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|Title:||Directing neuronal differentiation of primary neural progenitor cells by gene knockdown approach|
|Citation:||Low, W.C., Yau, W.W.Y., Stanton, L.W., Marcy, G., Goh, E., Chew, S.Y. (2012-07-01). Directing neuronal differentiation of primary neural progenitor cells by gene knockdown approach. DNA and Cell Biology 31 (7) : 1148-1160. ScholarBank@NUS Repository. https://doi.org/10.1089/dna.2011.1557|
|Abstract:||Directing differentiation of neural stem/progenitor cells (NPCs) to produce functional neurons is a promising remedy for neural pathological conditions. The major challenge, however, lies in the effective and efficient generation of a sizable population of neurons. A potential strategy is to incorporate RNA interference (RNAi) during directed stem cell differentiation to recapitulate the complex cell-signaling cascades that often occurs during the process. In this study, in vitro silencing of RE1-silencing transcription factor (REST) was carried out using small-interfering RNAs (siRNAs) to evaluate the efficacy of combining REST knockdown with conventional differentiation approaches to enhance neurogenesis. While earlier studies have demonstrated enhanced neuronal lineage commitment from embryonic stem cells and mesenchymal stem cells upon REST knockdown, the effects of REST silencing during other stages of neural development have not been extensively evaluated. We hypothesize that REST knockdown would enhance NPC development to mature neurons and that induced REST silencing can serve as a potential biochemical approach to direct cell fate. Under nonspecific induction conditions, REST knockdown induced eightfold higher Tuj1 mRNA expression at day 14 compared with untransfected cells and cells subjected to scrambled-siRNA treatment (controls). Immunostaining also revealed greater percentage of Tuj1 positive cells with REST knockdown. Combined with neuronal induction, REST silencing enhanced the kinetics of neuronal differentiation and the rate of maturation of committed neuronal cells. Specifically, upregulation of MAP2 occurred as early as 3 days after induction with REST silencing and the expression was comparable to the controls at day 14. Likewise, downregulation of REST generated more than twice the percentage of Tuj1 and MAP2 positive cells compared with controls at day 5 (p|
|Source Title:||DNA and Cell Biology|
|Appears in Collections:||Staff Publications|
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