Prominin-1 Allows Prospective Isolation of Neural Stem Cells from the Adult Murine Hippocampus

Walker, Tara L.; Wierick, Ann; Sykes, Alex M.; Waldau, Ben; Corbeil, Denis; Carmeliet, Peter; Kempermann, Gerd

doi:10.1523/jneurosci.3363-12.2013

Cited by 65 publications

(71 citation statements)

References 52 publications

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“…The precursor cells in the DG are activated by specific types of spatial learning and by stimuli such as environmental enrichment and physical activity, whereas the SVZ precursor cells are activated by olfactory learning and olfactory enrichment. Consistent with this, one of us (TLW) previously demonstrated that the DG contains a population of latent stem and progenitor cells that can be activated by neural excitation [15][16][17][18] . In contrast, we found that the SVZ precursor cells respond quite differently to this stimulus, with a decrease in neurosphere number in response to depolarizing levels of KCl 17.…”

Section: Representative Resultssupporting

confidence: 54%

One Mouse, Two Cultures: Isolation and Culture of Adult Neural Stem Cells from the Two Neurogenic Zones of Individual Mice

Walker

Kempermann

2014

JoVE

127

125

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The neurosphere assay and the adherent monolayer culture system are valuable tools to determine the potential (proliferation or differentiation) of adult neural stem cells in vitro. These assays can be used to compare the precursor potential of cells isolated from genetically different or differentially treated animals to determine the effects of exogenous factors on neural precursor cell proliferation and differentiation and to generate neural precursor cell lines that can be assayed over continuous passages. The neurosphere assay is traditionally used for the post-hoc identification of stem cells, primarily due to the lack of definitive markers with which they can be isolated from primary tissue and has the major advantage of giving a quick estimate of precursor cell numbers in brain tissue derived from individual animals. Adherent monolayer cultures, in contrast, are not traditionally used to compare proliferation between individual animals, as each culture is generally initiated from the combined tissue of between 5-8 animals. However, they have the major advantage that, unlike neurospheres, they consist of a mostly homogeneous population of precursor cells and are useful for following the differentiation process in single cells. Here, we describe, in detail, the generation of neurosphere cultures and, for the first time, adherent cultures from individual animals. This has many important implications including paired analysis of proliferation and/or differentiation potential in both the subventricular zone (SVZ) and dentate gyrus (DG) of treated or genetically different mouse lines, as well as a significant reduction in animal usage.

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Section: Representative Resultssupporting

confidence: 54%

One Mouse, Two Cultures: Isolation and Culture of Adult Neural Stem Cells from the Two Neurogenic Zones of Individual Mice

Walker

Kempermann

2014

JoVE

127

125

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“…No isoform-specific transcripts were detected in cell #6, but primers recognizing all isoforms produced a signal. Our results also show good concordance with previous studies that revealed the expression of only Prominin1 transcript isoforms 1, 2, and 6 in the subventricular zone (SVZ), and the absence of isoforms 3, 4, 5, and 8 (34,37). We also observed that isoform 1 was expressed exclusively in one of the GFP − cells, while other isoforms were present only in GFP + population.…”

Section: Figuresupporting

confidence: 93%

Application of an RNA Amplification Method For Reliable Single-Cell Transcriptome Analysis

et al. 2015

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Diverse cell types have unique transcriptional signatures that are best interrogated at single-cell resolution. Here we describe a novel RNA amplification approach that allows for high fidelity gene profiling of individual cells. This technique significantly diminishes the problem of 3′ bias, enabling detection of all regions of transcripts, including the recognition of mRNA with short or completely absent poly(A) tails, identification of noncoding RNAs, and discovery of the full array of splice isoforms from any given gene product. We assess this technique using statistical and bioinformatics analyses of microarray data to establish the limitations of the method. To demonstrate applicability, we profiled individual cells isolated from the mouse subventricular zone (SVZ)—a well-characterized, discrete yet highly heterogeneous neural structure involved in persistent neurogenesis. Importantly, this method revealed multiple splice variants of key germinal zone gene products within individual cells, as well as an unexpected coexpression of several mRNAs considered markers of distinct and separate SVZ cell types. These findings were independently confirmed using RNA-fluorescence in situ hybridization (RNA-FISH), contributing to the utility of this new technology that offers genomic and transcriptomic analysis of small numbers of dynamic and clinically relevant cells.

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“…Prom1 expression has been reported in ependymal cells and murine hippocampus [10], [11]. In transgenic Prom1-lacZ mice, Prom1/lacZ was co-expressed with Gfap in cells of the subventricular zone (SVZ) having properties of multi-potent self-renewing neural stem cells.…”

Section: Introductionmentioning

confidence: 99%

Prominin-1 (CD133) Defines Both Stem and Non-Stem Cell Populations in CNS Development and Gliomas

et al. 2014

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Prominin-1 (CD133) is a commonly used cancer stem cell marker in central nervous system (CNS) tumors including glioblastoma (GBM). Expression of Prom1 in cancer is thought to parallel expression and function in normal stem cells. Using RNA in situ hybridization and antibody tools capable of detecting multiple isoforms of Prom1, we find evidence for two distinct Prom1 cell populations in mouse brain. Prom1 RNA is first expressed in stem/progenitor cells of the ventricular zone in embryonic brain. Conversely, in adult mouse brain Prom1 RNA is low in SVZ/SGZ stem cell zones but high in a rare but widely distributed cell population (Prom1hi). Lineage marker analysis reveals Prom1hi cells are Olig2+Sox2+ glia but Olig1/2 knockout mice lacking oligodendroglia retain Prom1hi cells. Bromodeoxyuridine labeling identifies Prom1hi as slow-dividing distributed progenitors distinct from NG2+Olig2+ oligodendrocyte progenitors. In adult human brain, PROM1 cells are rarely positive for OLIG2, but express astroglial markers GFAP and SOX2. Variability of PROM1 expression levels in human GBM and patient-derived xenografts (PDX) – from no expression to strong, uniform expression – highlights that PROM1 may not always be associated with or restricted to cancer stem cells. TCGA and PDX data show that high expression of PROM1 correlates with poor overall survival. Within proneural subclass tumors, high PROM1 expression correlates inversely with IDH1 (R132H) mutation. These findings support PROM1 as a tumor cell-intrinsic marker related to GBM survival, independent of its stem cell properties, and highlight potentially divergent roles for this protein in normal mouse and human glia.

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Prominin-1 Allows Prospective Isolation of Neural Stem Cells from the Adult Murine Hippocampus

Cited by 65 publications

References 52 publications

One Mouse, Two Cultures: Isolation and Culture of Adult Neural Stem Cells from the Two Neurogenic Zones of Individual Mice

One Mouse, Two Cultures: Isolation and Culture of Adult Neural Stem Cells from the Two Neurogenic Zones of Individual Mice

Application of an RNA Amplification Method For Reliable Single-Cell Transcriptome Analysis

Prominin-1 (CD133) Defines Both Stem and Non-Stem Cell Populations in CNS Development and Gliomas

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