fibroblasts are hypersensitive to TOP1-induced DNA breakage, highlighting the possible relevance of these findings to neurodegeneration., This work was funded by an MRC project grant to KW Caldecott and M Hafezparast (MR/K01854X/1), and MRC and ERC Programme grants to KW Caldecott (ERC SIDSCA 694996, MR/P010121/1). MI Martinez-Macias was also supported by an European Molecular Biology Organisation (EMBO) Long-Term Fellowship ALTF 751-2013.

Neuroblasts born in the subventricular zone of adult mammals migrate via the rostral migratory stream into the granular cell layer or periglomerular layer of the olfactory bulb to differentiate into interneurons. To analyze if new neurons in the granular cell layer or periglomerular layer have different origins, we inserted a physical barrier into the rostral migratory stream, depleted cell proliferation with cytarabine infusions, labeled newborn cells with bromodeoxyuridine, and sacrificed mice after short-term (0, 2, or 14 days) or long-term (55 or 105 days) intervals. After short-term survival, the subventricular zone and rostral migratory stream rapidly repopulated with bromodeoxyuridine+ cells after cytarabine-induced depletion. Nestin, glial fibrillary acidic protein and the PAX6 were expressed in bromodeoxyuridine+ cells within the rostral migratory stream downstream of the physical barrier. After long-term survival after physical barrier implantation, bromodeoxyuridine+ neurons were significantly reduced in the granular cell layer, but bromodeoxyuridine+ and dopaminergic neurons in the periglomerular layer remained unaffected by the physical barrier. Thus, newborn neurons for the granular cell layer are mainly recruited from neural stem cells located in the subventricular zone, but new neurons for the periglomerular layer with dopaminergic predisposition can rise as well from neuronal stem or precursor cells in the rostral migratory stream.

In demyelinating diseases including multiple sclerosis (MS), neural stem cells (NSCs) can replace damaged oligodendrocytes if the local microenvironment supports the required differentiation process. Although chitinase-like proteins (CLPs) form part of this microenvironment, their function in this differentiation process is unknown. Here, we demonstrate that murine Chitinase 3-like-3 (Chi3l3/Ym1), human Chi3L1 and Chit1 induce oligodendrogenesis. In mice, Chi3l3 is highly expressed in the subventricular zone, a stem cell niche of the adult brain, and in inflammatory brain lesions during experimental autoimmune encephalomyelitis (EAE). We find that silencing Chi3l3 increases severity of EAE. We present evidence that in NSCs Chi3l3 activates the epidermal growth factor receptor (EGFR), thereby inducing Pyk2-and Erk1/2- dependent expression of a pro-oligodendrogenic transcription factor signature. Our results implicate CLP-EGFR-Pyk2-MEK-ERK as a key intrinsic pathway controlling oligodendrogenesis., Chitinase 3-like-3 (Chi3l3) is expressed in microglia, but its function is not clear. Here the authors show that Chi3l3 is expressed in the subventricular zone in mouse experimental immune encephalitis, which induces oligodendrogenesis.

The neural stem cell (NSC) reservoir can be harnessed for stem cell-based regenerative therapies. Zebrafish remarkably regenerate their brain by inducing NSC plasticity in a Amyloid-β-42 (Aβ42)-induced experimental Alzheimer's disease (AD) model. Interleukin-4 (IL-4) is also critical for AD-induced NSC proliferation. However, the mechanisms of this response have remained unknown. Using single-cell transcriptomics in the adult zebrafish brain, we identify distinct subtypes of NSCs and neurons and differentially regulated pathways and their gene ontologies and investigate how cell-cell communication is altered through ligand-receptor pairs in AD conditions. Our results propose the existence of heterogeneous and spatially organized stem cell populations that react distinctly to amyloid toxicity. This resource article provides an extensive database for the molecular basis of NSC plasticity in the AD model of the adult zebrafish brain. Further analyses of stem cell heterogeneity and neuro-regenerative ability at single-cell resolution could yield drug targets for mobilizing NSCs for endogenous neuro-regeneration in humans.

Cross-talk between the peripheral immune system and the central nervous system is important for physiological brain health. T cells are required to maintain normal baseline levels of neural precursor proliferation in the hippocampus of adult mice. We show here that neither T cells, B cells, natural killer cells nor natural killer T cells are required for the increase in hippocampal precursor proliferation that occurs in response to physical exercise. In addition, we demonstrate that a subpopulation of T cells, regulatory T cells, is not involved in maintaining baseline levels of neural precursor proliferation. Even when applied at supraphysiological numbers, populations of both naive and stimulated lymphocytes had no effect on hippocampal precursor proliferation in vitro. In addition, physical activity had no effect on peripheral immune cells in terms of distribution in the bone marrow, lymph nodes or spleen, activation state or chemokine receptor (CXCR4 and CCR9) expression. Together these results suggest that lymphocytes are not involved in translating the peripheral effects of exercise to the neurogenic niche in the hippocampus and further support the idea that the exercise-induced regulation of adult neurogenesis is mechanistically distinct from its baseline control., Frontiers in Immunology, 9, ISSN:1664-3224

Histone H3.3 glycine 34 to arginine/valine (G34R/V) mutations drive deadly gliomas and show exquisite regional and temporal specificity, suggesting a developmental context permissive to their effects. Here, we show that 50% of G34R/V-tumours (n=95) bear activating PDGFRA mutations that display strong selection pressure at recurrence. While considered gliomas, G34R/V-tumours actually arise in GSX2/DLX-expressing interneuron progenitors, where G34R/V-mutations impair neuronal differentiation. The lineage-of-origin may facilitate PDGFRA co-option through a chromatin loop connecting PDGFRA to GSX2 regulatory elements, promoting PDGFRA overexpression and mutation. At the single-cell level, G34R/V-tumours harbour dual neuronal/astroglial identity and lack oligodendroglial programs, actively repressed by GSX2/DLX-mediated cell-fate specification. G34R/V may become dispensable for tumour maintenance, while mutant-PDGFRA is potently oncogenic. Collectively, our results open novel research avenues in deadly tumours. G34R/V-gliomas are neuronal malignancies, where interneuron progenitors are stalled in differentiation by G34R/V-mutations, and malignant gliogenesis is promoted by co-option of a potentially targetable pathway, PDGFRA signalling.

Summary Tight regulation of the balance between self-renewal and differentiation of neural stem cells is crucial to assure proper neural development. In this context, Notch signaling is a well-known promoter of stemness. In contrast, the bifunctional brain-enriched microRNA miR-9/9∗ has been implicated in promoting neuronal differentiation. Therefore, we set out to explore the role of both regulators in human neural stem cells. We found that miR-9/9∗ decreases Notch activity by targeting NOTCH2 and HES1, resulting in an enhanced differentiation. Vice versa, expression levels of miR-9/9∗ depend on the activation status of Notch signaling. While Notch inhibits differentiation of neural stem cells, it also induces miR-9/9∗ via recruitment of the Notch intracellular domain (NICD)/RBPj transcriptional complex to the miR-9/9∗_2 genomic locus. Thus, our data reveal a mutual interaction between bifunctional miR-9/9∗ and the Notch signaling cascade, calibrating the delicate balance between self-renewal and differentiation of human neural stem cells., Graphical Abstract, Highlights • MiR-9/9∗ regulate Notch signaling by targeting NOTCH2 and HES1 • Notch directly regulates transcription of the miR-9_2 genomic locus • Notch-miR-9 reciprocal regulation calibrates NSC self-renewal and differentiation, In this article, Brüstle and colleagues show that a mutual interaction between microRNA-9 and Notch calibrates the delicate balance between self-renewal and differentiation of human neural stem cells (hNSC). While Notch maintains stemness and proliferation of hNSC, it also directly induces expression of miR-9, which in turn promotes hNSC differentiation by targeting NOTCH2 and HES1.

Summary: Physical exercise stimulates adult hippocampal neurogenesis and is considered a relevant strategy for preventing age-related cognitive decline in humans. The underlying mechanisms remains controversial. Here, we show that exercise increases proliferation of neural precursor cells (NPCs) of the mouse dentate gyrus (DG) via downregulation of microRNA 135a-5p (miR-135a). MiR-135a inhibition stimulates NPC proliferation leading to increased neurogenesis, but not astrogliogenesis, in DG of resting mice, and intriguingly it re-activates NPC proliferation in aged mice. We identify 17 proteins (11 putative targets) modulated by miR-135 in NPCs. Of note, inositol 1,4,5-trisphosphate (IP3) receptor 1 and inositol polyphosphate-4-phosphatase type I are among the modulated proteins, suggesting that IP3 signaling may act downstream miR-135. miR-135 is the first noncoding RNA essential modulator of the brain's response to physical exercise. Prospectively, the miR-135-IP3 axis might represent a novel target of therapeutic intervention to prevent pathological brain aging. : Pons-Espinal, Gasperini, and colleagues report that running induces NPC proliferation and neurogenesis via downregulation of miR-135a-5p in the mouse hippocampus. Remarkably, downregulation of miR-135a stimulates proliferation in the hippocampus of aged mice. ITPR1 and INPP4A, involved in IP3 signaling, are modulated by miR-135 in NPCs. Prospectively, therapeutic exploitation of the miR-135-IP3 axis might represent a novel intervention strategy for successful aging. Keywords: adult neurogenesis, running, aging, miR-135a, inositol 1,4,5-trisphosphate (IP3) pathway, ITPR1, INPP4A

Distinct micro-environmental properties have been reported to be essential for maintenance of neural precursor cells (NPCs) within the adult brain. Due to high complexity and technical limitations, the natural niche can barely be studied systematically in vivo. By reconstituting selected environmental properties (adhesiveness, proteolytic degradability, and elasticity) in geldrop cultures, we show that NPCs can be maintained stably at high density over an extended period of time (up to 8 days). In both conventional systems, neurospheres and monolayer cultures, they would expand and (in the case of neurospheres) differentiate rapidly. Further, we report a critical dualism between matrix adhesiveness and degradability. Only if both features are functional NPCs stay proliferative. Lastly, Rho-associated protein kinase was identified as part of a pivotal intracellular signaling cascade controlling cell morphology in response to environmental cues inside geldrop cultures. Our findings demonstrate that simple manipulations of the microenvironment in vitro result in an important preservation of stemness features in the cultured precursor cells.

Voltage-gated sodium and calcium channels as well as transient receptor potential (TRP) channels are expressed during the differentiation of human neural progenitor cells (hNPCs) and are likely to be involved in regulating neurogenesis. However, the molecular composition of these ion channels in proliferating and differentiating hNPCs is largely unknown. In this study, we investigated fetal mesencephalic hNPCs in respect to their sodium, calcium, and TRP channel subunit expression and function. Quantitative real-time polymerase chain reaction indicated a significant upregulation of voltage-gated sodium and calcium channel subunits in hNPCs after differentiation for 3 weeks in vitro. In contrast, the TRP channel expression did not increase significantly during hNPC maturation. Intracellular Ca2+ measurements showed the marked reduction of KCl-induced Ca2+ transients through inhibition of voltage-gated Ca2+ channels by verapamil and mibefradil in differentiated hNPCs. Application of TRP channel agonists induced intracellular Ca2+ peaks already in proliferating hNPCs without affecting their cell division. The coincubation of hNPCs with TRP channel agonists pregnenolone sulfate or RN1747 did not have any significant effect on their proliferation and differentiation. These data indicate that hNPCs derived from fetal midbrain tissue acquire essential voltage-gated sodium and calcium channel properties during neuronal maturation in vitro. An early role of TRP channels in neurogenesis which may be important for regenerative clinical applications or cellular models could not be elucidated using hNPCs.

The pathological cascade leading from primary storage to neural cell dysfunction and death in metachromatic leukodystrophy (MLD) has been poorly elucidated in human-derived neural cell systems. In the present study, we have modeled the progression of pathological events during the differentiation of patient-specific iPSCs to neuroepithelial progenitor cells (iPSC-NPCs) and mature neurons, astrocytes, and oligodendrocytes at the morphological, molecular, and biochemical level. We showed significant sulfatide accumulation and altered sulfatide composition during the differentiation of MLD iPSC-NPCs into neuronal and glial cells. Changes in sulfatide levels and composition were accompanied by the expansion of the lysosomal compartment, oxidative stress, and apoptosis. The neuronal and glial differentiation capacity of MLD iPSC-NPCs was significantly impaired. We showed delayed appearance and/or reduced levels of oligodendroglial and astroglial markers as well as reduced number of neurons and disorganized neuronal network. Restoration of a functional Arylsulfatase A (ARSA) enzyme in MLD cells using lentiviral-mediated gene transfer normalized sulfatide levels and composition, globally rescuing the pathological phenotype. Our study points to MLD iPSC-derived neural progeny as a useful in vitro model to assess the impact of ARSA deficiency along NPC differentiation into neurons and glial cells. In addition, iPSC-derived neural cultures allowed testing the impact of ARSA reconstitution/overexpression on disease correction and, importantly, on the biology and functional features of human NPCs, with important therapeutic implications.

Transcription factor Runx1 (Runt Related Transcription Factor 1), plays an important role in the differentiation of hematopoetic stem cells, angiogenesis and the development of nociceptive neurons. These known functions have in common that they relate to lineage decisions. We thus asked whether such role might also be found for Runx1 in adult hippocampal neurogenesis as a process, in which such decisions have to be regulated lifelong. Runx1 shows a widespread low expression in the adult mouse brain, not particularly prominent in the hippocampus and the resident neural precursor cells. Isoforms 1 and 2 of Runx1 (but not 3 to 5) driven by the proximal promoter were expressed in hippocampal precursor cells ex vivo, albeit again at very low levels, and were markedly increased after stimulation with TGF-β1. Under differentiation conditions (withdrawal of growth factors) Runx1 became down-regulated. Overexpression of Runx1 in vitro reduced proliferation, increased survival of precursor cells by reducing apoptosis, and increased neuronal differentiation, while slightly reducing dendritic morphology and complexity. Transfection with dominant-negative Runx1 in hippocampal precursor cells in vitro did not result in differences in neurogenesis. Hippocampal expression of Runx1 correlated with adult neurogenesis (precursor cell proliferation) across BXD recombinant strains of mice and covarying transcripts enriched in the GO categories "neural precursor cell proliferation" and "neuron differentiation". Runx1 is thus a plausible candidate gene to be involved in regulating initial differentiation-related steps of adult neurogenesis. It seems, however, that the relative contribution of Runx1 to such effect is complementary and will explain only small parts of the cell-autonomous pro-differentiation effect.

Background The microtubule-associated protein Tau plays a role in neurodegeneration as well as neurogenesis. Previous work has shown that the expression of the pro-aggregant mutant Tau repeat domain causes strong aggregation and pronounced neuronal loss in the hippocampus whereas the anti-aggregant form has no deleterious effects. These two proteins differ mainly in their propensity to form ß structure and hence to aggregate. Methods To elucidate the basis of these contrasting effects, we analyzed organotypic hippocampal slice cultures (OHSCs) from transgenic mice expressing the repeat domain (RD) of Tau with the anti-aggregant mutation (TauRDΔKPP) and compared them with slices containing pro-aggregant TauRDΔK. Transgene expression in the hippocampus was monitored via a sensitive bioluminescence reporter gene assay (luciferase). Results The expression of the anti-aggregant TauRDΔKPP leads to a larger volume of the hippocampus at a young age due to enhanced neurogenesis, resulting in an increase in neuronal number. There were no signs of activation of microglia and astrocytes, indicating the absence of an inflammatory reaction. Investigation of signaling pathways showed that Wnt-5a was strongly decreased whereas Wnt3 was increased. A pronounced increase in hippocampal stem cell proliferation (seen by BrdU) was observed as early as P8, in the CA regions where neurogenesis is normally not observed. The increase in neurons persisted up to 16 months of age. Conclusion The data suggest that the expression of anti-aggregant TauRDΔKPP enhances hippocampal neurogenesis mediated by the canonical Wnt signaling pathway, without an inflammatory reaction. This study points to a role of tau in brain development and neurogenesis, in contrast to its detrimental role in neurodegeneration at later age. Electronic supplementary material The online version of this article (10.1186/s13024-017-0230-8) contains supplementary material, which is available to authorized users.

Parkinson disease (PD) is a neurodegenerative disorder characterized by massive loss of midbrain dopaminergic neurons. Whereas onset of motor impairments reflects a rather advanced stage of the disorder, hyposmia often marks the beginning of the disease. Little is known about the role of the nigro-striatal system in olfaction under physiological conditions and the anatomical basis of hyposmia in PD. Yet, the early occurrence of olfactory dysfunction implies that pathogens such as environmental toxins could incite the disease via the olfactory system. In the present study, we demonstrate a dopaminergic innervation from neurons in the substantia nigra to the olfactory bulb by axonal tracing studies. Injection of two dopaminergic neurotoxins-1-methyl-4-phenylpyridinium and 6-hydroxydopamine-into the olfactory bulb induced a decrease in the number of dopaminergic neurons in the substantia nigra. In turn, ablation of the nigral projection led to impaired olfactory perception. Hyposmia following dopaminergic deafferentation was reversed by treatment with the D1/D2/D3 dopamine receptor agonist rotigotine. Hence, we demonstrate for the first time the existence of a direct dopaminergic projection into the olfactory bulb and identify its origin in the substantia nigra in rats. These observations may provide a neuroanatomical basis for invasion of environmental toxins into the basal ganglia and for hyposmia as frequent symptom in PD.

Inflammation entails a complex set of defense mechanisms acting in concert to restore the homeostatic balance in organisms after damage or pathogen invasion. This immune response consists of the activity of various immune cells in a highly complex manner. Inflammation is a double-edged sword as it is reported to have both detrimental and beneficial consequences. In this review, we discuss the effects of inflammation on stem cell activity, focusing primarily on neural stem/progenitor cells in mammals and zebrafish. We also give a brief overview of the effects of inflammation on other stem cell compartments, exemplifying the positive and negative role of inflammation on stemness. The majority of the chronic diseases involve an unremitting phase of inflammation due to improper resolution of the initial pro-inflammatory response that impinges on the stem cell behavior. Thus, understanding the mechanisms of crosstalk between the inflammatory milieu and tissue-resident stem cells is an important basis for clinical efforts. Not only is it important to understand the effect of inflammation on stem cell activity for further defining the etiology of the diseases, but also better mechanistic understanding is essential to design regenerative therapies that aim at micromanipulating the inflammatory milieu to offset the negative effects and maximize the beneficial outcomes.

Summary Evolutionary elaboration of tissues starts with changes in the genome and location of the stem cells. For example, GABAergic interneurons of the mammalian neocortex are generated in the ventral telencephalon and migrate tangentially to the neocortex, in contrast to the projection neurons originating in the ventricular/subventricular zone (VZ/SVZ) of the dorsal telencephalon. In human and nonhuman primates, evidence suggests that an additional subset of neocortical GABAergic interneurons is generated in the cortical VZ and a proliferative niche, the outer SVZ. The origin, magnitude, and significance of this species-specific difference are not known. We use a battery of assays applicable to the human, monkey, and mouse organotypic cultures and supravital tissue to identify neuronal progenitors in the cortical VZ/SVZ niche that produce a subset of GABAergic interneurons. Our findings suggest that these progenitors constitute an evolutionary novelty contributing to the elaboration of higher cognitive functions in primates.

Summary Adult neurogenesis requires the precise control of neuronal versus astrocyte lineage determination in neural stem cells. While microRNAs (miRNAs) are critically involved in this step during development, their actions in adult hippocampal neural stem cells (aNSCs) has been unclear. As entry point to address that question we chose DICER, an endoribonuclease essential for miRNA biogenesis and other RNAi-related processes. By specific ablation of Dicer in aNSCs in vivo and in vitro, we demonstrate that miRNAs are required for the generation of new neurons, but not astrocytes, in the adult murine hippocampus. Moreover, we identify 11 miRNAs, of which 9 have not been previously characterized in neurogenesis, that determine neurogenic lineage fate choice of aNSCs at the expense of astrogliogenesis. Finally, we propose that the 11 miRNAs sustain adult hippocampal neurogenesis through synergistic modulation of 26 putative targets from different pathways., Graphical Abstract, Highlights • Dicer depletion in aNSCs impairs neurogenesis and stimulates astrogliogenesis • Synergy of 11 miRNAs sustains neuronal fate of aNSCs • miRNA converge on multiple targets in different pathways to induce neurogenesis, In this article, the authors demonstrate that Dicer-dependent miRNAs are required for the generation of new neurons, but not astrocytes, in the adult hippocampus in vivo and in vitro. The authors identify a new set of 11 miRNAs that synergistically converge on multiple targets in different pathways to sustain neurogenic lineage fate commitment in aNSCs.

We have developed a good manufacturing practice for long-term cultivation of fetal human midbrain-derived neural progenitor cells. The generation of human dopaminergic neurons may serve as a tool of either restorative cell therapies or cellular models, particularly as a reference for phenotyping region-specific human neural stem cell lines such as human embryonic stem cells and human inducible pluripotent stem cells. We cultivated 3 different midbrain neural progenitor lines at 10, 12, and 14 weeks of gestation for more than a year and characterized them in great detail, as well as in comparison with Lund mesencephalic cells. The whole cultivation process of tissue preparation, cultivation, and cryopreservation was developed using strict serum-free conditions and standardized operating protocols under clean-room conditions. Long-term-cultivated midbrain-derived neural progenitor cells retained stemness, midbrain fate specificity, and floorplate markers. The potential to differentiate into authentic A9-specific dopaminergic neurons was markedly elevated after prolonged expansion, resulting in large quantities of functional dopaminergic neurons without genetic modification. In restorative cell therapeutic approaches, midbrain-derived neural progenitor cells reversed impaired motor function in rodents, survived well, and did not exhibit tumor formation in immunodeficient nude mice in the short or long term (8 and 30 weeks, respectively). We conclude that midbrain-derived neural progenitor cells are a promising source for human dopaminergic neurons and suitable for long-term expansion under good manufacturing practice, thus opening the avenue for restorative clinical applications or robust cellular models such as high-content or high-throughput screening.

The capsaicin receptor (TRPV1, transient receptor potential vanilloid type 1) was first discovered in the peripheral nervous system as a detector of noxious chemical and thermal stimuli including the irritant chili pepper. Recently, there has been increasing evidence of TRPV1 expression in the central nervous system. Here, we show that TRPV1 is expressed in neural precursor cells (NPCs) during postnatal development, but not in the adult. However, expression of TRPV1 is induced in the adult in paradigms linked to an increase in neurogenesis, such as spatial learning in the Morris water maze or voluntary exercise. Loss of TRPV1 expression in knockout mice leads to an increase in NPC proliferation. Functional TRPV1 expression has been confirmed in cultured NPCs. Our results indicate that TRPV1 expression influences both postnatal and activity-induced neurogenesis in adulthood. Stem Cells 2014;32:3183–3195

Adult neural progenitor cells (aNPC) are a potential autologous cell source for cell replacement in neurologic diseases or for cell-based gene therapy of neurometabolic diseases. Easy accessibility, long-term expandability, and detailed characterization of neural progenitor cell (NPC) properties are important requisites for their future translational/clinical applications. aNPC can be isolated from different regions of the adult human brain, including the accessible subcortical white matter (aNPCWM), but systematic studies comparing long-term expanded aNPCWM with aNPC from neurogenic brain regions are not available. Freshly isolated cells from subcortical white matter and hippocampus expressed oligodendrocyte progenitor cell markers such as A2B5, neuron-glial antigen 2 (NG2), and oligodendrocyte transcription factor 2 (OLIG2) in ∼20% of cells but no neural stem cell (NSC) markers such as CD133 (Prominin1), Nestin, SOX2, or PAX6. The epidermal growth factor receptor protein was expressed in 18% of aNPCWM and 7% of hippocampal aNPC (aNPCHIP), but only a small fraction of cells, 1 of 694 cells from white matter and 1 of 1,331 hippocampal cells, was able to generate neurospheres. Studies comparing subcortical aNPCWM with their hippocampal counterparts showed that both NPC types expressed mainly markers of glial origin such as NG2, A2B5, and OLIG2, and the NSC/NPC marker Nestin, but no pericyte markers. Both NPC types were able to produce neurons, astrocytes, and oligodendrocytes in amounts comparable to fetal NSC. Whole transcriptome analyses confirmed the strong similarity of aNPCWM to aNPCHIP. Our data show that aNPCWM are multipotent NPC with long-term expandability similar to NPC from hippocampus, making them a more easily accessible source for possible autologous NPC-based treatment strategies.

International audience; During embryonic development, the rostral neuroectoderm is regionalized into broad areas that are subsequently subdivided into progenitor compartments with specialized identity and fate. These events are controlled by signals emitted by organizing centers and interpreted by target progenitors, which activate superimposing waves of intrinsic factors restricting their identity and fate. The transcription factor Otx2 plays a crucial role in mesencephalic development by positioning the midbrain-hindbrain boundary (MHB) and its organizing activity. Here, we investigated whether Otx2 is cell-autonomously required to control identity and fate of dorsal mesencephalic progenitors. With this aim, we have inactivated Otx2 in the Pax7(+) dorsal mesencephalic domain, previously named m1, without affecting MHB integrity. We found that the Pax7(+) m1 domain can be further subdivided into a dorsal Zic1(+) m1a and a ventral Zic1(-) m1b sub-domain. Loss of Otx2 in the m1a (Pax7(+) Zic1(+)) sub-domain impairs the identity and fate of progenitors, which undergo a full switch into a coordinated cerebellum differentiation program. By contrast, in the m1b sub-domain (Pax7(+) Zic1(-)) Otx2 is prevalently required for post-mitotic transition of mesencephalic GABAergic precursors. Moreover, genetic cell fate, BrdU cell labeling and Otx2 conditional inactivation experiments indicate that in Otx2 mutants all ectopic cerebellar cell types, including external granule cell layer (EGL) precursors, originate from the m1a progenitor sub-domain and that reprogramming of mesencephalic precursors into EGL or cerebellar GABAergic progenitors depends on temporal sensitivity to Otx2 ablation. Together, these findings indicate that Otx2 intrinsically controls different aspects of dorsal mesencephalic neurogenesis. In this context, Otx2 is cell-autonomously required in the m1a sub-domain to suppress cerebellar fate and promote mesencephalic differentiation independently of the MHB organizing activity.

Wnt signaling is known to play crucial roles in the development of multiple organs as well as in cancer. In particular, constitutive activation of Wnt/β-Catenin signaling in distinct populations of forebrain or brainstem precursor cells has previously been shown to result in dramatic brain enlargement during embryonic stages of development as well as in the formation of medulloblastoma, a malignant brain tumor in childhood. In order to extend this knowledge to postnatal stages of both cerebral and cerebellar cortex development, we conditionally activated Wnt signaling by introducing a dominant active form of β-catenin in hGFAP-positive neural precursors. Such mutant mice survived up to 21 days postnatally. While the mice revealed enlarged ventricles and an initial expansion of the Pax6-positive ventricular zone, Pax6 expression and proliferative activity in the ventricular zone was virtually lost by embryonic day 16.5. Loss of Pax6 expression was not followed by expression of the subventricular zone marker Tbr2, indicating insufficient neuronal differentiation. In support of this finding, cortical thickness was severely diminished in all analyzed stages from embryonic day 14.5 to postnatal day 12, and appropriate layering was not detectable. Similarly, cerebella of hGFAP-cre::Ctnnb1(ex3)Fl/+ mice were hypoplastic and displayed severe lamination defects. Constitutively active β-Catenin induced inappropriate proliferation of granule neurons and inadequate development of Bergmann glia, thereby preventing regular migration of granule cells and normal cortical layering. We conclude that Wnt signaling has divergent roles in the central nervous system and that Wnt needs to be tightly controlled in a time- and cell type-specific manner.

Reprogramming of somatic cells into induced pluripotent stem cells (iPSCs) by retroviral overexpression of the transcription factors Oct4, Sox2, Klf4, and c-Myc holds great promise for the development of personalized cell replacement therapies. In an attempt to minimize the risk for chromosomal disruption and to simplify reprogramming, several studies demonstrated that a reduced set of reprogramming factors is sufficient to generate iPSC, albeit at lower efficiency. To elucidate the influence of factor reduction on subsequent differentiation, we compared the efficiency of neuronal differentiation in iPSC generated from postnatal murine neural stem cells with either one (Oct4; iPSC(1F-NSC) ), two (Oct4, Klf4; iPSC(2F-NSC) ), or all four factors (iPSC(4F-NSC) ) with those of embryonic stem cells (ESCs) and iPSC produced from fibroblasts with all four factors (iPSC(4F-MEF) ). After 2 weeks of coculture with PA6 stromal cells, neuronal differentiation of iPSC(1F-NSC) and iPSC(2F-NSC) was less efficient compared with iPSC(4F-NSC) and ESC, yielding lower proportions of colonies that stained positive for early and late neuronal markers. Electrophysiological analyses after 4 weeks of differentiation identified functional maturity in neurons differentiated from ESC, iPSC(2F-NSC) , iPSC(4F-NSC) , and iPSC(4F-MEF) but not in those from iPSC(1F-NSC) . Similar results were obtained after hematoendothelial differentiation on OP9 bone marrow stromal cells, where factor-reduced iPSC generated lower proportions of colonies with hematoendothelial progenitors than colonies of ESC, iPSC(4F-NSC) , and iPSC(4F-MEF) . We conclude that a reduction of reprogramming factors does not only reduce reprogramming efficiency but may also worsen subsequent differentiation and hinder future application of iPSC in cell replacement therapies.

SummaryRecent advances have suggested that direct induction of neural stem cells (NSCs) could provide an alternative to derivation from somatic tissues or pluripotent cells. Here we show direct derivation of stably expandable NSCs from mouse fibroblasts through a curtailed version of reprogramming to pluripotency. By constitutively inducing Sox2, Klf4, and c-Myc while strictly limiting Oct4 activity to the initial phase of reprogramming, we generated neurosphere-like colonies that could be expanded for more than 50 passages and do not depend on sustained expression of the reprogramming factors. These induced neural stem cells (iNSCs) uniformly display morphological and molecular features of NSCs, such as the expression of Nestin, Pax6, and Olig2, and have a genome-wide transcriptional profile similar to that of brain-derived NSCs. Moreover, iNSCs can differentiate into neurons, astrocytes, and oligodendrocytes. Our results demonstrate that functional NSCs can be generated from somatic cells by factor-driven induction.

The p53-family member, p73, plays a key role in the development of the central nervous system (CNS), in senescence, and in tumor formation. The role of p73 in neuronal differentiation is complex and involves several downstream pathways. Indeed, in the last few years, we have learnt that TAp73 directly or indirectly regulates several genes involved in neural biology. In particular, TAp73 is involved in the maintenance of neural stem/progenitor cell self-renewal and differentiation throughout the regulation of SOX-2, Hey-2, TRIM32 and Notch. In addition, TAp73 is also implicated in the regulation of the differentiation and function of postmitotic neurons by regulating the expression of p75NTR and GLS2 (glutamine metabolism). Further still, the regulation of miR-34a by TAp73 indicates that microRNAs can also participate in this multifunctional role of p73 in adult brain physiology. However, contradictory results still exist in the relationship between p73 and brain disorders, and this remains an important area for further investigation.

Alterations of mitochondrial metabolism and genomic instability have been implicated in tumorigenesis in multiple tissues. High-grade glioma (HGG), one of the most lethal human neoplasms, displays genetic modifications of Krebs cycle components as well as electron transport chain (ETC) alterations. Furthermore, the p53 tumor suppressor, which has emerged as a key regulator of mitochondrial respiration at the expense of glycolysis, is genetically inactivated in a large proportion of HGG cases. Therefore, it is becoming evident that genetic modifications can affect cell metabolism in HGG; however, it is currently unclear whether mitochondrial metabolism alterations could vice versa promote genomic instability as a mechanism for neoplastic transformation. Here, we show that, in neural progenitor/stem cells (NPCs), which can act as HGG cell of origin, inhibition of mitochondrial metabolism leads to p53 genetic inactivation. Impairment of respiration via inhibition of complex I or decreased mitochondrial DNA copy number leads to p53 genetic loss and a glycolytic switch. p53 genetic inactivation in ETC-impaired neural stem cells is caused by increased reactive oxygen species and associated oxidative DNA damage. ETC-impaired cells display a marked growth advantage in the presence or absence of oncogenic RAS, and form undifferentiated tumors when transplanted into the mouse brain. Finally, p53 mutations correlated with alterations in ETC subunit composition and activity in primary glioma-initiating neural stem cells. Together, these findings provide previously unidentified insights into the relationship between mitochondria, genomic stability, and tumor suppressive control, with implications for our understanding of brain cancer pathogenesis.

In recent years, tremendous progress has been made in identifying novel mechanisms and new medications that regulate immune cell function in multiple sclerosis (MS). However, a significant unmet need is the identification of the mechanisms underlying neurodegeneration, because patients continue to manifest brain atrophy and disability despite current therapies. Neural and mesenchymal stem cells have received considerable attention as therapeutic candidates to ameliorate the disease in preclinical and phase I clinical trials. More recently, progress in somatic cell reprogramming and induced pluripotent stem cell technology has allowed the generation of human “diseased” neurons in a patient-specific setting and has provided a unique biological tool that can be used to understand the cellular and molecular mechanisms of neurodegeneration. In the present review, we discuss the application and challenges of these technologies, including the generation of neurons, oligodendrocytes, and oligodendrocyte progenitor cells (OPCs) from patients and novel stem cell and OPC cellular arrays, in the discovery of new mechanistic insights and the future development of MS reparative therapies.

Reduced oxygen levels (1-5% O2, named herein 'physioxia') are beneficial for stem cell cultures leading to enhanced proliferation, better survival and higher differentiation potential, but the underlying molecular mechanisms remain elusive. A potential link between physioxia and the canonical Wnt pathway was found recently, but the differential involvement of this signalling pathway for the various stem cell properties such as proliferation, stem cell maintenance, and differentiation capacity remains enigmatic. We here demonstrate increased Wnt target gene transcription and stabilised active β-catenin upon physioxic cell culture in primary tissue-specific foetal mouse neural stem cells. Knock-out of the main oxygen sensing molecule, hypoxia-inducible factor-1α (Hif-1α), had no impact on Wnt activation assuming that physioxia induces the Wnt pathway independently of Hif-1α. To determine the physiological relevance of physioxia-induced Wnt/β-catenin signalling, we examined proliferation, cell cycle kinetics, survival and stem cell maintenance upon Wnt activation and inhibition. Whereas survival and stem cell maintenance seem to be independent of the Wnt pathway, our studies provide first evidence that Wnt/β-catenin signalling positively stimulates proliferation of physioxic cells by affecting cell cycle regulation. Together, our results provide mechanistic insight into oxygen-mediated regulation of the self-renewal activity of neural stem cells.

Gain-of-function mutations in histone 3 (H3) variants are found in a substantial proportion of pediatric high-grade gliomas (pHGG), often in association with TP53 loss and platelet-derived growth factor receptor alpha (PDGFRA) amplification. Here, we describe a somatic mouse model wherein H3.3K27M and Trp53 loss alone are sufficient for neoplastic transformation if introduced in utero. H3.3K27M-driven lesions are clonal, H3K27me3 depleted, Olig2 positive, highly proliferative, and diffusely spreading, thus recapitulating hallmark molecular and histopathological features of pHGG. Addition of wild-type PDGFRA decreases latency and increases tumor invasion, while ATRX knockdown is associated with more circumscribed tumors. H3.3K27M-tumor cells serially engraft in recipient mice, and preliminary drug screening reveals mutation-specific vulnerabilities. Overall, we provide a faithful H3.3K27M-pHGG model which enables insights into oncohistone pathogenesis and investigation of future therapies.

Mitochondrial DNA (mtDNA) mutations frequently cause neurological diseases. Modeling of these defects has been difficult because of the challenges associated with engineering mtDNA. We show here that neural progenitor cells (NPCs) derived from human induced pluripotent stem cells (iPSCs) retain the parental mtDNA profile and exhibit a metabolic switch toward oxidative phosphorylation. NPCs derived in this way from patients carrying a deleterious homoplasmic mutation in the mitochondrial gene MT-ATP6 (m.9185T>C) showed defective ATP production and abnormally high mitochondrial membrane potential (MMP), plus altered calcium homeostasis, which represents a potential cause of neural impairment. High-content screening of FDA-approved drugs using the MMP phenotype highlighted avanafil, which we found was able to partially rescue the calcium defect in patient NPCs and differentiated neurons. Overall, our results show that iPSC-derived NPCs provide an effective model for drug screening to target mtDNA disorders that affect the nervous system.

Neurogenesis in the adult dentate gyrus (DG) generates new granule neurons that differentiate in the inner one-third of the granule cell layer (GCL). The migrating precursors of these neurons arise from neural stem cells (NSCs) in the subgranular zone (SGZ). Although it is established that pathological conditions, including epilepsy and stroke, cause dispersion of granule neuron precursors, little is known about the factors that regulate their normal placement. Based on the high expression of the chemokine CXCL12 in the adult GCL and its role in guiding neuronal migration in development, we addressed the function of the CXCL12 receptor CXCR4 in adult neurogenesis. Using transgenic reporter mice, we detected Cxcr4-GFP expression in NSCs, neuronal-committed progenitors, and immature neurons of adult and aged mice. Analyses of hippocampal NSC cultures and hippocampal tissue by immunoblot and immunohistochemistry provided evidence for CXCL12-promoted phosphorylation/activation of CXCR4 receptors in NSCs in vivo and in vitro. Cxcr4 deletion in NSCs of the postnatal or mature DG using Cre technology reduced neurogenesis. Fifty days after Cxcr4 ablation in the mature DG, the SGZ showed a severe reduction of Sox2-positive neural stem/early progenitor cells, NeuroD-positive neuronal-committed progenitors, and DCX-positive immature neurons. Many immature neurons were ectopically placed in the hilus and inner molecular layer, and some developed an aberrant dendritic morphology. Only few misplaced cells survived permanently as ectopic neurons. Thus, CXCR4 signaling maintains the NSC pool in the DG and specifies the inner one-third of the GCL as differentiation area for immature granule neurons.

As a result of brain injury, astrocytes become activated and start to proliferate in the vicinity of the injury site. Recently, we had demonstrated that these reactive astrocytes, or glia, can form self-renewing and multipotent neurospheres in vitro. In the present study, we demonstrate that it is only invasive injury, such as stab wounding or cerebral ischemia, and not noninvasive injury conditions, such as chronic amyloidosis or induced neuronal death, that can elicit this increase in plasticity. Furthermore, we find that Sonic hedgehog (SHH) is the signal that acts directly on the astrocytes and is necessary and sufficient toelicit the stem cell response both in vitro and invivo. These findings provide a molecular basis for how cells with neural stem cell lineage emerge at sites of brain injury and imply that the high levels of SHH known to enter the brain from extraneural sources after invasive injury can trigger this response.

Summary Recent studies have shown that defined sets of transcription factors can directly reprogram differentiated somatic cells to a different differentiated cell type without passing through a pluripotent state, but the restricted proliferative and lineage potential of the resulting cells limits the scope of their potential applications. Here we show that a combination of transcription factors ( Brn4 / Pou3f4 , Sox2 , Klf4 , c-Myc , plus E47 / Tcf3 ) induces mouse fibroblasts to directly acquire a neural stem cell identity—which we term as induced neural stem cells (iNSCs). Direct reprogramming of fibroblasts into iNSCs is a gradual process in which the donor transcriptional program is silenced over time. iNSCs exhibit cell morphology, gene expression, epigenetic features, differentiation potential, and self-renewing capacity, as well as in vitro and in vivo functionality similar to those of wild-type NSCs. We conclude that differentiated cells can be reprogrammed directly into specific somatic stem cell types by defined sets of specific transcription factors.

Adult neurogenesis is an exceptional feature of the adult brain and in an intriguing way bridges between neuronal and glial neurobiology. Essentially, all classes of glial cells are directly or indirectly linked to this process. Cells with astrocytic features, for example, serve as radial glia-like stem cells in the two neurogenic regions of the adult brain, the hippocampal dentate gyrus and the subventricular zone of the lateral ventricles, producing new neurons, create a microenvironment permissive for neurogenesis, and are themselves generated alongside the new neurons in an associated but independently regulated process. Oligodendrocytes are generated from precursor cells intermingled with those generating neurons in an independent lineage. NG2 cells have certain precursor cell properties and are found throughout the brain parenchyma. They respond to extrinsic stimuli and injury but do not generate neurons even though they can express some preneuronal markers. Microglia have positive and negative regulatory effects as constituents of the ‘‘neurogenic niche’’. Ependymal cells play incompletely understood roles in adult neurogenesis, but under certain conditions might exert (back-up) precursor cell functions. Glial contributions to adult neurogenesis can be direct or indirect and are mediated by mechanisms ranging from gap-junctional to paracrine and endocrine. As the two neurogenic regions differ between each other and both from the non-neurogenic rest of the brain, the question arises in how far regionalization of both the glia-like precursor cells as well as of the glial cells determines sitespecific ‘‘neurogenic permissiveness.’’ In any case, however, ‘‘neurogenesis’’ appears to be an essentially glial achievement. V

Semaphorins and Plexins are cognate ligand-receptor families that regulate important steps during nervous system development. The Plexin-B2 receptor is critically involved in neural tube closure and cerebellar granule cell development, however, its specific ligands have only been suggested by in vitro studies. Here, we show by in vivo and in vitro analyses that the two Semaphorin-4 family members Sema4C and Sema4G are likely to be in vivo ligands of Plexin-B2. The Sema4C and Sema4G genes are expressed in the developing cerebellar cortex, and Sema4C and Sema4G proteins specifically bind to Plexin-B2 expressing cerebellar granule cells. To further elucidate their in vivo function, we have generated and analyzed Sema4C and Sema4G knock-out mouse mutants. Like Plexin-B2−/− mutants, Sema4C−/− mutants reveal exencephaly and subsequent neonatal lethality with partial penetrance. Sema4C−/− mutants that bypass exencephaly are viable and fertile, but display distinctive defects of the cerebellar granule cell layer, including gaps in rostral lobules, fusions of caudal lobules, and ectopic granule cells in the molecular layer. In addition to neuronal defects, we observed in Sema4C−/− mutants also ventral skin pigmentation defects that are similar to those found in Plexin-B2−/− mutants. The Sema4G gene deletion causes no overt phenotype by itself, but combined deletion of Sema4C and Sema4G revealed an enhanced cerebellar phenotype. However, Sema4C/Sema4G double mutants showed overall less severe cerebellar phenotypes than Plexin-B2−/− mutants, indicating that further ligands of Plexin-B2 exist. In explant cultures of the developing cerebellar cortex, Sema4C promoted migration of cerebellar granule cell precursors in a Plexin-B2-dependent manner, supporting the model that a reduced migration rate of granule cell precursors is the basis for the cerebellar defects of Sema4C−/− and Sema4C/Sema4G mutants.

Background Lack of appropriate tools and techniques to study fate and functional integration of newly generated neurons has so far hindered understanding of neurogenesis' relevance under physiological and pathological conditions. Current analyses are either dependent on mitotic labeling, for example BrdU-incorporation or retroviral infection, or on the detection of transient immature neuronal markers. Here, we report a transgenic mouse model (DCX-CreERT2) for time-resolved fate analysis of newly generated neurons. This model is based on the expression of a tamoxifen-inducible Cre recombinase under the control of a doublecortin (DCX) promoter, which is specific for immature neuronal cells in the CNS. Results In the DCX-CreERT2 transgenic mice, expression of CreERT2 was restricted to DCX+ cells. In the CNS of transgenic embryos and adult DCX-CreERT2 mice, tamoxifen administration caused the transient translocation of CreERT2 to the nucleus, allowing for the recombination of loxP-flanked sequences. In our system, tamoxifen administration at E14.5 resulted in reporter gene activation throughout the developing CNS of transgenic embryos. In the adult CNS, neurogenic regions were the primary sites of tamoxifen-induced reporter gene activation. In addition, reporter expression could also be detected outside of neurogenic regions in cells physiologically expressing DCX (e.g. piriform cortex, corpus callosum, hypothalamus). Four weeks after recombination, the vast majority of reporter-expressing cells were found to co-express NeuN, revealing the neuronal fate of DCX+ cells upon maturation. Conclusions This first validation demonstrates that our new DCX-CreERT2 transgenic mouse model constitutes a powerful tool to investigate neurogenesis, migration and their long-term fate of neuronal precursors. Moreover, it allows for a targeted activation or deletion of specific genes in neuronal precursors and will thereby contribute to unravel the molecular mechanisms controlling neurogenesis.