Methods to Culture, Differentiate, and Characterize Neural Stem Cells from the Adult and Embryonic Mouse Central Nervous System

Louis, Sharon A.; Mak, Carmen K. H.; Reynolds, Brent A.

doi:10.1007/978-1-62703-128-8_30

Cited by 47 publications

(57 citation statements)

References 13 publications

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“…ActB was included for comparison, although it does not overlap the periventricular zone in early embryogenesis. Adherent cultures were grown in media favoring proliferation (containing Egf ) or specification and differentiation (growth factor-free), in the presence of 1-100 ng/ml Tgfβ1, ActB or vehicle (Conti et al, 2005;Louis et al, 2013). Cells positive for the neural progenitor marker nestin, Olig2, caspase cleavage and BrdU-labeling were quantified at 12 h and 60 h in proliferation media and at 96 h in differentiation media.…”

Section: Tgfβ1 and Actb Promote Neural Progenitor Viability In Vitrosupporting

confidence: 43%

“…The protocol and proprietary reagents from STEMCELL Technologies were used to generate monolayer neural progenitor cultures (Conti et al, 2005;Louis et al, 2013). Ganglionic eminences from E14 mouse embryos were dissected and triturated, and dissociated cells plated into proprietary proliferation media containing 20 ng/ml recombinant human EGF (Invitrogen).…”

Section: Adherent Neural Progenitor Culturessupporting

confidence: 42%

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Combinatorial actions of Tgfβ and Activin ligands promote oligodendrocyte development and CNS myelination

et al. 2014

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In the embryonic CNS, development of myelin-forming oligodendrocytes is limited by bone morphogenetic proteins, which constitute one arm of the transforming growth factor-β (Tgfβ) family and signal canonically via Smads 1/5/8. Tgfβ ligands and Activins comprise the other arm and signal via Smads 2/3, but their roles in oligodendrocyte development are incompletely characterized. Here, we report that Tgfβ ligands and activin B (ActB) act in concert in the mammalian spinal cord to promote oligodendrocyte generation and myelination. In mouse neural tube, newly specified oligodendrocyte progenitors (OLPs) are first exposed to Tgfβ ligands in isolation, then later in combination with ActB during maturation. In primary OLP cultures, Tgfβ1 and ActB differentially activate canonical Smad3 and non-canonical MAP kinase signaling. Both ligands enhance viability, and Tgfβ1 promotes proliferation while ActB supports maturation. Importantly, co-treatment strongly activates both signaling pathways, producing an additive effect on viability and enhancing both proliferation and differentiation such that mature oligodendrocyte numbers are substantially increased. Co-treatment promotes myelination in OLPneuron co-cultures, and maturing oligodendrocytes in spinal cord white matter display strong Smad3 and MAP kinase activation. In spinal cords of ActB-deficient Inhbb −/− embryos, apoptosis in the oligodendrocyte lineage is increased and OLP numbers transiently reduced, but numbers, maturation and myelination recover during the first postnatal week. Smad3 −/− mice display a more severe phenotype, including diminished viability and proliferation, persistently reduced mature and immature cell numbers, and delayed myelination. Collectively, these findings suggest that, in mammalian spinal cord, Tgfβ ligands and ActB together support oligodendrocyte development and myelin formation.

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Section: Tgfβ1 and Actb Promote Neural Progenitor Viability In Vitrosupporting

confidence: 43%

Section: Adherent Neural Progenitor Culturessupporting

confidence: 42%

Combinatorial actions of Tgfβ and Activin ligands promote oligodendrocyte development and CNS myelination

et al. 2014

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“…The relative content of differentiating NSPC and proliferating NPC in neurospheres and primary neuronal cultures is estimated by using neural colony forming cell assay [11,30]. In addition, adhesion cell culturing on specifi c substrates or 3D scaffolds is sometimes used for solving special problems [22].…”

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confidence: 41%

Analysis of Neural Stem Cells from Human Cortical Brain Structures In Vitro

et al. 2016

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Comparative immunohistochemical analysis of the neocortex from human fetuses showed that neural stem and progenitor cells are present in the brain throughout the gestation period, at least from week 8 through 26. At the same time, neural stem cells from the first and second trimester fetuses differed by the distribution, morphology, growth, and quantity. Immunocytochemical analysis of neural stem cells derived from fetuses at different gestation terms and cultured under different conditions showed their differentiation capacity. Detailed analysis of neural stem cell populations derived from fetuses on gestation weeks 8-9, 18-20, and 26 expressing Lex/SSEA1 was performed.

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“…NS/PCs were prepared from the cortices of 1–2-day-old neonatal SD rats as previously described (27), with minor modifications. Single cells were plated in uncoated T50 flasks at a density of 1x10 5 cells/cm 2 with NS/PC culture medium composed of DMEM/F12 (Invitrogen) supplemented with 2% B27 supplement (Invitrogen), 20 ng/ml epidermal growth factor (EGF; Peprotech, Rocky Hill, NJ, USA) and 20 ng/ml basic fibroblast growth factor (bFGF; Peprotech).…”

Section: Methodsmentioning

confidence: 99%

High-mobility group box 1 released from astrocytes promotes the proliferation of cultured neural stem/progenitor cells

Sun

Luo

et al. 2014

International Journal of Molecular Medicine

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Astrocytes are major components of the adult neurogenic niche and play a crucial role in regulating neural stem cell proliferation and differentiation. Following brain injury, astrocytes become reactive and release high-mobility group box 1 (HMGB1), which plays a crucial role in the inflammatory process. However, although it has been reported that HMGB1 promotes neural stem/progenitor cell (NS/PC) proliferation in the developing brain, whether HMGB1 released by reactive astrocytes regulates NS/PC proliferation remains unknown. In this study, we aimed to investigate whether HMGB1 released from reactive astrocytes enhances NS/PC proliferation and to elucidate the possible mechanisms involved in this process. To evaluate the effects of HMGB1 on NS/PC proliferation, NS/PCs were cultured in HMGB1 culture medium and astrocyte-conditioned medium with or without reactive astrocyte-derived HMGB1 by RNA interference (RNAi). To explore the possible mechanisms, the HMGB1 receptor for advanced glycation endproducts (RAGE) in the NS/PCs was blocked with anti-RAGE antibody, and c-Jun N-terminal protein kinase (JNK) in the NS/PCs was inhibited using the potent JNK inhibitor, SP600125. Our results suggested that HMGB1 released from reactive astrocytes promoted NS/PC proliferation in vitro, and the blockade of RAGE or the inhibition of the JNK signaling pathway in the NS/PCs prevented the HMGB1-induced NS/PC proliferation. Our findings demonstrated that HMGB1 released by reactive astrocytes promoted NS/PC proliferation by binding RAGE and enhancing the phosphorylation of the JNK signaling pathway. These findings support a previously described mechanism of a crosstalk between astrocytes and NS/PCs, and suggest that reactive astrocyte-derived HMGB1 plays an important role in the repair of the central nervous system following brain injury.

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Methods to Culture, Differentiate, and Characterize Neural Stem Cells from the Adult and Embryonic Mouse Central Nervous System

Cited by 47 publications

References 13 publications

Combinatorial actions of Tgfβ and Activin ligands promote oligodendrocyte development and CNS myelination

Combinatorial actions of Tgfβ and Activin ligands promote oligodendrocyte development and CNS myelination

Analysis of Neural Stem Cells from Human Cortical Brain Structures In Vitro

High-mobility group box 1 released from astrocytes promotes the proliferation of cultured neural stem/progenitor cells

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