In Vivo Fate Mapping and Expression Analysis Reveals Molecular Hallmarks of Prospectively Isolated Adult Neural Stem Cells (original) (raw)
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Neuron, 2014
Adult neurogenic niches harbor quiescent neural stem cells; however, their in vivo identity has been elusive. Here, we prospectively isolate GFAP + CD133 + (quiescent neural stem cells [qNSCs]) and GFAP + CD133 + EGFR + (activated neural stem cells [aNSCs]) from the adult ventricular-subventricular zone. aNSCs are rapidly cycling, highly neurogenic in vivo, and enriched in colony-forming cells in vitro. In contrast, qNSCs are largely dormant in vivo, generate olfactory bulb interneurons with slower kinetics, and only rarely form colonies in vitro. Moreover, qNSCs are Nestin negative, a marker widely used for neural stem cells. Upon activation, qNSCs upregulate Nestin and EGFR and become highly proliferative. Notably, qNSCs and aNSCs can interconvert in vitro. Transcriptome analysis reveals that qNSCs share features with quiescent stem cells from other organs. Finally, small-molecule screening identified the GPCR ligands, S1P and PGD 2 , as factors that actively maintain the quiescent state of qNSCs.
Transcriptional Profiling of Adult Neural Stem-Like Cells from the Human Brain
PLoS ONE, 2014
There is a great potential for the development of new cell replacement strategies based on adult human neural stem-like cells. However, little is known about the hierarchy of cells and the unique molecular properties of stem-and progenitor cells of the nervous system. Stem cells from the adult human brain can be propagated and expanded in vitro as free floating neurospheres that are capable of self-renewal and differentiation into all three cell types of the central nervous system. Here we report the first global gene expression study of adult human neural stem-like cells originating from five human subventricular zone biopsies (mean age 42, range 33-60). Compared to adult human brain tissue, we identified 1,189 genes that were significantly up-and downregulated in adult human neural stem-like cells (1% false discovery rate). We found that adult human neural stem-like cells express stem cell markers and have reduced levels of markers that are typical of the mature cells in the nervous system. We report that the genes being highly expressed in adult human neural stem-like cells are associated with developmental processes and the extracellular region of the cell. The calcium signaling pathway and neuroactive ligand-receptor interactions are enriched among the most differentially regulated genes between adult human neural stem-like cells and adult human brain tissue. We confirmed the expression of 10 of the most up-regulated genes in adult human neural stem-like cells in an additional sample set that included adult human neural stem-like cells (n56), foetal human neural stem cells (n51) and human brain tissues (n512). The NGFR, SLITRK6 and KCNS3 receptors were further investigated by immunofluorescence and shown to be heterogeneously expressed in spheres. These receptors could potentially serve as new markers for the identification and characterisation of neural stem-and progenitor cells or as targets for manipulation of cellular fate.
Journal of molecular neuroscience : MN, 2015
Many genes are associated with the differentiation of neural stem cells (NSCs) into astrocytes, the most abundant and functionally diverse population of glial cells in the central nervous system, particularly in the brain. In the present study, we differentiated NSCs from the forebrain of embryonic day 14.5 mouse embryos into astrocytes over 1 and 7 days. We identified transcriptomes of NSCs and astrocytes using RNA sequencing and analyzed enriched gene networks, signal pathways, and ontology. To identify important regulators of differentiation, we performed gene clustering according to expression patterns and promoter CG types. Our data show that genes related to system development, including Fbln2, Bcan, Ncam1, Itih3, Tnr, and Vcan, regulate NSC differentiation through WNT/beta-catenin and epithelial to mesenchymal transition pathways. We identified many CG-rich promoter genes related to basic cellular maintenance such as transcription, translation, and structural components and C...
Stem cells and …, 2006
Multipotent neural stem/progenitor cells (NSPCs) can be isolated from many regions of the adult central nervous system (CNS), yet neurogenesis is restricted to the hippocampus and subventricular zone in vivo. Identification of the molecular cues that modulate NSPC fate choice is a prerequisite for their therapeutic applications. We previously demonstrated that primary astrocytes isolated from regions with higher neuroplasticity, such as newborn and adult hippocampus and newborn spinal cord, promoted neuronal differentiation of adult NSPCs, whereas astrocytes isolated from nonneurogenic of the adult spinal cord inhibited neural differentiation. To identify the factors expressed by these astrocytes that could modulate NSPC differentiation, we performed gene expression profiling analysis using Affymetrix rat genome arrays. Our results demonstrated that these astrocytes had distinct gene expression profiles. We further tested the functional effects of candidate factors that were differentially expressed in neurogenesis-promoting and -inhibiting astrocytes using in vitro NSPC differentiation assays. Our results indicated that two interleukins, IL-1β and IL-6, and a combination of factors that included these two interleukins could promote NSPC neuronal differentiation, whereas insulin-like growth factor binding protein 6 (IGFBP6) and decorin inhibited neuronal differentiation of adult NSPCs. Our results have provided further evidence to support the ongoing hypothesis that, in adult mammalian brains, astrocytes play critical roles in modulating NSPC differentiation. The finding that cytokines and chemokines expressed by astrocytes could promote NSPC neuronal differentiation may help us to understand how injuries induce neurogenesis in adult brains.
Proliferation and cilia dynamics in neural stem cells prospectively isolated from the SEZ
Scientific Reports, 2014
Neural stem cells (NSCs) generate new neurons in vivo and in vitro throughout adulthood and therefore are physiologically and clinically relevant. Unveiling the mechanisms regulating the lineage progression from NSCs to newborn neurons is critical for the transition from basic research to clinical application. However, the direct analysis of NSCs and their progeny is still elusive due to the problematic identification of the cells. We here describe the isolation of highly purified genetically unaltered NSCs and transit-amplifying precursors (TAPs) from the adult subependymal zone (SEZ). Using this approach we show that a primary cilium and high levels of epidermal growth factor receptor (EGFR) at the cell membrane characterize quiescent and cycling NSCs, respectively. However, we also observed non-ciliated quiescent NSCs and NSCs progressing into the cell cycle without up-regulating EGFR expression. Thus, the existence of NSCs displaying distinct molecular and structural conformations provides more flexibility to the regulation of quiescence and cell cycle progression. T he lateral subependymal zone (SEZ) is the largest germinal niche in the adult rodent brain giving rise thousands of new olfactory bulb interneurons every day 1. Nevertheless, neural stem cells (NSCs) in the adult SEZ are largely quiescent 2-4. Upon activation and cell division, NSCs generate rapidly proliferating transit amplifying precursors (TAPs) that progressively differentiate into neuroblasts 5,6. The expression of epidermal growth factor receptor (EGFR) at the cell membrane increases during the transition from NSCs to TAPs and rapidly decreases during neuronal differentiation and cell cycle exit 5. Previous studies have suggested that adult NSCs derive from a subset of embryonic radial glia (RG) precursors 7,8 and maintain RG characteristics such as the expression of Prominin-1 9 , apical/basal polarity, and a primary cilium contacting the lateral ventricle 10-12. Primary cilia are organelles essential for the transduction of key developmental signals 13. In RG their presence and length is negatively correlated with cell cycle progression 14,15. Ablation of primary cilia during development is also associated with an impairment of the transition from RG to adult NSCs 8,16 whereas, cilia deletion in the postnatal niche affects rapid proliferation and quiescence in the hippocampus 17 and in the SEZ 18 , respectively. However, the relationship between cilia, proliferation and lineage progression in the intact adult niche has not been directly investigated. Understanding the mechanisms underlying the progression through the neural lineage is a prerequisite for the manipulation of adult neurogenesis. An important step towards this goal is the purification of NSCs and more differentiated progenitors. We have previously shown that from late development onwards cells expressing high levels of EGFR at the cell membrane (E h) are enriched in clone forming NSCs and especially in TAPs 5,19. Indeed, analysis of EGFR and Prominin-1 expression has been used for the purification of NSCs from the SEZ of adult transgenic mice expressing GFP under the control of the GFAP promoter 18,20. However, it remains still unclear whether all adult NSCs are E h cells or if these cells represent the subset of cycling NSCs. Moreover, the use of reporter genes, which has been instrumental for NSC isolation, involves genetic manipulation, possible leaky reporter expression, and it does not allow the analysis of gene transcription in genetically unmodified cells.
Single-Cell Transcriptome Analyses Reveal Signals to Activate Dormant Neural Stem Cells
Cell, 2015
The scarcity of tissue-specific stem cells and the complexity of their surrounding environment have made molecular characterization of these cells particularly challenging. Through single-cell transcriptome and weighted gene co-expression network analysis (WGCNA), we uncovered molecular properties of CD133(+)/GFAP(-) ependymal (E) cells in the adult mouse forebrain neurogenic zone. Surprisingly, prominent hub genes of the gene network unique to ependymal CD133(+)/GFAP(-) quiescent cells were enriched for immune-responsive genes, as well as genes encoding receptors for angiogenic factors. Administration of vascular endothelial growth factor (VEGF) activated CD133(+) ependymal neural stem cells (NSCs), lining not only the lateral but also the fourth ventricles and, together with basic fibroblast growth factor (bFGF), elicited subsequent neural lineage differentiation and migration. This study revealed the existence of dormant ependymal NSCs throughout the ventricular surface of the CN...
Stem cell reports, 2018
Deciphering the mechanisms that regulate the quiescence of adult neural stem cells (NSCs) is crucial for the development of therapeutic strategies based on the stimulation of their endogenous regenerative potential in the damaged brain. We show that LeX cells sorted from the adult mouse subventricular zone exhibit all the characteristic features of quiescent NSCs. Indeed, they constitute a subpopulation of slowly dividing cells that is able to enter the cell cycle to regenerate the irradiated niche. Comparative transcriptomic analyses showed that they express hallmarks of NSCs but display a distinct molecular signature from activated NSCs (LeXEGFR cells). Particularly, numerous membrane receptors are expressed on quiescent NSCs. We further revealed a different expression pattern of Syndecan-1 between quiescent and activated NSCs and demonstrated its role in the proliferation of activated NSCs. Our data highlight the central role of the stem cell microenvironment in the regulation of...
Diversity of Adult Neural Stem and Progenitor Cells in Physiology and Disease
2021
Adult neural stem and progenitor cells (NSPCs) contribute to learning, memory, maintenance of homeostasis, energy metabolism and many other essential processes. They are highly heterogeneous populations that require input from a regionally distinct microenvironment including a mix of neurons, oligodendrocytes, astrocytes, ependymal cells, NG2+ glia, vasculature, cerebrospinal fluid (CSF), and others. The diversity of NSPCs is present in all three major parts of the CNS, i.e., the brain, spinal cord, and retina. Intrinsic and extrinsic signals, e.g., neurotrophic and growth factors, master transcription factors, and mechanical properties of the extracellular matrix (ECM), collectively regulate activities and characteristics of NSPCs: quiescence/survival, proliferation, migration, differentiation, and integration. This review discusses the heterogeneous NSPC populations in the normal physiology and highlights their potentials and roles in injured/diseased states for regenerative medic...