Niche-Independent Symmetrical Self-Renewal of a Mammalian Tissue Stem Cell

By L. Conti, S. Pollard, T. Gorba, Erika Reitano, M. Toselli, G. Biella, Yirui Sun, S. Sanzone, Qi-Long Ying, E. Cattaneo and 1 more. PLoS Biology, 2005

957 citations recorded by the source corpus.

Pluripotent mouse embryonic stem (ES) cells multiply in simple monoculture by symmetrical divisions. In vivo, however, stem cells are generally thought to depend on specialised cellular microenvironments and to undergo predominantly asymmetric divisions. Ex vivo expansion of pure populations of tissue stem cells has proven elusive. Neural progenitor cells are propagated in combination with differentiating progeny in floating clusters called neurospheres. The proportion of stem cells in neurospheres is low, however, and they cannot be directly observed or interrogated. Here we demonstrate that the complex neurosphere environment is dispensable for stem cell maintenance, and that the combination of fibroblast growth factor 2 (FGF-2) and epidermal growth factor (EGF) is sufficient for derivation and continuous expansion by symmetrical division of pure cultures of neural stem (NS) cells. NS cells were derived first from mouse ES cells. Neural lineage induction was followed by growth factor addition in basal culture media. In the presence of only EGF and FGF-2, resulting NS cells proliferate continuously, are diploid, and clonogenic. After prolonged expansion, they remain able to differentiate efficiently into neurons and astrocytes in vitro and upon transplantation into the adult brain. Colonies generated from single NS cells all produce neurons upon growth factor withdrawal. NS cells uniformly express morphological, cell biological, and molecular features of radial glia, developmental precursors of neurons and glia. Consistent with this profile, adherent NS cell lines can readily be established from foetal mouse brain. Similar NS cells can be generated from human ES cells and human foetal brain. The extrinsic factors EGF plus FGF-2 are sufficient to sustain pure symmetrical self-renewing divisions of NS cells. The resultant cultures constitute the first known example of tissue-specific stem cells that can be propagated without accompanying differentiation. These homogenous cultures will enable delineation of molecular mechanisms that define a tissue-specific stem cell and allow direct comparison with pluripotent ES cells.

Subfields: Biomedical Engineering, Developmental Neuroscience, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/6dc6f6d5743d243c09d49a5e918fba5ea7facaab

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Citations in the corpus

Lineage-specific polycomb targets and de novo DNA methylation define restriction and potential of neuronal progenitors. (2008, background): “However, this same study reports many fewer bivalent domains in hyperproliferative neuronal stem cells (Mikkelsen et al., 2007), which are derived by monolayer differentiation and result in a population clearly distinct from the radialglial cells we analyzed here (Conti et al., 2005).”
Ab initio reconstruction of transcriptomes of pluripotent and lineage committed cells reveals gene structures of thousands of lincRNAs (2010, methodology): “5 ES cells were differentiated into neural progenitor cells (NPC) through embryoid body formation for 4 days and selection in ITSFn media for 5–7 days, and maintained in FGF2 and EGF2 (R&D Systems) as described 27 .”
Oct4-induced pluripotency in adult neural stem cells. (2009, positive): “These cells were cultured in neural expansion medium as previously described (Conti et al., 2005; Kim et al., 2008).”
The Glial Nature of Embryonic and Adult Neural Stem Cells (2009, background): “INM may be a cell-autonomous feature of neuroepithelial or RG-like cells because NSCs derived from embryonic stem cell lines both express RG markers and undergo nucleokinesis as they progress through the cell cycle (Conti et al. 2005, Glaser & Brustle 2005, Elkabetz et al. 2008).”
Glioma stem cell lines expanded in adherent culture have tumor-specific phenotypes and are suitable for chemical and genetic screens. (2009, methodology): “We recently reported methodology for deriving and expanding mouse and human adherent neural stem (NS) cell lines (Conti et al., 2005; Pollard et al., 2006; Sun et al., 2008).”
Promotion of Reprogramming to Ground State Pluripotency by Signal Inhibition (2008, background): “However, in adherent culture in epidermal growth factor (EGF) plus fibroblast growth factor (FGF) the level of neuronal or glial differentiation is less than 1% [6,8].”
A feedback loop comprising lin-28 and let-7 controls pre-let-7 maturation during neural stem-cell commitment (2008, methodology): “NS cultures were prepared by adaptation of dissociated E12 forebrain cells to adherent growth after a brief period of culture as neurosphere”
Histone H3K27ac separates active from poised enhancers and predicts developmental state (2010, methodology): “Given the fact that neural progenitors function as precursors of the neural lineages that can give rise to many adult cell types in the brain, such as adult neurons, oligodendrocytes, and astrocytes (30), it fits that the genes proximal to active H3K27ac-positive enhancers reflect such a progenitor ”
Epigenetic memory in induced pluripotent stem cells (2010, methodology): “Neural Progenitor (NP) iPSC harboring integrated proviruses carrying the four reprogramming factrors 29 were differentiated to neural stem cells (NSC) as described 53 .”
Time-resolved structured illumination microscopy reveals key principles of Xist RNA spreading (2020, methodology): “In order to study Xist behavior in differentiated cells, WT and Ciz1 KO mESCs were differentiated into neural progenitor cells (NPCs) following a following a previously 45 adapted protocol (33,34).”
Distinct factors control histone variant H3.3 localization at specific genomic regions (2010, methodology): “3 remains enriched around the TSS and within the gene body of the housekeeping gene lactate dehydrogenase A Ldha (Figure 3C, Figure S3A).”
Nanog Is the Gateway to the Pluripotent Ground State (2009, positive): “Nanog / NS cells were derived using the established NS cell derivation protocol (Conti et al., 2005) from E12.”
A bivalent chromatin structure marks key developmental genes in embryonic stem cells. (2006, methodology): “These cells uniformly express nestin and Sox2 and upon growth factor withdrawal differentiate into neurons, astrocytes, and oligodendrocytes (Brustle et al., 1999; Conti et al., 2005).”
A combined chemical and genetic approach for the generation of induced pluripotent stem cells. (2008, positive): “For screening purposes, we chose to use mESC-derived NPCs containing a GFP-IRES-Puro/ GiP reporter under the control of the Oct3/ 4 regulatory elements, as mESCs can be grown in large quantities and their differentiation to a homogenous population of NPCs is well defined (Conti et al., 2005).”