In the field of developmental biology, there is compelling evidence for a network of activity of pluripotency and stem-associated genes comprising of Oct4, Nanog and nestin. During neurogenesis, the choice between enhancement versus suppression of transcriptional modulation of these identified genes determines the balance between self-renewal neural stem cells (NSC) and immature neuronal phenotypes. By using immunocytochemistry and RT-PCR techniques, our study aims to address the question whether and to what extent mRNA and protein profiles are expressed in human fetal neurospheres obtained from cortical and striatal brain regions, both in expansion (undifferentiated cells) and differentiation conditions monitored after 1 and 4 weeks in vitro culturing. Our results clearly demonstrate the sustained presence of opposite signals: strong downregulation of Oct4 and Nanog genes in cortical differentiating cells and significant up-regulation for nestin gene both in cortical and striatal differentiating cells. Notably, by immunostaining techniques, Oct4 and Nanog protein expression have indicated the presence of both nuclear and cytoplasmic content followed by their rapid turnover (immediately after 1 week). Moreover, during the differentiation process, dissociated neurospheres displayed unexpected number of nestin positive cells accompanied by a constant level of staining intensity. In conclusion, the present study provides new insights into brain region related features in terms of Oct4, Nanog and nestin expression both at cellular and molecular level.
Expression analysis of pluripotency-associated genes in human fetal cortical and striatal neural stem cells during differentiation
DIANA, ANDREA
2012-01-01
Abstract
In the field of developmental biology, there is compelling evidence for a network of activity of pluripotency and stem-associated genes comprising of Oct4, Nanog and nestin. During neurogenesis, the choice between enhancement versus suppression of transcriptional modulation of these identified genes determines the balance between self-renewal neural stem cells (NSC) and immature neuronal phenotypes. By using immunocytochemistry and RT-PCR techniques, our study aims to address the question whether and to what extent mRNA and protein profiles are expressed in human fetal neurospheres obtained from cortical and striatal brain regions, both in expansion (undifferentiated cells) and differentiation conditions monitored after 1 and 4 weeks in vitro culturing. Our results clearly demonstrate the sustained presence of opposite signals: strong downregulation of Oct4 and Nanog genes in cortical differentiating cells and significant up-regulation for nestin gene both in cortical and striatal differentiating cells. Notably, by immunostaining techniques, Oct4 and Nanog protein expression have indicated the presence of both nuclear and cytoplasmic content followed by their rapid turnover (immediately after 1 week). Moreover, during the differentiation process, dissociated neurospheres displayed unexpected number of nestin positive cells accompanied by a constant level of staining intensity. In conclusion, the present study provides new insights into brain region related features in terms of Oct4, Nanog and nestin expression both at cellular and molecular level.
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