Abstract
Neural stem cells (NSCs) have the ability to proliferate and differentiate into neurons and glia. Regulation of NSC fate by small molecules is important for the generation of a certain type of cell. The identification of small molecules that can induce new neurons from NSCs could facilitate regenerative medicine and drug development for neurodegenerative diseases. In this study, we screened natural compounds to identify molecules that are effective on NSC cell fate determination. We found that Kuwanon V (KWV), which was isolated from the mulberry tree (Morus bombycis) root, increased neurogenesis in rat NSCs. In addition, during NSC differentiation, KWV increased cell survival and inhibited cell proliferation as shown by 5-bromo-2-deoxyuridine pulse experiments, Ki67 immunostaining and neurosphere forming assays. Interestingly, KWV enhanced neuronal differentiation and decreased NSC proliferation even in the presence of mitogens such as epidermal growth factor and fibroblast growth factor 2. KWV treatment of NSCs reduced the phosphorylation of extracellular signal-regulated kinase 1/2, increased mRNA expression levels of the cyclin-dependent kinase inhibitor p21, down-regulated Notch/Hairy expression levels and up-regulated microRNA miR-9, miR-29a and miR-181a. Taken together, our data suggest that KWV modulates NSC fate to induce neurogenesis, and it may be considered as a new drug candidate that can regenerate or protect neurons in neurodegenerative diseases.
Highlights
Neural stem cells (NSCs) have proliferative properties and the ability to generate neurons, astrocytes, and oligodendrocytes [1, 2]
NSCs derived from the developing rat cortex at embryonic day 14 (E14) were maintained as free floating aggregates, termed neurospheres, which can be propagated for several months in the presence of growth factors such as epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF2) [39]
Neurospheres were expanded for one week in EGF and FGF2 and differentiated for 4 days in the absence of growth factors and in the presence of 14 separate phytochemicals at 0.5 μM [Kuwanon V (KWV), ent-16αH,17-hydroxy-kauran-19oic acid, ent-16β,17-dihydroxy-kauran-19-oic acid, kirenol, quercetin-3,40,7-O-trimethyl ether, hinokiflavone, idescarpine, idesolide, dehydrohirsutanonol, oregonin, 1,7-bis(3,4dihydroxyphenyl)-5-hydroxyheptan-3-one, curcumin, lup-20(29)en-2,28-diol-3-yl caffeate, and 20-O-acetylsalicortin] (Fig. 1A)
Summary
Neural stem cells (NSCs) have proliferative properties and the ability to generate neurons, astrocytes, and oligodendrocytes [1, 2]. Neural progenitor cells (NPCs) exist in the neurospheres. NPCs can’t be expanded for long period of time and show reduced neurogenesis in the later passages [3, 4]. PLOS ONE | DOI:10.1371/journal.pone.0118188 February 23, 2015
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