A Small-Molecule Inhibitor of Tgf-β Signaling Replaces Sox2 in Reprogramming by Inducing Nanog

Harvard Stem Cell Institute, Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA.
Cell stem cell (Impact Factor: 22.27). 10/2009; 5(5):491-503. DOI: 10.1016/j.stem.2009.09.012
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The combined activity of three transcription factors can reprogram adult cells into induced pluripotent stem cells (iPSCs). However, the transgenic methods used for delivering reprogramming factors have raised concerns regarding the future utility of the resulting stem cells. These uncertainties could be overcome if each transgenic factor were replaced with a small molecule that either directly activated its expression from the somatic genome or in some way compensated for its activity. To this end, we have used high-content chemical screening to identify small molecules that can replace Sox2 in reprogramming. We show that one of these molecules functions in reprogramming by inhibiting Tgf-beta signaling in a stable and trapped intermediate cell type that forms during the process. We find that this inhibition promotes the completion of reprogramming through induction of the transcription factor Nanog.

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Available from: Kyle M Loh, Jun 20, 2014
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    • "In addition, many epigenetic regulators have been showed to play critical roles during the reprogramming process such as enzymes that modulate histone modification and DNA (de) methylation (Apostolou and Hochedlinger, 2013; Buganim et al., 2013; Liang and Zhang, 2013; Theunissen and Jaenisch, 2014). Furthermore, previous studies have uncovered the important roles of multiple signaling pathways during reprogramming such as TGFb (Ichida et al., 2009; Maherali and Hochedlinger, 2009; Liu et al., 2013), BMP (Samavarchi-Tehrani et al., 2010; Chen et al., 2013), Wnt/b-catenin (Marson et al., 2008; Ho et al., 2013), p53-p21 (Hong et al., 2009; Kawamura et al., 2009; Marion et al., 2009), NF-kB (Lee et al., 2012), MAPK/ERK (Silva et al., 2008), mTOR (Wang et al., 2013) and Eras-Akt pathways (Tang et al., 2014; Yu et al., 2014). However, there might be additional factors and signals associated with the reprogramming process. "
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    • "However, we observed a significantly lower reprogramming efficiency compared with reprogramming using a feeder system. Therefore, further modifications were incorporated in the reprogramming process, including the use of hypoxic culture conditions (3%–5% O 2 ) and a transforming growth factor b (TGF-b) inhibitor (A83-01), a selective inhibitor of the TGF-b type I receptor ALK5, which has been shown to enhance reprogramming efficiency (Ichida et al., 2009). We also incorporated Alhydrogel (i.e., an aluminum hydroxide wet gel suspension), which enhanced integration-free reprogramming under defined and feeder-free conditions (Figure S1). "
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    • "This hypothesis was based on our data that HDDCs could upregulate their epithelial phenotype after EMT inhibition and that MET was required to reprogram fibroblasts into iPSCs (Li et al., 2010b). For this protocol, we incubated HDDCs with A83- 01 and VPA, a histone deacetylase inhibitor shown to synergize with RepSox EMT inhibitor (Ichida et al., 2009), for 3 days prior to our R protocol. Compared to the R protocol, these HDDCs had higher INS, PDX1, and NEUROD mRNA expression without significantly different expression of other analyzed markers, including glucagon (Fig. 5A). "
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