Number Of Neural Stem Cells Research Articles

Multi-Session Transcranial Direct Current Stimulation (tDCS) Elicits Inflammatory and Regenerative Processes in the Rat Brain

Transcranial direct current stimulation (tDCS) is increasingly being used in human studies as an adjuvant tool to promote recovery of function after stroke. However, its neurobiological effects are still largely unknown. Electric fields are known to influence the migration of various cell types in vitro, but effects in vivo remain to be shown. Hypothesizing that tDCS might elicit the recruitment of cells to the cortex, we here studied the effects of tDCS in the rat brain in vivo. Adult Wistar rats (n = 16) were randomized to either anodal or cathodal stimulation for either 5 or 10 consecutive days (500 µA, 15 min). Bromodeoxyuridine (BrdU) was given systemically to label dividing cells throughout the experiment. Immunohistochemical analyses ex vivo included stainings for activated microglia and endogenous neural stem cells (NSC). Multi-session tDCS with the chosen parameters did not cause a cortical lesion. An innate immune response with early upregulation of Iba1-positive activated microglia occurred after both cathodal and anodal tDCS. The involvement of adaptive immunity as assessed by ICAM1-immunoreactivity was less pronounced. Most interestingly, only cathodal tDCS increased the number of endogenous NSC in the stimulated cortex. After 10 days of cathodal stimulation, proliferating NSC increased by ∼60%, with a significant effect of both polarity and number of tDCS sessions on the recruitment of NSC. We demonstrate a pro-inflammatory effect of both cathodal and anodal tDCS, and a polarity-specific migratory effect on endogenous NSC in vivo. Our data suggest that tDCS in human stroke patients might also elicit NSC activation and modulate neuroinflammation.

Hormesis, Cell Death, and Regenerative Medicine for Neurodegenerative Diseases

Although the adult human brain has a small number of neural stem cells, they are insufficient to repair the damaged brain to achieve significant functional recovery for neurodegenerative diseases and stroke. Stem cell therapy, by either enhancing endogenous neurogenesis, or transplanting stem cells, has been regarded as a promising solution. However, the harsh environment of the diseased brain posts a severe threat to the survival and correct differentiation of those new stem cells. Hormesis (or preconditioning, stress adaptation) is an adaptation mechanism by which cells or organisms are potentiated to survive an otherwise lethal condition, such as the harsh oxidative stress in the stroke brain. Stem cells treated by low levels of chemical, physical, or pharmacological stimuli have been shown to survive better in the neurodegenerative brain. Thus combining hormesis and stem cell therapy might improve the outcome for treatment of these diseases. In addition, since the cell death patterns and their underlying molecular mechanism may vary in different neurodegenerative diseases, even in different progression stages of the same disease, it is essential to design a suitable and optimum hormetic strategy that is tailored to the individual patient.

Spatiotemporal Changes to the Subventricular Zone Stem Cell Pool through Aging

Through adulthood, the rodent subventricular zone (SVZ) stem cell niche generates new olfactory bulb interneurons. We had previously reported that the number of new neurons produced in the SVZ declines through aging; however, age-related changes attributable specifically to the SVZ neural stem cell (NSC) population have not been fully characterized. Here, we conducted a spatiotemporal evaluation of adult SVZ NSCs. We assessed ventricle-contacting NSCs, which together with ependymal cells form regenerative units (pinwheels) along the lateral wall of the lateral ventricle. Based on their apical GFAP-expressing process, individual NSCs were identified across the ventricle surface using serial reconstruction of the SVZ. We observed an 86% decline in total NSCs/mm² of intact ependyma in 2-year old versus 3-month-old mice, with fewer NSC processes within each aged pinwheel. This resulted in an associated 78% decline in total pinwheel units/mm². Regional analysis along the lateral ventricle surface revealed that the age-dependent decline of NSCs and pinwheels is spatially uniform and ultimately maintains the conserved ratio of olfactory bulb interneuron subtypes generated in young mice. However, the overall neurogenic output of the aged SVZ is reduced. Surprisingly, we found no significant change in the number of actively proliferating NSCs per mm² of ventricle surface. Instead, our data reveal that, although the total NSC number, pinwheel units and NSCs per pinwheel decline with age, the percentage of actively, mitotic NSCs increases, indicating that age-related declines in SVZ-mediated olfactory bulb neurogenesis occur downstream of NSC proliferation.

Reply: Quantitative evaluation of the human subventricular zone

ARTICLE Sir, We would like to thank Hoglinger and colleagues for their interest in our recent publication in Brain (van den Berge et al ., 2011). In this study, we showed that the number of adult neural stem cells and precursors in the main neurogenic niche of the human brain, the subventricular zone, is not significantly diminished in patients with Parkinson’s disease compared to age- and sex-matched controls. In addition, we provided evidence that two different human neural stem cell lines did not respond with an increase in cell proliferation following exposure to dopamine or dopamine agonists. We also examined the number of proliferating neural stem cells and precursors in the subventricular zone of mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a mouse model for Parkinson’s disease, and these results supported our findings in the human brain. Our data, however, are inconsistent with three previous studies (Hoglinger et al ., 2004; O'Keeffe et al ., 2009; O'Sullivan et al ., 2011). In a letter to the editor, Hoglinger and colleagues argue that our data should be interpreted with caution, and they state that our anatomical definition of the region of interest, the sampling procedure, some of the immunostaining procedures, and the quantification methods were of limited precision. Here, we would like to extend on the used methodologies as described in the methods section and supplementary material of our article (van den Berge et al ., 2011) and we would like to take the opportunity to clarify the issues raised by Hoglinger and colleagues. We agree that the human subventricular zone is highly variable in width, as we have shown before (van den Berge et al ., 2010), and that anatomical matching between donors is necessary for obtaining reproducible quantitative data. In our recent publication (van den Berge et al …

Effects of minocycline on endogenous neural stem cells after experimental stroke

Minocycline has been reported to reduce infarct size after focal cerebral ischemia, due to an attenuation of microglia activation and prevention of secondary damage from stroke-induced neuroinflammation. We here investigated the effects of minocycline on endogenous neural stem cells (NSCs) in vitro and in a rat stroke model.Primary cultures of fetal rat NSCs were exposed to minocycline to characterize its effects on cell survival and proliferation. To assess these effects in vivo, permanent cerebral ischemia was induced in adult rats, treated systemically with minocycline or placebo. Imaging 7days after ischemia comprised (i) Magnetic Resonance Imaging (MRI), assessing the extent of infarcts, (ii) Positron Emission Tomography (PET) with [11C]PK11195, characterizing neuroinflammation, and (iii) PET with 3’-deoxy-3’-[18F]fluoro-L-thymidine ([18F]FLT), detecting proliferating endogenous NSCs. Immunohistochemistry was used to verify ischemic damage and characterize cellular inflammatory and repair processes in more detail.In vitro, specific concentrations of minocycline significantly increased NSC numbers without increasing their proliferation, indicating a positive effect of minocycline on NSC survival. In vivo, endogenous NSC activation in the subventricular zone (SVZ) measured by [18F]FLT PET correlated well with infarct volumes. Similar to in vitro findings, minocycline led to a specific increase in endogenous NSC activity in both the SVZ as well as the hippocampus. [11C]PK11195 PET detected neuroinflammation in the infarct core as well as in peri-infarct regions, with both its extent and location independent of the infarct size. The data did not reveal an effect of minocycline on stroke-induced neuroinflammation.We show that multimodal PET imaging can be used to characterize and quantify complex cellular processes occurring after stroke, as well as their modulation by therapeutic agents. We found minocycline, previously implied in attenuating microglial activation, to have positive effects on endogenous NSC survival. These findings hold promise for the development of novel treatments in stroke therapy.

Neural stem cells: Brain building blocks and beyond

Neural stem cells are the origins of neurons and glia and generate all the differentiated neural cells of the mammalian central nervous system via the formation of intermediate precursors. Although less frequent, neural stem cells persevere in the postnatal brain where they generate neurons and glia. Adult neurogenesis occurs throughout life in a few limited brain regions. Regulation of neural stem cell number during central nervous system development and in adult life is associated with rigorous control. Failure in this regulation may lead to e.g. brain malformation, impaired learning and memory, or tumor development. Signaling pathways that are perturbed in glioma are the same that are important for neural stem cell self-renewal, differentiation, survival, and migration. The heterogeneity of human gliomas has impeded efficient treatment, but detailed molecular characterization together with novel stem cell-like glioma cell models that reflect the original tumor gives opportunities for research into new therapies. The observation that neural stem cells can be isolated and expanded in vitro has opened new avenues for medical research, with the hope that they could be used to compensate the loss of cells that features in several severe neurological diseases. Multipotent neural stem cells can be isolated from the embryonic and adult brain and maintained in culture in a defined medium. In addition, neural stem cells can be derived from embryonic stem cells and induced pluripotent stem cells by in vitro differentiation, thus adding to available models to study stem cells in health and disease.

Basic fibroblast growth factor increases the number of endogenous neural stem cells and inhibits the expression of amino methyl isoxazole propionic acid receptors in amyotrophic lateral sclerosis mice.

This study aimed to investigate the number of amino methyl isoxazole propionic acid (AMPA) receptors and production of endogenous neural stem cells in the SOD1G93AG1H transgenic mouse model of amyotrophic lateral sclerosis, at postnatal day 60 following administration of basic fibroblast growth factor (FGF-2). A radioligand binding assay and immunohistochemistry were used to estimate the number of AMPA receptors and endogenous neural stem cells respectively. Results showed that the number of AMPA receptors and endogenous neural stem cells in the brain stem and sensorimotor cortex were significantly increased, while motor function was significantly decreased at postnatal days 90 and 120. After administration of FGF-2 into mice, numbers of endogenous neural stem cells increased, while expression of AMPA receptors decreased, whilst motor functions were recovered. At postnatal day 120, the number of AMPA receptors was negatively correlated with the number of endogenous neural stem cells in model mice and FGF-2-treated mice. Our experimental findings indicate that FGF-2 can inhibit AMPA receptors and increase the number of endogenous neural stem cells, thus repairing neural injury in amyotrophic lateral sclerosis mice.

An oblique view on the role of spindle orientation in vertebrate neurogenesis

Neurogenesis is a dynamic process that produces a diverse number of glial and neural cell types from a limited number of neural stem cells throughout development and into adulthood. After an initial period of amplification through symmetric division, neural stem cells rely on asymmetric modes of division to self-renew while producing more committed progeny. Understanding the molecular mechanisms regulating the choice between symmetric and asymmetric modes of division is essential to understand human brain development and pathologies, and to explain the increasing cortical complexity observed in evolution. A popular model states the existence of a causal relationship between the orientation of the axis of division of stem cells and the fate of their progeny in many different tissues, but the validity of the model in neural stem cells is not clear. In this review, we briefly present the diversity of neural stem cells and intermediate progenitors in the developing central nervous system. We then draw a historic overview of the assumed causal relationship between spindle orientation and fate determination. We show how this prompted a search for regulators of spindle orientation, and present the current state of knowledge on the mechanism. Finally, we review data on the effect of defective spindle orientation and try to integrate conflicting observations by presenting alternative mechanisms that may regulate the choice between symmetric and asymmetric outcomes.

Effects of inflammatory cytokines and minocycline on cultured neural stem cells

Introduction: Manipulating endogenous neural stem cells (NSCs) in the adult brain has emerged as an experimental concept, aimed at enhancing the brain’s regenerative capacity after insults such as stroke. Neuroinflammatory processes occuring after stroke are mediated by pro-inflammatory cytokines such as TNF-alpha, interleukin–1ß (IL–1ß) and IL–6; however, their effects on NSCs are unknown to date. The tetracycline minocycline has neuroprotective effects in stroke, but – likewise – its effects on NSCs are unclear to date. Method: NSCs were dissected from fetal rat cortex (E13.5) and grown as monolayer cultures. To study the effects of pharmacological agents on NSCs, cultures were treated with various concentrations or combinations of TNF-alpha, IL–1ß, IL–6, and minocycline, and NSC numbers were assessed over time using a photometric assay (MTT). Furthermore, the effects of those drugs on NSC proliferation and differentiation potential were examined using immunocytochemical methods. Results: 10µM and 50µM of minocycline significantly increased NSC numbers compared to control conditions. This effect could not be attributed to an increase in NSC proliferation, thus indicating a positive effect of minocycline on cell survival. Minocycline alone did not affect the differentiation potential of NSCs. A cocktail of TNF-alpha, IL–1ß and IL–6 neither affected NSC numbers nor their proliferative activity. However, pro-inflammatory cytokines accelerated the differentiation of NSCs, promoting a glial fate. This effect was partly inhibited by co-treatment with minocycline. Conclusion: Our data suggest that minocycline positively affects NSC survival and promotes their tri-potential, undifferentiated state, while pro-inflammatory cytokines typically upregulated after stroke such as TNF-alpha, IL–1ß, and IL–6 accelerate NSC differentiation toward a glial fate. These results should help to establish novel (combinatory) treatments targeting the endogenous NSC niche in stroke.

(−)-Epigallocatechin-3-gallate increases the number of neural stem cells around the damaged area after rat traumatic brain injury

A major component of green tea is (-)-epigallocatechin gallate (EGCG), which has strong antioxidant properties. Here, we investigated the effect of EGCG on neural stem cell (NSC) proliferation around the damaged area following traumatic brain injury (TBI). In this study, male Wistar rats that had access to normal drinking water, or water containing 0.1% (w/v) EGCG, ad libitum received TBI at 10weeks of age. Immunohistochemistry revealed that the number of nestin-positive cells around the damaged area after TBI in the EGCG treatment group increased significantly compared with the normal water group (P<0.05). However, the number of 8-hydroxy-2'-deoxyguanosine-, 4-hydroxy-2-nonenal-, single-stranded DNA (ssDNA)-positive cells and the level of peroxidation around the damaged area after TBI significantly decreased in the EGCG treatment group when compared with the water group (P<0.05). Furthermore, in contrast to the EGCG group, almost all ssDNA-positive cells in the water group co-localized with NeuN and nestin-staining. Ex vivo studies revealed that spheres could only be isolated from injured brain tissue in the water group at 3days following TBI. However, in the EGCG group, spheres could be isolated at both 3 and 7days following TBI. A greater number of spheres could be isolated from the EGCG group, which differentiated into neurons and glia in culture without basic fibroblast growth factor. These results indicate that consumption of water containing EGCG pre- and post-TBI inhibits free radical-induced degradation of NSCs, which have the potential to differentiate into neurons and glia around the area of damage following TBI.

Ars2 maintains neural stem-cell identity through direct transcriptional activation of Sox2

SUMMARYFundamental questions concern the transcriptional networks that control the identity and self-renewal of neural stem cells (NSCs), a specialized subset of astroglial cells endowed with stem properties and neurogenic capacity. We observed that the zinc finger protein Ars2 is expressed by adult NSCs from the subventricular zone (SVZ). Selective knockdown of Ars2 in GFAP+ cells within the adult SVZ depleted NSC number and their neurogenic capacity. These phenotypes were recapitulated in the postnatal SVZ of hGFAP-Cre::Ars2fl/fl conditional knockouts, but were more severe. Ex vivo assays showed that Ars2 was necessary and sufficient to promote NSC self-renewal, by positively regulating the expression of Sox2. Although plant1–3 and animal4,5 orthologs of Ars2 are known for their conserved roles in microRNA biogenesis, we unexpectedly observed that Ars2 retained capacity to promote self-renewal in Drosha and Dicer knockout NSCs. Instead, chromatin immunoprecipitation revealed that Ars2 bound a specific region within the 6kb NSC enhancer of Sox2. This association was RNA-independent, and the bound region was required for Ars2-mediated activation of Sox2. We used gel-shift analysis to confirm direct interaction, and to refine the region bound by Ars2 to a specific conserved DNA sequence. The importance of Sox2 as a critical downstream effector was shown by its ability to restore the self-renewal and multipotency defects of Ars2 knockout NSCs. Altogether, we reveal Ars2 as a novel transcription factor that controls the multipotent progenitor state of NSCs via direct activation of the pluripotency factor Sox2.

Ethanol Affects Differentiation‐Related Pathways and Suppresses Wnt Signaling Protein Expression in Human Neural Stem Cells

Prenatal exposure of the fetus to ethanol (EtOH) can be teratogenic. We previously showed that EtOH alters the cell fate of human neural stem cells (NSC). As Wnt signaling plays an important role in fetal brain development, we hypothesized that EtOH suppresses Wnt signaling protein expression in differentiating NSC and thereby contributes to fetal alcohol spectrum disorder. NSC isolated from fetal human brains were cultured in mitogenic media to induce neurospheres, which were dissociated into single-cell suspensions and used for all experiments. Equal numbers of NSC were cultured on lysine/laminin-coated plates for 96 hours in differentiating media containing 0, 20, or 100 mM EtOH. Total mRNA was isolated from samples containing 0 or 100 mM EtOH and changes in expression of 263 genes associated with neurogenesis and NSC differentiation were determined by Oligo GEArray technology. The biological impact of gene changes was estimated using a systems biology approach with pathway express software and KEGG database. Based on the pathways identified, expression of Wnt proteins (Wnt3a and Wnt5a), Wnt-receptor complex proteins (p-LRP6, LRP6, DVL2, and DVL3), Wnt antagonist Naked-2 (NKD-2), and downstream Wnt proteins (β-catenin, Tyr-p-GSK3β, Ser-p-GSK3β) were analyzed by Western blot. Of the 263 genes examined, the expressions of 22 genes in differentiating NSC were either upwardly or downwardly affected by EtOH. These genes are associated with 5 pathways/cellular processes: axon guidance; hedgehog signaling; TGF-β signaling; cell adhesion molecules; and Wnt signaling. When compared to controls, EtOH, at both 20 and 100 mM concentrations, suppressed the expression of Wnt3a and Wnt5a, receptor complex proteins p-LRP6, LRP6 and DVL2, and cytoplasmic proteins Ser-p-GSK3β and β-catenin. Expression of NKD-2 and DVL3 remained unchanged and the expression of active Tyr-p-GSK3β increased significantly. EtOH can significantly alter neural differentiation pathway-related gene expression and suppress Wnt signaling proteins in differentiating human NSC.

The proliferative capacity of the subventricular zone is maintained in the parkinsonian brain

There are many indications that neurogenesis is impaired in Parkinson's disease, which might be due to a lack of dopamine in the subventricular zone. An impairment in neurogenesis may have negative consequences for the development of new therapeutic approaches in Parkinson's disease, as neural stem cells are a potential source for endogenous repair. In this study, we examined the subventricular zone of 10 patients with Parkinson's disease and 10 age- and sex-matched controls for proliferation and neural stem cell numbers. We also included five cases with incidental Lewy body disease, which showed Parkinson's disease pathology but no clinical symptoms and thus did not receive dopaminergic treatment. We quantified the neural stem cell number and proliferative capacity in the subventricular zone of these three donor groups. We found subventricular neural stem cells in each donor, with a high variation in number. We did not observe significant differences in neural stem cell number or in proliferation between the groups. Additionally, we were able to culture neural stem cells from post-mortem brain of several patients with Parkinson's disease, confirming the presence of viable neural stem cells in these brains. We have also examined the subventricular zone of a chronic, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinson's disease mouse model, and again found no effect of dopaminergic denervation on precursor proliferation. Lastly, we investigated the proliferation capacity of two different human neural stem cell lines in response to dopamine. Both cell lines did not respond with a change in proliferation to treatment with dopamine agonists and an antagonist. In summary, the adult neural stem cell pool in the subventricular zone was not clearly affected in the human parkinsonian brain or a Parkinson's disease mouse model. Furthermore, we did not find evidence that dopamine has a direct effect on human neural stem cell proliferation in vitro. Thus, we conclude that the number of adult neural stem cells is probably not diminished in the parkinsonian brain and that dopamine depletion most likely has no effect on human neural stem cells.

Leptin Induces Neuroprotection Neurogenesis and Angiogenesis after Stroke

Leptin is a potent AMP kinase (AMPK) inhibitor that is central to cell survival. Hence, we explored the effects of leptin on neurogenesis and angiogenesis after stroke. Neural stem cells (NSC) were grown as neurospheres in culture and treated with vehicle or leptin and neurosphere size and terminal differentiation were then determined. We then explored the effects of leptin on endogenous repair mechanisms in vivo. Sabra mice underwent photothrombotic stroke, were given vehicle or leptin and newborn cells were labeled with Bromo-deoxy-Uridine. Functional outcome was studied with the neurological severity score for 90 days post stroke and the brains were then evaluated with immunohistochemistry. In a subset of animals the brains were also evaluated for changes in the expression of leptin receptor and AMPK. In vitro, leptin led to a 2-fold increase in neurosphere size but did not change the differentiation of newborn cells. Following stroke, exogenous leptin led to a 4-fold increase in the number of NSC in the cortex abutting the lesion. There was a 1.5-fold increase in the number of newborn neurons and glia in leptin treated animals. Leptin also significantly increased the number of blood vessels in the peri-lesioned cortex. Leptin treated mice had increased expression of leptin receptor and increased phosphorylated AMPK concentration. Animals treated with leptin also had significantly better functional states. In conclusion, leptin induces neurogenesis and angiogenesis after stroke and leads to increased leptin receptor and pAMPK concentrations. This may explain at least in part the better functional outcome observed in leptin treated animals after stroke.

An Integrated In Vitro and In Vivo High-Throughput Screen Identifies Treatment Leads for Ependymoma

Using a mouse model of ependymoma-a chemoresistant brain tumor-we combined multicell high-throughput screening (HTS), kinome-wide binding assays, and invivo efficacy studies, to identify potential treatments with predicted toxicity against neural stem cells (NSC). We identified kinases within the insulin signaling pathway and centrosome cycle as regulators of ependymoma cell proliferation, and their corresponding inhibitors as potential therapies. FDA approved drugs not currently used to treat ependymoma were also identified that posses selective toxicity against ependymoma cells relative to normal NSCs both invitro and invivo, e.g., 5-fluorouracil. Our comprehensive approach advances understanding of the biology and treatment of ependymoma including the discovery of several treatment leads for immediate clinical translation.

Primary Cilia Regulate Proliferation of Amplifying Progenitors in Adult Hippocampus: Implications for Learning and Memory

Integration of new neurons into the adult hippocampus has been linked to specific types of learning. Primary cilia were found to be required for the formation of adult neural stem cells (NSCs) in the hippocampal dentate gyrus during development. However, the requirement of cilia in maintenance of adult NSCs is unknown. We developed a genetic mouse model in which fetal/perinatal brain development is unaffected, but adult hippocampal neurogenesis is constantly reduced by conditional ablation of primary cilia in adult GFAP(+) neural stem/progenitor cells. We found that this approach specifically reduces the number of hippocampal amplifying progenitors (also called type 2a cells) without affecting the number of radial NSCs (or type 1 cells). Constant reduction of adult hippocampal neurogenesis produced a delay rather than a permanent deficiency in spatial learning without affecting the retention of long-term memories. Decreased neurogenesis also altered spatial novelty recognition and hippocampus-independent cue conditioning. Here, we propose that adult hippocampal newborn neurons increase the efficiency of generating the new representations of spatial memories and that reduction of adult hippocampal neurogenesis may be biased toward cue-based strategies. This novel mouse model provides evidences that cognitive deficits associated with ciliary defects (ciliopathies) might be, in part, mediated by the deficiency of primary cilia in adult hippocampal stem/progenitor cells.

Evaluation of the targeted therapy everolimus on cognitive functions and cerebral plasticity in an animal model.

e13589 Background: Cancer and treatments can induce cognitive impairments in patients, called “chemofog” such as deficits of visual and spatial memories and decline of psychomotor processing speed. Although direct effect of chemotherapy on cognitive functions and neurogenesis has been shown, the impact of targeted therapies has never been investigated. Everolimus, which blocks the mTOR pathway, alters cell proliferation, metabolism and neoangiogenesis. Gene expression in hippocampus implicated in memory processes is regulated by the mTOR signalling pathway. Thus, we used a validated behavioral animal model to test the potential cognitive troubles induced by Everolimus. In addition, ex vivo analyses were conducted on neural cell plasticity involved in hippocampus-dependent behavioral tasks. Methods: Everolimus (5 mg/kg) was daily per os administered in adult C57BL/6J Rj mice for two weeks. Learning and memory processes were evaluated 7 days after the end of the treatment period by means of the Morris water maze and the object recognition tests. Ex vivo, hippocampal neurogenesis was investigated in mice that received BrdU during the 5 last days of the treatment period. Results: Everolimus slowed body weight gain from the last day of the treatment period until the end of behavioral sessions. One week after the end of the treatment, emotional reactivity, spontaneous activity, spatial learning and memory performances, as well as behavioral flexibility and object recognition memory were not impaired by Everolimus. Ex vivo analyses of the number of neural stem cells in neurogenic area showed no modification in treated mice at the end of the period of therapy. Conclusions: At short term, Everolimus did not modify cognitive functions evaluated by means of the hippocampal- and prefrontal-dependent behavioral tasks. These observations differ from our previous studies demonstrating that chemotherapy (5-fluorouracil) led to selective long-term cognitive deficits, i.e. behavioral flexibility and recognition memory, sustained by altered neurogenesis. These results reinforce the interest of developing targeted therapies for cancer treatment.

Division-Coupled Astrocytic Differentiation and Age-Related Depletion of Neural Stem Cells in the Adult Hippocampus

Production of new neurons in the adult hippocampus decreases with age; this decline may underlie age-related cognitive impairment. Here we show that continuous depletion of the neural stem cell pool, as a consequence of their division, may contribute to the age-related decrease in hippocampal neurogenesis. Our results indicate that adult hippocampal stem cells, upon exiting their quiescent state, rapidly undergo a series of asymmetric divisions to produce dividing progeny destined to become neurons and subsequently convert into mature astrocytes. Thus, the decrease in the number of neural stem cells is a division-coupled process and is directly related to their production of new neurons. We presenta scheme of the neurogenesis cascade in the adult hippocampus that includes a proposed "disposable stem cell" model and accounts for the disappearance of hippocampal neural stem cells, the appearance of new astrocytes, and the age-related decline in the production of new neurons.

Fbxw7-dependent Degradation of Notch Is Required for Control of “Stemness” and Neuronal-Glial Differentiation in Neural Stem Cells

Control of the growth and differentiation of neural stem cells is fundamental to brain development and is largely dependent on the Notch signaling pathway. The mechanism by which the activity of Notch is regulated during brain development has remained unclear, however. Fbxw7 (also known as Fbw7, SEL-10, hCdc4, or hAgo) is the F-box protein subunit of an Skp1-Cul1-F-box protein (SCF)-type ubiquitin ligase complex that plays a central role in the degradation of Notch family members. We now show that mice with brain-specific deletion of Fbxw7 (Nestin-Cre/Fbxw7(F/F) mice) die shortly after birth with morphological abnormalities of the brain and the absence of suckling behavior. The maintenance of neural stem cells was sustained in association with the accumulation of Notch1 and Notch3, as well as up-regulation of Notch target genes in the mutant mice. Astrogenesis was also enhanced in the mutant mice in vivo, and the differentiation of neural progenitor cells was skewed toward astrocytes rather than neurons in vitro, with the latter effect being reversed by treatment of the cells with a pharmacological inhibitor of the Notch signaling pathway. Our results thus implicate Fbxw7 as a key regulator of the maintenance and differentiation of neural stem cells in the brain.

Suppression of autophagy by FIP200 inactivation results in deficient self‐renewal of neural stem cells

Autophagy is an evolutionarily conserved process for bulk protein degradation. Recent studies suggested a critical role of basal autophagy to maintain neuronal homeostasis. However, it remains unknown about the functions of autophagy in neural stem cells (NSCs). FIP200 is a component of the ULK1‐Atg13‐FIP200‐Atg101 complex that is essential for the induction of autophagy. By conditional inactivation of FIP200 in adult NSCs using a hGFAP‐Cre, we prove that loss of autophagy causes deficient self‐renewal of NSCs. Aggregation of ubiquitinated proteins and p62 was observed in the subventricular zone (SVZ) and the subgranular zone (SGZ), two areas NSCs predominantly reside in adult brain, in the conditional knockout (CKO) mice. Moreover, CKO mice showed substantially reduced size of olfactory bulb and hippocampus. In contrast to the increased apoptosis of neurons in cerebellum and cortex, little difference in apoptosis was found in SVZ and hippocampus between CKO and control mice. Labeling of these regions with Ki67 or BrdU indicated decreased proliferation in CKO mice. Furthermore, BrdU retention experiments indicated reduced number of NSCs in these mice. Lastly, analysis of isolated NSCs by neurosphere assay showed impaired self‐renewal of NSCs from the CKO mice. Together, these results provide strong genetic evidence for a role of autophagy in the regulation of self renewal of NSCs.