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Neurosphere cell culture is a commonly used model to study the properties and potential applications of neural stem cells (NSCs). However, standard protocols to culture NSCs have yet to be established, and the mechanisms underlying NSC survival and maintenance of their undifferentiated state, in response to the growth factors FGF-2 and EGF are not fully understood. Using cultures of embryonic and adult olfactory bulb stem cells (eOBSCs and aOBSCs), we analyzed the consequences of FGF-2 and EGF addition at different intervals on proliferation, cell cycle progression, cell death and differentiation, as well as on global gene expression. As opposed to cultures supplemented daily, addition of FGF-2 and EGF every 4 days significantly reduced the neurosphere volume and the total number of cells in the spheres, mainly due to increased cell death. Moreover, partial FGF-2 and EGF deprivation produced an increase in OBSC differentiation during the proliferative phase. These changes were more evident in aOBSC than eOBSC cultures. Remarkably, these effects were accompanied by a significant upregulation in the expression of endogenous  Fgf-2  and genes involved in cell death and survival ( Cryab ), lipid catabolic processes ( Pla2g7 ), cell adhesion ( Dscaml1 ), cell differentiation ( Dscaml1 ,  Gpr17 ,  S100b, Ndrg2 ) and signal transduction ( Gpr17 ,  Ndrg2) . These findings support that a daily supply of FGF-2 and EGF is critical to maintain the viability and the undifferentiated state of NSCs in culture, and they reveal novel molecular hallmarks of NSC death, survival and the initiation of differentiation.

plos one

A Global Transcriptome Analysis Reveals Molecular Hallmarks of Neural Stem Cell Death, Survival, and Differentiation in Response to Partial FGF-2 and EGF Deprivation