Long-term molecular and cellular stability of human neural stem cell lines

Villa, Anna; Navarro-Galve, Beatriz; Bueno, Carlos; Franco, Sonia; Blasco, Marı́a A.; Martínez‐Serrano, Alberto

doi:10.1016/j.yexcr.2003.11.025

Cited by 79 publications

(67 citation statements)

References 65 publications

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“…Cross-talk between myc and telomerase has been demonstrated [24; 25], and retroviral over-expression of human TERT has been tried for immortalization of neuronal progenitor cells derived from the human fetal spinal cord [26]. Similar to previous observations in another cell line [25], our data indicate that when hc-NSCs differentiate, there is a parallel downregulation of v-myc and TERT expression.…”

Section: Discussionsupporting

confidence: 86%

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Generation of human cortical neurons from a new immortal fetal neural stem cell line

Cacci

Villa

Parmar

et al. 2007

Experimental Cell Research

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Isolation and expansion of neural stem cells (NSCs) of human origin are crucial for successful development of cell therapy approaches in neurodegenerative diseases. Different epigenetic and genetic immortalization strategies have been established for long-term maintenance and expansion of these cells in vitro. Here we report the generation of a new, clonal NSC (hc-NSC) line, derived from human fetal cortical tissue, based on v-myc immortalization. Using immunocytochemistry, we show that these cells retain the characteristics of NSCs after more than 50 passages. Under proliferation conditions, when supplemented with epidermal and basic fibroblast growth factors, the hc-NSCs expressed neural stem/progenitor cell markers like nestin, vimentin and Sox2. When growth factors were withdrawn, proliferation and expression of v-myc and telomerase were dramatically reduced, and the hc-NSCs differentiated into glia. and neurons (mostly glutamatergic and GABAergic, as well as tyrosine hydroxylase-positive, presumably dopaminergic neurons). RT-PCR analysis showed that the hc-NSCs retained expression of Pax6, Emx2 and Neurogenin2, which are genes associated with regionalization and cell commitment in cortical precursors during brain development. Our data indicate that this hc-NSC line could be useful for exploring the potential of human NSCs to replace dead or damaged cortical cells in animal models of acute and chronic neurodegenerative diseases. Taking advantage of its clonality and homogeneity, this cell line will also be a valuable experimental tool to study the regulatory role of intrinsic and extrinsic factors in human NSC biology. (c) 2006 Elsevier Inc. All rights reserved

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Section: Discussionsupporting

confidence: 86%

“…No numerical or structural chromosomal abnormalities were detected. Indirectly, this finding also speaks in favor of good telomere preservation as reported previously [25].…”

Section: Discussionsupporting

confidence: 82%

Generation of human cortical neurons from a new immortal fetal neural stem cell line

Cacci

Villa

Parmar

et al. 2007

Experimental Cell Research

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“…Quantification of the number of double BrdU þ -Nestin þ cells (Figure 7e) indicated that most of the BrdU þ cells were Nestin þ , while approximately one half of the Nestin þ cells incorporated BrdU. This was expected, considering that the hNS1 cells have a cell-cycle length of 40 h 27,28 and that they were exposed to BrdU for 24 h only. Bcl-X L did not influence these figures substantially, but at days 3-4 of differentiation, a two-fold increase in the number of Nestin þ cells also incorporating BrdU was detected.…”

Section: Resultsmentioning

confidence: 71%

“…In this study, we have used control and Bcl-X L overexpressing hNSCs derived from the hNS1 cell line (whose properties have been extensively described elsewhere). 8,27,28,30,37 This is a v-myc immortalized, nontransformed, human fetal forebrain-derived, multipotent, and clonal cell line of neural stem cells. hNS1 cell culture conditions are based on a chemically defined HSC-medium supplemented with 20 ng/ml of each epidermal growth factor (EGF) and basic fibroblast growth factor (FGF2).…”

Section: Methodsmentioning

confidence: 99%

Bcl-XL modulates the differentiation of immortalized human neural stem cells

et al. 2007

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Understanding basic processes of human neural stem cell (hNSC) biology and differentiation is crucial for the development of cell replacement therapies. Bcl-X L has been reported to enhance dopaminergic neuron generation from hNSCs and mouse embryonic stem cells. In this work, we wanted to study, at the cellular level, the effects that Bcl-X L may exert on cell death during differentiation of hNSCs, and also on cell fate decisions and differentiation. To this end, we have used both v-myc immortalized (hNS1 cell line) and non-immortalized neurosphere cultures of hNSCs. In culture, using different experimental settings, we have consistently found that Bcl-X L enhances neuron generation while precluding glia generation. These effects do not arise from a glia-to-neuron shift (changes in fate decisions taken by precursors) or by only cell death counteraction, but, rather, data point to Bcl-X L increasing proliferation of neuronal progenitors, and inhibiting the differentiation of glial precursors. In vivo, after transplantation into the aged rat striatum, Bcl-X L overexpressing hNS1 cells generated more neurons and less glia than the control ones, confirming the results obtained in vitro. These results indicate an action of Bcl-X L modulating hNSCs differentiation, and may be thus important for the future development of cell therapy strategies for the diseased mammalian brain. The efficient generation of human neurons and glia from stem cells is the object of intense investigation, in the context of basic and preclinical cell replacement research. 1,2 Different means of genetic and epigenetic manipulations of stem cell cultures are being tested, aiming at enhancing their ability to generate the desired cell types. [3][4][5][6] In previous studies, we and others have demonstrated that Bcl-X L overexpression has a major impact on the generation of dopaminergic neurons from human neural stem cells (hNSCs) and mouse embryonic stem cells (mES). 7,8 Also, recent data from conditional Bcl-X L mutant mice add support to these observations. 9,10 In the present study, we are extending these observations and analyzing in detail the effects of Bcl-X L on hNSCs differentiation, focusing on precursor cell fate choices, cell death, and precursor proliferation.Bcl-X L is the most potent antiapoptotic protein among Bcl-2 family members, both in vitro and in vivo. 11-14 More specifically, Bcl-X L is essential for neuronal survival during brain development and in the adult central nervous system (CNS) (knockout mice and gene-expression studies 15,16 ). In addition to its antiapoptotic role, recent studies have described new roles of Bcl-X L in cell physiology -effects on Ca 2 þ homeostasis and gene expression 17 and synaptic transmission regulation 18 -under not necessarily apoptotic conditions. Several other studies have also described the role of Bcl-X L in the control of cell cycle, 19,20 a process well known to be tightly linked to progression of precursor cells toward neuronal and glia differentiation. 21,22 In this work, we aimed...

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“…Immortalized human neural stem cell lines are suitable model systems for studying genetic and epigenetic influences on stability over passages, and on stem cell proliferation and differentiation in vitro and in vivo. Their advantage is multipotency within the neural lineage, expandability and stability, making them available over practically indefinite periods of time (Martinez-Serrano et al, 2001;Villa et al, 2002Villa et al, , 2004Kim, 2007). Several studies have described amelioration of behavioral deficits by grafting such immortalized human stem cells in rodent models of neurological disease (Lee et al, 2007), and have characterized their immunocytochemical and biochemical features.…”

mentioning

confidence: 99%

Functional properties of the human ventral mesencephalic neural stem cell line hVM1

Tønnesen

Seiz

Ramos

et al. 2010

Experimental Neurology

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The human fetal ventral mesencephalon-derived stem cell line, hVTVIl, yields high number of tyrosine hydroxylase-expressing presumed dopaminergic neurons upon in vitro differentiation. Here we report that cells generated from this line differentiate into a neuronal phenotype, express electrophysiological properties of functional neurons and respond to neurotransmitters in vitro. However, the electrophysiological properties are immature and the cells require longer maturation time than possible under in vitro

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Long-term molecular and cellular stability of human neural stem cell lines

Cited by 79 publications

References 65 publications

Generation of human cortical neurons from a new immortal fetal neural stem cell line

Generation of human cortical neurons from a new immortal fetal neural stem cell line

Bcl-XL modulates the differentiation of immortalized human neural stem cells

Functional properties of the human ventral mesencephalic neural stem cell line hVM1

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