elsevier induced pluripotent stem cells yamanaka

We found that promoters of 565 genes were co-bound by four Yamanaka factors in iPS cells, a 10-fold increase when compared with their binding in ES cells. AA treatment facilitates this step but may not be absolutely required ( Figure 2A). Here, we established completely reprogrammed induced pluripotent stem (iPS) cells and analyzed the global promoter occupancy of Yamanaka factors in these cells by ChIP-chip assays. Altogether, these results are consistent with the interpretation that Nanog is important during late stages of reprogramming by facilitating the transition to a stable self-sustaining pluripotency network (as indicated by PECAM1 and hence Oct4 positivity). Whereas nearly all SSEA1 + cells had turned on PECAM1 by day 12 of reprogramming in the presence of AA, a minor population of PECAM1 + cells was also detectable in the absence of AA, and these cultures gave rise to rare iPSC-like cells. Importantly, continuous AA treatment of Nanog –/– reprogramming cultures restored normal PECAM expression at day 9. However, PECAM1 expression was absent from Nanog –/– intermediates at day 9 under serum/LIF conditions. Surprisingly, EPCAM was expressed normally in Nanog –/– nascent iPSC cultures at day 6, indicating that Nanog deficiency affects neither Epcam transcription nor the mid stages of reprogramming. In contrast, PECAM1 expression is activated late (day 9) in iPSC formation and coincides with Oct4 expression in wild-type cells. Ascorbic Acid Rescues the Reprogramming Potential of Nanog-Deficient Cells Collectively, these data demonstrate that the reprogrammed Nanog –/– cells are pluripotent iPSCs and are thus functionally equivalent to Nanog –/– ESCs. We also found that Nanog –/– iPSC-like cells could contribute to adult chimeric mice ( Figure 1G), indicating that these progenitors have the capacity to fully mature and contribute to adult tissues. Additionally, immunohistochemistry of the chimeric embryos for GFP demonstrated that the Nanog –/– iPSC-like cells contributed to all three germ layers, including the neuroectoderm of the brain, the endoderm-derived lining of the gastrointestinal tract, and the mesoderm-derived smooth muscle layers of the gastrointestinal tract ( Figure S1E). These embryos gave rise to GFP + MEFs and GFP + neural progenitor cells (NPCs) in vitro, corroborating that the reprogrammed Nanog –/– cells had the potential to differentiate into mesodermal and a defined ectodermal lineage, respectively ( Figure S1D). We obtained 14 viable E13.5 embryos from 53 implanted blastocyts, of which 11 embryos had variable contributions of GFP chimerism ( Figure 1F). We therefore injected GFP + Nanog –/– iPSC-like cells into E3.5 wild-type blastocysts, transplanted them into the uterus of pseudopregnant recipient females, and isolated resultant fetuses at midgestation. Molecular Therapy Nucleic Acids ( partner)Ī more stringent assay of pluripotency is the ability of cells to contribute to chimeras.Molecular Therapy Methods & Clinical Development ( partner).Human Genetics and Genomics Advances ( partner).American Journal of Human Genetics ( partner).

Elsevier induced pluripotent stem cells yamanaka