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Isolation of primitive endoderm, mesoderm, vascular endothelial and trophoblast progenitors from human pluripotent stem cells

Nature Biotechnology volume 30, pages 531–542 (2012)Cite this article

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Abstract

To identify early populations of committed progenitors derived from human embryonic stem cells (hESCs), we screened self-renewing, BMP4-treated and retinoic acid–treated cultures with >400 antibodies recognizing cell-surface antigens. Sorting of >30 subpopulations followed by transcriptional analysis of developmental genes identified four distinct candidate progenitor groups. Subsets detected in self-renewing cultures, including CXCR4+ cells, expressed primitive endoderm genes. Expression of Cxcr4 in primitive endoderm was confirmed in visceral endoderm of mouse embryos. BMP4-induced progenitors exhibited gene signatures of mesoderm, trophoblast and vascular endothelium, suggesting correspondence to gastrulation-stage primitive streak, chorion and allantois precursors, respectively. Functional studies in vitro and in vivo confirmed that ROR2+ cells produce mesoderm progeny, APA+ cells generate syncytiotrophoblasts and CD87+ cells give rise to vasculature. The same progenitor classes emerged during the differentiation of human induced pluripotent stem cells (hiPSCs). These markers and progenitors provide tools for purifying human tissue-regenerating progenitors and for studying the commitment of pluripotent stem cells to lineage progenitors.

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Figure 1: Identification of cell surface markers expressed by candidate progenitor populations.
Figure 2: Analysis of gene expression profiles in sorted hESC-derived populations.
Figure 3: Primitive endoderm characteristics of CXCR4+ cells representing progenitor group no.1.
Figure 4: ROR2+ progenitors (representing group no.2) exhibit characteristics of embryonic mesoderm and generate mesoderm tissues in vivo.
Figure 5: CD87+ progenitors (representing group no.3) exhibit characteristics of endothelial microvasculature.
Figure 6: APA+ progenitors (representing group no.4) exhibit trophoblast characteristics and produce syncytiotrophoblasts by cell fusion.
Figure 7: Similarities between hESC- and hiPSC-derived progenitors and their suggested correspondence to pre- and gastrulation-stage mouse embryonic precursors.

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Acknowledgements

This work was supported by funds provided by the California Institute of Regenerative Medicine (CIRM) (Comprehensive grants RC1-00354-1, RT2-02060). Y.S. was supported by a CDA award from the Human Frontier Science Organization (HFSPO, CDA0063/2007-C). C.T. and A.S.L. are supported by the Howard Hughes Medical Institute Medical Fellows and the Stanford Medical Scholars Program. Y.R. is supported by a Human Frontier Science Program (HFSP) Long Term Fellowship. M.D., C.T. and R.A. are supported by CIRM (Comprehensive grants RC1-00354-1, RT2-02060). We would like to thank C. Chan for pentachrome stainings.

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  1. Micha Drukker

    Present address: Present address: Institute of Stem Cell Research, Helmholtz Zentrum Munich, German Research Center for Environmental Health, Neuherberg, Germany.,

Authors and Affiliations

  1. Institute of Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, California, USA

    Micha Drukker, Chad Tang, Reza Ardehali, Yuval Rinkevich, Jun Seita, Adriane R Mosley & Irving L Weissman

  2. Departments of Radiology and Medicine (Division of Cardiology), Stanford University School of Medicine, Stanford, California, USA

    Andrew S Lee

  3. Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel

    Yoav Soen

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M.D., I.L.W. and Y.S. designed the experiments and wrote the manuscript. M.D., Y.S., C.T., R.A., Y.R., J.S., A.S.L. and A.R.M. performed experiments and analyzed data. All authors endorse the full content of this work.

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Drukker, M., Tang, C., Ardehali, R. et al. Isolation of primitive endoderm, mesoderm, vascular endothelial and trophoblast progenitors from human pluripotent stem cells. Nat Biotechnol 30, 531–542 (2012). https://doi.org/10.1038/nbt.2239

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