Reference Maps of Human ES and iPS Cell Variation Enable High-Throughput Characterization of Pluripotent Cell Lines

Broad Institute, Cambridge, MA 02142, USA.
Cell (Impact Factor: 32.24). 02/2011; 144(3):439-52. DOI: 10.1016/j.cell.2010.12.032
Source: PubMed


The developmental potential of human pluripotent stem cells suggests that they can produce disease-relevant cell types for biomedical research. However, substantial variation has been reported among pluripotent cell lines, which could affect their utility and clinical safety. Such cell-line-specific differences must be better understood before one can confidently use embryonic stem (ES) or induced pluripotent stem (iPS) cells in translational research. Toward this goal we have established genome-wide reference maps of DNA methylation and gene expression for 20 previously derived human ES lines and 12 human iPS cell lines, and we have measured the in vitro differentiation propensity of these cell lines. This resource enabled us to assess the epigenetic and transcriptional similarity of ES and iPS cells and to predict the differentiation efficiency of individual cell lines. The combination of assays yields a scorecard for quick and comprehensive characterization of pluripotent cell lines.

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    • "Third, the reprogramming process itself involves a number of stochastic events, and only a small fraction of cells reaches a fully pluripotent stage (Bock et al., 2011; Sommer et al., 2013). Thus, a number of intermediate cell stages will appear during reprogramming , and cells may get trapped somewhere along the trajectory toward pluripotency (Bock et al., 2011; Boue et al., 2010; Polo et al., 2012). Consequently, the criteria as well as the methods used to define and demonstrate pluripotency must be considered carefully (Smith et al., 2009). "
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    ABSTRACT: Induced pluripotent stem cell (iPSC) technology has become an important tool for disease modeling. Insufficient data on the variability among iPSC lines derived from a single somatic parental cell line have in practice led to generation and analysis of several, usually three, iPSC sister lines from each parental cell line. We established iPSC lines from a human fibroblast line (HDF-K1) and used transcriptome sequencing to investigate the variation among three sister lines (iPSC-K1A, B, and C). For comparison, we analyzed the transcriptome of an iPSC line (iPSC-K5B) derived from a different fibroblast line (HDF-K5), a human embryonic stem cell (ESC) line (ESC-HS181), as well as the two parental fibroblast lines. All iPSC lines fulfilled stringent criteria for pluripotency. In an unbiased cluster analysis, all stem cell lines (four iPSCs and one ESC) clustered together as opposed to the parental fibroblasts. The transcriptome profiles of the three iPSC sister lines were indistinguishable from each other, and functional pathway analysis did not reveal any significant hits. In contrast, the expression profiles of the ESC line and the iPSC-K5B line were distinct from that of the sister lines iPSC-K1A, B, and C. Differentiation to embryoid bodies and subsequent analysis of germ layer markers in the five stem cell clones confirmed that the distribution of their expression profiles was retained. Taken together, our observations stress the importance of using iPSCs of different parental origin rather than several sister iPSC lines to distinguish disease-associated mechanisms from genetic background effects in disease modeling.
    09/2015; 17(5). DOI:10.1089/cell.2015.0009
    • "We demonstrated that certain hiPSC-derived cells such as RPE cells are much less immunogenic than other hiPSCderived cell types such as SMCs. The mechanisms underlying the differential immunogenicity can be very complex, depending on the intrinsic immune characteristics of the cell types and the impact of epigenetic abnormality of hiPSCs on differentiated cell types as previously noted (Bar-Nur et al., 2011; Bock et al., 2011; Kim et al., 2010, 2011; Polo et al., 2010; Ruiz et al., 2012). In support of this notion, we demonstrate that the abnormal expression of immunogenic antigens in some hiPSCderived cells, but not in others, contributes to the differential immunogenicity. "
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    Cell stem cell 08/2015; 17(3). DOI:10.1016/j.stem.2015.07.021 · 22.27 Impact Factor
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    • "Furthermore, by sequencing and combining dozens of low-input methylomes from the same sample (e.g., one-cell, four-cell, or 20-cell pools), it will be possible to create composite methylomes that provide excellent genomewide coverage based on relatively few cells and include an inherent assessment of variation. Essentially, composite methylomes redefine the concept of reference epigenome corridors (Bock et al., 2011) in the context of individual samples, thus providing a new type of reference methylome map that can account for tissue heterogeneity and cell-to-cell variation. "
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    ABSTRACT: Methods for single-cell genome and transcriptome sequencing have contributed to our understanding of cellular heterogeneity, whereas methods for single-cell epigenomics are much less established. Here, we describe a whole-genome bisulfite sequencing (WGBS) assay that enables DNA methylation mapping in very small cell populations (μWGBS) and single cells (scWGBS). Our assay is optimized for profiling many samples at low coverage, and we describe a bioinformatic method that analyzes collections of single-cell methylomes to infer cell-state dynamics. Using these technological advances, we studied epigenomic cell-state dynamics in three in vitro models of cellular differentiation and pluripotency, where we observed characteristic patterns of epigenome remodeling and cell-to-cell heterogeneity. The described method enables single-cell analysis of DNA methylation in a broad range of biological systems, including embryonic development, stem cell differentiation, and cancer. It can also be used to establish composite methylomes that account for cell-to-cell heterogeneity in complex tissue samples. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.
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